REPORT NO 555

Pathogens In Food Proficiency Testing

Program - Round 16

October 2007

ACKNOWLEDGMENTS

PTA wishes to gratefully acknowledge the technical assistance provided by Ms Vivianne Padgham EML Consulting Services Pty Ltd and Ms Kate Lewis IFM Quality Services Pty Ltd who supplied and distributed the samples

copy COPYRIGHT PROFICIENCY TESTING AUSTRALIA 2007

PO Box 7507 Silverwater NSW 2128 AUSTRALIA

TABLE OF CONTENTS

1 FOREWORD 1

2 FEATURES OF THE PROGRAM 1

3 FORMAT OF THE APPENDICES 1

4 DESIGN OF THE PROGRAM 2

5 HOMOGENEITY AND STABILITY TESTING 2

6 FALSE RESULTS 2

TABLE A False Results 3

7 TECHNICAL COMMENTS 3

Response Rate 3

Salmonella Results 4

Listeria Results 4

Listeria Speciation 4

Method Commentary 5

Overall Laboratory Performance 5

8 REFERENCE 6

APPENDICES

APPENDIX A

Summary of Results

Salmonella A11 - A12

Listeria A21 - A25

APPENDIX B

Summary of Methods

Salmonella B11 - B15

Listeria B21 - B24

APPENDIX C

Homogeneity and Stability Testing C11 - C13

APPENDIX D

Instructions to Participants D11

Results Sheets D21 - D23

-1shy

1 FOREWORD

This report summarises the results of round sixteen of a series of proficiency testing programs involving the analysis of different food types for the detection of a range of pathogens

Proficiency Testing Australia conducted the program in September 2007 The aim of the program was to assess laboratories ability to competently perform the nominated tests

2 FEATURES OF THE PROGRAM

(a) A total of 36 laboratories received samples Two laboratories (5 and 9) did not return results

(b) The results reported by participants are presented in Appendix A

(c) Laboratories were provided with five samples of flavoured pudding mix Instructions to Participants and a Results Sheet (see Appendix D)

(d) NATA accredited laboratories were requested to perform the tests according to their routine methods with the Australian Standard method being preferred

(e) Each laboratory was randomly allocated a unique code number for the program to ensure confidentiality of results Reference to each laboratory in this report is by its code number

Please note that some laboratories reported more than one set of results and therefore one code number (with letter) could appear several times in the same data set

3 FORMAT OF THE APPENDICES

APPENDIX AAppendix A contains the results reported by participating laboratories for each ofthe five samples

APPENDIX BAppendix B contains a summary of the methods used by each laboratory

APPENDIX CAppendix C contains the results of the homogeneity and stability testing

APPENDIX DAppendix D contains the ldquoInstructions to Participantsrdquo and pro-forma ldquoResultsSheetrdquo

-2shy

4 DESIGN OF THE PROGRAM

Participants were asked to determine the presence of Salmonella and Listeria in five samples of flavoured pudding mix

Each laboratory was provided with five samples labelled A B C D and E and was requested to test 25 grams of each sample for each analysis

bull Samples A and C were duplicates They contained Listeria monocytogenes and Citrobacter freundii plus the natural flora of the samples The levels of Listeria monocytogenes and Citrobacter freundii in each sample were approximately 250-500 cfu 25 g

bull Samples B and E were duplicates They contained Salmonella Senftenberg and the natural flora of the samples The level of Salmonella Senftenberg in each sample was approximately 150-200 cfu 25 g

bull Sample D contained Salmonella Adelaide and Listeria innocua as well as the natural flora in the sample The levels of Salmonella Adelaide and Listeria innocua in the sample were approximately 250-500 cfu 25 g

5 HOMOGENEITY AND STABILITY TESTING

Prior to sample distribution ten randomly selected samples from each matrix (AC BE and D) were analysed for homogeneity by IFM Quality Services Pty Ltd Based on the results of this testing the homogeneity of the samples was established

Stability testing was also performed on the samples by IFM Quality Services Pty Ltd The results showed that the samples were sufficiently stable for testing during the duration of the program

For more information on the homogeneity and stability testing see Appendix C

6 FALSE RESULTS

Testing methods were pooled and results examined for laboratories reporting false positives and false negatives The false positive and false negative results for this round of the program are summarised in the following table

-3shy

TABLE A FALSE RESULTS(by laboratory code number)

Presence Absence of Salmonella in Flavoured Pudding Mix

Sample False Positives False Negatives

A 35

B 23A 23B 35

C 15 29 35

D -

E 23A 23B 35

Presence Absence of Listeria in Flavoured Pudding Mix

Sample False Positives False Negatives

A -

B -

C -

D -

E -

7 TECHNICAL COMMENTS

Response Rate

Of the 36 laboratories that participated in the program 34 (94) submitted results for inclusion in the final report All of the 34 participants that submitted results for the program reported results for Salmonella Twenty seven of the 34 participants that submitted results for the program (79) reported results for Listeria One laboratory (23) submitted more than one set of results

-4shy

Salmonella Results

Of the 34 participants that submitted results for Salmonella four submitted incorrect results

Laboratories 15 and 29 reported false positive results for sample C while laboratory 35 reported false positive results for both sample A and sample C Samples A and C were duplicate samples that did not contain any Salmonella

Laboratories 23A 23B and 35 reported false negative results for both sample B and sample E Samples B and E were duplicate samples that contained Salmonella Senftenberg It should be noted that the Salmonella Senftenberg used in these samples is a weak H2S producer

Laboratory 35 reported 4 false results This laboratory should investigate their procedures for Salmonella recovery

The failure rates for the five different samples for Salmonella are calculated in Appendix A The overall failure rate for Salmonella is also calculated in Appendix A Here it was found that samples B C and E had the highest failure rate of all the samples for Salmonella (86) followed by sample A (29) There were no false results reported for sample D The overall failure rate for Salmonella was found to be 57

Listeria Results

All of the participants that submitted results for Listeria correctly reported that samples A C and D contained Listeria and that samples B and E did not contain Listeria However two laboratories had problems with Listeria speciation

Listeria Speciation

Of the 27 participants that tested the samples for Listeria 26 (96) submitted results for Listeria monocytogenes All 26 of these laboratories correctly reported the presence of Listeria monocytogenes in samples A and C

Twenty one of the 27 participants that submitted results for Listeria (78) also submitted results for species other than Listeria monocytogenes This includes laboratory 31 which did not complete the section for Other Listeria Species in Table A of the Results Sheet but did indicate that sample D contained Listeria innocua Of these 21 laboratories 18 (88) identified the species of Listeria detected All 18 of these laboratories correctly reported the presence of Listeria innocua in sample D Laboratories 25 and 30 correctly detected the presence of Listeria in sample D but incorrectly identified this as Listeria monocytogenes

-5shy

The failure rates for the five different samples for Listeria are calculated in Appendix A The overall failure rate for Listeria is also calculated in Appendix A

Method Commentary

Salmonella

As in the previous round of this program the majority of the participants reported using cultural methods for the recovery of Salmonella Of these 43 of the participants used the AS 5013 method and 16 used the AS 1766 method Some laboratories substituted RVS for RV in the AS 1766 method Cultural methods not based on the AS standards were used by 5 of laboratories

Immunoassay methods were used by 27 of the laboratories while 8 of the laboratories used PCR techniques

It should be noted that some participants used more than one method but reported only one result

A considerable range of confirmatory tests was reported by the participants with one laboratory not doing any confirmation

Detailed method information is provided in Appendix B1

Listeria

The previous round of this program showed that the majority of the participants used AS 17662262 It was reported by the participants that for this round 34 recoveries were obtained using AS 17662262 34 recoveries using immunoassay methods 26 using PCR and 7 using alternative methods

It should be noted that some participants used more than one method but reported only one result

Confirmation tests were more consistent between participants than the confirmation tests used for Salmonella confirmation

Detailed method information is provided in Appendix B2

Overall Laboratory Performance

The overall failure rate for Salmonella in this round of the program is higher than it has been in previous rounds The overall failure rate for Salmonella of 57 is higher than the 08 failure rate achieved in Round 15 It is also higher than the overall failure rate of 36 achieved in Round 14 for Salmonella (see Reports 520 and 539 for more details)

-6shy

There were no false results reported for Listeria in this round of the program For comparison the overall failure rate for Listeria in Round 15 of this program was 10 while the overall failure rate for Listeria in Round 14 of this program was 33 (see Reports 520 and 539 for more details)

In comparing the failure rates in different rounds of the program the levels of test organisms and the number of participants in the round should also be taken into consideration This information is available in the report issued for each round

8 REFERENCE

Guide to Proficiency Testing Australia (2006)

This document is located on the PTA website at wwwptaasnau under ldquoDocumentsrdquo

TABLE OF CONTENTS

1 FOREWORD 1

2 FEATURES OF THE PROGRAM 1

3 FORMAT OF THE APPENDICES 1

4 DESIGN OF THE PROGRAM 2

5 HOMOGENEITY AND STABILITY TESTING 2

6 FALSE RESULTS 2

TABLE A False Results 3

7 TECHNICAL COMMENTS 3

Response Rate 3

Salmonella Results 4

Listeria Results 4

Listeria Speciation 4

Method Commentary 5

Overall Laboratory Performance 5

8 REFERENCE 6

APPENDICES

APPENDIX A

Summary of Results

Salmonella A11 - A12

Listeria A21 - A25

APPENDIX B

Summary of Methods

Salmonella B11 - B15

Listeria B21 - B24

APPENDIX C

Homogeneity and Stability Testing C11 - C13

APPENDIX D

Instructions to Participants D11

Results Sheets D21 - D23

-1shy

1 FOREWORD

This report summarises the results of round sixteen of a series of proficiency testing programs involving the analysis of different food types for the detection of a range of pathogens

Proficiency Testing Australia conducted the program in September 2007 The aim of the program was to assess laboratories ability to competently perform the nominated tests

2 FEATURES OF THE PROGRAM

(a) A total of 36 laboratories received samples Two laboratories (5 and 9) did not return results

(b) The results reported by participants are presented in Appendix A

(c) Laboratories were provided with five samples of flavoured pudding mix Instructions to Participants and a Results Sheet (see Appendix D)

(d) NATA accredited laboratories were requested to perform the tests according to their routine methods with the Australian Standard method being preferred

(e) Each laboratory was randomly allocated a unique code number for the program to ensure confidentiality of results Reference to each laboratory in this report is by its code number

Please note that some laboratories reported more than one set of results and therefore one code number (with letter) could appear several times in the same data set

3 FORMAT OF THE APPENDICES

APPENDIX AAppendix A contains the results reported by participating laboratories for each ofthe five samples

APPENDIX BAppendix B contains a summary of the methods used by each laboratory

APPENDIX CAppendix C contains the results of the homogeneity and stability testing

APPENDIX DAppendix D contains the ldquoInstructions to Participantsrdquo and pro-forma ldquoResultsSheetrdquo

-2shy

4 DESIGN OF THE PROGRAM

Participants were asked to determine the presence of Salmonella and Listeria in five samples of flavoured pudding mix

