Study On Immune Effect Of Hand-foot-and-mouth Disease Virus VP1Protein And Chimeric Antigen Of Bacterial Ghost

Hand-food-and-mouth disease (HFMD) is a global infectious disease caused byintestinal viruses, which have different scale outbreak all over the world,In china, thisdisease occured since1981in Shanghai and in rescent years Beijing, Hebei, Tianjin,Fujian, Jilin, Shandong, Hubei, Anhui, Hubei, Guangdong and other more provinces andcities have reported。HFMD occurs more often in children under5years old, It is report acase to the infant, can cause hand, foot, mouth and other parts of the simplex, a fewpatients can cause myocarditis, pulmonary edema, aseptic meningitis and othercomplications, individual children with severe could even death, HFMD has become apublic health problem in the world, has attracted wide attention.In order to prevent andcontrol the foot and mouth disease, China’s Ministry of health list hand foot and mouthdisease in the infectious disease prevention law class C infectious disease management.At present, effective antiviral drugs still is lack in the whole world, developing aneffective vaccine is the most effective control strategy of HFMD infections. In this study,two kinds of pathogenic foot-and-mouth disease virus infection, design and evaluation ofa series of candidate vaccine molecules, which provid new strategy to prevent HFMD。1. The immune effect and cross protection of VP1antigenHFMD mainly caused by virus including enterovirus71and Coxsackievirusgroup.Studies have shown that VP1protein as antigen of the virus in the murine modelfor viral infections can provide a protective effect.In this study, we express recombinantenterovirus71and Coxsackievirus B3virus VP1protein by prokaryotic expressionsystem respectively. The EV71antigen VP1(EVP1), CB3antigen VP1(CVP1) as antigen single or mixed immunogen of adult BALB/C mice were immunized, inducethe specific antibody titer higher. Serum virus neutralizing activity tests showedneutralizing titer of EV71virus immune serum EVP1group is about1:832, CVP1immunoreactivity in blood to the CB3virus neutralization titer:1:1024;In the challengetest on the offspring neonatal rats, two VP1antigen can provide better protective effecton specific virus, EVP1immune group on160LD50EV71virus infection can providecomplete protection, CVP1immune group of150LD50CB3virus attack protection rateof88.9%; To improve toxic dose, the protective survival rate of70%of EVP1antigen forthe200LD50EV71virus, CVP1antigen of180LD50CB3virus attack protectionsurvival rate was75%, Further pathologic findings compared with the control group ofserious injury, immune protection group pathological organs injury in mice reduced ordisappear.In cross protection tests of two kinds of antigens, found CB3virus protectioneffect of EVP1immune group of150LD50up to60%, while the CVP1immune groupof high dose of EV71virus no protective effect, but at a low dose of EV71virus (15LD50) only27.3%protective effect of attack.In addition, we also successfully constructed with EVP1and the molecular structureof CVP1fusion protein ECVP1. The nickel column chromatography to obtain targetprotein, and verified the immune reaction, can also be antiserum identified two types ofviruses.2. Research on construction and immune effect of two VP1antigen ghostIn this study, using the laboratory of gram negative bacteria universal sandwichmembrane construction display vector pSOMPA, success induced EVP1or CVP1molecules in O157:H7ghost membrane anchor expression. In the newly constructedchimeric bacterial ghosts (EBGs, CBGs) structure, the O157:H7bacterial membranesurface antigen with the new EV71protective antigen EVP1or CB3protective antigenexpression of CVP1in bacterial ghost membrane.Two chimeric ghost mice immunized mice induced to produce not only specific antibodies against O157:H7bacteria (antiintimin antibody), also induces the production of specific antibodies against VP1antigen.As compared to the negative control, chimeric ghost can not only protect the miceagainst20MLD E.coli O157:H7live stomach attack by bacteria (70%and75%), theoffspring of mice against lethal dose produced by immune (2LD50) EV71or CB3infection have a protective effect, EBGs immune group can provide complete protection(100%), CBGs immune group also can provide a higher rate of protection (87.5%);Toimprove the EV71attack protection challenge dose of EBGs can resist the5LD50ratewas50%, CBGs immune group of CB3virus attacks to protect against5LD50rate was37.5%。Therefore, the VP1antigen expression of chimeric O157:H7immunizationstrategy can make the bacterium protective O157:H7pathogenic organism infection, butalso can provide immune protection against EV71and CB3viruses, can provide the basisfor the development of new vaccine.