Antibody signatures in hospitalized hand, foot and mouth disease patients with acute enterovirus A71 infection

Enterovirus A71 (EV-A71) infection is a major cause of severe hand, foot and mouth disease (HFMD) in young children. The characteristics of EV-A71 neutralizing antibodies in HFMD patients are not well understood. In this study, we identified and cloned EV-A71-neutralizing antibodies by single cell RNA and B cell receptor sequencing of peripheral blood mononuclear cells. From 145 plasmablasts, we identified two IgG1 monoclonal antibodies (mAbs) and six IgM mAbs that neutralized EV-A71. Four of the IgM mAbs harbor germline variable sequences and neutralize EV-A71 potently. Two genetically similar IgM antibodies from two patients have recurrent heavy chain variable domain gene usage and similar complementarity-determining region 3 sequences. We mapped the residues of EV-A71 critical for neutralization through selection of virus variants resistant to antibody neutralization in the presence of neutralizing mAbs. The residues critical for neutralization are conserved among EV-A71 genotypes. Epitopes for the two genetically similar antibodies overlap with the SCARB2 binding site of EV-A71. We used escape variants to measure the epitope-specific antibody response in acute phase serum samples from EV-A71 infected HFMD patients. We found that these epitopes are immunogenic and contributed to the neutralizing antibody response against the virus. Our findings advance understanding of antibody response to EV-A71 infection in young children and have translational potential: the IgM mAbs could potentially be used for prevention or treatment of EV-A71 infections. Author summaryHand, foot and mouth disease (HFMD) caused by enterovirus infections affects millions of young children annually. Enterovirus A71 (EV-A71) is a major causative pathogen of severe HFMD. EV-A71-specific antibody response is induced rapidly after illness onset, but the characteristics of primary antibody response to EV-A71 in young children are not well understood. Previous study demonstrated a dominant IgM producing EV-A71-specific antibody-secreting B cell response in children under 3 years old. However, no EV-A71-specific human IgM mAb has been reported. Here, we identified EV-A71 neutralizing mAbs, including two IgG1 and six IgM mAbs, from plasmablasts of EV-A71 infected children. IgM mAbs neutralize EV-A71 potently although possess few somatic mutations. We found sequence similarities between IgM antibodies from two patients. We selected EV-A71 variants in the presence of neutralizing mAbs and demonstrated that residues critical for neutralization are conserved among EV-A71 genotypes. We further explored epitope specific antibody responses in acute phase sera of EV-A71 infected patients and found that these neutralizing epitopes on the virus are immunogenic. Our findings advance understanding of antibody response in EV-A71 infected HFMD patients and can help inform the development of therapies against EV-A71 infections.