山东某地散养猪源大肠杆菌抗药性研究

Studies on Drug Resistance of Escherichia coli from Pigs Raise in Villages in Shandong

为调查养猪户各个生产环境中存在的多黏菌素抗药性情况。从山东某地规模较小的商品猪场及其周边散养猪户分别采集饲料、饮水、粪便、污水、猪鼻拭子等样品200个,分别制作含有药物(环丙沙星2μg/mL,黏杆菌素4μg/mL)的琼脂培养基,分离纯化细菌,并筛选抗药性菌株。将每户或商品猪场的样品合并为一个样品,用mcr-1基因和ndm-1基因扩增引物,进行PCR扩增。参照美国临床实验室标准化协会(CLSI)药敏试验标准,采用琼脂平板法和微量肉汤稀释法,测定分离菌株对强力霉素等8种抗生素的抗药性水平。结果显示,200个样本中分离到大肠杆菌50株,分离率为25%。从62个混合样品中检测到38个样品含有mcr-1基因(阳性率61.29%),6个样品含有ndm-1基因(阳性率9.68%)。细菌药敏试验结果显示,含有mcr-1基因和ndm-1基因的菌株对这8种抗生素的抗药率均较高。结果表明在猪体及其养殖环境中存在mcr-1和ndm-1这2种基因,散养猪模式和规模化模式导致的细菌抗药类型和水平存在差异。

To investigate the resistance of polymyxins in various aspects of pig production,samples of feed,drinking water,feces,sewage,nasal swabs from pigs were collected from one small scale commercial pig farm surrounding free-range raising pig farms and in several counties of Shandong province.MHB liquid medium and MHA solid medium were used respectively to prepare drug containing agar medium(ciprofloxacin 2μg/mL,colistin 4μg/mL),and from which bacteria were isolated.After isolation and purification,resistant strains were obtained.Samples from each pig farm were combined into a big one for detection,in which primers amplified by mcr-1 gene and ndm-1 gene were used for PCR amplification.8 antibiotics,including doxycycline,were tested by agar plate method and micro-broth dilution method in accordance with the American Clinical Laboratory Standardization Institute(CLSI)antimicrobial susceptibility test standard.The results showed that 50 strains of E.coli were isolated from 200 samples,and the isolation rate was 25%.In the 62 mixed samples,38 samples contained mcr-1 gene(positive rate was 61.29%),and 6 samples contained the ndm-1 gene(positive rate was 9.68%).According to antimicrobial susceptibility testing,bacteria containing the mcr-1 gene and the ndm-1 gene were more resistant to the eight antibiotics.The above results indicated that the discussed two genes existed both in the pig body and in their breeding environ ment,and differences in types and levels of bacterial resistance caused by free-range pig models and scale models existed.