Background

Since 1997, several countries within the Asian Pacific region have been affected by one or more massive outbreaks of Hand Foot and Mouth Disease (HFMD). Virus typing experiments revealed that these outbreaks were caused by strains of human enterovirus 71 (EV71) belonging to several different, recently emerged subgenogroups. In mainland China, a different situation was observed. The first outbreak, localized in Shangdong Province, was reported in 2007, and was followed by a wide-spread outbreak in mainland China in 2008. Since then, numbers of reported HFMD cases have been persistently high.

Methodology/Principal Findings

To gain insight in the epidemiological behavior of EV71 in China, we studied genetic diversity and EV71 population dynamics to address whether the increase in number of reported EV71 infections reflects a real increase in viral spread or is just the result of increased awareness and surveillance. We used systematically collected VP1 gene sequences of 257 EV71 strains collected in Guangdong province from 2008 to 2010 as part of HFMD surveillance activities, and supplemented them with 305 GenBank EV71 reference stains collected in China from 1998 to 2010. All isolates from Guangdong Province belonged to subgenogroup C4. Viral population dynamics indicated that the increased reporting of HFMD in China since 2007 reflects a real increase in viral spread and continued replacement of viral lineages through time. Amino acid sequence comparisons revealed substitution of amino acid in residues 22, 145 and 289 through time regularly with the VP1 gene of EV71 strains isolated in mainland China from 1998 to 2010.

Conclusions

EV71 strains isolated in mainland China mainly belonged to subgenogroup C4. There was exponential growth of the EV71 virus population in 2007 and 2008. There was amino acid substitution through time regularly with the VP1 gene which possibly increased viral spread and/or ability of the virus to circulate persistently among the Chinese population.

In China, rubella vaccination was introduced into the national immunization program in 2008, and a rubella epidemic occurred in the same year. In order to know whether changes in the genotypic distribution of rubella viruses have occurred in the postvaccination era, we investigate in detail the epidemiological profile of rubella in China and estimate the evolutionary rate, molecular clock phylogeny, and demographic history of the predominant rubella virus genotypes circulating in China using Bayesian Markov chain Monte Carlo phylodynamic analyses. 1E was found to be the predominant rubella virus genotype since its initial isolation in China in 2001, and no genotypic shift has occurred since then. The results suggest that the global 1E genotype may have diverged in 1995 and that it has evolved at a mutation rate of 1.65 × 10−3 per site per year. The Chinese 1E rubella virus isolates were grouped into either cluster 1 or cluster 2, which likely originated in 1997 and 2006, respectively. Cluster 1 viruses were found in all provinces examined in this study and had a mutation rate of 1.90 × 10−3 per site per year. The effective number of infections remained constant until 2007, and along with the introduction of rubella vaccine into the national immunization program, although the circulation of cluster 1 viruses has not been interrupted, some viral lineages have disappeared, and the epidemic started a decline that led to a decrease in the effective population size. Cluster 2 viruses were found only in Hainan Province, likely because of importation.

The incidence of measles in China from 1991 to 2008 was reviewed, and the nucleotide sequences from 1507 measles viruses (MeV) isolated during 1993 to 2008 were phylogenetically analyzed. The results showed that measles epidemics peaked approximately every 3 to 5 years with the range of measles cases detected between 56,850 and 140,048 per year. The Chinese MeV strains represented three genotypes; 1501 H1, 1 H2 and 5 A. Genotype H1 was the predominant genotype throughout China continuously circulating for at least 16 years. Genotype H1 sequences could be divided into two distinct clusters, H1a and H1b. A 4.2% average nucleotide divergence was found between the H1a and H1b clusters, and the nucleotide sequence and predicted amino acid homologies of H1a viruses were 92.3%–100% and 84.7%–100%, H1b were 97.1%–100% and 95.3%–100%, respectively. Viruses from both clusters were distributed throughout China with no apparent geographic restriction and multiple co-circulating lineages were present in many provinces. Cluster H1a and H1b viruses were co-circulating during 1993 to 2005, while no H1b viruses were detected after 2005 and the transmission of that cluster has presumably been interrupted. Analysis of the nucleotide and predicted amino acid changes in the N proteins of H1a and H1b viruses showed no evidence of selective pressure. This study investigated the genotype and cluster distribution of MeV in China over a 16-year period to establish a genetic baseline before MeV elimination in Western Pacific Region (WPR). Continuous and extensive MeV surveillance and the ability to quickly identify imported cases of measles will become more critical as measles elimination goals are achieved in China in the near future. This is the first report that a single endemic genotype of measles virus has been found to be continuously circulating in one country for at least 16 years.

Molecular characterization of wild-type measles viruses in China during 1995-2004 demonstrated that genotype H1 was endemic and widely distributed throughout the country. H1-associated cases and outbreaks caused a resurgence of measles beginning in 2005. A total of 210,094 measles cases and 101 deaths were reported by National Notifiable Diseases Reporting System (NNDRS) and Chinese Measles Laboratory Network (LabNet) from 2006 to 2007, and the incidences of measles were 6.8/100,000 population and 7.2/100,000 population in 2006 and 2007, respectively. Five hundred and sixty-five wild-type measles viruses were isolated from 24 of 31 provinces in mainland China during 2006 and 2007, and all of the wild type virus isolates belonged to cluster 1 of genotype H1. These results indicated that H1-cluster 1 viruses were the predominant viruses circulating in China from 2006 to 2007. This study contributes to previous efforts to generate critical baseline data about circulating wild-type measles viruses in China that will allow molecular epidemiologic studies to help measure the progress made toward China's goal of measles elimination by 2012.

Measles morbidity and mortality decreased significantly after measles vaccine was introduced into China in 1965. From 1995 to 2004, average annual measles incidence decreased to 5.6 cases per 100,000 population following the establishment of a national two-dose regimen. Molecular characterization of wild-type measles viruses demonstrated that genotype H1 was endemic and widely distributed throughout the country in China during 1995-2004. A total of 124,865 cases and 55 deaths were reported from the National Notifiable Diseases Reporting System (NNDRS) in 2005, which represented a 69.05% increase compared with 2004. Over 16,000 serum samples obtained from 914 measles outbreaks and the measles IgM positive rate was 81%. 213 wild-type measles viruses were isolated from 18 of 31 provinces in China during 2005, and all of the isolates belonged to genotype H1. The ranges of the nucleotide sequence and predicted amino acid sequence homologies of the 213 genotype H1 strains were 93.4%-100% and 90.0%-100%, respectively. H1-associated cases and outbreaks caused the measles resurgence in China in 2005. H1 genotype has the most inner variation within genotype, it could be divided into 2 clusters, and cluster 1 viruses were predominant in China throughout 2005.