ABSTRACT
Coxsackievirus A10 (CV-A10) infection, a prominent cause of childhood hand-foot-and-mouth disease (HFMD), frequently manifests with the intriguing phenomenon of onychomadesis, characterized by nail shedding. However, the underlying mechanism is elusive. Here, we found that CV-A10 infection in mice could suppress Wnt/ß-catenin signaling by restraining LDL receptor-related protein 6 (LRP6) phosphorylation and ß-catenin accumulation and lead to onychomadesis. Mechanistically, CV-A10 mimics Dickkopf-related protein 1 (DKK1) to interact with Kringle-containing transmembrane protein 1 (KRM1), the CV-A10 cellular receptor. We further found that Wnt agonist (GSK3ß inhibitor) CHIR99021 can restore nail stem cell differentiation and protect against nail shedding. These findings provide novel insights into the pathogenesis of CV-A10 and related viruses in onychomadesis and guide prognosis assessment and clinical treatment of the disease.
Subject(s)
Intercellular Signaling Peptides and Proteins , Low Density Lipoprotein Receptor-Related Protein-6 , Wnt Signaling Pathway , Animals , Wnt Signaling Pathway/drug effects , Low Density Lipoprotein Receptor-Related Protein-6/metabolism , Low Density Lipoprotein Receptor-Related Protein-6/genetics , Mice , Intercellular Signaling Peptides and Proteins/metabolism , Intercellular Signaling Peptides and Proteins/genetics , Humans , beta Catenin/metabolism , Nail Diseases/metabolism , Nail Diseases/virology , Nail Diseases/pathology , Nails/metabolism , Nails/pathology , Cell Differentiation/drug effects , Mice, Inbred C57BL , Hand, Foot and Mouth Disease/virology , Hand, Foot and Mouth Disease/metabolism , Hand, Foot and Mouth Disease/pathology , Hand, Foot and Mouth Disease/complications , Phosphorylation/drug effects , Coxsackievirus Infections/complications , Coxsackievirus Infections/metabolism , Glycogen Synthase Kinase 3 beta/metabolism , Pyridines/pharmacology , PyrimidinesABSTRACT
Coxsackievirus B1 (CVB1), an enterovirus with multiple clinical presentations, has been associated with potential long-term consequences, including hand, foot, and mouth disease (HFMD), in some patients. However, the related animal models, transmission dynamics, and long-term tissue tropism of CVB1 have not been systematically characterized. In this study, we established a model of CVB1 respiratory infection in rhesus macaques and evaluated the clinical symptoms, viral load, and immune levels during the acute phase (0-14 days) and long-term recovery phase (15-30 days). We also investigated the distribution, viral clearance, and pathology during the long-term recovery period using 35 postmortem rhesus macaque tissue samples collected at 30 days postinfection (d.p.i.). The results showed that the infected rhesus macaques were susceptible to CVB1 and exhibited HFMD symptoms, viral clearance, altered cytokine levels, and the presence of neutralizing antibodies. Autopsy revealed positive viral loads in the heart, spleen, pancreas, soft palate, and olfactory bulb tissues. HE staining demonstrated pathological damage to the liver, spleen, lung, soft palate, and tracheal epithelium. At 30 d.p.i., viral antigens were detected in visceral, immune, respiratory, and muscle tissues but not in intestinal or neural tissues. Brain tissue examination revealed viral meningitis-like changes, and CVB1 antigen expression was detected in occipital, pontine, cerebellar, and spinal cord tissues at 30 d.p.i. This study provides the first insights into CVB1 pathogenesis in a nonhuman primate model of HFMD and confirms that CVB1 exhibits tissue tropism following long-term infection.
Subject(s)
Disease Models, Animal , Enterovirus B, Human , Hand, Foot and Mouth Disease , Macaca mulatta , Viral Load , Viral Tropism , Animals , Hand, Foot and Mouth Disease/virology , Hand, Foot and Mouth Disease/pathology , Enterovirus B, Human/physiology , Enterovirus B, Human/pathogenicity , Antibodies, Viral/blood , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Animals, Newborn , Cytokines/metabolismABSTRACT
The majority of solid testicular tumors are treated with orchiectomy given the high risk of malignancy. We present a case of a testicular mass in an adult patient in the setting of recent hand, foot, and mouth disease that was managed conservatively with serial ultrasounds. Even though cases of viral-associated testicular masses are rare, this differential diagnosis should be considered in patients with a new testicular mass in the setting of recent viral infection and negative tumor markers. For these patients, observation may be an option instead of immediate orchiectomy.
