Transcriptome Analysis in Air-Liquid Interface Porcine Respiratory Epithelial Cell Cultures Reveals That the Betacoronavirus Porcine Encephalomyelitis Hemagglutinating Virus Induces a Robust Interferon Response to Infection.

Porcine hemagglutinating encephalomyelitis virus (PHEV) replicates in the upper respiratory tract and tonsils of pigs. Using an air-liquid interface porcine respiratory epithelial cells (ALI-PRECs) culture system, we demonstrated that PHEV disrupts respiratory epithelia homeostasis by impairing ciliary function and inducing antiviral, pro-inflammatory cytokine, and chemokine responses. This study explores the mechanisms driving early innate immune responses during PHEV infection through host transcriptome analysis. Total RNA was collected from ALI-PRECs at 24, 36, and 48 h post inoculation (hpi). RNA-seq analysis was performed using an Illumina Hiseq 600 to generate 100 bp paired-end reads. Differential gene expression was analyzed using DeSeq2. PHEV replicated actively in ALI-PRECs, causing cytopathic changes and progressive mucociliary disruption. Transcriptome analysis revealed downregulation of cilia-associated genes such as CILK1, DNAH11, LRRC-23, -49, and -51, and acidic sialomucin CD164L2. PHEV also activated antiviral signaling pathways, significantly increasing the expression of interferon-stimulated genes (RSAD2, MX1, IFIT, and ISG15) and chemokine genes (CCL5 and CXCL10), highlighting inflammatory regulation. This study contributes to elucidating the molecular mechanisms of the innate immune response to PHEV infection of the airway epithelium, emphasizing the critical roles of the mucociliary, interferon, and chemokine responses.

COVID-19 entre profissionais fisioterapeutas: uma análise segundo prevalência e fatores associados / Covid-19 among brazilian physiotherapists: analysis of prevalence and associated factors / Covid-19 entre los fisioterapeutas brasileños: análisis de prevalencia y factores asociados

Objetivo: analisar a prevalência da COVID-19 entre os fisioterapeutas brasileiros e os fatores associados segundo características demográficas e ocupacionais. Método: estudo transversal, analítico, segundo inquérito on-line, com a participação de 670 fisioterapeutas de todas as regiões do Brasil. Utilizou-se uma adaptação do método respondent driven sampling ao ambiente virtual para a coleta de dados. Análises bivariadas e de regressão logística múltipla foram utilizadas para identificar associação entre o diagnóstico da COVID-19 e variáveis demográficas e ocupacionais. Considerou-se variáveis estatisticamente significativas com base em um p<0,05. Resultados: a prevalência da COVID-19 foi de 30% (IC95%: 27,8-32,3). Fisioterapeutas da região Sudeste tiveram menores chances de ter diagnóstico da COVID-19. Fisioterapeutas que prestaram assistência em hospital de campanha, que ficaram isolados da família e que tem crianças menores de 12 em casa tiveram chances aumentadas para o diagnóstico da infecção. Conclusão: questões sociodemográficas e ocupacionais impactam no aumento do diagnóstico de Covid-19 entre profissionais fisioterapeutas, o que enfatiza a necessidade de um sistema de saúde de qualidade, igualitário nas diferentes regiões brasileiras.(AU)

Objective: to estimate the prevalence of COVID-19 among Brazilian physiotherapists and its associated factors. Method: cross-sectional study, according to an online survey, with the participation of 670 physiotherapists from all regions of Brazil. An adaptation of the respondent driven sampling method to the virtual environment was used to collect data. Bivariate and multiple logistic regression analyzes were used to identify associations between the diagnosis of COVID-19 and demographic and occupational variables. Variables were considered statistically significant based on p<0.05. Results: the prevalence of COVID-19 was 30% (95%CI: 27.8-32.3). In the Southeast region, physiotherapists were less likely to be diagnosed with COVID-19. Physiotherapists who provided care in a field hospital, who were isolated from their families and who have children under 12 years of age at home had an increased chance of being diagnosed with the infection.Conclusion: sociodemographic and occupational issues impact the increase in COVID-19 diagnoses among physiotherapists, which emphasizes the need for a quality and egalitarian health system in different Brazilian regions.(AU)

