Identification of genetic markers of resistance to macrolide class antibiotics in Mannheimia haemolytica isolates from a Saskatchewan feedlot.

Mannheimia haemolytica is a major contributor to bovine respiratory disease (BRD), which causes substantial economic losses to the beef industry, and there is an urgent need for rapid and accurate diagnostic tests to provide evidence for treatment decisions and support antimicrobial stewardship. Diagnostic sequencing can provide information about antimicrobial resistance genes in M. haemolytica more rapidly than conventional diagnostics. Realizing the full potential of diagnostic sequencing requires a comprehensive understanding of the genetic markers of antimicrobial resistance. We identified genetic markers of resistance in M. haemolytica to macrolide class antibiotics commonly used for control of BRD. Genome sequences were determined for 99 M. haemolytica isolates with six different susceptibility phenotypes collected over 2 years from a feedlot in Saskatchewan, Canada. Known macrolide resistance genes estT, msr(E), and mph(E) were identified in most resistant isolates within predicted integrative and conjugative elements (ICEs). ICE sequences lacking antibiotic resistance genes were detected in 10 of 47 susceptible isolates. No resistance-associated polymorphisms were detected in ribosomal RNA genes, although previously unreported mutations in the L22 and L23 ribosomal proteins were identified in 12 and 27 resistant isolates, respectively. Pangenome analysis led to the identification of 79 genes associated with resistance to gamithromycin, of which 95% (75 of 79) had no functional annotation. Most of the observed phenotypic resistance was explained by previously identified antibiotic resistance genes, although resistance to the macrolides gamithromycin and tulathromycin was not explained in 39 of 47 isolates, demonstrating the need for continued surveillance for novel determinants of macrolide resistance.IMPORTANCEBovine respiratory disease is the costliest disease of beef cattle in North America and the most common reason for injectable antibiotic use in beef cattle. Metagenomic sequencing offers the potential to make economically significant reductions in turnaround time for diagnostic information for evidence-based selection of antibiotics for use in the feedlot. The success of diagnostic sequencing depends on a comprehensive catalog of antimicrobial resistance genes and other genome features associated with reduced susceptibility. We analyzed the genome sequences of isolates of Mannheimia haemolytica, a major bovine respiratory disease pathogen, and identified both previously known and novel genes associated with reduced susceptibility to macrolide class antimicrobials. These findings reinforce the need for ongoing surveillance for markers of antimicrobial resistance to support improved diagnostics and antimicrobial stewardship.

Changes in the phenotypic susceptibility of Mannheimia haemolytica isolates to macrolide antimicrobials during the early feeding period following metaphylactic tulathromycin use in western Canadian feedlot calves.

Cattle at high-risk for bovine respiratory disease on entry to western Canadian feedlots are often treated metaphylactically with antimicrobials from the macrolide class. High levels of resistance to macrolides have been reported in Mannheimia haemolytica isolates from clinical samples, but it is less clear whether this trend extends to the broader feedlot population. The objective was to describe near-term [< 40 days on feed (DOF)] changes in the recovery and susceptibility of M. haemolytica isolates from healthy feedlot calves after metaphylactic exposure to tulathromycin. Eight cohorts of 100 calves (n = 800) were sampled via deep nasopharyngeal swab at entry processing (i.e., before metaphylaxis, at 1 DOF) and again at 13 DOF. Ten calves from each cohort (n = 80) were randomly sampled a third time at 36 DOF. Recovery of M. haemolytica isolates across all cohorts increased over the study period, from 33% (95% CI: 26.5 to 40.2%) at 1 DOF to 75% (95% CI: 71.4 to 78.3%) at 36 DOF. A significant shift in the minimum inhibitory concentration (MIC) distribution of tulathromycin from 1 DOF (MIC90 ≤ 8 µg/mL) to 13 DOF (MIC90 > 64 µg/mL) was observed. A subset of 36 isolates from 13 DOF screened for macrolide resistance genes via multiplex polymerase chain reaction all harbored the msrE and mphE genes. Recovery of M. haemolytica at 13 and 36 DOF did not decline in response to metaphylactic use of tulathromycin; conversely, we inferred the potential for rapid inter-pen spread of a macrolide-resistant clone by 13 DOF in 6 of 8 pens under selective pressure from antimicrobial use.

Changements dans la sensibilité phénotypique des isolats de Mannheimia haemolytica aux a ntibiotiques de la classe des macrolides au début de la période d'alimentation après l'utilisation m étaphylactique de tulathromycine chez les veaux des parcs d'engraissement de l'Ouest canadien. Les bovins à risque élevé de maladies respiratoires bovines à leur entrée dans les parcs d'engraissement de l'Ouest canadien sont souvent traités métaphylactiquement avec des antimicrobiens de la classe des macrolides. Des taux élevés de résistance aux macrolides ont été signalés chez les isolats de Mannheimia haemolytica provenant d'échantillons cliniques, mais il est moins clair si cette tendance s'étend à la population plus large des parcs d'engraissement. L'objectif était de décrire les changements à court terme [< 40 jours d'alimentation (DOF)] dans la récupération et la sensibilité des isolats de M. haemolytica provenant de veaux sains en parc d'engraissement après une exposition métaphylactique à la tulathromycine. Huit cohortes de 100 veaux (n = 800) ont été échantillonnées via un prélèvement nasopharyngé profond lors du traitement d'entrée (i.e., avant la métaphylaxie, à 1 DOF) et à nouveau à 13 DOF. Dix veaux de chaque cohorte (n = 80) ont été échantillonnés au hasard une troisième fois à 36 DOF. La récupération des isolats de M. haemolytica dans toutes les cohortes a augmenté au cours de la période d'étude, passant de 33 % (IC 95 % : 26,5 à 40,2 %) à 1 DOF à 75 % (IC 95 % : 71,4 à 78,3 %) à 36 DOF. Un changement significatif dans la distribution de la concentration minimale inhibitrice (MIC) de la tulathromycine de 1 DOF (MIC90 ≤ 8 µg/mL) à 13 DOF (MIC90 > 64 µg/mL) a été observé. Un sous-ensemble de 36 isolats de 13 DOF criblés pour les gènes de résistance aux macrolides via une réaction d'amplification en chaîne par polymérase multiplex hébergeaient tous les gènes msrE et mphE. L'isolement de M. haemolytica à 13 et 36 DOF n'a pas diminué en réponse à l'utilisation métaphylactique de la tulathromycine; à l'inverse, nous avons suggéré le potentiel de propagation rapide entre les enclos d'un clone résistant aux macrolides par 13 DOF dans 6 des 8 enclos sous la pression sélective de l'utilisation d'antimicrobiens.(Traduit par Dr Serge Messier).

Complete genome sequence of Mannheimia bovis isolated from a healthy feedlot calf in Saskatchewan, Canada.

A lack of whole genome sequences for Mannheimia spp. other than Mannheimia haemolytica complicates their identification. Here, we present the genome sequence of Mannheimia bovis 39324.S-11, isolated from a healthy calf on a feedlot in Saskatchewan, Canada, and compare it to ZY190616T, which is currently the only other isolate of M. bovis for which sequence is publicly available.