Emergence of genotype C1 Enterovirus A71 and its link with antigenic variation of virus in Taiwan

Author summary EV-A71 is a cause of hand-foot-mouth disease, epidemics of which still regularly occur around the globe. Given that EV-A71 immune protection from the disease correlates with neutralizing antibody responses, but the responses in humans prior to an outbreak are still poorly understood. An outbreak of hand-foot-mouth disease among children emerged in Taiwan from 2018 to 2019, and genotype C1 EV-A71 caused most of the cases. Here, we characterized EV-A71-neutralizing antibody profiles in details at both the serological and monoclonal levels and showed that antibodies generated by humans prior to the emergence of genotype C1 EV-A71 less effectively neutralize C1 compared to the prior circulating genotypes, which implies the presence of antigenic variation in the EV-A71 genotypes. We further identified and mapped critical neutralizing epitopes of 2018-2019 genotype C1 EV-A71 on the top and margin of the viral capsid pentamer and demonstrated thein vivoprotective effect of human monoclonal antibodies, which highlight the properties of human antibody-neutralizing sites on EV-A71 and the potential of human antibodies as antiviral agents. An outbreak of the hand-foot-mouth disease with severe neurological cases, mainly caused by the genotype C1 enterovirus A71 (EV-A71), occurred in Taiwan between 2018 and early 2019. In the recent decade, the most dominant EV-A71 genotypes in Taiwan were B5 and C4 but changed to C1 in 2018. Antibody-mediated immunity plays a key role in limiting the EV-A71 illness in humans. However, the level of neutralizing activities against genotype C1 virus by human polyclonal and monoclonal antibodies (MAbs) remains largely unclear. In the study, we demonstrated that that 39% (9 in 23) of post-infection sera from the genotype B5- or C4-infected patients in 2014-2017 exhibit reduced titers with the 2018-2019 genotype C1 viruses than with the earlier B5 and C4 viruses tested. This finding with polyclonal sera is confirmed with human MAbs derived from genotype B5 virus-infected individuals. The 2018-2019 genotype C1 virus is resistant to the majority of canyon-targeting human MAbs, which may be associated with the residue change near or at the bottom of the canyon region on the viral capsid. The remaining three antibodies (16-2-11B, 16-3-4D, and 17-1-12A), which target VP1 S241 on the 5-fold vertex, VP3 E81 on the 3-fold plateau and VP2 D84 on the 2-fold plateau of genotype C1 viral capsid, respectively, retained neutralizing activities with variable potencies. These neutralizing antibodies were also found to be protective against a lethal challenge of the 2018-2019 genotype C1 virus in an hSCARB2-transgenic mice model. These results indicate that the EV-A71-specific antibody response may consist of a fraction of poorly neutralizing antibodies against 2018-2019 genotype C1 viruses among a subset of previously infected individuals. Epitope mapping of protective antibodies that recognize the emerging genotype C1 virus has implications for anti-EV-A71 MAbs and the vaccine field.