Subject(s)
Hand, Foot and Mouth Disease , Testicular Neoplasms , Male , Adult , Humans , Hand, Foot and Mouth Disease/pathology , Testis , Testicular Neoplasms/diagnosis , Testicular Neoplasms/surgery , Testicular Neoplasms/pathology , Orchiectomy , UltrasonographyABSTRACT
Coxsackievirus B3 (CVB3) is the pathogen causing hand, foot and mouth disease (HFMD), which manifests across a spectrum of clinical severity from mild to severe. However, CVB3-infected mouse models mainly demonstrate viral myocarditis and pancreatitis, failing to replicate human HFMD symptoms. Although several enteroviruses have been evaluated in Syrian hamsters and rhesus monkeys, there is no comprehensive data on CVB3. In this study, we have first tested the susceptibility of Syrian hamsters to CVB3 infection via different routes. The results showed that Syrian hamsters were successfully infected with CVB3 by intraperitoneal injection or nasal drip, leading to nasopharyngeal colonization, acute severe pathological injury, and typical HFMD symptoms. Notably, the nasal drip group exhibited a longer viral excretion cycle and more severe pathological damage. In the subsequent study, rhesus monkeys infected with CVB3 through nasal drips also presented signs of HFMD symptoms, viral excretion, serum antibody conversion, viral nucleic acids and antigens, and the specific organ damages, particularly in the heart. Surprisingly, there were no significant differences in myocardial enzyme levels, and the clinical symptoms resembled those often associated with common, mild infections. In summary, the study successfully developed severe Syrian hamsters and mild rhesus monkey models for CVB3-induced HFMD. These models could serve as a basis for understanding the disease pathogenesis, conducting pre-trial prevention and evaluation, and implementing post-exposure intervention.
Subject(s)
Disease Models, Animal , Enterovirus B, Human , Hand, Foot and Mouth Disease , Macaca mulatta , Mesocricetus , Animals , Hand, Foot and Mouth Disease/virology , Hand, Foot and Mouth Disease/pathology , Enterovirus B, Human/pathogenicity , Antibodies, Viral/blood , Cricetinae , Female , Virus Shedding , Nasopharynx/virology , MaleABSTRACT
Hand, foot and mouth disease (HFMD) caused by Human Enterovirus A71 (HEVA71) infection is typically a benign infection. However, in minority of cases, children can develop severe neuropathology that culminate in fatality. Approximately 36.9% of HEVA71-related hospitalizations develop neurological complications, of which 10.5% are fatal. Yet, the mechanism by which HEVA71 induces these neurological deficits remain unclear. Here, we show that HEVA71-infected astrocytes release CXCL1 which supports viral replication in neurons by activating the CXCR2 receptor-associated ERK1/2 signaling pathway. Elevated CXCL1 levels correlates with disease severity in a HEVA71-infected mice model. In humans infected with HEVA71, high CXCL1 levels are only present in patients presenting neurological complications. CXCL1 release is specifically triggered by VP4 synthesis in HEVA71-infected astrocytes, which then acts via its receptor CXCR2 to enhance viral replication in neurons. Perturbing CXCL1 signaling or VP4 myristylation strongly attenuates viral replication. Treatment with AZD5069, a CXCL1-specific competitor, improves survival and lessens disease severity in infected animals. Collectively, these results highlight the CXCL1-CXCR2 signaling pathway as a potential target against HFMD neuropathogenesis.
