Hand, Foot And Mouth Disease Patients Research Articles

Genetic Diversity Analysis of Coxsackievirus A8 Circulating in China and Worldwide Reveals a Highly Divergent Genotype.

Coxsackievirus A8 (CV-A8) is one of the pathogens associated with hand, foot and mouth disease (HFMD) and herpangina (HA), occasionally leading to severe neurological disorders such as acute flaccid paralysis (AFP). Only one study aimed at CV-A8 has been published to date, and only 12 whole-genome sequences are publicly available. In this study, complete genome sequences from 11 CV-A8 strains isolated from HFMD patients in extensive regions from China between 2013 and 2018 were determined, and all sequences from GenBank were retrieved. A phylogenetic analysis based on a total of 34 complete VP1 sequences of CV-A8 revealed five genotypes: A, B, C, D and E. The newly emerging genotype E presented a highly phylogenetic divergence compared with the other genotypes and was composed of the majority of the strains sequenced in this study. Markov chain Monte Carlo (MCMC) analysis revealed that genotype E has been evolving for nearly a century and somehow arose in approximately 2010. The Bayesian skyline plot showed that the population size of CV-A8 has experienced three dynamic fluctuations since 2001. Amino acid residues of VP1100N, 103Y, 240T and 241V, which were embedded in the potential capsid loops of genotype E, might enhance genotype E adaption to the human hosts. The CV-A8 whole genomes displayed significant intra-genotypic genetic diversity in the non-capsid region, and a total of six recombinant lineages were detected. The Chinese viruses from genotype E might have emerged recently from recombining with European CV-A6 strains. CV-A8 is a less important HFMD pathogen, and the capsid gene diversity and non-capsid recombination variety observed in CV-A8 strains indicated that the constant generation of deleterious genomes and a constant selection pressure against these deleterious mutations is still ongoing within CV-A8 quasispecies. It is possible that CV-A8 could become an important pathogen in the HFMD spectrum in the future. Further surveillance of CV-A8 is greatly needed.

IFI27 as a potential indicator for severe Enterovirus 71-infected hand foot and mouth disease

The pathogenesis of Enterovirus 71 (EV71)-induced severe hand foot and mouth disease (HFMD) has not yet been clearly established. Further study into the mechanisms underlying host immune responses to EV71 infection and identifying important predictors will be crucial to antiviral treatment and early recognition of severe HFMD. The present study establishes that T help (Th)1 type, Th2 type, and Th17 type cytokine levels in serum of peripheral blood from patients with severe HFMD is higher than in peripheral blood from healthy subjects. The most significant increase occurred as the IL-6. In order to identify the important molecules in peripheral blood mononuclear cells (PBMCs) from severe HFMD patients, we performed transcriptome sequencing analysis of PBMC from severe HFMD patients and compared them to healthy controls. Interferon α-inducible protein 27 (IFI27) and cluster of differentiation 27 (CD27) were found to be the most significant differentially expressed gene. Finally, IFI27 was proved to be present at higher levels in patients with severe HFMD than in patients with mild HFMD. Our results suggest that IFI27 may be an indicator of the severity of cases EV71-induced HFMD.

Clinical, etiological and epidemiological investigations of hand, foot and mouth disease in southern Vietnam during 2015 - 2018.

Hand, foot and mouth disease (HFMD) continues to challenge Asia with pandemic potential. In Vietnam, there have been two major outbreaks occurring during 2011–2012 (>200,000 hospitalizations and >200 deaths) and more recently in 2018 (>130,000 hospitalizations and 17 deaths). Given the high burden and the complex epidemic dynamics of HFMD, synthesizing its clinical and epidemiological data remains essential to inform the development of appropriate interventions and design public health measures. We report the results of a hospital-based study conducted during 2015–2018, covering the severe HFMD outbreak recently documented in Vietnam in 2018. The study was conducted at three major hospitals responsible for receiving HFMD patients from southern Vietnam with a population of over 40 million. A total of 19 enterovirus serotypes were detected in 1196 HFMD patients enrolled in the clinical study during 2015–2018, with enterovirus A71 (EV-A71), coxsackievirus A6 (CV-A6), CV-A10 and CV-A16 being the major causes. Despite the emergence of coxsackieviruses, EV-A71 remains the leading cause of severe HFMD in Vietnam. EV-A71 was consistently detected at a higher frequency during the second half of the years. The emergence of EV-A71 subgenogroup C4 in late 2018 was preceded by its low activity during 2017–early 2018. Compared with EV-A71 subgenogroup B5, C4 was more likely to be associated with severe HFMD, representing the first report demonstrating the difference in clinical severity between subgenogroup C4 and B5, the two predominant EV-A71 subgenogroups causing HFMD worldwide. Our data have provided significant insights into important aspects of HFMD over four years (2015–2018) in Vietnam, and emphasize active surveillance for pathogen circulation remains essential to inform the local public health authorities in the development of appropriate intervention strategies to reduce the burden of this emerging infections. Multivalent vaccines are urgently needed to control HFMD.

