Review Book

Test portions (375 g) evaluated by the candidate method were enriched with 1125 mL of BPW (prewarmed to 42 ± 1 o C), homogenized and incubated for 10 h at 42 ± 1°C. Following incubation, a 20 μ L aliquot of each sample was lysed for 5 min at >2000 rpm. Lysed samples were transferred to PCR reaction tubes and assayed on the GENE-UP ® thermocycler. Regardless of presumptive results, all test portions were confirmed following two procedures: a traditional confirmation following the MLG 4.10 reference method beginning with a transfer to secondary enrichments and an alternative confirmation procedure by directly streaking the primary enrichment broth to ASAP and CHROMID Salmonella and incubating at 35 ± 1°C for 24 ± 2 h. Isolated colonies from both the traditional and alternative confirmation procedures were confirmed following the procedures outline in MLG 4.10 with biochemical confirmation conducted using VITEK 2 GN (OMA 2011.17) or API 20E (OMA 978.24). For test portions evaluated by the reference method, 25 g samples were enriched in 75 mL mTSB and incubated for 15–24 h at 42 ± 1°C. After incubation, an aliquot of the mTSB was transferred to secondary enrichments (Tetrathionate broth, Hajna, TT and Rappaport-Vassiliadis medium with soya, RVS). Secondary enrichment tubes were incubated at 42 ± 0.5 o C for 22-24 h in an incubator or for 18-24 h in a circulating water bath. After incubation, secondary enrichments were struck to selective agars (Brilliant green sulfa agar, BGS and double modified lysine iron agar, DMLIA) and incubated at 35 ± 1°C for 18-24 h. Regardless of the presence of typical colonies, all plates were reincubated for an additional 18-24 h. Isolated colonies from the selective agars were transferred to triple sugar iron agar (TSI) and lysine iron agar (LIA) slants and incubated at 35 ± 1°C for 24 ± 2 h. Final biochemical confirmation was conducted using VITEK 2 GN or API 20E.

Statistical Analysis

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