抗菌药物减量对规模奶牛场抗生素抗性基因多样性的影响

Effect of antimicrobial reduction on the diversity of antibiotic resistance genes in large scale dairy farms

为研究抗菌药物减量使用对河北省规模奶牛场大肠杆菌耐药性的影响及奶牛肠道菌群抗生素抗性基因(antibiotic resistance genes,ARGs)和可移动遗传元件(mobile genetic elements,MGEs,如整合子和转座子等)多样性的影响,评估抗菌药物减量化行动的实施效果,自河北省不同地区的3个达标奶牛场和2个非达标奶牛场采集肛拭子样品和粪便样品,肛拭子样品中分离出大肠杆菌并检测其对16种抗菌药物的耐药性,分析两类奶牛场大肠杆菌耐药率的差异性。采取SYBR荧光染料法高通量实时荧光定量PCR技术检测奶牛粪便中的ARGs和MGEs,计算其相对丰度,分析两类奶牛场之间ARGs和MGEs的差异性以及两者之间的相关性。结果显示,两类奶牛场来源的大肠杆菌耐药率差异性较小,非达标场更易产生多重耐药菌株。15个样品中共检出192个ARGs和7个MGEs;ARGs相对丰度最高的为β-内酰胺类抗性基因,其次为四环素类抗性基因。部分达标奶牛场检出的ARGs和MGEs低于非达标场,揭示抗菌药物的大量使用可能与ARGs的产生与传播有关;MGEs检出数量与ARGs检出数量具有相关性,ARGs可能通过水平基因转移进行传播。结果表明,实施抗菌药物减量使用对抗生素耐药基因的多样性具有一定程度的影响,但在短期内对奶牛场的大肠杆菌耐药率和ARGs多样性影响不显著,不同地区之间差异性不大,这也揭示了奶牛场抗生素耐药性的普遍性。

In order to study the effect of antimicrobial drug reduction on the resistance of E.coli and diversity of antibiotic resistance genes(ARGs)and mobile genetic elements(MGEs)in the intestinal microbiota of dairy cows in dairy farms in Hebei Province,and to evaluate the implementation effect of antimicrobial drug reduction actions,anal swab samples and fecal samples were collected from three qualified dairy farms and two non-standard dairy farms in different regions of Hebei Province,and E.coli was isolated from the anal swab samples and tested for their resistance to 16antimicrobial drugs,and the differences in the drug resistance rate of E.coli in the two types of dairy farms were analyzed.SYBR fluorescent dye method was used to detect ARGs and MGEs in dairy feces,their relative abundance was calculated,the differences and correlation between the two types of dairy farms were analyzed.The results showed that a total of 192 ARGs and 7 MGEs were detected in 15 samples,and the relative abundance of ARGs was the highest inβ-lactam resistance genes,followed by tetracycline resistance genes.The detection of ARGs and MGEs in some standard dairy farms was lower than that in non-standard farms,suggesting that the large use of antimicrobial drugs may be related to the production and transmission of ARGs,and the number of MGEs detected was correlated with the number of ARGs detected,and ARGs may be transmitted through horizontal gene transfer.The results showed that there was little difference in the drug resistance rate of E.coli from the two types of dairy farms,and multi-drug resistant strains were more likely to occur in non-standard farms.The implementation of antimicrobial reduction had a certain effect on the diversity of antibiotic resistance genes,but had no significant effect on the prevalence of E.coli drug resistance and the diversity of ARGs in dairy farms in the short term,and there was little difference between different regions,which also revealed the prevalence of antibiotic resistance in dairy farms.