Each laboratory was provided with five samples labelled A B C D and E and was requested to test 25 grams of each sample for each analysis

bull Samples A and C were duplicates They contained Listeria monocytogenes and Citrobacter freundii plus the natural flora of the samples The levels of Listeria monocytogenes and Citrobacter freundii in each sample were approximately 250-500 cfu 25 g

bull Samples B and E were duplicates They contained Salmonella Senftenberg and the natural flora of the samples The level of Salmonella Senftenberg in each sample was approximately 150-200 cfu 25 g

bull Sample D contained Salmonella Adelaide and Listeria innocua as well as the natural flora in the sample The levels of Salmonella Adelaide and Listeria innocua in the sample were approximately 250-500 cfu 25 g

5 HOMOGENEITY AND STABILITY TESTING

Prior to sample distribution ten randomly selected samples from each matrix (AC BE and D) were analysed for homogeneity by IFM Quality Services Pty Ltd Based on the results of this testing the homogeneity of the samples was established

Stability testing was also performed on the samples by IFM Quality Services Pty Ltd The results showed that the samples were sufficiently stable for testing during the duration of the program

For more information on the homogeneity and stability testing see Appendix C

6 FALSE RESULTS

Testing methods were pooled and results examined for laboratories reporting false positives and false negatives The false positive and false negative results for this round of the program are summarised in the following table

-3shy

TABLE A FALSE RESULTS(by laboratory code number)

Presence Absence of Salmonella in Flavoured Pudding Mix

Sample False Positives False Negatives

A 35

B 23A 23B 35

C 15 29 35

D -

E 23A 23B 35

Presence Absence of Listeria in Flavoured Pudding Mix

Sample False Positives False Negatives

A -

B -

C -

D -

E -

7 TECHNICAL COMMENTS

Response Rate

Of the 36 laboratories that participated in the program 34 (94) submitted results for inclusion in the final report All of the 34 participants that submitted results for the program reported results for Salmonella Twenty seven of the 34 participants that submitted results for the program (79) reported results for Listeria One laboratory (23) submitted more than one set of results

-4shy

Salmonella Results

Of the 34 participants that submitted results for Salmonella four submitted incorrect results

Laboratories 15 and 29 reported false positive results for sample C while laboratory 35 reported false positive results for both sample A and sample C Samples A and C were duplicate samples that did not contain any Salmonella

Laboratories 23A 23B and 35 reported false negative results for both sample B and sample E Samples B and E were duplicate samples that contained Salmonella Senftenberg It should be noted that the Salmonella Senftenberg used in these samples is a weak H2S producer

Laboratory 35 reported 4 false results This laboratory should investigate their procedures for Salmonella recovery

The failure rates for the five different samples for Salmonella are calculated in Appendix A The overall failure rate for Salmonella is also calculated in Appendix A Here it was found that samples B C and E had the highest failure rate of all the samples for Salmonella (86) followed by sample A (29) There were no false results reported for sample D The overall failure rate for Salmonella was found to be 57

Listeria Results

All of the participants that submitted results for Listeria correctly reported that samples A C and D contained Listeria and that samples B and E did not contain Listeria However two laboratories had problems with Listeria speciation

Listeria Speciation

Of the 27 participants that tested the samples for Listeria 26 (96) submitted results for Listeria monocytogenes All 26 of these laboratories correctly reported the presence of Listeria monocytogenes in samples A and C

Twenty one of the 27 participants that submitted results for Listeria (78) also submitted results for species other than Listeria monocytogenes This includes laboratory 31 which did not complete the section for Other Listeria Species in Table A of the Results Sheet but did indicate that sample D contained Listeria innocua Of these 21 laboratories 18 (88) identified the species of Listeria detected All 18 of these laboratories correctly reported the presence of Listeria innocua in sample D Laboratories 25 and 30 correctly detected the presence of Listeria in sample D but incorrectly identified this as Listeria monocytogenes

-5shy

The failure rates for the five different samples for Listeria are calculated in Appendix A The overall failure rate for Listeria is also calculated in Appendix A

Method Commentary

Salmonella

As in the previous round of this program the majority of the participants reported using cultural methods for the recovery of Salmonella Of these 43 of the participants used the AS 5013 method and 16 used the AS 1766 method Some laboratories substituted RVS for RV in the AS 1766 method Cultural methods not based on the AS standards were used by 5 of laboratories

Immunoassay methods were used by 27 of the laboratories while 8 of the laboratories used PCR techniques

It should be noted that some participants used more than one method but reported only one result

A considerable range of confirmatory tests was reported by the participants with one laboratory not doing any confirmation

Detailed method information is provided in Appendix B1

Listeria

The previous round of this program showed that the majority of the participants used AS 17662262 It was reported by the participants that for this round 34 recoveries were obtained using AS 17662262 34 recoveries using immunoassay methods 26 using PCR and 7 using alternative methods

It should be noted that some participants used more than one method but reported only one result

Confirmation tests were more consistent between participants than the confirmation tests used for Salmonella confirmation

Detailed method information is provided in Appendix B2

Overall Laboratory Performance

The overall failure rate for Salmonella in this round of the program is higher than it has been in previous rounds The overall failure rate for Salmonella of 57 is higher than the 08 failure rate achieved in Round 15 It is also higher than the overall failure rate of 36 achieved in Round 14 for Salmonella (see Reports 520 and 539 for more details)

-6shy

There were no false results reported for Listeria in this round of the program For comparison the overall failure rate for Listeria in Round 15 of this program was 10 while the overall failure rate for Listeria in Round 14 of this program was 33 (see Reports 520 and 539 for more details)

In comparing the failure rates in different rounds of the program the levels of test organisms and the number of participants in the round should also be taken into consideration This information is available in the report issued for each round

8 REFERENCE

Guide to Proficiency Testing Australia (2006)

This document is located on the PTA website at wwwptaasnau under ldquoDocumentsrdquo

-1shy

1 FOREWORD

This report summarises the results of round sixteen of a series of proficiency testing programs involving the analysis of different food types for the detection of a range of pathogens

Proficiency Testing Australia conducted the program in September 2007 The aim of the program was to assess laboratories ability to competently perform the nominated tests

2 FEATURES OF THE PROGRAM

(a) A total of 36 laboratories received samples Two laboratories (5 and 9) did not return results

(b) The results reported by participants are presented in Appendix A

(c) Laboratories were provided with five samples of flavoured pudding mix Instructions to Participants and a Results Sheet (see Appendix D)

(d) NATA accredited laboratories were requested to perform the tests according to their routine methods with the Australian Standard method being preferred

(e) Each laboratory was randomly allocated a unique code number for the program to ensure confidentiality of results Reference to each laboratory in this report is by its code number

Please note that some laboratories reported more than one set of results and therefore one code number (with letter) could appear several times in the same data set

3 FORMAT OF THE APPENDICES

APPENDIX AAppendix A contains the results reported by participating laboratories for each ofthe five samples

APPENDIX BAppendix B contains a summary of the methods used by each laboratory

APPENDIX CAppendix C contains the results of the homogeneity and stability testing

APPENDIX DAppendix D contains the ldquoInstructions to Participantsrdquo and pro-forma ldquoResultsSheetrdquo

-2shy

4 DESIGN OF THE PROGRAM

Participants were asked to determine the presence of Salmonella and Listeria in five samples of flavoured pudding mix

Each laboratory was provided with five samples labelled A B C D and E and was requested to test 25 grams of each sample for each analysis

bull Samples A and C were duplicates They contained Listeria monocytogenes and Citrobacter freundii plus the natural flora of the samples The levels of Listeria monocytogenes and Citrobacter freundii in each sample were approximately 250-500 cfu 25 g

bull Samples B and E were duplicates They contained Salmonella Senftenberg and the natural flora of the samples The level of Salmonella Senftenberg in each sample was approximately 150-200 cfu 25 g

bull Sample D contained Salmonella Adelaide and Listeria innocua as well as the natural flora in the sample The levels of Salmonella Adelaide and Listeria innocua in the sample were approximately 250-500 cfu 25 g

5 HOMOGENEITY AND STABILITY TESTING

Prior to sample distribution ten randomly selected samples from each matrix (AC BE and D) were analysed for homogeneity by IFM Quality Services Pty Ltd Based on the results of this testing the homogeneity of the samples was established

Stability testing was also performed on the samples by IFM Quality Services Pty Ltd The results showed that the samples were sufficiently stable for testing during the duration of the program

For more information on the homogeneity and stability testing see Appendix C

6 FALSE RESULTS

Testing methods were pooled and results examined for laboratories reporting false positives and false negatives The false positive and false negative results for this round of the program are summarised in the following table

-3shy

TABLE A FALSE RESULTS(by laboratory code number)

Presence Absence of Salmonella in Flavoured Pudding Mix

Sample False Positives False Negatives

A 35

B 23A 23B 35

C 15 29 35

D -

E 23A 23B 35

Presence Absence of Listeria in Flavoured Pudding Mix

Sample False Positives False Negatives

A -

B -

C -

D -

E -

7 TECHNICAL COMMENTS

Response Rate

Of the 36 laboratories that participated in the program 34 (94) submitted results for inclusion in the final report All of the 34 participants that submitted results for the program reported results for Salmonella Twenty seven of the 34 participants that submitted results for the program (79) reported results for Listeria One laboratory (23) submitted more than one set of results

-4shy

Salmonella Results

Of the 34 participants that submitted results for Salmonella four submitted incorrect results

Laboratories 15 and 29 reported false positive results for sample C while laboratory 35 reported false positive results for both sample A and sample C Samples A and C were duplicate samples that did not contain any Salmonella

Laboratories 23A 23B and 35 reported false negative results for both sample B and sample E Samples B and E were duplicate samples that contained Salmonella Senftenberg It should be noted that the Salmonella Senftenberg used in these samples is a weak H2S producer

Laboratory 35 reported 4 false results This laboratory should investigate their procedures for Salmonella recovery

The failure rates for the five different samples for Salmonella are calculated in Appendix A The overall failure rate for Salmonella is also calculated in Appendix A Here it was found that samples B C and E had the highest failure rate of all the samples for Salmonella (86) followed by sample A (29) There were no false results reported for sample D The overall failure rate for Salmonella was found to be 57

Listeria Results

All of the participants that submitted results for Listeria correctly reported that samples A C and D contained Listeria and that samples B and E did not contain Listeria However two laboratories had problems with Listeria speciation

Listeria Speciation

Of the 27 participants that tested the samples for Listeria 26 (96) submitted results for Listeria monocytogenes All 26 of these laboratories correctly reported the presence of Listeria monocytogenes in samples A and C

Twenty one of the 27 participants that submitted results for Listeria (78) also submitted results for species other than Listeria monocytogenes This includes laboratory 31 which did not complete the section for Other Listeria Species in Table A of the Results Sheet but did indicate that sample D contained Listeria innocua Of these 21 laboratories 18 (88) identified the species of Listeria detected All 18 of these laboratories correctly reported the presence of Listeria innocua in sample D Laboratories 25 and 30 correctly detected the presence of Listeria in sample D but incorrectly identified this as Listeria monocytogenes

