Coxsackievirus B3 Research Articles

Stabilin-1 mediates beneficial monocyte recruitment and tolerogenic macrophage programming during CVB3-induced viral myocarditis

Pathological innate and adaptive immune response upon viral infection may lead to cardiac injury and dysfunction. Stabilin-1 is a scavenger receptor that regulates several aspects of the innate immunity. Whether stabilin-1 affects the inflammatory response during viral myocarditis (VM) is entirely unknown. Here, we assess the role of stabilin-1 in the pathogenesis of VM and its suitability as a therapeutic target. Genetic loss of stabilin-1 increased mortality and cardiac necrosis in a mouse model of human Coxsackievirus B3 (CVB3)-induced myocarditis. Absence of stabilin-1 significantly reduced monocyte recruitment and strongly reduced the number of alternatively activated anti-inflammatory macrophages in the heart, enhancing a pro-inflammatory cardiac niche with a detrimental T lymphocyte response during VM. Yeast two-hybrid screening, confirmed by affinity chromatography, identified fibronectin as a stabilin-1 interacting partner. Absence of stabilin-1 specifically decreased monocyte adhesion on extracellular fibronectin in vitro. Loss of Type III repeats Extra Domain A (EDA) of fibronectin during VM also increased the mortality and cardiac necrosis as in stabilin-1 knockout mice, with reduced monocytic cardiac recruitment and increased T lymphocyte response.Collectively, stabilin-1 has an immune-suppressive role of limiting myocardial damage during VM, regulating anti-inflammatory monocyte-recruitment to the site of inflammation.

Persistent coxsackievirus B1 infection triggers extensive changes in the transcriptome of human pancreatic ductal cells

SummaryEnteroviruses, particularly the group B coxsackieviruses (CVBs), have been associated with the development of type 1 diabetes. Several CVB serotypes establish chronic infections in human cells in vivo and in vitro. However, the mechanisms leading to enterovirus persistency and, possibly, beta cell autoimmunity are not fully understood. We established a carrier-state-type persistent infection model in human pancreatic cell line PANC-1 using two distinct CVB1 strains and profiled the infection-induced changes in cellular transcriptome. In the current study, we observed clear changes in the gene expression of factors associated with the pancreatic microenvironment, the secretory pathway, and lysosomal biogenesis during persistent CVB1 infections. Moreover, we found that the antiviral response pathways were activated differently by the two CVB1 strains. Overall, our study reveals extensive transcriptional responses in persistently CVB1-infected pancreatic cells with strong opposite but also common changes between the two strains.

Primary Site of Coxsackievirus B Replication in the Small Intestines: No Proof of Peyer's Patches Involvement.

Background: Enterovirus (EV) infections are associated with a broad range of diseases. Since the first experimental infection of primates with poliovirus (PV), tonsils and the Peyer’s patches (PPs) have been believed to be the primary replication sites of EVs. Our aim was to localize different viral markers in the small intestines (SI) of coxsackievirus B (CVB) orally and intraperitoneally (i.p.) infected mice. Methods: Transverse sections of SIs of both infected and control male outbred mice were collected at different intervals post-infection (p.i) and analyzed for presence of interferon-alpha (IFN-α) and viral protein VP1 by immunohistochemistry and in situ hybridization (ISH). Fluorescent marker, eGFP, was identified in cryosections of mice infected with eGFP-CVB3. Results: In the infected SIs, we observed enlarged germinating centers (GCs) in the PPs; IFN-α was detected in the PPs and mucosal layer of the SIs. However, VP1, viral RNA and the eGFP were absent in the GCs of PPs at all stages of infection irrespective of the virus strains used. Conclusions: Virus was present in the epithelial cells but not in GCs of the PPs of the murine SIs. Our results do not support the hypothesis of EV replication in the PP especially in the GCs.