Objetivo: evaluar la tasa de prevalencia del COVID-19 en fisioterapeutas de Brasil y analizar sus factores asociados. Método: realizamos un estudio transversal mediante una encuesta on-line, en la que participaron 670 fisioterapeutas de todas las áreas de Brasil. Para la recogida de datos se utilizó una adaptación del método respondent driven sampling al entorno virtual. Se utilizaron análisis bivariados y de regresión logística múltiple para identificar la asociación entre el diagnóstico COVID-19 y variables demográficas y ocupacionales. Las variables se consideraron estadísticamente significativas en función de una p<0,05. Resultados: la prevalencia de COVID-19 fue del 30% (IC 95%: 27,8-32,3). Los fisioterapeutas del sudeste tenían menos probabilidades de ser diagnosticados de COVID-19. Los fisioterapeutas que prestaban asistencia en un hospital de campaña, que estaban aislados de sus familias y que tenían hijos menores de 12 años en casa tenían más probabilidades de que se les diagnosticara la infección. Conclusiones: aspectos sociodemográficos y ocupacionales inciden en el aumento del diagnóstico de COVID-19 entre los fisioterapeutas profesionales, lo que enfatiza la necesidad de un sistema de salud de calidad e igualitario en las diferentes regiones brasileñas.(AU)

Characteristics of endemic human Coronavirus infections during times of COVID-19 pandemic / Características de las infecciones causadas por Corona virus humanos endémicos durante la pandemia COVID-19

Abstract Introduction : After the implementation of mitigation strategies during the COVID-19 pandemic, the incidence of respiratory viruses, including human coronaviruses (HCoV), experienced a significant decrease. The aim of this study is to characterize the epidemiology and clinical aspects of HCoV infections in ambulatory adults during COVID-19 pandemic times. Methods : descriptive, prospective, longitudinal study performed in a private hospital in La Plata, Buenos Aires, Argentina between November 2020 and October 2022; 458 outpatient adults with upper respiratory tract infections (URTI) were studied undergoing clinical and microbiological follow-up. Results : 44 (9.6%) subjects were positive by multiplex PCR for HCoV. 14 of them for 229E (31.8%), 13 for OC43 (29.5%), 11 for HKU-1 (25.1%) and 6 for NL63 (13.6%). A repeated PCR was positive for the same HCoV in 19 (57%) of 33 patients on day 3-5. No hospitalizations or deaths were reported. Discussion : Endemic HCoV caused a significant pro portion of URTI among outpatient adults during COVID- 19-related restrictions times. An alternating pattern of circulation between alfa-HCoV and beta-HCoV was observed.

Resumen Introducción : Tras la implementación de estrate gias de mitigación durante la pandemia de COVID-19, la incidencia de virus respiratorios, incluyendo los coronavirus humanos (HCoV), disminuyó significati vamente. El objetivo de este estudio es caracterizar la epidemiología y los aspectos clínicos de las infecciones por HCoV en adultos ambulatorios durante la pandemia de COVID-19. Métodos : estudio descriptivo, prospectivo, longitudi nal, realizado en un hospital privado de La Plata, Buenos Aires, Argentina, entre noviembre de 2020 y octubre de 2022. Se estudiaron 458 pacientes adultos ambulatorios con infecciones del tracto respiratorio superior (ITRS) bajo seguimiento clínico y microbiológico. Resultados : 44 (9.6%) sujetos fueron positivos por PCR multiplex para HCoV. Se detectaron 14 229E (31.8%), 13 OC43 (29.5%), 11 HKU-1 (25.1%) y 6 NL63 (13.6%). Una segunda PCR fue positiva para el mismo HCoV en 19 (57 %) de 33 pacientes en los días 3-5. No se reportaron hospitalizaciones ni muertes. Discusión : los HCoV endémicos causaron una pro porción significativa de ITRS entre pacientes adultos ambulatorios durante los tiempos de restricciones rela cionados con COVID-19. Se observó un patrón alternante de circulación entre alfa-HCoV y beta-HCoV.

