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Yoke Fun Chan - One of the best experts on this subject based on the ideXlab platform.
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recent advances in the understanding of Enterovirus A71 infection a focus on neuropathogenesis
Expert Review of Anti-infective Therapy, 2021Co-Authors: Han Kang Tee, I-ching Sam, Mohd Izwan Zainol, Yoke Fun ChanAbstract:Introduction: Hand, foot, and mouth disease caused by Enterovirus A71 (EV-A71) is more frequently associated with neurological complications and deaths compared to other Enteroviruses.Areas covered...
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immune responses against Enterovirus A71 infection implications for vaccine success
Reviews in Medical Virology, 2019Co-Authors: Kam Leng Awyong, Chee Wah Tan, I-ching Sam, Nik Mohd Nasir Niknadia, Yoke Fun ChanAbstract:Enterovirus A71 (EV-A71) from the Picornaviridae family is an important emerging pathogen causing hand, foot, and mouth disease (HFMD) outbreaks worldwide. EV-A71 also caused fatal neurological complications in young children especially in Asia. On the basis of seroepidemiological studies from many Asian countries, EV-A71 infection is very common. Children of very young age are particularly vulnerable. Large-scale epidemics that occur every 3 to 4 years are associated with accumulation of an immunologically naive younger population. Capsid proteins especially VP1 with the presence of major B- and T-cell epitopes are the most antigenic proteins. The nonstructural proteins mainly contribute to T-cell epitopes that induce cross-reactive immune responses against other Enteroviruses. Dominant epitopes and their neutralization magnitudes differ in mice, rabbits, and humans. Neutralizing antibody is sufficient for immune protection, but poorer cellular immunity may lead to severe neurological complications and deaths. Some chemokines/cytokines are consistently found in severely ill patients, for example, IL-6, IL-10, IL-17A, MCP-1, IL-8, MIG, IP-10, IFN-γ, and G-CSF. An increase in white cell counts is a risk factor for severe HFMD. Recent clinical trials on EV-A71 inactivated vaccine showed >90% efficacy and a robust neutralization response that was protective, indicating neutralizing antibody correlates for protection. No protection against other Enteroviruses was observed. A comprehensive understanding of the immune responses to EV-A71 infection will benefit the development of diagnostic tools, potential therapeutics, and subunit vaccine candidates. Future development of a multivalent Enterovirus vaccine will require knowledge of correlates of protection, understanding of cross-protection and memory T-cell responses among Enteroviruses.
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2BC Non-Structural Protein of Enterovirus A71 Interacts with SNARE Proteins to Trigger Autolysosome Formation
Viruses, 2017Co-Authors: Jeffrey K. F. Lai, I-ching Sam, Pauline Verlhac, Joël Baguet, Eeva-liisa Eskelinen, Mathias Faure, Yoke Fun ChanAbstract:Viruses have evolved unique strategies to evade or subvert autophagy machinery. Enterovirus A71 (EV-A71) induces autophagy during infection in vitro and in vivo. In this study, we report that EV-A71 triggers autolysosome formation during infection in human rhabdomyosarcoma (RD) cells to facilitate its replication. Blocking autophagosome-lysosome fusion with chloroquine inhibited virus RNA replication, resulting in lower viral titres, viral RNA copies and viral proteins. Overexpression of the non-structural protein 2BC of EV-A71 induced autolysosome formation. Yeast 2-hybrid and co-affinity purification assays showed that 2BC physically and specifically interacted with a N-ethylmaleimide-sensitive factor attachment receptor (SNARE) protein, syntaxin-17 (STX17). Co-immunoprecipitation assay further showed that 2BC binds to SNARE proteins, STX17 and synaptosome associated protein 29 (SNAP29). Transient knockdown of STX17, SNAP29, and microtubule-associated protein 1 light chain 3B (LC3B), crucial proteins in the fusion between autophagosomes and lysosomes) as well as the lysosomal-associated membrane protein 1 (LAMP1) impaired production of infectious EV-A71 in RD cells. Collectively, these results demonstrate that the generation of autolysosomes triggered by the 2BC non-structural protein is important for EV-A71 replication, revealing a potential molecular pathway targeted by the virus to exploit autophagy. This study opens the possibility for the development of novel antivirals that specifically target 2BC to inhibit formation of autolysosomes during EV-A71 infection.