Risk factors for severe hand, foot, and mouth disease infected with Coxsackievirus A6: A hospital-based case-control study.

Coxsackievirus 6 (CV-A6) has been emerging as another predominant serotype for severe hand, foot, and mouth disease (HFMD) in China, after the introduction of enterovirus 71 inactivated vaccine (EV71 vaccine) for 3 years. Data on the risk factors for severe HFMD infected with CV-A6 are limited. We interviewed the caregivers to collected data on HFMD patients who sought medical care in the People's Hospital of Baoan district, Shenzhen, from 2015 to 2017. Totally, 131 severe patients were frequency-matched by age and gender with 174 mild patients infected with CV-A6. Univariable and multivariable logistic regression analyses were conducted to analyze the risk factors for severe CV-A6 HFMD. The average age was 20.62 ± 14.18 months and 20.52 ± 12.76 months for severe and mild patients, respectively. Multivariate analyses indicated complications at birth (odds ratio [OR], 4.18; 95% confidence interval [CI]: 1.64-10.63), peak body temperature over 39°C (OR, 4.04; 95% CI: 2.29-7.10) and first-born child (OR, 2.17; 95% CI: 1.27-3.70) increased the risk of severe HFMD infected with CV-A6. Breastfeeding (OR, 0.52; 95% CI: 0.32-0.87), and washing hands after playing frequently (OR, 0.58; 95% CI: 0.34-0.97) were negatively associated with severe illness. Compared with HFMD with infection of EV-A71, complications at birth and first-born child were newly found to be associated with severe illness in HFMD patients infected with CV-A6.

Genetic Variation of Multiple Serotypes of Enteroviruses Associated with Hand, Foot and Mouth Disease in Southern China

Enteroviruses (EVs) species A are a major public health issue in the Asia–Pacific region and cause frequent epidemics of hand, foot and mouth disease (HFMD) in China. Mild infections are common in children; however, HFMD can also cause severe illness that affects the central nervous system. To molecularly characterize EVs, a prospective HFMD virological surveillance program was performed in China between 2013 and 2016. Throat swabs, rectal swabs and stool samples were collected from suspected HFMD patients at participating hospitals. EVs were detected using generic real-time and nested reverse transcription-polymerase chain reactions (RT-PCRs). Then, the complete VP1 regions of enterovirus A71 (EV-A71), coxsackievirus A16 (CVA16) and CVA6 were sequenced to analyze amino acid changes and construct a viral molecular phylogeny. Of the 2836 enrolled HFMD patients, 2,517 (89%) were EV positive. The most frequently detected EVs were CVA16 (32.5%, 819), CVA6 (31.2%, 785), and EV-A71 (20.4%, 514). The subgenogroups CVA16_B1b, CVA6_D3a and EV-A71_C4a were predominant in China and recombination was not observed in the VP1 region. Sequence analysis revealed amino acid variations at the 30, 29 and 44 positions in the VP1 region of EV-A71, CVA16 and CVA6 (compared to the respective prototype strains BrCr, G10 and Gdula), respectively. Furthermore, in 21 of 24 (87.5%) identified EV-A71 samples, a known amino acid substitution (D31N) that may enhance neurovirulence was detected. Our study provides insights about the genetic characteristics of common HFMD-associated EVs. However, the emergence and virulence of the described mutations require further investigation.Electronic supplementary materialThe online version of this article (10.1007/s12250-020-00266-7) contains supplementary material, which is available to authorized users.

