Monitor and Control of Vertically Transmitted Poultry Diseases

Monitor and Control of Vertically Transmitted Poultry Diseases

Rafael Monleon, DVM, MSpVM, ACPV, PAS Business Unit Manager (Poultry)

Biochek Seminar – Manila, Philippines 29th March 2014

OUTLINE •  Types of Transmission

– Horizontal vs. Vertical

•  Bacterial Vertically Transmitted Diseases – Monitor & Control

•  Viral Vertically Transmitted Diseases – Monitor & Control

•  Summary

HORIZONTAL TRANSMISSION

VERTICAL TRANSMISSION

Important Poultry Vertically Transmitted Bacterial Diseases

•  Mycoplasma – Mycoplasma gallisepticum – Mycoplasma synoviae

•  Salmonella – Salmonella pullorum / S. gallinarum – Salmonella enteritidis / S. typhimurium

Important Poultry Vertically Transmitted Bacterial Diseases

•  Persistent infection (Chronic) makes bacterial VT diseases dentrimental

•  Life long carriers are observed even in well treated flocks

•  Ban on several antibiotic makes treatment difficult

•  Erradication or Vaccination are the most common options

Mycoplasma •  M. gallisepticum / M. synoviae •  Probably the most prevalent vertically

transmitted disease worldwide •  Losses in parents / progeny •  Hard to remove from farms

– Shed rate highest (30%) acute vs. decreases (to about 5%) chronic phase of the disease.

– Persistent problem. Carriers

7

Mycoplasma gallisepticum

•  In breeding chickens (GPs/PS) – Respiratory signs including sinusitis – Decreased egg production – Secondary Egg Yolk Peritonitis – Airsac in embryos

•  Causing late dead or first week mortalities/infections

•  In broilers – Mainly CRD

Mycoplasma Losses

9 Hy-line

MG/MS in Broilers Flock Av. Wt. Age % % Status (grams) FC Days Mort. Cond.

MG/MS Negative 2525 2.182 51.8 4.23 0.15 MG Positive 2378 2.205 52.9 7.05 1.47 MS Positive 2461 2.190 52.7 6.23 0.87

Source: LA Integration, 1991

Sinusitis

Sinusitis

Airsacculitis

Airsacculitis/Pericarditis

CRD

Mycoplasma synoviae (MS) (Infectious synovitis)

•  Mild respiratory disease •  Inflammation of the synovial sheaths. •  Chickens and Turkeys.

–  It is common in commercial laying flocks. –  In chickens mostly silent type infections

•  Synovitis rarely seen

•  Young birds - 4 to 12 weeks of age. •  Can also contribute to CRD in broilers •  Less virulent than MG •  Spreads faster than MG.

Swollen hock and foot pad

Synovi5s

Keel bursitis Synovial exudates

Car5lage erosions Chronic synovi5s

Airsacculitis

Mycoplasma synoviae

•  Some new conditions recently reported •  Eggshell Apex Abnormalities (EAA) since

2000. Also called Top Cone Abnormalities. – Mostly in Commercial Layers – Netherlands, South Africa and Japan – Estimate 2-3 egg losses / bird and –  5% loss in downgrades – MS with IBV D1466 > higher incidence

MS Egg Effects-EAA or Top Coning

•  PIP Analysis

•  Serology – RPA

– ELISA

– HI – mainly confirmation

•  PCR – qPCR

Monitoring

PIP Analysis

•  Routine observation •  Airsacculitis - Baby

Chicks or DIS pips •  When see in DIS or

day old chicks this is almost a pathognomic lesion

Mycoplasma Monitoring Farm MG MS

Day Old X X

6wks X X

16wks X X

24wks X X

34 wks X X

44wks X X

54wks X X

*testing every 3-4 weeks in production might be necessary

Serology Rapid Plate Agglutination

•  Sensitive. –  IgM antibody

•  Detects around – 7 to 10 days •  Screening – a flock test. •  Prone to false reactions •  May not detect atypical strains

Serology False Positives with RPA

•  Frozen sera. •  Too long in coldest area of refrigerator. •  Killed TC antigens/Oil Emulsion vaccines. •  Use of fetal calf serum – MG/MS antigens. •  Coryza bacterins.

Serology False Positive with RPA

•  Antigen too sensitive

•  Erysipelas infections

•  Staph bacterins and/or infections

•  Contaminated sera

•  Cross reaction with MS

Serology False Positive with MG RPA

•  Dilute serum 1:10.

•  Mix serum with equal volume of horse or swine sera.

•  Labs that use 2-fold dilutions consider > 1:8 as positive

Serology HI Test

•  More specific •  Positive 1:80, suspicious 1:40 •  Confirmatory •  Antigens not readily available •  Appears later than RPA (14 days) •  Antigen quality and variations •  Atypical strains

ELISA

•  Becoming the most common form of monitoring

–  Mass testing / Affordable (Combo)

•  In general a bit less sensitive but more specific

than RPA (IgM vs. IgG)

•  Less specific but more sensitive than HI

•  However Biochek MS test kit can detect 7d post

infection

–  Recombinant antigen

ELISA Ringtrial Results

MS Field Isolate in 4 Wk Old SPF Chickens , 7 DPI*

2010 2011

RPA Ms** 33% 17%

Mg/Ms(r) ELISA 100% 100%

Other Mg/Ms ELISAs** 17% 44%

% positivesAssay

GD Deventer

Results (MS WVU1853 Inoculated)

0

20

40

60

80

100

0 7 9 14 16

% Posi*ve

Days Post-‐Challenge

Mg/Ms(r) ELISA

Mg/Ms (c) ELISA

RPA (1&2)

RPA (3)

Results (MS K5664 Inoculated)

0

20

40

60

80

100

0 7 9 14 16

% Posi*ve

Days Post-‐Challenge

Mg/Ms(r) ELISA

Mg/Ms (c) ELISA

RPA (all 3)

Mycoplasma gallisepticum Day-old Chick Monitoring

36

0

2

4

6

8

10

12

14

16

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

# Samples Lot:

Mean Titer:

G.M.T.:

Titer Range Ref. Controls:

TiterS/P Ratio Titer Group

2 09/11/2007

Name : R1-IMPORTF.3-1ROSSCompany :

Code : S07-370Age : 01DType : GPHouse No. : L-1

Mg FS463305/11/2007 09/11/2007

5615282

R6 (700-2000)

Titer Group

ARBOR ACRES THAILAND CO.,LTD. 10/3 Soi Chuemsumphan25 Chuemsumphan Rd. Nongjok Bangkok 10530 THAILAND

A01 0.098A02 0.098A03 0.821A04 0.856A0501 0.096A0602 0.100A0703 0.101A0804 0.102A0905 0.099A1006 0.101A1107 0.099A1208 0.440B0109 0.104B0210 0.185B0311 0.118B0412 0.126B0513 0.095B0614 0.094B0715 0.100

613

NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -

- 0.000- 0.000+ 0.000+ 0.000

10.00020.00330.00440.00510.00130.00410.001

6130.46270.008

1350.117270.027390.03810.00010.00020.003

R6= 803Mean Titer Ref. Controls: 15 0 0150.50Positive Cutoff S/P >=

Histogram/BlockDiagram Page : Date :Report:

.

