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Genomics-based epidemiology and antimicrobial susceptibility of Mycoplasma bovis isolates from veal, dairy and beef herds
Bokma, J.; Vereecke, N.; Gille, L...
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Objectives: Mycoplasma bovis associated pneumonia is feared around the world, in particular because of its poor response to antimicrobial therapy. A rise in antimicrobial resistance of M. bovis is reported worldwide over the last two decades. Whether different, more resistant strains exist in industries such as the veal industry (more intensive antimicrobial use) compared to conventional dairy and beef farms, is currently unknown. Therefore, the objectives of this study were to compare strains originating from the veal, dairy and beef industry both on their genetic relatedness and antimicrobial susceptibility.
Materials and methods: MIC-values were determined for macrolides, tetracyclines, florfenicol, gentamicin, enrofloxacin and tiamulin with microbroth dilution on 144 epidemiologically independent Belgian M. bovis isolates (31 dairy, 70 beef, 11 dairy-beef mixed and 32 veal farms), mostly obtained from the respiratory tract. For a selection of 100 of these isolates (29 dairy, 41 beef, and 30 veal), the whole genome was sequenced by MinION Nanopore sequencing. The reference strain M. bovis PG45 was used as quality control in all experiments. Antimicrobial susceptibility data were analyzed using the epidemiological cut-off estimated by the visual eye-ball method to distinguish between wild type (WT) and non-wild type (nWT). Single Nucleotide Polymorphism (SNP) analysis was performed to type M. bovis strains, but also to pinpoint specific genetic markers in targeted genes, which were shown to associate with the observed phenotypic susceptibility results. Binary logistic regression (0: WT; 1: nWT) was performed on different sectors to compare antimicrobial susceptibility between sectors. In addition, a phylogenetic tree was developed using CSI Phylogeny (Center for Genomic Epidemiology) for SNP calling on consensus sequences to compare strains between sectors.
Results: Highest MIC-values were observed for macrolides, where almost all strains showed acquired resistance against 16-membered macrolides (tilmicosin and tylosin), and about 50% against the 15-membered macrolide, gamithromycin. A limited number of isolates showed acquired resistance against gentamicin, florfenicol, enrofloxacin and tiamulin. Almost all strains belonged to the wild type population for the tetracyclines (oxytetracycline and doxycycline). A remarkable difference between sectors was observed for gamithromycin, showing that beef herds (59% nWT) had a three times higher odds (95%CI: 1.23-7.35) for gamithromycin resistant M. bovis than dairy herds (32% nWT)(P = 0.02), whereas veal herds did not significantly differ from both sectors (47% nWT). The phylogenetic tree showed different clusters, although strains could not be associated with certain sectors. Specific genetic markers could be linked to acquired resistance for most strains.
Conclusions: This study shows that acquired resistance in Belgian M. bovis isolates is highest against macrolides, and minimal for tetracyclines. Secondly, no clear difference in acquired resistance (with the exception of gamithromycin) or strains between sectors were observed. This information could contribute to recommendations on antimicrobial therapy in case of M. bovis outbreaks and to further understanding of the epidemiology of this pathogen.
Keywords: Belgium, epidemiological cutoff, gamithromycin, tetracyclines.
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Affiliation of the authors at the time of publication
Department of Large Animal Internal Medicine, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium;
Department of Virology, Parasitology and Immunology, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium;
Clinical Department of Production Animals, Faculty of Veterinary Medicine, University of Liege, Liege, Belgium;
DGZ (Animal Health Service-Flanders), Torhout, Belgium;
Department of Pathology, Bacteriology and Avian diseases, Faculty of Veterinary Medicine, Merelbeke, Belgium.
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