Nested Multiplex Endpoint Polymerase Chain Reaction: An Alternative Method for Human Papilloma Virus Genotyping in Resource-limited Settings

E6, E7 messenger RNA (mRNA) PCR, and transcription-mediated amplification using Aptima assay]. 7,8 IntroductIonCervical cancer, caused by human papillomavirus (HPV), is the third-ranked cancer in India with 123,907 new cases and 77,348 deaths in 2020, with a 5-year prevalence of 42.82/100,000 population. 1,2 Screening methods have been developed to detect HPV infection and precancerous lesions at an early stage.These approaches include Pap smear, visual inspection of the cervix with acetic acid application (VIA), colposcopy followed by biopsy, and molecular methods polymerase chain reaction (PCR), hybrid capture 2 (HC2), transcription-mediated amplification on cervical swabs for the detection of various high-risk genotypes of HP. 3 Human papillomavirus is a double-stranded, nonenveloped DNA virus with >200 different genotypes. 4-6Among the genotypes, there are low-risk (nononcogenic) genotypes such as 6, 11, 42, 43, and 44, as well as 14 high-risk genotypes such as 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and 68.The genome of HPV is divided into three regions-(1) an upstream regulatory region (URR); (2) an early region, encoded by six genes (E1, E2, E4, E5a/5b, E6, E7); and (3) a late region, encoded by two genes for capsid proteins (L1, L2).The tests designed for HPV detection target L1 and L2 regions or the E6 and E7 genes for the oncoproteins. 4Laboratory methods for HPV detection include DNA detection (PCR, HC2, and fluorescence in situ hybridization (FISH) in formalin-fixed paraffin-embedded tissue) and RNA detection