Molecular diagnosis and phylogenetic analysis of human papillomavirus type-16 from suspected patients in Pakistan.

Abdullah Abdullah,Shahnaz Murtaza,Muhammad Qasim,Sajid Mehmood,Qamar Javed,Shamsa Parveen,Muhammad Zeeshan Hyder,Syed Muhammad Saqlan Naqvi,Salman Akbar Malik,Abdul Sami,Muhammad Ijaz,Shahida Husain Tarar,Muhammad Shafiq

doi:10.1186/s13027-016-0047-z

Abstract

BackgroundHuman Papillomavirus (HPV) is well known pathogen that can cause benign and malignant tumors in humans, yet there is very little information regarding HPV types prevalent in Pakistan.MethodsA total of 92 cervical secretions were collected from suspected married female patients and used for DNA isolation using a novel isolation method. The samples were tested through Polymerase Chain Reaction (PCR) using already reported primers MY09/MY11, GP5/GP6, GP5+/GP6+, CP65/CP70, CP66/CP69 and SPF1/SPF2 and with those developed in this study including HRT1 and HRT2 primer sets for typing HPV types and HACTB primer set for human beta actin gene as internal positive control. Sequencing and phylogenetic analyses were performed for two isolates to determine circulating HPV types.ResultsPCR with HRT1 and HRT2 indicated 2 (2.17 %) patients were positive for HPV type- 16 while 1 (1.08 %) with HPV type 18. Sequencing and phylogenetic analysis of isolates confirmed HPV type-16 in genus alpha 9 which have 99 % homology with already reported HPV from Japan and Costa Rica.ConclusionThis is the first report of HPV type-16 genus alpha 9 in Pakistan and the reported assay and sequence data will serve as valuable tools in further epidemiological studies for HPV surveillance to improve public health, especially of females in Pakistan.

Highlights

Human Papillomavirus (HPV) is well known pathogen that can cause benign and malignant tumors in humans, yet there is very little information regarding Human papillomaviruses (HPVs) types prevalent in Pakistan
The Polymerase Chain Reaction (PCR) analyses indicated that of 92 suspected married female patients of diverse age group and ethnicity, only 2 (2.17 %) patients tested positive for HPV type-16, while 1 (1.08 %) was tested positive with HPV Type 18
The PCR amplification with already reported primer sets MY09/MY11, GP5/GP6, GP5+/GP6+, CP65/CP70, CP66/CP69, and SPF1/SPF2 were performed but showed no positive result, the human actin beta gene was positively amplified using HACTB primer set. When these samples were tested with HRT1 and HRT2 novel primer sets, 3 out of 92 samples were found positive for HPV, two with HRT1 while other from HRT2 primer sets

Summary

Introduction

Human Papillomavirus (HPV) is well known pathogen that can cause benign and malignant tumors in humans, yet there is very little information regarding HPV types prevalent in Pakistan. Human papillomaviruses (HPVs) are diverse, nonenveloped, double stranded DNA viruses which are sexually transmitted and belong to family Papillomaviridae. The family Papillomaviridae is very large and contains 16 genera [1] with virus species which infects humans, non-human mammals and reptiles. More than 120 species of this family infect humans belonging to five genera including Alphapapillomavirus, Betapapillomavirus, Gammapapillomavirus, Mupapillomaviurs and Nupapillomavirus of which genus Alphapapillomavirus is the biggest. The HPV type-16, followed by HPV type-18 are reported to be associated with 70 % of invasive cervical cancer worldwide [4]. Infection with human papillomaviruses is directly associated with ano-genital cancer in both males and females [5, 6]. The cervical cancer associated with HPV infection is the second most common cancer in women worldwide, with 8.2 % mortality rate [7], and is the major cancer of women in most of the developing countries [8] where majority of the cervical cancer cases

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