Mutations in the non-structural protein region contribute to intra-genotypic evolution of enterovirus 71

Abstract

BackgroundClinical manifestations of enterovirus 71 (EV71) range from herpangina, hand-foot-and-mouth disease (HFMD), to severe neurological complications. Unlike the situation of switching genotypes seen in EV71 outbreaks during 1998–2008 in Taiwan, genotype B5 was responsible for two large outbreaks in 2008 and 2012, respectively. In China, by contrast, EV71 often persists as a single genotype in the population and causes frequent outbreaks. To investigate genetic changes in viral evolution, complete EV71 genome sequences were used to analyze the intra-genotypic evolution pattern in Taiwan, China, and the Netherlands.ResultsGenotype B5 was predominant in Taiwan’s 2008 outbreak and was re-emergent in 2012. EV71 strains from both outbreaks were phylogenetically segregated into two lineages containing fourteen non-synonymous substitutions predominantly in the non-structural protein coding region. In China, genotype C4 was first seen in 1998 and caused the latest large outbreak in 2008. Unlike shifting genotypes in Taiwan, genotype C4 persisted with progressive drift through time. A majority of non-synonymous mutations occurred in residues located in the non-structural coding region, showing annual increases. Interestingly, genotype B1/B2 in the Netherlands showed another stepwise evolution with dramatic EV71 activity increase in 1986. Phylogeny of the VP1 coding region in 1971–1986 exhibited similar lineage turnover with genotype C4 in China; however, phylogeny of the 3D-encoding region indicated separate lineage appearing after 1983, suggesting that the 3D-encoding region of genotype B2 was derived from an unidentified ancestor that contributed to intra-genotypic evolution in the Netherlands.ConclusionsUnlike VP1 coding sequences long used for phylogenetic study of enteroviruses due to expected host immune escape, our study emphasizes a dominant role of non-synonymous mutations in non-structural protein regions that contribute to (re-)emergent genotypes in continuous stepwise evolution. Dozens of amino acid substitutions, especially in non-structural proteins, were identified via genetic changes driven through intra-genotypic evolution worldwide. These identified substitutions appeared to increase viral fitness in the population, affording valuable insights not only for viral evolution but also for prevention, control, and vaccine against EV71 infection.

Is a multivalent hand, foot, and mouth disease vaccine feasible?

“…125 EV-A71 epidemics occur cyclically every 2-4 y with changes in the predominant genotype and subgenotype. These epidemics can be the result of infection by a single genotype/subgenotype, the co-circulation of divergent EV-A71 isolates, the emergence of variants or the unpredictable switching of genotypes and subgenotypes 126 ( Fig. 1).…”

“…29,30,[32][33][34]127,128 The EV-A71 C4 genotype has persisted with progressive drift through time in China. 9,117,126 Intra-genotype EV-A71 B shifts from B3 to B4 (1997)(1998)(1999)(2000) and B4 to B5 (2000)(2001)(2002)(2003) have occurred in Malaysia. Sequential inter-genotype shifts from C2 to B4 and subsequently from C4 to B5 were observed in Taiwan.…”

“…7,121,126 The co-circulation of several EV-A71 genotypes and CV-A16 during HFMD epidemics has been responsible for intra-typic genetic recombination between EV-A71 B and C genotypes in Taiwan and inter-typic recombination between EV-A71 and CV-A16 in China. 5,7,14,19,126,129 Another inter-serotypic recombination happened between EV-A71 genotype C2 and CV-A8 to create genotype B4, responsible for outbreaks in Japan and Taiwan in 1998. 7,126 The predominant C4a genotype may be a double recombinant virus among EV-A71 genotypes B, C and CV-A16.…”

“…5,7,14,19,126,129 Another inter-serotypic recombination happened between EV-A71 genotype C2 and CV-A8 to create genotype B4, responsible for outbreaks in Japan and Taiwan in 1998. 7,126 The predominant C4a genotype may be a double recombinant virus among EV-A71 genotypes B, C and CV-A16. 7,126 Almost each major HFMD outbreak was correlated to genetic variations caused by EV-A71 switches.…”

“…7,126 The predominant C4a genotype may be a double recombinant virus among EV-A71 genotypes B, C and CV-A16. 7,126 Almost each major HFMD outbreak was correlated to genetic variations caused by EV-A71 switches. 7 Phylogenetic analyses have revealed that CV-A16 B1a and B1b strains circulating in China were complex recombinant forms containing sequences from multiple EV-A donors.…”