A 5-h screening and 24-h confirmation procedure for detecting Escherichia coli O157 : H7 in beef using direct epifluorescent microscopy and immunomagnetic separation |
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Authors: | L Restaino E W Frampton R M Irbe & D R K Allison |
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Institution: | R &F Laboratories, West Chicago, IL, USA |
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Abstract: | An antibody-direct epifluorescent filter technique (Ab-DEFT) detected 100% of the raw ground beef samples inoculated with Escherichia coli O157 : H7 cells (0·15 cells g?1) and incubated in a prewarmed, modified buffered peptone water (mBPW) non-selective enrichment broth for 5 h at 42°C in an orbital shaking water bath (200 rev min?1). Over 50% of the microscopic fields viewed were positive (1–10 fluorescent cells field?1) in the Ab-DEFT. All positive screening results were confirmed within 24 h by subjecting 1 ml of the mBPW to the Dynabeads® anti- E. coli O157 immunomagnetic separation procedure, followed by plating on MacConkey sorbitol agar containing 5-bromo-4-chloro-3-indolyl-β- D -glucuronide. At this cell concentration, 41·7% of the inoculated samples were detected by the conventional method involving a 24-h selective enrichment. Exposure to viable cells before filtration was minimized by using a 0·58% formaldehyde concentration for 5 min at 50°C (killed >4·00 logs of E. coli O157 : H7 cells) without affecting cell fluorescence. |
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