Subject(s)
Central Nervous System Diseases/virology , Chemokine CXCL1/metabolism , Enterovirus A, Human/metabolism , Hand, Foot and Mouth Disease/pathology , Receptors, Interleukin-8B/metabolism , Animals , Astrocytes/metabolism , Astrocytes/virology , Cell Line , Central Nervous System Diseases/pathology , Disease Models, Animal , Female , HEK293 Cells , Hand, Foot and Mouth Disease/virology , Humans , MAP Kinase Signaling System/physiology , Mice , Mice, Inbred BALB C , Pyrimidines/pharmacology , Rats , Severity of Illness Index , Sulfonamides/pharmacologyABSTRACT
BACKGROUND: This research aimed to explore the association between the RIG-I-like receptor (RIG-I and MDA5 encoded by DDX58 and IFIH1, respectively) pathways and the risk or severity of hand, foot, and mouth disease caused by enterovirus 71 (EV71-HFMD). In this context, we explored the influence of gene methylation and polymorphism on EV71-HFMD. METHODOLOGY/PRINCIPAL FINDINGS: 60 healthy controls and 120 EV71-HFMD patients, including 60 mild EV71-HFMD and 60 severe EV71-HFMD patients, were enrolled. First, MiSeq was performed to explore the methylation of CpG islands in the DDX58 and IFIH1 promoter regions. Then, DDX58 and IFIH1 expression were detected in PBMCs using RT-qPCR. Finally, imLDR was used to detect DDX58 and IFIH1 single-nucleotide polymorphism (SNP) genotypes. Severe EV71-HFMD patients exhibited higher DDX58 promoter methylation levels than healthy controls and mild EV71-HFMD patients. DDX58 promoter methylation was significantly associated with severe HFMD, sex, vomiting, high fever, neutrophil abundance, and lymphocyte abundance. DDX58 expression levels were significantly lower in mild patients than in healthy controls and lower in severe patients than in mild patients. Binary logistic regression analysis revealed statistically significant differences in the genotype frequencies of DDX58 rs3739674 between the mild and severe groups. GeneMANIA revealed that 19 proteins displayed correlations with DDX58, including DHX58, HERC5, MAVS, RAI14, WRNIP1 and ISG15, and 19 proteins displayed correlations with IFIH1, including TKFC, IDE, MAVS, DHX58, NLRC5, TSPAN6, USP3 and DDX58. CONCLUSIONS/SIGNIFICANCE: DDX58 expression and promoter methylation were associated with EV71 infection progression, especially in severe EV71-HFMD patients. The effect of DDX58 in EV71-HFMD is worth further attention.
Subject(s)
DEAD Box Protein 58/genetics , DNA Methylation/genetics , Hand, Foot and Mouth Disease/pathology , Interferon-Induced Helicase, IFIH1/genetics , Receptors, Immunologic/genetics , Child , Child, Preschool , CpG Islands/genetics , DEAD Box Protein 58/metabolism , Enterovirus A, Human , Female , Genetic Predisposition to Disease/genetics , Hand, Foot and Mouth Disease/virology , Humans , Infant , Interferon-Induced Helicase, IFIH1/metabolism , Male , Polymorphism, Single Nucleotide/genetics , Promoter Regions, Genetic/genetics , Receptors, Immunologic/metabolism , Severity of Illness IndexABSTRACT
Sporadic occurrences and outbreaks of hand, foot, and mouth disease (HFMD) caused by Coxsackievirus A2 (CVA2) have frequently reported worldwide recently, which pose a great challenge to public health. Epidemiological studies have suggested that the main cause of death in critical patients is pulmonary edema. However, the pathogenesis of this underlying comorbidity remains unclear. In this study, we utilized the 5-day-old BALB/c mouse model of lethal CVA2 infection to evaluate lung damage. We found that the permeability of lung microvascular was significantly increased after CVA2 infection. We also observed the direct infection and apoptosis of lung endothelial cells as well as the destruction of tight junctions between endothelial cells. CVA2 infection led to the degradation of tight junction proteins (e.g., ZO-1, claudin-5, and occludin). The gene transcription levels of von Willebrand factor (vWF), endothelin (ET), thrombomodulin (THBD), granular membrane protein 140 (GMP140), and intercellular cell adhesion molecule-1 (ICAM-1) related to endothelial dysfunction were all significantly increased. Additionally, CVA2 infection induced the increased expression of inflammatory cytokines (IL-6, IL-1ß, and MCP-1) and the activation of p38 mitogen-activated protein kinase (MAPK). In conclusion, the disruption of the endothelial barrier contributes to acute lung injury induced by CVA2 infection; targeting p38-MAPK signaling may provide a therapeutic approach for pulmonary edema in critical infections of HFMD.
