Rescue Of Mutated Serotype O Cathay Topotype Foot And Mouth Disease Virus And Study Of Viral Particle Stability

Foot-and-Mouth Disease(FMD),caused by foot-and-mouth disease virus(FMDV),is an acute and fulminating infectious disease mainly infects cloven-hoofed animals such as cattle,sheep and swine.Vaccination,mostly use inactivated vaccines,is one of the critical preventive methods for controlling FMD in epidemic zones.The intact virus particle(146S particle)is essential for FMD inactivated vaccine to protected animal from disease.However,the 146 S particle stability is so poor that mild temperature or pH change will make 146 S particle disassemble into 12 S particles,leading inactivated vaccine failure.Considering 146 S particles have such shortcoming,researchers try to enhance acid,thermal stability of FMDV by amino acid residues substitution based on research results of the molecular structure of the 146 S and screen candidate vaccine strains.According to the result of previous work,this study select the vaccine strains O/GX/09-7 and try to replace amino acid residues of the structural protein VP2 and VP1.During the study,we amplified the P1 region gene of O/GX/09-7 by using the primer which designed for amino acid substitution,then constructed eight cloning plasmids and built up eight recombinant plasmids by inserting the gene into pGEM?-Teasy Vetor and p43-CHA90-Vector which has been built in early time.Using the recombinant plasmids and infectious clones pOZK/93,we transcript the genome RNA and transfected them into BHK-21 cell.Then we successful rescued two virus(vOZK/93 and vOS93H).After that,vOZK/93,vOS93 H and parental virus O/GX/09-7 were treated by heat and acid.Then we measured the TCID50 value and 146 S particle content of three virus and comparing with blank control respectively so that detected the effect of amino acid substitution on enhancing acid,thermal stability of FMDV.There is no obvious difference among vOZK/93,vOS93 H and parental virus O/GX/09-7 of thermal stability and vOZK/93 is similar to parental virus in acid stability.However,the results indicated that the acid stability of vOS93 H is increase compared with parental virus O/GX/09-7.