BACKGROUND : Experimental infection of horses with Bartonella species is not documented. OBJECTIVES : Determine clinical signs, hematologic changes, duration of bacteremia, and pattern of seroconversion in Bartonella henselae or Bartonella bovis-inoculated horses. ANIMALS : Twelve (2 groups of 6) randomly selected healthy adult horses seronegative and culture negative for Bartonella spp. METHODS :Experimental/observational study: Group I: B. henselae or saline control was inoculated intradermally into 4 naïve and 2 sentinel horses, respectively. Group II: same design was followed by means of B. bovis. Daily physical examinations, once weekly CBC, immunofluorescent antibody assay serology, real-time polymerase chain reaction (PCR), and twice weekly blood cultures were performed for 6 weeks and at postinoculation day 80 and 139. Bartonella alpha-Proteobacteria growth medium (BAPGM) enrichment blood culture was performed for horses that seroconverted to B. henselae antigens. RESULTS : Transient clinical signs consistent with bartonellosis occurred in some Bartonella-inoculated horses, but hematological alterations did not occur. Three B. henselae-inoculated horses seroconverted, whereas 1 B. bovis-inoculated horse was weakly seropositive. In Group I, B. henselae was amplified and sequenced from BAPGM blood culture as well as a subculture isolate from 1 horse, blood from a 2nd horse, and BAPGM blood culture from a 3rd horse although a subculture isolate was not obtained. All sentinels remained PCR, culture, and serology negative. CONCLUSIONS : Detection of Bartonella sp. in blood after experimental inoculation supports bacteremia and seroconversion. Culture with BAPGM may be required to detect Bartonella sp. Although mild clinical signs followed acute infection, no long-term effects were noted for 2 years postinoculation.