Bovine mastitis is the most common infectious disease in dairy cattle with major economic implications for the dairy industry worldwide. Continuous monitoring for the emergence of antimicrobial resistance (AMR) among bacterial isolates from dairy farms is vital not only for animal husbandry but also for public health. In this study, the prevalence of AMR in 113 Escherichia coli isolates from cases of bovine clinical mastitis in Canada was investigated. Kirby-Bauer disk diffusion test with 18 antibiotics and microdilution method with 3 heavy metals (copper, zinc, and silver) was performed to determine the antibiotic and heavy-metal susceptibility. Resistant strains were assessed for efflux and ß-lactamase activities besides assessing biofilm formation and hemolysis. Whole-genome sequences for each of the isolates were examined to detect the presence of genes corresponding to the observed AMR and virulence factors. Phenotypic analysis revealed that 32 isolates were resistant to one or more antibiotics and 107 showed resistance against at least one heavy metal. Quinolones and silver were the most efficient against the tested isolates. Among the AMR isolates, AcrAB-TolC efflux activity and ß-lactamase enzyme activities were detected in 13 and 14 isolates, respectively. All isolates produced biofilm but with different capacities, and 33 isolates showed α-hemolysin activity. A positive correlation (Pearson r = + 0.89) between efflux pump activity and quantity of biofilm was observed. Genes associated with aggregation, adhesion, cyclic di-GMP, quorum sensing were detected in the AMR isolates corroborating phenotype observations. This investigation showed the prevalence of AMR in E. coli isolates from bovine clinical mastitis. The results also suggest the inadequacy of antimicrobials with a single mode of action to curtail AMR bacteria with multiple mechanisms of resistance and virulence factors. Therefore, it calls for combinatorial therapy for the effective management of AMR infections in dairy farms and combats its potential transmission to the food supply chain through the milk and dairy products.

中文翻译：

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加拿大乳腺炎奶牛大肠埃希菌抗生素耐药性的流行情况和机制

牛乳腺炎是奶牛中最常见的传染病，对全球奶业具有重大的经济影响。持续监测来自奶牛场的细菌分离株中抗菌素耐药性 (AMR) 的出现，不仅对畜牧业而且对公众健康都至关重要。在这项研究中，调查了来自加拿大牛临床乳腺炎病例的 113 株大肠杆菌中 AMR 的流行情况。使用 18 种抗生素进行 Kirby-Bauer 圆盘扩散试验，并使用 3 种重金属（铜、锌和银）进行微量稀释，以确定抗生素和重金属敏感性。除了评估生物膜形成和溶血外，还评估了耐药菌株的外排和 ß-内酰胺酶活性。检查每个分离株的全基因组序列以检测与观察到的 AMR 和毒力因子相对应的基因的存在。表型分析显示，32 株分离株对一种或多种抗生素具有抗性，107 株对至少一种重金属具有抗性。喹诺酮类和银对测试的分离株最有效。在 AMR 分离株中，分别在 13 和 14 株分离物中检测到 AcrAB-TolC 外排活性和 ß-内酰胺酶活性。所有分离株都产生了生物膜，但能力不同，33 个分离株显示出 α-溶血素活性。观察到外排泵活性和生物膜数量之间呈正相关（Pearson r = + 0.89）。与聚集、粘附、环状双 GMP 相关的基因，在 AMR 分离株中检测到群体感应，证实了表型观察结果。该调查显示 AMR 在牛临床乳腺炎的大肠杆菌分离株中的流行。结果还表明，具有单一作用方式的抗菌剂不足以抑制具有多种耐药机制和毒力因子的 AMR 细菌。因此，它呼吁采用组合疗法来有效管理奶牛场中的 AMR 感染，并打击其通过牛奶和乳制品向食品供应链的潜在传播。