淋病奈瑟菌临床菌株PⅠ基因型及PⅠB基因点突变分析  

Genotyping of dominant outer membrane porinⅠgene of Neisseria gonorrhoeae isolates and analysis of me site mutation of PⅠB gene

作　　者：孙爱华[1] 罗冬娇[2] 胡美多[3] 周晓辉[3] 严杰[3] 

机构地区：[1]浙江医学高等专科学校 [2]杭州师范学院基础医学部 [3]浙江大学医学院病原生物学教研室,杭州310031

出　　处：《中华微生物学和免疫学杂志》2006年第9期790-794,共5页Chinese Journal of Microbiology and Immunology

基　　金：浙江省医药卫生科学研究基金(2004A018)

摘　　要：目的分析浙江地区淋病奈瑟菌株外膜孔蛋白PⅠ基因序列及其点突变与耐药性关系,了解本地区淋病奈瑟菌株PⅠA与PⅠB基因分布及其优势基因型。方法采用高保真PCR扩增34株淋病奈瑟菌全长PⅠ基因序列,T-A克隆后测序。根据测序结果,建立多重PCR同时检测113株淋病奈瑟菌PⅠA和PⅠB基因。分析测序菌株PⅠB基因中与耐药性密切相关的G120和A121变异情况,采用二倍琼脂稀释法确定PⅠB菌株的耐药性。结果11株PⅠA+淋病奈瑟菌均为ⅠA6血清型。23株PⅠB+淋病奈瑟菌中,6株(26．1％)为ⅠB3血清型、5株(21．7％)为ⅠB3／6血清型、2株(8．7％)为ⅠB4血清型、9株(39．1％)为ⅠB3／6-ⅠB2嵌合血清型、1株(4．3％)为ⅠB2-ⅠB4嵌合血清型。所建立的多重PCR可特异和准确地对PⅠA和PⅠB基因分型,检测灵敏度为10 ng DNA模板。113株淋病奈瑟菌中,26株(23．0％)和87株(77．0％)分别携带PⅠA和PⅠB基因。经测序的23株PⅠB+淋病奈瑟菌中,1株(4．3％)G120和A121均未突变,3株(13．0％)G120或A121突变,2株8．7％)G120突变但A121缺失,17株(73．9％)双位点突变。22株G120和／或A121突变的PⅠB+菌株均对青霉素和四环素耐药。结论所建立的多重PCR可用于淋病奈瑟菌PⅠA和PⅠB基因的分型检测。本地区流行的淋病奈瑟菌主要携带PⅠB基因。ⅠA6为PⅠA菌株优势血清型。PⅠB菌株以ⅠB3／6-ⅠB2嵌合、ⅠB3和ⅠB3／6血清型常见,但主要为耐药性G120和,或A121突变株。Objective To analyze the dominant genotypes of outer membrane porin Ⅰ （PⅠ） gene in the isolates of Neisseria gonorrhoeae from Zhejiang area and the correlation among the genetic site mutation and drug resistance, and to determine the distribution of PⅠA and PⅠB genes in the isolates and their dominant genotypes. Methods entire PⅠ gene of 34 N. gonorrhoeae isolates were amplified using high fidelity PCR. The target amplification fragments were sequenced after T-A cloning. The mutation sites G120 and A121 were analyzed. Results All the 11 PⅠA^＋ isolates were belonging to serovars ⅠA6. Of the 22 PⅠB^＋ isolates, 6 （26.1%）, 5（21.7%）, 2（8.7%）, 9（39.1%） and 1（4.3%） strains were identified as serovars ⅠB3, ⅠB3/6, ⅠB3/6-ⅠB2 and ⅠB2-ⅠB4 chimera, respectively. The sensitivity of the PCR was as high as 10 ng DNA template. 23.0%（26/113） and 77.0% （87/113） of the 113 N. gonorrhoeae isolates carry PⅠA and PⅠB genes. In the 23 sequenced PⅠB^＋ isolates, one strain（4.3%） had no mutation at the G120 and A121 sites, while 3 strains（13.0%） showed G120 or A121 mutation, 2 strains（8.7%） displayed G120 mutation but lack of A121, and 17 strains（73.9%） exhibited the double site mutation. All the 22 PⅠB^＋ isolates with mutation at G120 and/or A121 sites were resistant to penicillin and tetracycline. Conclusion The N. gonorrhoeae strains prevalent in the local area are mainly taking PⅠB gene. ⅠA6 is the dominant serovars of the PⅠA^＋ N. gonorrhoeae isolates. ⅠB3/6-ⅠB2 chimeric, ⅠB3 and ⅠB3/6 serovars are common in the PⅠB^＋ N. gonorrhoeae isolates and most of the isolates are the drug resistant G120 and/or A121 mutants.

关 键 词：淋病奈瑟菌 PorinⅠ基因 序列分析 多重PCR 突变 

分 类 号：R378[医药卫生—病原生物学]