Overuse of antibiotics in livestock and poultry feedlot has caused broad antibiotics resistance of microbes. Research on antibiotics-resistant microbes is helpful to reveal the status of antibiotics resistance and the transmission mechanism of antibiotics-resistant genes in the environment. In this study, samples were collected from sewage water near livestock and poultry feedlot, the abundance of both antibiotics-resistant bacteria and antibiotics-resistant Enterobacteria was analyzed, in the meantime, strain R11 with high polymyxin E resistance was isolated and studied. Firstly, 16S rDNA of strain R11was sequenced and antibiotics-resistance was analyzed, plasmids were extracted and digested with restriction enzyme, and then PFGE was used to detect the endogenous plasmids. Results showed that polymyxin E-resistant bacteria accounted for 0.14%~0.35% of all bacteria and polymyxin E-resistant Enterobacteria accounted for 1.50%~7.40% of all Enterobacteria. In addition, Enterobacteria occupied 1.13%~2.45% of all the bacteria, which showed that Enterobacteria may be the vital bacteria with polymyxin E resistance, and also occupied large amounts of bacteria in sewage water near livestock and poultry feedlot. Strain R11 was identified asEnterobacter cloacae by 16S rDNA sequence analysis. Strain R11 has multi-antibiotics resistance, which was only sensitive to lincomycin. The MICs of amikacin, tetracycline and ciprofloxacin of strain R11 were 10μg·mL-1, while the MICs of amoxicillin, sulfamethoxazole, ampicillin, streptomycin, polymyxin E was25 μg·mL-1, 32 μg·mL-1, 40 μg·mL-1, 90 μg·mL-1and 96 μg·mL-1, respectively. In addition, strain R11 carried several endogenous plasmids. All the results showed that strain R11 had a high level of polymyxin E resistance, and it was useful to study the mechanism of polymyxin E resistance in microbes.%畜禽养殖业大量使用抗生素引起了广泛的微生物耐药性问题,研究抗生素耐药菌株可以了解环境中细菌的耐药性情况和抗性基因的传播机制.采集畜禽养殖场周边的污水,分析了污水中多粘菌素E耐药性细菌和耐药性肠杆菌的丰度;同时以一株具有较高多粘菌素E耐药水平的肠杆菌R11为例,首先进行16S rDNA测序和抗生素耐药谱分析,然后通过质粒提取、酶切电泳以及脉冲场凝胶电泳(PFGE),分析其内源性质粒等情况.结果表明:在畜禽养殖场周边污水环境中,多粘菌素E耐药性细菌占细菌总数的0.14%~0.35%,多粘菌素E耐药性肠杆菌占肠杆菌总数的1.50%~7.40%,而肠杆菌占细菌总数的1.13%~2.45%,说明肠杆菌可能是携带多粘菌素E抗性基因的主要细菌种类,同时也是养殖业污水中细菌的主要组成部分.16S rDNA序列分析确定R11是一株Enterobacter cloacae,抗生素耐药谱分析R11具有多重耐药性,除对林可霉素敏感外,阿米卡星、四环素和环丙沙星的最小抑菌浓度(MICs)为10μg·mL-1,阿莫西林的MICs为25μg·mL-1,磺胺甲恶唑的MICs为32μg·mL-1,氨苄西林的MICs为40μg·mL-1,链霉素的MICs为90μg·mL-1,而多粘菌素E的MICs最高,达到96μg·mL-1.质粒提取酶切和PFGE分离鉴定结果显示,R11菌株可能有多个内源性质粒.综上所述,R11菌株具有较高水平的多粘菌素E耐药性,并且具有多重耐药性和多个内源性质粒,对于研究多粘菌素E耐药性机制具有一定的参考价值.
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