Bartonella spp have been implicated in various syndromes (eg, vasculitis, endocarditis, myocarditis) in dogs and humans. Few studies describe the prevalence of Bartonella spp in canids and/or the fleas infecting them. This study investigated the genera of fleas infesting dogs in two Florida animal shelters, determining the molecular prevalence of Bartonella spp in the fleas and the molecular prevalence of Bartonella spp in the 80 dogs from which the fleas were collected. Flea genera were determined for 43 dogs: 28 were infested with C felis alone, 8 were infested with Pulex spp alone, and 7 were infested by C felis and Pulex spp. Eighteen dogs were considered positive for Bartonella spp—via either whole blood testing or because infesting fleas tested positive for Bartonella DNA. The data supported the hypothesis that dogs may serve as a reservoir host for these fleas, presenting an important link in transmission between certain Bartonella spp pathogens implicated in human disease.