Iron: Innocent bystander or vicious culprit in COVID-19 pathogenesis?

Abstract: The coronavirus 2 (SARS-CoV-2) pandemic is viciously spreading through the continents with rapidly increasing mortality rates. Current management of COVID-19 is based on the premise that respiratory failure is the leading cause of mortality. However, mounting evidence links accelerated pathogenesis in gravely ill COVID-19 patients to a hyper-inflammatory state involving a cytokine storm. Several components of the heightened inflammatory state were addressed as therapeutic targets. Another key component of the heightened inflammatory state is hyper-ferritinemia which reportedly identifies patients with increased mortality risk. In spite of its strong association with mortality, it is not yet clear if hyper-ferritinemia in COVID-19 patients is merely a systemic marker of disease progression, or a key modulator in disease pathogenesis. Here we address implications of a possible role for hyper-ferritinemia, and altered iron homeostasis in COVID-19 pathogenesis, and potential therapeutic targets in this regard.


Hepatic complications of COVID-19 and its treatment

Abstract: Coronavirus disease 2019 (COVID-19) is highly contagious and has a variety of clinical manifestations, it can affect a number of other organs in addition to the lungs, and liver injury may occur. Severe acute respiratory syndrome coronavirus 2 can cause liver injury through systemic inflammatory response syndrome, cytokine storms, ischemia-reperfusion injury, side effects of treatment drugs, and underlying liver disease and can attack liver cells directly via angiotensin-converting enzyme 2. Clinical studies have found that liver injury in COVID-19 patients mainly manifests as abnormal liver biochemical indicators, but there have been no reports of liver failure caused by this disease. The number of COVID-19 patients with liver injury is increasing, and the incidence of liver injury in COVID-19 patients with severe disease are higher than in patients with mild disease. Liver injury may be a risk factor, which worsens in patients with COVID-19, and hence it is necessary to pay attention to the occurrence of liver injury in the diagnosis and treatment of COVID-19.


Heightened Innate Immune Responses in the Respiratory Tract of COVID-19 Patients

Abstract: The outbreaks of 2019 novel coronavirus disease (COVID-19) caused by SARS-CoV-2 infection have posed a severe threat to global public health. It is unclear how the human immune system responds to this infection. Here, we used metatranscriptomic sequencing to profile immune signatures in the bronchoalveolar lavage fluid of eight COVID-19 cases. The expression of proinflammatory genes, especially chemokines, was markedly elevated in COVID-19 cases compared to community-acquired pneumonia patients and healthy controls, suggesting that SARS-CoV-2 infection causes hypercytokinemia. Compared to SARS-CoV, which is thought to induce inadequate interferon (IFN) responses, SARS-CoV-2 robustly triggered expression of numerous IFN-stimulated genes (ISGs). These ISGs exhibit immunopathogenic potential, with overrepresentation of genes involved in inflammation. The transcriptome data was also used to estimate immune cell populations, revealing increases in activated dendritic cells and neutrophils. Collectively, these host responses to SARS-CoV-2 infection could further our understanding of disease pathogenesis and point toward antiviral strategies.


Novel COronaVirus Disease 2019 (COVID-19) epidemic: What are the risks for systemic sclerosis patients?


Weather the Cytokine Storm: A Report of Successful Management of a Colon Cancer Survivor and a Critically Ill Patient with COVID-19

Abstract: Patients with novel corona virus infection (COVID-19) can develop acute respiratory failure secondary to acute respiratory distress syndrome. Cytokine storm is suggested as one of underlying mechanisms for the rapid clinical decline. Immunocompromised patients and cancer patients are at particular risk for poor outcomes due to COVID-19 infection. This case report describes the presentation and clinical course of a cancer survivor who became critically ill and required mechanical ventilation. The patient was treated with hydroxychloroquine, azithromycin, and ceftriaxone; however, he remained febrile, hypoxemic, continued to require full mechanical ventilator support and his chest X-ray showed increased bilateral infiltrates. The patient was treated with tocilizumab, after which he improved and was successfully extubated. This report illustrates a possible role of tocilizumab in management of cytokine storm in critically ill patients with COVID-19 infection.


SARS-CoV-2: A Storm is Raging.

Abstract: The pandemic coronavirus infectious disease (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is rapidly spreading across the globe. In this issue of the JCI, Chen and colleagues compared the clinical and immunologic characteristics between moderate versus severe COVID-19. The authors found that respiratory distress on admission is associated with unfavorable outcomes. Increased cytokine levels (IL-6, IL-10 and TNFα), lymphopenia (in CD4+ and CD8+ T cells), and decreased IFNγ expression in CD4+ T cells are associated with severe COVID-19. Overall, this study characterized the cytokine storm in severe COVID-19 and provides insights into immune therapeutics and vaccine design.


Extracorporeal Hemoadsorption: An Option for COVID-19 Associated Cytokine Storm Syndrome.


Cytokine storm and the prospects for immunotherapy with COVID-19.

Abstract: Knowledge about the pathobiology of SARS-COV-2 as it interacts with immune defenses is limited. SARS-COV-2 is spread by droplets that come into contact with mucous membranes. COVID-19 is characterized by 3 stages: asymptomatic-paucisymptomatic incubation, nonsevere symptomatic illness for 80% of those infected, and severe respiratory illness. A syndrome characterized by hypercytokinemic inflammation referred to as a "cytokine storm" can occur in patients with advanced disease. Effective antiviral agents that can prevent viral infection in exposed individuals are needed.


Protective role of ACE2 and its downregulation in SARS-CoV-2 infection leading to Macrophage Activation Syndrome: Therapeutic implications.

Abstract: In light of the outbreak of the 2019 novel coronavirus disease (COVID-19), the international scientific community has joined forces to develop effective treatment strategies. The Angiotensin-Converting Enzyme (ACE) 2, is an essential receptor for cell fusion and engulfs the SARS coronavirus infections. ACE2 plays an important physiological role, practically in all the organs and systems. Also, ACE2 exerts protective functions in various models of pathologies with acute and chronic inflammation. While ACE2 downregulation by SARS-CoV-2 spike protein leads to an overactivation of Angiotensin (Ang) II/AT1R axis and the deleterious effects of Ang II may explain the multiorgan dysfunction seen in patients. Specifically, the role of Ang II leading to the appearance of Macrophage Activation Syndrome (MAS) and the cytokine storm in COVID-19 is discussed below. In this review, we summarized the latest research progress in the strategies of treatments that mainly focus on reducing the Ang II-induced deleterious effects rather than attenuating the virus replication.


Treat 2019 novel coronavirus (COVID-19) with IL-6 inhibitor: Are we already that far?

Abstract: The world is in the midst of the coronavirus disease 2019 (COVID-19) pandemic. Interleukin 6 (IL-6) inhibitor (tocilizumab) had been suggested for the treatment of acute respiratory distress syndrome (ARDS) patients based on the concept of "cytokine storm" in COVID-19. However, we still lack reliable studies to verify "cytokine storm" in COVID-19 pneumonia. Furthermore, IL-6 inhibitor has potential hazards of inducing infectious diseases. The efficacy of IL-6 monoclonal antibody-directed therapy remains to be fully evaluated.


Potential for jaktinib hydrochloride to treat cytokine storms in patients with COVID-19.

Abstract: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has become a serious public health threat to the whole world, and the number of infected is still rising dramatically at this moment. Several studies have confirmed that cytokine storms play a critical role in causing a case to worsen from mild to severe or critical. The current treatment for cytokine storms is limited, so the international medical community is focusing on a specific and effective remedy. Jaktinib hydrochloride is a broad spectrum JAK inhibitor. It can inhibit cytokine-induced immune activation by multiple mechanisms and also slow viral proliferation by inhibiting AAK1 without causing unacceptable toxicity. Jaktinib hydrochloride has great potential for the treatment of patients with coronavirus disease 2019 (COVID-19).


Brief annotations on cytokine release syndrome and interleukin-6 therapeutic blockage in SARS-CoV-2/COVID-19.


Preventing cytokine storm syndrome in COVID-19 using α-1 adrenergic receptor antagonists.

Abstract: Medications that target catecholamine-associated inflammation may prevent cytokine storm syndrome associated with COVID-19 and other diseases.


COVID-19 and the Heart.

Abstract: Infection with the novel coronavirus, SARS-CoV-2, produces a clinical syndrome known as COVID-19. When severe, COVID-19 is a systemic illness characterized by hyperinflammation, cytokine storm and elevations of cardiac injury biomarkers. Here we review what is known about the pathophysiology of COVID-19, its cardiovascular manifestations, and emerging therapeutic prospects. In this rapidly moving field, this review was comprehensive as of April 3, 2020.


Insights into the Use of C-Reactive Protein as a Diagnostic Index of Disease Severity in COVID-19 Infections.

Abstract: Approximately 20% of patients infected with SARS-CoV-2 (COVID-19) develop potentially life-threatening pathologies involving hyperinflammation, cytokine storm, septic shock complications, coagulation dysfunction, and multiple organ failure. Blood levels of the prototypic acute phase reactant, C-reactive protein (CRP), which is hepatically synthesized and released in response to interleukin-6 stimulation, is markedly elevated in patients with COVID-19. Markedly high CRP levels correlate with poor prognosis for survival. Insights into CRP structure-function relationships have uncovered both pro- and anti-inflammatory isoforms that may be used to monitor the extent of tissue damage associated with COVID-19 pathologies and prognoses. Herein, rationale is given for interpretation of CRP blood levels as a simple, rapid, and cost-effective way to assess disease severity and help guide therapeutic options in COVID-19 patients.


Tocilizumab in the treatment of a critical COVID-19 patient: a case report.

Abstract: In December 2019, Coronavirus disease 2019 (COVID-19) emerged in Wuhan and rapidly spread throughout China and the rest of the world. COVID-19 is currently a global pandemic. There are cytokine storms in severe COVID-19 patients. Interleukin-6 plays an important role in cytokine storm. Tocilizumab is a blocker of interleukin-6 receptor, which is likely to become an effective drug for patients with severe COVID-19. Here, we reported a case in which tocilizumab was effective for a critical COVID-19 patient.


