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AOAC INTERNATIONAL
For dry pet food, master samples were made at approximately 50, 100, and 1000 cfu per 25g
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portion using a wet spike on ground pet food* and stored at room temperature. Pet food was divided
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into portions of 25 g size and each was inoculated with 4 X 10 µL aliquots of an appropriately diluted
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suspension. Some portions were left un-spiked to serve as negative controls. Spiked portions at each
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target level were re-combined on an aluminum foil coated surface and mixed by hand to homogenize,
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placed into disinfected plastic tubs, and shaken vigorously to further homogenize. All samples were
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held at room temperature for two to four weeks to acclimate/stress the target cells, then range-finding
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studies were performed to determine the actual spike levels. Based on the results of these range-
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finding studies, pet food portions spiked at 100 and 1000 cfu/portion were combined in a 4:1 ratio
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(three parts middle level material to one part high level material) to create test portions with the
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appropriate inoculation level to produce fractional positive results. Analytical size portions (25 g spiked
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master sample and 350 g of unspiked product for each sample) were placed into sterile bags.
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* Note: Since
Salmonella
in real world dry pet food samples appears to originate as a post process
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event, it is felt that the wet spike and target stress by extended drying down/desiccation on the low A
w
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material is the most appropriate way to mimic a naturally occurring contamination event. Previous studies
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had indicated an approximately 2 log reduction in viable cells after
Salmonella
inoculated in this manner
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age for two weeks in dry pet food and comparable results were seen in this study. Since 2.5 X 10
2
cfu on
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average were inoculated into each sample portion, it can be inferred that the degree of injury for the target
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under these conditions is >99%.
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Sample Enrichment
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On the day of testing, an MPN was conducted for each sample matrix using 5 X 100g and 5 X 10g
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portions of the master sample. An aerobic plate count (APC) on un-spiked material was also
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conducted on the day enrichment commenced.
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For unpaired study designs, all alternative primary enrichments, without regard to presumptive
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alternative method result, were subjected to the secondary enrichments and subsequent plating,
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biochemical, and serological confirmatory steps of the appropriate reference method.
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Dupont BAX Salmonella PTM Report
Modification Approved 2012 / PTM Certification No. 081201
For Expert Review Panel Use Only
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