非O157产志贺毒素大肠杆菌分离株的志贺毒素基因变种及黏附相关基因分析

Analysis of stx gene variants and putative adhesin genes of non-O157 Shiga toxin-producing Escherichia coli isolates

目的了解我国部分地区非O157产志贺毒素大肠杆菌分离株的志贺毒素基因变种及其黏附相关基因,为进一步研究致病机制提供依据。方法采用聚合酶链反应(polymerase chain reaction,PCR)方法对29株分离菌株的stx1、stx2基因全长扩增并测序,通过与GenBank中已公布的变种序列比对确定菌株stx1、stx2基因的变种类型。对位于LEE毒力岛上的eaeA、escF、escC、tir、espA、espB、espD基因及LEE以外其他黏附相关基因iha、toxB、efa1、sfpA、lpfAO157/OI-141、lpfAO157/OI-154、saa、lpfAO113、eibG进行PCR检测。结果 25株stx1阳性菌株中有13株携带stx1a原毒素,12株携带stx1c变种;10株stx2阳性菌株中,7株携带stx2d变种,1株为stx2a原毒素,1株携带stx2g变种,1株携带与stx2eA亚单位、stx2dB亚单位最接近的stx2变种。LEE岛上的7个基因检测结果均为阴性,黏附相关基因iha阳性率为89.7%(26/29),saa阳性菌株3株、eibG阳性菌株1株,其余6个黏附相关基因均为阴性。结论我国部分非O157 STEC菌株的志贺毒素基因以stx1c、stx2d变种为主,LEE毒力岛不存在,而黏附相关基因iha广泛存在于不同血清型的产志贺毒素大肠杆菌菌株中。

Objective To understand the stx gene variants and putative adhesin genes of the non-O157 Shiga toxinproducing Escherichia coli （STEC） isolates collected from some areas in China. Methods Polymerase chain reaction （PCR） was performed to amplify the complete stxl and stx2 genes of 29 non-O157 STEC isolates, and PCR products were sequenced and compared with the published sequences of stx1 and stx2 variants in the GenBank. Genes of pathogenicity island called locus of enterocyte effacement （LEE） and other putative adhesin genes outside the LEE were also investigated by PCR. Results Among 25 StXl positive isolates, 13 carded stx1δ, 12 carried stx1c. Among 10 stx2 positive isolates, 7 carded stx2d, and stx2s, stX2a were identified from 1 isolate respectively, 1 isolate carried the stx2 variant that is similar to stx2e in A subunit and identical to StX2d in B subunit. Seven genes encoded in the LEE were all negative among 29 isolates; putative adhesin genes iha was identified in 26 isolates （89. 7% ） ; saa was detected in 3 isolates ; eibG was detected in 1 isolate; other 6 putative adhesin genes were all negative. Conclusion The predominant stx variants of non-O157 STEC isolates from some areas in China were stxlcand stx2d, the absence of the 7 genes in the LEE suggested that the 29 isolates were all LEE negative. The putative adhesin gene iha was widely distributed while saa and eibG genes were present only in some isolates. The mechanisms that mediate colonization of non-O157 STEC strains are needed to be further studied.