Network pharmacological analysis and mechanism prediction of Xiaochaihu Decoction in treatment of COVID-19 with syndrome of pathogenic heat lingering in lung and obstructive cardinalate/ 小柴胡汤治疗早期新型冠状病毒肺炎（COVID-19）邪热郁肺、枢机不利证功效网络分析与机制预测

Abstract: Objective: To study the efficacy network and potential mechanism of Xiaochaihu Decoction (XCHD) in the treatment of coronavirus disease 2019 (COVID-19) with syndrome of pathogenic heat lingering in the lung and obstructive cardinalat, and analyze the active ingredients of XCHD with anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) efficacy. Methods: The correspondence between COVID-19 and XCHD was analyzed by literature consulting. Based on network pharmacology, Cytoscape 3.6.0 and other software were then used to construct XCHD efficacy network of “Chinese medicine prescription-active ingredient-key target” for pneumonia and immune regulation, in order to confirm anti-SARS-CoV-2 active ingredients in the prescription. Some softwares were used to analyze XCHD for COVID-19 treatment in multiple aspects. Results: A total of 48 active ingredients with potential anti-SARS-CoV-2 effect in herbs were collected; 140 active ingredients in XCHD for pneumonia treatment and immune regulation were analyzed, of which 12 ingredients had direct anti-SARS-CoV-2 activity including baicalein, formononetin, quercetin, etc. The active ingredients in XCHD exerted efficacy for pneumonia treatment and immunoregulation through 95 key targets such as IL-6, NOS2, and ESR1, involving multiple pathways such as the TNF signaling pathway, IL-17 signaling pathway, and influenza A. Analysis of gene co-expression and PPI interaction analysis found that ACE2 only co-expressed with NOS2 in the above targets, and also interacted with only five targets in the PPI interaction network. It is speculated that the ACE2 target only plays an important role when SARS-CoV-2 invaded the human body, and had little effect in the treatment of pneumonia after viral infection. Conclusion: The active ingredients in XCHD play a role in treating COVID-19 by inhibiting SARS-CoV-2 activity, blocking the SARS-CoV-2 invasion pathway, inhibiting cytokine storm, and regulating immunity. It is worth noting that drugs designed for the ACE2 target can block virus invasion, but may not be effective for diseases such as alveolar inflammation, Therefore, this study also provides a multi-target and multi-directional space for XCHD for early COVID-19 treatment. In addition, when XCHD is used in the early treatment of COVID-19, we should pay attention to the precise use of drugs based on syndrome accurate identification, one is to avoid adverse reactions, the other is to avoid cytokine damage caused by re-crown disease.


The successful use of therapeutic plasma exchange for severe COVID-19 acute respiratory distress syndrome with multiple organ failure

Abstract: The COVID-19 pandemic has brought about an urgent need for effective treatment, while conserving vital resources such as intensive care unit beds and ventilators Antivirals, convalescent plasma, and biologics have been used with mixed results The profound "cytokine storm" induced endotheliopathy and microthrombotic disease in patients with COVID-19 may lead to acute respiratory distress syndrome, sepsis, and multi-organ failure We present a case of SARS-COV2 pneumonia with septic shock and multi-organ failure that demonstrated significant clinical improvement after therapeutic plasma exchange A 65-year-old female with multiple comorbidities presented with progressive dyspnea and dry cough She was found to be COVID-19 positive with pneumonia, and developed progressive hypoxemia and shock requiring vasopressors, cardioversion, and non-invasive positive pressure ventilation Given her worsening sepsis with multi-organ failure, she underwent therapeutic plasma exchange with rapid clinical improvement Her case supports the theory that plasma exchange may help abate the "cytokine storm" induced endotheliopathy and microthrombosis associated with COVID-19 Further studies are needed to identify markers of this pathway and the potential role of plasma exchange in these critically ill patients Copyright © The Author(s) 2020


Multiple expression assessments of ACE2 and TMPRSS2 SARS-CoV-2 entry molecules in the urinary tract and their associations with clinical manifestations of COVID-19

Abstract: Background Since December 2019, the novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), first spread quickly in Wuhan, China, then globally. From previously published evidence, ACE2 and TMPRSS2, are both pivotal entry molecules that enable cellular infection by SARS-CoV-2. Meanwhile, increased expression of pro-inflammatory cytokines, or a “cytokine storm,” is associated with multiple organ dysfunction syndrome that is often observed in critically ill patients. Methods We investigated the expression pattern of ACE2 and TMPRSS2 in major organs in the human body, especially under specific disease conditions. Multiple sequence alignment of ACE2 in different species was used to explain animal susceptibility. Moreover, the cell-specific expression patterns of ACE2 and cytokine receptors in the urinary tract were assessed using single-cell RNA sequencing (scRNA-seq). Additional biological relevance was determined through Gene Set Enrichment Analysis (GSEA) using an ACE2 specific signature. Results Our results revealed that ACE2 and TMPRSS2 were highly expressed in genitourinary organs. ACE2 was highly and significantly expressed in the kidney among individuals with chronic kidney diseases or diabetic nephropathy. In single cells, ACE2 was primarily enriched in gametocytes in the testis, and renal proximal tubules. The receptors for pro-inflammatory cytokines, especially IL6ST, were remarkably concentrated in endothelial cells, macrophages, and spermatogonial stem cells in the testis, and renal endothelial cells, which suggested the occurrence of alternative damaging mechanisms via autoimmune attacks. Conclusions This study provided new insights into the pathogenicity mechanisms of SARS-CoV-2 that underlie the clinical manifestations observed in the human testis and kidney. These observations might substantially facilitate the development of effective treatments for this rapidly spreading disease.


Comparative Survival Analysis of Immunomodulatory Therapy for COVID-19 'Cytokine Storm': A Retrospective Observational Cohort Study

Abstract: Background: Cytokine storm is a marker of COVID-19 illness severity and increased mortality. Immunomodulatory treatments have been repurposed to improve mortality outcomes. Methods: We conducted a retrospective analysis of electronic health records across the Northwell Health system. COVID-19 patients hospitalized between March 1, 2020 and April 15, 2020, were included. Cytokine storm was defined by inflammatory markers: ferritin >700ng/mL, C-reactive protein >30mg/dL, or lactate dehydrogenase >300U/L. Patients were subdivided into six groups -no immunomodulatory treatment (standard of care) and five groups that received either corticosteroids, anti-interleukin 6 (IL-6) antibody (tocilizumab) or anti-IL-1 therapy (anakinra) alone or in combination with corticosteroids. The primary outcome was hospital mortality. Results: There were 3,098 patients who met inclusion criteria. The most common comorbidities were hypertension (40-56%), diabetes (32-43%) and cardiovascular disease (2-15%). Patients most frequently met criteria with high lactate dehydrogenase (74.8%) alone, or in combination, followed by ferritin (71.4%) and C-reactive protein (9.4%). More than 80% of patients had an elevated D-dimer. Patients treated with a combination of tocilizumab and corticosteroids (Hazard Ratio [HR]: 0.459, 95% Confidence Interval [CI]: 0.295-0.714; p<0.0001) or corticosteroids alone (HR: 0.696, 95% CI: 0.512-0.946; p=0.01) had improved hospital survival compared to standard of care. Corticosteroids and tocilizumab was associated with increased survival when compared to corticosteroids and anakinra (HR: 0.612, 95% CI: 0.391-0.958; p-value=0.02). Conclusions: When compared to standard of care, corticosteroid and tocilizumab used in combination, or corticosteroids alone, was associated with reduced hospital mortality for patients with COVID-19 cytokine storm.


Dual-Histamine Blockade with Cetirizine - Famotidine Reduces Pulmonary Symptoms in COVID-19 Patients

Abstract: Background: The COVID-19 pandemic due to SARS-CoV-2 infection can produce Acute Respiratory Distress Syndrome as a result of a pulmonary cytokine storm. Antihistamines are safe and effective treatments for reducing inflammation and cytokine release. Combinations of Histamine-1 and Histamine-2 receptor antagonists have been effective in urticaria, and might reduce the histamine-mediated pulmonary cytokine storm in COVID-19. Can a combination of Histamine-1 and Histamine-2 blockers improve COVID-19 inpatient outcomes? Methods: A physician-sponsored cohort study of cetirizine and famotidine was performed in hospitalized patients with severe to critical pulmonary symptoms. Pulmonologists led the inpatient care in a single medical center of 110 high-acuity patients that were treated with cetirizine 10 mg and famotidine 20 mg b.i.d. plus standard-of-care. Results: Of all patients, including those with Do Not Resuscitate directives, receiving the dual-histamine blockade for at least 48 hours, the combination drug treatment resulted in a 16.4% rate of intubation, a 7.3% rate of intubation after a minimum of 48 hours of treatment, a 15.5% rate of inpatient mortality, and 11.0 days duration of hospitalization. The drug combination exhibited reductions in symptom progression when compared to published reports of COVID-19 patients. Concomitant medications were assessed and hydroxychloroquine was correlated with worse outcomes. Conclusions: This physician-sponsored cohort study of cetirizine and famotidine provides proof-of-concept of a new safe and effective method to reduce the progression in symptom severity, presumably by minimizing the histamine-mediated cytokine storm. Further clinical studies in COVID-19 are warranted of the repurposed off-label combination of two historically-safe histamine blockers.


Tocilizumab in Severe COVID-19 Pneumonia and Concomitant Cytokine Release Syndrome

Abstract: Younger patients with COVID-19 may experience an exaggerated immune response to SARS-CoV-2 infection and develop cytokine release syndrome (CRS), which may be life threatening There is no proven antiviral therapy for COVID-19 so far, but profound immunosuppression has recently been suggested as a treatment for COVID-19-associated CRS We present a case of life-threatening CRS caused by COVID-19 infection with a favourable response to immunosuppressive therapy with tocilizumab (TCZ) The rapid clinical and biochemical improvement following TCZ administration suggests that treatment with immunotherapy can be life-saving in selected patients with COVID-19-induced CRS LEARNING POINTS: Cytokine release syndrome may cause sudden and potentially life-threatening clinical deterioration in COVID-19 pneumonia, particularly in younger patients Immunosuppressive therapy may provide important additional therapeutic benefit in these patients Tocilizumab, a specific IL-6 inhibitor, led to dramatic clinical improvement in a young patient with severe COVID-19-associated cytokine release syndrome


Successful recovery of severe COVID-19 with cytokine storm treating with extracorporeal blood purification

Abstract: COVID-19 associated cytokine storm could induce ARDS rapidly and the patients would require the support of mechanic ventilation. However, the prognosis was not that optimistic. The outcome might be changed if the intervention of EBP was performed timely. We present a case of severe SARS-CoV-2 infection who recovered from cytokine storm.


