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Abstract
Bovine respiratory disease (BRD) is the largest health threat to the beef cattle industry. Many pathogens play a role in the pathogenesis of BRD, however, Mannhemia haemolytica (Mh) is the pathogen largely responsible for the pathology we observe in cases of BRD. Recent reports furthermore indicate that antimicrobial resistance is becoming more commonly encountered with this pathogen.
The objectives of the first study were to determine the genetic relationships between isolates collected at arrival and revaccination, to identify resistance genes present in these isolates, and to determine if they were carried on an integrative conjugative element (ICE). Twenty calves culture positive for Mh at arrival and revaccination were identified, and a total of 48 isolates were submitted for whole-genome sequencing (WGS). A phylogenetic tree was constructed, showing the arrival isolates falling into four clades, and all revaccination isolates within one clade. All revaccination isolates were positive for an ICE. Three different ICEs with resistance gene modules were identified.
The objective of the second study was to compare the ability of pulsed field gel electrophoresis (PFGE) and WGS to differentiate 48 Mh isolates. The tree produced by PFGE placed the isolates into 11 clusters, whereas the tree produced by WGS placed the isolates into 18 clusters. Multiple isolates that had been grouped together by WGS were not grouped together by PFGE. These results suggest that PFGE is less sensitive than WGS, and the two methodologies might not classify Mh identically.
My final objective was to assess concordance between different methods of assessing antimicrobial susceptibility. Concordance between disk diffusion and broth microdilution, between disk diffusion and WGS, and between broth microdilution and WGS were assessed. Total concordance between disk diffusion and broth microdilution for all comparisons was 90.8%, for disk diffusion and WGS was 94.5%, and for broth microdilution and WGS, was 92.8%. For individual antimicrobials, concordance between methods ranged from 60.4% to 100%, depending on the antimicrobial and testing methods evaluated. Very major errors were identified in 0 to 6.3% of classifications while minor errors were seen in up to 33% of classifications depending on the antimicrobial and testing method evaluated.
The objectives of the first study were to determine the genetic relationships between isolates collected at arrival and revaccination, to identify resistance genes present in these isolates, and to determine if they were carried on an integrative conjugative element (ICE). Twenty calves culture positive for Mh at arrival and revaccination were identified, and a total of 48 isolates were submitted for whole-genome sequencing (WGS). A phylogenetic tree was constructed, showing the arrival isolates falling into four clades, and all revaccination isolates within one clade. All revaccination isolates were positive for an ICE. Three different ICEs with resistance gene modules were identified.
The objective of the second study was to compare the ability of pulsed field gel electrophoresis (PFGE) and WGS to differentiate 48 Mh isolates. The tree produced by PFGE placed the isolates into 11 clusters, whereas the tree produced by WGS placed the isolates into 18 clusters. Multiple isolates that had been grouped together by WGS were not grouped together by PFGE. These results suggest that PFGE is less sensitive than WGS, and the two methodologies might not classify Mh identically.
My final objective was to assess concordance between different methods of assessing antimicrobial susceptibility. Concordance between disk diffusion and broth microdilution, between disk diffusion and WGS, and between broth microdilution and WGS were assessed. Total concordance between disk diffusion and broth microdilution for all comparisons was 90.8%, for disk diffusion and WGS was 94.5%, and for broth microdilution and WGS, was 92.8%. For individual antimicrobials, concordance between methods ranged from 60.4% to 100%, depending on the antimicrobial and testing methods evaluated. Very major errors were identified in 0 to 6.3% of classifications while minor errors were seen in up to 33% of classifications depending on the antimicrobial and testing method evaluated.