A Comparative Study of SARS, MERS with COVID-19

Background: B814, now's called Coronavirus first identified by Tyrrell and Bynoe in 1965 from the respiratory tract (embryonic tracheal) of an adult and later on during working on National Institutes of Health Robert Chanock used the term "OC" for same virus strain. After several years researchers reported that coronaviruses were caused disease in rats, mice, chickens, turkeys, calves, dogs, cats, rabbits etc. after effecting the enormous variety of animal, in year 2002-2003 it caused new respiratory disease named severe acute respiratory syndrome, (SARS) in southern China. Objective(s): The main objective of this article is to compare the status of various previous pandemics (i.e., SARS, MERS) with the current COVID-19 pandemic in terms of the life cycle, diagnosis process and prevention Results: On 31st December 2019, the World Health Organization (WHO) office in China received information regarding pneumonia cases of unknown etiology from the Wuhan district in central China. Subsequently, this new disease spread to China, and from there, to the rest of the world. By the end of March 2020, more than 2 million cases were confirmed of this new disease, with over 70000 deaths worldwide. After some time, researchers have identified that this new disease is caused by a novel beta-Coronavirus (virus SARS-CoV-2) and the new disease was named COVID-19. Since then, the Ministry of Health of various countries and WHO have been fighting this health emergency, which has not only affected public health, but also affected various economic sectors. Conclusion(s): The current outbreak SARS-CoV-2 phylogenetically resembled to Bat SARS, which was previously identified in year 2002 and 2012 having low mortality rate than MERS and SARS. However, SARS-CoV-2 and MERS having high virological similarity but both use different receptors to take entry in to the host cell via ACE-2 and DPP-4 respectively. Unfortunately, currently there is no approved treatment available worldwide. Currently, we can hope that together we will recover from this public health emergency very soon.Copyright © 2021 Bentham Science Publishers.

NSP4 and ORF9b of SARS-CoV-2 Induce Pro-Inflammatory Mitochondrial DNA Release in Inner Membrane-Derived Vesicles.

Circulating cell-free mitochondrial DNA (cf-mtDNA) has been found in the plasma of severely ill COVID-19 patients and is now known as a strong predictor of mortality. However, the underlying mechanism of mtDNA release is unexplored. Here, we show a novel mechanism of SARS-CoV-2-mediated pro-inflammatory/pro-apoptotic mtDNA release and a rational therapeutic stem cell-based approach to mitigate these effects. We systematically screened the effects of 29 SARS-CoV-2 proteins on mitochondrial damage and cell death and found that NSP4 and ORF9b caused extensive mitochondrial structural changes, outer membrane macropore formation, and the release of inner membrane vesicles loaded with mtDNA. The macropore-forming ability of NSP4 was mediated through its interaction with BCL2 antagonist/killer (BAK), whereas ORF9b was found to inhibit the anti-apoptotic member of the BCL2 family protein myeloid cell leukemia-1 (MCL1) and induce inner membrane vesicle formation containing mtDNA. Knockdown of BAK and/or overexpression of MCL1 significantly reversed SARS-CoV-2-mediated mitochondrial damage. Therapeutically, we engineered human mesenchymal stem cells (MSCs) with a simultaneous knockdown of BAK and overexpression of MCL1 (MSCshBAK+MCL1) and named these cells IMAT-MSCs (intercellular mitochondrial transfer-assisted therapeutic MSCs). Upon co-culture with SARS-CoV-2-infected or NSP4/ORF9b-transduced airway epithelial cells, IMAT-MSCs displayed functional intercellular mitochondrial transfer (IMT) via tunneling nanotubes (TNTs). The mitochondrial donation by IMAT-MSCs attenuated the pro-inflammatory and pro-apoptotic mtDNA release from co-cultured epithelial cells. Our findings thus provide a new mechanistic basis for SARS-CoV-2-induced cell death and a novel therapeutic approach to engineering MSCs for the treatment of COVID-19.

