UNASSIGNED: Hand, foot, and mouth disease (HFMD) is a global public health concern, notably within the Asia-Pacific region. Recently, the primary pathogen causing HFMD outbreaks across numerous countries, including China, is coxsackievirus (CV) A6, one of the most prevalent enteroviruses in the world. It is a new variant that has undergone genetic recombination and evolution, which might not only induce modifications in the clinical manifestations of HFMD but also heighten its pathogenicity because of nucleotide mutation accumulation.
UNASSIGNED: The study assessed the epidemiological characteristics of HFMD in China and characterized the molecular epidemiology of the major pathogen (CV-A6) causing HFMD. We attempted to establish the association between disease progression and viral genetic evolution through a molecular epidemiological study.
UNASSIGNED: Surveillance data from the Chinese Center for Disease Control and Prevention from 2021 to 2023 were used to analyze the epidemiological seasons and peaks of HFMD in Henan, China, and capture the results of HFMD pathogen typing. We analyzed the evolutionary characteristics of all full-length CV-A6 sequences in the NCBI database and the isolated sequences in Henan. To characterize the molecular evolution of CV-A6, time-scaled tree and historical population dynamics regarding CV-A6 sequences were estimated. Additionally, we analyzed the isolated strains for mutated or missing amino acid sites compared to the prototype CV-A6 strain.
UNASSIGNED: The 2021-2023 epidemic seasons for HFMD in Henan usually lasted from June to August, with peaks around June and July. The monthly case reporting rate during the peak period ranged from 20.7% (4854/23,440) to 35% (12,135/34,706) of the total annual number of cases. Analysis of the pathogen composition of 2850 laboratory-confirmed cases identified 8 enterovirus serotypes, among which CV-A6 accounted for the highest proportion (652/2850, 22.88%). CV-A6 emerged as the major pathogen for HFMD in 2022 (203/732, 27.73%) and 2023 (262/708, 37.01%). We analyzed all CV-A6 full-length sequences in the NCBI database and the evolutionary features of viruses isolated in Henan. In China, the D3 subtype gradually appeared from 2011, and by 2019, all CV-A6 virus strains belonged to the D3 subtype. The VP1 sequences analyzed in Henan showed that its subtypes were consistent with the national subtypes. Furthermore, we analyzed the molecular evolutionary features of CV-A6 using Bayesian phylogeny and found that the most recent common ancestor of CV-A6 D3 dates back to 2006 in China, earlier than the 2011 HFMD outbreak. Moreover, the strains isolated in 2023 had mutations at several amino acid sites compared to the original strain.
UNASSIGNED: The CV-A6 virus may have been introduced and circulating covertly within China prior to the large-scale HFMD outbreak. Our laboratory testing data confirmed the fluctuation and periodic patterns of CV-A6 prevalence. Our study provides valuable insights into understanding the evolutionary dynamics of CV-A6.

Hand, foot, and mouth disease (HFMD) is one of the most common class C infectious diseases, posing a serious threat to public health worldwide. Enterovirus A71 (EV-A71) and coxsackievirus A16 (CV-A16) have been regarded as the major pathogenic agents of HFMD; however, since an outbreak caused by coxsackievirus A6 (CV-A6) in France in 2008, CV-A6 has gradually become the predominant pathogen in many regions. CV-A6 infects not only children but also adults, and causes atypical clinical symptoms such as a more generalized rash, eczema herpeticum, high fever, and onychomadesis, which are different from the symptoms associated with EV-A71 and CV-A16. Importantly, the rate of genetic recombination of CV-A6 is high, which can lead to changes in virulence and the rapid evolution of other characteristics, thus posing a serious threat to public health. To date, no specific vaccines or therapeutics have been approved for CV-A6 prevention or treatment, hence it is essential to fully understand the relationship between recombination and evolution of this virus. Here, we systematically review the genetic recombination events of CV-A6 that have occurred worldwide and explore how these events have promoted virus evolution, thus providing important information regarding future HFMD surveillance and prevention.

UNASSIGNED: Hand, foot, and mouth disease (HFMD) is a common childhood infectious disease caused by a variety of enteroviruses (EVs). To explore the epidemiological characteristics and etiology of HFMD in Zhengzhou, China, we conducted a systematic analysis of HFMD surveillance data from Zhengzhou Center for Disease Control and Prevention from January 2009 to December 2021 (https://wjw.zhengzhou.gov.cn/).
UNASSIGNED: Surveillance data were collected from Zhengzhou Center for Disease Control and Prevention from January 2009 to December 2021 (https://wjw.zhengzhou.gov.cn/). Cases were analyzed according to the time of onset, type of diagnosis, characteristics, viral serotype, and epidemiological trends.
UNASSIGNED: We found that the primary causative agent responsible for the HFMD outbreaks in Zhengzhou was Enterovirus A71 (EVA-71) (48.56%) before 2014. After 2015, other EVs gradually became the dominant strains (57.68%). The data revealed that the HFMD epidemics in Zhengzhou displayed marked seasonality, with major peaks occurring from April to June, followed by secondary peaks from October to November, except in 2020. Both the severity and case-fatality ratio of HFMD decreased following the COVID-19 pandemic (severity ‰: 13.46 vs. 0.17; case-fatality ‰: 0.21 vs. 0, respectively). Most severe cases were observed in patients aged 1 year and below, accounting for 45.81%.
UNASSIGNED: Overall, the incidence rate of HFMD decreased in Zhengzhou following the introduction of the EVA-71 vaccine in 2016. However, it is crucial to acknowledge that HFMD prevalence continues to exhibit a distinct seasonal pattern and periodicity, and the occurrence of other EV infections poses a new challenge for children\'s health.

