Presence and characterisation of verotoxin producing E. coli in fresh Italian pork sausages, and preparation and use of an antibiotic-resistant strain for challenge studies

One hundred and twenty six samples of fresh pork sausages were analysed for the presence of verocytotoxigenic Escherichia coli (VTEC). Selective enrichment followed by DNA extraction and PCR amplification of the stx1 and stx2 genes highlighted the occurrence of the above mentioned genes in 20 out of 126 samples screened. From the stx positive enriched cultures, isolation was performed on CT-SMAC agar plates after immuno-magnetic separation of E. coli O157. Fifty three non-sorbitol fermenting isolates were obtained and further characterised, along with the reference strain E. coli ATCC 35150T. All the isolates were characterised by PCR assays, assessing the presence of stx1, stx2, rfbEO157:H7, eae and hlyA genes. The overall prevalence of VTEC was found to be 16%. VTEC strains were also characterised by plasmid profiling and REA-PFGE analysis, which allowed strain clustering into 5 and 8 groups, respectively. In addition, an antibiotic resistant E. coli O157:H7 strain was selected and used in challenge tests of raw pork at 4 °C. This strain could be selectively counted in the presence of a normal background microflora and it was shown that it could survive for 1 week at 4 °C in the raw food studied.