Microwave-Assisted Synthesis of Lactose Acetates with Antimicrobial, Cytotoxic, and Antiviral Properties

The task of this study was to perform the "green" synthesis of lactose octaacetate through microwave irradiation and to establish their biological activities. Lactose ester was prepared after microwave-assisted esterification of lactose with acetic anhydride (yield 85-90 %). Lactose octaacetate was characterized by a high degree of acetylation (DS 3.2-3.7). The lactose esters' structure was elucidated by infrared spectroscopy and nuclear magnetic resonance spectroscopy. Lactose octaacetate showed better antifungal activities than antibacterial activities. It possessed slight to moderate antifungal activities against Aspergillus niger ATCC 1015, Penicillium sp., Rhizopus sp., Fusarium moniliforme ATCC 38932. Lactose acetates demonstrated low cytotoxicity against three cell lines: Madin-Darby bovine kidney (MDBK) cells, human epithelial type 2 (HEp-2), and Madin-Darby canine kidney (MDCK) cells. This is the first report for antiviral activity of lactose acetates against herpes simplex virus type 1 (HSV-1), influenza virus A/Panama/2007/99/H3N2 - (IAV/H3N2), PV-1 and Coxsackievirus B1. It was found that this compound showed activity with SI = 2.4 only against PV-1, but against HSV-1, IAV/H3N2, and Coxsackievirus B1 was inactive. The current study demonstrated the applications of lactose acetates as antimicrobial and antiviral substances in food, pharmaceutical, agricultural and cosmetic preparations.

Identification of Immunogenic Epitopes That Permit the Detection of Antigen-Specific T Cell Reponses to Coxsackievirus B3

Background and Hypothesis:Coxsackievirus B3 (CVB3) is a non-enveloped RNA virus from the Picornaviridae family and is a primary cause of viral myocarditis in the United States. Approximately 5% of all symptomatic CVB3 infections are fatal. Therefore, there is a need to identify the mechanism(s) that regulate a protective immune response to CVB3. However, viral epitopes that stimulate T cell responses to CVB3 remain poorly characterized. To this end, we used a mouse model of CVB3 infection to identify the viral immunogenic CD8 T cell epitopes. We hypothesized that isolated antigen-experienced CD8 T cells from infected mice would be stimulated in the presence of predicted viral epitopes, confirming CVB3-specific T cells. 
 Experimental Design: To identify novel CD8 T cell epitopes, predicted 9-mer MHC binding peptides from the CVB3-Nancy polyprotein were identified using the Immune Epitope Database (IEDB) analysis resource consensus tool. The top ten predicted peptides were synthesized for our assays. Splenocytes from CVB3-infected male and female IFNAR -/- mice were stimulated with each peptide in the presence of brefeldin A for 6 hours at 37˚C. Following stimulation, cells were surfaced stained with antibodies specific for antigen-experienced CD8 T cells. Next, we performed intracellular staining for IFN-gamma. Cells were analyzed using flow cytometry. Candidate epitopes were identified as having results ≥2 standard deviations over the control. 
 Results: Thus far, our analysis has revealed responses to three novel CD8 T cell epitopes within the peptide library, including the viral epitopes within VP1 protein and the RNA-dependent RNA polymerase.
 Conclusion and Potential ImpactOverall, these data provide an advancement in CVB3 immunology. Further, these data generate new tools like MHC-tetramers to track endogenous T cell responses to CVB3 infection.

Immunohistochemical detection of enteroviruses in pancreatic tissues of patients with type 1 diabetes using a polyclonal antibody against 2A protease of Coxsackievirus.

