-
Pink orangey
colour
Opaque halo : Phospholipase activity
BACILLUS CEREUS RAPID AGAR (BACARA) Selective medium for
presumptive Bacillus of cereus group
For In Vitro use only To store between 2 and 8C
Page 1/2
PRINCIPLE Bacillus cereus is responsible for food-borne
outbreaks. It produces thermoresistant spores that make it
particularly adapted to foodstuffs submitted to a thermal
treatment. Some strains of B. cereus can grow at refrigeration
temperature, which is an emerging risk for ready-to-use products.
They represent by themselves 5% of the collective food poisoning in
France and are also involved in a lot of opportunistic infections
on in-patients. Bacillus cereus belongs to the Bacillus cereus
group within which can be found B. thuringiensis, B.
weihenstephanensis, B. mycoides, B. pseudo-mycoides et B.
anthracis. Except the last-mentioned, the phenotypic
differentiation between species of Bacillus cereus is impossible
with actual culture methods. BACARA agar is a selective chromogenic
medium that allows the enumeration of Bacillus of the cereus group
without confirmation. On this medium, typical colonies of B. cereus
show a pink / orangey colour due to the metabolism of the chromogen
substrate and are surrounded with an opaque halo due to the
phospholipase activity. The selectivity of BACARA agar has been
especially optimized to prevent growth of interfering flora and
thus to allow an easy interpretation of plates even when matrix
highly contaminated with competitive flora are analysed. FORMULA In
grams per litre of purified water Special mix of peptones 10.00
Yeast extract 4.00 Sodium chloride 4.00 Phosphate buffer 10.00 Agar
18.00 Mix of antibiotics 0.26 Chromogen substrate 0.05
Phospholipids Final pH : 7.2 + 0.2 at 25C
PREPARATION For the medium in flask, liquefy the medium in a
water-bath at 100C and cool to 44-47C. Per flask of 100 ml of
BACARA medium, add aseptically 4 ml of BACARA enrichment supplement
and 0.5 ml of BACARA selective supplement (chromogenic selective
mix). Homogeneize well the agar base and the supplements, avoiding
the incorporation of air bubbles. PROCEDURE (See also Annex I)
Surface method : After preparation and homogeneization of the
primary solution in the suitable diluent, spread 0.1ml of the
solution obtained onto the surface of a 90mm BACARA plate (to
increase the accuracy, spread 1ml onto a 140mm BACARA plate or
split onto 3 90mm BACARA plates). If necessary, repeat the step
with the decimal dilutions of the primary solution. Incubate the
BACARA plates for 24 2h at 30C 1C. Pour-plate method : After
preparation and homogeneization of the primary solution in the
suitable diluent, inoculate 1ml of the solution obtained in a
sterile Petri plate. Pour about 18 ml of BACARA medium. Homogeneize
well and let it solidify. If necessary, repeat the step with the
decimal dilutions of the primary solution. Incubate the BACARA
plates for 24 2h at 30C 1C. RESULTS After incubation, Bacillus from
cereus group grow as large pink/orangey colonies surrounded with an
opaque halo. Refer to the standard NF EN ISO 7218 for calculation
and expression of results. AES CHEMUNEX Rue Maryse Basti Ker Lann
CS 17219 35172 Bruz Cedex
Tl. 02 23 50 12 12 Fax. 02 23 50 12 00
http://www.aeschemunex.com - [email protected]
AES 10/10-07/10
BACARA Method
ALTERNATIVE METHODS FOR AGRIBUSINESS ANALYSIS
Certified by AFNOR Certification
www. afnor-validation.org
For human food and animal feeding stuffs. End of validity :
2014/07/02
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BACILLUS CEREUS RAPID AGAR (BACARA) Selective medium for
presumptive Bacillus of cereus group
For In Vitro use only Page 2/2
LIMITS AND PRECAUTIONS Comply with Good Laboratory Practice
(refer to the standard NF EN ISO 7218). Thanks to the high
specificity and selectivity of BACARA, it is not necessary to carry
out confirmation tests on typical colonies. All the strains
belonging to the Bacillus cereus group will give characteristic
colonies. Some bacteria can also grow as orangey coloured colonies
on BACARA but without expression of the phospholipase activity. The
absence of the opaque halo will make them easily distinguishable
from the Bacillus cereus. After opening, the BACARA enrichment
supplement can be used up to its expiry date if it was manipulated
according to good laboratory practice (ie: Use in aseptic
conditions and storage between 2 and 8C). After opening, the BACARA
selective supplement can be used up to 15 days maximum if it was
manipulated according to good laboratory practice. Warning: DO NOT
FLAME THE TUBE before closing it. This supplement contains a
flammable solvent. Supplements have to be stored away from light.
