Nested PCR followed by NGS: Validation and application for HPV genotyping of Tunisian cervical samples

dc.creatorArdhaoui, Monia
dc.creatorEnnaifer, Emna
dc.creatorSalim, Anna Christina de Matos
dc.creatorAraújo, Flávio Marcos Gomes de
dc.creatorLaasili, Thalja
dc.creatorBoubaker, Med Samir
dc.date2022-03-18T17:40:30Z
dc.date2022-03-18T17:40:30Z
dc.date2021
dc.date.accessioned2023-09-26T20:53:10Z
dc.date.available2023-09-26T20:53:10Z
dc.identifierARDHAOUI, Monia et al. Nested PCR followed by NGS: Validation and application for HPV genotyping of Tunisian cervical samples. PLoS one, v. 16, n. 8, e0255914, 2021. doi.org/10.1371/journal.pone.0255914
dc.identifier1932-6203
dc.identifierhttps://www.arca.fiocruz.br/handle/icict/51749
dc.identifier10.1371/journal.pone.0255914
dc.identifier.urihttps://repositorioslatinoamericanos.uchile.cl/handle/2250/8865599
dc.descriptionThe most used methodologies for HPV genotyping in Tunisian studies are based on hybridization that are limited to a restricted number of HPV types and to a lack of specificity and sensitivity for same types. Recently, Next-Generation sequencing (NGS) technology has been efficiently used for HPV genotyping. In this work we designed and validated a sensitive genotyping method based on nested PCR followed by NGS. Eighty-six samples were tested for the validation of an HPV genotyping assay based on Nested-PCR followed by NGS. These include, 43 references plasmids and 43 positive HPV clinical cervical specimens previously evaluated with the conventional genotyping method: Reverse Line Hybridization (RLH). Results of genotyping using NGS were compared to those of RLH. The analytical sensitivity of the NGS assay was 1GE/mu l per sample. The NGS allowed the detection of all HPV types presented in references plasmids. On the clinical samples, a total of 19 HPV types were detected versus 14 types using RLH. Besides the identification of more HPV types in multiple infection (6 types for NGS versus 4 for RLH), NGS allowed the identification of HPV types that were not detected by RLH. In addition, the NGS assay detected newly HPV types that were not described in Tunisia so far: HPV81, HPV43, HPV74, and HPV62. The high sensitivity and specificity of NGS for HPV genotyping in addition to the identification of new HPV types may justify the use of such technique to provide with high accuracy the profile of circulating types in epidemiological studies
dc.formatapplication/pdf
dc.languageeng
dc.publisherPublic Library of Science
dc.rightsopen access
dc.titleNested PCR followed by NGS: Validation and application for HPV genotyping of Tunisian cervical samples
dc.typeArticle


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