SARS-CoV-2: a storm is raging

Abstract: The pandemic coronavirus infectious disease (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is rapidly spreading across the globe. In this issue of the JCI, Chen and colleagues compared the clinical and immunological characteristics between moderate and severe COVID-19. The authors found that respiratory distress on admission is associated with unfavorable outcomes. Increased cytokine levels (IL-6, IL-10, and TNF-α), lymphopenia (in CD4+ and CD8+ T cells), and decreased IFN-Î³ expression in CD4+ T cells are associated with severe COVID-19. Overall, this study characterized the cytokine storm in severe COVID-19 and provides insights into immune therapeutics and vaccine design.


COVID-19: Consider IL-6 receptor antagonist for the therapy of cytokine storm syndrome in SARS-CoV-2 infected patients

Abstract: COVID-19 leads to mortality of several patients and the cytokine storm is reportedly critical in the patients. To reduce the cytokine storm, we would like to propose the interleukin (IL) 6 receptor (IL-6R) antagonist therapy for the COVID-19 patients. Two humanized monoclonal antibodies are in clinical trial following IL-6R antagonist therapies namely tocilizumab and sarilumab. However, researchers and physicians should look for more IL-6R antagonists for the therapy of cytokine storm syndrome severe acute respiratory syndrome coronavirus 2 infected persons to enhance the therapeutic options for cytokine storm.


Cytokine Release Syndrome (CRS) and Nicotine in COVID-19 Patients: Trying to Calm the Storm

Abstract: SARS-CoV-2 is a new coronavirus that has caused a worldwide pandemic. It causes severe acute respiratory syndrome (COVID-19), which is fatal in many cases, and is characterized by a cytokine release syndrome (CRS). Great efforts are currently being made to block the signal transduction pathway of pro-inflammatory cytokines in order to control this "cytokine storm" and rescue severely affected patients. Consequently, possible treatments for cytokine-mediated hyperinflammation, preferably within approved safe therapies, are urgently being researched to reduce rising mortality. One approach to inhibit proinflammatory cytokine release is to activate the cholinergic anti-inflammatory pathway through nicotinic acetylcholine receptors (α7nAchR). Nicotine, an exogenous α7nAchR agonist, is clinically used in ulcerative colitis to counteract inflammation. We have found epidemiological evidence, based on recent clinical SARS-CoV-2 studies in China, that suggest that smokers are statistically less likely to be hospitalized. In conclusion, our hypothesis proposes that nicotine could constitute a novel potential CRS therapy in severe SARS-CoV-2 patients.


Therapeutic blockade of granulocyte macrophage colony-stimulating factor in COVID-19-associated hyperinflammation: challenges and opportunities

Abstract: The COVID-19 pandemic is a global public health crisis, with considerable mortality and morbidity exerting pressure on health-care resources, including critical care. An excessive host inflammatory response in a subgroup of patients with severe COVID-19 might contribute to the development of acute respiratory distress syndrome (ARDS) and multiorgan failure. Timely therapeutic intervention with immunomodulation in patients with hyperinflammation could prevent disease progression to ARDS and obviate the need for invasive ventilation. Granulocyte macrophage colony-stimulating factor (GM-CSF) is an immunoregulatory cytokine with a pivotal role in initiation and perpetuation of inflammatory diseases. GM-CSF could link T-cell-driven acute pulmonary inflammation with an autocrine, self-amplifying cytokine loop leading to monocyte and macrophage activation. This axis has been targeted in cytokine storm syndromes and chronic inflammatory disorders. Here, we consider the scientific rationale for therapeutic targeting of GM-CSF in COVID-19-associated hyperinflammation. Since GM-CSF also has a key role in homoeostasis and host defence, we discuss potential risks associated with inhibition of GM-CSF in the context of viral infection and the challenges of doing clinical trials in this setting, highlighting in particular the need for a patient risk-stratification algorithm.


Understanding the COVID-19 coagulopathy spectrum


A Putative Role of de-Mono-ADP-Ribosylation of STAT1 by the SARS-CoV-2 Nsp3 Protein in the Cytokine Storm Syndrome of COVID-19

Abstract: As more cases of COVID-19 are studied and treated worldwide, it had become apparent that the lethal and most severe cases of pneumonia are due to an out-of-control inflammatory response to the SARS-CoV-2 virus. I explored the putative causes of this specific feature through a detailed genomic comparison with the closest SARS-CoV-2 relatives isolated from bats, as well as previous coronavirus strains responsible for the previous epidemics (SARS-CoV and MERS-CoV). The high variability region of the nsp3 protein was confirmed to exhibit the most variations between closest strains. It was then studied in the context of physiological and molecular data available in the literature. A number of convergent findings suggest de-mono-ADP-ribosylation (de-MARylation) of STAT1 by the SARS-CoV-2 nsp3 as a putative cause of the cytokine storm observed in the most severe cases of COVID-19. This may suggest new therapeutic approaches and help in designing assays to predict the virulence of naturally circulating SARS-like animal coronaviruses.


Colchicina: una herramienta terapéutica potencial frente a COVID-19. Experiencia en 5 pacientes./ [Colchicine: a potential therapeutic tool against COVID-19. Experience of 5 patients]

Abstract: COVID-19 is a newly emerged disease that has become a global public health challenge. Due to a lack of knowledge about the virus, a significant number of potential targets for using a particular drug have been proposed. Five cases with a clinical history of biopolymers in the gluteal region that developed iatrogenic allogenosis (IA) are presented here. The 5 cases were put under colchicine treatment for IA crisis and had non-specific symptoms (headache, cough without dyspnoea, and arthralgias) with a positive SARS-CoV-2 test. Their close contacts had mild to severe symptoms and three of them died. In the SARS-CoV-2 infection different inflammatory pathways are altered where colchicine reduces cytokine levels as well as the activation of macrophages, neutrophils, and the inflammasome. The possible mechanisms that colchicine may use to prevent acute respiratory distress syndrome (ARDS) in patients with COVID-19 infection are also reviewed in this article.


Tratamiento y evolución del síndrome de tormenta de citoquinas asociados a infección por SARS-CoV-2 en pacientes octogenarios./ [Evolution and treatment of storm cytoquine syndrome associated to SARS-CoV-2 infection among octogenarians]

Abstract: INTRODUCTION: Cytokine storm syndrome (CTS) is a serious complication of patients with SARS-CoV-2 infection. Treatment and evolution in octogenarians are not well defined. Our objective is to describe its clinical characteristics, the treatments and its clinical evolution. PATIENTS AND METHOD: Retrospective observational study of consecutive patients admitted in the period between March 23 and April 12, 2020 with confirmed SARS-CoV-2 infection, with pneumonia by radiological study or chest tomography, whith STC criteria and who received treatment. We classified patients as those who received only glucocorticoid (GC) pulses, or GC and tocilizumab pulses. We determined serum levels of ferritin, CRP and D-dimers. The final variable was survival. RESULTS: 21 patients, (80-88 years). The mean ferritin was 1056 microg/L (317-3,553), CRP 115.8mg/dL (22-306) and D-dimers 2.9m/L (0.45-17.5). All patients received GC pulses and in 2 cases simultaneously tocilizumab. The mean follow-up time was 13.7 days (8-21). The overall mortality was 38.1% (8/21 patients). The 2 patients who received tocilizumab died. The deceased had significantly higher levels of ferritin (1,254 vs. 925microg/L; P=.045) and CRP (197.6 vs. 76mg / dL; P=.007). At the end of the follow-up, a decrease in the biochemical parameters was observed with ferritin of 727microg/L, CRP of 27mg/dl and D-dimers of 1.18mg/L. In 13/21 patients (61.9%), the CTS was controlled without the need to add other treatments. CONCLUSIONS: STC mortality from SARS-CoV-2 is high despite treatment. A greater inflammatory response was associated with a higher mortality. Although it seems that the early use of GC pulses could control it, and the use of other treatments such as tocilizumab shouldo be, with the study design and its limitations, this conclusion cannot be stablished.


Cytokine storm and the prospects for immunotherapy with COVID-19

Abstract: Knowledge about the pathobiology of SARS-CoV-2 as it interacts with immune defenses is limited. SARS-CoV-2 is spread by droplets that come into contact with mucous membranes. COVID-19 is characterized by 2 or 3 stages: most patients who recover experience 2 stages of illness commencing with an asymptomatic or paucisymptomatic incubation period, followed by a nonsevere symptomatic illness lasting for several weeks, occurring in about 80% of those infected. In the remainder, a third phase marked by a severe respiratory illness, often accompanied by multisystem dysfunction, coagulopathy, and shock is observed. This phase of the illness is characterized by hypercytokinemic inflammation and is often referred to as "cytokine storm." While the immunopathogenesis remains unclear, prospects of treating this severe phase of the illness with immunotherapy are evolving, with some treatments showing promise.


Propuesta de uso de anakinra en el distrés respiratorio agudo secundario a COVID-19./ Propuesta de uso de anakinra en el distrés respiratorio agudo secundario a COVID-19./ Proposal for the use of anakinra in acute respiratory distress secondary to COVID-19

Abstract: The outcome of the SARS-CoV-2 (COVID-19) infection fundamentally affects the lung field, causing acute respiratory distress syndrome (ARDS). This process is an inflammatory picture, involving an NLRP3 inflamosome-triggered cytokine storm, the main player in alveolar destruction. IL-1 beta stands out among the cytokines that are triggered in this picture. Anakinra is a potent biological drug, capable of blocking this IL-1ß. We propose its use in controlling ARDS secondary to COVID-19 infection.


Dipeptidyl peptidase-4 (DPP4) inhibition in COVID-19

Abstract: AIMS: SARS-CoV-2 causes severe respiratory syndrome (COVID-19) with high mortality due to a direct cytotoxic viral effect and a severe systemic inflammation. We are herein discussing a possible novel therapeutic tool for COVID-19. METHODS: Virus binds to the cell surface receptor ACE2; indeed, recent evidences suggested that SARS-CoV-2 may be using as co-receptor, when entering the cells, the same one used by MERS-Co-V, namely the DPP4/CD26 receptor. The aforementioned observation underlined that mechanism of cell entry is supposedly similar among different coronavirus, that the co-expression of ACE2 and DPP4/CD26 could identify those cells targeted by different human coronaviruses and that clinical complications may be similar. RESULTS: The DPP4 family/system was implicated in various physiological processes and diseases of the immune system, and DPP4/CD26 is variously expressed on epithelia and endothelia of the systemic vasculature, lung, kidney, small intestine and heart. In particular, DPP4 distribution in the human respiratory tract may facilitate the entrance of the virus into the airway tract itself and could contribute to the development of cytokine storm and immunopathology in causing fatal COVID-19 pneumonia. CONCLUSIONS: The use of DPP4 inhibitors, such as gliptins, in patients with COVID-19 with, or even without, type 2 diabetes, may offer a simple way to reduce the virus entry and replication into the airways and to hamper the sustained cytokine storm and inflammation within the lung in patients diagnosed with COVID-19 infection.


