AOAC ERP MICRO AUGUST 2018

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

enrichments for each sample was adjusted to 6.8 ± 0.2. Test portions were incubated at 36 ± 1 o C

1 2 3 4 5 6 7 8 9

for 18-26 hours.

Following incubation 0.1 mL of primary enrichment was transferred into 10 mL of Rappaport- Vassiliadis medium (RV) and 1.0 mL into 10 mL of Tetrathionate (TT) broth. RV tubes were incubated at 42 ± 0.2 o C for 24 ± 2 hours and TT tubes were incubated at 43 ± 0.2 o C for 24 ± 2 hours due to high microbial background (>10 4 CFU/mL). Following incubation, a loop full of the secondary enrichments were streaked to BS, HE, and XLD and incubated at 35 ± 2 o C for 24 ± 2 hours. If no visible colonies were present after 24 hours of incubation on the BS plates, they were re-incubated for an additional 24 ± 2 hours at 35 ± 2 o C. A minimum of two suspect colonies from each selective agar were transferred to TSI and LIA and incubated at 35 ± 2°C for 24 ± 2 hours. Following incubation, TSI and LIA slants were examined for typical reactions. Slants producing typical reactions were streaked to TSA and incubated for 35 ± 2°C for 18-24 hours. Following incubation, isolates were serologically tested for both somatic O and flagella H agglutination. Additionally, purified TSA isolates were identified using the VITEK ® GN Biochemical Identification card following AOAC Official Method 2011.17.

10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46

3M ™ MDA 2 - Salmonella

All matrixes were enriched and incubated according to the AOAC protocol as described previously in “General Preparation”, subsection “ Performing Enrichments”. After incubation all

test portions were processed by the 3M ™ MDA 2 - Salmonella .

Molecular Detection System

The reaction tubes or strips containing samples were loaded into the MDS instrument and prompts were followed using the MDS software to identify samples and controls. The MDA 2 Salmonella assay was initiated and results were obtained within 75 minutes.

Interpretation of Results

Determining the presence or absence of pathogen DNA was carried out based on an algorithm that interprets the light output curve resulting from the detection of the nucleic acid amplification. Results were automatically analyzed by the software and were color-coded based on the result. A positive or negative result was determined by analysis of a number of unique curve parameters. Presumptive positive results were reported in real-time while negative results

AOAC Research Institute Expert Review Panel Use Only

were displayed after the run was complete.

Confirmation

All samples analyzed by the 3M ™ MDA 2 - Salmonella , regardless of presumptive result, were confirmed by procedures specified for each matrix in the reference methods. Final confirmation was achieved by VITEK ® 2 GN Biochemical Identification, AOAC OMA 2011.17

Matrix Study Results