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The mechanisms underlying pathological changes in the central nervous system (CNS) following Coxsackievirus A16 (CV-A16) infection have not yet been elucidated. IFN-γ-inducible protein-10 (IP-10) is often used as a predictive factor to monitor early virus infection. It has also been reported that IP-10 plays a pivotal role in neuroinflammation. In this study, we aimed to explore the role of IP-10 in the neuropathogenesis of CV-A16 infection. We observed that the level of IP-10, as well as the TLR3-TRIF-TRAF3-TBK1-NF-κB and RIG-I/MDA5-MAVS-TRAFS-TBK1-NF-κB pathways, which are the upstream of IP-10, were significantly elevated during the course of CV-A16 infection. This increase was accompanied by an increase in a series of inflammatory cytokines at different time-points during CV-A16 infection. To determine whether IP-10 influences BBB integrity, we examined junctional complexes. Our results revealed that the expression levels of Claudin5, Occludin, ZO-1 and VE-Cadherin were notably decreased in CV-A16-infected HUVECs, but these indicators were restored in CV-A16-infected HUVECs with Eldelumab treatment. Nevertheless, IP-10 is only a chemokine that primarily traffics CXCR3-positive immune cells to inflammatory sites or promotes the production of inflammatory cytokines. Therefore, the interactions between IP-10 and inflammatory cytokines were evaluated. Our data revealed that IP-10 mediated the production of TNF-α, which was also observed to change the junctional complexes. Moreover, in a suckling mouse model, IP-10 and TNF-α treatments exacerbated clinical symptoms, mortality and pathological changes in the brain of CV-A16-infected mice, but Anti-IP-10 and Anti-TNF-α treatments alleviated these changes. Our data also revealed that IP-10 may be detected early in CV-A16 infection, whereas TNF-α was detected late in CV-A16 infection, and the production of TNF-α was also found to be positively correlated with IP-10. In addition, IP-10 and TNF-α were observed to reduce junctional complexes and enhance virus entry into the CNS. Taken together, this study provides the first evidence that CV-A16 activates the IP-10/TNF-α regulatory axis to cause BBB damage and accelerate the formation of neuroinflammation in infected hosts, which not only provides a new understanding of the neuropathogenesis caused by CV-A16, but also offers a promising target for the development of CV-A16 antiviral drugs. [ABSTRACT FROM AUTHOR]

Introduction: Coxsackievirus A6 (CV-A6) has emerged as the predominant epidemic strain responsible for hand, foot and mouth disease (HFMD). CV-A6 infection can result in severe clinical manifestations, including encephalitis, meningitis, and potentially life-threatening central nervous system disorders. Our previous research findings demonstrated that neonatal mice infected with CV-A6 exhibited limb weakness, paralysis, and ultimately succumbed to death. However, the underlying mechanism of CV-A6-induced nervous system injury remains elusive. Numerous reports have highlighted the pivotal role of miRNAs in various viral infections. Methods: Separately established infection and control groups of mice were used to create miRNA profiles of the brain tissues before and after CV-A6 transfection, followed by experimental verification, prediction, and analysis of the results. Results: At 2 days post-infection (dpi), 4 dpi, and 2dpi vs 4dpi, we identified 175, 198 and 78 significantly differentially expressed miRNAs respectively using qRT-PCR for validation purposes. Subsequently, we predicted target genes of these differentially expressed miRNAs and determined their potential targets through GO (Gene Ontology) enrichment analysis and KEGG (Kyoto Encyclopedia of Genes and Genomes) enrichment analysis. Finally, we verified the miRNA-mRNA pairing via double luciferase experiments while confirming functional enrichment of target genes through Western Blotting analyses. Discussion: The results from this study suggest that transcriptional regulation, neuronal necrosis, pro-inflammatory cytokine release, and antiviral immunity are all implicated in the pathogenesis of central nervous systeminjury in mice infected with CV-A6. Brain injury resulting from CV-A6 infection may involve multiple pathways, including glial cell activation, neuronal necrosis, synaptic destruction, degenerative diseases of the nervous system. It can even encompass destruction of the blood-brain barrier, leading to central nervous system injury. The dysregulated miRNAs and signaling pathways discovered in this study provide valuable insights for further investigations into the pathogenesis of CV-A6. [ABSTRACT FROM AUTHOR]

