D68 Infection Research Articles

A self-amplifying RNA vaccine prevents enterovirus D68 infection and disease in preclinical models.

The recent emergence and rapid response to severe acute respiratory syndrome coronavirus 2 was enabled by prototype pathogen and vaccine platform approaches, driven by the preemptive application of RNA vaccine technology to the related Middle East respiratory syndrome coronavirus. Recently, the National Institutes of Allergy and Infectious Diseases identified nine virus families of concern, eight enveloped virus families and one nonenveloped virus family, for which vaccine generation is a priority. Although RNA vaccines have been described for a variety of enveloped viruses, a roadmap for their use against nonenveloped viruses is lacking. Enterovirus D68 was recently designated a prototype pathogen within the family Picornaviridae of nonenveloped viruses because of its rapid evolution and respiratory route of transmission, coupled with a lack of diverse anti-enterovirus vaccine approaches in development. Here, we describe a proof-of-concept approach using a clinical stage RNA vaccine platform that induced robust enterovirus D68-neutralizing antibody responses in mice and nonhuman primates and prevented upper and lower respiratory tract infections and neurological disease in mice. In addition, we used our platform to rapidly characterize the antigenic diversity within the six genotypes of enterovirus D68, providing the necessary data to inform multivalent vaccine compositions that can elicit optimal breadth of neutralizing responses. These results demonstrate that RNA vaccines can be used as tools in our pandemic-preparedness toolbox for nonenveloped viruses.

Real-Time Enterovirus D68 Outbreak Detection through Hospital Surveillance of Severe Acute Respiratory Infection, Senegal, 2023.

In December 2023, we observed through hospital-based surveillance a severe outbreak of enterovirus D68 infection in pediatric inpatients in Dakar, Senegal. Molecular characterization revealed that subclade B3, the dominant lineage in outbreaks worldwide, was responsible for the outbreak. Enhanced surveillance in inpatient settings, including among patients with neurologic illnesses, is needed.

Transcription factor EB (TFEB) interaction with RagC is disrupted during enterovirus D68 infection.

Enteroviruses are among the most significant causes of human disease. Some enteroviruses are responsible for severe paralytic diseases such as poliomyelitis or acute flaccid myelitis. The latter disease is associated with multiple non-polio enterovirus species, including enterovirus D68 (EV-D68), enterovirus 71, and coxsackievirus B3 (CVB3). Here, we demonstrate that EV-D68 interacts with a host transcription factor, transcription factor EB (TFEB), to promote viral RNA(vRNA) replication and regulate the egress of virions from cells. TFEB was previously implicated in the viral egress of CVB3, and the viral protease 3C cleaves TFEB during infection. Here, we show that EV-D68 3C protease also cleaves TFEB after the peak of vRNA replication. This cleavage disrupts TFEB interaction with the host protein RagC, which changes the localization and regulation of TFEB. TFEB lacking a RagC-binding domain inhibits autophagic flux and promotes virus egress. These mechanistic insights highlight how common host factors affect closely related, medically important viruses differently.

Pharmacological activation of TLR7 exerts inhibition on the replication of EV-D68 in respiratory cells.