Assay :Bleeding Date : Testing Date:

Min. - Max Titer : -

%CV :.

.

.

.

.

.

.

.

Neg/Sus/Pos = / /Total No. Samples: .

Sample ID Raw O.D. ResultWell

BioChek (c)

Very low amount of false positives in DOC

Mycoplasma synoviae Day-old Chick Monitoring

37

0

2

4

6

8

10

12

14

16

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

# Samples Lot:

Mean Titer:

G.M.T.:

Titer Range Ref. Controls:


1 09/11/2007

Name : R1-IMPORTF.3-1ROSSCompany :

Code : S07-370Age : 01DType : GPHouse No. : L-1

MS FS464605/11/2007 09/11/2007

211110

R6 (700-2000)

Titer Group


A01 0.146A02 0.142A03 0.884A04 0.974A0501 0.147A0602 0.147A0703 0.150A0804 0.159A0905 0.146A1006 0.149A1107 0.151A1208 0.144B0109 0.134B0210 0.137B0311 0.142B0412 0.141B0513 0.145B0614 0.147B0715 0.147

9

NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -

- 0.000- 0.000+ 0.000+ 0.000

10.00410.00430.00890.01910.00320.00640.00910.00010.00010.00010.00010.00010.00110.00410.004

R6= 1365Mean Titer Ref. Controls: 15 0 0150.50Positive Cutoff S/P >=

Histogram/BlockDiagram Page : Date :Report:

.



%CV :.

.

.

.

.

.

.

.



BioChek (c)

Very low amount of false positives in DOC

Mycoplasma Monitoring MG Positive

0

1

2

3

4

5

6

7

8

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

# Samples

Age : 34WType : BBBirth Date : 10/10/2008House No. : 2 NOReason for Testing : PROBLEM

A01 0.144A02 0.129A03 0.672A04 0.665B0801 2.334 6B0902 1.840 5B1003 1.805 5B1104 1.753 5B1205 1.346 4C0106 1.225 4C0207 1.063 3C0308 1.023 3C0409 1.465 4C0510 1.798 5C0611 1.824 5C0712 0.861 3C0813 1.736 5C0914 1.780 5C1015 1.704 5

Mean Titer:

G.M.T.:

0

Mg CH435011/06/2009 12/06/2009

4 2802 012 6 8184 07730

Titer Group

Lot:

S/P Ratio Titer Titer Group

BioChek B.V. Service Laboratory Burg. Bracklaan 57, 2811 BP , Reeuwijk, Holland Tel: +31 182 582 592 - Fax: +31 182 599 360

POS +POS +POS +POS +POS +POS +POS +POS +POS +POS +POS +POS +POS +POS +POS +

- 0.000- 0.000+ 0.000+ 0.000

6 8184.1315 1523.2025 0353.1364 8643.0393 5342.2733 1482.0462 6371.7422 5111.6663 9192.4975 0123.1235 0993.1722 0121.3624 8083.0074 9523.0894 7012.946

0 15150.50Positive Cutoff S/P >=

VI Index: 143

189 21/09/2010Histogram/BlockDiagram Page : Date :Report:

.



%CV :.

.

.

.

.

.

.

.

.

.

/ /Total No. Samples: Neg/Sus/Pos = .

Sample ID Well Raw O.D. Result

BioChek (c)

Case�History�1�Respiratory�Infection�BRserology�test�at�42D

Mycoplasma Monitoring Mg Suspect / MS Positive

PCR (qPCR)

•  Kits commercially available – Biochek MG/MS qPCR Q2 2014

•  Excellent, rapid method •  No interference with non-pathogenic

Mycoplasma •  Sensitive – Specific •  Very low % of false positives

PCR – False Positives

•  Laboratory contamination. •  Other Mycoplasma or bacteria with

similar DNA sequences. •  Reactions may be real.

PCR – False Positives

•  Laboratory contamination. •  Other Mycoplasma or bacteria with similar

DNA sequences. •  Reactions may be real.

Role of PCR •  Confirm diagnosis in serologically positive

flocks •  Check DOC for vertical transmission •  Test spike males or other bird moves •  Rapid ID of strains: vaccines vs. field •  May reduce need to culture •  Research

Control Mycoplasma •  Biosecurity

•  Antibiotics

•  Live vaccines

•  Bacterins

Control - Biosecurity

Hatchery People Source Flock

Insects

Rodents Wild Birds

Housing

Litter

Water

Feed Equipment

Eradication

•  Serological testing and elimination. •  Strict biosecurity •  Heating eggs to 41ºC •  Antibiotic injection. •  Antibiotic dip

Mycoplasma Eradication

•  Most economical in certain scenarios – Long term consequences are far too high

•  Easily done if truly desired •  Commitment / education.

Antibiotics

Tetracyclines Macrolides Quinolones Others

Oxyletracycline Tylosin Enrofloxacin Tiamulin

Chlortetracycline Lincomycin Danofloxacin

Tetracycline Kitasamicyn Sarafloxacin

Doxycycline Josamycin

Which one works?

Control – Antibiotics Native broiler chickens

•  Day 0 – MS Ab positive •  Tylosin @ 3d for 5 days •  Tylosin @ 3 wks every 4 weeks •  Results seems to have eliminated

serological evidence by 3 & 10 weeks

National Chiayi University

Report: Histogram/Blockdiagram

Dr. Kuo Lab

ResultSample ID Well Raw OD S/P Ratio Titer Titer Group

下午

2013/10/8Ms2013/10/8

2/0/18204218

403 - 12478269193

45

RF10 (2148)Meantiter Ref.ControlsRF10 (1500-4500)Titer Range Ref.Controls

Positive Cuttoff S/P: 0.5>=

4 pos3 pos1 pos1 pos0 neg1 pos0 neg8 pos7 pos4 pos5 pos3 pos2 pos3 pos5 pos2 pos9 pos7 pos2 pos8 pos

Comments:

Control on native broiler chickens DOC

National Chiayi University

Report: Histogram/Blockdiagram

Dr. Kuo Lab

ResultSample ID Well Raw OD S/P Ratio Titer Titer Group

Reason :

Housenumber :

smallCode :

Type :

21 Day(s)Age :

2013/10/30Samplingdate :

Company :

G.P.Customer-Name :

2013/10/30 07:24:37Lab code :

Assay:

Bleedingdate:

Lotnumber:

