断奶仔猪源大肠杆菌LEE及HPI毒力岛的检测

应用Duplex_PCR方法,对240株断奶仔猪源大肠杆菌分离株的LEE毒力岛的eaeA基因和耶尔森菌强毒力岛核心区的irp2基因进行了检测,并对HPI毒力岛的fyuA基因及其在大肠杆菌染色体中的插入位置进行了分析,以及随机选取部分PCR产物进行了克隆和序列分析。结果表明:其中29株(12.08%)为LEE HPI ,39株(16.25%)为LEE ,11株(4.58%)为HPI ;另外还发现:不同病例来源的分离株之间,两种毒力岛的携带率不同;在断奶仔猪腹泻源分离株中,29株(20.71%)为LEE HPI ,22株(15.71%)为LEE ,9株(6.43%)为HPI ;断奶仔猪水肿病源分离株中,仅5株(6.58%)为LEE ,2株(2.63%)为HPI ,未发现LEE HPI 菌株;断奶仔猪水肿病并发腹泻源分离株中,仅12株(50%)为LEE ,未发现HPI 及LEE HPI 菌株。本实验克隆的eaeA(425bp)与已发表序列完全一致,irp2(280bp)f、yuA(948bp)、asn_tRNA_intB(1391bp)均与已发表的序列高度同源,同源性分别在98.2%、98.3%、95.8%以上;40株LEE HPI 或HPI 分离株中,29株(72.5%)为fyuA ,且其HPI毒力岛位于大肠杆菌染色体asn_tRNA位点。

Prevalence of LEE and HPI pathogenicity islands of Escherichia coli isolates from weaned piglets in China

To investigate the distribution of LEE pathogenicity island and HPI of Yersinia entercolitica in Escherichia coli isolates from weaned piglets,PCR based on intimin gene(eaeA) of LEE pathogenicity island and high molecular weight protein 2(HMWP1) gene(irp2) of HPI was developed.A total of 240 isolates from 140 diarrheic,76 edematous and 24 edematous/diarrheic weaned piglets from different farms were tested for the presence of the two genes.Sequence analysis of randomly selected PCR products showed that eaeA gene of 5 isolates was 100%,irp2 gene of 7 isolates was 98.2%,fyuA gene of 5 isolates was 98.3% and Asn_tRNA_intB of 5 isolates was 95.8% identical to the published sequences.Isolates with LEE HPI gene were more frequently detected in diarrheic swine than in edematous swine and edematous/diarrheic swine,and isolates with LEE gene were more frequently detected in edematous/diarrheic piglets than in edematous and diarrheic piglets.Furthermore,isolates with LEE or HPI or LEE HPI were more frequently detected in diarrheic swine.72.5% of HPI~ isolates were fyuA positive and linked to asn_tRNA.