Acinetobacter is a bacteria found in the environment and clinical specimens, causing nosocomial infection and antimicrobial resistance (AMR) threats. This study examined the prevalence, species, and AMR characteristics of Acinetobacter isolated from surgical practice and the laboratory dog husbandry room environments (n = 235) at Rajamangala University of Technology Tawan-ok veterinary hospital during 2018-2019. The prevalence of Acinetobacter in the laboratory dog husbandry room and veterinary belongings were 2.55% and 0.43%, respectively. Species determination was Acinetobacter hemolyticus (2.1%) and Acinetobacter baumannii (0.4%) from environments in the laboratory dog husbandry room, and Acinetobacter junii (0.4%) from the shoes used in the surgical practice room. AMR was observed in both study environments and the specimens sent to the Veterinary Diagnostic Center. These isolates had a high resistant percentage to amoxicillin-clavulanic acid (84.62%), sulfamethoxazole-trimethoprim (61.54%), and cephalexin (53.85%) but were susceptible to imipenem. Compared to the isolates recovered from the clinical specimens, most isolates derived from environments exhibited multidrug resistance and shared correlated resistance patterns. These results highlight the need for sanitization in the dog husbandry room. Furthermore, the AMR results can be used as a preliminary baseline for studying AMR Acinetobacter contamination in animals and their environments.