Tracking verocytotoxigenic Escherichia coli O157, O26, O111, O103 and O145 in Irish cattle and sheep during slaughter

This study presents a duplex real-time PCR assay for the detection of vtl and vt2 genes in ovine carcass swabs, ovine fleece, bovine hide swabs and bovine faecal samples, using TaqMan probes. Sample matrices were spiked with a cocktail of VTEC (E. coli 0157, 026, 0111,0103 and 0145) and enriched for 6 h in mTSB. The resultant DNA was extracted and real-time PCR performed using specific primer/TaqMan probe sets for vti and vt2 genes. Detection was achieved for both low level (approximately 10 cfu/sample) and high level (approximately 10,000 cfu/sample) in all four matrices tested. This study also presents data on the carriage and transfer of verocytotoxigenic Escherichia coli (VTEC) 0157, 026, 0111, 0103 and 0145 from faeces and hide/fleece to dressed carcasses of Irish cattle and sheep as well as establishing the virulence potential of VTEC carried by these animals. Individual animals were tracked and faecal samples, hidelfleece and carcass swabs were analysed for verocytotoxin (vtl and vt2) genes using a duplex real-time PCR assay. Positive samples were screened for the five serogroups of interest by real-time PCR. Isolates were recovered from PCR positive samples using immunomagnetic separation and confirmed by latex agglutination and PCR. Isolates were subject to a virulence screen (vti, vt2, eaeA and hlyA) by PCR. VTEC isolates were examined by Pulsed-Field Gel Electrophoresis (PFGE). This study shows that while VTEC 0157 are being carried by cattle and sheep presented for slaughter in Ireland, a number of other verocytotoxin producing strains (particularly E. coli 026) are beginning to emerge and confirms that that emerging VTEC serogroups should be monitored throughout the food chain.