奶牛临床乳房炎源大肠杆菌主要毒力基因的克隆与序列分析

发布时间：2018-03-21 18:08

本文选题：奶牛临床乳房炎　切入点：大肠杆菌　出处：《宁夏大学》2015年硕士论文　论文类型：学位论文

【摘要】：为了研究宁夏地区奶牛临床乳房炎大肠杆菌分离株血清型分布和主要毒力基因,采集2013-2014年宁夏地区临床乳房炎病牛奶样作为大肠杆菌检测的样品,进行分离培养,采用细菌生化试验进行鉴定。使用大肠杆菌0抗原诊断血清对分离自奶牛临床乳房炎的120株大肠杆菌分离株进行血清学检测,并利用PCR方法扩增定居因子基因fimH、f17、f165、f41、 f5、eae、cs31a和转运蛋白trat基因。采集2013-2014年宁夏地区临床乳房炎病牛奶样作为大肠杆菌检测的样品,进行分离培养,采用细菌生化试验进行行鉴定,120株鉴定为大肠杆菌。大肠杆菌O抗原鉴定结果：经鉴定120株奶牛临床乳房炎大肠杆菌分离株中有71株检测出血清型,鉴定出08,010,015,018,022,O 26,053,055,068,086,091,092,093,0101,0107,0117,0126,0142,0148,0153,0158共21种血清型。优势血清型为0158,0101,0126,091,015,026,0107,占定型株的61.9%,49株未鉴定出血清型。选择trat和7种定居因子fimH、f17、f165、f41、f5、eae、cs31a进行检测。实验结果显示,120株大肠杆菌PCR扩增结果为trat基因分离率为24.2%,fimH基因分离率为85.8%,f17基因分离率为5%,f165基因分离率为3.3%,f41基因分离率为0,f5基因分离率,0,eae基因分离率为0,cs31a基因分离率0。实验结果表明肠道内感染和临床乳房炎大肠杆菌分离株血清型、毒力基因分离率不同。本实验选取奶牛临床乳房炎大肠杆菌分离株(0158)为样品,设计并合成引物,对fimH基因进行克隆和序列测定.测序结果显示fimh基因在第714碱基位点位点由T突变为A,在第717个碱基位点位点由A突变为G,第807个碱基位点位点由G突变为A,第831个碱基位点位点由T突变为C。
[Abstract]:In order to study the distribution of serotype and the main virulence genes of clinical mastitis Escherichia coli isolated from dairy cattle in Ningxia, samples of clinical mastitis in Ningxia from 2013 to 2014 were collected as samples for detection of Escherichia coli, and were isolated and cultured. The serological tests of 120 strains of Escherichia coli isolated from clinical mastitis of dairy cattle were carried out by using the diagnostic serum of Escherichia coli 0 antigen. The PCR method was used to amplify the gene fimHf17f165f41, f5eaeae cs31a and transporter trat. The samples of clinical mastitis in Ningxia from 2013 to 2014 were collected as samples for detection of Escherichia coli, and were isolated and cultured. A total of 120 strains of Escherichia coli were identified by bacterial biochemistry test. Results of the identification of Escherichia coli O antigen: 71 of 120 strains of clinical mastitis Escherichia coli isolated from dairy cattle were identified as serotype. 21 serotypes were identified from 080100 150 18022 O260530550680880860910920310101n010701177, 012614801480153150158. The dominant serotype was 0158O1010101260910260107, which accounted for 61.9107.The trat and the seven settlement factor fimHf17f165f41f41a were selected for detection. The results showed that the PCR amplification of 120 strains was trat based Escherichia coli. Because the isolation rate of fimH gene was 24.2and the isolation rate of f17 gene was 85.8.The isolation rate of 5f165 gene was 3.3f41 gene and the isolation rate of f41 gene was zero f5 gene. The isolation rate of 0eae gene was 0 cs31a gene. The results showed that intestinal tract infection and clinical mastitis were caused by intestinal infection and clinical mastitis. Serotype of Escherichia coli isolates, The isolation rate of virulence gene was different. In this experiment, primers were designed and synthesized by using clinical mastitis Escherichia coli strain 0158 as the sample. FimH gene was cloned and sequenced. The results of sequencing showed that fimh gene mutated from T to A at 714 base site, from A to G at 717 base site, from G to A at 807 base site, and from G to A at 714 base site. 831 base loci were mutated from T to C.
【学位授予单位】：宁夏大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S852.61

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