Testdate:

02013/10/30

Ms2013/10/30

10/0/0Neg/Sus/Pos:

10Mean Titer:

Min-Max Titer:

GMT:

%CV:

501 - 1382388

No. Samples:

Target Titer:

Target %CV:

VI Index: 0Target Range VI:

Interpretation VI:

RF10 (2910)Meantiter Ref.Controls

RF10 (1500-4500)Titer Range Ref.Controls

Positive Cuttoff S/P: 0.5>=

0.584D01+

0.608C01+

0.125B01-

0.104A01-

01 F01 0.191 0.159 138 0 neg

02 G01 0.157 0.088 65 0 neg

03 H01 0.164 0.103 79 0 neg

04 A02 0.170 0.115 91 0 neg

05 B02 0.141 0.055 35 0 neg

06 C02 0.129 0.030 16 0 neg

07 D02 0.151 0.076 54 0 neg

08 E02 0.132 0.036 21 0 neg

09 F02 0.114 0.000 1 0 neg

10 G02 0.113 0.000 1 0 neg

Comments:

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Control on native broiler chickens 3wks

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Control on native broiler chickens 10wks

Control – Antibiotics

•  Still WIP – Need more data

•  We see decrease in titers following a succesful use of antibiotics

•  Delay of 4-6 weeks

Serological Profile Following Treatment



Antibiotics Use

•  Loading dose •  Maintenance

–  Every 4-8 weeks

•  Helps egg production

•  It ill not stop shedding 100%

First three days. During vaccine reactions.

Production Broilers

Vaccination

•  Commercial Egg // Multiple Age Complexes – Currently also sometimes in single age

farms

•  Live

•  Killed (Bacterins)

Killed F 6/85 ts-11 Route s/c or i/m Various Spray Eye-drop Safety +++ ± +++ +++ Persistence - +++ - ++ Antibodies ++ ++ - ± Spread - ++ - ± Displacement - +++ + +

Vaccines for Mg

Vaccines for Ms

Killed MS-H Route s/c or i/m Eye-drop Safety +++ +++ Persistence - ++ Antibodies ++ ++ Spread - + Displacement - +

Baselines Live Mycoplasma vaccines

Differentiation of Vaccination Serology vs. Field Challenge Serology based on evaluation mean flock titers with baselines and evaluating % positives. Flocks are suspect of infection when mean titers > baseline and 100% positive.

Killed Vaccines Bacterins

•  Various manufacturers.

•  Some combinations. •  MG

•  MS

Killed Vaccines Bacterins

•  Expensive

•  Two applications

•  Need to inject every chicken

•  Protects partially against production losses

•  Decreases shed considerably –  Vertical Transmission

•  Does not spread

Salmonella •  S. pullorum / S. gallinarum

– Non-motile salmonella – Ser. Gr. D

•  S. enteritidis / S. typhimurium – Motile salmonella – Ser. Gr. D / Ser. Gr. B

•  Substantial losses due to mortality, egg contamination, public health significance

64

Salmonella pullorum

65 R.Monleon

Salmonella pullorum

66

Salmonella gallinarum

67

Salmonella gallinarum

68

Salmonella enteritidis

69

Type 2 Livers

Ruptured Tendons

R.Monleon

Monitoring

•  Serology – Agglutination (WBA, RPA)

– ELISA •  Bacterial Culture

– Enrichment – RV (soy peptone broth) & MK (tetrathionate broth) / MSRV / Others

– Brilliant Green / MacConkey / XLT / Others – Biochemistry – API Strips (RAPID ID 20 E)

Salmonella Monitoring Farm

SP/SG S. Enteri*dis S. typhimurium

Day Old X X

6wks X X

16wks X X X

24wks X X

34 wks X X

44wks X X

54wks X X

* Additional sampling might be needed on circumstances

Common Group B & D Salmonella spp. Serotyping Antigens

Serovar Group “O” Soma*c “O” An*gens Flagellar “H” An*gens

Heidelberg B 1, 4, 12 r

Agona B 1, 4, 12 r, g, s

Derby B 1, 4, 12 f, g

Typhimurium B 1, 4, 12 i

Kingston B 1,4,12,27 g, s, t

Gallinarum D 1, 9, 12 -‐

Pullorum D 1, 9, 12 -‐

Enteri5dis D 1, 9, 12 g, m

Berta D 1, 9, 12 f, g, t

Panama D 1, 9, 12 l, v

As “O” antigens 1 & 12 are common in both Group B & D, any representatives of either group may cause cross reactions when high amounts of antibodies are present.

Salmonella WBA

Pullorum Agglutination Test

Salmonella P/G RPA

ELISA Specificity Results-Group B, D

Commercial Broiler Breeders

Specificity >99.8%

ELISA Specificity Results – Group D

Commercial Layers

Sensitivity BioChek SE/ST ELISA Temporal Panel with samples of infected SPF birds with S. enteritidis

0.0000

0.2000

0.4000

0.6000

0.8000

1.0000

1.2000

1.4000

1.6000

00D 07D 14D 21D 28D 35D 42D

S/P

> 0.

5 =p

ositi

ve

DAYS P.I.

BIOCHEK SE/ST. ORAL S. enteritidis INFECTION at Day 00

MEAN S/P 10 CHICKENS

AHS Deventer Salmonella validation serum Panel

Salmonella Typhimurium Monitoring Negative Flock

0

5

10

15

20

25

30

35

40

45

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

# Samples Lot:

Mean Titer:

G.M.T.:


A01 0.081A02 0.082A03 0.453A04 0.493A0501 0.088A0602 0.091A0703 0.113A0804 0.128A0905 0.098A1006 0.119A1107 0.093A1208 0.103B0109 0.108B0210 0.102B0311 0.119B0412 0.097B0513 0.095B0614 0.120B0715 0.126B0816 0.109B0917 0.102B1018 0.090B1119 0.090B1220 0.094C0121 0.096C0222 0.108C0323 0.103C0424 0.120C0525 0.086C0626 0.101C0727 0.089C0828 0.102C0929 0.103C1030 0.108C1131 0.095C1232 0.102D0133 0.108D0234 0.104D0335 0.095D0436 0.101

Name : R2F.2ROSSCompany :

Code : S07-360Age : 40WType : GPHouse No. : 01

St FS458303/11/2007 04/11/2007

5194259


Titer Group

129

- 0.000- 0.000+ 0.000+ 0.000

140.017210.024830.080

1290.119400.042

1010.096260.029540.055690.068510.052

1010.096380.040310.034

1040.0981230.114710.070510.052190.022190.022290.032350.037690.068540.055

1040.09890.011

490.050160.019510.052540.055690.068310.034510.052690.068560.057310.034490.050

NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -

45 0 0450.50Positive Cutoff S/P >=


.



%CV :.

.

.

.

.

.

