Expression and evaluation of immune-potential of recombinant penton base and fibre proteins of Inclusion body Hepatitis-Hydropericardium Syndrome (IBH-HPS) virus of domestic fowl
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Date
2018-07
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G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand)
Abstract
Present study deals with epidemiology and phylogenetic analysis, expression and immune potential of recombinant penton base and fibre proteins of IBH-HPS virus isolates recovered from natural outbreaks. Epidemiological analysis revealed that mortality ranged from 0.5% to 20% and disease outbreak was mainly observed in 21 to 42 days old broiler chickens. Twenty samples, found positive for IBH-HPS virus antigens serologically by AGID, were subjected to virus isolation in CEL culture. Followed by DNA was extracted from infected CEL cells (4th passage). Amplification of L1 loop region (~900bp) of hexon gene was done followed by RFLP analysis with six restriction enzymes which revealed two RFLP pattern. L1 loop region of hexon gene amplicon was cloned and sequenced to determine serotype of IBH-HPS isolates, which indicated that nineteen isolates are FAdV- 11/2 while one is FAdV-8b. L1 loop sequence based phylogenetic analysis revealed that our sequences clustered into one major cluster having similarity with the Fowl adenovirus class D, E and C along with strains FAdV-2, 4, 10, 11, 8b. Penton base and fibre genes from selected isolate (Pantnagar/HA-14/R-21) were PCR amplified and amplicons of about 1713bp and 1668bp respectively were obtained. Cloning and sequencing of the penton base and fibre gene was done and checked for the presence of complete ORF. Both the genes were expressed in E.coli and expressed proteins were found to be immunoreactive in western blotting with known serum. Immune – potential of recombinant proteins was evaluated by inoculating with FCA in SPF chicks. Maximum protection (80%) was observed with fibre protein and antibody titre was maximum for fibre protein at every point of time. Virus shedding was observed in feces for shortest period i.e. upto 8th day post challenge in chicks receiving fibre protein.
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