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VP1 residues around the five-fold axis of Enterovirus A71 mediate heparan sulfate interaction
Virology, 2016Co-Authors: Chee Wah Tan, I-ching Sam, Vannajan Sanghiran Lee, Hui Vern Wong, Yoke Fun ChanAbstract:Enterovirus A71 (EV-A71) is a neurotropic Enterovirus that uses heparan sulfate as an attachment receptor. The molecular determinants of EV-A71-heparan sulfate interaction are unknown. With In silico heparin docking and mutagenesis of all possible lysine residues in VP1, we identified that K162, K242 and K244 are responsible for heparin interaction and inhibition. EV-A71 mutants with K242A and K244A rapidly acquired compensatory mutations, T100K or E98A, and Q145R-T237N respectively, which restored the heparin-binding phenotype. Both VP1-98 and VP1-145 modulates heparin binding. Heparin-binding phenotype was completely abolished with VP1-E98-E145, but was restored by an E98K or E145Q substitution. During cell culture adaptation, EV-A71 rapidly acquired K98 or Q/G145 to restore the heparin-binding phenotype. Together with next-generation sequencing analysis, our results implied that EV-A71 has high genetic plasticity by modulating positively-charged residues at the five-fold axis during in vitro heparin adaptation. Our finding has impact on EV-A71 vaccine production, evolutionary studies and pathogenesis.
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Enterovirus A71 dna launched infectious clone as a robust reverse genetic tool
PLOS ONE, 2016Co-Authors: Chee Wah Tan, I-ching Sam, Han Kang Tee, Michelle Hui Pheng Lee, Yoke Fun ChanAbstract:Enterovirus A71 (EV-A71) causes major outbreaks of hand, foot and mouth disease, and is occasionally associated with neurological complications and death in children. Reverse genetics is widely used in the field of virology for functional study of viral genes. For EV-A71, such tools are limited to clones that are transcriptionally controlled by T7/SP6 bacteriophage promoter. This is often time-consuming and expensive. Here, we describe the development of infectious plasmid DNA-based EV-A71 clones, for which EV-A71 genome expression is under transcriptional control by the CMV-intermediate early promoter and SV40 transcriptional-termination signal. Transfection of this EV-A71 infectious DNA produces good virus yield similar to in vitro-transcribed EV-A71 infectious RNA, 6.4 and 5.8 log10PFU/ml, respectively. Infectious plasmid with enhanced green fluorescence protein and Nano luciferase reporter genes also produced good virus titers, with 4.3 and 5.0 log10 PFU/ml, respectively. Another infectious plasmid with both CMV and T7 promoters was also developed for easy manipulation of in vitro transcription or direct plasmid transfection. Transfection with either dual-promoter infectious plasmid DNA or infectious RNA derived from this dual-promoter clone produced infectious viral particles. Incorporation of hepatitis delta virus ribozyme, which yields precise 3' ends of the DNA-launched EV-A71 genomic transcripts, increased infectious viral production. In contrast, the incorporation of hammerhead ribozyme in the DNA-launched EV-A71 resulted in lower virus yield, but improved the virus titers for T7 promoter-derived infectious RNA. This study describes rapid and robust reverse genetic tools for EV-A71.
Shin-ru Shih - One of the best experts on this subject based on the ideXlab platform.
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Enterovirus A71 Vaccines.
Vaccines, 2021Co-Authors: Shin-ru Shih, Blanton S. Tolbert, Gary BrewerAbstract:Enterovirus A71 (EV-A71) is a major causative agent of hand, foot, and mouth disease (HFMD) and herpangina. Moreover, EV-A71 infection can lead to neurological complications and death. Vaccination is the most efficient way to control virus infection. There are currently three inactivated, whole EV-A71 vaccines licensed by the China NMPA (National Medical Products Administration). Several other types of vaccines, such as virus-like particles and recombinant VP1 (capsid protein), are also under development. In this review, we discuss recent advances in the development of EV-A71 vaccines.
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effect of a neuropilin 1 derived virus receptor trap on Enterovirus A71 infection in vitro
Antimicrobial Agents and Chemotherapy, 2020Co-Authors: Hsiang Ching Wang, Pengnien Huang, Hui Chen Hung, Sung Nien Tseng, Chia Chia Chang, Ya Ru Tsai, Yunming Wang, Shin-ru ShihAbstract:We discovered that neuropilin 1 (NRP1) is a new receptor candidate to mediate Enterovirus A71 (EVA71) into cells. In the engineered form as a decoy receptor, NRP1 was able to recognize and neutralize EVA71 but not Enterovirus D68 or coxsackievirus B3 (CVB3). NRP1 recognizes EVA71 through a novel domain on the VP3 capsid protein. The principle in the design, engineering, and refinement of the NRP1-based decoy receptor described in this study represents a general and well-suited antiviral strategy.