.

.



BioChek (c)

Salmonella Typhimurium Monitoring Recently Contaminated Flock

0

5

10

15

20

25

30

35

40

45

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

# Samples Lot:

Mean Titer:

G.M.T.:


A01 0.084A02 0.084A03 0.565A04 0.586E0201 0.105E0302 0.154E0403 0.202E0504 0.127E0605 0.105E0706 0.228E0807 0.373 2E0908 0.115E1009 0.099E1110 0.091E1211 0.098F0112 0.099F0213 0.136F0314 0.128F0415 0.107F0516 0.113F0617 0.119F0718 0.123F0819 0.089F0920 0.115F1021 0.096F1122 0.108F1223 0.104G0124 0.108G0225 0.138G0326 0.098G0427 0.096G0528 0.107G0629 0.104G0730 0.138G0831 0.326 1G0932 0.137G1033 0.167G1134 0.096G1235 0.138H0136 0.151H0237 0.127

Name : R4F.1ROSSCompany :

Code : S07-361Age : 58WType : GPHouse No. : 02

St FS458303/11/2007 04/11/2007

142880131


Titer Group

828

- 0.000- 0.000+ 0.000+ 0.000

410.0431580.1422860.240910.087410.043

3580.2937860.588630.063280.031120.014250.028280.031

1130.106940.090450.047580.059720.071810.07980.010

630.063210.024470.049390.041470.049

1180.110250.028210.024450.047390.041

1180.1106430.4921160.1081920.169210.024

1180.1101500.136910.087

NEG -NEG -NEG -NEG -NEG -NEG -POS +NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -NEG -SUS -/+NEG -NEG -NEG -NEG -NEG -NEG -

42 1 2450.50Positive Cutoff S/P >=


.



%CV :.

.

.

.

.

.

.

.



BioChek (c)

Salmonella isolate from AA,Ross 14 - 25 October 2007

Lab code Date Farm Flock House Age(Wks.) Sock Dust Feces Remark

B07-2014 14 Oct 07 R3 2 1-2 24 2( C) - - Gr.C=S.Mbandaka

B07-2015 " R3 2 1-2 24 - 1(I),2(E) -B07-2053 21 Oct 07 KB 29 5 40 -ve - -B07-2055 " KB 29 5 40 - -ve -B07-2054 " KB 30 1-2 28 -ve - -B07-2056 " KB 30 1-2 28 - 1(B) - Gr.B=S.Stanley

B07-2062 " A1 36 4-6 52 4(B),5(G),6(E) - - Gr.B=S.Derby

B07-2063 " A1 36 4-6 52 - -ve -B07-2074 " A3 1 1-3 8 3(B) - - Gr.B=S.Stanley

B07-2075 " A3 1 1-3 8 - -ve -B07-2091 " R4 1 1-2 56 1(E) - -B07-2092 " R4 1 1-2 56 - 1(C),2(B) - Gr.C=S.Albany,Gr.B=Typhimurium

B07-2085 " R2 2 1-2 38 - - 1,2(B) H1-Gr.B=S.Stanley,H2-Gr.B=S.Typhimurium

Salmonella isolate from AA,Ross 14 - 25 October 2007

Lab code Date Farm Flock House Age(Wks.) Sock Dust Feces Remark

B07-2014 14 Oct 07 R3 2 1-2 24 2( C) - - Gr.C=S.Mbandaka

B07-2015 " R3 2 1-2 24 - 1(I),2(E) -B07-2053 21 Oct 07 KB 29 5 40 -ve - -B07-2055 " KB 29 5 40 - -ve -B07-2054 " KB 30 1-2 28 -ve - -B07-2056 " KB 30 1-2 28 - 1(B) - Gr.B=S.Stanley

B07-2062 " A1 36 4-6 52 4(B),5(G),6(E) - - Gr.B=S.Derby

B07-2063 " A1 36 4-6 52 - -ve -B07-2074 " A3 1 1-3 8 3(B) - - Gr.B=S.Stanley

B07-2075 " A3 1 1-3 8 - -ve -B07-2091 " R4 1 1-2 56 1(E) - -B07-2092 " R4 1 1-2 56 - 1(C),2(B) - Gr.C=S.Albany,Gr.B=Typhimurium

B07-2085 " R2 2 1-2 38 - - 1,2(B) H1-Gr.B=S.Stanley,H2-Gr.B=S.Typhimurium

Control

•  SP/SG –  HOST SPECIFIC – REQUIRES CHICKENS

•  SE/ST – Wide Range of hosts – Can come to the farm by multiple ways

•  Biosecurity is the most effective means to control disease

•  BESTEST – CULL PARENT FLOCK •  Stop Vertical Transmission

–  Do not Use Infected Parent Stock

Salmonella Control

•  Biosecurity •  Vaccination

– Live Vaccines •  SG •  SE / ST

– Killed Vaccines •  ST / SE / Others

•  Treatment

Control - Biosecurity

Hatchery People Source Flock

Insects

Rodents Wild Birds

Housing

Litter

Water

Feed Equipment

Control Biosecurity

•  Chicks must be obtained from flocks free of Salmonellas •  Salmonella Free flocks should not be mixed with other

birds •  C+D of premises should be done stringently – Houses

must be easy to be cleaned •  Use pelletized feed (thermal treatment) / Raw materials

free of Salmonella •  Avoid introduction of salmonellas by:

–  Poultry houses should be bird proof –  Houses must be vermin (Rats, mice, rabbits, cats, dogs) proof –  Insect control (flies, poultry mites, and the lesser mealworm) as

they may provide a means of survival in the environment

Control Biosecurity

•  Water should be sanitized –  (i.e. chlorinated water)

•  Control mechanical carriers –  footwear and clothing of humans, as well as

poultry equipment, processing trucks, and poultry crates

•  Effective disposal of dead birds is a must as they can be sources of infection to other birds

Control

•  Treatment – Not recommended, not 100% effective

•  Carriers - Shedding

– Enrofloxacin, Furazolidone, Sulfas, others – Treatment of eggs possile – After antibiotics C.E. possible

Control

•  Vaccination – SG9R (S. gallinarum)

•  Relative value •  6w / 14-16wk •  Safety / Potency

–  SE / ST •  Multiple applications (DOC + x2) •  Early protection •  Safe

– Bacterins

Live Vaccines •  Attenuated by modifications in genes coding for

metabolic/virulence functions •  Administration

–  Coarse spray or drinking water (most, SG 9R exception) –  Guidelines: day of age, 4-6 wks and 10-12 wks

•  Manufacturers/Products (most common)

Inactivated Vaccines •  Utilized in layers, broiler breeders, and

turkey breeders –  Approx. 90% shell egg producers

utilizing –  Layers utilize in combination with NDV

and IBV •  Manufacturers: Ceva, Lohman, Pfizer,

Merial (not US); Merck (not US) –  Gallimune – Merial –  Immunovac – BioVet Poland –  Nobilis Salenvac - Merck –  Poulvac SE - Pfizer

Baselines Test Vaccine Type Mean Titer Range

Wks aKer Vaccina*on to Test

D Inac5vated (2X) 3000-‐10000 4-‐6 wks a[er 2nd

Salenvac T, Gallimune Se+St

1000-‐5000 10-‐12 wks a[er 2nd

1X SE4 (preliminary) 1500-‐4000 4-‐6 wks a[er inact.

