Assessing genetic evolution and detecting human papillomavirus by matching two complementary highly sensitive approaches, nested-qPCR and sequencing.

Becoming armed with an appropriate strategy to isolate the minimum number of human papillomaviruses (HPV), regardless of DNA extraction method, can be a huge step in preventing false negative; it has a significant effect on the management and control of HPV infection among women's population. This study was conducted in Qom province, considering the risk factors associated with HPV. It was able to analyze genetic evolution in its genotypes and evaluated the limit of detection by a new diagnostic approach. Totally, 486 Pap smear samples were tested; then, the HPV DNA was developed by a semi-nested quantification PCR. Positive samples were sequenced and submitted to the GenBank (MG825048-MG825061). After alignment, phylogenetic and polymorphism analyses were performed on the sequenced samples with a number of GenBank sequences. The overall HPV prevalence among all women in Qom was 11.7%. HPV6 (43.24%) and HPV16 (6.75%) were the most frequent LR and HR genotypes, respectively. Although the Tajima's D of all genotypes was positive, it was negative individually. The position of genotypes 6, 11, and 73 was controversial on phylogenetic trees. Limit of detection (LOD) was obtained as about 10-100 copies per reaction in various genotypes of HPV by semi-nested qPCR. The nature of HPV could be preserved during natural selection. This research, through innovative usage of the primers, could detect different genotypes of the HPV, and inform the women society of the probable risk through its prevalence determination.