Aims: To determine the prevalence, seasonal variation and virulence characteristics of Escherichia coli O157:H7 in lambs presented for slaughter in Ireland. Methods and Results: Over a 13-month period, pre- and postchill carcass swabs, faeces and fleece samples from 1600 lambs were examined for the presence of E. coli O157:H7. Escherichia coli O157:H7 was isolated from 5.75% (23/400) of fleece samples, 1.5% (6/400) of pre- and 1% (4/400) of postchill carcass swabs but was not isolated in faeces (0/400). The present study detected no evidence of seasonal variation. Polymerase chain reaction analysis showed that both the vt1 and vt2 genes associated with clinical illness were carried by five of the E. coli O157:H7 isolates, while 24 of the remaining isolates carried the vt2 gene only. Phage typing detected four different subtypes: PT 32 (48.48%), PT 8 (12.12%), PT 31 (12.12%) and PT 21/28 (12.12%). Conclusions: Escherichia coli O157:H7 is present in lambs at slaughter in Irish abattoirs and the virulence profiles of these isolates reveals that they are potentially harmful to humans. Significance and Impact of the Study: The present study provides crucial information indicating that sheep may be a significant contributing source to human E. coli O157:H7 infection.