The globally reemerging respiratory pathogen enterovirus D68 (EV-D68) is implicated in outbreaks of severe respiratory illness and associated with acute flaccid myelitis. However, there remains a lack of effective treatments for EV-D68 infection. In this work, we found that the host Toll-like receptor 7 (TLR7) proteins, which function as powerful innate immune sensors, were selectively elevated in expression in response to EV-D68 infection. Subsequently, we investigated the impact of Vesatolimod (GS-9620), a Toll-like receptor 7 agonist, on EV-D68 replication. Our findings revealed that EV-D68 infection resulted in increased mRNA levels of TLR7. Treatment with Vesatolimod significantly inhibited EV-D68 replication [half maximal effective concentration (EC50) = 0.1427 µM] without inducing significant cytotoxicity at virucidal concentrations. Although Vesatolimod exhibited limited impact on EV-D68 attachment, it suppressed RNA replication and viral protein synthesis after virus entry. Vesatolimod broadly inhibited the replication of circulating isolated strains of EV-D68. Furthermore, our findings demonstrated that treatment with Vesatolimod conferred resistance to both respiratory and neural cells against EV-D68 infection. Overall, these results present a promising strategy for drug development by pharmacologically activating TLR7 to initiate an antiviral state in EV-D68-infected cells selectively.IMPORTANCEConventional strategies for antiviral drug development primarily focus on directly targeting viral proteases or key components, as well as host proteins involved in viral replication. In this study, based on our intriguing discovery that enterovirus D68 (EV-D68) infection specifically upregulates the expression of immune sensor Toll-like receptor 7 (TLR7) protein, which is either absent or expressed at low levels in respiratory cells, we propose a potential antiviral approach utilizing TLR7 agonists to activate EV-D68-infected cells into an anti-viral defense state. Notably, our findings demonstrate that pharmacological activation of TLR7 effectively suppresses EV-D68 replication in respiratory tract cells through a TLR7/MyD88-dependent mechanism. This study not only presents a promising drug candidate and target against EV-D68 dissemination but also highlights the potential to exploit unique alterations in cellular innate immune responses induced by viral infections, selectively inducing a defensive state in infected cells while safeguarding uninfected normal cells from potential adverse effects associated with therapeutic interventions.

Epidemiological and Clinical Insights into the Enterovirus D68 Upsurge in Europe 2021-2022 and Emergence of Novel B3-Derived Lineages, ENPEN Multicentre Study.

Enterovirus D68 (EV-D68) infections are associated with severe respiratory disease and acute flaccid myelitis (AFM). The European Non-Polio Enterovirus Network (ENPEN) aimed to investigate the epidemiological and genetic characteristics of EV-D68 infections and its clinical impact during the fall-winter season of 2021-2022. From 19 European countries, 58 institutes reported 10 481 (6.8%) EV-positive samples of which 1004 (9.6%) were identified as EV-D68 (including 852 respiratory samples). Clinical data were reported for 969 cases; 78.9% of infections were reported in children (0-5 years); and 37.9% of cases were hospitalized. Acute respiratory distress was commonly noted (93.1%) followed by fever (49.4%). Neurological problems were observed in 6.4% of cases including 6 diagnosed with AFM. Phylodynamic/Nextstrain and phylogenetic analyses based on 694 sequences showed the emergence of 2 novel B3-derived lineages, with no regional clustering. In conclusion, we describe a large-scale European EV-D68 upsurge with severe clinical impact and the emergence of B3-derived lineages.

Caspase-8 activation regulates enterovirus D68 infection-induced inflammatory response and cell death

Enterovirus D68 (EV-D68) infection causes severe acute respiratory infection and severe neurological complications, such as acute flaccid myelitis (AFM), in children. However, although EV-D68 has pandemic potential, no effective drugs or vaccines are currently clinically available. Furthermore, EV-D68 infection-induced inflammatory response and cell death are not fully understood. In this study, we demonstrated that several inflammatory cytokines were upregulated in a multiplicity of infection (MOI) dependent manner in EV-D68-infected human rhabdomyosarcoma (RD) cells. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) confirmed that tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), C-C motif chemokine ligand-5 (CCL-5), and CXC motif chemokine ligand-5 (CXCL-5) mRNA levels were highly upregulated after EV-D68 infection. IL-1β processing and maturation mediated by caspase-8 was inhibited by the caspase-8 inhibitor Z-IETD-FMK. EV-D68 infection activates caspase-8 to mediate IL-1β maturation and secretion. Additionally, EV-D68 activated cell death-related proteins such as caspase-3, poly (ADP-ribose) polymerase 1 (PARP-1), phosphorylation of Mixed Lineage Kinase domain-like protein (pMLKL), and gasdermin E (GSDME). Thus, EV-D68 infection activates caspase-8, which triggers the necroptosis and apoptosis pathways. Overall, our data suggest that caspase-8 activation is associated with the inflammatory response and cell death in EV-D68-infected RD cells. This mechanism represents a novel target for the treatment of EV-D68 infection by inhibiting caspase-8 activation.

Human B cells and dendritic cells are susceptible and permissive to enterovirus D68 infection.

