Critical amino acid changes in VP2 variable domain are associated with typical and atypical antigenicity in very virulent infectious bursal disease viruses

Abstract

Classical serotype 1 infectious bursal disease viruses (IBDV), but not very virulent (vv) isolates, react with neutralizing monoclonal antibody (NMab) 3 in virus neutralization tests or antigen-capture ELISA. Two other NMabs, 6 and 8, bind to both classical and most vv strains, but not to the atypical 94,432 and 91,168 vv strains, respectively. The basis for such reactivities was investigated by sequencing the genome region encoding the VP2 major immunogenic domain. In classical, variant, vaccine or vv IBDV strains, negative reactions with NMab3 were associated with changes in the Proline-Glycine pair at amino-acid (aa) positions 222-223 (hydrophilic peak A), and negative reactions with NMabs 6 and 8 with aa changes from positions 318 to 324 (hydrophilic peak B). The 91,168 and 94,432 viruses are the first vvIBDVs to present aa changes in peak B.

Extensive antigenic changes in an atypical isolate of very virulent infectious bursal disease virus and experimental clinical control of this virus with an antigenically classical live vaccine

“…(Bygrave & Faragher, 1970), used as a reference for the ''classical'' IBDVs, and with the 91168 and 94432 atypical vvIBDV strains (Eterradossi et al, 1997b(Eterradossi et al, , 1998. The Faragher 52/70 virus reacted strongly with all mAbs.…”

“…Dots indicate positions where the sequence is identical to F52/70. The sequence of the antigenically atypical vvIBDV isolates 91168 and 94432 (Eterradossi et al, 1998) has been indicated for comparative purposes. VP2 major and minor hydrophilic peaks are boxed with or without grey shading, respectively.…”

“…mAb pairs (3 and 4) and (6 and 7) are targeted at two overlapping antigenic sites, with the binding of mAbs 3 and 4 depending on the presence of Proline and Glycine residues at aa positions 222 to 223 and the binding of mAbs 6 and 7 depending on aa changes at positions 318 to 323. VP2 major hydrophilic peak B is recognized by mAb 8, the binding of which depends on a Glutamine residue at aa position 324 (Eterradossi et al, 1997a(Eterradossi et al, , 1998. mAb 5 probes an epitope involving the first minor hydrophilic peak 1 (249Q) (Eterradossi et al, unpublished results 1998(Eterradossi et al, unpublished results Á/2000.…”

“…Briefly, chimeric oligonucleotide primers, obtained by coupling the sequence of M13 and 21M13 standard primers to the IBDV-specific L2 and U2 sequences, respectively, were used to generate a 604 base pair (bp) reverse transcriptase-polymerase chain reaction (RT-PCR) product, 516 bp of which were IBDV-specific. The RT-PCR products were purified with the Qiaquick Gel purification kits (Qiagen, Chatsworth, CA, USA) and sequenced as described previously (Eterradossi et al, 1998. The resulting nt sequences were submitted to the EMBL database under accession numbers AJ583500 and AJ632141 for the 99323 and CEVAC IBD L viruses, respectively.…”

“…Micro virus neutralization (VN) tests were performed as described previously (Eterradossi et al, 1992): serial two-fold dilutions of the sera were mixed with 100 median tissue infective doses (TCID 50 ) of the CT IBDV strain (serotype 1 antigen, cell culture adapted; final volume of mix, 50 ml). The classical antigenicity of strain CT and the nt sequence encoding the variable domain of VP2 in this virus strain have been described previously (Eterradossi et al, 1997a(Eterradossi et al, , 1998. Virus neutralization was allowed for 45 min at room temperature.…”