Antimicrobial resistance is a global public health crisis. However, there is a paucity of data on anti-protozoal resistance (APR), especially in animals. Hence, we assessed the knowledge of prudent antiprotozoal drug usage (APU) and resistance among animal health practitioners (AHPs) in Kwara State, Nigeria.A cross-sectional survey of 435 AHPs was performed in Kwara State from the 5th of March to the 31st of July 2020 using a structured and validated questionnaire. We used logistic regression analysis to identify socio-demographic factors that are associated with the satisfactory perception of prudent APU and good knowledge of APR among AHPs.Our findings showed that 80.2% (n = 349) of the AHPs in Kwara state had a good knowledge of APR with a mean knowledge score of 5.8 ± 1.2. In the same vein, 75.6% (n = 329) of the AHPs had a satisfactory perception of prudent APU with a mean score of 3.84 ± 1.21. Only 10.1% of the AHPs had observed treatment failures after the use of anti-protozoal drugs. Most of the AHPs (75.6%) believed that APR poses a significant threat to animal production and health globally. Logistic regression analysis showed that female AHPs were more likely (OR: 2.17; 95% CI: 0.91, 5.20; p < 0.005) to have better knowledge of APR than their male counterparts. AHPs with tertiary education were likely (OR: 2.77; 95% CI: 0.96, 4.99; p < 0.05) to be more knowledgeable about APR and have satisfactory perceptions of APU (OR: 1.57; 95% CI: 1.16, 2.99; p = 0.07) respectively. Finally, veterinarians were 3.76 times (95% CI:1.26, 9.25; p < 0.001) more likely to have good knowledge of APR and better perceptions of APU (OR: 3.28; 95% CI: 1.89, 5.68; p < 0.001) than other AHPs respectively.To control antimicrobial resistance, continuous training of AHPs especially para-veterinary officers is essential to update their knowledge on prudent antimicrobial usage and prevent the emergence of resistant protozoan parasites.