ABSTRACTAims/IntroductionThe need for antiserum for immunohistochemical (IHC) detection of enterovirus (EV) in formaldehyde‐fixed and paraffin‐embedded samples is increasing. The gold standard monoclonal antibody (clone 5D8/1) against EV‐envelope protein (VP1) was proven to cross‐react with other proteins. Another candidate marker of EV proteins is 2A protease (2Apro), which is encoded by the EV gene and translated by the host cells during EV replication, and participates processing proproteins to viral capsid proteins.Materials and MethodsWe raised polyclonal antiserum by immunizing a rabbit with an 18‐mer peptide of Coxsackievirus B1 (CVB1)‐2Apro, and examined the specificity and sensitivity for EV on formaldehyde‐fixed and paraffin‐embedded tissue samples.ResultsEnzyme‐linked immunosorbent assay study showed a high titer of antibody for 18‐mer peptide of CVB1‐2Apro, cross‐reacting with CVB3‐2Apro peptide. IHC showed that antiserum against 2Apro reacted with CVB1‐infected and VP1‐positive Vero cells. Confocal laser scanning microscopy showed that antigen stained by the 2Apro antibody located in the same cell with VP1 stained by 5D8/1. IHC using 2Apro antiserum showed dense staining in the islets of EV‐associated fulminant type 1 diabetes pancreas and that located in the same cell stained positive for VP1 (5D8/1). Specificity of 2Apro antiserum by IHC staining was confirmed by negative 2Apro in 14 VP1‐negative non‐diabetes control pancreases.ConclusionsOur study provides a new polyclonal antiserum against CVB1‐2Apro, which might be useful for IHC of EV‐infected human tissues stored as archive of formaldehyde‐fixed and paraffin‐embedded tissue samples.

134. KRP-A218, an Orally Active and Selective PI4KB Inhibitor with Broad-Spectrum Anti-Rhinovirus Activity, Has Potent Therapeutic Antiviral Activity In vivo

BackgroundRhinovirus (RV) is a major respiratory virus that poses a threat to immunocompromised people and those with underlying disease. However, there are no approved therapies. Moreover, RV infection cannot be prevented by a vaccine because there are over 100 serotypes. Here we report the pharmacological profile of a novel small-molecule host-targeted antiviral (HTA), KRP-A218 (A218). A highly potent and selective inhibitor of phosphatidylinositol 4 kinase beta (PI4KB), a key host factor of RV replication, A218 is undergoing clinical study.Methods in vitro antiviral activities of A218 and Vapendavir (Vap), a virus-targeted antiviral, were examined by inhibition of CPE, viral load, or replication. in vivo antiviral activity and pathological analysis of A218 were examined in coxsackievirus B3 (CVB3; belong to the genus enterovirus as with RV)-infected mice as a surrogate model of RV infection as CVB3, unlike RV, replicates very well in both mouse and human tissue. Daily oral dosing of A218 (1-10 mg/kg) was started 2 days post intraperitoneal infection with CVB3. Tissue viral load, pancreas pathological change at 4 days post infection, and survival rate up to 14 days were evaluated. PI4KB heterozygous kinase-dead mice (PI4KB KD) were established by a CRISPR-Cas9 system. Viral load and survival rate following viral infection were evaluated in these mice.ResultsA218 showed broad antiviral activity for RV and enteroviruses (Table) and has a higher barrier to drug resistance than Vap. These results are consistent with expectations for HTAs. Repeated dosing of A218 starting 2 days post infection decreased viral load and improved acute pancreatitis, accompanied by decrease of inflammatory and pancreatitis markers in plasma. Moreover, therapeutic dosing of A218 improved survival rate in a CVB3-infected lethal mouse model (Figure). These results show the first evidence that a PI4KB inhibitor has potent therapeutic efficacy in a severe viral infection model. Similar effects were observed in PI4KB KD, supporting the on-target effect of A218.Table. Antiviral activity of A218 and Vap against RV/EV infection Figure. Therapeutic effect of A218 on survival rate in CVB3-infected miceConclusionA218 is a promising therapeutic agent for improving the exacerbation of pathological conditions caused by RV infection. Nonclinical package including GLP-Tox also supports the ongoing first-in-human study of A218.Disclosures Toshiyuki Matsui, MPharm, Kyorin pharmaceutical Co., LTD (Employee) Motomichi Fujita, PhD, Kyorin Pharmaceutical Co., Ltd. (Employee) Yuji Ishibashi, PhD, Kyorin Pharmaceutical co., ltd. (Employee) Tyzoon Nomanbhoy, PhD, ActivX Biosciences (Other Financial or Material Support, Full time employee of ActivX, a wholly owned subsidiary of Kyorin Pharmaceuticals) Jonathan S. Rosenblum, PhD, ActivX Biosciences (Employee) Michiaki Nagasawa, PhD, Kyorin Pharmaceutical Co., Ltd (Employee)