Enterovirus D68 (EV-D68) is predominantly associated with mild respiratory infections, but can also cause severe respiratory disease and extra-respiratory complications, including acute flaccid myelitis. Systemic dissemination of EV-D68 is crucial for the development of extra-respiratory diseases, but it is currently unclear how EV-D68 spreads systemically (viremia). We hypothesize that immune cells contribute to the systemic dissemination of EV-D68, as this is a mechanism commonly used by other enteroviruses. Therefore, we investigated the susceptibility and permissiveness of human primary immune cells for different EV-D68 isolates. In human peripheral blood mononuclear cells inoculated with EV-D68, only B cells were susceptible but virus replication was limited. However, in B cell-rich cultures, such as Epstein-Barr virus-transformed B-lymphoblastoid cell line (BLCL) and primary lentivirus-transduced B cells, which better represent lymphoid B cells, were productively infected. Subsequently, we showed that dendritic cells (DCs), particularly immature DCs, are susceptible and permissive for EV-D68 infection and that they can spread EV-D68 to autologous BLCL. Altogether, our findings suggest that immune cells, especially B cells and DCs, could play an important role in the pathogenesis of EV-D68 infection. Infection of these cells may contribute to systemic dissemination of EV-D68, which is an essential step toward the development of extra-respiratory complications.IMPORTANCEEnterovirus D68 (EV-D68) is an emerging respiratory virus that has caused outbreaks worldwide since 2014. EV-D68 infects primarily respiratory epithelial cells resulting in mild respiratory diseases. However, EV-D68 infection is also associated with extra-respiratory complications, including polio-like paralysis. It is unclear how EV-D68 spreads systemically and infects other organs. We hypothesized that immune cells could play a role in the extra-respiratory spread of EV-D68. We showed that EV-D68 can infect and replicate in specific immune cells, that is, B cells and dendritic cells (DCs), and that virus could be transferred from DCs to B cells. Our data reveal a potential role of immune cells in the pathogenesis of EV-D68 infection. Intervention strategies that prevent EV-D68 infection of immune cells will therefore potentially prevent systemic spread of virus and thereby severe extra-respiratory complications.

Predicting the impact of COVID-19 non-pharmaceutical intervention on short- and medium-term dynamics of enterovirus D68 in the US

Recent outbreaks of enterovirus D68 (EV-D68) infections, and their causal linkage with acute flaccid myelitis (AFM), continue to pose a serious public health concern. During 2020 and 2021, the dynamics of EV-D68 and other pathogens have been significantly perturbed by non-pharmaceutical interventions against COVID-19; this perturbation presents a powerful natural experiment for exploring the dynamics of these endemic infections. In this study, we analyzed publicly available data on EV-D68 infections, originally collected through the New Vaccine Surveillance Network, to predict their short- and long-term dynamics following the COVID-19 interventions. Although long-term predictions are sensitive to our assumptions about underlying dynamics and changes in contact rates during the NPI periods, the likelihood of a large outbreak in 2023 appears to be low. Comprehensive surveillance data are needed to accurately characterize future dynamics of EV-D68. The limited incidence of AFM cases in 2022, despite large EV-D68 outbreaks, poses further questions for the timing of the next AFM outbreaks.

Enterovirus D68 infection upregulates SOCS3 expression to inhibit JAK-STAT3 signaling and antagonize the innate interferon response of the host

Enterovirus D68 (EV-D68) can cause respiratory diseases and acute flaccid paralysis, posing a great threat to public health. Interferons are cytokines secreted by host cells that have broad-spectrum antiviral effects, inducing the expression of hundreds of interferon-stimulated genes (ISGs). EV-D68 activates ISG expression early in infection, but at a later stage, the virus suppresses ISG expression, a strategy evolved by EV-D68 to antagonize interferons. Here, we explore a host protein, suppressor of cytokine signaling 3 (SOCS3), is upregulated during EV-D68 infection and antagonizes the antiviral effects of type I interferon. We subsequently demonstrate that the structural protein of EV-D68 upregulated the expression of RFX7, a transcriptional regulator of SOCS3, leading to the upregulation of SOCS3 expression. Further exploration revealed that SOCS3 plays its role by inhibiting the phosphorylation of signal transducer and activator of transcription 3 (STAT3). The expression of SOCS3 inhibited the expression of ISG, thereby inhibiting the antiviral effect of type I interferon and promoting EV-D68 transcription, protein production, and viral titer. Notably, a truncated SOCS3, generated by deleting the kinase inhibitory region (KIR) domain, failed to promote replication and translation of EV-D68. Based on the above studies, we designed a short peptide named SOCS3 inhibitor, which can specifically bind and inhibit the KIR structural domain of SOCS3, significantly reducing the RNA and protein levels of EV-D68. In summary, our results demonstrated a novel mechanism by which EV-D68 inhibits ISG transcription and antagonizes the antiviral responses of host type I interferon.