B&D Inac5vated (2X) 3000-‐12000 4-‐6 wks a[er 2nd

Salenvac T, Gallimune Se+St

1000-‐5000 10-‐12 wks a[er 2nd

1x SE4 (preliminary) 2500-‐5000 4-‐6 wk a[er inact.

Salmonella Serology and Vaccination – experiences to date and expected results

Vaccines Application

ELISA KIT

SE/ST SE ST

SE inac + + -

SE+ST inac + + +

SG live + + -

SE live Oral +/- +/- -

+ + -

ST live Oral +/- +/- -

+ - +

EXAMPLE OF EXPECTED RESULTS

Se/St Results: Periodic Monitoring of Vaccinated Layers

Vaccinated with live S. typhimurium vaccine at 2 & 20 d, and inactivated S. enteritidis vaccine at 16 wk of age

SE/ST ELISA GMT Results Broiler breeders vaccinated with an autogenous salmonella vaccine

A- well vaccinated complex; B-poorly vaccinated complex

0

1000

2000

3000

4000

5000

6000

7000

A A A A B B B

GM

T

10W 30W 48W

Se/St ELISA Results Broiler Breeders – comparison of non-vaccinated with vaccinated (autogenous S. typhimurium, S. kentucky, and S. enteritidis at 10-12 wk,

and 17-18 wk SQ)

Typical Vaccination Curve 2x Inactivation: Peak response 4-6 wks post 2nd vaccination 100% positive

End of production 50- 85% positive

0

1000

2000

3000

4000

5000

6000

7000

19 24 31 38 46 61

ME

AN

EL

ISA

TIT

ER

AGE IN WEEKS

SE/ST: BROILER BREEDERS VACCINATED WITH INACTIVATED SE+ST VACINE

at 10W and 15W

Group B/D ELISA Results – Commercial Layers

Flock Age Mean Titer

GMT % CV % Pos

VacA (0.25 ml)

17 wk 8300 7879 27 100

Vac (0.5 ml) 17 wk 7937 7710 24 100

Control 17 wk 67 53 84 0 A Vaccinated flocks received an inactivated Se vaccine at 13 wks of age. Data provided by AviServe

Vaccination Programs •  Layers

– SE bacterin •  Usually 1x at 13 – 15 wk in layers

–  Live ST vaccines •  3 applications-2, 6, and 12 weeks •  Bacterin + Live vaccine

–  Live vaccine – 2 & 6 weeks –  Bacterin – 13-15 weeks

– Vaccine costs •  0.5 cents for live •  8 cents for bacterin (5 cents for handling and 2.5 cents

for vaccine)

Factors to Consider in Interpreting Serological Results

•  Use to identify infected flocks and not individual birds

•  Unvaccinated positive flocks may no longer be infected or excreting

•  Actively excreting flocks may be negative serologically

•  Chickens may acquire anti-Salmonella antibodies from parents via the yolk sac

•  Many live vaccines given orally do not provoke a significant antibody response

Commercial Layer Breeders (LB) vaccinated with 2 live ST vaccines at D7 and D28, and inactivated SE4 bacterin at 12W, and of 1D progeny (PR) derived from those flocks.

Samples provided by LAHI

Important Poultry Vertically Transmitted Viral Diseases

•  Chicken Anemia Virus •  Avian Encephalomyelitis •  Fowl Adenovirus •  Chicken Astrovirus

•  Lymphoid Leukosis (Not Covered Today) •  Newcastle Disease (Not Covered Today) •  ReoVirus (Not Covered Today)

Dynamics of VT Viral Infections Scenario I

16 W 40 W 25 W 8 W

Viral Infection

Point of Lay

NO SHEDDING OF ACTIVE VIRUS TO PROGENY

%

Positives (seroconversion)

Dynamics of VT Viral Infections Scenario II

16 W 40 W 25 W

%

Positives (seroconversion)

8 W

Viral

Infection

Point of Lay

SHEDDING OF ACTIVE VIRUS TO PROGENY FOR A PERIOD

OF ~4 – 8 WEEKS

Chicken Anemia Virus •  Discovered in late 70s •  CAA / CAV

– Circoviridae - Avian Gyrovirus 1 •  Induces vertical transmission in non-

seroconverted flocks •  Lesions on the bone marrow, thymus, etc •  Potent Immunosupressive Disease

– Blue-wing disease (Clostridium)

104

Post Mortem Findings

•  Pale organs

•  Pale fatty bone marrow (yellow or pink) •  Anaemic condition > low PCV (heamatogrit) scores ≤ 27

•  Thymus atrophy / Bursal atrophy

•  Watery blood

•  Subcutaneous and intramuscular hemorrhages

•  Proventricular hemorrhages

•  Skin lesions are prone to secondary bacterial infection leading to gangrenous dermatitis > blue-wing disease


Anorectic, lethargic, depressed, and pale

Gangrenous dermatitis


Atrophy of Thymus

Pale fatty bone marrow


Pale fatty bone marrow and bursal atrophy


Subcutaneous and intramuscular hemorrhages, gangrenous dermatitis


Anaemic condition indicated by low PCV (heamatogrit) scores ≤ 27

Post Mortem Findings Blood results of an field infected broiler flock,

indicating anemic condition of birds by low PCV (hematocrit) scores ≤ 27

Monitoring

• Histopathology

• Serology – VN, ELISA

• PCR – Thymus / Bone Marrow

Virus Neutralization •  VN is “gold standard”

•  Interpretation VN: § Log 2 VN < 5 negative result

§ Log 2 VN titers of ≥ 5 positive result

•  Log 2 VN titers of ≥ 8 prevent CAV infection / re-isolation

•  Log 2 VN titers of ≥ 11 are protective against clinical CAV

•  High MAB titers protect against clinical disease in broilers for 2-3 weeks

•  Once positive, naturally infected adult birds will seroconvert to protective VN ≥ 8 titers in 4-6 weeks post infection

CAV VN Serology •  Once positive, naturally infected adult birds will seroconvert

to protective VN ≥ 8 titers in 4-6 weeks post infection

Data courtesy of MSD, Boxmeer, Holland

4.0

5.0

6.0

7.0

8.0

9.0

10.0

11.0

12.0

14 21 31 38 49 55

Mea

n lo

g2 C

AV

VN

ant

ibod

y ti

tre

Weeks of age

Protective log 2 VN Titer

ELISA

• Accurate and inexpensive

• Easy interpretation

• Possibility of test a large number of samples and flocks in short periods of time

