In Ukraine, only 28 % of HIV-infected individuals are aware of their HIV status. Expansion of voluntary HIV counseling and testing (VCT) in primary and specialty health care settings holds promise for increasing the number of people who know their HIV status and can access care. To build capacity among health care providers to deliver VCT, we conducted two-day trainings on basic HIV knowledge and on VCT procedures in the Odessa region of Ukraine. The training program was developed by local trainers in collaboration with faculty from the US Southeast AIDS Training and Education Center and was delivered in the clinical settings where trainees worked (n = 392). We assessed training effectiveness in terms of change in knowledge among clinician trainees, comparing HIV specialists and nonspecialists and those working in urban and rural clinical settings. All else being equal, compared with their urban HIV-specialist peers, trainees who were rural nonspecialists demonstrated significantly greater increases in general HIV knowledge scores. This effort demonstrates that brief, on-site training programs support the expansion of VCT by increasing the knowledge and skills of rural nonspecialist clinicians to levels equal with urban HIV specialists.