DETECTION OF GENES FOR ENTEROTOXINS, EXFOLIATIVE TOXINS, AND TOXIC SHOCK SYNDROME TOXIN-1 IN STAPHYLOCOCCUS-AUREUS BY THE POLYMERASE CHAIN-REACTION
DETECTION OF GENES FOR ENTEROTOXINS, EXFOLIATIVE TOXINS, AND TOXIC SHOCK SYNDROME TOXIN-1 IN STAPHYLOCOCCUS-AUREUS BY THE POLYMERASE CHAIN-REACTION
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DOI：
10.1128/jcm.29.3.426-430.1991
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发表时间：
1991-03-01
影响因子：
9.4
通讯作者：
ROZEE, KR
中科院分区：
文献类型：
--
作者：
JOHNSON, WM;TYLER, SD;ROZEE, KR
应用合成的8对寡核苷酸引物，对临床标本和污染食品中分离的金黄色葡萄球菌肠毒素A~E、脱落毒素A、B和中毒性休克综合征毒素1的基因进行了检测。以毒素基因内部区域为靶点，用琼脂糖凝胶电泳法检测扩增片段。利用从88株金黄色葡萄球菌中提取的核酸对毒素基因进行了明确的识别，这些金黄色葡萄球菌的毒力从生物学和免疫学上都得到了证实。在免疫学检测中，两株金黄色葡萄球菌产生肠毒素C或中毒性休克综合征毒素1的结果不一致，表型鉴定与基因鉴定的总符合率为97.7%。通过使用已知产毒素细菌病原体和非产毒金黄色葡萄球菌的核酸，在聚合酶链式反应中建立了引物特异性。链球菌属的菌株，包括一些携带SPEA基因的热原外毒素A的产生菌，经设计用于检测葡萄球菌毒素的聚合酶链式反应呈阴性。所有葡萄球菌毒素基因的检测限值都是在各自的聚合酶链式反应中确定的。用聚合酶链式反应鉴定金黄色葡萄球菌菌株中的葡萄球菌毒素基因是一种非常特异、敏感、相对快速和廉价的方法，而传统的免疫学方法依赖于足够的基因表达来保证可靠性和敏感性。
Eight pairs of synthetic oligonucleotide primers were used in a polymerase chain reaction (PCR) protocol to detect genes for staphylococcal enterotoxins A to E, exfoliative toxins A and B, and toxic shock syndrome toxin 1 in Staphylococcus aureus strains isolated from clinical specimens and contaminated foods. Primers were targeted to internal regions of the toxin genes, and amplification fragments were detected after the PCR by agarose gel electrophoresis. Unequivocal discrimination of toxin genes was obtained by the PCR by using nucleic acids extracted from 88 strains of S. aureus whose toxigenicity was established biologically and immunologically. In immunological assays, two strains of S. aureus produced equivocal results for production of enterotoxin C or toxic shock syndrome toxin 1, giving an overall concordance between phenotypic and genotypic identification of 97.7%. Primer specificity was established in the PCR by using nucleic acids from known toxin-producing bacterial pathogens and from nontoxigenic S. aureus. Strains of Streptococcus spp., including some producers of pyrogenic exotoxin A carrying the speA gene, were negative by the PCR designed to detect staphylococcal toxins. The detection limits were established for all the staphylococcal toxin genes within their respective PCR protocols. The identification of staphylococcal toxin genes in strains of S. aureus by the PCR offers a very specific, sensitive, relatively rapid, and inexpensive alternative to traditional immunological assays which depend on adequate gene expression for reliability and sensitivity.