Serostatus of echovirus 11, coxsackievirus B3 and enterovirus D68 in cord blood: The implication of severe newborn enterovirus infection.

Maternal transplacental antibody is an important origins of passive immunity against neonatal enterovirus infection. Echovirus 11 (E11) and coxsackievirus B3 (CVB3) are important types causing neonatal infections. There were few investigations of enterovirus D68 (EVD68) infection in neonates. We aimed to investigate the serostatus of cord blood for these three enteroviruses and evaluate the factors associated with seropositivity. We enrolled 222 parturient (gestational age 34-42 weeks) women aged 20-46 years old between January and October 2021. All participants underwent questionnaire investigation and we collected the cord blood to measure the neutralization antibodies against E11, CVB3 and EVD68. The cord blood seropositive rates were 18% (41/222), 60% (134/232) and 95% (211/222) for E11, CVB3 and EVD68, respectively (p<0.001). Geometric mean titers were 3.3 (95% CI 2.9-3.8) for E11, 15.9 (95% CI 12.5-20.3) for CVB3 and 109.9 (95% CI 92.4-131.6) for EVD68. Younger parturient age (33.8±3.6 versus 35.2±4.4, p=0.04) was related to E11 seropositivity. Neonatal sex, gestational age and birth body weight were not significantly different between the seropositive group and the seronegative group. Cord blood seropositive rate and geometric mean titer of E11 were very low, so a large proportion of newborns are susceptible to E11. The circulation of E11 was low after 2019 in Taiwan. A large cohort of immune naïve newborns existed currently due to lack of protective maternal antibodies. It is imminent to monitor the epidemiology of neonates with enterovirus infections and strengthen the relevant preventive policies.

Clinical and molecular epidemiologic features of enterovirus D68 infection in children with acute lower respiratory tract infection in China.

Acute flaccid paralysis (AFP) associated with enterovirus D68 (EV-D68) infection has attracted much attention since an outbreak in the USA in 2014. Notably, EV-D68 was detected in a child with AFP for the first time in China in 2018. In a multicentre study from May 2017 to December 2019, we monitored EV-D68 infections in hospitalized children with acute lower respiratory tract infection (ALRTI) in China. Out of 3,071 samples collected from patients with ALRTI, ten were positive for EV-D68. All patients presented with mild diseases withno neurological symptoms or signs. Phylogenetic analysis based on the VP1 gene showed that all EV-D68 sequences obtained in this study belonged to subclade B3 and were close to sequences of EV-D68 strains obtained from patients with AFP in the USA. Four EV-D68 strains were isolated, and their complete genome sequences were determined. These sequences did not show any evidence of recombination events. To assess their neurotropism, the isolates were used to infect the "neuronal-like" cell line SH-SY5Y, and resulted in a cytopathic effect. We further analysed the structure and sites that may beassociated with neurovirulence, includingthe stem-loop structure in the untranslated region (3'UTR) and identified amino acid substitutions (M291T, V341A, T860N, D927N, S1108G, and R2005K) in the coding region and specific nucleotides (127T, 262C, and 339T) in the 5' UTR. In conclusion, EV-D68 infection was detected in a small number of children with ALRTI in China from 2017 to 2019. Disease symptoms in these children were relatively mild with no neurological complications, and all EV-D68 sequences belonged to subclade B3.

Enterovirus D68 Outbreak in Children, Finland, August–September 2022

We observed an intense enterovirus D68 outbreak in children in southwest Finland in August-September 2022. We confirmed enterovirus D68 infection in 56 children hospitalized for respiratory illnesses and in 1 child with encephalitis but were not able to test all suspected patients. Continuing surveillance for enterovirus D68 is needed.

