Genetic characterization of Escherichia coli O157:H7/- strains carrying the stx2 gene but not producing Shiga toxin 2
Genetic characterization of Escherichia coli O157:H7/- strains carrying the stx2 gene but not producing Shiga toxin 2
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DOI：
10.1111/j.1348-0421.2006.tb03779.x
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发表时间：
2006-01-01
影响因子：
2.6
通讯作者：
Nishibuchi, M
中科院分区：
文献类型：
--
作者：
Koitabashi, T;Vuddhakul, V;Nishibuchi, M
9株主要来自泰国和日本的非临床来源的大肠杆菌O157：H7/-菌株携带STX(2)基因，但在反向被动乳胶凝集试验中不产生Stx2毒素。一株含有STX(2)噬菌体(933W)的菌株(EDL933)与一株STX2阴性但含有STX(2)基因的菌株(泰国-12)进行了比较。为了研究STX2产生缺失的原因，对泰国-12STX(2)基因及其上游核苷酸序列进行了分析。用克隆的STX(2)基因构建表达载体，获得了完整的泰国-12STX(2)基因编码区，并用RPLA法进行了检测。LacZ融合分析发现，泰国-12的6个启动子不起作用。由于这6个基因位于STX(2)噬菌体基因组晚期启动子下游，Q蛋白的抗终止活性是STX(2)强转录所必需的。泰-12的Q基因与021噬菌体高度同源，而与933W噬菌体不具有高度同源性。外源Q基因的高水平表达表明，在TAI-12中Q的抗终止活性较弱。在STX2阴性菌株中未观察到STX(2)噬菌体的复制。在所有Stx2阴性株中，泰国-12株的Q-STX(2)基因序列都很保守。一项检测泰国-12 Q-STX(2)序列的聚合酶链式反应分析表明，来自市场上的马来西亚牛肉的O157菌株中有30%携带该序列，它们很少产生Stx2或根本没有产生Stx2。这些结果表明，STX(2)阳性的O157菌株产生很少或不产生Stx2，可能在亚洲环境中广泛分布。
Nine Escherichia coli O157:H7/- strains isolated primarily from non-clinical sources in Thailand and Japan carried the stx(2) gene but did not produce Stx2 toxin in a reversed passive latex agglutination (RPLA) assay. A strain (EDL933) bearing a stx(2) phage (933W) was compared to a strain (Thai-12) that was Stx2-negative but contained the stx(2) gene. To study the lack of Stx2 production, the Thai-12 stx(2) gene and its upstream nucleotide sequence were analyzed. The Thai-12 stx(2) coding region was intact and Stx2 was expressed from a cloned stx(2) gene using a plasmid vector and detected using RPLA. A lacZ fusion analysis found the Thai-12 six, promoter non-functional. Because the six, gene is downstream of the late promoter in the stx(2) phage genome, the antitermination activity of Q protein is essential for strong stx(2) transcription. Thai-12 had the q gene highly homologous to that of 021 phage but not to the 933W phage. High-level expression of exogenous q genes demonstrated Q antitermination activity was weak in Thai-12. Replication of stx(2) phage was not observed in Stx2-negative strains. The q-stx(2) gene sequence of Thai-12 was well conserved in all Stx2-negative strains. A PCR assay to detect the Thai-12 q-stx(2) sequence demonstrated that 30% of O157 strains from marketed Malaysian beef carried this sequence and they produced little or no Stx2. These results suggest that stx(2)-positive O157 strains that produce little or no Stx2 may be widely distributed in the Asian environment.