Bovine respiratory disease continues to be the most important ailment of feed yard cattle. While the disease is multifactorial in nature, therapy continues to target the primary bacterial pathogens, Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni. A survey of records from a single diagnostic laboratory was conducted to evaluate the percentage of M. haemolytica isolates that were resistant to multiple antimicrobials and if coresistance patterns could be detected. All susceptibility test results for M. haemolytica recovered from lung tissues of cattle were eligible for inclusion in the survey. There were no isolates over the course of the analysis that were resistant to all 6 antimicrobials, primarily due to a lack of resistance to ceftiofur. In 2009, just over 5% of isolates were resistant to 5 or more antimicrobials (pan-resistant). In 2011, more than 35% of the M. haemolytica isolates were characterized as pan-resistant. Significant antimicrobial coresistance patterns were only seen with oxytetracycline and tilmicosin; bacterial isolates that were resistant to either oxytetracycline or tilmicosin were more likely to be resistant to at least one other antimicrobial. The mechanisms by which M. haemolytica is developing multidrug resistance warrant investigation if antimicrobial utility in the therapy of bovine respiratory disease is to be preserved.

Mannheimia haemolytica typically resides in cattle as a commensal member of the upper respiratory tract microbiome. However, some strains can invade their lungs and cause respiratory disease and death, including those with multi-drug resistance. A nucleotide polymorphism typing system was developed for M. haemolytica from the genome sequences of 1133 North American isolates, and used to identify genetic differences between isolates from the lungs and upper respiratory tract of cattle with and without clinical signs of respiratory disease. A total of 26,081 nucleotide polymorphisms were characterized after quality control filtering of 48,403 putative polymorphisms. Phylogenetic analyses of nucleotide polymorphism genotypes split M. haemolytica into two major genotypes (1 and 2) that each were further divided into multiple subtypes. Multiple polymorphisms were identified with alleles that tagged genotypes 1 or 2, and their respective subtypes. Only genotype 2 M. haemolytica associated with the lungs of diseased cattle and the sequence of a particular integrative and conjugative element (ICE). Additionally, isolates belonging to one subtype of genotype 2 (2b), had the majority of antibiotic resistance genes detected in this study, which were assorted into seven combinations that ranged from 1 to 12 resistance genes. Typing of diverse M. haemolytica by nucleotide polymorphism genotypes successfully identified associations with diseased cattle lungs, ICE sequence, and antibiotic resistance genes. Management of cattle by their carriage of M. haemolytica could be an effective intervention strategy to reduce the prevalence of respiratory disease and supplemental needs for antibiotic treatments in North American herds.

Antimicrobial resistance (AMR) of bacterial pathogens is an emerging public health threat. This threat extends to pets as it also compromises our ability to treat their infections. Surveillance programs in the United States have traditionally focused on collecting data from food animals, foods, and people. The Veterinary Laboratory Investigation and Response Network (Vet-LIRN), a national network of 45 veterinary diagnostic laboratories, tested the antimicrobial susceptibility of clinically relevant bacterial isolates from animals, with companion animal species represented for the first time in a monitoring program. During 2017, we systematically collected and tested 1968 isolates. To identify genetic determinants associated with AMR and the potential genetic relatedness of animal and human strains, whole genome sequencing (WGS) was performed on 192 isolates: 69 Salmonella enterica (all animal sources), 63 Escherichia coli (dogs), and 60 Staphylococcus pseudintermedius (dogs). We found that most Salmonella isolates (46/69, 67%) had no known resistance genes. Several isolates from both food and companion animals, however, showed genetic relatedness to isolates from humans. For pathogenic E. coli, no resistance genes were identified in 60% (38/63) of the isolates. Diverse resistance patterns were observed, and one of the isolates had predicted resistance to fluoroquinolones and cephalosporins, important antibiotics in human and veterinary medicine. For S. pseudintermedius, we observed a bimodal distribution of resistance genes, with some isolates having a diverse array of resistance mechanisms, including the mecA gene (19/60, 32%). The findings from this study highlight the critical importance of veterinary diagnostic laboratory data as part of any national antimicrobial resistance surveillance program. The finding of some highly resistant bacteria from companion animals, and the observation of isolates related to those isolated from humans demonstrates the public health significance of incorporating companion animal data into surveillance systems. Vet-LIRN will continue to build the infrastructure to collect the data necessary to perform surveillance of resistant bacteria as part of fulfilling its mission to advance human and animal health. A One Health approach to AMR surveillance programs is crucial and must include data from humans, animals, and environmental sources to be effective.

Infectious bovine keratoconjunctivitis (IBK), also known as pinkeye, is one of the most common eye diseases in cattle. Several pathogens have been associated with IBK cases, however, Moraxella bovis, Moraxella bovoculi, Mycoplasma bovis, Mycoplasma bovoculi and bovine herpesvirus type 1 (BHV-1) are most frequently observed. A multiplex real-time PCR assay using two reactions was developed for the detection and differentiation of these five pathogens. Detection sensitivities of the multiplex assays were compared to singleplex reactions testing for the same targets. Correlation coefficients (R²) of >0.99, and PCR efficiencies between 92 and 106% were demonstrated in all singleplex and multiplex real-time PCR reactions. The limits of detection (LOD) of multiplex assays for Moraxella bovis, Moraxella bovoculi, Mycoplasma bovis, Mycoplasma bovoculi and BHV-1 were 19, 23, 25, 24 and 26 copies per reaction, respectively. No cross amplification was observed for specificity testing of 179 IBK positive clinical samples and 55 non-target clinical samples. Percentage of clinical samples positive for Mycoplasma bovoculi, Moraxella bovoculi, Moraxella bovis, BHV-1 and Mycoplasma bovis were 88.8% (159/179), 75.9% (136/179), 60.3% (108/179), 11.7% (21/179) and 10.0% (18/179), respectively. Moraxella bovis, Moraxella bovoculi and Mycoplasma bovoculi were more prevalent than Mycoplasma bovis and BHV-1 in IBK samples collected from animals in this study population. Our data indicates that the multiplex real-time PCR panel assay is highly sensitive and highly specific for the detection and differentiation of the five major pathogens associated with bovine pinkeye.