Cytokine release syndrome and the prospects for immunotherapy with COVID-19. Part 2: The role of interleukin 1

Abstract: Interleukin 1 (IL-1) is a potential target of therapy in COVID-19 during the severe respiratory-inflammatory phase ("cytokine release syndrome"), when pulmonary macrophages are hyperactivated, releasing IL-1 and other cytokines. Preliminary evidence indicates that anakinra and canakinumab, drugs that block the action of IL-1 and have a good safety profile, improve the outcomes of patients with COVID-19 cytokine release syndrome. Results from large, randomized clinical trials are pending.


Storm, typhoon, cyclone or hurricane in patients with COVID-19? Beware of the same storm that has a different origin

Abstract: Some of the articles being published during the severe acute respiratory syndrome-coronavirus (SARS-CoV)-2 pandemic highlight a link between severe forms of coronavirus disease 2019 (COVID-19) and the so-called cytokine storm, also with increased ferritin levels. However, this scenario is more complex than initially thought due to the heterogeneity of hyperinflammation. Some patients with coronavirus 2019 disease (COVID-19) develop a fully blown secondary haemophagocytic lymphohistiocytosis (sHLH), whereas others, despite a consistent release of pro-inflammatory cytokines, do not fulfil sHLH criteria but still show some features resembling the phenotype of the hyperferritinemic syndrome. Despite the final event (the cytokine storm) is shared by various conditions leading to sHLH, the aetiology, either infectious, autoimmune or neoplastic, accounts for the differences in the various phases of this process. Moreover, the evidence of a hyperinflammatory microenvironment provided the rationale to employ immunomodulating agents for therapeutic purposes in severe COVID-19. This viewpoint aims at discussing the pitfalls and issues to be considered with regard to the use of immunomodulating agents in COVID-19, such as timing of treatment based on the viral load and the extent of cytokine/ferritin overexpression. Furthermore, it encompasses recent findings in the paediatric field about a novel multisystem inflammatory disease resembling toxic shock syndrome and atypical Kawasaki disease observed in children with proven SARS-CoV2 infection. Finally, it includes arguments in favour of adding COVID-19 to the spectrum of the recently defined 'hyperferritinemic syndrome', which already includes adult-onset Still's disease, macrophage activation syndrome, septic shock and catastrophic anti-phospholipid syndrome.


Role of Interleukin-6 in Lung Complications in Patients With COVID-19: Therapeutic Implications

Abstract: COVID-19 is viral respiratory infection with frequently fatal lung complications in the elderly or in people with serious comorbidities. Lung destruction appears to be associated with a cytokine storm related to an increased level of interleukin-6 (IL6). Therapeutic targeting of the interleukin-6 signaling pathway can attenuate such a cytokine storm and can be beneficial for patients with COVID-19 in danger of pulmonary failure. This article demonstrates the importance of IL6 in progression of disease and the possibility of inhibition of IL6 signaling in COVID-19 therapy.


Endothelial progenitor cells and mesenchymal stem cells to overcome vascular deterioration and cytokine storm in critical patients with COVID-19


Tocilizumab challenge: A series of cytokine storm therapy experiences in hospitalized COVID-19 pneumonia patients

Abstract: To recognize the period of exaggerated cytokine response in patients with coronavirus disease 2019 (COVID-19) pneumonia, and to describe the clinical outcomes of using tocilizumab as a treatment option. The data of 12 adult COVID-19 pneumonia patients who were followed in the inpatient clinics of Biruni University Medical Faculty Hospital (Istanbul, Turkey) were retrospectively analyzed. Diagnostic tests, laboratory examinations, clinical findings, and computed tomography of the thorax imaging results were evaluated. A dramatic laboratory and clinical improvement was observed in 83% (10 out of 12) of patients after tocilizumab. In 17% (2 out of 12) of our patients, short-term ventilator support was required in the intensive care unit. The longest hospital stay was 18 days. However, in the end, all of our patients were discharged home with good health. Although arterial oxygen saturations (87.58 ± 3.12%) dropped in room air in the pre-tocilizumab period, post-tocilizumab they normalized in all patients (94.42 ± 1%). None of them had fever after tocilizumab treatment and the levels of C-reactive protein (13.08 ± 12.89) were almost within normal limits. Eosinophil values were quite low at the time of diagnosis (10 ± 17.06), but increased significantly post-tocilizumab (155.33 ± 192.69). There is currently no proven treatment for COVID-19 induced by novel coronavirus SARS-CoV-2. Based on our experience with twelve adult COVID-19 pneumonia patients,we can say that tocilizumab, an IL-6 inhibitor, is more beneficial in preventing the damage caused by excessive cytokine response in the body if administered at the right time and provides clinical and radiological recovery.


[Advances in the research of cytokine storm mechanism induced by Corona Virus Disease 2019 and the corresponding immunotherapies]

Abstract: Corona Virus Disease 2019 (COVID-19) has seriously affected the treatment of patients and social stability. In the later stage of disease, some COVID-19 patients may develop into acute respiratory distress syndrome or even multiple organ failure. However, one of the most important mechanism underlying the deterioration of disease is cytokine storm. At present, some therapies such as interleukin-6 antibody blocker, stem cell therapy, and transfusion of convalescent plasma have been applied to counteract the cytokine storm and have made some progress. This article reviews the influences of cytokine storm syndrome on the COVID-19 and the corresponding immunotherapies to resist cytokine storm.


Prevention of the cytokine storm in COVID-19


Hemophagocytic lymphohistiocytosis: a review inspired by the COVID-19 pandemic

Abstract: Hemophagocytic syndrome (HPS) or hemophagocytic lymphohistiocytosis (HLH) is an acute and rapidly progressive systemic inflammatory disorder characterized by cytopenia, excessive cytokine production, and hyperferritinemia. Common clinical manifestations of HLH are acute unremitting fever, lymphadenopathy, hepatosplenomegaly, and multiorgan failure. Due to a massive cytokine release, this clinical condition is considered as a cytokine storm syndrome. HPS has primary and acquired (secondary, reactive) forms. Its primary form is mostly seen in childhood and caused by various mutations with genetic inheritance and, therefore, is called familial HLH. Secondary HLH may be caused in the presence of an underlying disorder, that is, secondary to a malignant, infectious, or autoimmune/autoinflammatory stimulus. This paper aims to review the pathogenesis and the clinical picture of HLH, and its severe complication, the cytokine storm, with a special emphasis on the developed classification criteria sets for rheumatologists, since COVID-19 infection has clinical symptoms resembling those of the common rheumatologic conditions and possibly triggers HLH. MED-LINE/Pubmed was searched from inception to April 2020, and the following terms were used for data searching: "hemophagocytic syndrome" OR "macrophage activation syndrome" OR "hemophagocytic lymphohistiocytosis", OR "cytokine storm". Finally, AND "COVID-19" was included in this algorithm. The selection is restricted to the past 5 years and limited numbers of earlier key references were manually selected. Only full-text manuscripts, published in an English language peer-reviewed journal were included. Manuscript selection procedure and numbers are given in Fig. 2. Briefly, the database search with the following terms of "Hemophagocytic syndrome" OR "Macrophage activation syndrome" OR "Hemophagocytic lymphohistiocytosis" OR "Cytokine storm" yielded 6744 results from inception to April 2020. The selection is restricted to the past 5 years and only limited numbers of earlier key references were selected, and this algorithm resulted in 3080 manuscripts. The addition of (AND "COVID-19") resulted in 115 publications of which 47 studies, together with four sections of an online book were used in the final review. No statistical method was used. HLH is triggered by genetic conditions, infections, malignancies, autoimmune-autoinflammatory diseases, and some drugs. In COVID-19 patients, secondary HLH and cytokine storm may be responsible for unexplained progressive fever, cytopenia, ARDS, neurological and renal impairment. Differentiation between the primary and secondary forms of HLH is utterly important, since primary form of HLH requires complicated treatments such as hematopoietic stem cell transplantation. Further studies addressing the performance of HScore and other recommendations in the classification of these patients is necessary.


Potential for jaktinib hydrochloride to treat cytokine storms in patients with COVID-19

Abstract: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has become a serious public health threat to the whole world, and the number of infected is still rising dramatically at this moment. Several studies have confirmed that cytokine storms play a critical role in causing a case to worsen from mild to severe or critical. The current treatment for cytokine storms is limited, so the international medical community is focusing on a specific and effective remedy. Jaktinib hydrochloride is a broad spectrum JAK inhibitor. It can inhibit cytokine-induced immune activation by multiple mechanisms and also slow viral proliferation by inhibiting AAK1 without causing unacceptable toxicity. Jaktinib hydrochloride has great potential for the treatment of patients with coronavirus disease 2019 (COVID-19).


Utilizing tocilizumab for the treatment of cytokine release syndrome in COVID-19


Absence of severe complications from SARS-CoV-2 infection in children with rheumatic diseases treated with biologic drugs

Abstract: We read with interest the Editorial by Cron and Chatam (1) suggesting a cytokine storm syndrome (CSS) occurring in response to SARS-CoV-2 infection and, consequently, a possible role for targeted approaches to blocking inflammatory cytokines.


Weathering the COVID-19 storm: Lessons from hematologic cytokine syndromes

Abstract: A subset of patients with severe COVID-19 develop profound inflammation and multi-organ dysfunction consistent with a "Cytokine Storm Syndrome" (CSS). In this review we compare the clinical features, diagnosis, and pathogenesis of COVID-CSS with other hematological CSS, namely secondary hemophagocytic lymphohistiocytosis (sHLH), idiopathic multicentric Castleman disease (iMCD), and CAR-T cell therapy associated Cytokine Release Syndrome (CRS). Novel therapeutics targeting cytokines or inhibiting cell signaling pathways have now become the mainstay of treatment in these CSS. We review the evidence for cytokine blockade and attenuation in these known CSS as well as the emerging literature and clinical trials pertaining to COVID-CSS. Established markers of inflammation as well as cytokine levels are compared and contrasted between these four entities in order to establish a foundation for future diagnostic criteria of COVID-CSS.


