| Influenza is an acute respiratory infectious disease caused by three types ofinfluenza virus infectied A (A), B (B), propylene (C), respectively,. RNA segments ofinfluenza A and B virus genome consists of eight single-stranded， virus C RNA islack of NA segment.. Different RNA segments encoded different proteins withspecific biological functions, thereby determined the genetic characteristics ofinfluenza viruses.Variation of influenza viruses, included virulence, antigenic variation, and thephase variation. Virulence, included host adaptive mutation, conditional pathogenicmutation and cold-adapted variation, antigenic variation mainly refered to thevariation of HA and NA antigens, mainly included two form antigenic drift andantigenic shift. Because antigens of influenza viruses was mutated easily, which hasrepeatedly aroused a worldwide pandemic catastrophe for humanity.The most effective measures to prevent and control the epidemic of influenza isto inoculate the flu vaccine which can be divided into whole virus influenza vaccine,influenza virus vaccine, influenza virus subunit vaccine. Inactivated influenza vaccinecurrently used mainly for the H1N1, H3N2and influenza B viruses triple vaccine, thatwas safe, and had good immunity.Live attenuated influenza vaccine was obtained after less parental strain (oftencold-adapted strains) of the six internal genes (PB2, PB1, PA, NP, influenza virusattenuated strain M, NS) and the current pandemic strain of two genes (HA and NA)reassortanted, They has the feature both cold-adapted strains of temperature-sensitive,cold-adapted, attenuated features, of the donor but also antigenic characteristics of theepidemic strains. The live attenuated influenza vaccine approved were the UnitedStates FluMistt and the Russia TM LAIV.Influenza viruses are RNA segmental viruseswith high recombinant rate, whichis very easy to change its antigenic mutation. Attenuated strain of influenza vaccinesused to the crowd must ensure the genetic stability of its genes in order to ensure thatthe vaccine is still able to maintain its basic features without making people sick and can produce good immune protection. Therefore, it is very important for production ofattenuated strain influenza virus to its genetic stability of vaccine strain, geneticstability can only ensure the safety and effectiveness of vaccines.The purpose of this experimental study is to investigate genetic stability of liveattenuated influenza vaccine strains usde to the production.We will inoculate A/17/California/2009/38(H1N1), A/17/Perth/09/87(H3N2)and B/56/Brisbane/60/08three influenza virus strains in the chick embryoreduction.All RNA of the strain2,3,5,10were extracted and reversed transcriptioninto cDNA, amplified by PCR to connecte to the vector and construct the plasmid, theplasmid transferred into E. coli clones were cultured, and then sequenced plasmiddelivery.We blasted the genes sequence of PB1, PB2,PA, NP, M, NS six genes fromA/17/California/2009/38, A/17/Perth/09/87, PB2B/56/Brisbane/60/08viruses fourgenerations of12strains, and the genes sequence of their parents attenuated strainsA/Leningrad/134/17/57(H2N2), B/USSR/60/69f and amino acid sequence alignment,We blasted the A/17/California/2009/38, A/17/Perth/09/87, B/56/Brisbane/60/08these three viruses four generations of HA and NA of12strains of these two geneswith WHO Recommended2011to2012corresponding attenuated influenza virusstrains suitable for the northern hemisphere an A/California/07/2009(H1N1)-likevirus, an A/Perth/16/2009(H3N2)-like virus, a B/Brisbane/60/2008-like virus forthe full gene sequence and amino acid sequence alignment. The result showed thatthere occurred mutations of nucleotide and amino acid in the process of passage, butthe mutation rate is very low, the total mutation rate of influenza virus was0.035%,the total mutation rate of influenza B virus was0.022%, this indicated that homologyof the virus strain and strain Outward was greater than99%, and gene keep stabilityafter serially passaged10generations.We blasted the attenuated key sites of nucleotide and amino acid sequences onA/17/California/2009/38, A/17/Perth/09/87, B/56/Brisbane/60/08and attenuatedstraincorresponding sites of their parents A/Leningrad/134/17/57(H2N2), the nucleotideand amino acid sequences B/USSR/60/69. Than the results show that the key sites of various generations attenuatedinfluenza virus strains the nucleotide and amino acid change does not appear. Geneticinstructions within this three strains of the virus implicated resume generations10generations stable, attenuated characteristics have not changed.This experiment can prove A/17/California/2009/38, A/17/Perth/09/87,B/56/Brisbane/60/08these three viruses are genetically stable virus gene, in line withnational influenza live attenuated vaccine virus seed production requirements, theycan guarantee the safety and efficacy of the vaccine in genetic stability. |
PDF Full Text Request