Subject(s)
Acute Lung Injury/genetics , Coxsackievirus Infections/genetics , Hand, Foot and Mouth Disease/genetics , Pulmonary Edema/genetics , Acute Lung Injury/complications , Acute Lung Injury/pathology , Acute Lung Injury/virology , Animals , Apoptosis/genetics , Claudin-5/genetics , Coxsackievirus Infections/complications , Coxsackievirus Infections/pathology , Coxsackievirus Infections/virology , Cytokines/genetics , Disease Models, Animal , Endothelial Cells/pathology , Endothelial Cells/virology , Hand, Foot and Mouth Disease/complications , Hand, Foot and Mouth Disease/pathology , Hand, Foot and Mouth Disease/virology , Humans , Mice , Occludin/genetics , Pulmonary Edema/complications , Pulmonary Edema/pathology , Pulmonary Edema/virology , Tight Junctions/genetics , Tight Junctions/pathology , Zonula Occludens-1 Protein/genetics , p38 Mitogen-Activated Protein Kinases/geneticsABSTRACT
Hand, foot, and mouth disease (HFMD) caused by enterovirus A71 (EV-A71) is a contagious viral disease, and toll-like receptors (TLRs) play essential roles in resisting the pathogen. The aim of this study was to assess the potential relationship between several TLRs polymorphisms and the HFMD severity in a Chinese children population. A total of 328 Chinese children with HFMD were included in the present study. The polymorphisms of TLR3 (rs3775290, rs3775291, rs3775296, rs1879026, rs5743312, rs5743313, rs5743303, rs13126816, and rs3775292), TLR4 (rs4986790, rs4986791, rs2149356, rs11536889, and rs41426344), TLR7 (rs179009, rs179010, rs179016, rs3853839, rs2302267, rs1634323, and rs5741880), and TLR8 (rs3764880, rs2159377, rs2407992, rs5744080, rs3747414, rs3764879, and rs5744069) genes were selected. The study indicated that individuals with the GG genotype of TLR3 single-nucleotide polymorphism rs1879026 had a higher risk of developing severe cases (GG vs. GT: OR = 1.875; 95% CI, 1.183-2.971; p = .007). Meanwhile, TLR3 rs3775290 CC genotype and C allele were associated with lower disease severity in females (CC vs. CT: OR = 0.350; 95% CI, 0.163-0.751; p = .006; C vs. T: OR = 0.566; 95% CI, 0.332-0.965; p = .036). TLR3 rs3775291 CC genotype showed 2.537 folds higher risk of developing severe cases in females (CC vs. CT: OR = 2.537; 95% CI, 1.108-5.806; p = .026). Moreover, TLR3 rs1879026 GG genotype was found to be related to increased risk of severe cases in males (GG vs. GT: OR = 2.076; 95% CI, 1.144-3.768; p = .016). The current findings show that the genetic variants of TLR3 rs1879026, rs3775290, and rs3775291 are associated with the severity of EV-A71-associated HFMD in a Chinese children population.
Subject(s)
Enterovirus A, Human , Genetic Predisposition to Disease , Hand, Foot and Mouth Disease/genetics , Polymorphism, Single Nucleotide , Toll-Like Receptor 3/genetics , Alleles , Asian People/genetics , Case-Control Studies , Child, Preschool , China , Female , Hand, Foot and Mouth Disease/pathology , Hand, Foot and Mouth Disease/virology , Humans , Infant , Infant, Newborn , MaleABSTRACT
Severe hand, foot, and mouth disease (HFMD) caused by enterovirus 71 (EV71) infection is associated with high mortality and disability. DC-SIGN, a receptor for EV71, is widely distributed in dendritic cells and may influence the severity of HFMD caused by EV71 infection. This observational study attempts to explore whether single-nucleotide polymorphisms (SNPs) in DC-SIGN are related to the severity of EV71-associated HFMD. Based on linkage disequilibrium and functional predictions, two DC-SIGN SNPs were selected and tested to explore their potential association with the severity of HFMD caused by EV71 infection. Two hundred sixteen Han Chinese children with HFMD caused by EV71 were enrolled to obtain clinical data, including the severity of HFMD, serum DC-SIGN levels, and DC-SIGN SNPs. We found a significant association between the rs7248637 polymorphism (A vs. G: OR = 0.644, 95% CI = 0.515-0.806) and the severity of HFMD caused by EV71 infection, as well as the rs4804800 polymorphism (A vs. G: OR = 1.539, 95% CI =1.229-1.928). These two DC-SIGN SNPs may have an effect on the severity of HFMD caused by EV71 infection.