Viral shedding in patients with hand, foot and mouth disease induced by EV71, CA16, or CA6: A protocol for systematic review and meta analysis.

Introduction:Hand, foot, and mouth disease (HFMD) has been an important public health concern worldwide, especially in the Asia-Pacific region. Unfortunately, the effect of current measures on preventing and controlling HFMD may be limited. Isolation of infectious sources is reported as an important way to prevent and control this disease. The isolation period is determined on the basis of duration of viral shedding in patients with HFMD. However, the results of previous researches on duration of viral shedding remain controversial. Here, we present a protocol of a systematic review and single-arm meta-analysis for assessing the duration of viral shedding in patients with HFMD induced by Enterovirus 71 (EV71), Coxsackievirus A16 (CA16), or Coxsackievirus A6 (CA6).Methods and analysis:A comprehensive literature search will be performed in PubMed, EMBASE, the Cochrane library, Chinese National Knowledge Infrastructure (CNKI), Chinese Biomedical Literature Database (CBM), and Wanfang Database, covering the period from inception to May 1, 2019. Point estimate of positive rate with corresponding 95% confidence intervals (CIs) of EV71, CA16, or CA6 in HFMD patients’ fecal or throat samples will be carried out using STATA 14.0. Subgroup analyses will be performed for mild cases, severe cases, and close contacts. Sensitive analysis will also be performed to evaluate the influences of individual studies on the final effect by exclusion of a few articles of poor quality. We will assess the risk of bias for the final studies included in our meta-analysis using previously available tools and the modified risk of bias tool.Results:The results of this systematic review and meta-analysis will be published in a peer-reviewed journal.Conclusion:To the best of our knowledge, this paper will be the first systematic review and meta-analysis for assessing the duration of viral shedding in patients with HFMD induced by EV71, CA16, or CA6. The conclusions drawn from this review will provide the scientific basis to formulate the isolation period of HFMD.Ethics and dissemination:Ethical review is not required as this article is for a systematic review since there is no direct involvement of patients in the whole process. We will publish the results of this systematic review and meta-analysis of single-arm studies in a peer-reviewed journal.Registration number:Prospero CRD42020139999.

Genome-wide association study identifies TPH2 variant as a novel locus for severe CV-A6-associated hand, foot, and mouth disease in Han Chinese

BackgroundLittle is known about the association between genetic susceptibility and the severity of hand, foot, and mouth disease (HFMD) infected with coxsackievirus A6 (CV-A6). MethodsThree hundred and sixty-four CV-A6 HFMD patients were enrolled, including 115 severe and 249 mild patients. A genome-wide association study (GWAS) was performed involving eight DNA pools of 115 severe and 115 mild CV-A6 HFMD patients pair-matched by age and gender. Differences in relative allele signal scores of SNPs in Illumina Human OmniZhongHua-8 BeadChips were compared between the two groups. The tag SNPS for potentially functional SNPs or their high linked SNPs were selected for individual genotyping in all 364 patients and assessed for their associations with severe CV-A6 HFMD using multivariable logistic regression analyses. ResultsThe top 30 significant SNPs obtained from pooled DNA GWAS analysis were checked for biological functions and their high linkage disequilibrium (LD) SNPs. Four tag SNPs (rs1558206, rs6927647, rs9375728 and rs10879355) were selected for further individual genotyping in 364 CV-A6 patients. Only SNP rs10879355 was associated with severe CV-A6 HFMD, with CC genotype having a greater risk of severe illness than TT+TC genotypes (OR=2.48, 95%CI: 1.34, 4.56). SNP rs4290270 is in complete LD with rs10879355 in Chinese Han children. ConclusionsThis is the first report that one potentially functional SNP rs4290270 in the TPH2 gene may be associated with the risk of severe CV-A6 HFMD.

Severity detection tool for patients with infectious disease.

