Genetic variation, recombinant characteristics, and seroprevalence analysis of echovirus 3 causing severe and mild cases of hand, foot, and mouth disease in Guizhou Province

ABSTRACT Echovirus 3 (E-3) has recently been identified as an emerging pathogen associated with both severe and mild cases of hand, foot, and mouth disease (HFMD) in Guizhou Province, China. Although these strains exhibit distinct clinical phenotypes, their underlying molecular mechanisms remain unclear. This study reports the phylogenetic and genetic features of eight E-3 strains from severe and mild HFMD cases and investigates E-3 seroprevalence in Guizhou. Eight shared nucleotide substitutions in the untranslated regions (UTRs) and six shared amino acid substitutions in the open reading frame (ORF) were identified between E-3 strains from severe and mild case groups. Nine variant-specific amino acid mutations were under positive selection and exhibited high variability. Among the 115 nucleotide substitutions in the ORF regions, 31 were specific to the severe case group and 41 to the mild case group. Recombination analysis revealed that the Guizhou E-3 strains underwent complex interserotypic recombination with echovirus 21 (E-21), enterovirus B80 (EV-B80), and echovirus 30 (E-30). E-3 strains from the severe case group demonstrated enhanced replication efficiency, higher viral titers, and greater cytopathogenicity than those from the mild case group. Analysis of 178 samples revealed a seropositivity rate of 52.25% and a geometric mean titer (GMT) of 1:28.23 for E-3, indicating the prevalence of asymptomatic infections in Guizhou Province. All Guizhou E-3 strains exhibited non-temperature-sensitive characteristics, suggesting their potential for transmissibility within the population. This study provides valuable insights for global studies on E-3-related diseases and molecular epidemiology, establishing a foundation for further investigation into the pathogenicity of E-3 variants. IMPORTANCE This study is the first report of echovirus 3 (E-3) infections associated with severe hand, foot, and mouth disease (HFMD) cases in Guizhou Province, China. All E-3 strains showed thermotolerance and higher seroprevalence, suggesting potentially enhanced transmissibility. Several shared nucleotide substitutions and amino acid substitutions were identified in Guizhou strains from both severe and mild case groups, indicating that E-3 may have evolved through continuous genetic adaptation, potentially driving the emergence of more transmissible epidemic variants. This study revealed that the E-3 strains in the severe case group exhibited enhanced replication efficiency, higher viral titers, and increased cytopathogenicity in vitro , thereby providing evidence to support a potential association between genetic variations and clinical severity. Notably, all E-3-infected patients in the severe case group exhibited more severe clinical manifestations. These findings provide critical updates on the epidemiological trends of enteroviruses associated with HFMD and establish a foundation for a better understanding of genetic diversity, evolutionary patterns, and molecular mechanisms of enterovirus infections.