Guglielmetti P, Bravo L, Zanchi A, Montè R, Lombardi G, Rossolini G M
Istituto di Clinica delle Malattie Infettive, Università di Siena, Italy.
Mol Cell Probes. 1994 Feb;8(1):39-44. doi: 10.1006/mcpr.1994.1005.
A polymerase chain reaction assay was developed for detection of the Vibrio cholerae heat-stable enterotoxin gene (sto). The assay is based on two oligonucleotide primers suitable for amplification of the entire sto open reading frame. Reaction conditions were defined to obtain optimal results in terms of specificity and sensitivity. Under these conditions the assay was highly sensitive and enabled good results to be obtained using, as a template, crude DNA preparations from single bacterial colonies. A rapid protocol for direct sequence analysis of the amplification product, which may be used in combination with the PCR assay to confirm the product identity and analyse sequence polymorphisms within the sto gene, was also developed. Twenty-two V. cholerae non-O1 isolates from sporadic cases of V. cholerae non-O1-associated gastroenteritis from Cuba were analysed by this PCR assay. Four strains (18.2%) were found to carry the sto gene. The prevalence of V. cholerae non-O1 strains carrying the sto gene among clinical isolates is higher than that reported for other geographical areas, except in the case of epidemics.
开发了一种聚合酶链反应检测法，用于检测霍乱弧菌热稳定肠毒素基因（sto）。该检测法基于两个适合扩增整个sto开放阅读框的寡核苷酸引物。确定了反应条件，以便在特异性和灵敏度方面获得最佳结果。在这些条件下，该检测法高度灵敏，使用单个细菌菌落的粗制DNA制剂作为模板即可获得良好结果。还开发了一种用于扩增产物直接序列分析的快速方案，该方案可与PCR检测法结合使用，以确认产物身份并分析sto基因内的序列多态性。通过该PCR检测法对来自古巴散发性非O1群霍乱弧菌相关肠胃炎病例的22株非O1群霍乱弧菌分离株进行了分析。发现4株菌株（18.2%）携带sto基因。除了在流行情况下，临床分离株中携带sto基因的非O1群霍乱弧菌菌株的流行率高于其他地理区域报告的流行率。