Hand foot and mouth disease (HFMD) and tetanus are serious infectious diseases in low- and middle-income countries. Tetanus, in particular, has a high mortality rate and its treatment is resource-demanding. Furthermore, HFMD often affects a large number of infants and young children. As a result, its treatment consumes enormous healthcare resources, especially when outbreaks occur. Autonomic nervous system dysfunction (ANSD) is the main cause of death for both HFMD and tetanus patients. However, early detection of ANSD is a difficult and challenging problem. The authors aim to provide a proof-of-principle to detect the ANSD level automatically by applying machine learning techniques to physiological patient data, such as electrocardiogram waveforms, which can be collected using low-cost wearable sensors. Efficient features are extracted that encode variations in the waveforms in the time and frequency domains. The proposed approach is validated on multiple datasets of HFMD and tetanus patients in Vietnam. Results show that encouraging performance is achieved. Moreover, the proposed features are simple, more generalisable and outperformed the standard heart rate variability analysis. The proposed approach would facilitate both the diagnosis and treatment of infectious diseases in low- and middle-income countries, and thereby improve patient care.

Heparan sulfate attachment receptor is a major selection factor for attenuated enterovirus 71 mutants during cell culture adaptation.

Enterovirus 71 (EV71) is a causative agent of hand, foot, and mouth disease (HFMD). However, this infection is sometimes associated with severe neurological complications. Identification of neurovirulence determinants is important to understand the pathogenesis of EV71. One of the problems in evaluating EV71 virulence is that its genome sequence changes rapidly during replication in cultured cells. The factors that induce rapid mutations in the EV71 genome in cultured cells are unclear. Here, we illustrate the population dynamics during adaptation to RD-A cells using EV71 strains isolated from HFMD patients. We identified a reproducible amino acid substitution from glutamic acid (E) to glycine (G) or glutamine (Q) in residue 145 of the VP1 protein (VP1-145) after adaptation to RD-A cells, which was associated with attenuation in human scavenger receptor B2 transgenic (hSCARB2 tg) mice. Because previous reports demonstrated that VP1-145G and Q mutants efficiently infect cultured cells by binding to heparan sulfate (HS), we hypothesized that HS expressed on the cell surface is a major factor for this selection. Supporting this hypothesis, selection of the VP1-145 mutant was prevented by depletion of HS and overexpression of hSCARB2 in RD-A cells. In addition, this mutation promotes the acquisition of secondary amino acid substitutions at various positions of the EV71 capsid to increase its fitness in cultured cells. These results indicate that attachment receptors, especially HS, are important factors for selection of VP1-145 mutants and subsequent capsid mutations. Moreover, we offer an efficient method for isolation and propagation of EV71 virulent strains with minimal selection pressure for attenuation.

Sequence analysis of VP1 and VP4 genes of enterovirus 71 strains isolated from children with severe and mild hand-foot and mouth disease

Objective To analyze and compare VP1 and VP4 genetic characteristics of human enterovirus 71(EV71) isolated from children with severe and mild hand-foot and mouth disease(HFMD). Methods EV71 strains isolated from severe HFMD patients (6 cases) and mild HFMD patients (6 cases) in Taizhou district, Zhejiang province were included during 2016.Total virus RNA was extracted by Viral RNA Mini Extraction Kit, and reverse transcription polymerase chain reaction(RT-PCR) was used to amplify the sequence of the VP1 and VP4 genes of EV71.And then the sequencing results were compared with those of A, B, C genotype reference EV71 strains from GenBank by nucleotide alignment and amino acid alignment analysis. Results There were no statistically significant differences in sex and age between the two groups(χ2=14.51, t=2.82, all P<0.05). The homogeneity between EV71 strains from severe patients and mild patients was 95.8%~99.6% and 99.1%~100.0% for VP1 nucleotide sequences and amino acid sequences, respectively, while 95.0%~99.9% and 99.0%~100.0% for VP4 nucleotide sequences and amino acid sequences, respectively.The twelve EV71 strains isolated from HFMD patients in Taizhou shared the highest identity with EV71-genotype C, especially genotype C4a.In addition, compared with the mild patients, three respective strains from severe HFMD patients showed mutations at the residue 170 in the VP1 protein(V→L), residue 293 in the VP1 protein(A→S) and residue 7 in the VP4 protein(T→A). Conclusion All EV71 strains isolated from severe and mild HFMD patients in Taizhou district share high homology of nucleotide and amino sequence, and all of them belong to subgenogroup C4a.The mutations in the VP1 and VP4 of EV71 might be related to HFMD disease severity. Key words: Hand-foot and mouth disease; Enterovirus 71; Genotype; Disease severity; Mutation

Neutralizing Antibodies against Enteroviruses in Patients with Hand, Foot and Mouth Disease

