This study was aimed to assess zoonotic disease knowledge, risk perceptions, and preventive practices of animal health professionals in Nigeria. Cross-sectional questionnaire survey was conducted on 582 participants and 529 responded. Collected data were analyzed by descriptive statistics and multivariate logistic regression models. The proportion of veterinarians (92.0%) which knowledge about zoonosis was much higher (P < 0.001) than that of para-veterinarians (32.4%). In contrast to para-veterinarians (46.2%), the majority of veterinarians (76.7%) perceived high risk of zoonotic infections during necropsy/tissue collections. Similarly, a much higher (P < 0.001) proportion of veterinarians (54.0%) considered hand washing before eating at work as effective way of risk mitigation, compared to para-veterinarians (25.0%). Professionals in large animal practice were less likely (OR 0.35; 95% CI: 0.16, 0.77) not to be engaged in satisfactory protective measures. These results constitute public health contributions to the risk mitigation information that may support measures for zoonosis prevention in Nigeria.