-5shy

The failure rates for the five different samples for Listeria are calculated in Appendix A The overall failure rate for Listeria is also calculated in Appendix A

Method Commentary

Salmonella

As in the previous round of this program the majority of the participants reported using cultural methods for the recovery of Salmonella Of these 43 of the participants used the AS 5013 method and 16 used the AS 1766 method Some laboratories substituted RVS for RV in the AS 1766 method Cultural methods not based on the AS standards were used by 5 of laboratories

Immunoassay methods were used by 27 of the laboratories while 8 of the laboratories used PCR techniques

It should be noted that some participants used more than one method but reported only one result

A considerable range of confirmatory tests was reported by the participants with one laboratory not doing any confirmation

Detailed method information is provided in Appendix B1

Listeria

The previous round of this program showed that the majority of the participants used AS 17662262 It was reported by the participants that for this round 34 recoveries were obtained using AS 17662262 34 recoveries using immunoassay methods 26 using PCR and 7 using alternative methods

It should be noted that some participants used more than one method but reported only one result

Confirmation tests were more consistent between participants than the confirmation tests used for Salmonella confirmation

Detailed method information is provided in Appendix B2

Overall Laboratory Performance

The overall failure rate for Salmonella in this round of the program is higher than it has been in previous rounds The overall failure rate for Salmonella of 57 is higher than the 08 failure rate achieved in Round 15 It is also higher than the overall failure rate of 36 achieved in Round 14 for Salmonella (see Reports 520 and 539 for more details)

-6shy

There were no false results reported for Listeria in this round of the program For comparison the overall failure rate for Listeria in Round 15 of this program was 10 while the overall failure rate for Listeria in Round 14 of this program was 33 (see Reports 520 and 539 for more details)

In comparing the failure rates in different rounds of the program the levels of test organisms and the number of participants in the round should also be taken into consideration This information is available in the report issued for each round

8 REFERENCE

Guide to Proficiency Testing Australia (2006)

This document is located on the PTA website at wwwptaasnau under ldquoDocumentsrdquo

-2shy

4 DESIGN OF THE PROGRAM

Participants were asked to determine the presence of Salmonella and Listeria in five samples of flavoured pudding mix

Each laboratory was provided with five samples labelled A B C D and E and was requested to test 25 grams of each sample for each analysis

bull Samples A and C were duplicates They contained Listeria monocytogenes and Citrobacter freundii plus the natural flora of the samples The levels of Listeria monocytogenes and Citrobacter freundii in each sample were approximately 250-500 cfu 25 g

bull Samples B and E were duplicates They contained Salmonella Senftenberg and the natural flora of the samples The level of Salmonella Senftenberg in each sample was approximately 150-200 cfu 25 g

bull Sample D contained Salmonella Adelaide and Listeria innocua as well as the natural flora in the sample The levels of Salmonella Adelaide and Listeria innocua in the sample were approximately 250-500 cfu 25 g

5 HOMOGENEITY AND STABILITY TESTING

Prior to sample distribution ten randomly selected samples from each matrix (AC BE and D) were analysed for homogeneity by IFM Quality Services Pty Ltd Based on the results of this testing the homogeneity of the samples was established

Stability testing was also performed on the samples by IFM Quality Services Pty Ltd The results showed that the samples were sufficiently stable for testing during the duration of the program

For more information on the homogeneity and stability testing see Appendix C

6 FALSE RESULTS

Testing methods were pooled and results examined for laboratories reporting false positives and false negatives The false positive and false negative results for this round of the program are summarised in the following table

-3shy

TABLE A FALSE RESULTS(by laboratory code number)

Presence Absence of Salmonella in Flavoured Pudding Mix

Sample False Positives False Negatives

A 35

B 23A 23B 35

C 15 29 35

D -

E 23A 23B 35

Presence Absence of Listeria in Flavoured Pudding Mix

Sample False Positives False Negatives

A -

B -

C -

D -

E -

7 TECHNICAL COMMENTS

Response Rate

Of the 36 laboratories that participated in the program 34 (94) submitted results for inclusion in the final report All of the 34 participants that submitted results for the program reported results for Salmonella Twenty seven of the 34 participants that submitted results for the program (79) reported results for Listeria One laboratory (23) submitted more than one set of results

-4shy

Salmonella Results

Of the 34 participants that submitted results for Salmonella four submitted incorrect results

Laboratories 15 and 29 reported false positive results for sample C while laboratory 35 reported false positive results for both sample A and sample C Samples A and C were duplicate samples that did not contain any Salmonella

Laboratories 23A 23B and 35 reported false negative results for both sample B and sample E Samples B and E were duplicate samples that contained Salmonella Senftenberg It should be noted that the Salmonella Senftenberg used in these samples is a weak H2S producer

Laboratory 35 reported 4 false results This laboratory should investigate their procedures for Salmonella recovery

The failure rates for the five different samples for Salmonella are calculated in Appendix A The overall failure rate for Salmonella is also calculated in Appendix A Here it was found that samples B C and E had the highest failure rate of all the samples for Salmonella (86) followed by sample A (29) There were no false results reported for sample D The overall failure rate for Salmonella was found to be 57

Listeria Results

All of the participants that submitted results for Listeria correctly reported that samples A C and D contained Listeria and that samples B and E did not contain Listeria However two laboratories had problems with Listeria speciation

Listeria Speciation

Of the 27 participants that tested the samples for Listeria 26 (96) submitted results for Listeria monocytogenes All 26 of these laboratories correctly reported the presence of Listeria monocytogenes in samples A and C

Twenty one of the 27 participants that submitted results for Listeria (78) also submitted results for species other than Listeria monocytogenes This includes laboratory 31 which did not complete the section for Other Listeria Species in Table A of the Results Sheet but did indicate that sample D contained Listeria innocua Of these 21 laboratories 18 (88) identified the species of Listeria detected All 18 of these laboratories correctly reported the presence of Listeria innocua in sample D Laboratories 25 and 30 correctly detected the presence of Listeria in sample D but incorrectly identified this as Listeria monocytogenes

-5shy

The failure rates for the five different samples for Listeria are calculated in Appendix A The overall failure rate for Listeria is also calculated in Appendix A

Method Commentary

Salmonella

As in the previous round of this program the majority of the participants reported using cultural methods for the recovery of Salmonella Of these 43 of the participants used the AS 5013 method and 16 used the AS 1766 method Some laboratories substituted RVS for RV in the AS 1766 method Cultural methods not based on the AS standards were used by 5 of laboratories

Immunoassay methods were used by 27 of the laboratories while 8 of the laboratories used PCR techniques

It should be noted that some participants used more than one method but reported only one result

A considerable range of confirmatory tests was reported by the participants with one laboratory not doing any confirmation

Detailed method information is provided in Appendix B1

Listeria

The previous round of this program showed that the majority of the participants used AS 17662262 It was reported by the participants that for this round 34 recoveries were obtained using AS 17662262 34 recoveries using immunoassay methods 26 using PCR and 7 using alternative methods

It should be noted that some participants used more than one method but reported only one result

Confirmation tests were more consistent between participants than the confirmation tests used for Salmonella confirmation

Detailed method information is provided in Appendix B2

Overall Laboratory Performance

The overall failure rate for Salmonella in this round of the program is higher than it has been in previous rounds The overall failure rate for Salmonella of 57 is higher than the 08 failure rate achieved in Round 15 It is also higher than the overall failure rate of 36 achieved in Round 14 for Salmonella (see Reports 520 and 539 for more details)

-6shy

There were no false results reported for Listeria in this round of the program For comparison the overall failure rate for Listeria in Round 15 of this program was 10 while the overall failure rate for Listeria in Round 14 of this program was 33 (see Reports 520 and 539 for more details)

In comparing the failure rates in different rounds of the program the levels of test organisms and the number of participants in the round should also be taken into consideration This information is available in the report issued for each round

8 REFERENCE

Guide to Proficiency Testing Australia (2006)

This document is located on the PTA website at wwwptaasnau under ldquoDocumentsrdquo

-3shy

TABLE A FALSE RESULTS(by laboratory code number)

Presence Absence of Salmonella in Flavoured Pudding Mix

Sample False Positives False Negatives

A 35

B 23A 23B 35

C 15 29 35

D -

E 23A 23B 35

Presence Absence of Listeria in Flavoured Pudding Mix

Sample False Positives False Negatives

A -

B -

C -

D -

E -

7 TECHNICAL COMMENTS

Response Rate

Of the 36 laboratories that participated in the program 34 (94) submitted results for inclusion in the final report All of the 34 participants that submitted results for the program reported results for Salmonella Twenty seven of the 34 participants that submitted results for the program (79) reported results for Listeria One laboratory (23) submitted more than one set of results

-4shy

Salmonella Results

Of the 34 participants that submitted results for Salmonella four submitted incorrect results

Laboratories 15 and 29 reported false positive results for sample C while laboratory 35 reported false positive results for both sample A and sample C Samples A and C were duplicate samples that did not contain any Salmonella

Laboratories 23A 23B and 35 reported false negative results for both sample B and sample E Samples B and E were duplicate samples that contained Salmonella Senftenberg It should be noted that the Salmonella Senftenberg used in these samples is a weak H2S producer

Laboratory 35 reported 4 false results This laboratory should investigate their procedures for Salmonella recovery

The failure rates for the five different samples for Salmonella are calculated in Appendix A The overall failure rate for Salmonella is also calculated in Appendix A Here it was found that samples B C and E had the highest failure rate of all the samples for Salmonella (86) followed by sample A (29) There were no false results reported for sample D The overall failure rate for Salmonella was found to be 57

Listeria Results

All of the participants that submitted results for Listeria correctly reported that samples A C and D contained Listeria and that samples B and E did not contain Listeria However two laboratories had problems with Listeria speciation

Listeria Speciation

Of the 27 participants that tested the samples for Listeria 26 (96) submitted results for Listeria monocytogenes All 26 of these laboratories correctly reported the presence of Listeria monocytogenes in samples A and C

Twenty one of the 27 participants that submitted results for Listeria (78) also submitted results for species other than Listeria monocytogenes This includes laboratory 31 which did not complete the section for Other Listeria Species in Table A of the Results Sheet but did indicate that sample D contained Listeria innocua Of these 21 laboratories 18 (88) identified the species of Listeria detected All 18 of these laboratories correctly reported the presence of Listeria innocua in sample D Laboratories 25 and 30 correctly detected the presence of Listeria in sample D but incorrectly identified this as Listeria monocytogenes

-5shy

The failure rates for the five different samples for Listeria are calculated in Appendix A The overall failure rate for Listeria is also calculated in Appendix A

Method Commentary

Salmonella

As in the previous round of this program the majority of the participants reported using cultural methods for the recovery of Salmonella Of these 43 of the participants used the AS 5013 method and 16 used the AS 1766 method Some laboratories substituted RVS for RV in the AS 1766 method Cultural methods not based on the AS standards were used by 5 of laboratories

Immunoassay methods were used by 27 of the laboratories while 8 of the laboratories used PCR techniques

