Phagedx (Tm) E. Coli O157:H7 Assay In Ground Beef Level 3 Modification To Add 375 G Beef Trim And New Method Procedure For 25 G Ground Beef: Aoac Performance Tested Method(Sm) 081601

Background: ThePhageDx (TM) E. coli O157:H7 Assay originally required a 5h enrichment period and a 10mL test sample for 25g ground beef. The proposed method modifications seek to include beef trim (375g) matrix and a new method procedure for 25g ground beef comprised of 6-7 h enrichment and 1mL test sample.Objective: To validate the method modifications for PhageDx E. coli O157:H7 Assay to include beef trim (375g) matrix and modify enrichment time and test sample size for 25g ground beef.Method: For both matrixes, pre-warmed tryptic soya broth (TSB; 42 +/- 1 degrees C) was added in a 3:1 (media:sample size) ratio, blended, and enriched for 9-10 h (beef trim, 375g) or 6h (ground beef, 25g) at 42 +/- 1 degrees C. One milliliter samples were transferred to a microfuge tube, centrifuged, and the supernatant removed. The pellet was resuspended in 0.2mL of TSB and phage reagent was added. Samples were incubated for 2h at 37 +/- 1 degrees C. After infection, the sample was centrifuged, and 150 mu l of the supernatant was transferred to a 96-well plate. Then, lysis buffer and luciferase substrate were added and the sample read on a luminometer to determine the presence or absence of E. coli O157:H7.Results: No significant differences were found between the PhageDx E. coli O157:H7 Assay method modifications and the U.S. Department of Agriculture Food Safety and Inspection Service (FSIS) Microbiology Laboratory Guidebook (MLG) reference method.Conclusions: The independent study demonstrated that the PhageDx E. coli O157:H7 Assay method modifications meet the qualifications for Performance Tested Method(SM) status.Highlights: The PhageDx E. coli O157:H7 Assay is a rapid detection method capable of detecting E. coli O157:H7 in 25 g ground beef and 375 g beef trim samples in as little as 8.5 h and 11.5 h total turn around time, respectively.