Oncogenic human papillomavirus genotyping by multiplex PCR and fragment analysis

Abstract

Human papillomavirus (HPV) detection and genotyping are determinant in cervical cancer prevention. They help to identify women at risk and allow the establishment of epidemiologic profiles. Many PCR-based genotyping methods have been developed. They require many steps or specialized equipment increasing the assay duration and cost. This affects their routine use, especially in developing countries. Therefore, the aim of this study was to develop a new HPV genotyping method that can be routinely used also in low incomes countries. For this purpose, fifteen high risk HPV type specific reverse primers were designed on L1 gene and fluorescently labeled. These primers were used on two multiplex PCR with one common forward primer (MY11). The lengths of products were revealed by capillary electrophoresis. This technique identifies sixteen high risk HPVs (types 16, 18, 31, 33, 35, 39, 45, 51, 52, 53, 56, 58, 59, 68, 73 and 82). It was optimized on HPV genome plasmids and evaluated on artificial and cervical samples. All the sixteen targeted genotypes were identified specifically and repeatedly in simple and multiple infections in both artificial and clinical samples. The developed technique is sensitive, specific, easy to perform and appropriate for routine laboratory use and high throughput screening programs.