It should be noted that some participants used more than one method but reported only one result

A considerable range of confirmatory tests was reported by the participants with one laboratory not doing any confirmation

Detailed method information is provided in Appendix B1

Listeria

The previous round of this program showed that the majority of the participants used AS 17662262 It was reported by the participants that for this round 34 recoveries were obtained using AS 17662262 34 recoveries using immunoassay methods 26 using PCR and 7 using alternative methods

It should be noted that some participants used more than one method but reported only one result

Confirmation tests were more consistent between participants than the confirmation tests used for Salmonella confirmation

Detailed method information is provided in Appendix B2

Overall Laboratory Performance

The overall failure rate for Salmonella in this round of the program is higher than it has been in previous rounds The overall failure rate for Salmonella of 57 is higher than the 08 failure rate achieved in Round 15 It is also higher than the overall failure rate of 36 achieved in Round 14 for Salmonella (see Reports 520 and 539 for more details)

-6shy

There were no false results reported for Listeria in this round of the program For comparison the overall failure rate for Listeria in Round 15 of this program was 10 while the overall failure rate for Listeria in Round 14 of this program was 33 (see Reports 520 and 539 for more details)

In comparing the failure rates in different rounds of the program the levels of test organisms and the number of participants in the round should also be taken into consideration This information is available in the report issued for each round

8 REFERENCE

Guide to Proficiency Testing Australia (2006)

This document is located on the PTA website at wwwptaasnau under ldquoDocumentsrdquo

-4shy

Salmonella Results

Of the 34 participants that submitted results for Salmonella four submitted incorrect results

Laboratories 15 and 29 reported false positive results for sample C while laboratory 35 reported false positive results for both sample A and sample C Samples A and C were duplicate samples that did not contain any Salmonella

Laboratories 23A 23B and 35 reported false negative results for both sample B and sample E Samples B and E were duplicate samples that contained Salmonella Senftenberg It should be noted that the Salmonella Senftenberg used in these samples is a weak H2S producer

Laboratory 35 reported 4 false results This laboratory should investigate their procedures for Salmonella recovery

The failure rates for the five different samples for Salmonella are calculated in Appendix A The overall failure rate for Salmonella is also calculated in Appendix A Here it was found that samples B C and E had the highest failure rate of all the samples for Salmonella (86) followed by sample A (29) There were no false results reported for sample D The overall failure rate for Salmonella was found to be 57

Listeria Results

All of the participants that submitted results for Listeria correctly reported that samples A C and D contained Listeria and that samples B and E did not contain Listeria However two laboratories had problems with Listeria speciation

Listeria Speciation

Of the 27 participants that tested the samples for Listeria 26 (96) submitted results for Listeria monocytogenes All 26 of these laboratories correctly reported the presence of Listeria monocytogenes in samples A and C

Twenty one of the 27 participants that submitted results for Listeria (78) also submitted results for species other than Listeria monocytogenes This includes laboratory 31 which did not complete the section for Other Listeria Species in Table A of the Results Sheet but did indicate that sample D contained Listeria innocua Of these 21 laboratories 18 (88) identified the species of Listeria detected All 18 of these laboratories correctly reported the presence of Listeria innocua in sample D Laboratories 25 and 30 correctly detected the presence of Listeria in sample D but incorrectly identified this as Listeria monocytogenes

-5shy

The failure rates for the five different samples for Listeria are calculated in Appendix A The overall failure rate for Listeria is also calculated in Appendix A

Method Commentary

Salmonella

As in the previous round of this program the majority of the participants reported using cultural methods for the recovery of Salmonella Of these 43 of the participants used the AS 5013 method and 16 used the AS 1766 method Some laboratories substituted RVS for RV in the AS 1766 method Cultural methods not based on the AS standards were used by 5 of laboratories

Immunoassay methods were used by 27 of the laboratories while 8 of the laboratories used PCR techniques

It should be noted that some participants used more than one method but reported only one result

A considerable range of confirmatory tests was reported by the participants with one laboratory not doing any confirmation

Detailed method information is provided in Appendix B1

Listeria

The previous round of this program showed that the majority of the participants used AS 17662262 It was reported by the participants that for this round 34 recoveries were obtained using AS 17662262 34 recoveries using immunoassay methods 26 using PCR and 7 using alternative methods

It should be noted that some participants used more than one method but reported only one result

Confirmation tests were more consistent between participants than the confirmation tests used for Salmonella confirmation

Detailed method information is provided in Appendix B2

Overall Laboratory Performance

The overall failure rate for Salmonella in this round of the program is higher than it has been in previous rounds The overall failure rate for Salmonella of 57 is higher than the 08 failure rate achieved in Round 15 It is also higher than the overall failure rate of 36 achieved in Round 14 for Salmonella (see Reports 520 and 539 for more details)

-6shy

There were no false results reported for Listeria in this round of the program For comparison the overall failure rate for Listeria in Round 15 of this program was 10 while the overall failure rate for Listeria in Round 14 of this program was 33 (see Reports 520 and 539 for more details)

In comparing the failure rates in different rounds of the program the levels of test organisms and the number of participants in the round should also be taken into consideration This information is available in the report issued for each round

8 REFERENCE

Guide to Proficiency Testing Australia (2006)

This document is located on the PTA website at wwwptaasnau under ldquoDocumentsrdquo

-5shy

The failure rates for the five different samples for Listeria are calculated in Appendix A The overall failure rate for Listeria is also calculated in Appendix A

Method Commentary

Salmonella

As in the previous round of this program the majority of the participants reported using cultural methods for the recovery of Salmonella Of these 43 of the participants used the AS 5013 method and 16 used the AS 1766 method Some laboratories substituted RVS for RV in the AS 1766 method Cultural methods not based on the AS standards were used by 5 of laboratories

Immunoassay methods were used by 27 of the laboratories while 8 of the laboratories used PCR techniques

It should be noted that some participants used more than one method but reported only one result

A considerable range of confirmatory tests was reported by the participants with one laboratory not doing any confirmation

Detailed method information is provided in Appendix B1

Listeria

The previous round of this program showed that the majority of the participants used AS 17662262 It was reported by the participants that for this round 34 recoveries were obtained using AS 17662262 34 recoveries using immunoassay methods 26 using PCR and 7 using alternative methods

It should be noted that some participants used more than one method but reported only one result

Confirmation tests were more consistent between participants than the confirmation tests used for Salmonella confirmation

Detailed method information is provided in Appendix B2

Overall Laboratory Performance

The overall failure rate for Salmonella in this round of the program is higher than it has been in previous rounds The overall failure rate for Salmonella of 57 is higher than the 08 failure rate achieved in Round 15 It is also higher than the overall failure rate of 36 achieved in Round 14 for Salmonella (see Reports 520 and 539 for more details)

-6shy

There were no false results reported for Listeria in this round of the program For comparison the overall failure rate for Listeria in Round 15 of this program was 10 while the overall failure rate for Listeria in Round 14 of this program was 33 (see Reports 520 and 539 for more details)

In comparing the failure rates in different rounds of the program the levels of test organisms and the number of participants in the round should also be taken into consideration This information is available in the report issued for each round

8 REFERENCE

Guide to Proficiency Testing Australia (2006)

This document is located on the PTA website at wwwptaasnau under ldquoDocumentsrdquo

-6shy

There were no false results reported for Listeria in this round of the program For comparison the overall failure rate for Listeria in Round 15 of this program was 10 while the overall failure rate for Listeria in Round 14 of this program was 33 (see Reports 520 and 539 for more details)

In comparing the failure rates in different rounds of the program the levels of test organisms and the number of participants in the round should also be taken into consideration This information is available in the report issued for each round

8 REFERENCE

Guide to Proficiency Testing Australia (2006)

This document is located on the PTA website at wwwptaasnau under ldquoDocumentsrdquo

APPENDIX A

Summary of Results

Section A1

Salmonella

A11

Salmonella Results

Lab Code A B C D E False

Results 1 Absent Present Absent Present Present 2 Absent Present Absent Present Present 3 Absent Present Absent Present Present 4 Absent Present Absent Present Present 6 Absent Present Absent Present Present 7 Absent Present Absent Present Present 8 Absent Present Absent Present Present

10 Absent Present Absent Present Present 11 Absent Present Absent Present Present 12 Absent Present Absent Present Present 13 Absent Present Absent Present Present 14 Absent Present Absent Present Present 15 Absent Present Present Present Present 1 16 Absent Present Absent Present Present 17 Absent Present Absent Present Present 18 Absent Present Absent Present Present 19 Absent Present Absent Present Present 20 Absent Present Absent Present Present 21 Absent Present Absent Present Present 22 Absent Present Absent Present Present

23A Absent Absent Absent Present Absent 2 23B Absent Absent Absent Present Absent 2 24 Absent Present Absent Present Present 25 Absent Present Absent Present Present 26 Absent Present Absent Present Present 27 Absent Present Absent Present Present 28 Absent Present Absent Present Present 29 Absent Present Present Present Present 1 30 Absent Present Absent Present Present 31 Absent Present Absent Present Present 32 Absent Present Absent Present Present 33 Absent Present Absent Present Present 34 Absent Present Absent Present Present 35 Present Absent Present Present Absent 4 36 Absent Present Absent Present Present

Note

A highlighted result (ie bold print) is a false result and should be investigated

A12

Salmonella Failure Rate

No of Results Sample

Total A B C D E

False Results 1 3 3 0 3 10

Total Results 35 35 35 35 35 175

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 1 35

= 29

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 3 35

= 86

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 3 35

= 86

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 35

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 3 35

= 86

Overall failure rate Total No of False Results = (Salmonella) Total No of Results

= 10 175

= 57

Section A2

Listeria

12

3456

A21

Listeria Results

Lab Code A B C D E False

Results 2 Present Absent Present Present Absent 3 Present Absent Present Present Absent 6 Present Absent Present Present Absent 8 Present Absent Present Present Absent

10 Present Absent Present Present Absent 11 Present Absent Present Present Absent 12 Present Absent Present Present Absent 13 Present Absent Present Present Absent 14 Present Absent Present Present Absent 16 Present Absent Present Present Absent 18 Present Absent Present Present Absent 19 Present Absent Present Present Absent 20 Present Absent Present Present Absent 21 Present Absent Present Present Absent 24 Present Absent Present Present Absent 25 Present Absent Present Present Absent 26 Present Absent Present Present Absent 27 Present Absent Present Present Absent 28 Present Absent Present Present Absent 29 Present Absent Present Present Absent 30 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

A22

Listeria Failure Rate

No of Results Sample

Total A B C D E

False Results 0 0 0 0 0 0

Total Results 27 27 27 27 27 135

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 0 27

= 0

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 0 27

= 0

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 0 27

= 0

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 27

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 0 27

= 0

Overall failure rate Total No of False Results = (Listeria) Total No of Results

= 0 135

= 0

A23

Listeria Speciation ndash Listeria monocytogenes

Lab Code A B C D E False

Results 2 Present Absent Present Absent Absent 3 Present - Present Absent -6 Present - Present Absent -8 Present - Present - Absent