Hand, foot and mouth disease (HFMD) is an emerging infection with pandemic potential. Knowledge of neutralizing antibody responses among its pathogens is essential to inform vaccine development and epidemiologic research. We used 120 paired-plasma samples collected at enrollment and >7 days after the onset of illness from HFMD patients infected with enterovirus A71 (EV-A71), coxsackievirus A (CVA) 6, CVA10, and CVA16 to study cross neutralization. For homotypic viruses, seropositivity increased from <60% at enrollment to 97%–100% at follow-up, corresponding to seroconversion rates of 57%–93%. Seroconversion for heterotypic viruses was recorded in only 3%–23% of patients. All plasma samples from patients infected with EV-A71 subgenogroup B5 could neutralize the emerging EV-A71 subgenogroup C4. Collectively, our results support previous reports about the potential benefit of EV-A71 vaccine but highlight the necessity of multivalent vaccines to control HFMD.

Association between platelet count and the risk and progression of hand, foot, and mouth disease among children

OBJECTIVE:We aimed to evaluate the association between platelet (PLT) count and the risk and progression of hand, foot, and mouth disease (HFMD).METHODS:In total, 122 HFMD patients and 40 healthy controls were enrolled in the study. The differences between variables among the different subgroups were compared. Logistic regression analyses were performed to assess the relationship between various parameters and HFMD risk/progression. Sensitivity analysis was conducted by detecting the trend of the association between PLT count quartiles and HFMD risk/progression. A generalized additive model was used to identify the nonlinear relationship between PLT count and HFMD risk/progression. The relationship between gender and PLT count as well as the risk/progression of HFMD was detected using a stratified logistic regression model. RESULTS:Significant differences were observed in terms of age, male/female ratio, white blood cell (WBC) count, and PLT count between patients with stage I-II, III-IV HFMD and healthy controls. Moreover, the alanine aminotransferase and magnesium levels between patients with stage I-II and III-IV HFMD significantly differed. Moreover, a significant difference was noted in the male/female ratio among the different PLT groups. The group with a low PLT count had a lower risk of HFMD progression than the group with a high PLT count (Q4) (p=0.039). Lower age, male gender, and WBC count were found to be associated with HFMD risk. Meanwhile, PLT count was correlated to HFMD progression. The sensitivity analysis yielded a similar result using the minimally adjusted model (p for trend=0.037), and minimal changes were observed using the crude and fully adjusted model (p for trend=0.054; 0.090). A significant nonlinear relationship was observed between PLT count and HFMD progression after adjusting for age, gender, and WBC (p=0.039).CONCLUSIONS:PLT was independently associated with HFMD progression in a nonlinear manner.

An infectious clone of enterovirus 71(EV71) that is capable of infecting neonatal immune competent mice without adaptive mutations

ABSTRACT Enterovirus 71 (EV71) is a major pathogen that causes hand, foot and mouth disease (HFMD), which is a life threatening disease in certain children. The pathogenesis of EV71-caused HFMD is poorly defined due to the lack of simple and robust animal models with severe phenotypes that recapitulate symptoms observed in humans. Here, we generated the infectious clone of a clinical isolate from a severe HFMD patient. Virus rescued from the cDNA clone was infectious in cell lines. When administrated intraperitoneally to neonatal ICR, BALB/c and C57 immune competent mice at a dosage of1.4 × 104 pfu per mouse, the virus caused weight loss, paralysis and death in the infected mice after 4–5 days of infection. In the infected mice, detectable viral replication was detected in various tissues such as heart, liver, brain, lung, kidney, small intestine, leg skeletal muscle and medulla oblongata. The histology of the infected mice included massive myolysis, glomerular atrophy, villous blunting in small intestine, widened alveolar septum, diminished alveolar spaces and lymphocytes infiltration into the lung. By using the UV-inactivated virus as a control, we elucidated that the virus first amplified in the leg skeletal muscle tissue and the muscle tissue served as a primary viral replication site. In summary, we generated a stable EV71 infectious clone that is capable of infecting neonatal immune competent mice without adaptive mutations and provide a simple, valuable animal model for the studies of EV71pathogenesis and therapy.

Meteorological factors associated with hand, foot and mouth disease in a Central Highlands province in Viet Nam: an ecological study.