A complex COVID-19 case with rheumatoid arthritis treated with tocilizumab

Abstract: Recurrences of COVID-19 were observed in a patient with long-term usage of hydroxychloroquine, leflunomide, and glucocorticoids due to her 30-year history of rheumatoid arthritis (RA). Tocilizumab was applied and intended to target both COVID-19 and RA. However, disease of this patient aggravated after usage of tocilizumab. After the discussion of a multiple disciplinary team (MDT) including rheumatologists, antimicrobial treatments were applied to target the potential opportunistic infections (Pneumocystis jirovecii and Aspergillus fumigatus), which were authenticated several days later via high throughput sequencing. As an important cytokine in immune responses, IL-6 can be a double-edged sword: interference in the IL-6-IL-6 receptor signaling may save patients from cytokine release storm (CRS), but can also weaken the anti-infectious immunity, particularly in rheumatic patients, who may have received a long-term treatment with immunosuppressive/modulatory agents. Thus, we suggest careful considerations before and close monitoring in the administration of tocilizumab in rheumatic patients with COVID-19. Besides tocilizumab, several disease-modifying antirheumatic drugs (DMARDs) can also be applied in the treatment of COVID-19. Therefore, we also reviewed and discussed the application of these DMARDs in COVID-19 condition.


Tocilizumab's efficacy in patients with Coronavirus Disease 2019 (COVID-19) is determined by the presence of cytokine storm


COVID 19: a clue from innate immunity

Abstract: The recent COVID-19 pandemic has had a significant impact on our lives and has rapidly expanded to reach more than 4 million cases worldwide by May 2020. These cases are characterized by extreme variability, from a mild or asymptomatic form lasting for a few days up to severe forms of interstitial pneumonia that may require ventilatory therapy and can lead to patient death.Several hypotheses have been drawn up to understand the role of the interaction between the infectious agent and the immune system in the development of the disease and the most severe forms; the role of the cytokine storm seems important.Innate immunity, as one of the first elements of guest interaction with different infectious agents, could play an important role in the development of the cytokine storm and be responsible for boosting more severe forms. Therefore, it seems important to study also this important arm of the immune system to adequately understand the pathogenesis of the disease. Research on this topic is also needed to develop therapeutic strategies for treatment of this disease.


Inhibition of cytokine signaling by ruxolitinib and implications for COVID-19 treatment

Abstract: Approximately 15% of patients with coronavirus disease 2019 (COVID-19) experience severe disease, and 5% progress to critical stage that can result in rapid death. No vaccines or antiviral treatments have yet proven effective against COVID-19. Patients with severe COVID-19 experience elevated plasma levels of pro-inflammatory cytokines, which can result in cytokine storm, followed by massive immune cell infiltration into the lungs leading to alveolar damage, decreased lung function, and rapid progression to death. As many of the elevated cytokines signal through Janus kinase (JAK)1/JAK2, inhibition of these pathways with ruxolitinib has the potential to mitigate the COVID-19-associated cytokine storm and reduce mortality. This is supported by preclinical and clinical data from other diseases with hyperinflammatory states, where ruxolitinib has been shown to reduce cytokine levels and improve outcomes. The urgent need for treatments for patients with severe disease support expedited investigation of ruxolitinib for patients with COVID-19.


Targeting JAK-STAT Signaling to Control Cytokine Release Syndrome in COVID-19

Abstract: Recent advances in the pathophysiologic understanding of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has indicated that patients with severe coronavirus disease 2019 (COVID-19) might experience cytokine release syndrome (CRS), characterized by increased interleukin (IL)-6, IL-2, IL-7, IL-10, etc. Therefore, the treatment of cytokine storm has been proposed as a critical part of rescuing severe COVID-19. Several of the cytokines involved in COVID-19 employ a distinct intracellular signaling pathway mediated by Janus kinases (JAKs). JAK inhibition, therefore, presents an attractive therapeutic strategy for CRS, which is a common cause of adverse clinical outcomes in COVID-19. Below, we review the possibilities and challenges of targeting the pathway in COVID-19.


Melatonin Inhibits COVID-19-induced Cytokine Storm by Reversing Aerobic Glycolysis in Immune Cells: A Mechanistic Analysis


The COVID-19 Cytokine Storm; What We Know So Far

Abstract: COVID-19 is a rapidly spreading global threat that has been declared as a pandemic by the WHO. COVID-19 is transmitted via droplets or direct contact and infects the respiratory tract resulting in pneumonia in most of the cases and acute respiratory distress syndrome (ARDS) in about 15 % of the cases. Mortality in COVID-19 patients has been linked to the presence of the so-called "cytokine storm" induced by the virus. Excessive production of proinflammatory cytokines leads to ARDS aggravation and widespread tissue damage resulting in multi-organ failure and death. Targeting cytokines during the management of COVID-19 patients could improve survival rates and reduce mortality.


COVID-19: a recommendation to examine the effect of hydroxychloroquine in preventing infection and progression ;Journal of Antimicrobial Chemotherapy ;Oxford Academic

Abstract: A novel coronavirus disease (COVID-19), caused by infection with SARS-CoV-2, has swept across 31 provinces in China and over 40 countries worldwide The transition from first symptoms to acute respiratory distress syndrome (ARDS) is highly likely to be due to uncontrolled cytokine release There is an urgent need to identify safe and effective drugs for treatment Chloroquine (CQ) exhibits a promising inhibitory effect However, the clinical use of CQ can cause severe side effects We propose that hydroxychloroquine (HCQ), which exhibits an antiviral effect highly similar to that of CQ, could serve as a better therapeutic approach HCQ is likely to attenuate the severe progression of COVID-19, inhibiting the cytokine storm by suppressing T cell activation It has a safer clinical profile and is suitable for those who are pregnant It is cheaper and more readily available in China We herein strongly urge that clinical trials are performed to assess the preventive effects of HCQ in both disease infection and progression


Molecular characterization of SARS-CoV-2 in the first COVID-19 cluster in France reveals an amino acid deletion in nsp2 (Asp268del)


Conserved High Free Energy Sites in Human Coronavirus Spike Glycoprotein Backbones.

Abstract: Methods previously developed by the author are applied to uncover several sites of interest in the spike glycoproteins of all known human coronaviruses (hCoVs), including SARS-CoV-2 that causes COVID-19. The sites comprise three-dimensional neighborhoods of peptides characterized by four key properties: (1) they pinpoint regions of high free energy in the backbone whose obstruction might interrupt function; (2) by their very definition, they occur rarely in the universe of all gene-encoded proteins that could obviate host response to compounds designed for their interference; (3) they are common to all seven hCoV spikes, possibly retaining activity in light of inevitable viral mutation; and (4) they are exposed in the molecular surface of the glycoprotein. These peptides in SARS-CoV-2 are given by the triples of residues (131, 117, 134), (203, 227, 228), and (1058, 730, 731) in its spike.


An overview of the genetic variations of the SARS-CoV-2 genomes isolated in Southeast Asian countries.

Abstract: Monitoring the mutation dynamics of human severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is critical in understanding its infectivity, virulence and pathogenicity for development of a vaccine. In an "age of mobility", the pandemic highlights the importance and vulnerability of regionalisation and labour market interdependence in Southeast Asia. We intend to characterise the genetic variability of viral populations within the region to provide preliminary information for regional surveillance in the future. By analysing 142 complete genomes from South East Asian (SEA) countries, we identified three central variants distinguished by nucleotide and amino acid changes.


SARS-CoV-2 (COVID-19) structural and evolutionary dynamicome: Insights into functional evolution and human genomics.

Abstract: The pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has challenged the speed at which laboratories discover the viral composition and study health outcomes. The small ~30-kb ssRNA genome of coronaviruses makes them adept at cross-species spread, but also enable a robust understanding of all the proteins the viral genome encodes. We have employed protein modeling, molecular dynamic simulations, evolutionary mapping, and 3D printing to gain a full proteome- and dynamicome-level understanding of SARS-CoV-2. We established the Viral Integrated Structural Evolution Dynamic Database (VIStEDD at prokoplab.com/vistedd) to facilitate future discoveries and educational use. Here, we highlight the use of VIStEDD for nsp6, nucleocapsid (N), and spike (S) surface glycoprotein. For both nsp6 and N, we found highly conserved surface amino acids that likely drive protein-protein interactions. In characterizing viral S protein, we developed a quantitative dynamics cross-correlation matrix to gain insights into its interactions with the angiotensin I-converting enzyme 2 (ACE2)-solute carrier family 6 member 19 (SLC6A19) dimer. Using this quantitative matrix, we elucidated 47 potential functional missense variants from genomic databases within ACE2/SLC6A19/transmembrane serine protease 2 (TMPRSS2), warranting genomic enrichment analyses in SARS-CoV-2 patients. These variants had ultralow frequency but existed in males hemizygous for ACE2. Two ACE2 noncoding variants (rs4646118 and rs143185769) present in ~9% of individuals of African descent may regulate ACE2 expression and may be associated with increased susceptibility of African Americans to SARS-CoV-2. We propose that this SARS-CoV-2 database may aid research into the ongoing pandemic.


Implications of SARS-CoV-2 Mutations for Genomic RNA Structure and Host microRNA Targeting.

Abstract: The SARS-CoV-2 virus is a recently-emerged zoonotic pathogen already well adapted to transmission and replication in humans. Although the mutation rate is limited, recently introduced mutations in SARS-CoV-2 have the potential to alter viral fitness. In addition to amino acid changes, mutations could affect RNA secondary structure critical to viral life cycle, or interfere with sequences targeted by host miRNAs. We have analysed subsets of genomes from SARS-CoV-2 isolates from around the globe and show that several mutations introduce changes in Watson-Crick pairing, with resultant changes in predicted secondary structure. Filtering to targets matching miRNAs expressed in SARS-CoV-2-permissive host cells, we identified ten separate target sequences in the SARS-CoV-2 genome; three of these targets have been lost through conserved mutations. A genomic site targeted by the highly abundant miR-197-5p, overexpressed in patients with cardiovascular disease, is lost by a conserved mutation. Our results are compatible with a model that SARS-CoV-2 replication within the human host is constrained by host miRNA defences. The impact of these and further mutations on secondary structures, miRNA targets or potential splice sites offers a new context in which to view future SARS-CoV-2 evolution, and a potential platform for engineering conditional attenuation to vaccine development, as well as providing a better understanding of viral tropism and pathogenesis.


