Molecular Evolution Of Coxsachievirus A16 From Hand, Foot And Mouth Disease Based On Bayesian Evolutionary Method

Background and Objective:Hand-foot-mouth disease(HFMD) is a C class infectious disease with the highest morbidity and mortality in China in recent years. Minority patients have severe neurologic complications such as acute flaccid paralysis, encephalitis, and pulmonary edema, individual severe patients even death. As the second generation sequencing technology and the rapid development of computer science and technology, bioinformatics is widely used to identify and recognize the evolution of the virus. The best nucleotide substitution model can be obtained according to the prior probability based on Bayesian-Markov chain method. With application of posterior probability optimization standard, the rapidly process of large data and complex model will be done. Currently, this method has been used on study of virus’ system development by many researchers. The viruses include EVA71, JEV, HBV, MV, AV and etc. Coxsackie virus A16(CVA16) accounts for about 50% of the pathogens in China according to the continuous monitoring. It’s one of the major pathogens of HFMD and has been reported in clinical death. Relative to the situation that EVA71 vaccine has entered the clinical application, current research on CVA16 is not enough. Therefore, we applied the Bayesian method for molecular evolution of CVA16, so that scientific and reasonable explanation can be made on CVA16’s genotyping and regions epidemic at molecular level.It also provide a new perspective for the development of vaccine.Material and Method: This study isolated and identified 20 strain enteric viruses, and 6 strains of CVA16 VP1 gene were amplified and sequenced. We downloaded and sorted 708 currently published CVA16 VP1 gene from the Genbank database and constructed the largest CVA16 VP1 gene data set by far. The restructure detection was done by recombination detection program(RDP3) and the saturation was detected by DAMBE software. The best nucleotide substitution model was calculated by JModeltest. Bayesian-Markov analyses was performed to construct a maximum clade credibility(MCC) tree for CVA16 VP1 gene data set. The evolution characteristics of CVA16 VP1 gene was also analysed dynamicly through Bayesian skyline plot.Results and Conclusion : Among the 6 CVA16 VP1 genes we sequenced, the variation rate of 891 nucleotide is 3.36% and the variation rate of 296 amino acid is 1.68%. It can be seen that the rate of amino acid variation rate is slightly lower than the level of the nucleotide. The reconstruction detection results showed that there was no recombination between the strains, and the saturation monitoring results showed that the sequence replacement was not saturated. Therefore, it’s suitable for the construction of phylogenetic tree. After calculation, the optimal nucleotide substitution model was GTR. Data preprocessing results show that our data set is suitable for the next evolutionary analysis.All CVA16 strains identified could be classified into five major genogroups, denoted by GI–GV. Further, the genogroup GIV could be divided into sub-genogroups GIV-1, GIV-2, and GIV-3; All of CVA16 epidemic strains belonged to genogroup GII in 1981-1993 in Japan. Thereafter, genogroup GIII became the new epidemic strain until 1995. Later, genogroups GIV and GV replaced GIII and became the predominant genogroups since 1997; 81 CVA16 epidemic strains from Malaysia were collected, of which genogroup GV was the most dominant, accounting for 83%(68/82, 83%); GIV(195/315,62%) and GV(118/315,37%) have co-circulated in China since 2007, and the CVA16 epidemic strain extant in the Jilin province, China, can be classified as GIV-3. The CVA16 serotypes circulating recently in China have the same ancestor, suggesting that the epidemic strains are native and have not invaded the region since 2007.The genetic diversity dynamics of the CVA16 VP1 gene shows a continuous increase since the mid-1990 s, with sharp increases in genetic diversity in 1997 and 2007 and reached peak in 2007. Very low genetic diversity existed after 2010. The related literature reports and MCC are all shown that the increase of genetic diversity corresponding to the emergence of new genogroup. And the emergence of a new genogroup always leads to a large-scale HFMD outbreak. So genetic diversity was associated with the HFMD outbreak trends which infected by CVA16.The CVA16 VP1 gene evolutionary rate was estimated to be 4.0718E-3 substitutions per site per year inferred by the models of GTR. All three codon positions of the CVA16 VP1 gene had different relative substitution rates, and relative substitution rate of the third codons was highest.