Coxsackie virus B infection and pregnancy

Frequently, enteroviral infection with Coxsackie virus B (CV-B) goes unnoticed, without or only with mild symptoms. The infection is well known for the gastro-intestinal transmission, but recently, the severe forms of neonate’s infection, suggested that vertical transmission should not be neglected and moreover the fetal consequences should be studied. This review relates the most important findings regarding infection with CV-B during pregnancy, most of the information being based on murine studies. It seems that CV-B infection is associated with high rate of abortion and could also impair fertility. Regarding long term effects, CV-B might cause autoimmune diseases, congenital heart defects and neurologic disorders. Severe acute disease in new-born is generally acquired from a symptomatic mother that develop a febrile illness during the last week of pregnancy.

Analysis of the long noncoding RNA profiles of RD and SH-SY5Y cells infected with coxsackievirus B5, using RNA sequencing.

Hand, foot, and mouth disease caused by coxsackievirus B5 (CV-B5) is a considerable threat to infant health, especially with regard to neurological damage. Long noncoding RNAs (lncRNAs) are known to play pivotal roles in virus-host interactions. However, the roles of lncRNAs in CV-B5-host interactions have not yet been elucidated. In the current study, we used RNA sequencing to determine the expression profiles of lncRNAs in CV-B5-infected human rhabdomyosarcoma (RD) and SH-SY5Y cells. Our results showed that, of the differentially expressed lncRNAs, 508 were upregulated and 760 were downregulated in RD cells. Of these, 46.2% were long noncoding intergenic RNAs (lincRNAs), 28.6% were antisense lncRNAs, 24.1% were sense overlapping lncRNAs, and 1.0% were sense intronic lncRNAs. Moreover, 792 lncRNAs were upregulated and 811 lncRNAs were downregulated in SH-SY5Y cells, 48.6% of which were lincRNAs, 34.7% were antisense lncRNAs, 16.0% were sense overlapping lncRNAs, and 0.8% were sense intronic lncRNAs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that lncRNAs that were differentially expressed in CV-B5-infected RD cells were associated with disease, and those differentially expressed in SH-SY5Y cells were involved in signaling pathways. RT-qPCR analysis of seven lncRNAs supported these results. Moreover, our study revealed that lncRNA-IL12A inhibits viral replication. We conclude that lncRNAs constitute potential novel molecular targets for the prevention and treatment of CV-B5 infection and also may serve to distinguish neurogenic diseases caused by CV-B5 infection.

Virus-Host Interactions of Enteroviruses and Parvovirus B19 in Myocarditis

Viral diseases are a major threat to modern society and the global health system. It is therefore of utter relevance to understand the way viruses affect the host as a basis to find new treatment solutions. The understanding of viral myocarditis (VMC) is incomplete and effective treatment options are lacking. This review will discuss the mechanism, effects, and treatment options of the most frequent myocarditis-causing viruses namely enteroviruses such as Coxsackievirus B3 (CVB3) and Parvovirus B19 (PVB19) on the human heart. Thereby, we focus on: 1. Viral entry: CVB3 use Coxsackievirus-Adenovirus-Receptor (CAR) and Decay Accelerating Factor (DAF) to enter cardiac myocytes while PVB19 use the receptor globoside (Gb4) to enter cardiac endothelial cells. 2. Immune system responses: The innate immune system mediated by activated cardiac toll-like receptors (TLRs) worsen inflammation in CVB3-infected mouse hearts. Different types of cells of the adaptive immune system are recruited to the site of inflammation that have either protective or adverse effects during VMC. 3. Autophagy: CVB3 evades autophagosomal degradation and misuses the autophasomal pathway for viral replication and release. 4. Viral replication sites: CVB3 promotes the formation of double membrane vesicles (DMVs), which it uses as replication sites. PVB19 uses the host cell nucleus as the replication site and uses the host cell DNA replication system. 5. Cell cycle manipulation: CVB3 attenuates the cell cycle at the G1/S phase, which promotes viral transcription and replication. PVB19 exerts cell cycle arrest in the S phase using its viral endonuclease activity. 6. Regulation of apoptosis: Enteroviruses prevent apoptosis during early stages of infection and promote cell death during later stages by using the viral proteases 2A and 3C, and viroporin 2B. PVB19 promotes apoptosis using the non-structural proteins NS1 and the 11 kDa protein. 7. Energy metabolism: Dysregulation of respiratory chain complex expression, activity and ROS production may be altered in CVB3- and PVB19-mediated myocarditis. 8. Ion channel modulation: CVB3-expression was indicated to alter calcium and potassium currents in Xenopus laevis oocytes and rodent cardiomyocytes. The phospholipase 2-like activity of PVB19 may alter several calcium, potassium and sodium channels. By understanding the general pathophysiological mechanisms of well-studied myocarditis-linked viruses, we might be provided with a guideline to handle other less-studied human viruses.