SARS-CoV-2 receptor mutation in Egyptian population

Abstract: The Coronavirus disease 2019 (COVID-19) is a respiratory tract infectious disease caused by Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2). SARS-CoV-2 triggers severe pneumonia leading to acute respiratory distress syndrome and death in severe cases. According to WHO reported, Egypt is among the countries with low confirmed SARS CoV2 infected symptomatic cases and death. We postulate that one of the reasons for this may be due mutations in the viral receptor. Therefore this study was conducted to confirm or reject this postulation.


International expansion of a novel SARS-CoV-2 mutant

Abstract: Letter to the editor. There is no abstract. The summary was showed: SARS-CoV-2 has inevitably mutated during its pandemic spread to cause unpredictable effects on COVID-19 and complicate epidemic control efforts. Here we report that a novel SARS-CoV-2 mutation (ORF3a) appears to be spreading worldwide, which deserves close attention.


Identification of unique mutations in SARS-CoV-2 strains isolated from India suggests its attenuated pathotype

Abstract: Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), which was first reported in Wuhan, China in November 2019 has developed into a pandemic since March 2020, causing substantial human casualties and economic losses. Studies on SARS-CoV-2 are being carried out at an unprecedented rate to tackle this threat. Genomics studies, in particular, are indispensable to elucidate the dynamic nature of the RNA genome of SARS-CoV-2. RNA viruses are marked by their unique ability to undergo high rates of mutation in their genome, much more frequently than their hosts, which diversifies their strengths qualifying them to elude host immune response and amplify drug resistance. In this study, we sequenced and analyzed the genomic information of the SARS-CoV-2 isolates from two infected Indian patients and explored the possible implications of point mutations in its biology. In addition to multiple point mutations, we found a remarkable similarity between relatively common mutations of 36-nucleotide deletion in ORF8 of SARS-CoV-2. Our results corroborate with the earlier reported 29-nucleotide deletion in SARS, which was frequent during the early stage of human-to-human transmission. The results will be useful to understand the biology of SARS-CoV-2 and itsattenuation for vaccine development.


ACE2 diversity in placental mammals reveals the evolutionary strategy of SARS-CoV-2

Abstract: The recent emergence of SARS-CoV-2 is responsible for the current pandemic of COVID-19, which uses the human membrane protein ACE2 as a gateway to host-cell infection We performed a comparative genomic analysis of 70 ACE2 placental mammal orthologues to identify variations and contribute to the understanding of evolutionary dynamics behind this successful adaptation to infect humans Our results reveal that 4% of the ACE2 sites are under positive selection, all located in the catalytic domain, suggesting possibly taxon-specific adaptations related to the ACE2 function, such as cardiovascular physiology Considering all variable sites, we selected 30 of them located at the critical ACE2 binding sites to the SARS-CoV-like viruses for analysis in more detail Our results reveal a relatively high diversity of ACE2 between placental mammal species, while showing no polymorphism within human populations, at least considering the 30 inter-species variable sites A perfect scenario for natural selection favored this opportunistic new coronavirus in its trajectory of infecting humans We suggest that SARS-CoV-2 became a specialist coronavirus for human hosts Differences in the rate of infection and mortality could be related to the innate immune responses, other unknown genetic factors, as well as non-biological factors


Molecular analysis of several in-house rRT-PCR protocols for SARS-CoV-2 detection in the context of genetic variability of the virus in Colombia.

Abstract: The COVID-19 pandemic caused by SARS-CoV-2 is a public health problem unprecedented in the recent history of humanity. Different in-house real-time RT-PCR (rRT-PCR) methods for SARS-CoV-2 diagnosis and the appearance of genomes with mutations in primer regions have been reported. Hence, whole-genome data from locally-circulating SARS-CoV-2 strains contribute to the knowledge of its global variability and the development and fine-tuning of diagnostic protocols. To describe the genetic variability of Colombian SARS-CoV-2 genomes in hybridization regions of oligonucleotides of the main in-house methods for SARS-CoV-2 detection, RNA samples with confirmed SARS-CoV-2 molecular diagnosis were processed through next-generation sequencing. Primers/probes sequences from 13 target regions for SARS-CoV-2 detection suggested by 7 institutions and consolidated by WHO during the early stage of the pandemic were aligned with Muscle tool to assess the genetic variability potentially affecting their performance. Finally, the corresponding codon positions at the 3' end of each primer, the open reading frame inspection was identified for each gene/protein product. Complete SARS-CoV-2 genomes were obtained from 30 COVID-19 cases, representative of the current epidemiology in the country. Mismatches between at least one Colombian sequence and five oligonucleotides targeting the RdRP and N genes were observed. The 3' end of 4 primers aligned to the third codon position, showed high risk of nucleotide substitution and potential mismatches at this critical position. Genetic variability was detected in Colombian SARS-CoV-2 sequences in some of the primer/probe regions for in-house rRT-PCR diagnostic tests available at WHO COVID-19 technical guidelines; its impact on the performance and rates of false-negative results should be experimentally evaluated. The genomic surveillance of SARS-CoV-2 is highly recommended for the early identification of mutations in critical regions and to issue recommendations on specific diagnostic tests to ensure the coverage of locally-circulating genetic variants.


Genetic Variant of SARS-CoV-2 Isolates in Indonesia: Spike Glycoprotein Gene

Abstract: Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), a novel coronavirus and the primary causative agent of coronavirus disease 2019 (COVID-19), first occurred in China and rapidly spread worldwide The government of the Republic of Indonesia confirmed its first two cases of COVID-19 in March 2020 COVID-19 is a serious illness with no efficacious antiviral medication or approved vaccine currently available Therefore, there is a need to investigate the genome of SARS-CoV-2 In this study, we characterized SARS-CoV-2 spike glycoprotein genes from Indonesia to investigate their genetic composition and variability Overall, ten SARS-CoV-2 spike glycoprotein gene sequences retrieved from GenBank (National Center for Biotechnology Information, USA) and the GISAID EpiCoV database (Germany) were compared We analyzed nucleotide variants and amino acid changes using Molecular Evolutionary Genetics Analysis (MEGA) X and analyzed gene similarity using the LALIGN web server Interestingly, we revealed several specific mutation sites, however, there were no significant changes in the genetic composition of SARS-CoV-2 spike glycoprotein genes, when compared to the WuhanHu-1 isolate from China However, this is a preliminary study and we recommend that molecular epidemiology and surveillance programs against COVID-19 in Indonesia be improved


Impact of emerging mutations on the dynamic properties the SARS-CoV-2 main protease: an in silico investigation

Abstract: The new coronavirus (SARS-CoV-2) is a global threat to world health and its economy. Its main protease (Mpro), which functions as a dimer, cleaves viral precursor proteins in the process of viral maturation. It is a good candidate for drug development owing to its conservation and the absence of a human homolog. An improved understanding of the protein behaviour can accelerate the discovery of effective therapies in order to reduce mortality. 100 ns all-atom molecular dynamics simulations of 50 homology modelled mutant Mpro dimers were performed at pH 7 from filtered sequences obtained from the GISAID database. Protease dynamics were analysed using RMSD, RMSF, Rg, the averaged betweenness centrality and geometry calculations. Domains from each Mpro protomer were found to generally have independent motions, while the dimer-stabilising N-finger region was found to be flexible in most mutants. A mirrored interprotomer pocket was found to be correlated to the catalytic site using compaction dynamics, and can be a potential allosteric target. The high number of titratable amino acids of Mpro may indicate an important role of pH on enzyme dynamics, as previously reported for SARS-CoV. Independent coarse-grained Monte Carlo simulations suggest a link between rigidity/mutability and enzymatic function.


COVID-3D: An online resource to explore the structural distribution of genetic variation in SARS-CoV-2 and its implication on therapeutic development

Abstract: The emergence of the COVID-19 pandemic has spurred a global rush to uncover basic biological mechanisms, to inform effective vaccine and drug development. Despite viral novelty, global sequencing efforts have already identified genomic variation across isolates. To enable easy exploration and spatial visualization of the potential implications of SARS-CoV-2 mutations on infection, host immunity and drug development we have developed COVID-3D (http://biosig.unimelb.edu.au/covid3d/).


Cross-neutralization antibodies against SARS-CoV-2 and RBD mutations from convalescent patient antibody libraries

Abstract: The emergence of coronavirus disease 2019 (COVID-19) pandemic led to an urgent need to develop therapeutic interventions. Among them, neutralizing antibodies play crucial roles for preventing viral infections and contribute to resolution of infection. Here, we describe the generation of antibody libraries from 17 different COVID-19 recovered patients and screening of neutralizing antibodies to SARS-CoV-2. After 3 rounds of panning, 456 positive phage clones were obtained with high affinity to RBD (receptor binding domain). Then the positive clones were sequenced and reconstituted into whole human IgG for epitope binning assays. After that, all 19 IgG were classified into 6 different epitope groups or Bins. Although all these antibodies were shown to have ability to bind RBD, the antibodies in Bin2 have more superiority to inhibit the interaction between spike protein and angiotensin converting enzyme 2 receptor (ACE2). Most importantly, the antibodies from Bin2 can also strongly bind with mutant RBDs (W463R, R408I, N354D, V367F and N354D/D364Y) derived from SARS-CoV-2 strain with increased infectivity, suggesting the great potential of these antibodies in preventing infection of SARS-CoV-2 and its mutations. Furthermore, these neutralizing antibodies strongly restrict the binding of RBD to hACE2 overexpressed 293T cells. Consistently, these antibodies effectively neutralized pseudovirus entry into hACE2 overexpressed 293T cells. In Vero-E6 cells, these antibodies can even block the entry of live SARS-CoV-2 into cells at only 12.5 nM. These results suggest that these neutralizing human antibodies from the patient-derived antibody libraries have the potential to become therapeutic agents against SARS-CoV-2 and its mutants in this global pandemic.


Ethnically diverse mutations in PIEZO1 associate with SARS-CoV-2 positivity

Abstract: COVID-19, caused by the SARS-CoV-2 virus, carries significant risk of mortality and has spread globally with devastating societal consequences. Endothelial infection has been identified as a feature of the disease and so there is motivation to determine the relevance of endothelial membrane mechanisms affecting viral entry and response. Here, through a study of patient data in UK Biobank released on 16 April 2020, we suggest relevance of PIEZO1, a non-selective cation channel protein that both mediates endothelial responses to mechanical force and unusually indents the cell membrane. PIEZO1 notably has roles that may also be relevant in red blood cell function, pulmonary inflammation, bacterial infection and fibrotic auto-inflammation. We provide evidence that single nucleotide polymorphisms (SNPs) in the gene encoding PIEZO1 are more common in individuals who test positive for SARS-CoV-2 regardless of pre-existing hypertension, myocardial infarction, stroke, diabetes mellitus or arthritis. Some of these SNPs are more common in African and Caribbean populations, which are groups that were recently shown to have greater susceptibility to infection. One of the SNPs is a missense mutation that results in an amino acid change in an evolutionarily conserved and previously unexplored N-terminal region PIEZO1. The data support the notion of genetic factors influencing SARS-CoV-2 infection and suggest a specific role for PIEZO1.


