Detection and typing of human papillomavirus by E6 nested multiplex PCR
Detection and typing of human papillomavirus by E6 nested multiplex PCR
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DOI：
10.1128/jcm.42.7.3176-3184.2004
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发表时间：
2004-07-01
影响因子：
9.4
通讯作者：
Bültmann, B
中科院分区：
文献类型：
--
作者：
Sotlar, K;Diemer, D;Bültmann, B
应用套式多重聚合酶链式反应(NMPCR)技术，结合简并的E6/E7通用引物和型特异性引物，检测和分型人乳头瘤病毒(HPV)基因6/11、16、18、31、33、35、39、42、43、44、45、51、52、56、58、59、66和68型(n=1525)。用MY09-MY11和GP5+-GP6+对495例宫颈刮片标本进行NMPCR检测，并与MY09-MY11和GP5+-GP6+的常规聚合酶链式反应(MY/GP)进行比较。无宫颈上皮内瘤变(CIN 0)组HPV感染率为34.7%(102/294)，轻度不典型增生组(CIN 1/11)HPV感染率为94.2%(113/120)，重度不典型增生组(CIN III)HPV感染率为97.8%(44/45)。将所有四种HPV检测方法的组合作为“金标准”：在所有三种形态亚型中，NMPCR法的灵敏度均显著高于MY09-MY11和Gp5+-Gp6+法(P<；0.0001)，而灵敏度与My/GP法相当或相当。临床标本中均检出18种HPV基因分型。高危型与低危型HPV的比例从CIN 0的4：1(80/103)上升到CIN I~III的19：1(149/157)。47.9%(124/259)的患者出现多重感染。总之，新的NMPCR方法是一种灵敏而有用的检测HPV DNA的工具，特别是当需要准确的HPV基因分型和识别多个HPV感染时。
A nested multiplex PCR (NMPCR) assay that combines degenerate E6/E7 consensus primers and type-specific primers was evaluated for the detection and typing of human papillomavirus (HPV) genotypes 6/11, 16, 18, 31, 33, 35, 39, 42, 43, 44, 45, 51, 52, 56, 58, 59, 66, and 68 using HPV DNA-containing plasmids and cervical scrapes (n = 1,525). The performance of the NMPCR assay relative to that of conventional PCR with MY09-MY11 and GP5+-GP6+ primers, and nested PCR with these two primer sets (MY/GP) was evaluated in 495 cervical scrapes with corresponding histologic and cytologic findings. HPV prevalence rates determined with the NMPCR assay were 34.7% (102 of 294) in the absence of cervical intraepithelial neoplasia (CIN 0), 94.2% (113 of 120) in the presence of mild or moderate dysplasia (CIN 1/11), and 97.8% (44 of 45) in the presence of severe dysplasia (CIN III). The combination of all four HPV detection methods applied in the study was taken as "gold standard": in all three morphological subgroups the NMPCR assay had significantly (P < 0.0001) higher sensitivities than the MY09-MY11 and GP5+-GP6+ assays and sensitivities comparable or equal to those of the MY/GP assay. All 18 HPV genotypes investigated were detected among the clinical samples. The ratio of high- to low-risk HPV genotypes increased from 4:1 (80 of 103) in CIN 0 to 19:1 (149 of 157) in CIN I to III. Multiple infections were detected in 47.9% (124 of 259) of the patients. In conclusion, the novel NMPCR method is a sensitive and useful tool for HPV DNA detection, especially when exact HPV genotyping and the identification of multiple HPV infections are required.