Computational Prediction of RNA-RNA Interactions between Small RNA Tracks from Betacoronavirus Nonstructural Protein 3 and Neurotrophin Genes during Infection of an Epithelial Lung Cancer Cell Line: Potential Role of Novel Small Regulatory RNA.

Whether RNA-RNA interactions of cytoplasmic RNA viruses, such as Betacoronavirus, might end in the biogenesis of putative virus-derived small RNAs as miRNA-like molecules has been controversial. Even more, whether RNA-RNA interactions of wild animal viruses may act as virus-derived small RNAs is unknown. Here, we address these issues in four ways. First, we use conserved RNA structures undergoing negative selection in the genomes of SARS-CoV, MERS-CoV, and SARS-CoV-2 circulating in different bat species, intermediate animals, and human hosts. Second, a systematic literature review was conducted to identify Betacoronavirus-targeting hsa-miRNAs involved in lung cell infection. Third, we employed sophisticated long-range RNA-RNA interactions to refine the seed sequence homology of hsa-miRNAs with conserved RNA structures. Fourth, we used high-throughput RNA sequencing of a Betacoronavirus-infected epithelial lung cancer cell line (Calu-3) to validate the results. We proposed nine potential virus-derived small RNAs: two vsRNAs in SARS-CoV (Bats: SB-vsRNA-ORF1a-3p; SB-vsRNA-S-5p), one vsRNA in MERS-CoV (Bats: MB-vsRNA-ORF1b-3p), and six vsRNAs in SARS-CoV-2 (Bats: S2B-vsRNA-ORF1a-5p; intermediate animals: S2I-vsRNA-ORF1a-5p; and humans: S2H-vsRNA-ORF1a-5p, S2H-vsRNA-ORF1a-3p, S2H-vsRNA-ORF1b-3p, S2H-vsRNA-ORF3a-3p), mainly encoded by nonstructural protein 3. Notably, Betacoronavirus-derived small RNAs targeted 74 differentially expressed genes in infected human cells, of which 55 upregulate the molecular mechanisms underlying acute respiratory distress syndrome (ARDS), and the 19 downregulated genes might be implicated in neurotrophin signaling impairment. These results reveal a novel small RNA-based regulatory mechanism involved in neuropathogenesis that must be further studied to validate its therapeutic use.

SARS-CoV-2 Is More Efficient than HCoV-NL63 in Infecting a Small Subpopulation of ACE2+ Human Respiratory Epithelial Cells.

Human coronavirus (HCoV)-NL63 is an important contributor to upper and lower respiratory tract infections, mainly in children, while severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of COVID-19, can cause lower respiratory tract infections, and more severe, respiratory and systemic disease, which leads to fatal consequences in many cases. Using microscopy, immunohistochemistry (IHC), virus-binding assay, reverse transcriptase qPCR (RT-qPCR) assay, and flow cytometry, we compared the characteristics of the susceptibility, replication dynamics, and morphogenesis of HCoV-NL63 and SARS-CoV-2 in monolayer cultures of primary human respiratory epithelial cells (HRECs). Less than 10% HRECs expressed ACE2, and SARS-CoV-2 seemed much more efficient than HCoV-NL63 at infecting the very small proportion of HRECs expressing the ACE2 receptors. Furthermore, SARS-CoV-2 replicated more efficiently than HCoV-NL63 in HREC, which correlates with the cumulative evidence of the differences in their transmissibility.

