ARS | Publication request: Inoculation of beef with low concentrations of Escherichia coli O157:H7 and examination of factors that interfere with its detection by culture isolation and rapid methods

Submitted to: Journal of Food Protection
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 30, 2010
Publication Date: December 1, 2010
Citation: Bosilevac, J.M., Kalchayanand, N., Schmidt, J.W., Shackelford, S.D., Wheeler, T.L., Koohmaraie, M. 2010. Inoculation of beef with low concentrations of Escherichia coli O157:H7 and examination of factors that interfere with its detection by culture isolation and rapid methods. Journal of Food Protection. 73:2180-2188.

Interpretive Summary: Tests for E. coli O157:H7 need to detect one cell in a beef sample that can be 375 grams in size. However, in order to determine if this is possible, first one cell must be placed into a sample of that size, then the detection tests need to be verified at that level of bacteria, and finally problems that may interfere need to be examined. Preparing and dispensing one cell of E. coli O157:H7 is difficult. We provide a protocol for preparing a one-cell suspension and determined that half of samples do not receive any E. coli when this is done. When samples were inoculated with 5 cells of E. coli O157:H7, detection tests identified it was present approximately 95 percent of the time. It was found that extra bacteria that may be present in a beef sample could interfere with detection, also the fat in the sample could reduce the recovery of magnetic beads used to detect E. coli O157:H7.

Technical Abstract: Reliable detection of Escherichia coli O157:H7 in test-and-hold programs requires the detection of as little as 1 CFU E. coli O157:H7 in a sample of beef trim or ground beef that is up to 375 grams in size. We present a reliable protocol for generating a control inoculum for verification testing at this low level and evaluate its use. Results show that half of all samples received no cells when 1 CFU was the target concentration, and that targets greater than 3 CFU were much more reliable. Detection by culture isolation and two commercial assays, Qualicon BAX-MP and BioControl GDS, identified 94±11%, 92±10% and 92±7% of samples inoculated with 5.4 CFU (range 1 to 9 CFU), respectively. We also examined the effect of background flora and fat content effects on the detection of E. coli O157:H7. At levels of background flora below 6 log CFU/g APC, the rapid methods identified all samples inoculated with 26 CFU E. coli O157:H7 per 65g. At APC of 6.7 log CFU/g, culture, BAX-MP and GDS did not detect 0%, 25% and 87% respectively, of inoculated samples, and neither commercial method detected E. coli O157:H7 when APC was 7.7 log CFU/g while culture did not detect 37%. Increased fat content correlated with decreasing recovery of immunomagnetic separation (IMS) beads but this was not observed to interfere with detection of E. coli O157:H7.