BackgroundHand, foot and mouth disease (HFMD) is a public health problem in Viet Nam, and studies have reported seasonal fluctuation in the occurrence of HFMD. This study sought to describe the occurrence of HFMD and its associated meteorological factors in Dak Lak province, Viet Nam.MethodsMonthly data on HFMD cases were collected from all commune health stations in Dak Lak province from 2012 through 2013. An HFMD case was defined as a brief febrile illness accompanied by a typical skin rash with or without mouth ulcers. Average temperature, maximum temperature, minimum temperature, humidity, rainfall, evaporation, sunshine duration and wind speed were recorded monthly at five local meteorological stations throughout Dak Lak.Data were aggregated at the district level, and the association between these meteorological factors and HFMD cases were examined by Poisson regression.ResultsIn 2012 through 2013, there were 7128 HFMD patients in Dak Lak. The number of HFMD cases increased during the rainy season. An increased risk of HFMD was associated with higher average temperature (risk ratio and 95% confidence interval: 1.06; 1.03–1.08 per 1 °C increase), higher rainfall (1.19; 1.14–1.24 per 200 mm increase) and longer sunshine duration (1.14; 1.07–1.22 per 60 hours increase). The risk of HFMD was inversely associated with wind speed (0.77; 0.73–0.81 per 1 m/s increase).ConclusionThis study suggests that there is a significant association between HFMD occurrence and climate. Temperature, rainfall, wind speed and sunshine duration could be used as meteorological predictors of HFMD occurrence in Viet Nam’s Central Highlands region. Intensified surveillance for HFMD during the rainy season is recommended.

Value of abnormal HLA-DR expression on CD14+ monocytes in estimating immune function status and clinical prognosis of patients with hand, foot and mouth disease

Objective To investigate the value of abnormal expression of HLA-DR on peripheral blood monocytes in evaluating the immune function status, clinical prognosis and severity of patients with hand, foot and mouth disease (HFMD). Methods From June 2017 to October 2018, 100 cases of mild HFMD, 80 cases of severe HFMD, 32 cases of critical HFMD and 40 healthy children (control group) were recruited in this study. The patients were divided into two groups, lower DR group (DR-L, HLA-DR expression 30%) according to the HLA-DR expression on monocytes. Flow cytometry was used to detect the CD14+ monocytes expressing HLA-DR and the absolute count of lymphocyte subsets. Immunoturbidimetry was used to detect the levels of IgG, IgM and IgA in plasma samples. Enzyme-linked immunosorbent assay (ELISA) was performed to detect the levels of IFN-γ and IL-10 in plasma samples. Pediatric critical illness score (PCIS) and the pediatric risk of mortality Ⅲ (PRISM Ⅲ) were used to estimate the severity of HFMD. Results ① There were significant differences in HLA-DR expression on monocytes among children with mild, severe and critical HFMD (F=47.102, P 0.05). ④ Compared with the DR-N group, the DR-L group showed decreased IFN-γ level and increased IL-10 level in plasma (P<0.05). The ratio of IFN-γ/IL-10 of the DR-L group was lower than that of the DR-N group and control group (P<0.05). HLA-DR expression was negatively correlated with the concentration of IL-10 in plasma (r=-0.704, P<0.05), and positively correlated with the IFN-γ/IL-10 ratio (r=0.773, P<0.05). ⑤ Compared with the DR-N group, the DR-L group showed lower PCIS and higher PRISM Ⅲ. HLA-DR expression was positively correlated with PCIS (r=0.715, P=0.00) and negatively correlated with PRISM Ⅲ (r=-0.610, P=0.00). ⑥ The incidence of pulmonary edema, pulmonary hemorrhage and cardiopulmonary failure and the mortality of HFMD patients in the DR-L group were significantly higher than those in the DR-N group (P<0.05). Conclusions Patients with severe or critical HFMD had cellular immune dysfunction and abnormal HLA-DR expression on CD14+ monocytes. Assessing the expression of HLA-DR on monocytes could be used to evaluate the cellular immunity of patients with severe or critical HFMD. Lower expression of HLA-DR on CD14+ monocytes might be associated with severe HFMD and poor prognosis. Key words: Hand, foot and mouth disease; HLA-DR; Cellular immunity

Scavenger receptor class a, member 3 is associated with severity of hand, foot, and mouth disease in a case-control study.