The mechanisms underlying pathological changes in the central nervous system (CNS) following Coxsackievirus A16 (CV-A16) infection have not yet been elucidated. IFN-γ-inducible protein-10 (IP-10) is often used as a predictive factor to monitor early virus infection. It has also been reported that IP-10 plays a pivotal role in neuroinflammation. In this study, we aimed to explore the role of IP-10 in the neuropathogenesis of CV-A16 infection. We observed that the level of IP-10, as well as the TLR3-TRIF-TRAF3-TBK1-NF-κB and RIG-I/MDA5-MAVS-TRAFS-TBK1-NF-κB pathways, which are the upstream of IP-10, were significantly elevated during the course of CV-A16 infection. This increase was accompanied by an increase in a series of inflammatory cytokines at different time-points during CV-A16 infection. To determine whether IP-10 influences BBB integrity, we examined junctional complexes. Our results revealed that the expression levels of Claudin5, Occludin, ZO-1 and VE-Cadherin were notably decreased in CV-A16-infected HUVECs, but these indicators were restored in CV-A16-infected HUVECs with Eldelumab treatment. Nevertheless, IP-10 is only a chemokine that primarily traffics CXCR3-positive immune cells to inflammatory sites or promotes the production of inflammatory cytokines. Therefore, the interactions between IP-10 and inflammatory cytokines were evaluated. Our data revealed that IP-10 mediated the production of TNF-α, which was also observed to change the junctional complexes. Moreover, in a suckling mouse model, IP-10 and TNF-α treatments exacerbated clinical symptoms, mortality and pathological changes in the brain of CV-A16-infected mice, but Anti-IP-10 and Anti-TNF-α treatments alleviated these changes. Our data also revealed that IP-10 may be detected early in CV-A16 infection, whereas TNF-α was detected late in CV-A16 infection, and the production of TNF-α was also found to be positively correlated with IP-10. In addition, IP-10 and TNF-α were observed to reduce junctional complexes and enhance virus entry into the CNS. Taken together, this study provides the first evidence that CV-A16 activates the IP-10/TNF-α regulatory axis to cause BBB damage and accelerate the formation of neuroinflammation in infected hosts, which not only provides a new understanding of the neuropathogenesis caused by CV-A16, but also offers a promising target for the development of CV-A16 antiviral drugs.

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. [ABSTRACT FROM AUTHOR]

BackgroundHand, foot, and mouth disease (HFMD) is endemic in Singapore. Prevention efforts have been ramped up since major outbreaks in the early 2000's. This study aims to assess the current knowledge, and attitudes towards and practise (KAP) levels of HFMD prevention strategies (HFMD-PS) amongst parents and teachers of children under 5 years amidst the COVID-19 pandemic.Methods and resultsA convenience sample of 240 teachers and 404 parents responded to a self-administered standardised questionnaire between mid-October and December 2020. A scoring framework was used to assess responses in the ‘knowledge', ‘attitude', and ‘practice' domains. A multivariable analysis was adjusted for ethnicity and attitudes towards getting children to follow proper handwashing steps and regularly disinfecting children's toys amongst parents, knowledge about HFMD's infectious period, and the responses to a child turning symptomatic in the childcare centre amongst teachers. Existing levels of knowledge and attitudes of parents and teachers were not high, and only a small proportion practised high levels of prevention measures (99 parents and 28 teachers). Key facilitators for a higher practise level in parents include the following: (1) awareness of regular liquid soap's efficacy as a disinfectant, (2) toy cleaning before and after playtime, and (3) the cleaning agent used for this practise. Teachers had no significant factors associated with higher practise levels.ConclusionThis study suggested potential gaps between positive knowledge and attitudes towards prevention strategies and their actual adoption levels in homes and childcare centres during COVID-19 pandemic. These evidences suggest the importance of continuous promotion of HFMD prevention practise in homes and childcare centres, even amidst pandemics.

Enterovirus 71 (EV71) infection causes hand, foot, and mouth disease (HFMD), and even fatal neurological complications. However, the mechanisms underlying EV71 neurological pathogeneses are largely unknown. This study reveals a distinct mechanism by which EV71 induces apoptosis and autophagy in neural cells. EV71 non-structure protein 3D (also known as RNA-dependent RNA polymerase, RdRp) interacts with the peroxisomal protein acyl-CoA oxidase 1 (ACOX1), and contributes to ACOX1 downregulation. Further studies demonstrate that EV71 reduces peroxisome numbers. Additionally, knockdown of ACOX1 or peroxin 19 (PEX19) induces apoptosis and autophagy in neural cells including human neuroblastoma (SK-N-SH) cells and human astrocytoma (U251) cells, and EV71 infection induces neural cell death through attenuating ACOX1 production. Moreover, EV71 infection and ACOX1 knockdown facilitate reactive oxygen species (ROS) production and attenuate the cytoprotective protein deglycase (DJ-1)/Nuclear factor erythroid 2-related factor 2 (NRF2)/Heme oxygenase 1 (HO-1) pathway (DJ-1/NRF2/HO-1), which collectively result in ROS accumulation in neural cells. In conclusion, EV71 downregulates ACOX1 protein expression, reduces peroxisome numbers, enhances ROS generation, and attenuates the DJ-1/NRF2/HO-1 pathway, thereby inducing apoptosis and autophagy in neural cells. These findings provide new insights into the mechanism underlying EV71-induced neural pathogenesis, and suggest potential treatments for EV71-associated diseases.