Primer aislamiento y caracterización de la cepa prototipo del virus SARS-CoV-2 a inicios de la pandemia de la COVID-19 en el Perú

Introducción: Actualmente los contagios por el virus del SARS-CoV-2 supera los 600 millones de casos en el mundo. Objetivo: Aislar y caracterizar el virus SARS-CoV-2 causante de la COVID-19 a inicios de la pandemia en el Perú. Materiales y métodos: Se realizó el aislamiento viral a partir de 20 muestras de hisopado nasal y faríngeo positivas a SARS-CoV-2 por RT-PCR. El aislamiento se realizó en las líneas celulares Vero ATCC CCL-81 y Vero E6, evaluando el efecto citopático, la presencia del virus por RT-PCR, inmunofluorescencia indirecta (IFI) y posterior identificación por secuenciación genómica. Posteriormente, uno de los aislamientos de mayor circulación fue seleccionado y denominado cepa prototipo (PE/B.1.1/28549/2020), realizándose 10 pasajes sucesivos en células Vero ATCC CCL-81 para evaluar la dinámica de mutaciones. Resultados: Se observaron 11 aislamientos de virus por efecto citopático confirmándose por RT-PCR e IFI, de los cuales 6 fueron secuenciados identificándose los linajes B.1, B.1.1, B.1.1.1 y B.1.205, según el comité Pango de los genomas. La cepa prototipo corresponde a la variante B.1.1 y el análisis de las secuencias de los pasajes sucesivos mostró mutaciones a nivel de la proteína de la espiga (S) del virus, sin variación en la identidad del linaje. Conclusiones: Se aislaron 4 linajes en la línea celular Vero ATCC CCL-81. Los subcultivos en la misma línea celular muestran mutaciones en la proteína de la espiga, lo que indica mayor adaptabilidad a la célula hospedera y variación de la patogenicidad in vitro, comportamiento que le permite tener más éxito de supervivencia.

Introduction: Currently, infections caused by the SARS-CoV-2 virus exceed 600 million cases in the world. Objective: Isolation and characterization of the SARS-CoV-2 virus causing COVID-19 at the beginning of the pandemic in Peru. Materials and methods: Twenty nasal and pharyngeal swab samples were isolated from SARS-CoV-2 using two cell lines, Vero ATCC CCL-81 and Vero E-6; virus identification was performed by RT-PCR and the onset of cytopathic effect (CPE) was evaluated by indirect immunofluorescence and subsequent identification by genomic sequencing. One of the most widely circulating isolates were selected and named the prototype strain (PE/B.1.1/28549/2020). Then 10 successive passages were performed on Vero ATCC CCL-81 cells to assess mutation dynamics. Results: We detected 11 virus isolates by cytopathic effect, and subsequently confirmed by RT-PCR and indirect immunofluorescence. Of these, six were sequenced and identified as the lineages B.1, B.1.1, B.1.1.1, and B.1.205 according to the Pango lineage nomenclature. The prototype strain corresponded to lineage B.1.1. The analysis of the strains from the successive passages showed mutations mainly at in the spike (S) protein of the virus without variation in the identity of the lineage. Conclusions: Four lineages were isolated in the Vero ATCC CCL-81 cell line. Subcultures in the same cell line show mutations in the spike protein indicating greater adaptability to the host cell and variation in pathogenicity in vitro, a behavior that allows it to have more survival success.

Avaliação do uso de estratégias não invasivas em pacientes com insuficiência respiratória por COVID-19 / Evaluation of the use of non-invasive strategies in patients with respiratory failure by COVID-19