PC-578-02 JUNCTIONAL TACHYCARDIA

Background: Junctional ectopic tachycardia (JET) is a rare tachyarrhythmia in adults. The precise site of origin within the AV junction is unknown. Objective: N/A Methods: N/A Results: A 71-year-old male presented with dyspnea on exertion and recently diagnosed tachycardia in March 2021. He had a history of diabetes mellitus, obesity, hypertension, obstructive sleep apnea, and COVID-19 in 2020. A 14-day monitor demonstrated 43% supraventricular ectopy SVE burden and short runs of SVT. He presented for an electrophysiology (EP) study. He presented to the EP lab in sinus rhythm with frequent SVE. Multipolar catheters were placed in the His bundle region, right atrium, coronary sinus, and right ventricle. The SVE beats had the same QRS morphology, and an identical HV interval and His-right bundle activation sequence as in sinus rhythm and no retrograde conduction, consistent with premature junctional complexes (PJCs). Occasional short bursts of junctional tachycardia were noted. Isoproterenol was titrated to a maximum dose of 8 mcg/min. No other SVT was inducible with atrial overdrive pacing or programmed stimulation or with isoproterenol infusion. A 6 mm tip cryoablation catheter was advanced to the right atrium to the anatomical location of the slow pathway in the inferior triangle of Koch using an electroanatomic mapping system (EnSite NavX). Signals immediately prior to ablation (Figure 1) were notable for a pre-potential 26 ms prior to the His with PJCs. Cryoablation was performed at this site (Figure 2) with resolution of the PJCs at the onset of the freeze. After thawing, a second freeze was administered. No further PJCs were noted at baseline or with isoproterenol infusion. Conclusion: JET could originate from anywhere within the AV node or proximal His bundle. The application of cryoablation at a typical AV nodal slow pathway location with a preceding pre- potential and immediate obliteration of PJCs suggests that the origin in this case was from this region rather than a true His bundle extrasystole. Identification of pre-potentials to the junctional ectopy can guide safe ablation of this dysrhythmia. [Formula presented] [Formula presented]

A phytochemical-based medication search for the SARS-CoV-2 infection by molecular docking models towards spike glycoproteins and main proteases† † Electronic supplementary information (ESI) available. See DOI: 10.1039/d0ra10458b

Identifying best bioactive phytochemicals from different medicinal plants using molecular docking techniques demonstrates a potential pre-clinical compound discovery against SARS-CoV-2 viral infection. The in silico screening of bioactive phytochemicals with the two druggable targets of SARS-CoV-2 by simple precision/extra precision molecular docking methods was used to compute binding affinity at its active sites. phyllaemblicin and cinnamtannin class of phytocompounds showed a better binding affinity range (−9.0 to −8.0 kcal mol−1) towards both these SARS-CoV-2 targets;the corresponding active site residues in the spike protein were predicted as: Y453, Q496, Q498, N501, Y449, Q493, G496, T500, Y505, L455, Q493, and K417;and Mpro: Q189, H164, H163, P168, H41, L167, Q192, M165, C145, Y54, M49, and Q189. Molecular dynamics simulation further established the structural and energetic stability of protein–phytocompound complexes and their interactions with their key residues supporting the molecular docking analysis. Protein–protein docking using ZDOCK and Prodigy server predicted the binding pose and affinity (−13.8 kcal mol−1) of the spike glycoprotein towards the human ACE2 enzyme and also showed significant structural variations in the ACE2 recognition site upon the binding of phyllaemblicin C compound at their binding interface. The phyllaemblicin and cinnamtannin class of phytochemicals can be potential inhibitors of both the spike and Mpro proteins of SARS-CoV-2;furthermore, its pharmacology and clinical optimization would lead towards novel COVID-19 small-molecule therapy. Identifying best bioactive phytochemicals from different medicinal plants using molecular docking techniques demonstrates a potential pre-clinical compound discovery against SARS-CoV-2 viral infection.

A phytochemical-based medication search for the SARS-CoV-2 infection by molecular docking models towards spike glycoproteins and main proteases.

Identifying best bioactive phytochemicals from different medicinal plants using molecular docking techniques demonstrates a potential pre-clinical compound discovery against SARS-CoV-2 viral infection. The in silico screening of bioactive phytochemicals with the two druggable targets of SARS-CoV-2 by simple precision/extra precision molecular docking methods was used to compute binding affinity at its active sites. phyllaemblicin and cinnamtannin class of phytocompounds showed a better binding affinity range (-9.0 to -8.0 kcal mol-1) towards both these SARS-CoV-2 targets; the corresponding active site residues in the spike protein were predicted as: Y453, Q496, Q498, N501, Y449, Q493, G496, T500, Y505, L455, Q493, and K417; and Mpro: Q189, H164, H163, P168, H41, L167, Q192, M165, C145, Y54, M49, and Q189. Molecular dynamics simulation further established the structural and energetic stability of protein-phytocompound complexes and their interactions with their key residues supporting the molecular docking analysis. Protein-protein docking using ZDOCK and Prodigy server predicted the binding pose and affinity (-13.8 kcal mol-1) of the spike glycoprotein towards the human ACE2 enzyme and also showed significant structural variations in the ACE2 recognition site upon the binding of phyllaemblicin C compound at their binding interface. The phyllaemblicin and cinnamtannin class of phytochemicals can be potential inhibitors of both the spike and Mpro proteins of SARS-CoV-2; furthermore, its pharmacology and clinical optimization would lead towards novel COVID-19 small-molecule therapy.

