The strain-specific epitopes identified on FMDV serotype A A/AF72 strain and A/GDMM/2013 strain.

The antibody-driven variations on A/AF72 strain (A) and A/GDMM/2013 strain (B) were separately determined by selection of neutralization-escape mutants using 20 A/AF72-specific and A/GDMM/2013-specific neutralizing mAbs individually. The proportion of pin chart indicated each mAb-driven variation accounted for the five known antigen sites encompassing VP1 GH loop and C-terminus (site 1/5), VP1 B-C loop (site 3), VP2 B-C loop (site 2) and VP3 B-B knob (site 4), as well as the other unidentified site. (C) Amino acid sequence alignment of VP3 of A/AF72, A/WH/CHA/09 and A/GDMM/2013 strains. The VP3 68 and 175 positions that formed strain-specific epitopes were framed with black oval circles. (D) Immunofluorescence analysis of rescued VP3 68 (T→A) mutant that was constructed basis on entire P1 gene of A/GDMM/2013 strain. BHK-21 cells were infected with the rescue mutant or wildtype virus (A/GDMM/2013) at an MOI of 10 for 4 h. FMDV protein 3A was detected using mouse mAb 3A24 and an Alexa Fluor 561-conjugated secondary antibody. (E) The neutralization efficacy of the A/AF72-specific mAbs W3 and W72 against wildtype (A/GDMM/2013) and its mutant (VP3 T68A) was evaluated using a microneutralization assay. The neutralization concentration represents the lowest antibody required to fully prevent CPE. ** indicates a significant difference compared to wildtype at P<0.01.