Feoli-Fonseca J C, Oligny L L, Brochu P, Simard P, Falconi S, Yotov W V
Department of Pathology, Ste-Justine Hospital, Women and Children University Hospital Center, University of Montreal, Montreal, Quebec, Canada.
J Med Virol. 2001 Apr;63(4):284-92. doi: 10.1002/1096-9071(200104)63:4<284::aid-jmv1003>3.0.co;2-h.
Human papillomaviruses (HPV) are etiological agents of cervical cancer. In order to address clinical demand for HPV detection and sequence typing, mostly in pre-cancerous cervical lesions, we applied our two-tier PCR-direct sequencing (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 intraepithelial 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 isolates: HPV6, 16, 11, and 31 (21.5%, 20.0%, 7.0%, and 4.5%, respectively). Some interesting results are the following: all currently known high-risk HPV (14 types) and low-risk HPV (6 types) were detected; HPV18 was not the 1st or 2nd but rather the 4th-5th most frequent high-risk HPV type; the highest 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 represented 19% and of the total HPV isolates but were strongly increased in co-infections (36.5% of all isolates). To our knowledge, this is the largest sequencing-based study of HPV. The HPV types of unknown cancer risk, representing the majority of the known HPV types, 27 of the 47 types detected in this study, are not likely to play a major role in cervical cancer because their prevalence in CIN-I, II, and III declines from 16% to 8% to 2.5%. The two-tier PCR-DS method provides greater sensitivity than cycle sequencing using only one pair of primers. It could be used in a simple laboratory setting for quick and reliable typing of known and novel HPV from clinical specimens with fine sequence precision. It could also be applied to anti-cancer vaccine development.
人乳头瘤病毒（HPV）是宫颈癌的病原体。为满足HPV检测和序列分型的临床需求，主要针对癌前宫颈病变，我们应用了基于MY09/MY11和GP5 + /GP6 +引物组的两层PCR直接测序（PCR-DS）方法。我们检测了691份病理标本，均为活检标本，其中75%经组织学诊断为宫颈上皮内瘤变（CIN）I-III级。总共484个样本（70%）检测为HPV阳性，产生了来自47种HPV类型的531条HPV序列，包括两种新型。四种最常见的HPV类型占所有分离株的52.9%：HPV6、16、11和31（分别为21.5%、20.0%、7.0%和4.5%）。一些有趣的结果如下：检测到了所有目前已知的高危HPV（14种）和低危HPV（6种）；HPV18不是最常见的前两种高危HPV类型，而是第四至第五常见的高危HPV类型；在疑似HPV感染的样本中，HPV检测率最高（86%）的是最年轻的年龄组（0-10岁），其中包括70%（44/63）的 “生殖道” HPV类型；风险未确定的HPV类型占所有HPV分离株的19%，但在合并感染中显著增加（占所有分离株的36.5%）。据我们所知，这是基于测序的最大规模的HPV研究。风险未知的HPV类型占已知HPV类型的大多数，在本研究中检测到的47种类型中有27种，它们在宫颈癌中不太可能起主要作用因为它们在CIN-I、II和III中的患病率从16%降至8%再降至2.5%。两层PCR-DS方法比仅使用一对引物的循环测序具有更高的灵敏度。它可用于简单的实验室环境，以快速、可靠地对临床标本中的已知和新型HPV进行分型，序列精度高。它也可应用于抗癌疫苗的开发。
