荧光定量聚合酶链反应分型检测单纯疱疹病毒方法的评价与应用  

Evaluation and application of fluorescence quantitative polymerase chain reaction for detection and typing of herpes simplex virus

作　　者：刘军连[1] 司少艳[2] 常国辉[3] 杜海平[1] 徐冰心[2] 吕世超[1] 敬华[2] 易勇[2] 

机构地区：[1]中国人民解放军第306医院皮肤科,北京100101 [2]中国人民解放军第306医院检验科 [3]中国人民解放军军事医学科学院病原微生物生物安全国家重点实验室

出　　处：《实用预防医学》2014年第8期897-899,共3页Practical Preventive Medicine

基　　金：国家自然科学基金(30872284;81071402);病原微生物生物安全国家重点实验室开放基金(SKLPBS0913)

摘　　要：目的 荧光定量聚合酶链反应（FQ PCR）用于单纯疱疹病毒（HSV）分型检测的方法学评价及其初步应用.方法 以HSV-1 DNA多聚酶基因和HSV-2糖蛋白D基因为靶基因区,设计合成正向引物、反向引物和探针,采用FQPCR分别检测HSV-1和HSV-2标准株,对FQ-PCR进行方法学评价,并与病毒分离培养法相比较. 结果 建立的FQ-PCR对HSV检测和分型具有特异性;线性范围好（标准品的浓度在5×10^2 ～5×10^8 copies/ml,r=0.998）;灵敏度达到5×10^2 copies/ml;重复性较好,批内CV值为2.3％,批间CV值为4.8％.对120份生殖器棉拭标本用FQ-PCR和分离培养法进行分型检测,FQ-PCR的阳性率为25.8％,高于分离培养法的阳性率20.0％ （P＜0.05）. 结论 FQ-PCR方法具有快速、特异性好、敏感性高的优点,且分型准确,可用于HSV感染的早期诊断和分型.Objective To develop a fluorescence quantitative polymerase chain reaction （FQ-PCR） method for detection and typing of herpes simplex virus （HSV） and to evaluate its initial application.Methods By employing the FQ-PCR technique,the primers and probes targeted at HSV-1 DNA polymerase gene and HSV-2 glycoprotein D gene fraction were designed and applied to amplify DNA from HSV-1 or HSV-2,and then the PCR reaction system was optimized and evaluated.HSV in swab specimens from patients with genital herpes was detected by FQ PCR-and viral culture,and the detection results of the two methods were compared.Results The FQ-PCR assay showed good specificity for detection and typing of HSV,with a good linear range （5 × 10^2-5 × 10^8copies/ml,r =0.998）,a sensitivity of 5 × 10^2 copies/ml and a good reproducibility （the intra-and inter-assay coefficients of variation being 2.3% and 4.8%,respectively）.120 genital swab specimens were tested for HSV by both FQ-PCR and viral culture.The positive rate by FQ-PCR was higher than that by viral culture （25.8 % vs.20.0 %,P 〈 0.05）.Conclusions FQ-PCR is a rapid,specific,sensitive and accurate method for the detec tion and typing of HSV,which can be used in early clinical diagnnois and typing of HSV infection.

关 键 词：单纯疱疹病毒 荧光定量聚合酶链反应 检测 分型 

分 类 号：R373.1[医药卫生—病原生物学]