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to 10-mL FDA RV broth (prepared from constituent components on the day of testing) and
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incubated for 24
±
2 hr at 42±0.2°C. An additional 1-mL aliquot of each enrichment was added to 10
312
mL TT broth and incubated for 24
±
2 hr at 35±2°C. All samples were then struck in 10-μL amounts
313
onto BS agar, XLD agar and HE agar. Agars were incubated at 35°C for 24±2 hr after which plates
314
were examined for colonies with typical characteristics of
Salmonella
. Suspect colonies were
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confirmed using the biochemical and serological methods described in the FDA-BAM [2].
316
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Twenty spiked and five unspiked samples tested with the BAX® System method (test method)
318
were enriched at 39-42°C in pre-warmed (42°C) BAX® System MP media (without pH adjustment)
319
for 24 hours and tested by BAX® System assay at 10 and 24 hours before secondary enrichment
320
in RV and TT broth as for reference method enriched samples. All samples were then struck in 10-
321
μL amounts on BS agar, XLD agar and HE agar. Agars were incubated at 35°C for 24±2 hr after
322
which plates were examined for colonies with typical characteristics of
Salmonella
. Suspect
323
colonies were confirmed using the biochemical and serological methods described in the FDA-
324
BAM [3].
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Dry Kibble Type Pet Food
- Master samples were made at ~ 50, 100, and 1000 cfu per 25g
327
portion using a wet spike on ground pet food and stored at room temperature. Two to four weeks
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prior to enrichment and assay, pet food was divided into portions of 25g size and each was
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inoculated with 4 X 10µL aliquots of an approximately 2, 5, 20, and 50 cfu / 40µL suspension. The
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portions for each level were re-combined on an aluminum foil coated surface and mixed by hand
331
to homogenize, placed into disinfected plastic tubs, and shaken vigorously to further homogenize.
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Samples were held at room temperature for two to four weeks to acclimate/stress the target cells.
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Prior to testing, range finding studies were conducted on spiked material to determine the sample
334
spike level that would be likely to give fractional recovery. Because range finding studies indicated
335
above fractional levels in the high spiked samples, and below fractional levels in the middle spiked
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level samples, the high spike level material was homogenized with the middle spike level material
337
in a 1:4 ratio (one part high level material to three parts middle level material). As during initial
338
master sample preparation, the portions for the two levels were combined and mixed by hand to
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Dupont BAX Salmonella PTM Report
PTM Certification No. 081201 (08/07/2012)
For Expert Review Panel Use Only
Do Not Distribute