Yeung P S Marie, Hayes Micaela C, DePaola Angelo, Kaysner Charles A, Kornstein Laura, Boor Kathryn J
Department of Food Science, Cornell University, Ithaca, New York 14853, USA.
Appl Environ Microbiol. 2002 Jun;68(6):2901-9. doi: 10.1128/AEM.68.6.2901-2909.2002.
Historically, Vibrio parahaemolyticus infections have been characterized by sporadic cases caused by multiple, diverse serotypes. However, since 1996, V. parahaemolyticus serotype O3:K6 strains have been associated with several large-scale outbreaks of illness, suggesting the emergence of a "new" group of organisms with enhanced virulence. We have applied three different molecular subtyping techniques to identify an appropriate method for differentiating O3:K6 isolates from other serotypes. Pulsed-field gel electrophoresis (PFGE) following NotI digestion differentiated seven closely related subtypes among O3:K6 and related strains, which were distinct from PFGE patterns for non-O3:K6 isolates. Ribotyping and tdh sequencing were less discriminatory than PFGE, but further confirmed close genetic relationships among recent O3:K6 isolates. In vitro adherence and cytotoxicity studies with human epithelial cells showed that O3:K6 isolates exhibited statistically higher levels of adherence and cytotoxicity to host cells than non-O3:K6 isolates. Epithelial cell cytotoxicity patterns were determined with a lactate dehydrogenase release assay. At 3 h postinfection, high relative cytotoxicities (>50% maximum lactate dehydrogenase activity) were found among a greater proportion of recently isolated O3:K6 and closely related strains (75%) than among the non-O3:K6 isolates (23%). A statistically significant relationship between adherence and cytotoxicity suggests that the pathogenic potential of some isolates may be associated with increased adherence to epithelial cells. Our findings suggest that enhanced adherence and cytotoxicity may contribute to the apparent unique pathogenic potential of V. parahaemolyticus O3:K6 strains.
从历史上看，副溶血性弧菌感染的特征是由多种不同血清型引起的散发病例。然而，自1996年以来，副溶血性弧菌O3:K6血清型菌株已与几起大规模疾病暴发相关联，这表明出现了一群具有增强毒力的“新型”生物体。我们应用了三种不同的分子分型技术来确定一种区分O3:K6分离株与其他血清型的合适方法。NotI消化后的脉冲场凝胶电泳（PFGE）在O3:K6及相关菌株中区分出七个密切相关的亚型，这些亚型与非O3:K6分离株的PFGE模式不同。核糖体分型和tdh测序的区分能力不如PFGE，但进一步证实了近期O3:K6分离株之间密切的遗传关系。对人上皮细胞进行的体外黏附及细胞毒性研究表明，与非O3:K6分离株相比，O3:K6分离株对宿主细胞的黏附及细胞毒性在统计学上更高。用乳酸脱氢酶释放试验确定上皮细胞毒性模式。感染后3小时，在近期分离的O3:K6及密切相关菌株中，较高比例（75%）的菌株具有高相对细胞毒性（>最大乳酸脱氢酶活性的50%），而非O3:K6分离株中的这一比例为23%。黏附与细胞毒性之间具有统计学意义的关系表明，一些分离株的致病潜力可能与对上皮细胞黏附增加有关。我们的研究结果表明，增强的黏附及细胞毒性可能有助于副溶血性弧菌O3:K6菌株明显独特的致病潜力。