10 Present Absent Present Absent Absent 11 Present Absent Present Absent Absent 12 Present Absent Present Absent Absent 13 Present Absent Present Absent Absent 14 Present Absent Present Absent Absent 16 Present Not done Present Absent Not done 18 Present Absent Present Absent Absent 19 Present - Present Absent -20 Present Absent Present Absent Absent 21 Present Absent Present Absent Absent 24 Present Absent Present Absent Absent 25 Present Absent Present Present Absent 1 26 Present - Present Absent -27 Present Absent Present Absent Absent 28 Present - Present Absent Absent 30 Present Absent Present Present Absent 1 31 Present - Present Absent Absent 32 Present Absent Present Absent Absent 33 Present Absent Present Absent Absent 34 Present Absent Present Absent Absent 35 Present Absent Present Absent Absent 36 Present Absent Present Absent Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 8 did not report whether or not sample D contained Listeria

monocytogenes This laboratory did however report that sample D contained another Listeria species (see the table on page A24) It is possible therefore that the ldquoabsentrdquo result reported by laboratory 8 for sample E should have been reported for sample D

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Section A1

Salmonella

A11

Salmonella Results

Lab Code A B C D E False

Results 1 Absent Present Absent Present Present 2 Absent Present Absent Present Present 3 Absent Present Absent Present Present 4 Absent Present Absent Present Present 6 Absent Present Absent Present Present 7 Absent Present Absent Present Present 8 Absent Present Absent Present Present

10 Absent Present Absent Present Present 11 Absent Present Absent Present Present 12 Absent Present Absent Present Present 13 Absent Present Absent Present Present 14 Absent Present Absent Present Present 15 Absent Present Present Present Present 1 16 Absent Present Absent Present Present 17 Absent Present Absent Present Present 18 Absent Present Absent Present Present 19 Absent Present Absent Present Present 20 Absent Present Absent Present Present 21 Absent Present Absent Present Present 22 Absent Present Absent Present Present

23A Absent Absent Absent Present Absent 2 23B Absent Absent Absent Present Absent 2 24 Absent Present Absent Present Present 25 Absent Present Absent Present Present 26 Absent Present Absent Present Present 27 Absent Present Absent Present Present 28 Absent Present Absent Present Present 29 Absent Present Present Present Present 1 30 Absent Present Absent Present Present 31 Absent Present Absent Present Present 32 Absent Present Absent Present Present 33 Absent Present Absent Present Present 34 Absent Present Absent Present Present 35 Present Absent Present Present Absent 4 36 Absent Present Absent Present Present

Note

A highlighted result (ie bold print) is a false result and should be investigated

A12

Salmonella Failure Rate

No of Results Sample

Total A B C D E

False Results 1 3 3 0 3 10

Total Results 35 35 35 35 35 175

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 1 35

= 29

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 3 35

= 86

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 3 35

= 86

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 35

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 3 35

= 86

Overall failure rate Total No of False Results = (Salmonella) Total No of Results

= 10 175

= 57

Section A2

Listeria

12

3456

A21

Listeria Results

Lab Code A B C D E False

Results 2 Present Absent Present Present Absent 3 Present Absent Present Present Absent 6 Present Absent Present Present Absent 8 Present Absent Present Present Absent

10 Present Absent Present Present Absent 11 Present Absent Present Present Absent 12 Present Absent Present Present Absent 13 Present Absent Present Present Absent 14 Present Absent Present Present Absent 16 Present Absent Present Present Absent 18 Present Absent Present Present Absent 19 Present Absent Present Present Absent 20 Present Absent Present Present Absent 21 Present Absent Present Present Absent 24 Present Absent Present Present Absent 25 Present Absent Present Present Absent 26 Present Absent Present Present Absent 27 Present Absent Present Present Absent 28 Present Absent Present Present Absent 29 Present Absent Present Present Absent 30 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

A22

Listeria Failure Rate

No of Results Sample

Total A B C D E

False Results 0 0 0 0 0 0

Total Results 27 27 27 27 27 135

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 0 27

= 0

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 0 27

= 0

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 0 27

= 0

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 27

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 0 27

= 0

Overall failure rate Total No of False Results = (Listeria) Total No of Results

= 0 135

= 0

A23

Listeria Speciation ndash Listeria monocytogenes

Lab Code A B C D E False

Results 2 Present Absent Present Absent Absent 3 Present - Present Absent -6 Present - Present Absent -8 Present - Present - Absent

10 Present Absent Present Absent Absent 11 Present Absent Present Absent Absent 12 Present Absent Present Absent Absent 13 Present Absent Present Absent Absent 14 Present Absent Present Absent Absent 16 Present Not done Present Absent Not done 18 Present Absent Present Absent Absent 19 Present - Present Absent -20 Present Absent Present Absent Absent 21 Present Absent Present Absent Absent 24 Present Absent Present Absent Absent 25 Present Absent Present Present Absent 1 26 Present - Present Absent -27 Present Absent Present Absent Absent 28 Present - Present Absent Absent 30 Present Absent Present Present Absent 1 31 Present - Present Absent Absent 32 Present Absent Present Absent Absent 33 Present Absent Present Absent Absent 34 Present Absent Present Absent Absent 35 Present Absent Present Absent Absent 36 Present Absent Present Absent Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 8 did not report whether or not sample D contained Listeria

monocytogenes This laboratory did however report that sample D contained another Listeria species (see the table on page A24) It is possible therefore that the ldquoabsentrdquo result reported by laboratory 8 for sample E should have been reported for sample D

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

A11

Salmonella Results

Lab Code A B C D E False

Results 1 Absent Present Absent Present Present 2 Absent Present Absent Present Present 3 Absent Present Absent Present Present 4 Absent Present Absent Present Present 6 Absent Present Absent Present Present 7 Absent Present Absent Present Present 8 Absent Present Absent Present Present

10 Absent Present Absent Present Present 11 Absent Present Absent Present Present 12 Absent Present Absent Present Present 13 Absent Present Absent Present Present 14 Absent Present Absent Present Present 15 Absent Present Present Present Present 1 16 Absent Present Absent Present Present 17 Absent Present Absent Present Present 18 Absent Present Absent Present Present 19 Absent Present Absent Present Present 20 Absent Present Absent Present Present 21 Absent Present Absent Present Present 22 Absent Present Absent Present Present

23A Absent Absent Absent Present Absent 2 23B Absent Absent Absent Present Absent 2 24 Absent Present Absent Present Present 25 Absent Present Absent Present Present 26 Absent Present Absent Present Present 27 Absent Present Absent Present Present 28 Absent Present Absent Present Present 29 Absent Present Present Present Present 1 30 Absent Present Absent Present Present 31 Absent Present Absent Present Present 32 Absent Present Absent Present Present 33 Absent Present Absent Present Present 34 Absent Present Absent Present Present 35 Present Absent Present Present Absent 4 36 Absent Present Absent Present Present

Note

A highlighted result (ie bold print) is a false result and should be investigated

A12

Salmonella Failure Rate

No of Results Sample

Total A B C D E

False Results 1 3 3 0 3 10

Total Results 35 35 35 35 35 175

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 1 35

= 29

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 3 35

= 86

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 3 35

= 86

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 35

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 3 35

= 86

Overall failure rate Total No of False Results = (Salmonella) Total No of Results

= 10 175

= 57

Section A2

Listeria

12

3456

A21

Listeria Results

Lab Code A B C D E False

Results 2 Present Absent Present Present Absent 3 Present Absent Present Present Absent 6 Present Absent Present Present Absent 8 Present Absent Present Present Absent

10 Present Absent Present Present Absent 11 Present Absent Present Present Absent 12 Present Absent Present Present Absent 13 Present Absent Present Present Absent 14 Present Absent Present Present Absent 16 Present Absent Present Present Absent 18 Present Absent Present Present Absent 19 Present Absent Present Present Absent 20 Present Absent Present Present Absent 21 Present Absent Present Present Absent 24 Present Absent Present Present Absent 25 Present Absent Present Present Absent 26 Present Absent Present Present Absent 27 Present Absent Present Present Absent 28 Present Absent Present Present Absent 29 Present Absent Present Present Absent 30 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

A22

Listeria Failure Rate

No of Results Sample

Total A B C D E

False Results 0 0 0 0 0 0

Total Results 27 27 27 27 27 135

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 0 27

= 0

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 0 27

= 0

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 0 27

= 0

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 27

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 0 27

= 0

Overall failure rate Total No of False Results = (Listeria) Total No of Results

= 0 135

= 0

A23

Listeria Speciation ndash Listeria monocytogenes

Lab Code A B C D E False

Results 2 Present Absent Present Absent Absent 3 Present - Present Absent -6 Present - Present Absent -8 Present - Present - Absent

10 Present Absent Present Absent Absent 11 Present Absent Present Absent Absent 12 Present Absent Present Absent Absent 13 Present Absent Present Absent Absent 14 Present Absent Present Absent Absent 16 Present Not done Present Absent Not done 18 Present Absent Present Absent Absent 19 Present - Present Absent -20 Present Absent Present Absent Absent 21 Present Absent Present Absent Absent 24 Present Absent Present Absent Absent 25 Present Absent Present Present Absent 1 26 Present - Present Absent -27 Present Absent Present Absent Absent 28 Present - Present Absent Absent 30 Present Absent Present Present Absent 1 31 Present - Present Absent Absent 32 Present Absent Present Absent Absent 33 Present Absent Present Absent Absent 34 Present Absent Present Absent Absent 35 Present Absent Present Absent Absent 36 Present Absent Present Absent Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 8 did not report whether or not sample D contained Listeria

monocytogenes This laboratory did however report that sample D contained another Listeria species (see the table on page A24) It is possible therefore that the ldquoabsentrdquo result reported by laboratory 8 for sample E should have been reported for sample D

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

A12

Salmonella Failure Rate

No of Results Sample

Total A B C D E

False Results 1 3 3 0 3 10

Total Results 35 35 35 35 35 175

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 1 35

= 29

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 3 35

= 86

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 3 35

= 86

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 35

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 3 35

= 86

Overall failure rate Total No of False Results = (Salmonella) Total No of Results

= 10 175

= 57

Section A2

Listeria

12

3456

A21

Listeria Results

Lab Code A B C D E False

Results 2 Present Absent Present Present Absent 3 Present Absent Present Present Absent 6 Present Absent Present Present Absent 8 Present Absent Present Present Absent

10 Present Absent Present Present Absent 11 Present Absent Present Present Absent 12 Present Absent Present Present Absent 13 Present Absent Present Present Absent 14 Present Absent Present Present Absent 16 Present Absent Present Present Absent 18 Present Absent Present Present Absent 19 Present Absent Present Present Absent 20 Present Absent Present Present Absent 21 Present Absent Present Present Absent 24 Present Absent Present Present Absent 25 Present Absent Present Present Absent 26 Present Absent Present Present Absent 27 Present Absent Present Present Absent 28 Present Absent Present Present Absent 29 Present Absent Present Present Absent 30 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

