title: Production of recombinant non-structural protein-3 hydrophobic domain deletion (NS3ΔHD) protein of bluetongue virus from prokaryotic expression system as an efficient diagnostic reagent.
abstract: Serological diagnostics for bluetongue (BT), which is an infectious, non-contagious and arthropod-borne virus disease of ruminants, are primarily dependent on availability of high quality native or recombinant antigen(s) based on either structural/non-structural proteins in sufficient quantity. Non-structural proteins (NS1-NS4) of BT virus are presumed candidate antigens in development of DIVA diagnostics. In the present study, NS3 fusion gene encoding for NS3 protein containing the N- and C-termini with a deletion of two hydrophobic domains (118A to S141 aa and 162S to A182 aa) and intervening variable central domain (142D to K161 aa) of bluetongue virus 23 was constructed, cloned and over-expressed using prokaryotic expression system. The recombinant NS3ΔHD fusion protein (∼38 kDa) including hexa-histidine tag on its both termini was found to be non-cytotoxic to recombinant Escherichia coli cells and purified by affinity chromatography. The purified rNS3ΔHD fusion protein was found to efficiently detect BTV-NS3 specific antibodies in indirect-ELISA format with diagnostic sensitivity (DSn = 94.4%) and specificity (DSp = 93.9%). The study indicated the potential utility of rNS3ΔHD fusion protein as candidate diagnostic reagent in developing an indirect-ELISA for sero-surveillance of animals for BTV antibodies under DIVA strategy, wherever monovalent/polyvalent killed BT vaccine formulations devoid of NS proteins are being practiced for immunization.
authors.affiliation: Division of Virology, ICAR-Indian Veterinary Research Institute (IVRI), Regional Campus, Mukteswar, 263 138, Nainital, Uttarakhand (UK), India.
authors.forename: Nihar Nalini
authors.initials: NN
authors.lastname: Mohanty
authors.affiliation: Division of Virology, ICAR-Indian Veterinary Research Institute (IVRI), Regional Campus, Mukteswar, 263 138, Nainital, Uttarakhand (UK), India.
authors.forename: Nirmal
authors.initials: N
authors.lastname: Chacko
authors.affiliation: Division of Virology, ICAR-Indian Veterinary Research Institute (IVRI), Regional Campus, Mukteswar, 263 138, Nainital, Uttarakhand (UK), India.
authors.forename: Sanchay Kumar
authors.initials: SK
authors.lastname: Biswas
authors.affiliation: Division of Virology, ICAR-Indian Veterinary Research Institute (IVRI), Regional Campus, Mukteswar, 263 138, Nainital, Uttarakhand (UK), India.
authors.forename: Karam
authors.initials: K
authors.lastname: Chand
authors.affiliation: Division of Virology, ICAR-Indian Veterinary Research Institute (IVRI), Regional Campus, Mukteswar, 263 138, Nainital, Uttarakhand (UK), India.
authors.forename: Awadh Bihari
authors.initials: AB
authors.lastname: Pandey
authors.affiliation: Division of Virology, ICAR-Indian Veterinary Research Institute (IVRI), Regional Campus, Mukteswar, 263 138, Nainital, Uttarakhand (UK), India.
authors.forename: Bimalendu
authors.initials: B
authors.lastname: Mondal
authors.affiliation: Division of Virology, ICAR-Indian Veterinary Research Institute (IVRI), Regional Campus, Mukteswar, 263 138, Nainital, Uttarakhand (UK), India.
authors.forename: Divakar
authors.initials: D
authors.lastname: Hemadri
authors.affiliation: Division of Virology, ICAR-Indian Veterinary Research Institute (IVRI), Regional Campus, Mukteswar, 263 138, Nainital, Uttarakhand (UK), India. Electronic address: sb.shivachandra@icar.gov.in.
authors.forename: Sathish Bhadravati
authors.initials: SB
authors.lastname: Shivachandra
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001819:Bluetongue / Q000175:diagnosis / Q000276:immunology; D001820:Bluetongue virus / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D004926:Escherichia coli; D015870:Gene Expression; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D012418:Ruminants / Q000276:immunology / Q000821:virology; D017384:Sequence Deletion; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology
chemical_list: D011993:Recombinant Fusion Proteins; C075080:S10 protein, Bluetongue virus; D017361:Viral Nonstructural Proteins
keywords: Bluetongue virus; ELISA; Hydrophobic domain deletion; Non-structural protein 3 (NS3); Prokaryotic expression; Recombinant protein