SESAME Salmonella TEST

SESAME Salmonella TEST
INTENDED USE
SESAME Salmonella TEST represents an alternative method for the detection of Salmonella in
human and animal products, as well as environmental samples (with the exception of those from
animal production). The analyses can be declared negative in 48 hours after only two steps of preenrichment (SESAME Salmonella Enrichment) and differentiation (SESAME Salmonella
Detection). The confirmation of presumptive positive samples is achieved using COMPASS
Salmonella Agar, which requires an additional 24 hours incubation.
SESAME Salmonella TEST is officially certified by AFNOR Certification, under the reference
number BKR 23/04-12/07, of which the validity runs until December 4th, 2011.
PRINCIPLES
- Pre-enrichment is performed by inoculating into SESAME Salmonella Enrichment and
incubating for (18 ± 2) hours. This liquid medium allows a proper osmotic balance and its
buffered formula enables an optimal resuscitation of Salmonella strains. The SESAME
Salmonella Enrichment formula is compliant with Buffered Peptone Water.
- The differentiation or detection stage is possible through the inoculation of the enrichment
media
into
SESAME
Salmonella
Detection,
followed
by
incubation
for
(24 ± 3) hours. The excellent capacity of this semi-solid media to promote migration of
Salmonella, combined with judicious selective agents allows the rapid identification of
presumptive positive samples by simple visualization on the surface of the Petri plate. This step
is nevertheless inappropriate for non-mobile Salmonella (Salmonella Gallinarum, S.
Pullorum,…) although growth can be observed at the inoculation point.
- Any needed confirmation steps are performed by inoculation onto COMPASS Salmonella Agar
with incubation requiring (24 ± 3) hours. The principle of this solid chromogenic, selective
culture media is based on the specific revelation of esterase and β-glucosidase, which for the
former is exclusive to Salmonella and for the latter which is lacking for the vast majority of
strains. Rare strains of Enterobacter have been observed to express both an esterase activity
and β-glucosidase, resulting in blue-green colonies surrounded by a halo of magenta. Certain
strains of Salmonella belonging to the serovars Dublin and Atento, as well as a few strains from
the subspecies S. houtenae, S. bongori and S. diarizonae may also present a weak to nil
magenta pigmentation, due to a poor expression of the esterase activity in these strains.
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TYPICAL COMPOSITION AND PREPARATION OF SESAME Salmonella TEST
(can be adjusted to obtain optimal performance)
SESAME Salmonella Enrichment
(BK194, BM136, BM137 or BM144)
Pour 1 liter of medium :
SESAME Salmonella Detection
(BK195, BM138)
Pour 1 liter of medium :
- Peptone .............................................. 10.0 g
- Sodium chloride .................................... 5.0 g
- Phosphate buffers ............................... 5.06 g
-
pH 7,0 ± 0,2 (at 25°C)
Peptone................................................4,59 g
Acid hydrolysate of casein ...................4,59 g
Sodium chloride ...................................7,34 g
Potassium phosphate ..........................1,47 g
Selective agents................................ 10,98 g
Bacteriological agar .............................2,70 g
pH 5,2 ± 0,2 (at 25°C)
PREPARATION
PREPARATION
- Suspend 20,0 g of dehydrated medium
(BK194) in 1 liter of distilled or deionized
water.
- Mix well, until complete dissolution.
- Divide into tubes or vials.
- Sterilize in an autoclave at 121°C for 15
minutes.
- Suspend
31,7 g of dehydrated medium
(BK195) in 1 liter distilled or deionized water.
- Slowly bring to boiling, stirring with constant
agitation until complete dissolution.
- DO NOT autoclave.
- If the ready-to-melt format (BM138) is used,
heat the medium with the least amount of time
necessary to achieve complete and
homogeneous melting.
- Do not repeat this operation more than once.
- Cool and maintain the medium at 44-47°C.
- Pour into empty, sterile Petri dishes.
- Let solidify on a cool surface.
- Do not dry the plates after cooling.
- The references BM136, BM137 and BM144
are ready-to-use (see § PRESENTATION).
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INSTRUCTIONS FOR USE / RESULTS
- Introduce aseptically 25 g of the sample to be tested into ready-to-use SESAME Salmonella
Enrichment, in order to achieve a 1:10 dilution.
- Mix well.
- Incubate at (37 ± 1)°C for (18 ± 2) hours.
- Inoculate three drops (approximately 0.1 mL) of the enrichment broth from SESAME
Salmonella Enrichment, to the center of a plate of SESAME Salmonella Detection.
- Incubate at (41,5 ± 1,0)°C, for (24 ± 3) hours, without inverting the plates.
The appearance of a white, opaque halo with a diameter equal or superior to 30 mm, at the
inoculation point indicates the presumptive presence of Salmonella.
NOTE 1 :
In the context of AFNOR VALIDATION, test portions weighing more than 25 g have not been
tested.
NOTE 2 :
The use of SESAME Salmonella TEST is not appropriate for immobile Salmonella (Salmonella
Gallinarum, S. Pullorum, …).
NOTE 3 :
SESAME Salmonella Detection can be stored for 72 h at 4°C after inoculation and incubation
before performing subsequent confirmations.
CONFIRMATION
In the context of the AFNOR VALIDATION, positive results must be confirmed by one of several
methods :
ƒ
performing classical tests described in standardized methods such as CEN, ISO or
AFNOR (including purifications steps)
ƒ
by using COMPASS Salmonella Agar
ƒ
though the use of another certified AFNOR VALIDATION method, that uses a different
principle of detection from that used in SESAME Salmonella TEST (the validated
protocol used in this second method must be followed in its entirety ; all the steps
preceding the start off point for the confirmation step must be common to the two
methods : for example, an enrichment medium common to both).
or
or
In the event of discordant results (positive with the alternative method, but lacking confirmation with
one of the options listed above), the laboratory must perform the necessary steps to assure the
validity of the results.
