Molecular Epidemiology Of Newcastle Disease Viruses Isolated In China During 2018-2020 And Development Of New Vaccines Against The Prevalent Virulent Viruses

Newcastle disease(ND)is an acute and highly contagious disease caused by a virulent strain of Newcastle disease virus(NDV)and mainly infects birds.The Office Internationale Des Epizooties(OIE)lists it as a notifiable disease,and the Ministry of Agriculture of China lists it as the first class animal infectious disease.Since its first outbreak in 1926,NDV has already caused at least four panzootics worldwide,which not only posed a seriously threat to global poultry industries,but also caused huge economic losses.In recent years,due to the wide use of ND vaccine and the continuous improvement of poultry industry scale in China,ND has been effectively controlled throughout the country.However,due to the high genetic diversity,the antigenic variation and the wide host range of NDV,the prevention and control of ND in China is still in a severe status.In order to elucidate the current molecular epidemiological characteristics of NDV,we conducted a molecular epidemiological study on NDVs isolated in China during 2018 to 2020.Additionally,we developed the ND vaccines against the sub-genotype ?h,genotype ? and genotype ?? NDVs,which are circulating in Asian presently,to prevent the emergence of these genotypes in China.1.Molecular epidemiology of Newcastle disease viruses isolated in ChinaDuring 2018 to 2020,a total of 37 NDV strains were isolated from the live poultry markets and clinical samples in china.Thirty NDVs were isolated from the live poultry markets including 25 NDVs from chicken flocks,2 from goose and 3 from duck flocks.Among these isolates,26 lentogenic NDVs belongs to class ? genotype 1.1.2;five lentogenic strains belongs to class ?genotype ? and five lentogenic strains belongs to class ? genotype ?.Notably,one NDV strain,isolated from geese in 2020,belongs to class ? genotype ?,and this genotype was first isolated from Guangdong in 2010.The result of cross hemagglutination inhibition test showed significant antigenic difference between the genotype ? NDV isolate and the existing ND vaccine strains.The results mentioned above indicate that although the main NDV isolates in China are lentogenic strains,the virulent NDV could still be isolated.Remarkably,these virulent NDV isolates show different genotype and antigenicity from the commericial NDV vaccines,which are used widely in China.Therefore,it is necessary to develop new vaccines to prevent these genotype NDVs spread in China.2.Development of vaccines against the prevalent NDV infectionAccording to the phylogenetic analysis of NDV prevalent strains in Asia in recent years,the gene sequences encoding the ectodomain of F and HN protein from NDV LX strain were replaced with the corresponding genomic regions from genotype ?h,genotype ? and genotype ?? respectively.Three attenuated recombinant NDVs,NDV/LX-?hFHN,NDV/LX-?FHN and NDV/LX-??FHN,with high propagation ability were generated successfully by reverse genetics.The inactivated and live vaccines were made based on these three recombinant NDVs respectively and were injected into 3-week-old SPF chickens.Blood samples were collected regularly for hemagglutination inhibition(HI)test.Three weeks post-immunization,the groups vaccinated with NDV/LX-?hFHN,NDV/LX-?FHN were challenged with the genotype ?h and ? virulent NDV strains,respectively(no genotype ??NDV strain available).On the day 3,5,and 7 post-challenge,clocal and orpharygeal swabs were collected for virus isolation.The results showed that the titers of HI antibody induced by the live and inactivated vaccines were 6 log2 and 8 log2,respectively,and all of the immunized chickens were completely protected from mortality following virulent strains challenge.Compared with the commercial vaccines,the novel vaccines made of NDV/LX-?hFHN and NDV/LX-?FHN could provide better protection following the genotype ?h and ? virulent NDVs challenge,respectively,interms of the virus sheding.Taken together,the three recombinant viruses are suitable for vaccine preparation and can effectively eliminate the risk of ND spread caused by these genotype viruses which are circulating in Asia at present.