Efficacy of enrichment broths in the recovery of freeze-injured Escherichia coli O157:H7 in inoculated ground beef by PCR.
Escherichia coli O157:H7 strains ATCC 35150 and ATCC 43894 and five pooled isolates from beef and pork freeze injured at -25 degrees C in beef infusion were used to inoculate ground beef. Samples (25 g each) were added to 225 ml of buffered peptone water with vancomycin, cefsulodin, and cefixime (BPW-VCC), 225 ml of modified EC broth plus novobiocin (mEC+n), and 225 ml of R&F enrichment broth (R&F-EB) and aerobically incubated at 41 to 42 degrees C. After 6, 7, 8, and 24 h of incubation, levels of E. coli O157:H7 recovered from each broth by a PCR assay with the BAX automated system as well as by conventional enrichment with the use of nonaerated mEC+n incubated at 35 degrees C for 24 h were compared with levels recovered by cultural isolation with immunomagnetic separation and plating on BCM E. coli O157:H7 chromogenic agar. For ground beef inoculated with a mean of 4.23 +/- 1.00 total cells (74% freeze injured) per 25 g, after 6 h the PCR assay identified 72.7, 57.6, and 66% of the samples for R&F-EB, BPW-VCC, and mEC+n, respectively, as presumptive positive, whereas the recovery rates after 7 and 8 h exceeded 90%, with the rate for R&F-EB being 100%. For ground beef inoculated with a mean of 1.50 +/- 0.56 total cells (80% freeze injured) per 25 g, after 6 h the PCR assay identified 47.6, 19.1, and 9.5% of the samples for R&F-EB, BPW-VCC, and mEC+n, respectively, as presumptive positive. These values increased to 81.0, 61.9, and 52.4% after 7 h and to 95.2, 61.9, and 71.4% after 8 h. After 24 h, only 55 to 60% of the samples at both inoculum levels tested positive by PCR with conventional enrichment and incubation, whereas >95% of the samples tested positive with R&F-EB aerated at 41 to 42 degrees C. Culture results for R&F-EB and mEC+n after 7 and 8 h of incubation were closely correlated with presumptive positive PCR results.
Lionberg
W C
WC
R&F Laboratories, 245 West Roosevelt Road, Building 3, Unit 16, West Chicago, Illinois 60185, USA. rf_labs@worldnet.att.net
Restaino
L
L
Frampton
E W
EW
Barbour
W M
WM
D000818:Animals; D002417:Cattle; D015169:Colony Count, Microbial / Q000379:methods; D003470:Culture Media / Q000737:chemistry*; D019453:Escherichia coli O157 / Q000254:growth & development / Q000302:isolation & purification*; D005615:Freezing; D018189:Immunomagnetic Separation / Q000379:methods; D007700:Kinetics; D008461:Meat Products / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D013552:Swine; D013997:Time Factors
D003470:Culture Media