Abstract 9991: Can Isosorbide Dinitrate Save Lives of Hospitalized COVID Patients with Hypertension and a Defined Cardiac Injury?

Introduction: As defined by MB-CPK, hsTn-I, MB, or EKG and/or cardiac echo abnormalities, cardiac injury (CI) determines a median survival time (MST) of 10 days for hospitalized COVID patients (HCPs). HCPs without defined CI have an MST beyond 39 days (Huang, et al ., see Figure). Hypertension (HTN) presents in 17-41% of HCPs in various studies, with COVID mortality independent of HTN. Therefore, a randomized clinical trial (RCT) is proposed for HCPs with CI and incidental BP elevation to compare IV isosorbide dinitrate (ISDN) with usual anti-HTN care (UC). Vasodilatory ISDN lowers BP, and has been proposed an an anti-SARS-CoV-2 drug. Others report improved survival of ISDN treated Coxsackie B3 virus-infected mice as evidence for anti-viral activity. The endpoint for this CI RCT pilot of 100 ISDN-treat HCPs and 100 UC controls is mortality. Secondary endpoints are interval biomarkers to dissect ISDN anti-viral action. Methods: Log-rank analysis was performed with 1:1 allocation. Accrual time was 180 days with a 60-day follow-up. Power was 0.8 with a type I error of 0.05. Results: For 200 total subjects, an MST greater than 14.9 days in the ISDN arm was significant if UC stays at 10 days (see Figure). Conclusions: Testing repurposed ISDN as a COVID drug is feasible. A successful pilot with improved MST suggests ISDN has anti-SARS-CoV-2 action, since COVID mortality is independent of HTN. Biomarkers could include viral clearance, oxygenation, D-dimer, IL-6, LDH, as well as platelet and lymphocyte counts. The need to treat HCPs with elevated BP per guidelines permits study entry. Cross-over treatment occurs if a regimen fails. Immunomodulators and remdesivir are administered per COVID treatment guidelines for all HCPs. IV ISDN, bolused and/or infused, avoids 1 st pass hepatic effects. IV IDSN is approved in most countries, but not in the USA or Canada. A successful pilot would permit larger IV ISDN RCTs as IND RCTs, and can serve as a template for other treatments for HCPs with defined CI.

Consequences of Both Coxsackievirus B4 and Type 1 Diabetes on Female Non-Obese Diabetic Mouse Kidneys.

Despite the 2019 Executive Order on Advancing American Kidney Health Initiative, kidney disease has moved up in rank from the 9th to the 8th leading cause of death in the United States. A recent push in the field of nephrology has been to identify molecular markers and/or molecular profiles involved in kidney disease process or injury that can help identify the cause of injury and predict patient outcomes. While these studies have had moderate success, they have not yet considered that many of the health conditions that cause kidney disease (diabetes, hypertension, etc.) can also be caused by environmental factors (such as viruses), which in and of themselves can cause kidney disease. Thus, the goal of this study was to identify molecular and phenotypic profiles that can differentiate kidney injury caused by diabetes (a health condition resulting in kidney disease) and coxsackievirus B4 (CVB4) exposure (which can cause diabetes and/or kidney disease), both alone and together. Non-obese diabetic (NOD) mice were used for this study due to their susceptibility to both type 1 diabetes (T1D)- and CVB4-mediated kidney injury, in order to glean a better understanding of how hyperglycemia and viral exposure, when occurring on their own and in combination, may alter the kidneys’ molecular and phenotypic profiles. While no changes in kidney function were observed, molecular biomarkers of kidney injury were significantly up- and downregulated based on T1D and CVB4 exposure, both alone and together, but not in a predictable pattern. By combining individual biomarkers with function and phenotypic measurements (i.e., urinary albumin creatinine ratio, serum creatinine, kidney weight, and body weight), we were able to perform an unbiased separation of injury group based on the type of injury. This study provides evidence that unique kidney injury profiles within a kidney disease health condition are identifiable, and will help us to identify the causes of kidney injury in the future.