Introdução: Durante a pandemia de COVID-19, tornou-se necessário o uso de estratégias no tratamento dos pacientes que evoluem com insuficiência respiratória aguda. Objetivo: Avaliar o uso de estratégias não invasivas no desfecho de pacientes com insuficiência respiratória aguda ou crônica agudizada por COVID-19. Métodos: Pesquisa de caráter observacional e retrospectivo por meio da coleta de dados em prontuário eletrônico com pacientes submetidos ao uso de cânula nasal de alto fluxo (CNAF) e/ou ventilação mecânica não invasiva (VNI). Resultados: 81 pacientes, sendo 70,4% (57) do sexo masculino, com 56,5 ± 14,6 anos. 49,4% (40) dos indivíduos fizeram uso de CNAF e VNI, 9,9% (8) e 40,7% (33) apenas VNI ou CNAF, respectivamente. O tempo médio de uso da CNAF foi de 4,4 ± 3,7 dias e de VNI foi de 2,7 ± 3,4 dias. Observou-se que 43 (53,1%) dos pacientes pesquisados evoluíram para intubação orotraqueal (IOT) e 40 (49,4%) para óbito. Destes, 22 encontravam-se em IOT. Houve diferença estatística quando comparados idade entre os grupos IOT e não IOT, 60,5 ± 13,9 anos vs 52,1 ± 14,2 anos (p = 0,012), respectivamente. Conclusão: O uso de VNI e/ou CNAF pode ser considerado como alternativa no tratamento de pacientes com COVID-19. Contudo, os diversos fatores intrínsecos e extrínsecos ao paciente ainda contribuem para a alta taxa de IOT e de mortalidade.

COVID-19 entre los fisioterapeutas brasileños: análisis de prevalencia y factores asociados / Covid-19 entre profissionais fisioterapeutas brasileiros: análise de prevalência e fatores associados / Covid-19 among brazilian physiotherapists: analysis of prevalence and associated factors

RESUMO Objetivo: estimar a prevalência de COVID-19 entre os fisioterapeutas brasileiros e seus fatores associados. Método: estudo transversal, segundo inquérito on-line, com a participação de 670 fisioterapeutas de todas as regiões do Brasil. Utilizou-se uma adaptação do método respondent driven sampling ao ambiente virtual para a coleta de dados. Análises bivariadas e de regressão logística múltipla foram utilizadas para identificar associação entre o diagnóstico de COVID-19 e variáveis demográficas e ocupacionais. Consideraram-se variáveis estatisticamente significativas com base em um p<0,05. Resultados: a prevalência de COVID-19 foi de 30% (IC95%: 27,8-32,3). Fisioterapeutas da região Sudeste tiveram menores chances de receber diagnóstico de covid-19. Fisioterapeutas que prestaram assistência em hospital de campanha, que ficaram isolados da família e que têm crianças menores de 12 anos em casa tiveram chances aumentadas de diagnóstico da infecção. Conclusão: questões sociodemográficas e ocupacionais impactam no aumento do diagnóstico de COVID-19 entre profissionais fisioterapeutas, o que enfatiza a necessidade de um sistema de saúde de qualidade e igualitário nas diferentes regiões brasileiras.

RESUMEN Objetivo: evaluar la tasa de prevalencia del COVID-19 en fisioterapeutas de Brasil y analizar sus factores asociados. Método: realizamos un estudio transversal mediante una encuesta on-line, en la que participaron 670 fisioterapeutas de todas las áreas de Brasil. Para la recogida de datos se utilizó una adaptación del método respondent driven sampling al entorno virtual. Se utilizaron análisis bivariados y de regresión logística múltiple para identificar la asociación entre el diagnóstico COVID-19 y variables demográficas y ocupacionales. Las variables se consideraron estadísticamente significativas en función de una p<0,05. Resultados: la prevalencia de COVID-19 fue del 30% (IC 95%: 27,8-32,3). Los fisioterapeutas del sudeste tenían menos probabilidades de ser diagnosticados de COVID-19. Los fisioterapeutas que prestaban asistencia en un hospital de campaña, que estaban aislados de sus familias y que tenían hijos menores de 12 años en casa tenían más probabilidades de que se les diagnosticara la infección. Conclusiones: aspectos sociodemográficos y ocupacionales inciden en el aumento del diagnóstico de COVID-19 entre los fisioterapeutas profesionales, lo que enfatiza la necesidad de un sistema de salud de calidad e igualitario en las diferentes regiones brasileñas.

