The prevalence and antimicrobial resistance of respiratory pathogens isolated from feedlot cattle in Canada. Front Microbiol 2025 [doi: https://doi.org/0.3389/fmicb.2025.1497402]. OBJECTIVES: This study investigated in vitro antimicrobial resistance (AMR) classification in bovine respiratory disease (BRD) pathogens recovered from healthy feedlot cattle enrolled over 2 years, specifically Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni. METHODS: Deep-guarded nasopharyngeal (NP) swabs were used to sample feedlot cattle in multiple (2019 n=21, 2020 n=26) feedlots at 2 timepoints. NP swabs were collected from 16 animals upon entry processing and later in the feeding period at randomly selected feedlots. Cattle from the same groups (not necessarily the same animals) were sampled at both timepoints. Susceptibility testing was performed using the broth microdilution method. RESULTS: Forty-seven groups of cattle and 1,392 cattle were sampled over 2 years, providing 625 bacterial isolates for investigation. Pasteurella multocida (27.4%; 381/1392) was the most frequently isolated BRD bacterium, followed by H. somni (9%; 125/1392) and M. haemolytica (8.5%; 119/1392). Resistance to ≥ 3 antimicrobial classes was detected in 2.4% (95% CI 0.8% - 7.1%) of M. haemolytica, 3.4% (95% CI 0.7% - 14.8%) of H. somni, and 21.3% (95% CI 12.5% - 33.9%) of P. multocida isolates. Potential associations were investigated between recovery of resistant bacteria and sampling quarter, sampling timepoint (arrival or second sample), days on feed (DOF), animal age categories, and BRD risk categories. There was a significant (p<0.05) increase in resistance prevalence after arrival for macrolide drugs in M. haemolytica, and for ampicillin, danofloxacin, enrofloxacin, spectinomycin, gamithromycin, tildipirosin, tulathromycin and tetracycline in P. multocida isolates. Histophilus somni isolates recovered from calves were more likely to be resistant to tulathromycin than those recovered from yearlings (p<0.05), while P. multocida isolates recovered from calves were more likely to be resistant to gamithromycin, spectinomycin, tulathromycin, tildipirosin, and tetracycline than those recovered from yearlings (p<0.05). Resistance to tetracycline, tildipirosin, and tulathromycin decreased between 61-80 and 81-100 DOF when compared to 20-40 DOF, whereas, spectinomycin resistance was lower in cattle sampled between 61-80 DOF than those sampled at 20-40 DOF for P. multocida. DISCUSSION: The diversity of AMR profiles and associated risk factors among BRD pathogens underscores the importance of including all 3 bacteria in future AMR studies in beef cattle.