The putative role of biting flies in Bartonella transmission among ruminants was investigated. Amplification of the Bartonella citrate synthase gene from 83 Hippoboscidae was detected in 94% of 48 adult Lipoptena cervi flies, 71% of 17 adult Hippobosca equina flies, 100% of 20 adult Melophagus ovinus flies, and 100% of 10 M. ovinus pupae. Our findings suggest that Hippoboscidae play a role in the transmission of Bartonella among ruminants. The vertical transmission of Bartonella in M. ovinus and the presence of Bartonella DNA in all samples suggest a symbiotic association between Bartonella and M. ovinus.Bartonella spp. are intracellular small gram-negative bacteria transmitted by blood-sucking arthropods and considered to be emerging pathogens in humans and animals (1,8,9,22). In recent years, these organisms have been identified in a wide range of wild and domestic mammals (4, 6, 7, 11), some of which have been associated with zoonoses. Recently, four new Bartonella species have been isolated from ruminants: B. schoenbuchensis and B. capreoli were recovered from wild roe deer (Capreolus capreolus) (3, 10), whereas B. bovis (3) and B. chomelii (20) were recovered from domestic cattle. There are no pathological outcomes associated with Bartonella infection in ruminants (5, 7).Arthropod vectors involved in the transmission of Bartonella spp. among ruminants are still unknown. As blood-sucking ectoparasites of ruminants, flies of the family Hippoboscidae are good candidates for the transmission of Bartonella. Among Hippoboscidae, Lipoptena, Hippobosca, and Melophagus are the three main genera which parasitize mammals (14, 18). The deer ked (Lipoptena cervi), the predominant Lipoptena species in Europe, parasitizes cervids (15), whereas the louse fly (Hippobosca equina) parasitizes cows and horses and the sheep ked (Melophagus ovinus) is a permanent ectoparasite of sheep (Ovis aries) (18).The aim of this study was to determine if Hippoboscidae could be putative vectors of Bartonella spp. in ruminants. We investigated whether Bartonella DNA could be detected in adult and pupal stages of Hippoboscidae collected from ruminants (domestic cattle and roe deer) known to be naturally infected with Bartonella spp. We extended the study to Hippoboscidae collected from sheep and horses for which no evidence of Bartonella infection had ever been demonstrated.Collection and identification of Hippoboscidae. Eighty-three Hippoboscidae flies of different species, stages, and genders were collected and taxonomically identified under a binocular lens (18). Samples were obtained from the hosts or from our parasitology collection (Table 1). Each sample was stored in absolute ethanol.DNA extraction and PCR. Each fly and pupa were washed three times in sterile water baths and once in a 70% ethanol bath and then dried. DNA was then extracted after the flies were crushed with a bead beater as previously described (16).A 380-bp fragment of the citrate synthase (gltA) gene of the genus Bartonella was amplified in fly DNA extracts by PCR, us...