Human papillomavirus (HPV) study of 691 pathological specimens from Quebecby PCR-direct sequencing approach

Human papillomaviruses (HPV) are etiological agents of cervical cancer. In order to address clinical demand for HPV detection and sequence typing, mos tly in pre-cancerous cervical lesions, we applied our two-tier PCR-direct s equencing (PCR-DS) approach based on the use of both MY09/MY11 and GP5+/GP6 + sets of primers. We tested 691 pathological specimens, all of which were biopsies, 75% of which were diagnosed histologically as cervical intraepith elial neoplasia (CIN) grades I-III. In total, 484 samples (70%) tested HPV- positive, yielding 531 HPV sequences from 47 HPV types, including two novel types. Four most frequently found HPV types accounted for 52.9% of all iso lates: HPVG, 16, 11, and 31 (21.5%, 20.0%, 7.0%, and 4.5%, respectively). S ome interesting results are the following: all currently known high-risk HP V (14 types) and low-risk HPV (6 types) were detected; HPV18 was not the 1s t or 2nd but rather the 4th-5th most frequent high-risk HPV type; the highe st detection rate for HPV (86%) among samples suspected to be HPV-infected was found in the youngest age group (0-10 years old), including 70% (44/63) "genital" HPV types; HPV types of undetermined cervical cancer risk repres ented 19% and of the total HPV isolates but were strongly increased in co-i nfections (36.5% of all isolates). To our knowledge, this is the largest se quencing-based study of HPV. The HPV types of unknown cancer risk, represen ting the majority of the known HPV types, 27 of the 47 types detected in th is study, are not likely to play a major role in cervical cancer because th eir prevalence in GIN-I, II, and III declines from 16% to 8% to 2.5%. The t wo-tier PCR-DS method provides greater sensitivity than cycle sequencing us ing only one pair of primers. It could be used in a simple laboratory setti ng for quick and reliable typing of known and novel HPV from clinical speci mens with fine sequence precision. It could also be applied to anti-cancer vaccine development. (C) 2001 Wiley-Liss, Inc.