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

Abstract

This study presents a duplex real-time PCR assay for the detection of vt1 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 O157, O26, O111, O103 and O145) and enriched for 6 h in mTSB. The resultant DNA was extracted and real-time PCR performed using specific primer/TaqMan probe sets for vt1 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) O157, O26, O111, O103 and O145 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, hide/fleece and carcass swabs were analysed for verocytotoxin (vt1 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 (vt1, vt2, eaeA and hlyA) by PCR. VTEC isolates were examined by Pulsed-Field Gel Electrophoresis (PFGE). This study shows that while VTEC O157 are being carried by cattle and sheep presented for slaughter in Ireland, a number of other verocytotoxin producing strains (particularly E. coli O26) are beginning to emerge and confirms that that emerging VTEC serogroups should be monitored throughout the food chain.

VTEC O157 was the most frequently isolated serotype from the cattle tracking study and was, to a much lesser extent, isolated from sheep fleece and carcass samples. The most frequently isolated non-O157 VTEC from both cattle and sheep tracking studies was VTEC O26. VTEC O103 and O145 were isolated at a frequency no greater than 1% in each of the sample types in cattle with no VTEC O145 isolates found in sheep samples. VTEC O111 were not isolated from cattle or sheep tracking studies. This work concluded that as the majority of VTEC recovered carried eaeA and hlyA genes, they are potentially of clinical significance because they carry the genetic factors linked to severe human illness. This is a significant finding as it highlights the importance of monitoring emerging VTEC serogroups to aid in epidemiologic investigations and prevention of clinical illness.

This study provides information that fills a knowledge gap in regards to potentially significant E. coli O26, O111, O103 and O145 as well as updated information in regards to E. coli O157 in Irish cattle and sheep. Further investigation and continued surveillance of VTEC (and emerging E. coli serogroups of clinical interest) in Irish cattle and sheep is necessary to help investigations into the routes of transmission of VTEC to prevent future outbreaks.

Date of Award	Oct 2013
Original language	English
Supervisor	DAVID MCDOWELL (Supervisor)