COVID-19 genomic susceptibility: Definition of ACE2 variants relevant to human infection with SARS-CoV-2 in the context of ACMG/AMP Guidance

Abstract: BACKGROUND: Mortality remains very high and unpredictable in COVID-19, with intense public protection strategies tailored to preceived risk. Genomic studies are in process to identify differences in host susceptibility to infection by the SARS-CoV-2 virus. Open source research publications are accessible to pre-genotyped individuals. Males are more at risk from severe COVID-19. METHODS: To facilitate development of Clinical Genetics support to the public and healthcare professionals, genomic structure and variants in 213,158 exomes/genomes were integrated for ACE2 encoding the SARS-CoV-2 receptor. ACMG/AMP-based pathogenicity criteria were applied. RESULTS: Across the 19 ACE2 exons on the X chromosome, 9 of 3596 (0.25%) nucleotides were homozygous variant in females compared to 262/3596 (7.3%) hemizygous variant in males (p< 0.0001). 90% of variants were very rare, although K26R affecting a SARS-CoV-2-interacting amino acid is present in ~1/239 people. Modelling the ''COVID-resistant '' state where pathogenic alleles would be beneficial, nine null alleles met PVS1. Thirty-seven variants met PM1 based on critical location +/-PP3 based on computational modelling. Modelling a ''COVID-susceptible '' state, 31 variants in four upstream open reading frames and 5' untranslated regions could meet PM1, and may have differential effects if aminoglycoside antibiotics were prescribed for pneumonia and sepsis. CONCLUSIONS: Males are more likely to exhibit consequences from a single variant ACE2 allele. Differential allele frequencies in COVID-19 susceptible and resistant individuals are likely to emerge before variants meet ACMG/AMP criteria for actionable results in patients. Prioritising genomic regions for functional study and ACMG/AMP-structured approaches to research-based presentation of COVID-19 susceptibility variant results are encouraged.


Mutation density changes in SARS-CoV-2 are related to the pandemic stage but to a lesser extent in the dominant strain with mutations in spike and RdRp

Abstract: Since its emergence in Wuhan, China in late 2019, the origin and evolution of SARS-CoV-2 have been among the most debated issues related to COVID-19. Throughout its spread around the world, the viral genome continued acquiring new mutations and some of them became widespread. Among them, 14408 C>T and 23403 A>G mutations in RdRp and S, respectively, became dominant in Europe and the US, which led to debates regarding their effects on the mutability and transmissibility of the virus. In this study, we aimed to investigate possible differences between time-dependent variation of mutation densities (MDe) of viral strains that carry these two mutations and those that do not. Our analyses at the genome and gene level led to two important findings: First, time-dependent changes in the average MDe of circulating SARS-CoV-2 genomes showed different characteristics before and after the beginning of April, when daily new case numbers started levelling off. Second, this pattern was much delayed or even non-existent for the “mutant” (MT) strain that harbored both 14408 C>T and 23403 A>G mutations. Although these differences were not limited to a few hotspots, it is intriguing that the MDe increase is most evident in two critical genes, S and Orf1ab, which are also the genes that harbor the defining mutations of the MT genotype. The nature of these unexpected relationships warrant further research.


Mutations can reveal how the coronavirus moves—but they’re easy to overinterpret

Abstract: Immediately after Christian Drosten published a genetic sequence of the novel coronavirus online on 28 February, he took to Twitter to issue a warning As the virus has raced around the world, more than 350 genome sequences have been shared on the online platform GISAID They hold clues to how the new virus, named severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is spreading and evolving But because the sequences represent a tiny fraction of cases and show few telltale differences, they are easy to overinterpret, as Drosten realized A virologist at the Charité University Hospital in Berlin, he had sequenced the virus from a German patient infected with COVID-19 in Italy The genome looked similar to that of a virus found in a patient in Munich, the capital of Bavaria, more than 1 month earlier;both shared three mutations not seen in early sequences from China Drosten realized this could give rise to the idea that the Italian outbreak was “seeded” by the one in Bavaria, which state public health officials said had been quashed by tracing and quarantining all contacts of the 14 confirmed cases But he thought it was just as likely that a Chinese variant carrying the three mutations had taken independent routes to both countries The newly sequenced genome “is not sufficient to claim a link between Munich and Italy,” Drosten tweeted


Mutational Signatures and Heterogeneous Host Response Revealed Via Large-Scale Characterization of SARS-COV-2 Genomic Diversity

Abstract: To dissect the mechanisms underlying the observed inflation of variants in SARS-CoV-2 genome, we present the largest up-to-date analysis of intra-host genomic diversity, which reveals that the majority of samples present a complex sublineage architecture, due to the interplay between host-related mutational processes and transmission dynamics. Strikingly, the deconvolution of the entire set of intra-host variants reveals the existence of mutually exclusive viral mutational signatures, which prove that distinct hosts differently respond to SARS-CoV-2 infections. In particular, two signatures are likely ruled by APOBEC and Reactive Oxygen Species (ROS), which induce hypermutation in a significant number of samples, and appear to be affected by severe purifying selection. Conversely, several mutations linked to low-rate mutational processes appear to transit to clonality in the population, eventually leading to the definition of new viral genotypes and to an increase of overall genomic diversity. Finally, we demonstrate that a high number of variants are observed in samples associated to independent lineages, likely due to signature-related mutational hotspots or to positive selection.


Intra-host site-specific polymorphisms of SARS-CoV-2 is consistent across multiple samples and methodologies

Abstract: Despite the potential relevance to clinical outcome, intra-host dynamics of SARS-CoV-2 are unclear. Here, we quantify and characterize intra-host variation in SARS-CoV-2 raw sequence data uploaded to SRA as of 14 April 2020, and compare results between two sequencing methods (amplicon and RNA-Seq). Raw fastq files were quality filtered and trimmed using Trimmomatic, mapped to the WuhanHu1 reference genome using Bowtie2, and variants called with bcftools mpileup. To ensure sufficient coverage, we only included samples with 10X coverage for >90% of the genome (n=406 samples), and only variants with a depth >=10. Derived (i.e. non-reference) alleles were found at 408 sites. The number of polymorphic sites (i.e. sites with multiple alleles) within samples ranged from 0-13, with 72% of samples (295/406) having at least one polymorphic site. Correlation between number of polymorphic sites and coverage was very low for both sequencing methods (R2 < 0.1, p < 0.05). Polymorphisms were observed >1 sample at 66 sites (range: 2-38 samples). The minor allele frequency (MAF) at each shared polymorphic site was 0.03% - 48.5%. 33/66 sites occurred in ORF1a1b, and 37/66 changes were non-synonymous. At 10/66 sites, derived alleles were found in samples sequenced using both methods. Polymorphic amplicon samples were found at 10/10 positions, while polymorphic RNA-Seq samples were found at 7/10 positions. In conclusion, our results suggest that intra-host variation is prevalent among clinical samples. While mutations resulting from amplification and/or sequencing errors cannot be excluded, the observation of shared polymorphic sites with high MAF across multiple samples and sequencing methods is consistent with true underlying variation. Further investigation into intra-host evolutionary dynamics, particularly with longitudinal sampling, is critical for broader understanding of disease progression.


Anti-frameshifting ligand active against SARS coronavirus-2 is resistant to natural mutations of the frameshift-stimulatory pseudoknot

Abstract: The coronavirus SARS-CoV-2 causing the COVID-19 pandemic uses −1 programmed ribosomal frameshifting (−1 PRF) to control the expression levels of key viral proteins. Because modulating −1 PRF can attenuate viral propagation, ligands binding to the viral RNA pseudoknot that stimulates −1 PRF may prove useful as therapeutics. Mutations in the pseudoknot have been observed over the course of the pandemic, but how they affect −1 PRF and the activity of inhibitors is unknown. Cataloguing natural mutations in all parts of the SARS-CoV-2 pseudoknot, we studied a panel of 6 mutations in key structural regions. Most mutations left the −1 PRF efficiency unchanged, even when base-pairing was disrupted, but one led to a remarkable three-fold decrease, suggesting that SARS-CoV-2 propagation may be less sensitive to modulation of −1 PRF efficiency than some other viruses. Examining the effects of one of the few small-molecule ligands known to suppress −1 PRF significantly in SARS-CoV, we found that it did so by similar amounts in all SARS-CoV-2 mutants tested, regardless of the basal −1 PRF efficiency, indicating that the activity of anti-frameshifting ligands can be resistant to natural pseudoknot mutations. These results have important implications for therapeutic strategies targeting SARS-CoV-2 through modulation of −1 PRF.


Pervasive RNA secondary structure in the genomes of SARS-CoV-2 and other coronaviruses – an endeavour to understand its biological purpose

Abstract: The ultimate outcome of the COVID-19 pandemic is unknown and is dependent on a complex interplay of its pathogenicity, transmissibility and population immunity. In the current study, SARS coronavirus 2 (SARS-CoV-2) was investigated for the presence of large scale internal RNA base pairing in its genome. This property, termed genome scale ordered RNA structure (GORS) has been previously associated with host persistence in other positive-strand RNA viruses, potentially through its shielding effect on viral RNA recognition in the cell. Genomes of SARS-CoV-2 were remarkably structured, with minimum folding energy differences (MFEDs) of 15%, substantially greater than previously examined viruses such as HCV (MFED 7-9%). High MFED values were shared with all coronavirus genomes analysed created by several hundred consecutive energetically favoured stem-loops throughout the genome. In contrast to replication-association RNA structure, GORS was poorly conserved in the positions and identities of base pairing with other sarbecoviruses – even similarly positioned stem-loops in SARS-CoV-2 and SARS-CoV rarely shared homologous pairings, indicative of more rapid evolutionary change in RNA structure than in the underlying coding sequences. Sites predicted to be base-paired in SARS-CoV-2 showed substantially less sequence diversity than unpaired sites, suggesting that disruption of RNA structure by mutation imposes a fitness cost on the virus which is potentially restrictive to its longer evolution. Although functionally uncharacterised, GORS in SARS-CoV-2 and other coronaviruses represent important elements in their cellular interactions that may contribute to their persistence and transmissibility.