Subject(s)
Cell Adhesion Molecules/genetics , Enterovirus Infections/genetics , Genetic Predisposition to Disease/genetics , Hand, Foot and Mouth Disease/genetics , Lectins, C-Type/genetics , Receptors, Cell Surface/genetics , Asian People/genetics , Cell Adhesion Molecules/blood , Child , Child, Preschool , China/epidemiology , Enterovirus A, Human , Enterovirus Infections/epidemiology , Enterovirus Infections/pathology , Female , Genetic Association Studies , Genotype , Hand, Foot and Mouth Disease/epidemiology , Hand, Foot and Mouth Disease/pathology , Humans , Infant , Lectins, C-Type/blood , Male , Polymorphism, Single Nucleotide , Receptors, Cell Surface/blood , Severity of Illness IndexABSTRACT
Children with HFMD due to EV71 infection are more likely to suffer from neurogenic complications, leading to higher morbidity and mortality. ILCs play crucial roles in the initiation of host immunity. However, the contribution of ILCs to the occurrence and development of HFMD due to EV71 infection remains to be explored. The results of our study showed that the levels of peripheral ILC1s and Th1 cells were increased in children with severe HFMD compared to healthy children, as were ILC1- and Th1-related cytokines and transcription factors. Furthermore, HFMD children with a higher frequency of circulating ILC1s exhibited a 2.9-fold greater risk of severity when HFMD was accompanied by VEM. Our study is the first to show that ILC1 abnormalities contribute to the pathogenesis of the severity of HFMD, in which ILC1s are aberrant increased and affect the cellular and humoral immunity. ILC1s could be used in the diagnosis of HFMD.
Subject(s)
Enterovirus A, Human/immunology , Hand, Foot and Mouth Disease/immunology , Lymphocytes/immunology , Th1 Cells/immunology , Antibodies, Viral/immunology , B-Lymphocytes/cytology , B-Lymphocytes/immunology , Child, Preschool , Female , Hand, Foot and Mouth Disease/complications , Hand, Foot and Mouth Disease/pathology , Humans , Immunoglobulin M/immunology , Lymphocytes/cytology , Male , Severity of Illness Index , Th1 Cells/cytologyABSTRACT
Enterovirus 71 (EV-A71) and coxsackievirus A16 (CV-A16) are the major pathogens responsible for hand, foot and mouth disease (HFMD), but the mechanism by which these viruses cause disease remains unclear. In this study, we used transcriptome sequencing technology to investigate changes in the transcriptome profiles after infection with EV-A71 and CV-A16 in human bronchial epithelial (16HBE) cells. Using systematic bioinformatics analysis, we then searched for useful clues regarding the pathogenesis of HFMD. As a result, a total of 111 common differentially expressed genes were present in both EV-A71- and CV-A16-infected cells. A trend analysis of these 111 genes showed that 91 of them displayed the same trend in EV-A71 and CV-A16 infection, including 49 upregulated genes and 42 downregulated genes. These 91 genes were further used to conduct GO, pathway, and coexpression network analysis. It was discovered that enriched GO terms (such as histone acetylation and positive regulation of phosphorylation) and pathways (such as glycosylphosphatidylinositol (GPI)-anchor biosynthesis and DNA replication) might be closely associated with the pathogenic mechanism of these two viruses, and key genes (such as TBCK and GPC) might be involved in the progression of HFMD. Finally, we randomly selected 10 differentially expressed genes for qRT-PCR to validate the transcriptome sequencing data. The experimental qRT-PCR results were roughly in agreement with the results of transcriptome sequencing. Collectively, our results provide clues to the mechanism of pathogenesis of HFMD induced by EV-A71 and CV-A16.
Subject(s)
Enterovirus A, Human/pathogenicity , Epithelial Cells/virology , Gene Expression Profiling , Hand, Foot and Mouth Disease/pathology , Cell Line , DNA Replication , Epithelial Cells/physiology , Gene Expression Regulation , Hand, Foot and Mouth Disease/virology , HumansABSTRACT
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.