A22

Listeria Failure Rate

No of Results Sample

Total A B C D E

False Results 0 0 0 0 0 0

Total Results 27 27 27 27 27 135

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 0 27

= 0

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 0 27

= 0

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 0 27

= 0

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 27

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 0 27

= 0

Overall failure rate Total No of False Results = (Listeria) Total No of Results

= 0 135

= 0

A23

Listeria Speciation ndash Listeria monocytogenes

Lab Code A B C D E False

Results 2 Present Absent Present Absent Absent 3 Present - Present Absent -6 Present - Present Absent -8 Present - Present - Absent

10 Present Absent Present Absent Absent 11 Present Absent Present Absent Absent 12 Present Absent Present Absent Absent 13 Present Absent Present Absent Absent 14 Present Absent Present Absent Absent 16 Present Not done Present Absent Not done 18 Present Absent Present Absent Absent 19 Present - Present Absent -20 Present Absent Present Absent Absent 21 Present Absent Present Absent Absent 24 Present Absent Present Absent Absent 25 Present Absent Present Present Absent 1 26 Present - Present Absent -27 Present Absent Present Absent Absent 28 Present - Present Absent Absent 30 Present Absent Present Present Absent 1 31 Present - Present Absent Absent 32 Present Absent Present Absent Absent 33 Present Absent Present Absent Absent 34 Present Absent Present Absent Absent 35 Present Absent Present Absent Absent 36 Present Absent Present Absent Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 8 did not report whether or not sample D contained Listeria

monocytogenes This laboratory did however report that sample D contained another Listeria species (see the table on page A24) It is possible therefore that the ldquoabsentrdquo result reported by laboratory 8 for sample E should have been reported for sample D

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Section A2

Listeria

12

3456

A21

Listeria Results

Lab Code A B C D E False

Results 2 Present Absent Present Present Absent 3 Present Absent Present Present Absent 6 Present Absent Present Present Absent 8 Present Absent Present Present Absent

10 Present Absent Present Present Absent 11 Present Absent Present Present Absent 12 Present Absent Present Present Absent 13 Present Absent Present Present Absent 14 Present Absent Present Present Absent 16 Present Absent Present Present Absent 18 Present Absent Present Present Absent 19 Present Absent Present Present Absent 20 Present Absent Present Present Absent 21 Present Absent Present Present Absent 24 Present Absent Present Present Absent 25 Present Absent Present Present Absent 26 Present Absent Present Present Absent 27 Present Absent Present Present Absent 28 Present Absent Present Present Absent 29 Present Absent Present Present Absent 30 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

A22

Listeria Failure Rate

No of Results Sample

Total A B C D E

False Results 0 0 0 0 0 0

Total Results 27 27 27 27 27 135

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 0 27

= 0

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 0 27

= 0

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 0 27

= 0

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 27

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 0 27

= 0

Overall failure rate Total No of False Results = (Listeria) Total No of Results

= 0 135

= 0

A23

Listeria Speciation ndash Listeria monocytogenes

Lab Code A B C D E False

Results 2 Present Absent Present Absent Absent 3 Present - Present Absent -6 Present - Present Absent -8 Present - Present - Absent

10 Present Absent Present Absent Absent 11 Present Absent Present Absent Absent 12 Present Absent Present Absent Absent 13 Present Absent Present Absent Absent 14 Present Absent Present Absent Absent 16 Present Not done Present Absent Not done 18 Present Absent Present Absent Absent 19 Present - Present Absent -20 Present Absent Present Absent Absent 21 Present Absent Present Absent Absent 24 Present Absent Present Absent Absent 25 Present Absent Present Present Absent 1 26 Present - Present Absent -27 Present Absent Present Absent Absent 28 Present - Present Absent Absent 30 Present Absent Present Present Absent 1 31 Present - Present Absent Absent 32 Present Absent Present Absent Absent 33 Present Absent Present Absent Absent 34 Present Absent Present Absent Absent 35 Present Absent Present Absent Absent 36 Present Absent Present Absent Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 8 did not report whether or not sample D contained Listeria

monocytogenes This laboratory did however report that sample D contained another Listeria species (see the table on page A24) It is possible therefore that the ldquoabsentrdquo result reported by laboratory 8 for sample E should have been reported for sample D

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

12

3456

A21

Listeria Results

Lab Code A B C D E False

Results 2 Present Absent Present Present Absent 3 Present Absent Present Present Absent 6 Present Absent Present Present Absent 8 Present Absent Present Present Absent

10 Present Absent Present Present Absent 11 Present Absent Present Present Absent 12 Present Absent Present Present Absent 13 Present Absent Present Present Absent 14 Present Absent Present Present Absent 16 Present Absent Present Present Absent 18 Present Absent Present Present Absent 19 Present Absent Present Present Absent 20 Present Absent Present Present Absent 21 Present Absent Present Present Absent 24 Present Absent Present Present Absent 25 Present Absent Present Present Absent 26 Present Absent Present Present Absent 27 Present Absent Present Present Absent 28 Present Absent Present Present Absent 29 Present Absent Present Present Absent 30 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

3 Present Absent Present Present Absent 3 Present Absent Present Present Absent 3 Present Absent Present Present Absent

A22

Listeria Failure Rate

No of Results Sample

Total A B C D E

False Results 0 0 0 0 0 0

Total Results 27 27 27 27 27 135

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 0 27

= 0

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 0 27

= 0

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 0 27

= 0

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 27

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 0 27

= 0

Overall failure rate Total No of False Results = (Listeria) Total No of Results

= 0 135

= 0

A23

Listeria Speciation ndash Listeria monocytogenes

Lab Code A B C D E False

Results 2 Present Absent Present Absent Absent 3 Present - Present Absent -6 Present - Present Absent -8 Present - Present - Absent

10 Present Absent Present Absent Absent 11 Present Absent Present Absent Absent 12 Present Absent Present Absent Absent 13 Present Absent Present Absent Absent 14 Present Absent Present Absent Absent 16 Present Not done Present Absent Not done 18 Present Absent Present Absent Absent 19 Present - Present Absent -20 Present Absent Present Absent Absent 21 Present Absent Present Absent Absent 24 Present Absent Present Absent Absent 25 Present Absent Present Present Absent 1 26 Present - Present Absent -27 Present Absent Present Absent Absent 28 Present - Present Absent Absent 30 Present Absent Present Present Absent 1 31 Present - Present Absent Absent 32 Present Absent Present Absent Absent 33 Present Absent Present Absent Absent 34 Present Absent Present Absent Absent 35 Present Absent Present Absent Absent 36 Present Absent Present Absent Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 8 did not report whether or not sample D contained Listeria

monocytogenes This laboratory did however report that sample D contained another Listeria species (see the table on page A24) It is possible therefore that the ldquoabsentrdquo result reported by laboratory 8 for sample E should have been reported for sample D

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

A22

Listeria Failure Rate

No of Results Sample

Total A B C D E

False Results 0 0 0 0 0 0

Total Results 27 27 27 27 27 135

No of False Results (A) Failure rate (Sample A) = Total No of Results (A) = 0 27

= 0

Failure rate (Sample B) = No of False Results (B) Total No of Results (B)

= 0 27

= 0

No of False Results (C) Failure rate (Sample C) = Total No of Results (C) = 0 27

= 0

Failure rate (Sample D) = No of False Results (D) Total No of Results (D)

= 0 27

= 0

Failure rate (Sample E) = No of False Results (E) Total No of Results (E)

= 0 27

= 0

Overall failure rate Total No of False Results = (Listeria) Total No of Results

= 0 135

= 0

A23

Listeria Speciation ndash Listeria monocytogenes

Lab Code A B C D E False

Results 2 Present Absent Present Absent Absent 3 Present - Present Absent -6 Present - Present Absent -8 Present - Present - Absent

10 Present Absent Present Absent Absent 11 Present Absent Present Absent Absent 12 Present Absent Present Absent Absent 13 Present Absent Present Absent Absent 14 Present Absent Present Absent Absent 16 Present Not done Present Absent Not done 18 Present Absent Present Absent Absent 19 Present - Present Absent -20 Present Absent Present Absent Absent 21 Present Absent Present Absent Absent 24 Present Absent Present Absent Absent 25 Present Absent Present Present Absent 1 26 Present - Present Absent -27 Present Absent Present Absent Absent 28 Present - Present Absent Absent 30 Present Absent Present Present Absent 1 31 Present - Present Absent Absent 32 Present Absent Present Absent Absent 33 Present Absent Present Absent Absent 34 Present Absent Present Absent Absent 35 Present Absent Present Absent Absent 36 Present Absent Present Absent Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 8 did not report whether or not sample D contained Listeria

monocytogenes This laboratory did however report that sample D contained another Listeria species (see the table on page A24) It is possible therefore that the ldquoabsentrdquo result reported by laboratory 8 for sample E should have been reported for sample D

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

A23

Listeria Speciation ndash Listeria monocytogenes

Lab Code A B C D E False

Results 2 Present Absent Present Absent Absent 3 Present - Present Absent -6 Present - Present Absent -8 Present - Present - Absent

10 Present Absent Present Absent Absent 11 Present Absent Present Absent Absent 12 Present Absent Present Absent Absent 13 Present Absent Present Absent Absent 14 Present Absent Present Absent Absent 16 Present Not done Present Absent Not done 18 Present Absent Present Absent Absent 19 Present - Present Absent -20 Present Absent Present Absent Absent 21 Present Absent Present Absent Absent 24 Present Absent Present Absent Absent 25 Present Absent Present Present Absent 1 26 Present - Present Absent -27 Present Absent Present Absent Absent 28 Present - Present Absent Absent 30 Present Absent Present Present Absent 1 31 Present - Present Absent Absent 32 Present Absent Present Absent Absent 33 Present Absent Present Absent Absent 34 Present Absent Present Absent Absent 35 Present Absent Present Absent Absent 36 Present Absent Present Absent Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 8 did not report whether or not sample D contained Listeria

monocytogenes This laboratory did however report that sample D contained another Listeria species (see the table on page A24) It is possible therefore that the ldquoabsentrdquo result reported by laboratory 8 for sample E should have been reported for sample D

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

A24

Listeria Speciation ndash Other Listeria Species

Lab Code A B C D E False

Results 2 Absent - Absent Present -3 - - - Present -8 - - - Present -

10 Absent Not done Absent Present Not done 11 Absent Absent Absent Present Absent 12 - Absent - Present Absent 13 Absent Absent Absent Present Absent 19 Absent - Absent Present -20 - - - Present -21 Absent - Absent Present -24 Absent Absent Absent Present Absent 25 Absent Absent Absent Absent Absent 1 26 Absent - Absent Present -27 Absent Absent Absent Present Absent 28 - - - Present -31 - - - - -32 Absent Absent Absent Present Absent 33 Absent Absent Absent Present Absent 34 Absent Absent Absent Present Absent 35 Absent - Absent Present -36 Not done Absent Not done Present Absent

Notes

1 A highlighted result (ie bold print) is a false result and should be investigated 2 Laboratory 31 did not record if other Listeria species were present in the

samples on Table A of the Results Sheet but did indicate that sample D contained Listeria innocua (see the table on page A25)