ABSTRACT Objective: to estimate the prevalence of COVID-19 among Brazilian physiotherapists and its associated factors. Method: cross-sectional study, according to an online survey, with the participation of 670 physiotherapists from all regions of Brazil. An adaptation of the respondent driven sampling method to the virtual environment was used to collect data. Bivariate and multiple logistic regression analyzes were used to identify associations between the diagnosis of COVID-19 and demographic and occupational variables. Variables were considered statistically significant based on p<0.05. Results: the prevalence of COVID-19 was 30% (95%CI: 27.8-32.3). In the Southeast region, physiotherapists were less likely to be diagnosed with COVID-19. Physiotherapists who provided care in a field hospital, who were isolated from their families and who have children under 12 years of age at home had an increased chance of being diagnosed with the infection. Conclusion: sociodemographic and occupational issues impact the increase in COVID-19 diagnoses among physiotherapists, which emphasizes the need for a quality and egalitarian health system in different Brazilian regions.

In silico and in vitro evaluation of imatinib as an inhibitor for SARS-CoV-2.

The rapid geographic expansion of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the infectious agent of Coronavirus Disease 2019 (COVID-19) pandemic, poses an immediate need for potent drugs. Enveloped viruses infect the host cell by cellular membrane fusion, a crucial mechanism required for virus replication. The SARS-CoV-2 spike glycoprotein, due to its primary interaction with the human angiotensin-converting enzyme 2 (ACE2) cell-surface receptor, is considered a potential target for drug development. In this study, around 5,800 molecules were virtually screened using molecular docking. Five molecules were selected for in vitro experiments from those that reported docking scores lower than -6 kcal/mol. Imatinib, a Bcr-Abl tyrosine kinase inhibitor, showed maximum antiviral activity in Vero cells. We further investigated the interaction of imatinib, a compound under clinical trials for the treatment of COVID-19, with SARS-CoV-2 RBD, using in silico methods. Molecular dynamics simulations verified that imatinib interacts with RBD residues that are critical for ACE2 binding. This study also provides significant molecular insights on potential repurposable small-molecule drugs and chemical scaffolds for the development of novel drugs targeting the SARS-CoV-2 spike RBD.Communicated by Ramaswamy H. Sarma.

Mesenchymal Stromal Cells and their EVs as Potential Leads for SARSCoV2 Treatment.

In December 2019, a betacoronavirus was isolated from pneumonia cases in China and rapidly turned into a pandemic of COVID-19. The virus is an enveloped positive-sense ssRNA and causes a severe respiratory syndrome along with a cytokine storm, which is the main cause of most complications. Therefore, treatments that can effectively control the inflammatory reactions are necessary. Mesenchymal Stromal Cells and their EVs are well-known for their immunomodulatory effects, inflammation reduction, and regenerative potentials. These effects are exerted through paracrine secretion of various factors. Their EVs also transport various molecules such as microRNAs to other cells and affect recipient cells' behavior. Scores of research and clinical trials have indicated the therapeutic potential of EVs in various diseases. EVs also seem to be a promising approach for severe COVID-19 treatment. EVs have also been used to develop vaccines since EVs are biocompatible nanoparticles that can be easily isolated and engineered. In this review, we have focused on the use of Mesenchymal Stromal Cells and their EVs for the treatment of COVID-19, their therapeutic capabilities, and vaccine development.

Cardiac Manifestations Associated With Coronavirus Disease: A Case Report

Abstract This study presents a 47-year-old female patient, with a history of diabetes, who contracted SARS-CoV-2 and exhibited cardiovascular complications.

Antiviral effect of lysosomotropic disaccharide trehalose on porcine hemagglutinating encephalomyelitis virus, a highly neurotropic betacoronavirus.

