Antimicrobials are widely used to maintain the health and productivity of animals. However, the misuse of antimicrobials in animals contributes to the development of antimicrobial resistance (AMR), an emerging One Health issue and a growing global health challenge.

This study assessed animal health practitioners’ knowledge, awareness, and practices regarding antimicrobial use and antimicrobial resistance, as well as the frequency of antimicrobial usage in animals in Niger State. A questionnaire-based cross-sectional survey was conducted, employing a purposive sampling method to select animal health practitioners across the state. Descriptive and analytical statistical analyses were performed at a 95% confidence level.

A high proportion (62.8%) of the targeted animal health practitioners participated in the survey. Concerning the uses of antimicrobials in animals, 42.3% of respondents indicated they used antimicrobials for the treatment of infections, while 12.2% mentioned growth promotion as a reason for use. Only 19.5% correctly identified antimicrobial misuse as administering an under-dose; 16.3% considered it as giving an over-dose, and 13.0% did not know what misuse entailed.

Regarding disease status determination before administering antimicrobials, 10.6% of practitioners did not assess the disease status before treatment, while only 1.6% reported carrying out confirmatory diagnosis prior to antimicrobial administration. A high proportion (69.1%) of practitioners administered daily dosages of antimicrobials to sick animals as prescribed on the product labels.

The most frequently used antimicrobials were tetracycline (99.2%) and penicillin-streptomycin (82.1%). Satisfactory knowledge about antimicrobial use was significantly associated with being within the age group of 50–59 years (p = 0.002) and being a veterinarian (p = 0.001).

Factors that significantly influenced antimicrobial misuse included improper antimicrobial usage (p < 0.001), lack of enforcement of regulatory laws (p = 0.005), weak financial status of clients (p = 0.001), low education and limited expertise of some practitioners (p = 0.001), and the proliferation of counterfeit antimicrobials (p = 0.001).