Enterovirus D68 Infection Induces TDP-43 Cleavage, Aggregation, and Neurotoxicity.

Enterovirus D68 (EV-D68), which causes severe respiratory diseases and irreversible central nervous system damage, has become a serious public health problem worldwide. However, the mechanisms by which EV-D68 exerts neurotoxicity remain unclear. Thus, we aimed to analyze the effects of EV-D68 infection on the cleavage, subcellular translocation, and pathogenic aggregation of TAR DNA-binding protein 43 kDa (TDP-43) in respiratory or neural cells. The results showed that EV-D68-encoded proteases 2A and 3C induced TDP-43 translocation and cleavage, respectively. Specifically, 3C cleaved residue 327Q of TDP-43. The 3C-mediated cleaved TDP-43 fragments had substantially decreased protein solubility compared with the wild-type TDP-43. Hence, 3C activity promoted TDP-43 aggregation, which exerted cytotoxicity to diverse human cells, including glioblastoma T98G cells. The effects of commercially available antiviral drugs on 3C-mediated TDP-43 cleavage were screened, and the results revealed lopinavir as a potent inhibitor of EV-D68 3C protease. Overall, these results suggested TDP-43 as a conserved host target of EV-D68 3C. This study is the first to provide evidence on the involvement of TDP-43 dysregulation in EV-D68 pathogenesis. IMPORTANCE Over the past decade, the incidence of enterovirus D68 (EV-D68) infection has increased worldwide. EV-D68 infection can cause different respiratory symptoms and severe neurological complications, including acute flaccid myelitis. Thus, elucidating the mechanisms underlying EV-D68 toxicity is important to develop novel methods to prevent EV-D68 infection-associated diseases. This study shows that EV-D68 infection triggers the translocalization, cleavage, and aggregation of TDP-43, an intracellular protein closely related to degenerative neurological disorders. The viral protease 3C decreased TDP-43 solubility, thereby exerting cytotoxicity to host cells, including human glioblastoma cells. Thus, counteracting 3C activity is an effective strategy to relieve EV-D68-triggered cell death. Cytoplasmic aggregation of TDP-43 is a hallmark of degenerative diseases, contributing to neural cell damage and central nervous system (CNS) disorders. The findings of this study on EV-D68-induced TDP-43 formation extend our understanding of virus-mediated cytotoxicity and the potential risks of TDP-43 dysfunction-related cognitive impairment and neurological symptoms in infected patients.

Pterostilbene, an active constituent of blueberries, enhances innate immune activation and restricts enterovirus D68 infection.

Enterovirus D68 (EV-D68) is a globally re-emerging respiratory pathogen implicated in outbreaks of severe respiratory illnesses and associated with acute flaccid myelitis. However, effective vaccines or treatments for EV-D68 infections remain scarce. We demonstrated that the active constituent of blueberries, pterostilbene (Pte), and its major metabolite, pinostilbene (Pin), facilitated innate immune responses in EV-D68-infected human respiratory cells. Pte and Pin treatment clearly relieved EV-D68-triggered cytopathic effects. Importantly, both Pte and Pin disrupted viral RNA replication (EC50 rank from 1.336 to 4.997 µM) and infectious virion production in a dose-dependent manner, without cytotoxicity at virucidal concentrations. Pte- or Pin-treated respiratory cells did not show any influences on EV-D68 entry but showed substantially decreased viral RNA replication and protein synthesis. Finally, we showed that Pte and Pin broadly suppressed the replication capacity of circulating EV-D68 strains isolated from recent pandemics. In summary, our results suggest that Pte and its derivative, Pin, enhance host immune recognition of EV-D68 and suppress EV-D68 replication, which represents a promising strategy for antiviral drug development.