• Correlation of ELISA vs. VN is critical

ELISA (BioChek) –  Detects antibodies against CAV in serum of chickens –  Indirect ELISA, dilution 1:500 –  Specificity > 98 % –  Sensitivity: will give positive results 10-20 days p.i –  Filter 405 nm –  Positive Cuttof S/P ≥ 0.35 –  Log 10 titer = 1.1 x log (s/p) + 3.361

S/P Titer Interpretation ≤ 0.349 ≤ 724 NEGATIVE ≥ 0.350 ≥ 725 POSITIVE

ELISA (BioChek) vs. VN

– Biochek ELISA correlates well with VN –  Interpretation of results when compared to VN

Positive BC Titers > 2296 correspond to protective VN titers ≥ 11

Positive BC Titers > 724-2295 correspond to VN titers ≥ 8-10

CAV ELISA KITS ON MARKET

BIOCHEK Comp. I Comp. S

DILUTION 1:500 1:10 detection 1:100

1:100 vaccine response

Interpretation 3000-8000 80-100% (positive)

Complicated 3000-5000 CV <50%

Application of CAV ELISA •  SPF Screening for Negative CAV status

•  Vaccination Monitoring §  Confirm success of vaccination prior to lay and during Lay.

•  Non-vaccinated breeders: §  Check at 12-16W to determine need of vaccination.

-  Criteria: vaccinate when birds have < 80% positive titers

•  Disease Monitoring BR > 35D of age

PCR

Control

• Biosecurity

•  Induced Exposure

• Vaccination

Biosecurity

•  Virus Ubiquitous

•  Cleaning and Disinfection + Downtime

– Beware “Clean / New House Syndrome”

– Some benefit with formaldehyde

•  Prevent Immunosuppressive Disease § IBD, MDV, REV, Others

Induced Exposure

•  Seroconversion § Once neutralizing antibodies are present shedding

generally stops

•  Natural Exposure – Litter, Faeces § Beware what you bring in (i.e. Mycoplasma / Salmo)

•  Liver Homogenates

Promoting Sero-conversion

124

Vaccination CAV

• Live Vaccines

§ Circomune, CAV-Vac, P4, Thymovac • WW / Injection / Drinking Water

§ Rate of spread minimize ->

Missed bird might be negative

Vaccination CAV

•  Initial vaccination between 8 and 12 weeks

§ Monitoring (14-16 weeks) > Seroconversion

§ Revaccination if needed

§ High uniform levels of antibody to CAV are detected from 4 weeks post vaccination onwards and are maintained throughout the laying period

BioChek CAV ELISA Vaccination Monitoring

•  Objective: Confirm vaccination success

•  Test 4 - 6 weeks after vaccination

•  Vaccinate at age ( 8-12 W) so that confirmation testing and revaccination can be done

•  Test at an age (14-16W) that revaccination is possible (revaccination during lay not possible)

Vaccination and Serological Confirmation

Criteria for successful vaccination §  80- 100 % of birds must have positive

protective titers > 2000 at least 4 weeks prior to onset of lay

§  When < 80% positive protective titers revaccinate immediately

CAV Vaccination Baselines BioChek

BIOCHEK VACCINATION BASELINES LAYERS/BREEDERS (Continued) Titer values may vary according to age & type of bird , vaccine type, vaccination program, and other factors such as placement programs. You may find different results under different circumstances. TEST VACCINE MEAN TITER WKS AFTER VAC. SUSPECT TITER TYPE RANGE TO TEST % POS VI Index INFECTION

MS live MS-H (eye drop) 500 - 3 000 6 -12 wks 30- 70% Pos > 5000 and > 90% AI Inact 2x H5N2 1 000 - 4 000 6 – 10 wks 100% Pos Inact 2x H9N2 2 000 - 6 000 6 – 10 wks 100% Pos EDS inact. 1x 1 000 - 4 000 4 – 6 wks SE live 3x DW (Salmonella Vac E) 100 - 500 5 - 6 wks < 15% Pos (Salm D)

inact. 2x 3 000 - 10 000 4 - 6 wks after 2nd 90-100% Pos 50 - 500

Salenvac T, Gallimune Se +St Talovac 109 SE, Poulvac SE, 1 000 - 5 000 10 -12 wks after 2nd 50-100% Pos 10 - 100 Layermune SE, Avipro SE4

SE/ST live 3x DW (Salmonella Vac E+T) 500 - 1 500 5 - 6 wks < 70% Pos (Salm B&D)

inact. 2x 3 000 - 12 000 4 - 6 wks after 2nd 90-100% Pos 50 - 500 Salenvac T, Gallimune Se +St Talovac 109 SE, Poulvac SE, 1000 - 6 000 10 -12 wks after 2nd 50-100% Pos 10 - 100 Layermune SE, Avipro SE4 CAV live (Tymovac, PG4, CAV-Vac, 3 000 - 8 000 4 - 6 wks 80-100% Pos 100 - 300 Circomune)

* ORT: Titers > 10 000 often correlate with clinical disease These guidelines are based on our experience and information from our clients. BioChek does not accept any responsibility for the results using these guidelines.

CAV: Case History Serological profile of vaccinated BB flock

BB vaccinated at 15W (IM) with live Nobilis CAV P4 CAV vaccine

Vaccinated flocks have persistant high and uniform protective titers


98% Seroconversion 4 W post vac.


98% Seroconversion 9W post vac.


100% Seroconversion 16W post vac.

CAV: Case History

Monitoring of immune status of non-vaccinated BB flocks at 13W

Poor immune status 36% POS Poor uniformity Vaccination needed

Good immune status 100% POS Good Uniformity No vaccination necessary

CAV: MAB transfer Parent to Progeny 01D

AGE W Parent 01D ChickMA Transfer Rate

49 5188 3254 63%

49 5013 2570 51%

Average 57%

MEAN BIOCHEK CAV ELISA TITER (n=20)

CAV: MAB Transfer Parent – Chick 01D

Parent 49W 100% POS 40 wks post vac

Chick 01D 100% POS 51% transfer Parent to Chick

100% POS

100% POS

MDA Negative Broilers: Field Case

Intervet

MDA Negative Broilers: Field Case

Intervet

Vaccination CAV Broilers

•  Seroconversion of parents induce transfer of antibodies (MAB)

•  Progeny with high level MAB is protected against clinical CAV

•  High MAB titers protect against clinical disease in broilers for 2-3 weeks

•  UNIFORMITY of titers is KEY for PROTECTION

CAV BioChek Serology Broilers

• Limited data available

• Most healthy broiler flocks will test negative to low positive (MT < 2000) at > 35D