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

A25

Listeria Speciation

Lab Code A C D

2 3 6 8 10 11 12 13 14 16 18 19 20 21 24 25 26 27 28 30 31 32 33 34 35 36

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes L monocytogenes

L innocua L innocua

--

L innocua L innocua L innocua L innocua

----

L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua

L monocytogenes L innocua L innocua L innocua L innocua L innocua L innocua

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

APPENDIX B

Summary of Methods

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

SECTION B1

Salmonella

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Code

Salmonella ndash Media

Non-selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 Buffered peptone

water

ICS Strip -Immunoconcentration

(VIDAS) ICS-F broth VIDAS SLM Not done Not done

2 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG

3 Buffered peptone

water MSC broth RV broth XLD BSA TSI urea L-lysineLDC B-galactosidaseONPG O amp H serotyping

4 Buffered peptone

water RVS broth MKTTn XLD BGA TSI Urea L-lysineLDC VP Indole Oxoid Salmonella antigen kit rapid 20E kit

6 Buffered peptone

water RVS broth M-broth XLD VIDAS shy

Salmonella (Biomerieux)

BSA BAX (PCR) Salmonella (Dupont)

kit

Plate onto non-selective agar Microbact 24E (Oxoid) kit O amp H serotyping

7 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Enterotube kit

8 Buffered peptone

water MKTTn RVS broth XLD BGS Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

10 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping Remel Europe Ltd agglutination serum

11 Buffered peptone

water MSC broth RVS broth XLD BSA Urea L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

12 Buffered peptone

water

RVS broth SX broth (both AS Std amp Vidas SLM SX performed)

MKTTn XLD BGA SMID2 L-lysineLDC B-galactosidaseONPG O amp Vi amp H serotyping

13 Buffered peptone

water RVS broth MKTTn XLD BSA Urea L-lysineLDC B-galactosidaseONPG Microbact kit Serobact ID kit

14 Buffered peptone

water Salmonella X-press

broth XLD Salmonella chromogenic medium Plate onto non-selective agar API 20E kit oxidase

15 Buffered peptone

water RVS broth MKTTn XLD Brilliant green agar Plate onto non-selective agar Microbact 24E kit Salmonella (Oxoid) ID kit

16 Buffered peptone

water Not done Not done ABAX PCR Not done Plate onto non-selective agar API 20E kit O Vi amp H serotyping

17 Buffered peptone

water Not done Not done XLD Brilliant Green Agar

with Sulphamandelate

Plate onto non-selective agar L-lysineLDC B-galactosidaseONPG O serotyping

18 Buffered peptone

water MSC broth RVS broth XLD BSA Plate onto non-selective agar Microbact 24E kit O amp H serotyping Oxoid Salmonella serology test kit BAX

B11

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

191 Buffered peptone

water RV MSC broth XLD BSA ID 20E kit O amp H serotyping

192 Buffered peptone water

Not done Not done BAX PCR Plate onto non-selective agar ID 20E kit

20 Buffered peptone

water RVS broth MKTTn XLD Salmonella Chromogenic Agar

Plate onto non-selective agar TSI urea L-lysineLDC B-galactosidaseONPG VP indole OBIS Salmonella kit O amp H serotyping Microbact MB 12E ID kit

21 Buffered peptone water

RVS broth MKTTn M-broth XLD BSA L-lysineLDC B-galactosidaseONPG Microbact GNB 24E kit O Vi amp H serotyping

22 Buffered peptone water

MKTTn RVS broth XLD Chromogenic Salmonella

TSI API 20E kit O amp H serotyping (Oxoid) Latex ID kit

23A Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

23B Not done Modified buffered peptone water

Not done Tecra Unique Not done Not done

24 Buffered peptone water

MSC broth RV broth M-broth Tecra performed

XLD BSA XLD BSA Plate onto non-selective agar TSI Microbact 12A kit

25 Buffered peptone water

RVS broth Not done Tecra Ultima XLD Chromogenic Salmonella

Microbact 12E amp Latex

26 Modified buffered peptone

Tecra Unique Not done XLD MLCB Plate onto non-selective agar API 20E kit gram stain amp oxidase

27 Buffered peptone water

MSC broth RV broth M-broth XLD VIDAS SLM HEA (Hektoen Enteric Agar)

Plate onto non-selective agar API 10S (Biomerieux) kit BAX system PCR assay

28 Buffered peptone water

RVS broth MKTTn BSA XLD Microbact GNB 12A 12B kit

29 Buffered peptone water

MKTTn RVS broth XLD BGA TSI L-lysineLDC B-galactosidaseONPG indole O amp H serotyping

30 Buffered peptone water

MKTTn RVS broth XLD BGA Urea B-galactosidaseONPG Vitek kit

31 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar TSI Microbact kit O Vi amp H serotyping Oxoid biological sera ID kit

32 Buffered peptone water

RVS broth MKTTn XLDHK agar SMID2 Plate onto non-selective agar O amp H serotyping

33 Buffered peptone water

MKTTn RVS broth XLD BSA Plate onto non-selective agar commercial kit O amp H serotyping Microbact 12A12E ID kit

B12

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Salmonella ndash Media

Code Non-selective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation

XLD VIDAS

34 Buffered peptone

water RVS broth M-broth performed then

plated onto XLD SMID2 Plate onto non-selective agar API 20E

SMID2

35 Buffered peptone

water RVS broth MKTTn XLD BSA Plate onto non-selective agar (NAgar) L-lysineLDC B-galactosidaseONPG indole Microbact (Enterobacteria 24E) kit O Vi amp H serotyping

36 Buffered peptone

water MKTTn RVS broth XLD BSA O amp H serotyping Glissuda tube

Abbreviations BGA brilliant green agar BHI brain-heart infusion broth BSA bismuth sulfite agar CLED cystine lactose electrolyte deficient agar DCA desoxycholate agar LDC lysine decarboxylase MSC mannitol selenite cystine MSRV mannitol selenite Rappaport-Vassiliadis ONPG o-nitrophenyl-β-D-galactopyranoside RV Rappaport Vassiliadis TSI triple sugar iron agar XLD xylose lysine desoxycholate agar

B13

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

1 37 41 24 5

2 37 37 42 37 37 37 18 24 24 48 24

3 37 37 42 37 37 37 18 24 24 24 48 24

4 37 42 37 37 37 37 24 24 24 24 24 24

6 37 42 42 37 37 37 18 6 18 24 24 48 18

7 37 415 37 37 35 35 18 24 24 24 24 24

8 37 37 42 37 37 37 24 24 24 24 24 24

10 37 37 42 37 37 37 24 24 24 24 48 24

11 37 37 42 37 37 37 24 24 24 24 48 24

12 37 42 37 37 37 18 18 18 24 24

13 37 42 37 37 37 37 24 24 24 24 48 24

14 37 42 37 37 37 24 24 24 24 48 24

15 37 42 37 37 37 37 18 24 24 24 24 24

16 37 37 18 24

17 35 35 35 35 18 24 24 24

18 35 35 42 35 35 35 24 24 24 24 48 24

191 37 42 37 37 37 37 24 24 24 24 48 24

192 37 37 24 24

B14

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Code

Salmonella ndash Incubation Temperature (degdegdegdegC) Salmonella ndash Duration of Incubation (hours)

Nonshyselective Selective 1 Selective 2 Detection

Isolation 1 Detection Isolation 2 Confirmation Nonshy

selective Selective 1 Selective 2 Detection Isolation 1

Detection Isolation 2 Confirmation

20 37 42 37 37 37 37 24 24 24 24 24 24

21 37 37 42 37 37 37 37 10 7 18 24 48 24

22 37 37 42 37 37 37 18 24 24 24 24 24

23A 37 37 16 5

23B 37 37 18 5

24 37 MSB 37 RVB42 37 37 37 37 18 6 18 24 48 24

25 37 42 37 24 24 24 48

26 37 37 37 37 18 as per kit inst 24 24

27 37 37 42 42 37 37 37 18 8 18 24 24 24

28 37 42 37 37 37 37 18 24 24 24 24 24

29 37 37 42 37 37 37 24 24 24 24 24 24

30 35 35 42 35 18 24 18 24 18 24 18 24

31 37 37 42 37 37 37 18 18 18 24 48 18

32 37 42 37 37 37 37 24 24 24 24 24 24

33 37 37 42 37 37 37 18 24 24 24 48 24

34 37 42 42 37 37 37 24 6-8 16-20 16-20 24 24 24

35 37 42 37 37 37 37 24 24 24 24 48 24

36 37 37 42 37 37 37 24 24 24 24 48 24

B15

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

SECTION B2

Listeria

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

2 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Plate onto non-selective agar motility test API Listeria ID kit gram stain

3 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

VIDAS - Listeria

6 Not done Half Fraser broth Full strength Fraser broth

Oxford agar Palcam agar

(Biomerieux) Elisa kit BAX (PCR) Genus

Listeria (Dupont) kit

Plate onto non-selective agar CAMP test motility test Microbact 12L (Oxoid) ID kit BAX (PCR) L monocytogenes kit (Dupont)

8 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar DNA probe BAX

Plate onto non-selective agar β haemolysis CAMP test sugar fermentation tests API ID kit

10 Not done Buffered Listeria enrichment broth Not done Oxford agar VIDAS Elisa kit β haemolysis CAMP test motility test sugar fermentation tests

11 Buffered Listeria enrichment broth Oxford agar Palcam agar VIDAS

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

12 Half Fraser broth Full strength Fraser broth Oxford agar Vidas Lis Elisa kit β haemolysis motility test API ID kit

13 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar β haemolysis API ID kit

14 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

Chromogenic Listeria agar Plate onto non-selective agar motility test gram stain amp catalase

16 Not done Half Fraser broth MOPS - BLEB Not done Not done ABAX PCR Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

18 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX DNA probe

Plate onto non-selective agar β haemolysis RLM gram stain catalase

191 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done Plate onto non-selective agar β haemolysis CAMP test latex test

192 Half Fraser broth MOPS - BLEB Not done Not done BAX PCR β haemolysis CAMP test

20 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests

21 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX

β haemolysis CAMP test motility test sugar fermentation tests Microbact 12L ID kit

24 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact 12L ID kit

B21

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Code

Listeria ndash Media

Non-selective Primary Second Selective Agar Plating 1 Agar Plating 2 Other Detection Confirmation

25 Not done Buffered Listeria enrichment broth

Full strength Fraser broth Not done Not done

Tecra VIA Elisa kit

Plate onto non-selective agar β haemolysis CAMP test motility test Microbact 12L ID kit

26 Not done Buffered Listeria enrichment broth Not done Oxford agar

Rapid LMD by Bio-rad Elisa kit

Plate onto non-selective agar β haemolysis CAMP test API Listeria ID kit gram stain amp catalase

27 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS LIS

Plate onto non-selective agar motility test API Listeria ID kit BAX system PCR assay

28 Half Fraser broth Full strength Fraser broth Oxford agar Not done

Plate onto non-selective agar β haemolysis motility test Microbact Listeria ID kit