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., Competing Interests: Conflict of interest The authors of this study declared that they do not have any conflict of interest., (Copyright © 2024 The Authors. Publishing services by Elsevier B.V. All rights reserved.)

Enterovirus 71 (EV-A71) and coxsackievirus A16 (CV-A16) remain the predominant etiological agents of hand, foot, and mouth disease (HFMD). The clinical manifestations caused by the two viruses are obviously different. CV-A16 usually triggers a repeated infection, and airway epithelial integrity is often the potential causative factor of respiratory repeated infections. Our previous studies have demonstrated that there were some differentially expressed miRNAs involved in the regulation of adhesion function of epithelial barrier in EV-A71 and CV-A16 infections. In this study, we compared the differences between EV-A71 and CV-A16 infections on the airway epithelial barrier function in human bronchial epithelial (16HBE) cells and further screened the key miRNA which leaded to the formation of these differences. Our results showed that more rapid proliferation, more serious destruction of 16HBE cells permeability, more apoptosis and disruption of intercellular adhesion-associated molecules were found in CV-A16 infection as compared to EV-A71 infection. Furthermore, we also identified that microRNA-4516 (miR-4516), which presented down-regulation in EV-A71 infection and up-regulation in CV-A16 infection was an important regulator of intercellular junctions by targeting Poliovirus receptor related protein 1(PVRL1). The expressions of PVRL1, claudin4, ZO-1 and E-cadherin in CV-A16-infected cells were significantly less than those in EV-A71-infected cells, while the expressions of these proteins were subverted when pre-treated with miR-4516-overexpression plasmid in EV-A71 infected and miR-4516-knockdown plasmid in CV-A16 infected 16HBE cells. Thus, these data suggested that the opposite expression of miR-4516 in EV-A71 and CV-A16 infections might be the initial steps leading to different epithelial impairments of 16HBE cells by destroying intercellular adhesion, which finally resulted in different outcomes of EV-A71 and CV-A16 infections.

Enterovirus 71 (EV-A71) and coxsackievirus A16 (CV-A16) remain the predominant etiological agents of hand, foot, and mouth disease (HFMD). The clinical manifestations caused by the two viruses are obviously different. CV-A16 usually triggers a repeated infection, and airway epithelial integrity is often the potential causative factor of respiratory repeated infections. Our previous studies have demonstrated that there were some differentially expressed miRNAs involved in the regulation of adhesion function of epithelial barrier in EV-A71 and CV-A16 infections. In this study, we compared the differences between EV-A71 and CV-A16 infections on the airway epithelial barrier function in human bronchial epithelial (16HBE) cells and further screened the key miRNA which leaded to the formation of these differences. Our results showed that more rapid proliferation, more serious destruction of 16HBE cells permeability, more apoptosis and disruption of intercellular adhesion-associated molecules were found in CV-A16 infection as compared to EV-A71 infection. Furthermore, we also identified that microRNA-4516 (miR-4516), which presented down-regulation in EV-A71 infection and up-regulation in CV-A16 infection was an important regulator of intercellular junctions by targeting Poliovirus receptor related protein 1(PVRL1). The expressions of PVRL1, claudin4, ZO-1 and E-cadherin in CV-A16-infected cells were significantly less than those in EV-A71-infected cells, while the expressions of these proteins were subverted when pre-treated with miR-4516-overexpression plasmid in EV-A71 infected and miR-4516-knockdown plasmid in CV-A16 infected 16HBE cells. Thus, these data suggested that the opposite expression of miR-4516 in EV-A71 and CV-A16 infections might be the initial steps leading to different epithelial impairments of 16HBE cells by destroying intercellular adhesion, which finally resulted in different outcomes of EV-A71 and CV-A16 infections. [ABSTRACT FROM AUTHOR]

