Genotype Distribution Of High-risk Human Papillomavirus And Polymorphism Analysis Of 52 Type E6/E7 Genes In Jingzhou Region

The incidence of cervical cancer ranks second among female malignant tumors in China,and it is association with the persistent human papillomavirus（HPV）infection.Up to now,more than 200 HPV types have been identified,which can be divided into high-risk and low-risk types according to the clinical progression of the disease.High-risk types,such as HPV16,18,31,33,35,39,45,51,52,56,58,66 and 68,etc,are closely relevant to the occurrence of cervical cancer;low-risk types,such as HPV6and HPV11,mainly cause benign lesions such as condyloma acuminatum.The global infection rate of high-risk HPV is relatively high,but the epidemiological characteristics are different due to regions and ethnics.High-risk HPV screening based on molecular biology techniques and HPV vaccination can reduce the incidence of cervical cancer.The E6 and E7 genes are the main oncogenes of HPV,which are sustained high expression in cancer cancer and HPV related diseases.The amino acid changes caused by genetic variation can change or affect the infection efficiency,pathogenicity and the host immune response,so E6 and E7 are ideal targets for research and development of HPV therapeutic vaccines.Vaccine research should focus on HPV types with high detection rate in the region.In this study,through molecular epidemiological investigation,the infection rate of high-risk HPV,dominant type and distribution characteristics of HPV infection in different age groups were clarified in Jingzhou region.Analyses of the E6 and E7 gene polymorphisms,the lineage distribution of variant strains and the positive selection sites,and the prediction of the T cell and B cell epitopes of the E6 and E7 protein of the high-risk HPV dominant types in Jingzhou region were done,in order to provide experimental data for the development of diagnosis and therapeutic vaccines against the virus in Jingzhou region.Objective:The epidemiological characteristics of high-risk human papillomavirus infection in Jingzhou region from 2018 to 2020 were retrospectively analyzed.The variation characteristics of E6 gene and E7 gene of HPV52 type were analyzed,and the T cell and B cell epitopes of E6 and E7 protein were predicted.Methods:20147 cases of high-risk HPV typing data were collected in Jingzhou Central Hospital from January 2018 to December 2020,in order to analyze the high-risk HPV infection rate,genotype distribution characteristics and HPV infection rates of different age groups in the region.The E6 gene and E7 gene of HPV52 single positive samples were amplified by PCR,sequenced,compared with Gen Bank reference sequence and analyzed sequence variability.MEGA-X and PAML4.9 were used to construct phylogenetic tree and estimate the selection pressure of E6-E7 genes,respectively.IEDB and ABCpred were used to predict E6 and E7 protein T cell and B cell epitopes.Results:1.The high-risk HPV infection rate in Jingzhou region was 20.13%,and the infection rate in 2018,2019 and 2020 were 20.83%,20.07%and 19.53%,respectively.Single HPV infection was dominated,the infection rate was 14.34%,and multiple infections account for 5.79%.HPV52,HPV58 and HPV16 were the virus types with relatively high infection rate,accounting for 6.61%,3.49%and 2.79%,respectively.The infection rate of HPV51 had showed the increasing trend during the 2018-2020 period.2.The infection rate of high-risk HPV were higher in women aged≤24 and≥55 years,with the infection rates being 22.78%and 30.83%,respectively.The infection rate of women aged 25-34 years was the lowest,which was 17.39%.Among single and multiple HPV type infections,the infection rate was highest in the age group of≥55 years.From 2018 to 2020,the infection rate of women in the 35-44 age group had been decreasing year by year over time.3.Fifteen single-nucleotide mutations were detected in the HPV52 E6 gene variant sequences,including six non-synonymous mutations and nine synonymous mutations.Nineteen single nucleotide mutations were detected in the E7 gene variant sequences,including ten non-synonymous mutations and nine synonymous mutations.There were three non-synonymous mutations in the encodingα-helix andβ-sheet regions of E6 and E7 protein,respectively.Phylogenetic analysis showed that the HPV52 E6-E7 gene variant sequences were mainly distributed in the B lineage in Jingzhou region.The selection pressure analysis showed that there was no positive selection site in E6-E7 variant sequences.4.Epitope prediction analyses were done.To HLA-A*11:01,the T cell epitope _86-94KTLEERVKK of E6 protein showed the strongest affinity.To HLA-A*02:01,the T cell epitope _84-92MLLGTLQVV of E7 protein showed the strongest affinity.To HLA-DRB1*12:02,the T cell epitope _63-77CIMCLRFLSKISEYR of E6 protein had the strongest affinity.To HLA-DQB1*02:01,the T cell epitope _6-20ATIKDYILDLQPETT of E7 protein had the strongest affinity.The best B cell epitopes for E6 protein were_129-144MGRWTGRCSECWRPRP and _108-123TPLCPEEKERHVNANK,and _23-38HCYEQ LGDSSDEEDTD for E7 protein.Amino acid variation affected the affinity of the predicted epitopes.Conclusion:1.The infection rate of high-risk HPV was 20.13%in Jingzhou region,and the three HPV types with high infection rate were HPV52,HPV58 and HPV16.2.The HPV infection rate of women aged≤24 years and≥55 years were high,and high-risk HPV typing tests should be implemented for women in these age group.If necessary,clinical intervention can be carried out in time.3.There were abundant variation sites of HPV52 E6 and E7 gene sequences in Jingzhou region.The discovery of single nucleotide mutations and the lineage distribution of mutation strains can provide basic data for further understanding the characteristics of the virus variation and for diagnosis and prevention of HPV52 in this region.4.Predicting T cell and B cell epitopes with strong affinity through bioinformatics can provide important information for subsequent study on therapeutic HPV vaccine.