• Clinical flocks have MT > 5000

• Serology only meaningful when comparing healthy non-clinical birds with clinical birds

CAV BioChek Serology Broilers

Non-Clinical BR 42D

Clinical CAV BR 42D

CAV Vaccination Baselines BioChek

Avian Encephalomyelitis •  Old Disease; Hard to See in the Field

– Vaccine (Vaccination) failures •  Picornavirus •  Epidemic Tremor •  Drops in egg production in affected

parents •  Up to 100% mortality in affected vertically

transmitted chicks

143

Avian Encephalomyelitis

144

Karki

AE: Clinical Signs Adult Birds Egg and hatchability drop during 2-5 wks of production due to AE infection of BB

Avian Encephalomyelitis

146 Hong, 2012

Hatchability Loss (AE)

　 9/12 9/15 9/19 9/22 9/26 9/29 Diff

(Candling – Setting) 0 3.9 2.9 3.3 2.3 4.1

Hatchability after setting and candling

0.020.040.060.080.0

100.0

9/12 9/15 9/19 9/22 9/26 9/29Date

Hat

chbi

lity(

%)

-10.00.010.020.030.040.0 Hatchability

from the settingHatchabilityafter candlingDiff(Setting)

Diff(Candling)

Hong, 2012

ELISA found AE –ve at 33 weeks old Farm manager forgot to vaccinate Vaccination carried out at 34 weeks old (Calnek 1143 strain)

Sample Titer 1 120 2 115 3 209 4 115 5 279 6 307 7 151 8 188 9 318 10 220 11 241 12 198 13 258 14 374 15 105 양성율 0/15 평균 213

BioChek kit, cutoff 1071

Losses in Broiler

Hong, 2012

Date Customer No of chicks from affected B

No of total chicks

No of dead bird

Mortality (%)

12 J 29,500 29,500 21,500 73 12 P 11,700 30,000 3,454 30 15 K 10,600 29,000 10,600 100 15 P 13,600 36,500 2,283 17 19 J 27,000 27,000 20,000 74 19 K 2,800 8,000 1,400 50 19 P 7,400 38,000 5,206 70 22 M 20,800 32,000 20,800 100 26 J 6,000 13,000 4,000 67 26 J 4,200 17,000 2,000 48

Total 133,600 260,000 91,243 68(Ave)

Diagnosis Histology and serology

•  Clinical signs if present •  Histology : microscopic lesions in gut and brain

tissues and brain •  Non vaccinated birds: positive serology •  Vaccinated birds: detection of abnormal serology

through Monitoring

AE ELISA (BioChek)

–  Indirect ELISA, dilution 1:500 – Specificity > 98 % – Sensitivity 10-14 days post

infection – Filter 405 nm – Positive Cuttof S/P ≥ 0.5 – Log 10 titer = 1.1 x log (s/p) +

3.361

S/P Titer Interpretation ≤ 0.499 ≤ 1070 NEGATIVE ≥ 0.5 ≥ 1071 POSITIVE

AE and Immunity §  Humoral immunity but not cellular immunity is the

most important factor for protection §  Immunized Adults with positive serology are

effectively protected from egg drops and vertical transmission.

§  Young birds with maternal immunity are effectively protected against clinical AE.

§  Vaccination goal is to prevent AE outbreak by making sure 100% of birds are positive prior to entering production

AE Prevention Vaccination

Breeders §  Vaccination with live or inactivated vaccine §  Live vaccination at least 4 weeks prior to onset of

lay §  Inactivated vaccination possibly during

production period §  Lifetime immunity is acquired through vaccination Young chicks §  protection through maternal antibodies

AE Prevention Vaccination and Serological Confirmation

§  Live vaccines application through drinking water or wing web method

§  Wing-web more prone to misapplication §  Most AE outbreaks of vaccinated flocks due to poor

vaccine handling and/or vaccine application. §  Vaccination must give at least 80% or more positives

before lay to provide effective 100% immunity during lay

§  Application methods never give 100% delivery, monitoring of success of vaccination with ELISA is recommended

AE Prevention Vaccination and Serological Confirmation

§  Serological test to confirm vaccine take §  Test 4 - 6 weeks after vaccination §  Vaccinate at age ( 8-12 W) so that confirmation

testing and revaccination can be done §  Test at an age (14-16W) that revaccination is

possible §  Inactivated vaccination possibly during production

period

Prevention Vaccination and Serological Confirmation

Criteria for successful vaccination •  80% of birds must have positive titers at

least 4 weeks prior to onset of lay •  When < 80% positive titers revaccinate

immediately •  1x confirmation testing and taking action

on results can prevent AE infection ever from occurring !

AE Prevention Vaccination Baselines BioChek

50 - 500

VI Index

AE: Case History Serological profile of a natural infection in BB

24W 0% POS

28W 64% POS

31W 73% POS

AE: Case History Serological profile of vaccinated BB flock

BB vaccinated at 14W with live AE vaccine through drinking water

Vaccination monitoring and revaccination could have prevented this AE outbreak !

No vaccination monitoring at 16W

Example Natural Infection

0

1

2

3

4

5

6

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

# Samples

0

1

2

3

4

5

6

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

# Samples

0

2

4

6

8

10

0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18

# Samples

AE 2 03824-08-2010 1 58430-09-2010 5715GREECECEVA

MONITORINGD50

BB 31WTiter Group

AE 1 44906-08-2010 1 15730-09-2010 5814GREECECEVA

MONITORINGD50

BB 28WTiter Group

AE 34809-07-2010 22330-09-2010 8710GREECECEVA

MONITORINGD50

BB 24WTiter Group

G.M.T.:

G.M.T.:

G.M.T.:

BioChek B.V. Service Laboratory Burg. Bracklaan 57, 2811 BP , Reeuwijk, Holland Tel: +31 182 582 592 - Fax: +31 182 599 360

24-08-20121 Date :Page :

Assay : Mean Titer Bleeding Date

%CV :Testing Date:#Samples :Name:Company:Code:Reason:

House:Complex:Type: Age:

.




.




.