Seroprevalence of Enterovirus D68 Infection among Humans: A Systematic Review and Meta-Analysis

Introduction: Human enterovirus D68 (EV-D68), which belongs to enteroviruses of the small RNA family, is a type of enterovirus that can cause acute respiratory tract infection and central nervous system diseases. This study systematically analysed and summarized EV-D68 antibody studies in databases and identified the seropositivity rates of different regions, ages, and sexes. Methods: Meta-analysis was performed using STATA 16.0 software. I² and Q tests were used to analyse the heterogeneity of the included studies. Meta-regression analysis was performed for different groups, and Egger’s linear regression analysis was used to evaluate publication bias. Results: The results of multiple studies indicated that the serological prevalence range of EV-D68 antibody was 17.78–96.69%. The results of the meta-analysis showed that the seropositivity rate of EV-D68 antibody was 76% (95% confidence interval [CI]: 67–84%), among which that of the Chinese population was 74% (95% CI: 61–86%) and that of other countries was 79% (95% CI: 65–91%). At the same time, a subgroup analysis was conducted. The seroprevalence of EV-D68 antibody was related to age but not sex or region. Conclusion: The seropositivity rate was lower in the below 5-year age group; however, it gradually increased with age. The results of this study showed that EV-D68 infection was widespread in the population, and the current clinical infection situation could not reflect the actual epidemic situation of the virus, among which children under 5 years old were vulnerable to infection, which should be given greater attention for epidemic prevention and control.

Increase in Acute Respiratory Illnesses Among Children and Adolescents Associated with Rhinoviruses and Enteroviruses, Including Enterovirus D68 - United States, July-September 2022.

Increases in severe respiratory illness and acute flaccid myelitis (AFM) among children and adolescents resulting from enterovirus D68 (EV-D68) infections occurred biennially in the United States during 2014, 2016, and 2018, primarily in late summer and fall. Although EV-D68 annual trends are not fully understood, EV-D68 levels were lower than expected in 2020, potentially because of implementation of COVID-19 mitigation measures (e.g., wearing face masks, enhanced hand hygiene, and physical distancing) (1). In August 2022, clinicians in several geographic areas notified CDC of an increase in hospitalizations of pediatric patients with severe respiratory illness and positive rhinovirus/enterovirus (RV/EV) test results.* Surveillance data were analyzed from multiple national data sources to characterize reported trends in acute respiratory illness (ARI), asthma/reactive airway disease (RAD) exacerbations, and the percentage of positive RV/EV and EV-D68 test results during 2022 compared with previous years. These data demonstrated an increase in emergency department (ED) visits by children and adolescents with ARI and asthma/RAD in late summer 2022. The percentage of positive RV/EV test results in national laboratory-based surveillance and the percentage of positive EV-D68 test results in pediatric sentinel surveillance also increased during this time. Previous increases in EV-D68 respiratory illness have led to substantial resource demands in some hospitals and have also coincided with increases in cases of AFM (2), a rare but serious neurologic disease affecting the spinal cord. Therefore, clinicians should consider AFM in patients with acute flaccid limb weakness, especially after respiratory illness or fever, and ensure prompt hospitalization and referral to specialty care for such cases. Clinicians should also test for poliovirus infection in patients suspected of having AFM because of the clinical similarity to acute flaccid paralysis caused by poliovirus. Ongoing surveillance for EV-D68 is critical to ensuring preparedness for possible future increases in ARI and AFM.

Enterovirus D68 Infection in Human Primary Airway and Brain Organoids: No Additional Role for Heparan Sulfate Binding for Neurotropism.

ABSTRACTEnterovirus D68 (EV-D68) is an RNA virus that can cause outbreaks of acute flaccid paralysis (AFP), a polio-like disease. Before 2010, EV-D68 was a rare pathogen associated with mild respiratory symptoms, but the recent EV-D68 related increase in severe respiratory illness and outbreaks of AFP is not yet understood. An explanation for the rise in severe disease is that it may be due to changes in the viral genome resulting in neurotropism. In this regard, in addition to sialic acid, binding to heparan sulfate proteoglycans (HSPGs) has been identified as a feature for viral entry of some EV-D68 strains in cell lines. Studies in human primary organotypic cultures that recapitulate human physiology will address the relevance of these HSPG-binding mutations for EV-D68 infection in vivo. Therefore, in this work, we studied the replication and neurotropism of previously determined sialic acid-dependent and HSPG-dependent strains using primary human airway epithelial (HAE) cultures and induced human pluripotent stem cell (iPSC)-derived brain organoids. All three strains (B2/2042, B2/947, and A1/1348) used in this study infected HAE cultures and human brain organoids (shown for the first time). Receptor-blocking experiments in both cultures confirm that B2/2042 infection is solely dependent on sialic acid, while B2/947 and A1/1348 (HSPG to a lesser extent) binds to sialic acid and HSPG for cell entry. Our data suggest that HSPG-binding can be used by EV-D68 for entry in human physiological models but offers no advantage for EV-D68 infection of brain cells.IMPORTANCE Recent outbreaks of enterovirus D68, a nonpolio enterovirus, is associated with a serious neurological condition in young children, acute flaccid myelitis (AFM). As there is no antiviral treatment or vaccine available for EV-D68 it is important to better understand how EV-D68 causes AFM and why only recent outbreaks are associated with AFM. We investigated if a change in receptor usage of EV-D68 increases the virulence of EV-D68 in the airway or the central nervous system and thus could explain the increase in AFM cases. We studied this using physiologically relevant human airway epithelium and cerebral organoid cultures that are physiologically relevant human models. Our data suggest that heparan sulfate proteoglycans can be used by EV-D68 as an additional entry receptor in human physiological models but offers no advantage for EV-D68 infection of brain cells, and our data show the potential of these 46 innovative models for virology.