Hand, foot and mouth disease (HFMD) was one of the most common infectious disease in the past few decades. After 2013, coxsackievirus A6 (CVA6) has replaced enterovirus 71 (EV-A71) and coxsackievirus A16 (CVA16), becoming the predominant pathogen responsible for HFMD in many areas in China. The objective of this study is to investigate the genetic characteristics and molecular epidemiology of CVA6 in Linyi from 2022 to 2023. A total of 965 HFMD cases were enrolled in this study and analyses based on VP1 nucleotide sequences were performed to determine the evolutionary trajectory of CVA6. In 2022, 281 (281/386, 72.8%) were positive for enterovirus (EVs), and 217 (217/281, 77.2%) were CVA6 positive. In 2023, 398 (398/579, 68.7%) samples were positive for EVs, and 243 (243/398, 61.1%) were CVA6 positive. Six sequences were selected from each year for the homology analysis. The results showed that 12 strains isolated in Linyi were far from the prototype strain (AY421764) and the first CVA6 strain reported in China (JQ364886). Phylogenetic analysis showed that the CVA6 strains isolated in Linyi all belonged to D3 subgenotype. CVA6 is emerging as a common pathogen causing HFMD in Linyi, and continuous surveillance of HFMD etiological agents is necessary.

Hand, foot, and mouth disease (HFMD) is a contagious viral infection predominantly affecting infants and young children, caused by multiple enteroviruses, including Enterovirus 71 (EV71), Coxsackievirus A16 (CA16), Coxsackievirus A10 (CA10), and Coxsackievirus A6 (CA6). The high pathogenicity of HFMD has garnered significant attention. Currently, there is no specific treatment or broad-spectrum preventive measure available for HFMD, and existing monovalent vaccines have limited impact on the overall incidence or prevalence of the disease. Consequently, with the emergence of new viral strains driven by vaccine pressure, there is an urgent need to develop strategies for the rapid response and control of new outbreaks. In this study, we demonstrated the broad protective effect of maternal antibodies against three types of HFMD by immunizing mother mice with a trivalent inactivated vaccine targeting EV71, CA16, and CA10, using a neonatal mouse challenge model. Based on the feasibility of maternal antibodies as a form of passive immunization to prevent HFMD, we prepared a multivalent antiviral milk by immunizing dairy cows with the trivalent inactivated vaccine to target multiple HFMD viruses. In the neonatal mouse challenge model, this immunized milk exhibited extensive passive protection against oral infections caused by the three HFMD viruses. Compared to vaccines, this strategy may offer a rapid and broadly applicable approach to providing passive immunity for the prevention of HFMD, particularly in response to the swift emergence and spread of new variants.

This study was conducted to efficiently produce virus-like particles (VLPs) of enterovirus 71 (EV71), a causative virus of hand, foot, and mouth disease (HFMD). The expression level of the P1 precursor, a structural protein of EV71, was modified to increase VLP production, and the optimal expression level and duration of the 3CD protein for P1 cleavage were determined. The expression level and duration of 3CD were controlled by the p10 promoter, which was weakened by repeated burst sequence (BS) applications, as well as the OpIE2 promoter, which was weakened by the insertion of random untranslated region sequences of various lengths. The cleavage and production efficiency of the P1 precursor were compared based on the expression time and level of 3CD, revealing that the p10-BS5 promoter with four repeated BSs was the most effective. When P1 and 3CD were expressed using the hyperexpression vector and the p10-BS5 promoter, high levels of structural protein production and normal HFMD-VLP formation were observed, respectively. This study suggests that the production efficiency of HFMD-VLPs can be significantly enhanced by increasing the expression of the P1 precursor and controlling the amount and duration of 3CD expression.

