犬源大肠杆菌部分生物学特性研究及喹诺酮类耐药基因检测

发布时间：2018-02-03 01:38

本文关键词： 大肠杆菌犬源生物学特性系统发育组 PMQR基因　出处：《石河子大学》2017年硕士论文　论文类型：学位论文

【摘要】：为了解新疆石河子地区的犬源大肠杆菌质粒介导喹诺酮类耐药(PMQR)基因的流行现状与分布,无菌采集三家动物医院腹泻与健康宠物犬的肛拭子,分别进行细菌分离鉴定、生化鉴定、16SrDNA扩增,并对14种临床常用抗菌药物进行耐药性检测。同时利用PCR技术对系统发育组分群以及筛查犬源大肠杆菌喹诺酮类耐药基因。研究结果如下:1.对新疆石河子三家动物医院的75份腹泻与健康宠物犬的肛拭子样品进行分离鉴定与16SrDNA扩增,共获得63株大肠杆菌。其中来自腹泻犬有45株,健康犬18株,总分离率达84.0%。溶血性与致病性试验表明,具有β型溶血的分离株主要来自于腹泻犬的样品。大肠杆菌纯培养物经腹腔接种于小鼠后发现,呈β型溶血的分离株能使小鼠在50 h之内全部死亡。2.对63株犬源大肠杆菌使用多重PCR进行系统发育组分群,发现分离株主要分布在A组(28株,44.44%),其次是B2组(22株,34.92%)和D组(10株,15.87%),极少数在B1组(3株,4.76%)。同时分布在B2与D组的犬源大肠杆菌(32株,50.79%)几乎全部具有致病性。3.选择14种常用抗生素进行耐药性检测。结果表明,对四环素的耐药率最高,达85.71%(54/63);其次是林可霉素和恩诺沙星,耐药率分别为73.02%(46/63)和68.25%(43/63);分离株对氟苯尼考最敏感,仅占4.76%(3/63)。其它抗菌药物的耐药率分布在17.46%~65.10%。耐4种药物及以上的分离株占80.95%(51/63);对9种抗菌药物有耐药性的菌株数量最多,占44.44%(28/63);其次是耐6种药物的菌株占36.51%(23/63)。对14种药物均耐药的比例为3.17%(2/63),对14种药物均敏感的比例为4.76%(3/63)。4.对7种喹诺酮类耐药基因进行PCR检测,qnr基因检出率为36.51%(23/63)。其中qnr A基因检出率最多,占20.63%(13/63);其次是qnr S基因,检出率为11.11%(7/63);qnr C和qnr D基因的检出率分别为3.17%(2/63)和1.59%(1/63)。对aac(6’)-Ib基因的检出率为12.70%(8/63)。未检出qnr B基因和qeq A基因。其中含有2种基因的分离株有11株(17.46%);同时含有3种PMQR基因的分离株有1株,检出率占1.59%。本研究首次对新疆石河子地区犬源大肠杆菌的PMQR基因进行检测,扩展了质粒介导的犬源大肠杆菌喹诺酮类耐药基因的流行病学资料,为研究犬源大肠杆菌耐药性提供数据支持,同时也为兽医临床上正确选择喹诺酮类药物防治犬细菌性疾病提供科学依据。
[Abstract]:In order to understand the prevalence and distribution of PMQR gene mediated by plasmid in canine Escherichia coli (E.coli) in Shihezi area Xinjiang the anal swabs of diarrhea and healthy pet dogs in three animal hospitals were collected. Isolation and identification of bacteria, biochemical identification of 16s rDNA amplification. At the same time, PCR technique was used to detect phylogenetic groups and to screen canine Escherichia coli quinolones resistance genes. The results were as follows:. 1. 75 rectal swab samples of diarrhea and healthy pet dogs from three animal hospitals in Shihezi, Xinjiang were isolated and identified and 16s rDNA amplification was performed. A total of 63 strains of Escherichia coli were obtained, of which 45 were from diarrhea dogs and 18 from healthy dogs. The total isolation rate was 84.0. Hemolytic and pathogenicity tests showed. The isolated strains with beta hemolysis were mainly from the samples of diarrhea dogs. The pure culture of Escherichia coli was found in mice after intraperitoneal inoculation. 尾-hemolysis strain could make all mice die within 50 hours. 63 strains of Escherichia coli were divided by multiple PCR. It was found that the isolates were mainly distributed in group A (28 strains), followed by B 2 group (22 strains) and D group (10 strains), and a few in group B1 (3 strains). 4.76. 32 strains of Escherichia coli were distributed in group B2 and group D at the same time. The results showed that the resistance rate of tetracycline to tetracycline was the highest (85.71% 54 / 63). The resistance rates of lincomycin and enrofloxacin were 73.022 / 63 and 68.25 / 63 / 63, respectively. The isolates were most sensitive to florfenicol. The drug resistance rate of other antimicrobial agents was 17.46 and 65.103.The isolates with resistance to 4 or more drugs accounted for 80.955.51 / 63; The number of strains resistant to 9 antimicrobial agents was the most, accounting for 44.44% 28 / 63%; The next one was resistant to 6 drugs (36.51%). The proportion of drug resistance to 14 drugs was 3.17% (2 / 63). The rate of sensitivity to all 14 drugs was 4.76%. 7 quinolones resistant genes were detected by PCR. The detection rate of qnr gene was 36.51 / 23 / 63. The detection rate of qnr A gene was the highest (20.63%). The detection rate of qnr S gene was 11.11%. The detection rate of qnr C gene and qnr D gene were 3.17 and 1.590.The detection rate of aac(6')-Ib gene was 12.70% (P < 0.05). Qnr B gene and qeq A gene were not detected. At the same time, there was one isolate containing three PMQR genes, the detection rate was 1.59.The PMQR gene of canine Escherichia coli in Shihezi area of Xinjiang was detected for the first time in this study. The epidemiological data of canine Escherichia coli quinolones resistance genes mediated by plasmid were expanded to provide data support for the study of canine Escherichia coli resistance. It also provides scientific basis for the correct selection of quinolones in veterinary clinic for the prevention and treatment of canine bacterial diseases.
【学位授予单位】：石河子大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S852.61

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