Subject(s)
Adaptation, Physiological , Capsid Proteins , Enterovirus A, Human , Genome, Viral , Mutation, Missense , Receptors, Virus , Amino Acid Substitution , Animals , Capsid Proteins/genetics , Capsid Proteins/metabolism , Chlorocebus aethiops , Enterovirus A, Human/genetics , Enterovirus A, Human/metabolism , Hand, Foot and Mouth Disease/genetics , Hand, Foot and Mouth Disease/metabolism , Hand, Foot and Mouth Disease/pathology , Humans , Receptors, Virus/metabolism , Vero CellsABSTRACT
Hand, foot, and mouth disease (HFMD) is most frequently caused by several serotypes of human enterovirus (EV) including Enterovirus 71 (EV-A71), coxsackievirus A16 (CV-A16), or other types of EV. The aim of this study was to determine the epidemiological characteristics of HFMD and to describe the epidemiologic characteristics of HFMD among severe and mild cases. We collected 4760 HFMD cases in Hangzhou from 2016 to 2018. Specimens from these cases were collected and tested for EV-A71, CV-A16, CV-A6, CV-A10, CV-A2, and CV-A5 by reverse transcriptase polymerase chain reaction. From 2016 to 2018, the prevalence of HFMD was seasonal each year. Among the 4760 probable HFMD cases, 3559 cases were confirmed (74.8%), including 426 cases of EV-A71 infections (8.9%), 249 cases of CV-A16 infections (5.2%), and 2884 cases of other EV infections (60.6%). The percentage of other EV infections was more than 80%, which increased year by year. Random selection of samples for detection of other EV infections in 2017 and 2018, among the 1297 cases, showed there were 835 (64.4%) cases of CV-A6 infections, 177 (13.6%) cases of CV-A10 infections, 100 (7.7%) cases of CV-A2 infections, 40 (3.1%) cases of CV-A5 infections, 3 (0.02 %) cases of mixed infections, and 11.0% untyped EV infections. Preschool children were still the primary population susceptible to HFMD. In severe cases, EV-A71 infection was the main cause. Characterizing the epidemiology and the relationship between severe and common cases of HFMD would provide relevant evidences for the prevention and treatment of HFMD.
Subject(s)
Hand, Foot and Mouth Disease/epidemiology , Hand, Foot and Mouth Disease/pathology , Age Factors , Child, Preschool , China/epidemiology , Female , Hand, Foot and Mouth Disease/physiopathology , Humans , Infant , Male , Real-Time Polymerase Chain ReactionABSTRACT
In recent years, outbreaks of hand-foot-mouth disease (HFMD) in China, Singapore and other Western Pacific Region, involving millions of children, have become a big threat to public health. This study aimed to quantitatively assess all qualified studies and identify the risk factors for HFMD death. A systematic search of the databases PubMed, Medline, Embase and the Cochrane Library was performed. Study heterogeneity and publication bias were estimated. Seven case-control studies involving 1641 participants (634 died and 1007 survived) were included in the meta-analysis. Human enterovirus 71 infection, male, age ⩽3 years, vomiting, cyanosis, convulsion, duration of fever ⩾3 days, atypical rashes and abdominal distention were not significantly related to HFMD death (P ⩽ 0.05). Lethargy (odds ratio (OR) = 6.62; 95% CI 3.61-12.14; I2 = 0%; P < 0.0001), pneumonoedema/pneumorrhagia (OR = 4.09; 95% CI 2.44-6.87; I2 = 0%; P < 0.0001), seizures (OR = 6.85; 95% CI 2.37-19.74; I2 = 0%; P = 0.0004), dyspnoea (OR = 8.24; 95% CI 2.05-33.19; I2 = 83%; P = 0.003) and coma (OR = 3.76; 95% CI 1.85-7.67; I2 = 0%; P = 0.0003) were significantly associated with HFMD death, which were risk factors for HFMD death.
