Preliminary characterization of an epitope involved in neutralization and cell attachment that is located on the major bovine rotavirus glycoprotein

Abstract

The 38,200-molecular weight (unreduced)/41,900-molecular-weight (reduced) glycoprotein of bovine rotavirus, isolate C486, was identified as the major neutralizing antigen. This glycoprotein as well as the corresponding glycoprotein of another bovine rotavirus serotype also specifically attached to cell monolayers under normal conditions for virus adsorption in vitro. Further support for this glycoprotein being directly responsible for virus attachment to cells was that (i) infectious virus of both serotypes could compete with the C486 glycoprotein for cell surface receptors, and (ii) neutralizing monospecific antiserum and neutralizing monoclonal antibodies directed toward the glycoprotein could block this virus-cell interaction. Preliminary epitope mapping of the glycoprotein with monoclonal antibodies further localized the neutralization-adsorption domain to a peptide with an approximate molecular weight of 14,000. The effect of two protein modifications, glycosylation and disulfide bridging, on the reactivity of this peptide with antibodies and cell surface receptors was investigated. It was demonstrated that, whereas glycosylation did not appear to affect these reactivities, disulfide bridging seemed to be essential.

Establishment of fetal bovine intestinal epithelial cell cultures susceptible to bovine rotavirus infection

“…This suggests that mild trypsin-EDTA treatment did not have a significant effect on epithelial cell infection by rotavirus. Furthermore, rotavirus infection of epithelial cells requires that rotavirus be cultured in the presence of trypsin or the virus must be treated with trypsin prior to addition to the culture (Sabara et al, 1985;Macartney et al, 2000;Lee et al, 1998;Vijay-Kumar et al, 2005;Cuadras et al, 2002;Bugarcic and Taylor, 2006). These studies indicate that trypsin treatment is required for epithelial cell infection by rotavirus.…”

“…In the present study we used a BRV lab-adapted strain C486, in contrast to the wild-type strains used by others, to infect the cultured bovine intestinal epithelial cell. This strain has previously been used at a high MOI to infect epithelial cultures (Lee et al, 1998;Sabara et al, 1985). Therefore, we used BRV strain C486 at MOIs of 30 and 50 for infectivity and cytopathic effect (CPE) studies, respectively.…”

“…BRV infected and mock-infected cell cultures were examined with an inverted light microscope at 24 and 48 h pi to determine if there were focal areas of cell destruction which would be indicative of viral-induced cytolysis. At 48 h pi culture supernatants from both jejunal and ileal cultures were collected and the amount of infectious virus in the culture supernatant was determined with a plaque assay which used MA104 cells (Sabara et al, 1985).…”

Entry kinetics and mouse virulence of Ross River virus mutants altered in neutralization epitopes

“…Studies with picornaviruses (3,8,32), orthomyxoviruses (44), and rotaviruses (34) suggest that the neutralization antigenic sites of these viruses are involved in virus-cell interaction. Recent studies with a number of alphaviruses also indicate a role of the neutralization domain in early virus-cell interactions, receptor recognition, and pathogenesis (21,26,36,38).…”

Applications of Monoclonal Antibodies in Animal Health and Production

“…MAb technology has been applied to most of the major pathogens that cause scours in neonatal calves, piglets and lambs. Greenberg et al (1983) demonstrated that MAbs against the 42kDa major structural protein of rotavirus could be used for preliminary serotyping of strains from different animal species, and Sabara et al (1985) used MAbs to map the epitope on this glycoprotein that is involved in neutralization of bovine rotavirus and virus attachment to cells during infection. Sonza et al (1983) and Thiriart et al (1986) also produced neutralizing MAbs against rotavirus, whilst direct detection of rotavirus in porcine faecal specimens was achieved by Liprandi et al (1986), who used their MAb against the 45 kDa group specific antigen in a double sandwich indirect ELISA system.…”