Photoelectrochemical biosensor for Coxsackievirus B3 detection with recombinase polymerase amplification coupled with ZnSeNSs/AuNPs/BNNSs modified electrode

Coxsackievirus B3 (CVB3), a member of the genus Enterovirus, has high risks for incidence and mortality. Developing speedy and sensitive molecular diagnostics for CVB3 detection is challenging. Herein, a novel highly sensitive photoelectrochemical (PEC) biosensor based on nanocomposite (ZnSeNSs/AuNPs/BNNSs) modified electrode was fabricated. This biosensor relied on reverse transcription of CVB3 RNA and the rapid isothermal amplication by recombinase polymerase amplification (RPA). The amplified product was identified and linked to photoelectrode. Then biotin on the amplified product was connected to strepavidin-alkaline phosphatase (SA-ALP). The alkaline phosphatase catalyzed the hydrolysis of substrate 4-aminophenylphosphate monosodium (4-APP) to generate free 4-aminophenol which was a electron donor. An enhancing PEC signal was obtained. Therefore, a relationship between photocurrent response and CVB3 concentration was established. Under the optimal conditions, this PEC biosensor displayed a liner photocurrent response in the range of 0.1 fM ∼100 fM with a low detection limit of 30 aM. Furthermore, this strategy with satisfactory stability and selectivity provides an innovative way for CVB3 detection and lays a foundation for the research of portable devices for clinical sample processing and analysis in the further.

Neutralization of interleukin-38 exacerbates coxsackievirus B3-induced acute myocarditis in mice

BackgroundInterleukin (IL)-38, a novel member of the IL-1 family, has been reported to be involved in several diseases associated with viral infection. However, the expression and functional role of IL-38 in acute viral myocarditis (AVMC) have not been investigated.MethodsMale BALB/c mice were treated with intraperitoneal (i.p.) injection of coxsackievirus B3 (CVB3) for establishing AVMC models. On day 7 post-injection, the expression of IL-38 and IL-36R (IL-36 receptor) were measured. Mice were then treated with i.p. injection of mouse Anti-IL-38 Antibodies (Abs) for neutralization of IL-38. The survival, bodyweight loss, cardiac function, and myocarditis severity of mice were recorded. The percentages of splenic Th1 and Th17 cells, the expression levels of Th1/Th17-related master transcription factors (T-bet and RORγt) and cytokines were determined by flow cytometry, RT-qPCR, and ELISA, respectively. Cardiac viral replication was further detected.ResultsThe mRNA and protein expression levels of IL-38 in myocardium and serum, as well as cardiac IL-36R mRNA levels were significantly elevated in mice with AVMC. Increased IL-38 levels were negatively correlated with the severity of AVMC. Neutralization of IL-38 exacerbated CVB3-induced AVMC, as verified by the lower survival rate, impaired cardiac function, continuous bodyweight loss, and higher values of HW/BW and cardiac pathological scores. In addition, neutralization of IL-38 suppressed Th1 cells differentiation while promoted Th17 cells differentiation, accompanied by decreased T-bet mRNA expression and increased RORγt expression. Down-regulation of IFN-γ and up-regulation of IL-17, TNF-α, and IL-6 mRNA and protein expression levels in myocardium and serum were also observed in the IL-38 neutralization group. Furthermore, neutralization of IL-38 markedly promoted cardiac viral replication.ConclusionsNeutralization of IL-38 exacerbates CVB3-induced AVMC in mice, which may be attributable to the imbalance of Th1/Th17 cells and increased CVB3 replication. Thus, IL-38 can be considered as a potential therapeutic target for AVMC.

