An atypical winter outbreak of hand, foot, and mouth disease associated with human enterovirus 71, 2010

Hand, foot, and mouth disease (HFMD), primarily a disease of young children, is caused by a virus belonging to the group enteroviruses. EV71 is known to be a causative agent of HFMD, herpangina, aseptic meningitis, paralysis, and meningoencephalitis [1]. HFMD usually presents with symptoms including fever and a characteristic rash associated with the limbs, mouth and skin. In severe disease such as caused by EV71, however, the patients may develop nervous system diseases such as aseptic meningitis, encephalitis, brainstem encephalitis, encephalomyelitis, and poliomyelitis-like syndrome as well as neurogenic pulmonary edema and myocarditis, resulting in high morbidity and mortality [2].

EV71 was first isolated in the United States (California) in 1969 and by the mid-1970s, EV71 outbreaks, characterized by central nervous system complications occurred in succession in Bulgaria and Hungary [3]. In the late 1990s, EV71 emerged in East Asia. In 1997, 2628 cases in Malaysia were reported, with 34 deaths [4]. One year later, an estimated 1.5 million cases occurred in Taiwan; 405 cases developed severe neuropathic complications with 78 deaths [5]. In 1999, an EV71 pandemic occurred in Perth, Australia; 6000 cases were reported in 6 months; 29 cases developed severe disease [6]. Subsequently, numerous outbreaks have been documented in eastern and southeastern Asia, including Singapore [7], South Korea [8], Malaysia [9], Japan [10], Vietnam [11] and mainland China. In 2008, a large scale outbreak of HFMD occurred in Anhui Province, China; 3321 cases were reported with 22 deaths [12].

HFMD outbreaks associated with EV71 exhibit a significant seasonal pattern with a peak in summer and low incidence in winter [13, 14], however the number of HFMD cases increase significantly with increasing temperature and relative humidity [13, 15]. In this study, we describe an atypical winter outbreak of HFMD from November 30 to December 28, 2010, in Linyi City, Shandong Province, China, during which 173 cases were admitted to the Linyi People's Hospital. In this study, we analyzed the epidemiological characteristics of this outbreak and the molecular epidemiology of EV71, with an attempt to provide scientific evidence for the prevention and control of HFMD.

HFMD epidemics in China mainly occur in the spring and summer [20‐22], when hot and humid weather is conducive to the propagation and spread of the virus [23]. However, the Linyi 2010 HFMD outbreak occurred in December, which is relatively uncommon. According to meteorological parameters provided by the Meteorological Bureau of Shandong Province, the mean temperature of Linyi City during the winter of 2010 was 0.6°C higher than that in 2009. Flett et al. suggested that atypical seasonality of HFMD outbreaks during winter might be related to unusually mild temperatures [24]. Research in Hong Kong has also indicated that altered disease etiology of HFMD might be explained by increased winter temperatures [25]. It is reasonable to speculate that the short-term changes in weather variables may affect the seasonality of HFMD. However, cold weather increases the probability that populations will gather in confined spaces such as houses and indoor play areas; increasing the likelihood of unwanted exposures. In addition, more upper respiratory tract infections occur during the winter, resulting in a decrease in immune functions among pediatric patients, which might contribute to increasing incidence of HFMD.

The epidemiology of EV71 in China since this virus was first detected in 1998 has shown that only subgenotype C4 is in endemic circulation [19]. Phylogenetic analysis suggests that three EV71 variants contributed to the 2010 Linyi outbreak and share strong similarities to some EV71 viruses circulating previously in Shandong Province. However, no obvious evidence revealed the transmission relationship between strains from this outbreak and other provinces in China. Therefore, we speculate that this uncommon winter outbreak did not occur in isolation, but was related with local epidemic occurring in the region during the spring and summer of 2010. It is probable that the change of climatic parameters contributed to the occurrence of this atypical seasonal outbreak but the extent of this contribution needs to be further examined.

Subgenotype C4a is geographically broadly distributed and genetically variable; it is estimated to undergo 4.97 × 10⁻³ substitutions per nucleotide per year, a rapid rate of change [26]. Phylogenetic analysis shows that subgenotype C4 should be re-designated as a novel genotype, D [27, 28]. Thus, the three lineages may represent different EV71 transmission modes in this region; this hypothesis should be clarified by further molecular epidemiological studies.

Amino sequence analysis showed one primary variation between viruses isolated from severe and mild cases of HFMD, substitution of the amino acid residue at position 145. Several studies have revealed that this change in the VP1 region of EV71 may play an important role in virulence. Residue 145 is reported to be located in the DE loop which contains neutralizing antigenic sites; this loop is located in the canyon rim, a region involved in the receptor binding for enterovirus and rhinovirus [29‐32]. Selection pressure analysis of many VP1 sequences of EV71 showed that amino acid 145 was a positive selection site [33, 34]. Mutation of glutamine to glutamic acid on VP1 145 region cooperatively promote viral binding and RNA accumulation of EV71, contributing to viral infectivity in vitro and mouse lethality in vivo[35]. Substitution of glycine to glutamic acid at the same position might increase the efficiency of uncoating upon specific binding of the virion to the receptor molecule on the target cells in NOD/SCID mice and subsequently facilitate the infection of a mouse adapted EV71 strain [36]. Although it has been demonstrated that the amino acid substitution of Gly to Glu at position 145 of VP1 could increase EV71 virulence in mice, Glu to Gly/Gln/Arg substitutions may enhance virulence in humans [37]. Consistent with this result, isolates from severe cases in our investigation were significantly more likely to carry the Gly/Gln at amino acid 145 than those from mild cases. This substitution may influence the virus binding to antibody or receptor, thereby affecting the virulence of the virus [37].

In conclusion, HFMD caused by EV71 has become a global concern in recent years. In February 2008, HFMD was officially categorized as a Class C infectious disease in China. Further studies on the transmission patterns of EV71 using approaches in molecular biology are therefore warranted, and recognizing EV71 isolates with increased virulence should be a priority for HFMD prevention and control efforts. Moreover, monitoring the genetic variation of EV71 may be useful in facilitating the development of effective EV71 vaccine candidates on Chinese market and evaluating their effects of prevention and control of the EV71 infections.