A nested multiplex PCR (NMPCR) assay that combines degenerate E6/E7 consensus primers and typespecific 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 Human papillomomaviruses (HPVs) constitute a group of more than 100 different genotypes associated with benign and malignant neoplasms of skin and mucous membranes (5, 34). Approximately 40 different HPV genotypes have been detected in the anogenital mucosa (34). On the basis of their epidemiological association with the development of cervical carcinoma, a group of so-called high-risk HPV genotypes has been defined. These include HPV genotype 16 (HPV-16),

The oncogenic potential of the high-risk human papillomavirus (HPV) genotypes (types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68) depends on the expression of the two viral oncogenes E6 and E7. Thus, the detection of HPV E6/E7 oncogene transcripts could serve as a factor in the evaluation of a risk of development of cervical intraepithelial neoplasia (CIN) and its progression to cervical cancer. A nested RT-PCR assay for the detection of E6/E7 oncogene transcripts of all known high-risk HPV genotypes was established. In the study described, 779 high-risk HPV-DNA-positive cervical scrapes exhibiting all grades of CIN, including non-dysplastic cervical mucosa (CIN 0), were examined. Spliced E6/E7 oncogene transcripts of all the high-risk HPVs were detected in numerous samples, with an overall detection rate of 47%. In 227 cases with agreement between the cytologic and histologic findings, the prevalence increased with lesion severity: CIN 0, 18%; CIN I, 58%; CIN II, 77%; CIN III, 84%. Multiple transcriptionally active high-risk HPVs were detected in 12% (33/279) of patients with multiple high-risk HPV infections. This work sets the stage for a prospective follow-up study currently being undertaken to evaluate the prognostic relevance of the detection of high-risk HPV E6/E7 oncogene transcripts for the persistence of a high risk HPV infection, and the possible evolution and further development of a CIN. Future applications of the assay described may include the monitoring of women in studies investigating antiviral treatment or vaccination.