A new PCR-based assay amplifies the E6-E7 genes of most mucosal human papillomaviruses (HPV)

We established a new assay to detect the E6-E7 DNA of mucosal human papillo maviruses (HPV) by a PCR-based method using four pairs of degenerate LCR an d E7 primers (LCR-E7 PCR), This assay amplifies the full length of E6 and t he N-terminal part of E7. HPV typing was performed using restriction-fragme nt-length polymorphism (RFLP), and by analyzing the sequences of cloned PCR products. We compared this assay with the first generation hybrid captured assay (HCA-I) and the MY09/11-PCR method. LCR-E7 PCR was able to detect mo re than 34 mucosal HPV types and theoretically should detect two additional types. LCR-157 PCR and HCA-I detected HPV DNA in 70% (69/99) and 55% (54/9 9) of low-grade cervical intraepithelial lesions (LSIL), 89% (105/118) and 76% (90/118) of high-grade cervical intraepithelial lesions (HSIL), and 90% (56/62) and 79% (49/62) of invasive squamous cell carcinomas (SCC), respec tively. LCR-E7 PCR was more sensitive than the HCA-I Lest. Discordant resul ts between the LCR-E7 and MY 11/09-PCR tests were observed in one of 185 (0 .5%) normal samples, seven of 85 (8.2%) LSIL samples, seven of 82 (8.5%) HS IL samples, and four of 72 (5.6%) SCC samples. The discordant results were mostly observed in samples with a low-copy number of the HPV genome or with multiple HPV infection. The sensitivity of LCR-E7 PCR was equivalent to th at of MY 11/09 ECR, and false positives were less frequent in LCR-E7 PCR. L CR-E7 PCR may be useful for determining the biological activity of detected HPV types, since this method amplifies the entire E6 gene. (C) 2000 Elsevi er Science B.V. All rights reserved.