Purpose: Hand, Foot and Mouth disease (HFMD) is a contagious pediatric viral disease caused due to enteroviruses (EV) of the family Picornaviridae. Cases of HFMD were reported from a tertiary care health centre, Udhampur, (Jammu and Kashmir), Northern India. The present study highlights the clinical and molecular virological aspects of HFMD cases., Material and Methods: Cases reported during August 2016-September 2017, and clinically diagnosed as HFMD of all age groups were included. Clinical, Biochemical and molecular virology aspects were compared. Clinical samples (n ​= ​50) such as vesicle swab, buccal and throat swabs were collected for enterovirus detection. EV-RNA was detected by 5'NCR based RT-PCR and genotyping by VP1 gene amplification and cycle sequencing., Results: Of the cases of HFMD enrolled (n ​= ​50), highest (84%) were of children aged <5 years, presented either or both anathemas and exanthemas with prodromal symptoms (fever, irritability). Clinical presentations involved mainly oral ulcers on lips and tongue (48%). Oral erosions were either single or multiple in numbers. Exanthemas were seen on hand and palm, widely spread up to buttocks, legs, arms and trunk. Of these, six patients were found anemic. Complete blood count (CBC) indicated lymphocytosis and C-reactive protein (n ​= ​10) in children aged <5 years. EV-RNA was detected in 78% (39/50) of the clinical samples. VP1 gene based typing indicated the presence of CV-A16, CVA6 and EV-A71 types., Conclusions: The study highlights association of EVs in HFMD cases in the reported region. CV-A16, CV-A6 and EV-A71 types were reported for the first time from Udhampur (J&K), Northern India. No differences were observed in the clinical profile of EV strains detected. Circulation of the strains warrant and alarm outbreaks. More focused studies on HFMD and monitoring of viral strains is mandatory., Competing Interests: Conflicts of interest There are no conflicts of interest., (Copyright © 2022 Indian Association of Medical Microbiologists. Published by Elsevier B.V. All rights reserved.)

Background A rapid expansion of hand, foot, and mouth disease (HFMD) outbreaks has occurred and caused deaths in China in recent years, but little is known about the other etiologic agents except enterovirus 71 (EV71) and coxsackievirus A 16 (CA16). The objective of this study is to determine the genotype compositions of enterovirus causing HFMD in Shanghai and identify any associations between enterovirus types and clinical manifestations. Methods Stool specimens were collected from patients hospitalized for treatment of HFMD, from May 2010 to April 2011. Enterovirus was detected by reverse transcription PCR and directly genotyped by sequencing the PCR products. Phylogenetic analysis was based on the VP1 partial gene. Results Of 290 specimens, 277 (95.5%) tested positive for enterovirus. The major genotypes were EV71 (63.8%), CA10 (9.0%), CA6 (8.3%), CA16 (6.9%), CA12 (2.4%), and CA4 (1.4%). The EV71 strains belonged to the C4a subtype and CA16 belonged to the B subtype. CA6 was closely related to strains detected in Japan, Taiwan and China, and CA10, CA12 and CA4 were phylogenetically similar to other strains circulating in China. Mean hospital stays and the prevalence of complications in patients with EV71 infection were higher than those in patients in CA6, CA10 or CA16 infection (P < 0.05 for all comparisons). Children with CA12 infection were the youngest, and most likely have the highest risk of complications when compared to the other non-EV71 infection groups. Conclusions This study demonstrated a diversified pathogen compositions attributing to HFMD and clinical symptoms differing in enterovirus genotypes. It deserves our attention as early identification of enterovirus genotypes is important for diagnosis and treatment of HFMD patients. [ABSTRACT FROM AUTHOR]

Hand, foot, and mouth disease (HFMD), with vesiculae on the hands, feet and mouth, is an infectious disease caused by many viral pathogens. However, the differences of immune response induced by these pathogens are unclear. We compared the clinical manifestations and the levels of immunologic indicators from 60 HFMD patients caused by different viral pathogens to analyze the differences in the immune response. It was shown that Th2 cytokines (IL-4 and IL-10) increased significantly in EV71-infected children; Th1 cytokines (IL-2 and IFN-γ) rose in CA16-infected children; both Th1 and Th2 cytokines elevated in non-EVG-infected children; only individual cytokines (such as IL-10) went up in EVG-infected children. Meanwhile, the antibodies induced by viral infection could not cross-interfere between the different pathogens. These differences might be due to variations in the immune response induced by the individual pathogens or to the pathogenesis of the infections by the individual pathogens.