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exosomes facilitate transmission of Enterovirus A71 from human intestinal epithelial cells
The Journal of Infectious Diseases, 2020Co-Authors: Hsingi Huang, Jhaoyin Lin, Hsiaochu Chiang, Pennien Huang, Qingdong Lin, Shin-ru ShihAbstract:Background Enterovirus A71 (EV-A71) has been noted for its tendency to lead to neurological manifestations in young children and infants. Although the alimentary tract has been identified as the primary replication site of this virus, how EV-A71 replicates in the gut and is transmitted to other organs remains unclear. Methods By using differentiated C2BBe1 cells as a model, we observed that intestinal epithelial cells (IECs) were permissive to EV-A71 infection, and viral particles were released in a nonlytic manner. Results The coexistence of active caspase 3 and EV-A71 protein was observed in the infected undifferentiated C2BBe1 and RD cells but not in the infected differentiated C2BBe1 cells. Furthermore, EV-A71 infection caused differentiated C2BBe1 and intestinal organoids to secrete exosomes containing viral components and have the ability to establish active infection. Inhibition of the exosome pathway decreased EV-A71 replication and release in IECs and increased the survival rates of infected animals. Conclusions Our findings showed that EV-A71 is able to be actively replicated in enterocytes, and that the exosome pathway is involved in the nonlytic release of viral particles, which may be useful for developing antiviral strategies.
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antiviral activity of a llama derived single domain antibody against Enterovirus A71
Antimicrobial Agents and Chemotherapy, 2020Co-Authors: Pengnien Huang, Shin-ru Shih, Hsiang Ching Wang, Hui Chen Hung, Yunming Wang, S Y Tseng, Tengyuan Chang, Minyuan Chou, Yu Jen Chen, John T A HsuAbstract:In the past few decades, Enterovirus A71 (EVA71) has caused devastating outbreaks in the Asia-Pacific region, resulting in serious sequelae in infected young children. No preventive or therapeutic interventions are currently available for curing EVA71 infection, highlighting a great unmet medical need for this disease. Here, we showed that one novel single-domain antibody (sdAb), F1, isolated from an immunized llama, could alleviate EVA71 infection both in vitro and in vivo We also confirmed that the sdAb clone F1 recognizes EVA71 through a novel conformational epitope comprising the highly conserved region of VP3 capsid protein by using competitive-binding and overlapping-peptide enzyme-linked immunosorbent assays (ELISAs). Because of the virion's icosahedral structure, we reasoned that adjacent epitopes must be clustered within molecular ranges that may be simultaneously bound by an engineered antibody with multiple valency. Therefore, two single-domain binding modules (F1) were fused to generate an sdAb-in-tandem design so that the capture of viral antigens could be further increased by valency effects. We showed that the tetravalent construct F1×F1-hFc, containing two sdAb-in-tandem on a fragment crystallizable (Fc) scaffold, exhibits more potent neutralization activity against EVA71 than does the bivalent sdAb F1-hFc by at least 5.8-fold. We also demonstrated that, using a human scavenger receptor class B member 2 (hSCARB2) transgenic mouse model, a half dose of the F1×F1-hFc provided better protection against EVA71 infection than did the F1-hFc. Thus, our study furnishes important insights into multivalent sdAb engineering against viral infection and provides a novel strategic deployment approach for preparedness of emerging infectious diseases such as EVA71.
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Antivirals and vaccines for Enterovirus A71
Journal of Biomedical Science, 2019Co-Authors: Jing-yi Lin, Yu-an Kung, Shin-ru ShihAbstract:Enterovirus A71 (EV-A71) is an important emerging virus posing a threat to children under five years old. EV-A71 infection in infants or young children can cause hand-foot-and-mouth disease, herpangina, or severe neurological complications. However, there are still no effective antivirals for treatment of these infections. In this review, we summarize the antiviral compounds developed to date based on various targets of the EV-A71 life cycle. Moreover, development of a vaccine would be the most effective approach to prevent EV-A71 infection. Therefore, we also summarize the development and clinical progress of various candidate EV-A71 vaccines, including inactivated whole virus, recombinant VP1 protein, synthetic peptides, viral-like particles, and live attenuated vaccines.
