Bovine Rotavirus and Bovine Coronavirus are the most important causes of diarrhea in newborn calves and in some other species such as pigs and sheep. VP8 subunit of rotavirus is the major determinant of the viral infectivity and neutralization. Spike glycoprotein of coronavirus is responsible for induction of neutralizing antibody response. Studies showed that immunoglobulin of egg yolk (IgY) from immunized hens has been identified to be a convenient source for specific antibodies for using in immunotherapy and immunodiagnostic to limit the infections.
In this study, chimeric VP8-S2 gene was designed using by computational techniques. The chimeric VP8-S2 gene was cloned and sub-cloned into pGH and pET32a (+) vectors. Then, recombinant pET32a-VP8-S2 vector was transferred into E. coli BL21 CodonPlus (DE3). The expressed protein was purified by Ni-NTA chromatography column. Hens were immunized with the purified VP8-S2 protein three times. IgY was purified from egg yolks using polyethylene glycol precipitation method. Activity and specificity of anti-VP8-S2 IgY were detected by dot-blotting, Western-blotting and indirect ELISA.
We obtained anti-VP8-S2 IgY by immunizing hens with the recombinant VP8-S2 protein. The anti-VP8-S2 IgY was showed to bind specifically to the chimeric VP8-S2 protein by dot-blotting, Western-blotting analyses and indirect ELISA. The result of this study indicated that such construction can be useful to investigate as candidates for development of detection methods for simultaneous diagnosis of both infections. Specific IgY against the recombinant VP8-S2 could be recommended as a candidate for passive immunization against bovine rotavirus and bovine coronavirus.
Abraham S, Kienzle TE, Lapps W, Brian DA (1990) Deduced sequence of the bovine coronavirus spike protein and identification of the internal proteolytic cleavage site. Virology 176: 296-301.
Acres SD, Babiuk LA (1978) Studies on rotaviral antibody in bovine serum and lacteal secretions, using radioimmunoassay. J Am Vet Med Associ 173: 555-559.
Bradford MM (1976) A rapid and sensitive for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248-254.
Favacho AR, Kurtenbach E, Sardi SI, Gouvea VS (2006) Cloning, expression and purification of recombinant bovine rotavirus hemagglutinin, VP8*, in Escherichia coli. Protein Expr Purif 46: 196-203.
Fernandez FM, Conner ME, Hodgins DC, Parwani AV, Nielsen PR, Crawford SE, Estes MK, Saif LJ (1998) Passive immunity to bovine rotavirus in newborn calves fed colostrum supplements from cows immunized with recombinant SA11 rotavirus core-like particle (CLP) or virus-like particle (VLP) vaccines. Vaccine 16: 507-516.
Fiore L, Greenberg HB, Mackow ER (1991) The VP8 fragment of VP4 is the rhesus rotavirus hemagglutinin. Virology 181: 553-563.
Han S, Zhang X, Zhao J (2012) Production of Egg Yolk Antibody (IgY) against Recombinant Canine Parvovirus VP2 Protein. Acta Scientiae Veterinariae 40: 1029.
Ikemori Y, Ohta M, Umeda K, Icatlo FC Jr, Kuroki M, Yokoyama H, Kodama Y (1997) Passive protection of neonatal calves against bovine coronavirus-induced diarrhea by administration of egg yolk or colostrum antibody powder. Vet Microbiol 58: 105-111.
Jayaram H, Estes MK, Prasad BV (2004) Emerging themes in rotavirus cell entry, genome organization, transcription and replication. Virus Res 101: 67-81.
Kapikian AZ, Shope RE (1996) Rotaviruses, Reoviruses, Coltiviruses, and Orbiviruses. In: Baron S (ed.) Medical Microbiology. Galveston (TX) University of Texas Medical Branch at Galveston, chapter 63.
Kirkwood CD (1985) Genetic and antigenic diversity of human rotaviruses: potential impact on vaccination programs. J Infect Dis 202 (Suppl): S43-S48.
Kobayashi C, Yokoyama H, Nguyen Sa Van, Kodama Y, Kimata T, Izeki M (2004) Effect of egg yolk antibody on experimental Cryptosporidium parvum infection in scid mice. Vaccine 23: 232-235.
Kovacs-Nolan J, Sasaki E, Yoo D, Mine Y (2001) Cloning and expression of human rotavirus spike protein, VP8*, in Escherichia coli. Biochem Biophys Res Commun 282: 1183-1188.
Larralde G, Gorziglia M (1992) Distribution of conserved and specific epitopes on the VP8 subunit of rotavirus VP4. J Virol 66: 7438-7443.
Liou JF, Shiau JW, Tai C, Chen LR (2011) Production of egg yolk immunoglobulin against Escherichia coli from white Leghorn and Lohmann chickens. J Anim Vet Adv 10: 2349-2356.
Mine Y, Kovacs-Nolan J (2002) Chicken egg yolk antibodies as therapeutics in enteric infectious disease: a review. J Med Food 5: 159-169.
Parreño V, Marcoppido G, Vega C, Garaicoechea L, Rodriguez D, Saif L, Fernhndez F (2010) Milk supplemented with immune colostrum: protection against rotavirus diarrhea and modulatory effect on the systemic and mucosal antibody responses in calves experimentally challenged with bovine rotavirus. Vet Immunol Immunopathol 136: 12-27.
Polson A, Von Wechmar MB, Van Regenmortel MH (1980) Isolation of viral IgY antibodies from yolks of immunized hens. Immunol Commun 9: 475-493.
Saif LD, Redman DR, Brock KV, Kohler EM, Heckert RA (1988) Winter dysentery in adult dairy cattle: detection of coronavirus in the faeces. Vet Rec 123: 300-301.
Takase-Yoden S, Kikuchi T, Siddell SG, Taguchi F (1991) Localization of major neutralizing epitopes on the S1 polypeptide of the murine coronavirus peplomer glycoprotein. Virus Res 18: 99-107.
Vega C, Bok M, Chacana P, Saif L, Fernandez F, Parreño V (2011) Egg yolk IgY: protection against rotavirus induced diarrhea and modulatory effect on the systemic and mucosal antibody responses in newborn calves. Vet Immunol Immunopathol 142: 156-169.
Yoo D, Lee J, Harland R, Gibbons E, Elazhary Y, Babiuk LA (1997). Maternal immunization of pregnant cattle with recombinant vp8 protein of bovine rotavirus elicits neutralizing antibodies to multiple serotypes. Colostral neutralizing antibody by rotavirus vp8*. Adv Exp Med Biol 412: 405-411.