A Saliva-Based RNA Extraction-Free Workflow Integrated With Cas13a for SARS-CoV-2 Detection.

A major bottleneck in scaling-up COVID-19 testing is the need for sophisticated instruments and well-trained healthcare professionals, which are already overwhelmed due to the pandemic. Moreover, the high-sensitive SARS-CoV-2 diagnostics are contingent on an RNA extraction step, which, in turn, is restricted by constraints in the supply chain. Here, we present CASSPIT (Cas13 Assisted Saliva-based & Smartphone Integrated Testing), which will allow direct use of saliva samples without the need for an extra RNA extraction step for SARS-CoV-2 detection. CASSPIT utilizes CRISPR-Cas13a based SARS-CoV-2 RNA detection, and lateral-flow assay (LFA) readout of the test results. The sample preparation workflow includes an optimized chemical treatment and heat inactivation method, which, when applied to COVID-19 clinical samples, showed a 97% positive agreement with the RNA extraction method. With CASSPIT, LFA based visual limit of detection (LoD) for a given SARS-CoV-2 RNA spiked into the saliva samples was ~200 copies; image analysis-based quantification further improved the analytical sensitivity to ~100 copies. Upon validation of clinical sensitivity on RNA extraction-free saliva samples (n = 76), a 98% agreement between the lateral-flow readout and RT-qPCR data was found (Ct<35). To enable user-friendly test results with provision for data storage and online consultation, we subsequently integrated lateral-flow strips with a smartphone application. We believe CASSPIT will eliminate our reliance on RT-qPCR by providing comparable sensitivity and will be a step toward establishing nucleic acid-based point-of-care (POC) testing for COVID-19.

COVID-19: The Emerging Immunopathological Determinants for Recovery or Death.

Hyperactivation of the host immune system during infection by SARS-CoV-2 is the leading cause of death in COVID-19 patients. It is also evident that patients who develop mild/moderate symptoms and successfully recover display functional and well-regulated immune response. Whereas a delayed initial interferon response is associated with severe disease outcome and can be the tipping point towards immunopathological deterioration, often preceding death in COVID-19 patients. Further, adaptive immune response during COVID-19 is heterogeneous and poorly understood. At the same time, some studies suggest activated T and B cell response in severe and critically ill patients and the presence of SARS-CoV2-specific antibodies. Thus, understanding this problem and the underlying molecular pathways implicated in host immune function/dysfunction is imperative to devise effective therapeutic interventions. In this comprehensive review, we discuss the emerging immunopathological determinants and the mechanism of virus evasion by the host cell immune system. Using the knowledge gained from previous respiratory viruses and the emerging clinical and molecular findings on SARS-CoV-2, we have tried to provide a holistic understanding of the host innate and adaptive immune response that may determine disease outcome. Considering the critical role of the adaptive immune system during the viral clearance, we have presented the molecular insights of the plausible mechanisms involved in impaired T cell function/dysfunction during various stages of COVID-19.

A saliva-based RNA extraction-free workflow integrated with Cas13a for SARS-CoV-2 detection (preprint)

A major bottleneck in scaling-up COVID-19 testing is the need for sophisticated instruments and well-trained healthcare professionals, which are already overwhelmed due to the pandemic. Moreover, the high-sensitive SARS-CoV-2 diagnostics are contingent on an RNA extraction step, which, in turn, is restricted by constraints in the supply chain. Here, we present CASSPIT (Cas13 Assisted Saliva-based & Smartphone Integrated Testing), which will allow direct use of saliva samples without the need for an extra RNA extraction step for SARS-CoV-2 detection. CASSPIT utilizes CRISPR-Cas13a based SARS-CoV-2 RNA detection, and lateral-flow assay (LFA) readout of the test results. The sample preparation workflow includes an optimized chemical treatment and heat inactivation method, which, when applied to COVID-19 clinical samples, showed a 97% positive agreement with the RNA extraction method. With CASSPIT, LFA based visual limit of detection (LoD) for a given SARS-CoV-2 RNA spiked into the saliva samples was [~]200 copies; image analysis-based quantification further improved the analytical sensitivity to [~]100 copies. Upon validation of clinical sensitivity on RNA extraction-free saliva samples (n=76), a 98% agreement between the lateral-flow readout and RT-qPCR data was found (Ct<35). To enable user-friendly test results with provision for data storage and online consultation, we subsequently integrated lateral-flow strips with a smartphone application. We believe CASSPIT will eliminate our reliance on RT-qPCR by providing comparable sensitivity and will be a step toward establishing nucleic acid-based point-of-care (POC) testing for COVID-19.