Shiga toxin-producing Escherichia coli (STEC) are one of the emerging zoonotic pathogens with cattle being considered as the major reservoirs of infection. Outbreaks of human disease are often traced to contacts with cattle or their products. The present study was, thus, undertaken to isolate and characterize STEC from bovines and their handlers. Faecal samples from bovines (n=103) and their handlers’ stool and fingertip rinses (n=70) were collected over an eight month from August, 2011 to March, 2012. Multiplex polymerase chain reaction (mPCR) for stx₁, stx₂, eaeA and hlyA genes detected STEC in 15 (11.90%) of the 126 E. coli isolates with the most frequent virulence gene combination as stx₁, stx₂ and hlyA. Both stx₁ and stx₂ genes in combination were found to be more frequent (66.6%) than only stx_2­ (26.6%) and stx₁ (6.66%) genes among the STEC isolates. The intimin and enterohaemolysin encoding genes, eaeA and hlyA were detected in 6 (40%) and 11 (73.3%) of the STEC isolates. The overall prevalence of STEC in cattle, buffalo and bovine handlers was 15%, 6.7% and 4.28% respectively. Though there were no similar serogroups of STEC isolated from bovines and their handlers, their virulence gene profile was similar in one farm. A positive correlation (r=0.958) of STEC prevalence of bovines and their handlers existed in different localities. The study indicates that bovines in the region harbour STEC, and are the probable source of STEC transmission to humans especially occupationally exposed groups.