Analysis Of Complete Genome Characterization Of Shandong Local Strains Of Human Papillomavirus Type 6

BackgroundHPV6 is the most common gene subtype in the prevalence of HPV,which is distributed all over the world.Among HPV positive people,the infection rate of HPV6 is the highest,and the infection rate of HPV6 in women with normal cytology is about 0.8%.HPV6 often causes condyloma acuminatum in the external genitalia and perianal area,and is also related to Buschke Lowenstein tumor and.In addition,some HPV6 variants are classified as"carcinogens",such as tonsil cancer,lung cancer,rare Verrucous Cancer of vulva and penis,and other respiratory and reproductive tract cancer and/or malignant laryngeal papilloma lacking specific promoters.Due to the importance of HPV6,it has been successively covered in tetravalent and nine valent HPV vaccines.So far,the research on the whole genome of HPV6 is still in its infancy,many countries,including China,lack big data and high-quality and complete information reports on the whole genome sequence of HPV6.Objectives1.The HPV6 genome sequence information of patients with condyloma acuminatum in Shandong Province from 2019 to 2021 was obtained,and the variation and homology were analyzed at the molecular level.2.To explore the variation lineage composition of HPV6 in Shandong Province and clarify the dominant(sub)lineage of prevalence.3.According to the molecular variation of HPV6 genome and the distribution characteristics of corresponding polymorphic sites,the differences between different(non)coding regions were analyzed.4.To analyze the sequence variation between different lineages of HPV6 Shandong local strains and the variation characteristics of representative sequences,and understand the variation characteristics of HPV6 prevalent in Shandong in recent years,so as to provide basis for ensuring the sequence correctness of vaccine VLP.Methods1.Collection and preservation of HPV samplesThe samples were from the pathological tissues of Condyloma Acuminatum Patients in three sentinel hospitals in Shandong Province from 2019 to 2021.2.HPV DNA extractionVirus DNA is extracted by Tianlong technology automatic nucleic acid extractor and supporting virus DNA extraction reagent3.PCR amplification and electrophoresis detectionThe extracted virus DNA was amplified by PCR and the PCR product was agarose gel electrophoresis of 1%.4.Sequencing and gene analysis of PCR productsThe positive product was sent to Beijing LiuheHuada gene Biotechnology Co.,Ltd.for sequencing to obtain the full length of HPV6 genome.Sequence alignment and homology analysis were carried out,and the phylogenetic tree was constructed.Results1.Epidemiological data show that the patients with HPV6 condyloma acuminatum are mainly male(male to female ratio 2.6:1),and the age is mainly 20～40 years old.2.Sixty-four full-length HPV6 sequences were successfully obtained,belonging to lineages A,B1 and B3,with the number of 25,38 and 1 respectively.3.There were 935 SNVs in 64 sequences at 177 locations,and the missense mutation rate was 27.40%.455 SNVs occurred in 25 lineage A sequences,and the missense mutation rate was 15.40%(70/455);471 SNVs occurred in 38 lineages B1 sequences,and the missense mutation rate was 38.85%(183/471);Nine SNVs occurred in B3 sequence of one lineage,and the missense mutation rate was 22.22%(2/9).4.The maximum pairing difference level is the highest in E5B and E5 A and the lowest in L1.The number of SNV occurred was the most in E1 and the least in E7 and E4.The number of amino acid variants L1 and E1 was the most,and E6,E4 and E7 were the least.Conclusions1.In recent years,there are three variation lineages of HPV6 Shandong local strains.The most popular is sublineage B1,followed by variant lineage A,while the prevalence of sublineage B3 is low.2.Among ORFs of HPV6,the variability of E1,L1,L2 and LCR was high,E2,E5A and E5B were in the middle level,and E6,E7 and E4 were relatively conservative.3.The nucleotide mutation rate of single sequence and SNV site lineage A is higher than B1,but the amino acid mutation level A is more conservative than B1,that is,the missense mutation rate B1 is higher than A.4.Representative variant sites of HPV6 Shandong popular strains were obtained through the representative variant sites identified in the actual cases of lineages A and B1.