Buffalo fly (Haematobia irritans exigua) is a major pest of beef and dairy cattle in northern Australia. Global warming is expected to increase the southern range of buffalo fly. Chemical control is compromised by resistance and may not be feasible in extensive production systems and there is rapidly growing market preference for beef and dairy products produced in low-chemical systems. Wolbachia are vertically transmitted intracellular bacteria that can profoundly influence host reproduction and fitness and are of increasing interest for use in biocontrol programs. To determine whether Australian flies are infected with Wolbachia, buffalo flies were collected from 12 cattle herds around Australia and assayed by standard PCR for the Wolbachia wsp gene. H. i. exigua from Indonesia and horn fly (H. i. irritans) from Canada were also tested. All H. i. exigua samples tested were negative for Wolbachia infection whereas a very strong signal for Wolbachia was obtained from H. i. irritans.