The Detection And Preliminary Purification Of Enterovirus 71

Hand-foot-mouth disease (HFMD) is a commom enteroviral infection, mostly caused by coxsackievirus A16 and enterovirus 71(EV71).Young childern are the most frequent shedders of enterovirus.Expect HFMD , EV71 may also cause severe neurological disease, such as encephalitis and poliomyelitis-like paralysis.Since 2008, our country has several large outbreaks of hand, foot, mouth disease,and it seriously affected people's health and lives.There are no diagnostic reagents and effective vaccine against EV7l,so the research of EV7l is very important to the development of EV7l diagnostic reagent and vaccine.The purpose of this study is to explore the preparation course of inactiated accine of EV71, work are as follows: we buiId a panel of enterovirus through virus isolation and culture,and establish the technology platform of virus isolation and identification; To bulid the metheds of Nested PCR, which is to identify EV71 and detect the mycoplasma; The preliminary purification of virus from the harvest of EV71 . The results of this study will provide a important tool and a foundation for the prevention and control of EV71.Virus were isolated from clinical samples stool. Complete VP1 gene sequence (the frist nucleotide sequences for coding capsid proteins of EV71) for recent 9 EV71 isolates of Anhui Province and Hennan Province were determined and compared with that of A,B,C genotype reference strains and some strains anciently isolated from China available from GeneBank by homogeneity and phylogenetic tree analyses. 9 strains isolated by us are all belong to C4 subgenogroup of EV71.Three strains of Anhui Province(H7,H8,H9)share high nucleotide homology with one another(≥98.8%), and H7,H9 are the same with eath other. Three strains of Henan Province(H3,H4,H6)also share high nucleotide homology(≥99.6%), two of them(H3,H4)have a homology of 100%, they also have high homology with H1 and H5 that belong to the strains of Henan Province .One of Henan Province strains(H2)has high homology with the other strains of Henan Province strains, but the phylogenetic analysis based on VP1 demonstrates that H2 also has high homology with the strains of Anhui. Maybe the increasing incidence of mutation of Anhui strains induces the frist outbreak and prevalence of HFMD, and H2 may drives from Anhui Province.In the course of the isolation and indetifiation of EV71 , we bulid the metheds of Nested PCR, which is to identify EV71 and detect the mycoplasma. In this reSearch,we buiId a panel of enterovirus through virus isolation and culture,which are all isolated from our country. We can do a large scale of EV7l production,concentration and purincation.Now we have achieved EV7l vims with ahigh titIer and purity.This basic work is useful for the later EV7l research and vaccine development.