Enterovirus D68 in Hospitalized Children, Barcelona, Spain, 2014-2021.

To determine molecular epidemiology and clinical features of enterovirus D68 (EV-D68) infections, we reviewed EV-D68–associated respiratory cases at a hospital in Barcelona, Spain, during 2014–2021. Respiratory samples were collected from hospitalized patients or outpatients with symptoms of acute respiratory tract infection or suggestive of enterovirus infection. Enterovirus detection was performed by real-time multiplex reverse transcription PCR and characterization by phylogenetic analysis of the partial viral protein 1 coding region sequences. From 184 patients with EV-D68 infection, circulating subclades were B3 (80%), D1 (17%), B2 (1%), and A (<1%); clade proportions shifted over time. EV-D68 was detected mostly in children (86%) and biennially (2016, 2018, 2021). In patients <16 years of age, the most common sign/symptom was lower respiratory tract infection, for which 11.8% required pediatric intensive care unit admission and 2.3% required invasive mechanical ventilation; neurologic complications developed in 1. The potential neurotropism indicates that enterovirus surveillance should be mandatory.

Global prevalence and case fatality rate of Enterovirus D68 infections, a systematic review and meta-analysis.

A substantial amount of epidemiological data has been reported on Enterovirus D68 (EV-D68) infections after the 2014 outbreak. Our goal was to map the case fatality rate (CFR) and prevalence of current and past EV-D68 infections. We conducted a systematic review (PROSPERO, CRD42021229255) with published articles on EV-68 infections in PubMed, Embase, Web of Science and Global Index Medicus up to January 2021. We determined prevalences using a model random effect. Of the 4,329 articles retrieved from the databases, 89 studies that met the inclusion criteria were from 39 different countries with apparently healthy individuals and patients with acute respiratory infections, acute flaccid myelitis and asthma-related diseases. The CFR estimate revealed occasional deaths (7/1353) related to EV-D68 infections in patients with severe acute respiratory infections. Analyses showed that the combined prevalence of current and past EV-D68 infections was 4% (95% CI = 3.1-5.0) and 66.3% (95% CI = 40.0-88.2), respectively. The highest prevalences were in hospital outbreaks, developed countries, children under 5, after 2014, and in patients with acute flaccid myelitis and asthma-related diseases. The present study shows sporadic deaths linked to severe respiratory EV-D68 infections. The study also highlights a low prevalence of current EV-D68 infections as opposed to the existence of EV-D68 antibodies in almost all participants of the included studies. These findings therefore highlight the need to implement and/or strengthen continuous surveillance of EV-D68 infections in hospitals and in the community for the anticipation of the response to future epidemics.

Re-emergence of enterovirus D68 in Europe after easing the COVID-19 lockdown, September 2021.

We report a rapid increase in enterovirus D68 (EV-D68) infections, with 139 cases reported from eight European countries between 31 July and 14 October 2021. This upsurge is in line with the seasonality of EV-D68 and was presumably stimulated by the widespread reopening after COVID-19 lockdown. Most cases were identified in September, but more are to be expected in the coming months. Reinforcement of clinical awareness, diagnostic capacities and surveillance of EV-D68 is urgently needed in Europe.