Subject(s)
Enterovirus A, Human/isolation & purification , Hand, Foot and Mouth Disease/mortality , Asia/epidemiology , Child, Preschool , Female , Hand, Foot and Mouth Disease/pathology , Humans , Incidence , Infant , Male , Risk Factors , Survival AnalysisABSTRACT
Enterovirus 71 (EV71) is a causative agent of hand-foot-mouth disease, and it sometimes causes severe neurological disease. Development of effective vaccines and animal models to evaluate vaccine candidates are needed. However, the animal models currently used for vaccine efficacy testing, monkeys and neonatal mice, have economic, ethical, and practical drawbacks. In addition, EV71 strains prepared for lethal challenge often develop decreased virulence during propagation in cell culture. To overcome these problems, we used a mouse model expressing human scavenger receptor B2 (hSCARB2) that showed lifelong susceptibility to EV71. We selected virulent EV71 strains belonging to the subgenogroups B4, B5, C1, C2, and C4 and propagated them using a culture method for EV71 without an apparent reduction in virulence. Here, we describe a novel EV71 vaccine efficacy test based on these hSCARB2 transgenic (Tg) mice and these virulent viruses. Adult Tg mice were immunized subcutaneously with formalin-inactivated EV71. The vaccine elicited sufficient levels of neutralizing antibodies in the immunized mice. The mice were subjected to lethal challenge with virulent viruses via intravenous injection. Survival, clinical signs, and body weight changes were observed for 2 weeks. Most immunized mice survived without clinical signs or histopathological lesions. The viral replication in immunized mice was much lower than that in nonimmunized mice. Mice immunized with the EV71 vaccine were only partially protected against lethal challenge with coxsackievirus A16. These results indicate that this new model is useful for in vivo EV71 vaccine efficacy testing.IMPORTANCE The development of new vaccines for EV71 relies on the availability of small animal models suitable for in vivo efficacy testing. Monkeys and neonatal mice have been used, but the use of these animals has several drawbacks, including high costs, limited susceptibility, and poor experimental reproducibility. In addition, the related ethical issues are considerable. The new efficacy test based on hSCARB2 Tg mice and virulent EV71 strains propagated in genetically modified cell lines presented here can overcome these disadvantages and is expected to accelerate the development of new EV71 vaccines.
Subject(s)
Enterovirus A, Human/immunology , Hand, Foot and Mouth Disease/prevention & control , Lysosomal Membrane Proteins/immunology , Receptors, Scavenger/immunology , Viral Vaccines/pharmacology , Animals , Cell Line , Disease Models, Animal , Drug Evaluation , Enterovirus A, Human/genetics , Enterovirus A, Human/pathogenicity , Hand, Foot and Mouth Disease/genetics , Hand, Foot and Mouth Disease/immunology , Hand, Foot and Mouth Disease/pathology , Humans , Lysosomal Membrane Proteins/genetics , Mice , Mice, Transgenic , Receptors, Scavenger/genetics , Vaccines, Inactivated/genetics , Vaccines, Inactivated/immunology , Vaccines, Inactivated/pharmacology , Viral Vaccines/genetics , Viral Vaccines/immunologyABSTRACT
Enterovirus 71 (EV-A71) is one of the major causative agents of hand, foot, and mouth disease. EV-A71 infection is sometimes associated with severe neurological diseases such as acute encephalitis, acute flaccid paralysis, and cardiopulmonary failure. Therefore, EV-A71 is a serious public health concern. Scavenger receptor class B, member 2 (SCARB2) is a type III transmembrane protein that belongs to the CD36 family and is a major receptor for EV-A71. SCARB2 supports attachment and internalization of the virus and initiates conformational changes that lead to uncoating of viral RNA in the cytoplasm. The three-dimensional structure of the virus-receptor complex was elucidated by cryo-electron microscopy. Two α-helices in the head domain of SCARB2 bind to the G-H loop of VP1 and the E-F loop of VP2 capsid proteins of EV-A71. Uncoating takes place in a SCARB2- and low pH-dependent manner. In addition to SCARB2, other molecules support cell surface binding of EV-A71. Heparan sulfate proteoglycans, P-selectin glycoprotein ligand-1, sialylated glycan, annexin II, vimentin, fibronectin, and prohibitin enhance viral infection by retaining the virus on the cell surface. These molecules are known as "attachment receptors" because they cannot initiate uncoating. In vivo, SCARB2 expression was observed in EV-A71 antigen-positive neurons and epithelial cells in the crypts of the palatine tonsils in patients that died of EV-A71 infection. Adult mice are not susceptible to infection by EV-A71, but transgenic mice that express human SCARB2 become susceptible to EV-A71 infection and develop neurological diseases similar to those observed in humans. Attachment receptors may also be involved in EV-A71 infection in vivo. Although heparan sulfate proteoglycans are expressed by many cultured cell lines and enhance infection by a subset of EV-A71 strains, they are not expressed by cells that express SCARB2 at high levels in vivo. Thus, heparan sulfate-positive cells merely adsorb the virus and do not contribute to replication or dissemination of the virus in vivo. In addition to these attachment receptors, cyclophilin A and human tryptophanyl aminoacyl-tRNA synthetase act as an uncoating regulator and an entry mediator that can confer susceptibility to non-susceptibile cells in the absence of SCARB2, respectively. The roles of attachment receptors and other molecules in EV-A71 pathogenesis remain to be elucidated.