Influence of cold stress on the preliminary enrichment time needed for detection of enterohemorrhagic Escherichia coli in ground beef by PCR

Abstract

The influence of cold stress at 4 and 0°C on the detection time as assessed by impedance technology (Bactometer; Biomerieux, Marcy I'Etoile, France) of different enterohemorrhagic Escherichia coli (EHEC) strains was determined. Although there is some variation in susceptibility among EHEC strains, prolonged exposure of EHEC to cold stress, i.e., 4 and 5 days at 4 and 0°C, respectively, in general significantly increased their detection time. This reflects an increase of the lag-phase time caused by cold stress. Two EHEC strains were selected to determine the minimum preliminary enrichment time that would ensure a positive PCR detection of low numbers of verotoxin-producing E. coil (VTEC; 2 to 2 x 105 CFU/25 g) inoculated into ground beef (25 g) and stored at 4 or -20°C for 8 and 14 days, respectively. Incubation times of 6 and 9 h of 1 to 10 CFU/g and I to 10 CFU/25 g, respectively, were sufficient for PCR detection of VTEC in ground beef when analysis was performed immediately after inoculation (no cold stress). When cells are exposed to cold stress (4 or -20°C) a 24-h enrichment period is recommended. Restriction of enrichment time to 9 h under these circumstances decreases the sensitivity of PCR detection to 80 CFU/g. Hence, to obtain maximum sensitivity, PCR detection of VTEC in naturally contaminated ground beef should be performed after 24 h of enrichment.