Hsiang Ching Wang - One of the best experts on this subject based on the ideXlab platform.
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effect of a neuropilin 1 derived virus receptor trap on Enterovirus A71 infection in vitro
Antimicrobial Agents and Chemotherapy, 2020Co-Authors: Hsiang Ching Wang, Pengnien Huang, Hui Chen Hung, Sung Nien Tseng, Chia Chia Chang, Ya Ru Tsai, Yunming Wang, Shin-ru ShihAbstract:We discovered that neuropilin 1 (NRP1) is a new receptor candidate to mediate Enterovirus A71 (EVA71) into cells. In the engineered form as a decoy receptor, NRP1 was able to recognize and neutralize EVA71 but not Enterovirus D68 or coxsackievirus B3 (CVB3). NRP1 recognizes EVA71 through a novel domain on the VP3 capsid protein. The principle in the design, engineering, and refinement of the NRP1-based decoy receptor described in this study represents a general and well-suited antiviral strategy.
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antiviral activity of a llama derived single domain antibody against Enterovirus A71
Antimicrobial Agents and Chemotherapy, 2020Co-Authors: Pengnien Huang, Shin-ru Shih, Hsiang Ching Wang, Hui Chen Hung, Yunming Wang, S Y Tseng, Tengyuan Chang, Minyuan Chou, Yu Jen Chen, John T A HsuAbstract:In the past few decades, Enterovirus A71 (EVA71) has caused devastating outbreaks in the Asia-Pacific region, resulting in serious sequelae in infected young children. No preventive or therapeutic interventions are currently available for curing EVA71 infection, highlighting a great unmet medical need for this disease. Here, we showed that one novel single-domain antibody (sdAb), F1, isolated from an immunized llama, could alleviate EVA71 infection both in vitro and in vivo We also confirmed that the sdAb clone F1 recognizes EVA71 through a novel conformational epitope comprising the highly conserved region of VP3 capsid protein by using competitive-binding and overlapping-peptide enzyme-linked immunosorbent assays (ELISAs). Because of the virion's icosahedral structure, we reasoned that adjacent epitopes must be clustered within molecular ranges that may be simultaneously bound by an engineered antibody with multiple valency. Therefore, two single-domain binding modules (F1) were fused to generate an sdAb-in-tandem design so that the capture of viral antigens could be further increased by valency effects. We showed that the tetravalent construct F1×F1-hFc, containing two sdAb-in-tandem on a fragment crystallizable (Fc) scaffold, exhibits more potent neutralization activity against EVA71 than does the bivalent sdAb F1-hFc by at least 5.8-fold. We also demonstrated that, using a human scavenger receptor class B member 2 (hSCARB2) transgenic mouse model, a half dose of the F1×F1-hFc provided better protection against EVA71 infection than did the F1-hFc. Thus, our study furnishes important insights into multivalent sdAb engineering against viral infection and provides a novel strategic deployment approach for preparedness of emerging infectious diseases such as EVA71.
Sabine Diedrich - One of the best experts on this subject based on the ideXlab platform.
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increased detection of Enterovirus A71 infections germany 2019
Eurosurveillance, 2019Co-Authors: Sindy Bottcher, Sabine Diedrich, Kathrin KeerenAbstract:We report on the increased circulation of Enterovirus A71 in Germany in 2019. Strains were mainly identified in hospitalised patients with suspected aseptic meningitis/encephalitis. Molecular analysis showed co-circulation of EV-A71 sub-genogroups C1 and C4, a signal for physicians and public health authorities to include/intensify EV diagnostic in patients showing signs of aseptic meningitis, encephalitis or acute flaccid paralysis/myelitis.
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a severe pediatric infection with a novel Enterovirus A71 strain thuringia germany
Journal of Clinical Virology, 2016Co-Authors: Matthias Karrasch, Sindy Bottcher, Elisabeth Fischer, Martin Scholten, Andreas Sauerbrei, Andreas Henke, Diane M Renz, H J Mentzel, Klas Boer, Sabine DiedrichAbstract:Infection by Enterovirus A71 (EV-A71) is an important cause of hand, foot, and mouth disease (HFMD). Outbreaks including severe cases with neurological and cardiopulmonary complications have been reported particularly from Southeast Asia. In Europe, the epidemiology of EV-A71 is not well understood. In summer 2015, a two-year-old girl from Thuringia, Germany, presented with rhombencephalitis/brainstem encephalitis associated with severe neurological and cardiopulmonary complications. EV-A71 was detected in stool and almost the entire viral genome was amplified and sequenced. While the capsid protein VP1-encoding region belongs to the EV-A71 subgenogroup C1, the 3D polymerase encoding region represents a unique lineage. Thus, the data suggest that the Thuringian EV-A71 sequence likely represents a recombinant. The case underlines the importance of intensified EV-A71 surveillance in Germany and Europe including analysis of full-genome data.