BACKGROUND: Hand, foot and mouth disease (HFMD) is a common infectious disease caused by viral infection by a variety of enteroviruses, with coxsackievirus A 10 (CA10) having become more prevalent in recent years.
METHODS: In this study, models of CA10 infection were established in 7-day-old Institute of Cancer Research (ICR) mice by intraperitoneal injection to analyze the pathogenicity of the virus. RNA sequencing analysis was used to screen the differentially expressed genes (DEGs) after CA10 infection. Coxsackievirus A 16 (CA16) and enterovirus 71 (EV71) infections were also compared with CA10.
RESULTS: After CA10 virus infection, the mice showed paralysis of the hind limbs at 3 days post infection and weight loss at 5 days post infection. We observed viral replication in various tissues and severe inflammatory cell infiltration in skeletal muscle. The RNA-sequencing analysis showed that the DEGs in blood, muscle, thymus and spleen showed heterogeneity after CA10 infection and the most up-regulated DEGs in muscle were enriched in immune-related pathways. Compared with CA16 and EV71 infection, CA10 may have an inhibitory effect on T helper (Th) cell differentiation and cell growth. Additionally, the common DEGs in the three viruses were most enriched in the immune system response, including the Toll-like receptor pathway and the nucleotide-binding and oligomerization domain (NOD)-like pathway.
CONCLUSIONS: Our findings revealed a group of genes that coordinate in response to CA10 infection, which increases our understanding of the pathological mechanism of HFMD.

Coxsackievirus A6 (CV-A6) has emerged as the predominant causative agent of hand, foot, and mouth disease (HFMD) in young children. Since the declaration of coronavirus disease 2019 (COVID-19) as a global pandemic, the incidence of infectious diseases, including HFMD, has decreased markedly. When social mitigation was relaxed during the COVID-19 pandemic in 2022, the re-emergence of HFMD was observed in Gwangju, South Korea, and seasonal characteristics of the disease appeared to have changed. To investigate the molecular characteristics of enterovirus (EV) associated with HFMD during 2022, 277 specimens were collected. Children aged younger than 5 years accounted for the majority of affected individuals. EV detection and genotyping were performed using real-time RT-PCR and nested RT-PCR followed by sequence analysis. The EV detection rate was found to be 82.3%, and the main genotype identified was CV-A6. Sixteen CV-A6 samples were selected for whole genome sequencing. According to phylogenetic analysis, all CV-A6 strains from this study belonged to the sub-genotype D3 clade based on VP1 sequences. Analysis of 3D polymerase phylogeny showed that only the recombinant RF-A group was identified. In conclusion, circulating EV types should be continuously monitored to understand pathogen emergence and evolution during the post-pandemic era.

Coxsackievirus-A6 (CV-A6) is responsible for more severe dermatological manifestations compared to other enteroviruses such as CV-A10, CV-A16, and EV-A71, causing HFMD in children and adults. Between 2005 and 2007, the recombinant subclade D3/RF-A started to expand globally, and a CV-A6 pandemic started. The study aimed to conduct whole-genome sequencing (WGS) of an isolated CV-A6 strain from currently circulating HFMD cases from India in 2022. Gene-specific RT-PCR and sequencing were used to perform molecular characterization of the isolated virus. Confirmation of these isolates was also performed by transmission electron microscopy and WGS. Among eleven positive clinical enterovirus specimens, eight CV-A6 strains were successfully isolated in the RD cell line. Isolates confirmed the presence of the CV-A6 strain based on VP1 and VP2 gene-specific RT-PCR. Sequences of isolates were clustered and identified as the novel CV-A6 strain of the D3/Y sub-genotype in India. The studies revealed that the D3/Y sub-genotype is being introduced into Indian circulation. The predicted putative functional loops found in VP1 of CV-A6 showed that the nucleotide sequences of the amino acid were a remarkably conserved loop prediction compatible with neutralizing linear epitopes. Therefore, this strain represents a potential candidate for vaccine development and antiviral studies.

This study aimed to determine the intention and willingness-to-pay (WTP) of Chinese parents/guardians to vaccinate their children with the EV-71 vaccine. Knowledge levels about hand, foot, and mouth disease (HFMD) and the EV-71 vaccine were also investigated.
A cross-sectional, self-administered online survey was conducted between November 2022 and March 2023. A stratified multi-stage random sampling method was used to recruit parents/guardians of children aged 0-5 years in southeastern China.
A total of 3,626 complete responses were received. The mean knowledge score of HFMD was 9.99 (±4.23) out of a total of 14 points. The majority of the participants reported a somewhat willing intent (58.8%), followed by an extremely willing intent (28.9%). Participants who did not consider the EV-71 vaccine expensive (OR = 2.94, 95%CI 2.45-3.53) perceived that the EV-71 vaccine is effective (OR = 2.73, 95%CI 1.52-4.90), and a high knowledge level of HFMD (OR = 1.90, 95%CI 1.57-2.29) had the highest significant odds of having an extremely willing intent to vaccinate their children with the EV-71 vaccine. The median (interquartile range [IQR]) of WTP for the EV-71 vaccine was CNY¥200/USD$28 (IQR CNY¥100-400/USD$14-56). The highest marginal WTP for the vaccine was mainly influenced by the perceived high cost of the vaccine. Those participants who did not consider the EV-71 vaccine expensive had more than 10 times higher odds of vaccinating their children (OR = 10.86, 95%CI 8.49-13.88). Perceived susceptibility, perceived benefits, and perceived barriers were also significant influencing factors in the highest marginal WTP.
The findings demonstrate the importance of improving health promotion and reducing the barriers to EV-71 vaccination. Therefore, it is important to improve health promotion and reduce the barriers to EV-71 vaccination.