Many members of the genus Betacoronavirus are neurotropic viruses that frequently cause serious harm to humans or animals, including highly neurotropic porcine hemagglutinating encephalomyelitis virus (PHEV). Nevertheless, very few approved treatments exist to combat these viruses. Lysosomotropic trehalose, a widely used, nontoxic, natural disaccharide that can traverse the blood-brain barrier, has been proposed as a potential antiviral agent for use in prevention or treatment of betacoronavirus-associated infections. The purpose of this study was to determine if trehalose could inhibit PHEV infection of cells of a mouse central nervous system-derived neuroblastoma cell line in vitro or brain cells in vivo. Our results demonstrated that treatment of PHEV-infected mouse neuroblastoma cells and mice with trehalose reduced viral replication and that these trehalose antiviral effects were dependent on expression of lysosomal protein progranulin. Collectively, these results indicated that trehalose holds promise as a new antiviral agent for use in controlling neurotropic betacoronavirus infections.

Clinical management of COVID-19: living guideline - 15 September 2022

The WHO COVID-19 Clinical management: living guidance contains the Organization's most up-to-date recommendations for the clinical management of people with COVID-19. Providing guidance that is comprehensive and holistic for the optimal care of COVID-19 patients throughout their entire illness is important. The latest version of this living guideline is available in pdf format (via the 'Download' button) and via an online platform, and is updated regularly as new evidence emerges. No further updates to the previous existing recommendations were made in this latest version. This updated (fifth) version contains 16 new recommendations for the rehabilitation of adults with post COVID-19 condition (see Chapter 24), which includes: strong recommendation that exertional desaturation and cardiac impairment following COVID-19 should be ruled out and managed before consideration of physical exercise training

Individual and Synergistic Anti-Coronavirus Activities of SOCS1/3 Antagonist and Interferon α1 Peptides.

Suppressors of Cytokine Signaling (SOCS) are intracellular proteins that negatively regulate the induction of cytokines. Amongst these, SOCS1 and SOCS3 are particularly involved in inhibition of various interferons. Several viruses have hijacked this regulatory pathway: by inducing SOCS1and 3 early in infection, they suppress the host immune response. Within the cell, SOCS1/3 binds and inhibits tyrosine kinases, such as JAK2 and TYK2. We have developed a cell penetrating peptide from the activation loop of the tyrosine kinase, JAK2 (residues 1001-1013), denoted as pJAK2 that acts as a decoy and suppresses SOCS1 and 3 activity. This peptide thereby protects against several viruses in cell culture and mouse models. Herein, we show that treatment with pJAK2 inhibited the replication and release of the beta coronavirus HuCoV-OC43 and reduced production of the viral RNA, as measured by RT-qPCR, Western blot and by immunohistochemistry. We confirmed induction of SOCS1 and 3 in rhabdomyosarcoma (RD) cells, and this induction was suppressed by pJAK2 peptide. A peptide derived from the C-terminus of IFNα (IFNα-C) also inhibited replication of OC43. Furthermore, IFNα-C plus pJAK2 provided more potent inhibition than either peptide alone. To extend this study to a pandemic beta-coronavirus, we determined that treatment of cells with pJAK2 inhibited replication and release of SARS-CoV-2 in Calu-3 cells. We propose that these peptides offer a new approach to therapy against the rapidly evolving strains of beta-coronaviruses.

Inhibition of Human Respiratory Influenza A Virus and Human Betacoronavirus-1 by the Blend of Double-Standardized Extracts of Aronia melanocarpa (Michx.) Elliot and Sambucus nigra L.