Hand, foot, and mouth disease (HFMD) spreads rapidly and has been recognized as a public health problem in recent years in China. Unfortunately, there is no effective vaccine or antiviral drug currently for EV71 infection. In this study, we aim to identify biomarker which are associated with for severity of EV71 infection cases using high-throughput RNA sequencing approach.RNA sequencing of samples from severe HFMD (S) patients group (n = 10) and control HFMD (C) patients group (n = 10) were performed and the results were verified by qPCR. mRNA with the highest expression level was selected to be validated in an independent cohort comprising of 45 severe EV71 infected patients and 45 control by qPCR assay.Seventeen significant differentially expressed genes were identified. Scavenger receptor class A, member 3 (SCARA3) was one of the significantly upregulated genes with the highest expression level and was selected for validation. The mean relative expression levels in severe HFMD and control HFMD patients were 10.1-fold and 5.0-fold, respectively, P value <.001.We found that SCARA3 is associated with severity of HFMD, and it may be a potential prognostic marker to predict the HFMD progression in EV71 infected patients.

Assessing the efficacy and safety of magnesium sulfate for management of autonomic nervous system dysregulation in Vietnamese children with severe hand foot and mouth disease

BackgroundBrainstem encephalitis is a serious complication of hand foot and mouth disease (HFMD) in children. Autonomic nervous system (ANS) dysregulation and hypertension may occur, sometimes progressing to cardiopulmonary failure and death. Vietnamese national guidelines recommend use of milrinone if ANS dysregulation with Stage 2 hypertension develops. We wished to investigate whether magnesium sulfate (MgSO4) improved outcomes in children with HFMD if used earlier in the evolution of the ANS dysregulation (Stage 1 hypertension).MethodsDuring a regional epidemic we conducted a randomized, double-blind, placebo-controlled trial of MgSO4 in children with HFMD, ANS dysregulation and Stage 1 hypertension, at the Hospital for Tropical Diseases in Ho Chi Minh city. Study participants received an infusion of MgSO4 or matched placebo for 72 h. We also reviewed data from non-trial HFMD patients in whom milrinone failed to control hypertension, some of whom received MgSO4 as second line therapy. The primary outcome for both analyses was a composite of disease progression within 72 h - addition of milrinone (trial participants only), need for ventilation, shock, or death.ResultsBetween June 2014 and September 2016, 14 and 12 participants received MgSO4 or placebo respectively, before the trial was stopped due to futility. Among 45 non-trial cases with poorly controlled hypertension despite high-dose milrinone, 33 received MgSO4 while 12 did not. There were no statistically significant differences in the composite outcome between the MgSO4 and the placebo/control groups in either study (adjusted relative risk (95%CI) of [6/14 (43%) vs. 6/12 (50%)], 0.84 (0.37, 1.92), p = 0.682 in the trial and [1/33 (3%) vs. 2/12 (17%)], 0.16 (0.01, 1.79), p = 0.132 in the observational cohort). The incidence of adverse events was similar between the groups. Potentially toxic magnesium levels occurred very rarely with the infusion regime used.ConclusionAlthough we could not demonstrate efficacy in these studies, there were no safety signals associated with use of 30-50 mg/kg/hr. MgSO4 in severe HFMD. Intermittent outbreaks of HFMD are likely to continue across the region, and an adequately powered trial is still needed to evaluate use of MgSO4 in controlling hypertension in severe HFMD, potentially involving a higher dose regimen.Trial registrationClinicalTrials.gov Identifier: NCT01940250 (Registered 22 AUG 2013).Trial sponsor: University of Oxford

Global cytokine/chemokine profile identifies potential progression prediction indicators in hand-foot-and-mouth disease patients with Enterovirus A71 infections