BioChek (c)

24 WKS

28 WKS

31 WKS

Fowl Adenovirus •  Inclusion Body Hepatitis / Gizzard Erosion •  Hydropericardium Syndrome •  12 serotypes / 5 species •  Generally no drop in egg production

– However some reports mortality and drops •  Vertically infected chicks develop disease

in first 3 weeks •  Severe morbidity / Mortality

162

Fowl Adenovirus (FAdV)

308 Production Производство к.20

0.00

10.00

20.00

30.00

40.00

50.00

60.00

70.00

80.00

90.00

22 27 32 37 42 47 52 57 62

Weeks of Age

% P

rodu

ctio

n

0

50

100

150

200

250

300hw%

hh%

hh std

cum fert

норма вывода,Std H %

hh std СПК

норма инк.яйц,CV

Средний вывод

АВИАГЕН

He HH

Px feed

He Std СПК

Klausz, 2009

Weekly hen mortality %

0.00

5.00

10.00

15.00

20.00

25.00

23 24 25 26 27 28 29 30 31 32 33 34 35 36 37Age / weeks

% m

orta

lity

Weekly hen mortality %

Klausz, 2009

Hen mortality

0

20

40

60

80

100

120

140

160

129

133

137

141

145

149

153

157

161

165

169

173

177

181

185

189

193

197

201

205

209

213

217

221

225

229

233

237

241

245

249

253

257

261

Age, days

mor

talit

y, b

irds

Klausz, 2009

0

1

2

3

4

5

6

7

8

9

10

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17อายุไก่(วัน)

% M

orta

lity

B9A7A8A2A3A4A5

FAdV Offspring Mortality – Vertical

Transmission

AGE

Chicken Astrovirus •  New Emerging Disease •  Worldwide Distribution •  Chick Nephropathy

–  “Gout” •  White Chicks

– Green Livers •  Astrovirus suspected

– Affected chicks have same source flocks

168

CAstV-B •  Chicken Astrovirus

– Subgroup B •  Middle East / India •  Visceral gout in very young chicks aged 4

– 7 days •  5-15% mortality •  Flocks coming from same source in

multiple occasions

169

Chicken Astrovirus

170 Klausz, 2012

Chicken Astrovirus

171 Bulbule, 2013

Chicken Astrovirus

172 Bulbule, 2013

not shown). Water deprivation, nutritional factors and othermanagement errors were ruled out as being the causes ofthis outbreak of gout. The kidney samples from thisparticular flock were positive for CAstV and negative forIBV, ANV, chicken anaemia virus (CAV) and IBDV, asdetected by qRT-PCR and qPCR and confirmed by nucle-otide sequencing (data not shown) .We then collected 894 kidney samples from gout-affected

chicks from different regions of India and tested them forthe presence of CAstV, ANV, IBV, IBDV and CAV. It wasobserved that 373 (41.7%) samples were positive for CAstValone, while 326 (36.4%) were positive for both CAstV andANV and 18 (2.0%) were positive for CAstV, ANV and

IBV (Table 3). Overall 717 (80.2%) samples were positivefor CAstV. All samples were negative for IBDV and CAV.

Virus isolation. Kidney samples that were only positive forCAstV were randomly selected for virus isolation andmolecular characterization. Eighteen samples were inocu-lated into SPF embryonated chicken eggs via the allantoicroute. At 5 days p.i., all samples caused stunting of embryoswith yellowish discolouration, necrosis of the livers, and paleand swollen kidneys (Figure 2). A significant difference inweight was observed between infected and control embryos.The average weight for embryonated eggs that had beeninoculated with the 18 CAstV isolates was 11.25 ± 1.0 g

Figure 1. Gross and histopathological lesions from a gout-affected commercial broiler chick. 1a: Prominent ureter and urate deposition inthe kidney. 1b: Urate deposition on the heart. 1c: Interstitial nephritis and urate deposits in the kidney (arrows). 1d: Infiltration ofinflammatory cells in the myocardium (arrow). 1c and 1d: bar =100 µm, haematoxylin and eosin staining.

Table 3. Quantitative RT-PCR and qPCR detection of CAstV, ANV and IBV in kidney samples collected from commercial broiler flockssuffering from gout.a

Incidence of the following pathogens in the samples testedb

Year ofcollection

Region ofIndia CAstV ANV IBV

CAstV + ANV combinedinfection

CAstV + ANV + IBV combinedinfection

2011 Northern 55/154 (35.7) 0/154 (0) 0/154 (0) 65/154 (42.2) 3/154 (1.9)Southern 77/167 (46.1) 0/167 (0) 0/167 (0) 61/167 (36.5) 1/167 (0.6)Western 55/185 (29.7) 0/185 (0) 0/185 (0) 83/185 (44.9) 4/185 (2.2)Eastern 10/10 (100) 0/10 (0) 0/10 (0) 0/10 (0) 0/10 (0)

2012 Northern 42/90 (46.6) 0/90 (0) 0/90 (0) 27/90 (30) 4/90 (4.4)Southern 71/140 (50.7) 0/140 (0) 0/140 (0) 46/140 (32.8) 2/140 (1.4)Western 46/128 (35.9) 0/128 (0) 0/128 (0) 44/128 (34.3) 4/128 (3.1)Eastern 17/20 (85) 0/20 (0) 0/20 (0) 0/20 (0) 0/20 (0)Totalc 373 (41.7) 0 (0) 0 (0) 326 (36.4) 18 (2.0)

aAll samples were negative for IBDV and CAV. bNumber of positive samples/number of samples tested (% positive). cNumber (%) positive of 894samples examined.

Astrovirus-induced gout in broilers in India 467

Dow

nloa

ded

by [D

r Raf

ael M

onle

on] a

t 13:

45 2

7 N

ovem

ber 2

013

White Chick Syndrome •  Astrovirus related disease

– Todd, 2013 •  Sporadic Worldwide Distribution •  Drops in egg production, hatchability •  Severe lesions in baby chicks

173

Chicken Astrovirus White Chick Syndrome

174


175 Soares, 2012


176

White chick syndrome 30-34 week flock October 2011

0.0

10.0

20.0

30.0

40.0

50.0

60.0

70.0

80.0

90.0

100.09/

12/1

1

9/14

/11

9/16

/11

9/18

/11

9/20

/11

9/22

/11

9/24

/11

9/26

/11

9/28

/11

9/30

/11

10/0

2/11

10/0

4/11

10/0

6/11

10/0

8/11

10/1

0/11

10/1

2/11

10/1

4/11

10/1

6/11

perc

ent Pen1 eggs

Pen2 eggshatch %

Soares, 2012

Chicken Astrovirus White Chick Syndrome( Green Livers)

177 Soares, 2012

Chicken Astrovirus White Chick Syndrome( Green Livers)

178 Undisclosed, 2010

Compare Normal chick (Left side) and affected chick.

Chicken Astrovirus White Chick Syndrome (Green Livers)

179 Soares, 2012

Chicken Astrovirus Green Livers

180 Todd, 2013

•  IFA

•  VN

•  ELISA – Commercially Available from Biochek Soon

•  PCR

Monitoring

•  Vertically transmitted diseases represent a great threat to any poultry operation due to its economic significance

•  Monitoring of VT diseases allows to take preventive measures that improve productivity and generates return of invest.

•  Among others ELISA is one of the tools of choice for its relatively low cost / performance ratio

Summary

THANK YOU [email protected]

Health & Medicine

Monitor and Control of Vertically Transmitted Poultry Diseases