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.

3D-QSAR, ADMET and Molecular Docking Study of a Series of 2-Substituted 1H-Benzimidazole-4-Carboxamide Derivatives Inhibitors Against Enteroviruses

A series of bioactive benzimidazoles were theoretically evaluated for their effect against enteroviruses, particularly coxsackievirus B and coxsackievirus A16. The structure-activity relationship allowed to deduce a very efficient model that can be used as a reference for the synthesis of compounds against this type of virus. The model used is essentially based on the RML and NN method, and its validation was carried out using cross-validation. The results obtained by molecular docking and ADMET of compound 9 as an inhibitor of enterovirus A16 give the proposal of the drug of this series.

Coxsackievirus B4 Transplacental Infection Severely Disturbs Central Tolerogenic Mechanisms in the Fetal Thymus

Thymus plays a fundamental role in central tolerance establishment, especially during fetal life, through the generation of self-tolerant T cells. This process consists in T cells education by presenting them tissue-restricted autoantigens promiscuously expressed by thymic epithelial cells (TECs), thus preventing autoimmunity. Thymus infection by Coxsackievirus B (CV-B) during fetal life is supposed to disturb thymic functions and, hence, to be an inducing or accelerating factor in the genesis of autoimmunity. To further investigate this hypothesis, in our current study, we analyzed thymic expression of autoantigens, at the transcriptional and protein level, following in utero infection by CV-B4. mRNA expression levels of Igf2 and Myo7, major autoantigens of pancreas and heart, respectively, were analyzed in whole thymus and in enriched TECs together along with both transcription factors, Aire and Fezf2, involved in autoantigens expression in the thymus. Results show that in utero infection by CV-B4 induces a significant decrease in Igf2 and Myo7 expression at both mRNA and protein level in whole thymus and in enriched TECs as well. Moreover, a correlation between viral load and autoantigens expression can be observed in the whole thymus, indicating a direct effect of in utero infection by CV-B4 on autoantigens expression. Together, these results indicate that an in utero infection of the thymus by CV-B4 may interfere with self-tolerance establishment in TECs by decreasing autoantigen expression at both mRNA and protein level and thereby increase the risk of autoimmunity onset.

Time-Series Analysis of Coxsackievirus B Serotype Surveillance Data in Japan

ObjectiveStudies have identified serotypes of Coxsackievirus B (CVB) enterovirus as a cause of type 1 diabetes. Studies have also identified cyclical variations in type 1 diabetes incidence—peak incidences occurring in 4- to 6-years periods in two regions in England, a 5-year period in Western Australia, and 5.33-year period in Poland. To date, no studies have investigated whether CVB infection rates demonstrate similar cyclical variation characteristics. The purpose of this study was to characterize periodicity in CVB surveillance data.ResultsMaximum entropy spectral analysis was performed on monthly CVB surveillance data. In addition to demonstrating a 1-year cycle for all serotypes, spectral peaks demonstrated dominant cycles—6.9-, 3.8-, 4.3-, 9.5-, and 7.8- year periods for CVB1, CVB2, CVB3, CVB4, and CVB5, respectively. Pearson correlation was used to compare the least-squares fit curves based on periods estimated from the analysis with the original data. The results for all five serotypes—CVB1, CVB2, CVB3, CVB4, and CVB5—demonstrated good correlation—ρ = 0.96, ρ = 0.60, ρ = 0.90, ρ = 0.88, and ρ = 0.67, respectively. This method could be a useful tool for the efficient investigation of CVB as a pathogen of type 1 diabetes.

Abstract 16811: In-Utero Coxsackievirus Type B Infection is Associated With Congenital Pulmonary Atresia

Introduction: Maternal exposure to various pathogens during pregnancy has been well established as a cause of congenital heart defects (CHD). Suggestive, although inconclusive, data exists of an association between Coxsackievirus B (CVB) infection during pregnancy and CHD. Therefore, this study was undertaken to examine association between perinatal infection with CVB and CHD. Hypothesis: CVB infection during pregnancy may play a role in the pathogenesis of CHD. Methods: In a prospective study of 122 pregnant women with pregnancies affected with CHD, circulating CVB IgG and IgM antibodies in the maternal serum were quantified. To determine specific CVB serotypes involved, Neutralizing antibodies (NA) were measured against three CVB serotypes: CVB1-Chi07, CVB3, Nancy, and CVB4, J.V.B. Results: A significant number of the women were positive for CVB antibodies; 31.1% for IgG and 6.6% for IgM. Elevated levels of IgG were seen in 50% of women carrying babies with Pulmonary Atresia (PA) (p=0.04). In addition, 55.6% (p=0.100) of moms carrying babies with Hypoplastic Left Heart Syndrome (HLHS), aortic atresia/mitral stenosis (AA/MS) variant and 47.1% (p=0.127) of HLHS AA/mitral atresia (MA) had positive IgG levels. Among babies of women with positive IgM titers, 75% (p=0.001) had isolated Ventricular Septal Defect (VSD), comprising 26.1% (6/23) of all babies with isolated VSD in the cohort.We also show a possible association between CVB4 and CHD in the cohort. All mothers with positive IgM titers whose babies were born with a VSD had elevated CVB4 NA (p=0.001). Among women with elevated IgG having babies with PA, 40.0% (p=0.061) had elevated CVB1 NA, 70.0% (p=0.014) had elevated CVB3 NA, and 40.0% (p=0.876) had elevated CVB4 NA. Conclusion: We present data from a clinical study suggesting that CVB infection during pregnancy may contribute to the burden of CHD in affected babies. Our data demonstrates not only an association of CVB with CHD in pregnant women but this is the first report showing significant association between in-utero CVB infection and PA. Our study has broad implications for the future understanding and potential management of CHD as well as introduces a new avenue of research for cardiac defects.

Preliminary Anti-Coxsackie Activity of Novel 1-[4-(5,6-dimethyl(H)- 1H(2H)-benzotriazol-1(2)-yl)phenyl]-3-alkyl(aryl)ureas

Background: Coxsackievirus infections are associated with cases of aseptic meningitis, encephalitis, myocarditis, and some chronic disease. Methods: A series of benzo[d][1,2,3]triazol-1(2)-yl derivatives (here named benzotriazol-1(2)-yl) (4a-i, 5a-h, 6a-e, g, i, j and 7a-f, h-j) were designed, synthesized and in vitro evaluated for cytotoxicity and antiviral activity against two important human enteroviruses (HEVs) members of the Picornaviridae family [Coxsackievirus B 5 (CVB-5) and Poliovirus 1 (Sb-1)]. Results: Compounds 4c (CC50 >100 μM; EC50 = 9 μM), 5g (CC50 >100 μM; EC50 = 8 μM), and 6a (CC50 >100 μM; EC50 = 10 μM) were found active against CVB-5. With the aim of evaluating the selectivity of action of this class of compounds, a wide spectrum of RNA (positive- and negativesense), double-stranded (dsRNA) or DNA viruses were also assayed. For none of them, significant antiviral activity was determined. Conclusion: These results point towards a selective activity against CVB-5, an important human pathogen that causes both acute and chronic diseases in infants, young children, and immunocompromised patients.