Isolation And Identification Of Genotype VII NDV Isolates And Cross-protective Immunity Test With Lasotastrain

Newcastle disease is a potent, highly contagious disease, which is caused by theNewcastle disease virus against the global poultry industry. Although NDV only has oneserotype, Newcastle disease outbreaks still occur frequently in flocks vaccinated with NDVlive vaccines (e.g. La Sota, Clone30) and inactivated vaccines in China. Immune failure isconsidered to be related to the genetic variation of NDV epidemic strains.On the basis of restriction fragment length polymorphism analysis and partial nucleotidesequencing of the cleavage recognition sequence of the F gene, NDV strains can be classifiedinto ten genogroups (named, I~X), among which genogroups VI and VII can be furtherdivided into several sub-genotypes. On the basis of pathogenicity, NDVs have beencategorized into velogenic, mesogenic, and lentogenic pathotypes. The F protein mediates thefusion of viral and cellular membranes during penetration and spread between infected andadjacent cells. The important F gene areas (1~389bp) nucleotide sequence of NDV F geneare related to viral pathogenicity and through the analysis of cleavage site sequence of Fprotein the virulence of Newcastle disease virus can be predicted. The viral HN proteinmediates attachment to sialic acid-containing receptor(s) and, via its neuraminidase (NA)activity, the apparently opposing activity of release of sialic acid from soluble andmembrane-associated glycoconjugates. HN is a type II membrane glycoprotein, existing onthe surface of virions and infected cells as a tetrameric spike. The ectodomain of the HNglycoprotein consists of a membrane-proximal, stalk-like segment supporting a terminalglobular domain. The antigenic, receptor recognition, and NA active sites all reside in thelatter. Thus, analysis of genetic variation of NDV’ F and HN genes can serve as thetheoretical basis to help predict the prevalence of ND effectively. The main results of thisstudy as follow:1. Several ND-suspected field lungs, tracheal rings and other organizations of the deadchickens from nine different poultry farms were collected from Shaanxi Province, China,from September,2011to December,2011. Nine Shannxi field isolates were classified as VIIdby serological and molecular methods. 2. On the basis of nucleotide sequences of the F gene among NDV isolates, aphylogenetic tree was constructed. Nine Shannxi field isolates had distant kinship with thetraditional vaccine strains, such as Mukteswar, Clone30, La Sota, B1, Sato/30and Herts/33.Analysis of the deduced amino acid sequences of the F protein of recent Chinese NDVisolates and the traditional vaccine strains revealed many amino acid residue substitutions atneutralizing epitopes on F protein. Additionally, compared with the pandemic strains in Chinanearly10years, the deduced amino acid sequences of the F protein of recent Shannxi fieldNDV isolates revealed some amino acid residue substitutions at neutralizing epitopes on Fprotein.3. Nine NDV isolates belonged to amino acid residues of571NDVs. Besides, nine NDVisolates were all clustered together with BP01, JSD0812, sh09, chicken/China/JSX1/2010,JS-12-11-Ch and Ch/SD672/12which were representatives of genotype VIId isolates.Comparison of sequences at the cleavage site of the F protein and the C-terminal extension ofthe HN protein indicated that HN gene relationships revealed by phylogenetic analyses werealso maintained in comparisons between the F gene cleavage. Additionally, compared withthe traditional vaccine strains, the deduced amino acid sequences of the HN protein of recentShannxi field NDV isolates revealed many amino acid residue substitutions at neutralizingepitopes on HN protein. This divergence further elucidated that amino acid residuesubstitutions may lead to the change of antibody recognition capabilities and types.4. By comparison of the genome of NDV/Chicken/TC/1/2011with reference strains,shared31.9%~32.0%and89.1%~99.1%identities with genotype I-IV NDV (e.g. La Sota,Clone30, B1, Mukteswar) and genotype V-IX NDV, respectively. Among these results, thehighest homology percent was between NDV/Chicken/TC/1/2011and JSD0812, which was99.1%.5. Cross-protectivity test showed that the isolate aluminum hydroxide gel typeinactivated vaccine with lower antibody level and the resultant isolate inactivatedvaccine+Lasota attenuated vaccine conferred clinical protection to100%of chicks challengedwith NDV/Chicken/TC/1/2011while the single Lasota attenuated vaccine and Lasotainactivated vaccine only confered75%and83.3%clinical protection. It revealed that therewas antigen diversity between NDV vaccine strains and epidemic strains.