重庆地区肺炎衣原体感染株的部分基因分析

PARTIAL GENETIC ANALYSIS OF CHLAMYDIA PNEUMONIAE IN CHONGQING

目的 用两对引物对临床标本中肺炎衣原体进行检测 ,对阳性PCR产物部分序列进行测定和同源性分析 ,探讨肺炎衣原体的基因分型。方法 用根据肺炎衣原体 16srRNA及种特异性 5 3kDa蛋白抗原基因序列设计的两对引物Cpnl～Cpn2 ,5 3A～ 5 3B对 1994年 5月～ 1999年 3月间采集的有呼吸道疾患病人的鼻咽拭子标本 182份进行PCR检测 ,从阳性标本中随意挑取 13 6和 14 4号标本 ,纯化PCR产物 ,进行核苷酸测序 ,并将测序结果与已报道的肺炎衣原体分离株进行同源性比较分析。结果 标准AR - 3 9株及 2 1例标本均扩增得到约 4 60nts的 16srRNA基因片段 ,2 5例标本扩增得约 83 0nts的5 3kDa蛋白基因核酸片段。对 13 6、14 4号标本的PCR产物测序结果显示 ,2 4 4nts的 16srRNA基因序列完全相同 ,且与标准肺炎衣原体AR - 3 9、TW 183、IOL2 0 7及CWL0 2 9的对应序列 10 0 %同源 ,而与马肺炎衣原体分离株N16相比较有 3处突变 ,与非洲热带蟾蜍分离株CPXT1比较有 3处突变 ,与小鼠肺炎衣原体分离株J13 8相比有一处不同 ,表明人的肺炎衣原体株 16rRNA基因较为保守 ,与动物分离株有差异 ;标本 14 5nts的 5 3kDa蛋白基因也 10 0 %同源 ,该序列与从人体分离的AR - 3 9株以及从小鼠分离的J13 8株序列完全一致 ,表明肺炎衣原体 5

Aim To explore the difference among the clinical strains of Chlamydia pneumoniae by genetic analysis with PCR.Methods 182 people suffered with respiratory problems between 1994 ～1999 were investigated.Nasopharyngeal swabs were collected and PCR was performed with two primer pairs based on 16s rRNA gene and 53kDa antigenic gene of Chlamydia pneumoniae.PCR products of two random clinical samples No.136 and 144 were partial sequenced.Nucleic sequence difference were analyzed by computer.Results An about 460nts fragment was amplified in 21 out of 182 samples with the 16s rRNA gene primer pair.While 25 samples were amplified out 830nts fragment by the other primer pair.The sequences of no.136 and 144 were identical in both of the 16s rRND and 53kDa protein gene.Among the 244nts of the amplified fragment of 16s rRNA gene,there were 3nts different compared to a horse isolate N16 and aTropicalis xenopus isolate CPXT1, there was 1nt different between no.136 and a mouse isolate J138,but these sequences were 100% identical to human isolates AR-39,TW-183,CWL-029and IOL-207.Among 145nts of the amplified fragment of 53kDa protein gene,no difference was found compared to strain AR-39 and J138.Conclusions The genotype analysis of Chlamydia is of a help to epidemiology.But performing this program in Chlamydia pneumoniae may rely on the development of other typing gene analysis and the investigation of more isolates.