Transcriptomic analysis reveals that coxsackievirus B3 Woodruff and GD strains use similar key genes to induce FoxO signaling pathway activation in HeLa cells.

Coxsackievirus B3 (CVB3) is a major cause of viral myocarditis in humans. Although there have been studies on CVB3 infection and pathogenesis, the precise disease mechanism is still not clear. In this study, we used RNA-seq technology to compare the transcriptomic profile of virus-infected HeLa cells to that of uninfected cells to identify key genes involved in host-virus interaction. For this, two CVB3 strains, CVB3 Woodruff, an experimental strain, and GD16-69/GD/CHN/2016, a clinical strain, were selected to examine the common mechanisms underlying their infection. Transcriptomic profiles revealed increased expression of the cell cycle genes CCNG2, GADD45B, PIM1, RBM15, KLF10, and RIOK3 and decreased expression of CYBA. The autophagy-related genes ATG12 and YOD1 were found to be upregulated, while the expression of SOD2 and XPO1 increased slightly in infected cells, and only a minor change was observed in GABARAP expression. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed the FoxO signaling pathway to be enriched and showed a close interaction with differentially expressed genes (DEGs) in the protein-protein interaction network. DEGs associated with related pathways such as cell cycle, autophagy, and oxidative stress resistance were also confirmed by qRT-PCR. In summary, the FoxO signaling pathway was activated during infection with both CVB3 strains and was found to have a regulatory role in downstream pathways such as cell cycle, autophagy, oxidative stress resistance, and the antiviral immune response.

SARS-CoV-2-induced autophagy dysregulation may cause neuronal dysfunction in COVID-19.

SARS-CoV-2-induced autophagy dysregulation may cause neuronal dysfunction in COVID-19.

Spectrum of Viral Pathogens Identified in Children with Clinical Myocarditis (Pre-Coronavirus Disease-2019, 2000-2018): Etiologic Agent Versus Innocent Bystander

To identify the etiologies of viral myocarditis in children in the pre-coronavirus disease 2019 era. This was a retrospective review of all patients (age <18years) diagnosed with myocarditis and hospitalized at Rady Children's Hospital San Diego between 2000 and2018. Twenty-nine patients met inclusion criteria. Of 28 (97%) patients who underwent testing for viruses, polymerase chain reaction was used in 24 of 28 (86% of cases), and 16 of 24 (67%) detected a virus. Pathogens were rhinovirus (6), influenza A/B (4), respiratory syncytial virus (RSV) (3), coronavirus (3), parvovirus B19 (2), adenovirus (2), and coxsackie B5 virus, enterovirus, and parainfluenza virus type 2 in one case each. Six (21%) patients had no pathogen detected but imaging and other laboratory test results were compatible with myocarditis. Age 0-2years was associated with RSV, influenza A/B, coronavirus, and enteroviruses (P<.001). Twenty-one patients (72%) experienced full clinical recovery. Three patients (10%) required venoarterial extracorporeal membrane oxygenation (VA-ECMO), and all 3 recovered. Three others (10%) required and underwent successful cardiac transplantation without complications. Two patients (7%) died 9-10days after hospitalization (1 had RSV and 1 had influenza A/B). Two other patients presented with complete atrioventricular block; 1 case (rhinovirus) resolved spontaneously, and 1 (coronavirus) resolved after support with VA-ECMO. Age <2years, female sex, lower ejection fraction at admission, and greater initial and peak levels of brain natriuretic peptide were significant predictors of critical outcomes (use of VA-ECMO, listing for cardiac transplantation, and death). Viral nucleic acid-based testing revealed a wider spectrum of viruses that could be associated with myocarditis in children than previously reported and traditionally anticipated. A predilection of certain pathogens in the very young patients was observed. Whether the observed range of viral agents reflects an undercurrent of change in viral etiology or viral detection methods is unclear, but the wider spectrum of viral pathogens found underscores the usefulness of polymerase chain reaction testing to explore possible viral etiologies of myocarditis in children.

