Department of Histology, Microbiology, and Medical Biotechnologies, University of Padova, Padova, Italy.
J Clin Virol. 2011 Oct;52(2):93-7. doi: 10.1016/j.jcv.2011.07.006. Epub 2011 Jul 29.
An accurate tool for human papillomavirus (HPV) typing is important both for management of patients with HPV infection and for surveillance studies.
Design and evaluation of an HPV typing method based on 454 next generation sequencing (NGS) technology.
Development of an HPV typing method based on 454 NGS of HPV L1 amplicons generated with MY09/11-based primers. Evaluation of the NGS method in control samples and in a panel of cervical cytological samples. Comparison of the NGS typing method with cycle sequencing and with the reverse hybridization-based INNO-LiPA HPV Genotyping Extra assay (LiPA).
In control samples carrying mixtures of HPV16 and HPV18 DNA, the NGS method could reliably detect genotype sequences occurring at a frequency of 1% in multiple infections with a sensitivity of 100 genome equivalents/μL. In cervical cytology samples, comparison with cycle sequencing demonstrated accuracy of HPV typing by NGS. The NGS method had however lower sensitivity for some HPV types than LiPA, conceivably due to the poor sensitivity of the MY09/11-based primers. At variance, LiPA could not detect HPV types which were present in low proportion in multiple infections (<10% of HPV reads obtained by NGS). In addition, NGS allowed identifying the presence of different variants of the same HPV type in a specimen.
NGS is a promising method for HPV typing because of its high sensitivity in multiple infection and its potential ability to detect a broad spectrum of HPV types, subtypes, and variants.
准确的人乳头瘤病毒（HPV）分型工具对于 HPV 感染患者的管理和监测研究都很重要。
设计和评估一种基于 454 高通量测序（NGS）技术的 HPV 分型方法。
基于 MY09/11 引物扩增的 HPV L1 扩增子的 454 NGS 开发 HPV 分型方法。在对照样本和宫颈细胞学样本中评估 NGS 方法。将 NGS 分型方法与循环测序和基于反向杂交的 INNO-LiPA HPV Genotyping Extra 检测（LiPA）进行比较。
在携带 HPV16 和 HPV18 DNA 混合物的对照样本中，该 NGS 方法能够可靠地检测到多重感染中频率为 1%的基因型序列，灵敏度为 100 个基因组当量/μL。在宫颈细胞学样本中，与循环测序比较表明 NGS 进行 HPV 分型的准确性。然而，与 LiPA 相比，NGS 对某些 HPV 型的敏感性较低，这可能是由于 MY09/11 引物的敏感性较差。相比之下，LiPA 无法检测到在多重感染中存在比例较低（通过 NGS 获得的 HPV 读取的<10%）的 HPV 型。此外，NGS 还可以在标本中识别同一 HPV 型的不同变体的存在。
NGS 是一种很有前途的 HPV 分型方法，因为它在多重感染中具有很高的灵敏度，并且具有检测广泛 HPV 型、亚型和变体的潜力。