Viral and bacterial diseases are among the greatest concerns of humankind since ancient times. Despite tremendous pharmacological progress, there is still a need to search for new drugs that could treat or support the healing processes. A rich source of bioactive compounds with antiviral potency include plants such as black chokeberry and elderberry. The aim of this study was to assess the in vitro antiviral ability of an originally designed double-standardized blend of extracts from Aronia melanocarpa (Michx.) Elliot and Sambucus nigra L. (EAM-ESN) or separated extracts of A. melanocarpa (EAM) or S. nigra (ESN) against four human respiratory tract viruses: influenza A virus (A/H1N1), betacoronavirus-1 (HCoV-OC43) belonging to the same ß-coronaviruses as the current pandemic SARS-CoV-2, human herpesvirus type 1 (HHV-1), and human adenovirus type 5 (HAdV-5). Antiviral assays (AVAs) were used to evaluate the antiviral activity of the plant extracts in a cell-present environment with extracts tested before, simultaneously, or after viral infection. The virus replication was assessed using the CPE scale or luminescent assay. The EAM-ESN blend strongly inhibited A/H1N1 replication as well as HCoV-OC43, while having a limited effect against HHV-1 and HAdV-5. This activity likely depends mostly on the presence of the extract of S. nigra. However, the EAM-ESN blend possesses more effective inhibitory activity toward virus replication than its constituent extracts. A post-infection mechanism of action of the EAM-ESN make this blend the most relevant for potential drugs and supportive treatments; thus, the EAM-ESN blend might be considered as a natural remedy in mild, seasonal respiratory viral infections.

Targeting of Silver Cations, Silver-Cystine Complexes, Ag Nanoclusters, and Nanoparticles towards SARS-CoV-2 RNA and Recombinant Virion Proteins.

Background: Nanosilver possesses antiviral, antibacterial, anti-inflammatory, anti-angiogenesis, antiplatelet, and anticancer properties. The development of disinfectants, inactivated vaccines, and combined etiotropic and immunomodulation therapy against respiratory viral infections, including COVID-19, remains urgent. Aim: Our goal was to determine the SARS-CoV-2 molecular targets (genomic RNA and the structural virion proteins S and N) for silver-containing nanomaterials. Methods: SARS-CoV-2 gene cloning, purification of S2 and N recombinant proteins, viral RNA isolation from patients' blood samples, reverse transcription with quantitative real-time PCR ((RT)2-PCR), ELISA, and multiplex immunofluorescent analysis with magnetic beads (xMAP) for detection of 17 inflammation markers. Results: Fluorescent Ag nanoclusters (NCs) less than 2 nm with a few recovered silver atoms, citrate coated Ag nanoparticles (NPs) with diameters of 20-120 nm, and nanoconjugates of 50-150 nm consisting of Ag NPs with different protein envelopes were constructed from AgNO3 and analyzed by means of transmission electron microscopy (TEM), atomic force microscopy (AFM), ultraviolet-visible light absorption, and fluorescent spectroscopy. SARS-CoV-2 RNA isolated from COVID-19 patients' blood samples was completely cleaved with the artificial RNase complex compound Li+[Ag+2Cys2-(OH-)2(NH3)2] (Ag-2S), whereas other Ag-containing materials provided partial RNA degradation only. Treatment of the SARS-CoV-2 S2 and N recombinant antigens with AgNO3 and Ag NPs inhibited their binding with specific polyclonal antibodies, as shown by ELISA. Fluorescent Ag NCs with albumin or immunoglobulins, Ag-2S complex, and nanoconjugates of Ag NPs with protein shells had no effect on the interaction between coronavirus recombinant antigens and antibodies. Reduced production of a majority of the 17 inflammation biomarkers after treatment of three human cell lines with nanosilver was demonstrated by xMAP. Conclusion: The antiviral properties of the silver nanomaterials against SARS-CoV-2 coronavirus differed. The small-molecular-weight artificial RNase Ag-2S provided exhaustive RNA destruction but could not bind with the SARS-CoV-2 recombinant antigens. On the contrary, Ag+ ions and Ag NPs interacted with the SARS-CoV-2 recombinant antigens N and S but were less efficient at performing viral RNA cleavage. One should note that SARS-CoV-2 RNA was more stable than MS2 phage RNA. The isolated RNA of both the MS2 phage and SARS-CoV-2 were more degradable than the MS2 phage and coronavirus particles in patients' blood, due to the protection with structural proteins. To reduce the risk of the virus resistance, a combined treatment with Ag-2S and Ag NPs could be used. To prevent cytokine storm during the early stages of respiratory infections with RNA-containing viruses, nanoconjugates of Ag NPs with surface proteins could be recommended.