三基因分型法检测梅毒螺旋体基因型别  

Molecular typing of Treponema pallidum by analysis of three target genes

作　　者：彭锐锐[1] 尹跃平[1] 魏万惠[1] 王红春[1] 张津萍[2] 陈祥生[1] 

机构地区：[1]中国医学科学院北京协和医学院皮肤病研究所、中国疾病预防控制中心性病控制中心,南京210042 [2]中国医学科学院北京协和医学院性病检验科,南京210042

出　　处：《中华皮肤科杂志》2011年第11期779-782,共4页Chinese Journal of Dermatology

基　　金：国家十一五科技重大专项（2008ZX10001-005）;美国北卡大学医学院研究基金（5U19A1031496-18）,志谢 美国华盛顿大学Sheila A.Lukehart和Lauren C.Tantalo教授

摘　　要：目的 分析运用最新的三基因定位分型方法检测梅毒螺旋体基因型别的敏感性与特异性。方法 分别采用Nichols标准株和快速血浆反应素环状卡片试验（RPR）、梅毒螺旋体明胶凝集试验（TPPA）均阴性的生殖器疱疹患者湿润性溃疡皮损DNA提取液作为阳性和阴性标本。分析arp基因60个碱基对重复序列的数目、tprEGJ基因MseI酶切后限制性片段长度多态性的型别和tp0548基因序列的型别，根据上述三基因的分析结果，分析梅毒螺旋体基因型别。临床标本来自一期或二期梅毒患者的湿润性皮损。先经梅毒螺旋体特异的polA基因扩增，阳性者用建立的分型方法进行分析。结果 Nichols标准株的基因型别为14a/a，阴性对照的三基因均无扩增。临床标本中三个基因的扩增敏感性分别是94.1%、91.2%和94.1%；91.2%的临床标本检测出完整的基因型别；3个基因的优势型别分别是14型、d型和f型。结论 改良的三基因分型方法检测梅毒螺旋体基因型别具有敏感性高、特异性好、区分度强的特点。Objective To evaluate the performance of a three-gene typing system in the determination of Treponema pallidum （Tp） genotypes. Methods To determine the genotypes of Tp, three targets were assessed, including the number of 60 base-pair repeats, restriction fragment length polymorphism （RFLP） pattern of tprEGJ gene after MseI digestion and the sequence of tp0548 gene. The DNA extracted from the Nichols strain of Tp served as the positive control, and that from the moist ulcer of patients with genital herpes and negative RPR or TPPA test results served as the negative control. To validate the typing method, clinical specimens were collected from the moist skin lesions of patients with primary or secondary syphilis, and subjected to the amplification of polA gene by PCR. The enhanced molecular typing system was used to determine the genotypes of Tp in Tp DNA-positive specimens. Results The Nichols strain harbored a genotype of 14a/a. No amplification of any of the three target genes was found in the negative control. The arp gene, tprEGJ gene and tp0548 gene were amplified from 94.1%, 91.2% and 94.1% of the 40 clinical specimens, and the genotype was successfully determined by the three-gene typing system for 91.2% of the clinical Tp strains. The predominant type of arp, tprEGJ and tp0548 genes was 14 repeats, d and f, respectively in these clinical Tp isolates. Conclusion The enhanced molecular tying method for Tp exhibits high sensitivity, specificity and discrimination potential.

关 键 词：梅毒 密螺旋体 苍白 基因分型 

分 类 号：R759.1[医药卫生—皮肤病学与性病学]