Genetic diversity of enteroviruses in patients with respiratory infection

Enterovirus genetic variability underlies the variety of clinical forms of diseases they cause. The aim of the presented study was to establish the genetic diversity of enteroviruses (EVs) that caused acute respiratory infection (ARI) in 2016–2019. Biological samples were obtained from 203 patients with various forms of ARI, EV detection was carried out by RT-PCR, followed by sequencing of the main capsid protein gene and phylogenetic reconstruction. EV RNA was detected in 34.4 % of samples, most often in children aged 1–6 years (53.1–54.8 %). Coxsackieviruses B were found in patients with respiratory enterovirus infection (EVI) significantly more often than other EVs, the dominant serotypes were Coxsackievirus B4, B5. Despite the significant genetic diversity of EVs identified in patients with ARI (three genetic lines of Coxsackievirus B5, two genotypes of Coxsackievirus B2, one genotype of Coxsackievirus B3, three genovariant Coxsackievirus B4, one genovariant Coxsackievirus B1), there is no evidence of their connection with the formation of the respiratory form of EVI.The high level of genetic variability of EVs requires regular molecular-epidemiological surveillance for the identification of emerging genetic variants and assessment of their epidemic potential.

Stability of SARS-CoV-2 and other airborne viruses under different stress conditions.

Viral stability under stress conditions may directly affect viral dissemination, seasonality, and pathogenesis. We exposed airborne viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), mumps virus, coxsackievirus B5, human rhinovirus A16, and respiratory syncytial virus, to different temperatures, UV light exposure time, pH values, and osmotic pressures and measured the remaining viral infectivity. Reduced thermal stability was observed for coxsackievirus B5 at 45 °C, while SARS-CoV-2 demonstrated residual infectivity at 55 °C. UV light exposure was an efficient means of viral inactivation but was less efficient for non-enveloped viruses. Rhinovirus A16 and respiratory syncytial virus demonstrated extreme sensitivity to acid conditions, while SARS-CoV-2, rhinovirus A16, and respiratory syncytial virus were unstable in an alkaline environment. The information obtained in this study will be useful for the development of viral inactivation methods and may be correlated with epidemiological and seasonal viral characteristics.

Viral pathogens in urban stormwater runoff: Occurrence and removal via vegetated biochar-amended biofilters

Urban runoff is one of the greatest sources of microbial pollution to surface waters. Biofilters can limit the impact of stormwater runoff on surface water quality by diverting runoff from receiving waters. However, our understanding of how biofilter design choices, including the addition of vegetation and geomedia, may impact the removal of pathogens is lacking. In this study, we characterized viruses (adenovirus, enterovirus, norovirus GII, crAssphage) in San Francisco Bay area urban runoff and assessed the removal of lab-cultured viruses (MS2, adenovirus 2, coxsackievirus B5) from biochar-amended biofilter mesocosms during challenge testing. We quantified viruses using (RT-)qPCR and F+ coliphage plaque assays. We found that all the pathogenic viruses targeted were found at low concentrations (adenovirus: all positive samples were <limit of quantification, enterovirus: <limit of quantification-1.9 × 102 gc/L, norovirus GII: <limit of quantification-1.2 × 102 gc/L) in San Francisco Bay area urban runoff and the presence of norovirus GII in runoff was associated with developed land use and decreased precipitation. Biofilters had variable success in removing adenovirus, enterovirus, and MS2 from runoff in laboratory-scale column experiments. In addition, there was no significant difference in the removal of each virus in vegetated versus non-vegetated biofilters, with the exception of MS2 which had slightly higher removal in vegetated biofilters (0.40 log10 units, Welch's t-test, p = 0.004). When comparing removal of human viruses and viral indicators, adenovirus and enterovirus were removed more efficiently (log10-removal adenovirus = 3.2; log10-removal enterovirus = 1.1) than indicator virus MS2 (log10-removal by RT-qPCR = 0.36, log10-removal by plaque assay = 0.36). These results provide evidence that MS2 may be a conservative indicator for human virus removal in biofiltration systems, but more work is needed to examine this relationship. Results from this study can help inform design choices regarding biofilters intended to improve water quality and our understanding of virus attenuation in biofiltration systems.