Since World War II, the military has experienced outbreaks of Q fever among deploying units including recent case reports of Q fever in US military personnel returning from serving in the Middle East during Operation Iraqi Freedom and Operation Enduring Freedom. Occupational exposure and prevalence of Q fever among US Army Veterinary Corps officers have not been examined. A retrospective serosurvey and observational study of 500 military veterinarians were conducted using archived serum specimens from military veterinarians who entered and served between 1989 and 2008 and were tested for exposure to Coxiella burnetii. Corresponding longitudinal health-related, demographic, medical and deployment data were examined. A total of 69 (13.8%) individuals at military entry and 85 (17%) had late career positive titres. A total of 18 (3.6%) individuals showed seroconversion. Women were more likely to be seropositive after military service [prevalence ratio (PR) 1.96; 95% confidence interval (CI) 1.15–3.35] and were also more likely to seroconvert (incidence rate ratio 3.55; 95% CI 1.19–12.7). Women who deployed to Operation Iraqi Freedom were more likely to be seropositive (PR 3.17; 95% CI 1.03–9.71). Veterinarians with field service and pathology specialties had the highest incidence rates (7.0/1000 PY; 95% CI 4–12 and 3–19, respectively). This is the first report documenting US military veterinarians' exposure to C. burnetii. Military veterinarians are at risk prior to service, with moderate number of new cases developing during service and most maintaining titres for long periods of time. Women consistently demonstrated higher seroprevalence and incidence levels. As increasing numbers of women enter the veterinary profession and subsequently the US Army, this may warrant close monitoring. This study likely underestimates exposure and risk and does not address chronic health effects, which may be valuable to explore in future health studies.