Sequence Analysis of HN Protein Gene of Mesogenic Newcastle Disease Virus from Recent Outbreaks in Namakkal
R. Vetrivel K. Shoba G. Ravikumar S. Vairamuthu T. R. Gopalakrishnamurthy
Vol 8(5), 302-312
Proventriculus, caecal tonsil, spleen, brain were collected from twelve Newcastle disease (ND) suspected layer flocks from Namakkal. The tissue samples were processed and Haemagglutination test (HA) was conducted. The tissue sample were subjected to RT-PCR targeting M gene of RT-PCR (product size-145bp) and those found positive were used for virus isolation. Virus isolation was done in 9-11 day old embryonated chicken eggs and the allantoic fluid of the embryos that showed characteristic lesion NDV like haemorrhage in occipital region and extremities were harvested and was screened by HA, HI and FPCS for confirmation of the virus. Out of eight isolates, three isolates were characterised as mesogenic by Intra cerebral Pathogenecity index and mean death time. Two mesogenic isolates TN/NKL/F1, TN/NKL/S2 were subjected to detail study. Based on the fusion protein cleavage site sequence the virulence nature of the NDV was confirmed. The coding sequence (cds) of FPCS , mesogenic viruses had an amino acid sequence of 112 RRQKRF 117. For HN gene study the allantoic fluid was ultra centrifuged at 32000 rpm for 3h and the viral pellet was used for RNA extraction. The amplified product size of 1892bp was purified and cds was subjected to sequencing which comprised of 577 amino acids. The two mesogenic isolates were found to have 99% homology with Komarov and 93% homology with R2B. The two isolates had Arginine at 362 position whereas Komarov has Glycine at that position.
Keywords : HN ICPI FPCS NDV MDT Mesogenic Sequencing
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