Simple SummaryMannheimia haemolyticais a key bacterial pathogen contributing to important ruminant diseases and accounting for a large proportion of overall antimicrobial use in cattle and sheep. The recent emergence of ovine strains with reduced susceptibility to antimicrobials—which could lead to treatment failure, increased costs of livestock production, and the dissemination of antimicrobial-resistant genes to other bacteria and possibly the environment—is of concern. This study investigated the levels of antimicrobial resistance ofM. haemolyticaisolated from clinically healthy sheep at slaughter and the genetic bacterial resistance mechanisms. Low levels of phenotypic resistance were detected for most of the antimicrobials tested except for tetracycline (4.3%) and tylosin (89.1%). A few antimicrobial resistance determinants were found in the genome of six out of nine isolates, consisting of genes conferring resistance to tetracyclines (tetH), aminoglycosides (strA), and sulfonamides (sul2), which were sometimes linked to the presence of plasmids but did not always lead to resistance phenotypes. Our results suggest that there is limited resistance inM. haemolyticastrains of veterinary origin, but the presence of several resistance genes, some of which were found in mobile genetic elements that play a major role in the dissemination of antimicrobial resistance in members of thePasteurellaceaefamily, deserves further consideration.AbstractMannheimia haemolyticais the main pathogen contributing to pneumonic pasteurellosis in sheep. The aim of this study was to investigate the antimicrobial resistance levels inM. haemolyticaisolates from the lungs of slaughtered sheep and to examine the genetic resistance mechanisms involved. A total of 256M. haemolyticaisolates, 169 from lungs with pneumonic lesions and 87 from lungs without lesions, were analyzed by the disk diffusion method for 12 antimicrobials, and the whole genome of 14 isolates was sequenced to identify antimicrobial resistance determinants. Levels of phenotypic resistance ranged from <2% for 10 antimicrobials (amoxicillin, amoxicillin-clavulanic, ceftiofur, cefquinome, lincomycin/spectinomycin, gentamicin, erythromycin, florfenicol, enrofloxacin, and doxycycline) to 4.3% for tetracycline and 89.1% for tylosin. Six isolates carriedtetHgenes and four isolates carried, in addition, thestrAandsul2genes in putative plasmid sequences. No mutations associated with macrolide resistance were identified in 23 rDNA sequences, suggesting that theM. haemolyticaphenotypic results for tylosin should be interpreted with care in the absence of well-established epidemiological and clinical breakpoints. The identification of strains phenotypically resistant to tetracycline and of several resistance genes, some of which were present in plasmids, highlights the need for continuous monitoring of susceptibility patterns inPasteurellaceaeisolates from livestock.