用聚合酶链反应和限制性片段长度多态性分析进行沙眼衣原体的基因分型

Gene Typing of Chlamydia Trachomatis by Polymerase Chain Reaction and Restriction Fragment Length Polymorphism

为了研究泌尿生殖道沙眼衣原体（ＣＴ）感染的分子流行病学，我们建立了一种不需做培养的ＣＴ血清型鉴定方法。用聚合酶链反应（ＰＣＲ）扩增ＣＴ大部分主要外膜蛋白基因（ｏｍｐｌ），将扩增产物做限制性片段长度多态性（ＲＦＬＰ）分析，即用限制性内切酶ＡｌｕⅠ和ＭｓｐⅠ同时消化，产生的片段通过１０％聚丙烯酰胺凝胶电泳分离。经硝酸银染色后，１５个ＣＴ血清型标准株产生１３个特征性带谱。血清型Ｃ和Ｊ带型相似，用ＨｉｎｆⅠ酶切后区分；Ｈ和Ｌ３带型相同，用ＤｄｅⅠ和ＥｃｏＲⅠ同时酶切后区分。对１２份ＣＴ培养阳性的临床标本行ＰＣＲＲＦＬＰ分型检测。结果：Ｄ血清型为６份标本，Ｅ型２份，Ｆ型２份，Ｆ／Ｇ混合型１份，Ｂ型１份。

In order to facilitate molecular epidemiologic study of chlamydial urogenital infections,a method that aviods culture was developed to determine the serovars of Chlamydia trachomatis(CT). Polymerase chain reaction was used first to amplify a large part of the major outer membrane protien gene(omp1). The amplified DNA was then digested simultaneously with restriction endonuclease Alu I and Msp I and the resulting fragments were separated on 10% polyacrylamide gels. After silver nitrate staining, a total of 13 characteristic patterns were observed for the 15 serovars. Serovar C and J were similar in pattern, but could be distinguished by cleavage with HinfⅠ. Using Dde I and EcoRI, the patterns of serovar H and L3 could be differentiated from each other. Analysis of 12 CT positive clinical specimens gave the following results: 6D,2E,2F,1F/G and 1B strains. This study indicates that the omp1 based genotyping is useful for large scale epidemiologic studies of chlamydial infections.