29 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Motility test

30 UVM-I Full strength Fraser broth Oxford agar

β haemolysis CAMP test motility test sugar fermentation tests MB 12L ID kit

31 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar

VIDAS LIS Elisa kit

Plate onto non-selective agar β haemolysis motility test Microbact ID kit

32 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Plate onto non-selective agar API Listeria ID kit

33 Not done Demi Fraser

broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis Microbact 12L Listeria ID kit

34 Not done Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar VIDAS Elisa kit

Plate onto non-selective agar motility test Listeria API ID kit catalase gram stain Henry illumination

35 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar Not done

Plate onto non-selective agar β haemolysis CAMP test motility test sugar fermentation tests Microbact (Listeria 12L) ID kit

36 Half Fraser broth Full strength Fraser broth Oxford agar Palcam agar BAX PCR

Plate onto non-selective agar β haemolysis serological test Listeria API ID kit

B22

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

2 30 37 37 37 37 24 48 48 48 24

3 30 37 37 37 37 24 48 48 48 24 sugars up

to 7 days

6 30 30 30 37 37 24 24 48 48 24

8 30 37 37 37 37 24 48 48 48 24

10 30 37 37 motility room temp 48 48

24 motility 5 days

11 30 37 37

motility 25 sugars β

haemolysis CAMP 37

24 48 48

CAMP 24 β haemolysis

48 sugars amp

motility 5 days

12 30 30 37 24 24 24

13 30 37 37 37 37 24 48 48 48 24

14 30 30 37 37 37 30 24 24 48 48 24 48 24

16 30 35 25 37 24 24 24 48

18 30 35 35 35 24 48 48 48

nonshy

191 30 37 37 37 selective 30 CAMP

24 48 48 48 24

37

192 30 37 37 24 24 24

20 30 30 37 37 37 24 24 48 48 48

21 30 30 30 30 30 24 24 48 48 24

24 30 30 37 37 motility 25

37 24 24 24 24 24 motility 72

B23

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Lab Code

Listeria ndash Incubation Temperature (degdegdegdegC) Listeria ndash Duration of Incubation (hours)

Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm Nonshyselective Primary Second

Selective Agar

Plating 1 Agar

Plating 2 Other

Detection Confirm

25 30 30 24 24

26 30 48

27 30 30 37 37 37 24 24 48 48 24

28 30 30 37 25 37 24 24 48 24

29 30 37 37 37 25 24 48 48 48 24

30 30 37 37 24 48 24 48

31 30 37 37 37 30 37 24 48 48 48 48 24

32 30 30 37 37 37 24 24 48 48 24

33 30 37 37 37 37 24 48 48 48 24

34 30 30 37 37 37 24 24 48 48 24

35 30 37 37 37 37 24 48 48 48 24

36 30 37 37 37 37 24 48 48 48 24

B24

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

APPENDIX C

Homogeneity and

Stability Testing

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

C11

HOMOGENEITY TESTING RESULTS

Ten samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd to confirm that the samples were homogeneous Homogeneity testing was completed and the results were analysed prior to sample dispatch For Salmonella CCFRA Method 312 2003 Detection of Salmonella Cultural Method and CCFRA 315 2003 Confirmation of Salmonella were followed For Listeria CCFRA Method 322 Detection of Listeria Species Cultural Method Using Fraser Broth and Two Stage Enrichment and CCFRA 327 2003 Confirmation of Listeria Species were followed The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 20807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072324 Not detected Detected Not detected 072325 Not detected Detected Not detected 072326 Not detected Detected Not detected 072327 Not detected Detected Not detected 072328 Not detected Detected Not detected 072329 Not detected Detected Not detected 072330 Not detected Detected Not detected 072331 Not detected Detected Not detected 072332 Not detected Detected Not detected 072333 Not detected Detected Not detected

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 23807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072334 Detected Not detected Not detected 072335 Detected Not detected Not detected 072336 Detected Not detected Not detected 072337 Detected Not detected Not detected 072338 Detected Not detected Not detected 072339 Detected Not detected Not detected 072340 Detected Not detected Not detected 072341 Detected Not detected Not detected 072342 Detected Not detected Not detected 072343 Detected Not detected Not detected

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

C12

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 27807

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072344 Detected Not detected Detected 072345 Detected Not detected Detected 072346 Detected Not detected Detected 072347 Detected Not detected Detected 072348 Detected Not detected Detected 072349 Detected Not detected Detected 072350 Detected Not detected Detected 072351 Detected Not detected Detected 072352 Detected Not detected Detected 072353 Detected Not detected Detected

Based on the above testing results the homogeneity of the samples was established

STABILITY TESTING RESULTS

To determine whether the samples used for this program were stable three samples from each matrix (AC BE and D) were randomly chosen and set aside for testing by IFM Quality Services Pty Ltd The stability testing took place on 10 September 2007 The results are tabulated below

Note In the following tables ldquoListeria spprdquo results mean Listeria species other than Listeria monocytogenes

Samples AC (containing Listeria monocytogenes)

Samples AC ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072354 Not detected Detected Not detected 072355 Not detected Detected Not detected 072356 Not detected Detected Not detected

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

C13

Samples BE (containing Salmonella Senftenberg)

Samples BE ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072357 Detected Not detected Not detected 072358 Detected Not detected Not detected 072359 Detected Not detected Not detected

Sample D (containing Salmonella Adelaide and Listeria innocua)

Sample D ndash tested 10907

IFM Lab No Salmonella 25 g L monocytogenes 25 g Listeria spp 25 g (see Note)

072360 Detected Not detected Detected 072361 Detected Not detected Detected 072362 Detected Not detected Detected

Based on these results the samples were considered to be stable during the period that this proficiency testing program was conducted

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

APPENDIX D

Instructions to Participants

and

Results Sheets

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

D11

PROFICIENCY TESTING AUSTRALIAPATHOGENS IN FOOD PROGRAM ndash ROUND 16

INSTRUCTIONS TO PARTICIPANTS

On receipt of samples

Open the container immediately and check the contents are in order

bull Record the temperature of the samples bull Return the contents to the original packaging bull Transfer the samples to a refrigerator (2-5degC) for storage prior to testing bull Protect the samples from light

Prior to testing please note

bull Five samples (labelled A B C D E) each containing 60 g of flavoured pudding mix are to be tested for the presenceabsence of Salmonella and Listeria as per instructions below

bull Samples are for laboratory use only bull Store your samples in the original packaging between 2-5degC until testing commences bull Testing is to commence on 4 September 2007 or as soon as possible after this date Testing

must not commence after 7 September 2007 bull Where practical your laboratory is encouraged to test different samples using different analysts bull Laboratories are required to perform all tests for which NATA accreditation is held or sought and

are welcome to report results for any remaining tests bull Listeria speciation is not mandatory but is encouraged bull Salmonella serotyping is not required bull Your laboratory has been allocated the code number shown on the attached Results and Method

Information Sheets

Instructions

You have been supplied with flavoured pudding mix in plastic sachets

1 Aseptically open the sachets Weigh out 25 g for each test to be performed (Salmonella Listeria)

2 Proceed as per your laboratory test method 3 The Australian Standard Methods are the preferred test methods 4 Report results as presence or absence per 25 gram of sample in Table A of the supplied Results

Sheets by filling in (bull) in the appropriate circles If Salmonella or Listeria are not detected in a sample then this should be indicated by filling in (bull) in the circle alongside ldquoabsentrdquo

5 Report all method information in Tables B and C of the supplied Results Sheets by filling in (bull) in the appropriate circles If more than one method is used for a test report each result separately (copy and use a separate Results Sheet for each method)

Please return results no later than Friday 14 September 2007 to

Mark Bunt Proficiency Testing Australia PO Box 7507 Silverwater NSW 2128 AUSTRALIA

Telephone + 61 2 9736 8397 (1300 782 867) Fax +61 2 9743 6664

Pathogens in Food 16 September 2007 Page 1 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

PTA ldquoPathogens in Foodrdquo Round 16 Proficiency Testing Program

RESULTS SHEET

Date samples arrived Sample temperature Date testing began Signature Laboratory Code

Table A Results

Test Sample A Sample B Sample C Sample D Sample E

Salmonella

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Listeria

monocytogenes

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Present

Absent

Not done

Other

Listeria

Species

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

Present

Indicate species

Absent

Not done

____________

D21

Pathogens in Food 16 September 2007 Page 2 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Table B Method Information Salmonella Laboratory Code Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Enrichment

Selective Enrichment Medium 1

Selective Enrichment Medium 2

DetectionIsolation Confirmation of suspect Salmonella

Media Not done Not done Not done Not done Not done Not done

Buffered

peptone water

MSC broth

RVS broth

MSC broth

RVS broth PLATING Medium 1

XLD

PLATING Medium 2

XLD

Plate onto non-selective agar

BIOCHEMICAL Other (specify) MKTTn MKTTn

BSA BSA TSI ______________ M-broth

Other

(specify)

M-broth

Other

(specify)

Other (specify)

_____________

Other (specify)

________________

Urea L-lysine LDC Β-galactosidase ONPG VP Indole _____________ _____________ Commercial kit (specify)

_ _ ________________

SEROTYPING

O Vi H Commercial ID kit (specify)

________________

Other (specify) __________________

Incubation 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC 35ordmC Temperature 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC

42ordmC 42ordmC 42ordmC 42ordmC 42ordmC 42ordmC Other ____ ordmC Other ____ ordmC Other ____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

18 hours 24 hours 48 hours

72 hours 72 hours 72 hours 72 hours 72 hours 72 hours Other_______ Other_______ Other_______ Other________ Other________ Other __________

D22

Pathogens in Food 16 September 2007 Page 3 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4

Table C Method Information Listeria Laboratory Code

Please indicate the methodology used by filling in (bull) in the appropriate circles below

Method Non-selective Primary Secondary Selective Agar Plating Other Detection Confirmation of Enrichment Selective

Enrichment Selective

Enrichment Medium 1 Medium 2 Methods suspect Listeria

Media Not done Not done Not done Not done Not done Not done Not done Tryptone soy broth with yeast extract Other (specify)

_____________

Buffered Listeria enrichment broth Half Fraser broth UVM-I Other (specify)

_____________

Full strength Fraser broth UVM-II Other (specify)

_____________

Oxford agar Other agar (specify)

_____________

Palcam agar Other agar (specify)

_____________

Elisa kit (specify type)

DNA probe (specify type)

_____________

Plate onto nonshyselective agar β haemolysis CAMP test Motility test Growth in anaerobic conditions

_____________ Sugar fermentation tests

Other detection Latex test methods (specify type)

DNA probe Serological test Commercial ID

_____________ kit (specify)

_____________

Other (specify)

_____________

Incubation 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC 25ordmC Temperature 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC 30ordmC

37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC 37ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC Other _____ ordmC

Duration of Incubation

4 hours 24 hours 48 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

24 hours 48 hours 72 hours Other

__________ __________ __________ __________ __________ __________ __________

D23

Pathogens in Food 16 September 2007 Page 4 of 4