Molecular epidemiology and evolution of enterovirus 71 strains isolated from 1970 to 1998

Abstract

Enterovirus 71 (EV71) (genus Enterovirus, family Picornaviridae), a common cause of hand, foot, and mouth disease (HFMD), may also cause severe neurological diseases, such as encephalitis and poliomyelitis-like paralysis. To examine the genetic diversity and rate of evolution of EV71, we have determined and analyzed complete VP1 sequences (891 nucleotides) for 113 EV71 strains isolated in the United States and five other countries from 1970 to 1998. Nucleotide sequence comparisons demonstrated three distinct EV71 genotypes, designated A, B, and C. The genetic variation within genotypes (12% or fewer nucleotide differences) was less than the variation between genotypes (16.5 to 19.7%). Strains of all three genotypes were at least 94% identical to one another in deduced amino acid sequence. The EV71 prototype strain, BrCr-CA-70, isolated in California in 1970, is the sole member of genotype A. Strains isolated in the United States and Australia during the period from 1972 to 1988, a 1994 Colombian isolate, and isolates from a large HFMD outbreak in Malaysia in 1997 are all members of genotype B. Although strains of genotype B continue to circulate in other parts of the world, none have been isolated in the United States since 1988. Genotype C contains strains isolated in 1985 or later in the United States, Canada, Australia, and the Republic of China. The annual rate of evolution within both the B and C genotypes was estimated to be approximately 1.35 x 10(-2) substitutions per nucleotide and is similar to the rate observed for poliovirus. The results indicate that EV71 is a genetically diverse, rapidly evolving virus. Its worldwide circulation and potential to cause severe disease underscore the need for additional surveillance and improved methods to identify EV71 in human disease.

FIG. 1

Dendrogram generated by the neighbor-joining method with the DNADIST distance measure program (PHYLIP, version 3.5). The phylogram was calculated based on the nucleotide divergence of the VP1 gene (position 2442 to 3332). The last four or five characters of each strain name indicate the state or country and year of isolation. Branch lengths are proportional to the number of nucleotide differences; the frequencies with which the branches for genotypes A, B, and C appeared in 1,000 bootstrap replications were 898, 543, and 999, respectively. Clades with bootstrap numbers are expressed in percentile. The marker denotes a measurement of the relative phylogenetic distance. (A) The branch length for the outgroup, CA16-G10-51, was reduced by 0.75 to save space.

FIG. 2

Alignment of genotype consensus VP1 amino acid sequences. The EV71 consensus sequence shows amino acid residues that are identical in at least 85% of all strains and those that are identical in at least 50% but less than 85% of all strains. Sites that are identical in all strains of all genotypes are double underlined; those that are identical in all strains of genotypes B and C, but different in BrCr-CA-70, are single underlined. The genotype consensus sequences indicate sites of at least 85% consensus among all strains of a given genotype (hyphens) and sites that are characteristic of one or more genotypes (uppercase letter, 85% consensus within genotype; lowercase letter, 50 to 85% consensus within genotype).