Development of a PCR method for rapid identification of new Streptococcus mutans serotype k strains
Development of a PCR method for rapid identification of new Streptococcus mutans serotype k strains
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DOI：
10.1128/jcm.42.11.4925-4930.2004
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发表时间：
2004-11-01
影响因子：
9.4
通讯作者：
Ooshima, T
中科院分区：
文献类型：
--
作者：
Nakano, K;Nomura, R;Ooshima, T
在之前的研究中，我们从人的血液和口腔中分离并鉴定了一种新的k型变形链球菌。通过对k型菌株rgpF基因5‘端rgpF基因5’端序列与参考菌株的比较分析，发现k型菌株rgpF基因5‘端与参考菌株rgpF基因5’端核苷酸序列具有较高的同源性，并根据该可变区序列设计了一套快速鉴定k型菌株的方法。用特异性扩增k型菌株的引物进行聚合酶链式反应，对c、e、f型菌株呈阴性反应，对k型菌株呈阳性反应，对k型菌株鉴定的敏感性为5~50个细胞。其次，用200例2~18岁受试者唾液DNA检测了血清k型特异性引物的阳性反应频率，其中10例出现阳性反应，高于我们以前用血清学方法进行的调查。此外，我们先前研究中所有k型菌株的受试者的唾液样本都显示出对k型特异性引物的阳性。这些结果表明，这种新的聚合酶链式反应方法是一种有效的检测变形链球菌k型的方法。
In a previous study, we isolated and characterized a new serotype k of Streptococcus mutans from human blood and oral cavities. Analysis of the genes involved in biosynthesis of the serotype-specific polysaccharide of serotype k strains revealed that the serotype k-specific nucleotide alignment was commonly present in the 5' region of the rgpF gene (350 bp from the initial sequence) compared to the reference strains, and then a method for rapid identification of serotype k strains was developed by use of PCR with primers designed on the basis of the sequence of the variable region. PCR assays with primers specific for amplification of serotype k strains showed a negative reaction with serotype c, e, and f strains and a positive reaction with serotype k strains, with the sensitivity for identification of the serotype k strains shown to range from 5 to 50 cells. Next, the frequency of positive reactions for serotype k-specific primers was surveyed with DNA taken from saliva samples from 200 subjects (2 to 18 years of age), and 10 of those showed a positive reaction, which was higher than the frequency in our previous survey with a serological method. In addition, all saliva samples from subjects with serotype k strains in our previous study were shown to be positive with the serotype k-specific primers. These results indicate that this new PCR method is effective for identification of subjects with S. mutans serotype k.