ObjectiveNew clinical indicators are urgently needed for predicting the progression and complications of hand-foot-and-mouth disease (HFMD) caused by EV-A71 infections. Materials and methodsSerum specimens from 132 EV-A71 HFMD patients and 73 health children were collected during 2012–2014 in Shenzhen, China. The specific cytokines/chemokines were detected with a 274-human cytokine antibody array, followed by a 38-inflammation cytokine array, and further validated by ELISA. ResultsCytokines varied in different severity of EV-A71 HFMD patients. The ROC curve analysis revealed 5 serum cytokines with high sensitivity and specificity in predicting the disease progression. Eotaxin, IL-8 and IP-10 have showed high AUC values (0.90–0.95) for discrimination between the health controls and the patient group. The three cytokines showed high sensitivity (80–91%) and specificity (88–95%). MMP-8 had a high sensitivity and specificity to predict mild HFMD (100%, 100%). IL-1b and leptin discriminated the severe/critical group from the mild group (79% and 69% in sensitivity, 73% and 63% in specificity). ConclusionsEotaxin, IP-10 and IL-8 could be potential indicators for predicting HFMD progression with EV-A71 infection. MMP-8 is a specific indicator for mild infection, while IL-1b and leptin display potential for predicting the severity and criticality.

Increased effector γδ T cells with enhanced cytokine production are associated with inflammatory abnormalities in severe hand, foot, and mouth disease

BackgroundAlthough γδ T cells have been reported to be closely related to the immunopathogenesis of some viral infectious diseases, the changes or roles of γδ T cells in the development of hand, foot, and mouth disease (HFMD) remain unclear. MethodsPeripheral γδ T cells and their subsets were determined by surface (γδ TCR, Vδ1 TCR, Vδ2 TCR, CD45RA, and CD27) or intracellular (IFN-γ, TNF-α, CD107a, and Granzyme B) markers in healthy controls (HCs) and HFMD patients with FACS. The plasma levels of IFN-γ, TNF-α, IL-6, and MCP-1 were measured by ELISA. Differences in γδ T cells or their subsets and correlations between γδ T cells and inflammation indicators were statistically analyzed. ResultsCompared to HCs, HFMD patients showed increased effector γδ T and TNF-α+γδ T cells and plasma TNF-α levels, especially in severe cases. In addition, significantly increased Vδ1 T and IFN-γ+γδ T cells and other plasma inflammatory cytokines were further found in severe patients. Furthermore, EV71+ severe patients showed significantly increased effector and cytokine-producing γδ T cells, while the EV71− severe patients displayed significantly greater plasma cytokine levels. The percentage of IFN-γ+γδ T or TNF-α+γδ T cells was positively correlated with that of effector γδ T cells. There was a positive correlation between the proportion of Vδ1 T cells and white blood cell (WBC) count or the proportion of IFN-γ+γδ T or TNF-α+γδ T cells and neutrophil (N) count, while there was a negative correlation between Vδ2 T cells and WBC or N count. Moreover, the percentages of Vδ1 T and effector γδ T cells in the acute phase of disease declined significantly to normal levels during the recovery phase. ConclusionsIncreased effector γδ T cells with enhanced cytokine production were remarkably observed in severe HFMD patients, which was also associated with clinical inflammation parameters. These data indicated that γδ T cells might be involved in inflammatory abnormalities in severe HFMD.

Interleukin-37 Inhibits the Imbalance Between T Helper 17 Cells and Regulatory T Cells in Hand, Foot, and Mouth Disease.

The aim of this study was to explore the role of interleukin-37 (IL-37) in imbalance of T helper (Th)17/regulatory T cells (Tregs) in hand, foot, and mouth disease (HFMD). The proportions of CD4+ IL-17A+ Th17 cells and CD4+ CD25+Foxp3+ Tregs in peripheral blood or peripheral blood mononuclear cells (PBMCs) from HFMD patients and healthy controls were measured by fluorescence activated cell sorter. The level of IL-37, IL-10, IL-17A, IL-23, and transforming growth factor β1 (TGF-β1) in serum or PBMCs of HFMD patients and control subjects were detected using enzyme-linked immunosorbent assay. Results showed that Th17 cells proportion and IL-17A and IL-23 levels were highly increased, whereas Tregs proportion and IL-10 and TGF-β1 levels were significantly decreased in HFMD patients. Moreover, IL-37 stimulation elevated Tregs proportion but reduced Th17 cell proportion in subjects with HFMD. On the contrary, we found methylprednisolone pulse therapy/methylprednisolone combinated with intravenous gamma globulin inhibits Th17/Treg imbalance through upregulation of IL-37 in HFMD. In conclusion, IL-37 inhibits the imbalance of Th17/Tregs in HFMD.