某规模化猪场猪流行性腹泻病毒检测及主要毒力基因克隆分析

发布时间：2018-08-02 10:12

【摘要】：猪流行性腹泻(Porcine epidemic diarrhea,PED)是由猪流行性腹泻病毒(Porcine epidemic diarrhea virus,PEDV)引起的以呕吐、腹泻和脱水为主要特征的猪急性接触性肠道传染病,对新生仔猪的致死率可达100%。该病在世界范围内频发,给养猪业造成了巨大的经济损失。本研究以2014年11~12月份在陕西渭南某规模化养猪场发生的疑似PEDV感染仔猪为研究对象,经临床症状观察和病理剖检,初步诊断为猪流行性腹泻;通过对20份小肠病料进行PEDV M基因的RT-PCR检测,检测出了9份猪流行性腹泻病毒核酸,PEDV检测阳性率为45.0%。参考Gen Bank中CV777经典毒株全基因组序列(AF353511.1),设计了扩增PEDV ORF3基因、M基因和S基因部分序列的引物,对相应基因进行了克隆和序列分析,通过与CV777株对比分析,发现分离株的M基因具有高度保守性,可以作为检测PEDV的靶基因;WN毒株与经典毒株CV777相比,ORF3基因共存在11处碱基突变,这些碱基突变是否导致病毒毒力的改变还有待研究。S基因变异较大,S蛋白内分的核心中和表位(CEO,499-638aa)共有9个氨基酸位点发生了突变共有9个位点氨基酸发生了突变这些突变,S基因的突变可能是导致PEDV疫苗免疫效果下降的一个原因。
[Abstract]:Porcine epidemic diarrhea (Porcine epidemic) is an acute contact infectious disease caused by porcine epidemic diarrhea virus (Porcine epidemic diarrhea virus), which is characterized by vomiting, diarrhea and dehydration. The disease occurs frequently all over the world, causing huge economic loss to pig industry. In this study, suspected PEDV infected piglets in Weinan Pig Farm in Weinan, Shaanxi Province, in November and December 2014, were selected as the research objects. After clinical symptom observation and pathological dissection, it was preliminarily diagnosed as epidemic diarrhea in pigs. By RT-PCR detection of PEDV M gene in 20 samples of small intestine disease, 9 porcine epidemic diarrhea virus nucleic acid (PEDV) positive rate was 45.0%. Referring to the whole genome sequence (AF353511.1) of the classical CV777 strain in Gen Bank, primers were designed to amplify the M gene and the S gene sequence of the PEDV ORF3 gene. The corresponding genes were cloned and sequenced. The results were compared with those of the CV777 strain. It was found that the M gene of the isolated strain was highly conserved and could be used as a target gene for the detection of PEDV. Compared with the classical strain CV777, there were 11 base mutations in the ORF3 gene. Whether these base mutations lead to the virulence of the virus remains to be studied. The core neutralizing epitope of the S protein (CEOO 499-638aa) has 9 amino acid loci mutated and 9 amino acid loci mutated. These mutations in the S gene may be one of the reasons for the decrease in the immune response of the PEDV vaccine.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S858.28

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