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NOTE 4 :
It is reminded that after migration on SESAME Salmonella Detection and isolation on COMPASS
Salmonella Agar, there exists rare strains of Enterobacter that can express esterase activity and
produce blue-green colonies surrounded by a magenta halo. Certain strains of Salmonella
belonging to the serovars Dublin and Atento, as well as a few strains from the subspecies S.
houtenae, S. bongori and S. diarizonae may also present a weak to nil magenta pigmentation, due
to a poor expression of the esterase activity in these strains.
QUALITY CONTROL
SESAME Salmonella Enrichment :
- Prepared medium : amber solution, limpid, may present a slight precipitate after prolonged
storage.
SESAME Salmonella Detection :
- Prepared medium : blue semi-solid agar.
Typical cultural response after (18 ± 2) hours of incubation at (37 ± 1)°C on SESAME Salmonella
Enrichment, followed by (24 ± 3) hours of incubation at (41,5 ± 1,0)°C on SESAME Salmonella
Detection, with an ultimate subculture on COMPASS Salmonella Agar for (24 ± 3) hours at
(37 ± 1)°C :
Growth on SESAME
Salmonella Detection
Microorganisms
Cultural characteristics after
subculture on COMPASS
Salmonella Agar
Salmonella Enteritidis
CIP 82.97
+ Escherichia coli
ATCC® 25922
+ Pseudomonas aeruginosa ATCC 27853
white, opaque halo
≥ 30mm
magenta colonies
Salmonella Typhimurium
ATCC 14028
+ Escherichia coli
ATCC 8739
+ Pseudomonas aeruginosa ATCC 27853
white, opaque halo
≥ 30mm
magenta colonies
Escherichia coli
Enterococcus faecalis
ATCC 25922
ATCC 29212
inhibited
Inhibited
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STORAGE / SHELF LIFE
ƒ SESAME Salmonella Enrichment
Dehydrated medium : 2-30°C.
- The expiration date is indicated on the label.
Prepared medium (benchmark value*) :
- Prepared media in vials or tubes : 6 months at 2-25°C.
Ready-to-use media in vials or in flexible bags :
- Store between 2 and 25°C, shielded from light.
- The expiration dates are indicated on the labels.
ƒ SESAME Salmonella Detection
Dehydrated medium : 2-25°C, shielded from light.
- The expiration date is indicated on the label.
Prepared medium (benchmark value*) :
- Prepared media in plates : 15 days at 2-8°C, shielded from light.
Ready-to-melt media in vials :
- Store between 2-8°C, shielded from light.
- The expiration date is indicated on the label.
Pre-poured media in Petri dishes :
- Store between 2-8°C, shielded from light.
- The expiration date is indicated on the label.
ƒ SESAME Salmonella TEST Kit :
Ready-to-use media and ready-to-melt media in vials :
- Store between 2-8°C, shielded from light.
- The expiration date is indicated on the label.
PRESENTATION
Code
SESAME Salmonella TEST kit :
- Kit composed of
9 x 225 mL vials SESAME Salmonella Enrichment
and
1 x 200 mL vial of SESAME Salmonella Detection
BT00908
SESAME Salmonella Enrichment :
- 500 g bottle
- 10 x 225 mL
- 3 x 3 liter flexible bags
- 2 x 5 liter flexible bags
BK194HA
BM13608
BM13708
BM14408
SESAME Salmonella Detection :
- 500 g bottle
- 10 x 200 mL
- 20 pre-poured plates in Petri dishes (∅ 90 mm)
- 120 pre-poured plates in Petri dishes (∅ 90 mm)
BK195HA
BM13808
BM14108
BM15008
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BIBLIOGRAPHY
PERRY, D.F., and QUIRING, C.. 1997. Fundamental aspects of enzyme/chromogenic substrate interactions in agar
media formulations for esterase and glycosidase detection in Salmonella. In Salmonella and Salmonellosis-Proceedings.
Ploufragan-France, 63-70.
HUMBERT, F., LALANDE, F., ROSE, V., et SALVAT, G.. 1998. Evaluation d’un nouveau milieu d’isolement pour la mise
en évidence des salmonelles dans les élevages et les denrées d’origine animale. 5ème congrès de la Société Française
de Microbiologie. 128.
NF EN ISO 6579 (V 08-013). Décembre 2002. Microbiologie des aliments. Méthode horizontale pour la recherche des
Salmonella spp.
NF EN ISO 16140 (V 08-103). Octobre 2003. Microbiologie des aliments. Protocole pour la validation des méthodes
alternatives.
XP CEN ISO/TS 11133-2 (V 08-104-2). Janvier 2004. Microbiologie des aliments. Guide pour la préparation et la
production des milieux de culture. Partie 2 : Guide général pour les essais de performance des milieux de culture.
NF EN ISO 6579 (V 08-013). Mars 2007. Microbiologie des aliments. Méthode horizontale pour la recherche des
Salmonella spp.
BKR 23/04-12/07
ALTERNATIVE ANALYTICAL METHODS FOR AGRIBUSINESS
Certified by AFNOR Certification
www.afnor-validation.org
*Benchmark value refers to the expected value under standard laboratory conditions following manufacturer’s instructions. It is provided as a guide only and
no warranty, implied or otherwise is associated with this information.
The information provided on the package take precedence over the formulations or instructions described in this document.
The information and specifications contained in this technical data sheet date from 2009-09-08.
They are susceptible to modification at any time, without warning.
Code document : BT009/A/2008-02 : 4
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