AOAC ERP MICRO AUGUST 2018

OMAMAN-31 E: AOAC RI PTM Report 091501 ERP Use Only - March 2016

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For instant non-fat dry milk, cocoa powder, black pepper, and dry dog food lyophilized cultures were used in the validation. Lyophilized cultures were prepared by transferring a single Salmonella colony from SBA into BHI broth and incubating the culture at 35 ± 2 o C for 18-24 hours. The culture was then diluted in a sterile cryoprotectant, reconstituted non-fat dry milk (NFDM), and freeze dried for 48-72 hours. A bulk lot of the matrixes were inoculated with a lyophilized culture that was diluted in NFDM to a low level expected to yield fractional positive results and a high level expected to yield all positive results. After inoculation matrixes were All raw or fresh food matrixes were held for 48-72 hours post-inoculation at 2-8 °C to allow for equilibration of the organism as per AOAC guidelines. For all 325 gram test portions, 25 grams of inoculated matrix was mixed with 300 grams of uninoculated matrix. For the 375 gram test portions of dry dog food, creamy peanut butter, and spent sprout irrigation water, 25 grams of inoculated matrix was mixed with 350 grams of uninoculated matrix. For pasteurized liquid whole egg, a 100 gram test portion was directly sampled from the bulk inoculated matrix. For all 25 gram test portions, a 25 gram test portion was directly sampled from the bulk inoculated For sealed ceramic tile and sealed concrete surfaces, 4” x 4” areas were inoculated with 0.25 mL of diluted Salmonella culture and sampled using sampling sponges. For the uninoculated test portions, sterile BHI broth was applied to the test area. Sealed concrete was also inoculated with a competitor organism, Proteus vulgaris ATCC 6380, at 10 x the level of the target organism. For stainless steel, 1” x 1” areas were inoculated with 0.10 mL of diluted culture and sampled using sampling swabs. For the uninoculated test portions, sterile BHI broth was applied to the test area. Each surface was dried for 16-24 hours at room temperature (24 ± 2 o C) prior to The 3M ™ hydrated sampling sponges (pre-moistened with Dey-Engley broth) (sealed ceramic tile and sealed concrete) and 3M ™ Tecra Enviro Swabs (pre-moistened with Letheen broth) (stainless steel) were utilized to sample by using horizontal and vertical sweeping motions. The sponges and the swabs were held at room temperature (24 ± 2 °C) for 2 hours prior to enrichment. To determine the inoculation level for the environmental surfaces, aliquots of each The level of Salmonella in the low level inoculum for all 25 g test portions was determined by Most Probable Number (MPN) on the day of analysis by evaluating 5 x 50 g, 20 x 25 g (reference method test portions), and 5 x 10 g inoculated test samples. The level of Salmonella in the low level inoculum for all 100 g test portions was determined by MPN by evaluating 5 x 200 g, 20 x 100 g (reference method test portions), and 5 x 50 g inoculated test samples. The level of Salmonella in the low level inoculum for all 325 g test portions was determined by MPN by evaluating 5 x 650 g, 20 x 325 g (reference method test portions), and 5 x 160 g inoculated test samples. The level of Salmonella in the low level inoculum for all 375 g test portions was determined by MPN by evaluating 5 x 750 g, 20 x 375 g (reference method test portions), and 5 held for 2 weeks at room temperature (24 ± 2 o C). matrix. sampling. inoculum were plated onto Tryptic Soy agar (TSA) in triplicate.

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AOAC Research Institute Expert Review Panel Use Only

x 180 g inoculated test samples.