The supplemented agar base has to be used immediately or poured
into sterile Petri plates. The plates poured from flasks can be
kept up to 2 weeks at 2-8C in a hermetically sealed package. For
the modes of storage, refer to the standard NF EN ISO 7218. BACARA
plates can be placed in the refrigerator after incubation up to 48
hours. The halo and the colour of the colonies will not be altered
at those low temperatures. BIBLIOGRAPHY 1- AFNOR NF EN ISO 7932
Juillet 2005. Microbiologie des
aliments Mthode horizontale pour le dnombrement de Bacillus
cereus prsomptifs. Technique par comptage des colonies 30C.
2- N.A LOGAN, P.C.B. TURNBULL : Actualit permanente en
Bactriologie Clinique Bactries arobies sporules
3- F.A DROBNIEWSKI : Bacillus cereus and related species Clin
Microbiol Rev 1993 October Species ; 6 (4): 324-338.
4- NF EN ISO 7218. : Microbiologie des aliments - Exigences
gnrales et recommandations.
PACKAGING : Ready to use base medium To store between 2 and 8C
AEB620106 : Kit BACARA base + supplements for 5x100 ml Pre-poured
complete medium To store between 2 and 8C AEB520100 : Pack of 20
plates of 90 mm AEB120102: Pack of 10 plates of 140 mm Manufactured
by AES CHEMUNEX - Combourg France 520100 : 02/07/10- B AES CHEMUNEX
Rue Maryse Basti Ker Lann CS 17219 35172 Bruz Cedex
Tl. 02 23 50 12 12 Fax. 02 23 50 12 00
http://www.aeschemunex.com - [email protected]
-
AES CHEMUNEX Rue Maryse Basti Ker Lann CS17219 35172 Bruz cedex
Tel : 02 23 50 12 12 fax : 02 23 50 12 00 -
http://www.aeschemunex.com [email protected]
AES 10/10-07/10
ALTERNATIVE METHODS FOR AGRIBUSINESS ANALYSIS
Certified by AFNOR Certification
www. afnor-validation.org
Culture media for food industry
Annex I Enumeration of presumptive Bacillus cereus
BACARA METHOD
Prepare and dilute the sample according to the specifications of
NF EN ISO 6887, NF EN ISO 8261 standards and the specific standard
of the product.
Spreading of 0.1 ml* onto 1 90 mm plate of BACARA medium
(AEB520100 Pack of 20 plates 90 mm) previously dried in an
incubator. To increase the accuracy, 1 ml* onto 1 140 mm plate
(AEB120102 Pack of 10 plates 140 mm) or 1 ml* onto 3 90 mm plates.
Incubation 24 2 hours at 30 1C
SURFACE METHOD POUR-PLATE METHOD
Inoculate 1 ml* in 1 sterile Petri plate, pour
about 18 ml of BACARA medium (AEB620106 Kit base + supplements
for
5x100ml) cooled at 44-47C.
Incubation 24 2 hours at 30 1C
520100 : 02/07/10 B
Enumerate the plates containing less than 150 characteristic
colonies.
Characteristic colonies of presumptive Bacillus of cereus
group : large pink / orangey surrounded with an opaque halo.
* : repeat the step with the decimal dilutions of the primary
solution if necessary.