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recombinant Enterovirus A71 subgenogroup c1 strains germany 2015
Emerging Infectious Diseases, 2016Co-Authors: Sindy Bottcher, Patrick Obermeier, Katrin Neubauer, Sabine DiedrichAbstract:To the Editor: Enterovirus A71 (EV-A71) strains circulate worldwide, and numerous outbreaks have been reported from Asia, Australia, Europe, and America (1). Symptomatic infections range from mild febrile illness or characteristic diseases such as hand, foot and mouth disease to severe neurologic disorders such as meningitis/encephalitis and acute flaccid paralysis. EV-A71 infections are usually asymptomatic and self-limiting but can also result in life-threatening complications such as pulmonary edema and cause death, predominantly in children <5 years of age. On the basis of viral protein 1 (VP1) sequences, 3 genogroups (A, B, C), including different subgenogroups (B0–B5, C1–C5), have been defined (2,3). Additional genogroups (D, E, F, G) have been proposed (4,5). In Europe, C1 and C2 strains have circulated predominantly within the past 2 decades, and recent introduction of C4 strains has been reported (6,7). Within subgenogroup C1, a lineage is replaced by the subsequent lineage over time (8).
Lunbiao Cui - One of the best experts on this subject based on the ideXlab platform.
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microrna 195 suppresses Enterovirus A71 induced pyroptosis in human neuroblastoma cells through targeting nlrx1
Virus Research, 2021Co-Authors: Xiaojuan Zhu, Ying Chi, Fengcai Zhu, Yongjun Jiao, Lunbiao CuiAbstract:Enterovirus A71 (EV-A71) emerged as a leading cause of virus derived infant encephalitis in most Asian countries. Some recent studies point out the critical role of microRNA (miRNA) in the regulation of pyroptosis. However, the role of miRNAs in the regulation of EV-A71 infection-induced pyroptosis was not previously explored. In this study, we utilized microRNA array and real-time PCR to verify that miR-195 significantly down-regulate in EV-A71-infected SH-SY5Y human neuroblastoma cells. An inverse correlation of NLRX1 with miR-195 expression in EV-A71-infected SH-SY5Y cells was found. Target prediction of miR-195 showed that NLRX1 could directly interact with miR-195. Results from luciferase reporter assays, qRT-PCR and western blotting demonstrated the negative regulation between miR-195 and NLRX1. Silencing NLRX1 expression with small interfering RNAs (siRNAs-NLRX1) and over-expression of miR-195 also attenuate the EV-A71 associated pyroptosis. Our findings provided evidence showed that miR-195 can regulate EV-A71 infection-induced pyroptosis, by directly targeting NLRX1.
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pyroptosis induced by Enterovirus A71 infection in cultured human neuroblastoma cells
Virology, 2018Co-Authors: Xiaojuan Zhu, Ying Chi, Minghao Zhou, Fengcai Zhu, Lunbiao CuiAbstract:Abstract Enterovirus A71 (EV-A71) infection can cause hand, foot and mouth disease (HFMD), and even fatal meningoencephalitis. Unfortunately, there is currently no effective treatment for EV-A71 infection due to the lack of understanding of the mechanism of neurological diseases. In this study, we employed SH-SY5Y human neuroblastoma cells to explore the roles of caspase-1 in neuropathogenesis. The expression and activity of caspase-1 were analyzed. The potential immuneconsequences mediated by caspase-1 including cell death, lysis, DNA degradation, and secretion of pro-inflammatory were also examined. We found the gene expression levels of caspase-1, IL-1β, IL-18 and active caspase-1 were markedly increased in the SH-SY5Y cells at 48 h post EV-A71 infection. The cell death, lysis, and DNA degradation were also increased during infection, which could be significantly alleviated by caspase-1 inhibition. These observations provided additional experimental evidence supporting caspase-1-mediated pyroptosis as a novel pathway of inflammatory programmed cell death.