Discovery of a 382-nt deletion during the early evolution of SARS-CoV-2

Abstract: To date, the SARS-CoV-2 genome has been considered genetically more stable than SARS-CoV or MERS-CoV. Here we report a 382-nt deletion covering almost the entire open reading frame 8 (ORF8) of SARS-CoV-2 obtained from eight hospitalized patients in Singapore. The deletion also removes the ORF8 transcription-regulatory sequence (TRS), which in turn enhances the downstream transcription of the N gene. We also found that viruses with the deletion have been circulating for at least four weeks. During the SARS-CoV outbreak in 2003, a number of genetic variants were observed in the human population [1], and similar variation has since been observed across SARS-related CoVs in humans and bats. Overwhelmingly these viruses had mutations or deletions in ORF8, that have been associated with reduced replicative fitness of the virus [2]. This is also consistent with the observation that towards the end of the outbreak sequences obtained from human SARS cases possessed an ORF8 deletion that may be associated with host adaptation [1]. We therefore hypothesise that the major deletion revealed in this study may lead to an attenuated phenotype of SARS-CoV-2.


SARS-CoV-2 and ORF3a: Non-Synonymous Mutations and Polyproline Regions

Abstract: The effect of the rapid accumulation of non-synonymous mutations on the pathogenesis of SARS-CoV-2 is not yet known. To predict the impact of non-synonymous mutations and polyproline regions identified in ORF3a on the formation of B-cell epitopes and their role in evading the immune response, nucleotide and protein sequences of 537 available SARS-CoV-2 genomes were analyzed for the presence of non-synonymous mutations and polyproline regions. Mutations were correlated with changes in epitope formation. A total of 19 different non-synonymous amino acids substitutions were detected in ORF3a among 537 SARS-CoV-2 strains. G251V was the most common and identified in 9.9% (n=53) of the strains and was predicted to lead to the loss of a B-cell like epitope in ORF3a. Polyproline regions were detected in two strains (EPI_ISL_410486, France and EPI_ISL_407079, Finland) and affected epitopes formation. The accumulation of non-synonymous mutations and detected polyproline regions in ORF3a of SARS-CoV-2 could be driving the evasion of the host immune response thus favoring viral spread. Rapid mutations accumulating in ORF3a should be closely monitored throughout the COVID-19 pandemic. Importance At the surge of the COVID-19 pandemic and after three months of the identification of SARS-CoV-2 as the disease-causing pathogen, nucleic acid changes due to host-pathogen interactions are insightful into the evolution of this virus. In this paper, we have identified a set of non-synonymous mutations in ORF3a and predicted their impact on B-cell like epitope formation. The accumulation of non-synonymous mutations in ORF3a could be driving protein changes that mediate immune evasion and favoring viral spread.


Variation of 2019 novel coronavirus complete genomes recorded in the 1stmonth of outbreak: Implication for mutation


Translation-associated mutational U-pressure in the first ORF of SARS-CoV-2 and other coronaviruses

Abstract: Within four months of the ongoing COVID-19 pandemic caused by SARS-CoV-2, more than 250 nucleotide mutations have been detected in the ORF1 of the virus isolated from different parts of the globe. These observations open up an obvious question about the rate and direction of mutational pressure for further vaccine and therapeutics designing. In this study, we did a comparative analysis of ORF1a and ORF1b by using the first isolate (Wuhan strain) as the parent sequence. We observed that most of the nucleotide mutations are C to U transitions. The rate of synonymous C to U transitions is significantly higher than the rate of nonsynonymous ones, indicating negative selection on amino acid substitutions. Further, trends in nucleotide usage bias have been investigated in 49 coronaviruses species. A strong bias in nucleotide usage in fourfold degenerated sites towards uracil residues is seen in ORF1 of all the studied coronaviruses. A more substantial mutational U pressure is observed in ORF1a than in ORF1b owing to the translation of ORF1ab via programmed ribosomal frameshifting. Unlike other nucleotide mutations, mutational U pressure caused by cytosine deamination, mostly occurring in the RNA-plus strand, cannot be corrected by the proof-reading machinery of coronaviruses. The knowledge generated on the direction of mutational pressure during translation of viral RNA-plus strands has implications for vaccine and nucleoside analogue development for treating covid-19 and other coronavirus infections.


Evidence for strong mutation bias towards, and selection against, T/U content in SARS-CoV2: implications for attenuated vaccine design

Abstract: Large-scale re-engineering of synonymous sites is a promising strategy to generate attenuated viruses for vaccines. Attenuation typically relies on de-optimisation of codon pairs and maximization of CpG dinculeotide frequencies. So as to formulate evolutionarily-informed attenuation strategies, that aim to force nucleotide usage against the estimated direction favoured by selection, here we examine available whole-genome sequences of SARS-CoV2 to infer patterns of mutation and selection on synonymous sites. Analysis of mutational profiles indicates a strong mutation bias towards T with concomitant selection against T. Accounting for dinucleotide effects reinforces this conclusion, observed TT content being a quarter of that expected under neutrality. A significantly different mutational profile at CDS sites that are not 4-fold degenerate is consistent with contemporaneous selection against T mutations more widely. Although selection against CpG dinucleotides is expected to drive synonymous site G+C content below mutational equilibrium, observed G+C content is slightly above equilibrium, possibly because of selection for higher expression. Consistent with gene-specific selection against CpG dinucleotides, we observe systematic differences of CpG content between SARS-CoV2 genes. We propose an evolutionarily informed gene-bespoke approach to attenuation that, unusually, seeks to increase usage of the already most common synonymous codons. Comparable analysis of H1N1 and Ebola finds that GC3 deviated from neutral equilibrium is not a universal feature, cautioning against generalization of results.


SARS-CoV-2 mutations altering regulatory properties: deciphering host’s and virus’s perspectives

Abstract: Since the first recorded case of the SARS-CoV-2, it has acquired several mutations in its genome while spreading throughout the globe. However, apart from some changes in protein coding, functional importance of these mutations in disease pathophysiology are still largely unknown. In this study, we investigated the significance of these mutations both from the host’s and virus’s perspective by analyzing the host miRNA binding and virus’s internal ribosome entry site (IRES), respectively. Strikingly, we observed that due to the acquired mutations, host miRNAs bind differently compared to the reference; where few of the miRNAs lost and few gained the binding affinity for targeting the viral genome. Moreover, functional enrichment analysis suggests that targets of both of these gained and lost miRNAs might be involved in various host immune signaling pathways. Also, we sought to shed some insights on the impacts of mutations on the IRES structure of SARS-CoV-2. Remarkably, we detected that three particular mutations in the IRES can disrupt its secondary structure which can further make the virus less functional. These results could be valuable in exploring the functional importance of the mutations of SARS-CoV-2 and could provide novel insights into the differences observed different parts of the world.


Two mutations P/L and Y/C in SARS-CoV-2 helicase domain exist together and influence helicase RNA binding

Abstract: RNA helicases play pivotal role in RNA replication by catalysing the unwinding of complex RNA duplex structures into single strands in ATP/NTP dependent manner. SARS coronavirus 2 (SARS-CoV-2) is a single stranded positive sense RNA virus belonging to the family Coronaviridae. The viral RNA encodes non structural protein Nsp13 or the viral helicase protein that helps the viral RNA dependent RNA polymerase (RdRp) to execute RNA replication by unwinding the RNA duplexes. In this study we identified a novel mutation at position 541of the helicase where the tyrosine (Y) got substituted with cytosine (C). We found that Y541C is a destabilizing mutation increasing the molecular flexibility and leading to decreased affinity of helicase binding with RNA. Earlier we had reported a mutation P504L in the helicase protein for which had not performed RNA binding study. Here we report that P504L mutation leads to increased affinity of helicase RNA interaction. So, both these mutations have opposite effects on RNA binding. Moreover, we found a significant fraction of isolate population where both P504L and Y541C mutations were co-existing.


SARS-CoV2 envelope protein: Non-synonymous mutations and its consequences

Abstract: In the NCBI database, as on June 6, 2020, total number of available complete genome sequences of SARS-CoV2 across the world is 3617. The envelope protein of SARS-CoV2 possesses several non-synonymous mutations over the transmembrane domain and (C)-terminus in 15 (0.414%) genomes among 3617 available SARS-CoV2 genomes. More precisely, it is to be mentioned that 10(0.386%) out of 2588 genomes from the USA, 3(0.806%) from Asia, 1 (0.348%) from Europe and 1 (0.274%) from Oceania contain the missense mutations over the envelope proteins of SARS-CoV2 genomes. The C-terminus motif DLLV has been changed to DFLV and YLLV in the proteins QJR88103 (Australia: Victoria) and QKI36831 (China: Guangzhou) respectively, which might affect the binding of this motif with the host protein PALS1.


Conserved High Free Energy Sites in Human Coronavirus Spike Glycoprotein Backbones

Abstract: Methods previously developed by the author are applied to uncover several sites of interest in the spike glycoproteins of all known human coronaviruses (hCoVs), including SARS-CoV-2 that causes COVID-19. The sites comprise three-dimensional neighborhoods of peptides characterized by four key properties: (1) they pinpoint regions of high free energy in the backbone whose obstruction might interrupt function; (2) by their very definition, they occur rarely in the universe of all gene-encoded proteins that could obviate host response to compounds designed for their interference; (3) they are common to all known hCoV spikes, possibly retaining activity in light of inevitable viral mutation; and (4) they are exposed in the molecular surface of the glycoprotein. These peptides in SARS-CoV-2 are given by the triples of residues (131, 117, 134), (203, 227, 228), and (1058, 730, 731) in its spike.


An overview of the genetic variations of the SARS-CoV-2 genomes isolated in Southeast Asian countries

Abstract: Monitoring the mutation dynamics of human severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is critical in understanding its infectivity, virulence and pathogenicity for development of a vaccine. In an "age of mobility", the pandemic highlights the importance and vulnerability of regionalisation and labour market interdependence in Southeast Asia. We intend to characterise the genetic variability of viral populations within the region to provide preliminary information for regional surveillance in the future. By analysing 142 complete genomes from South East Asian (SEA) countries, we identified three central variants distinguished by nucleotide and amino acid changes.


Comparison five primer sets from different genome region of covid-1for detection of virus infection by conventional rt-pcr

Abstract: Background and Objectives: The new beta-coronavirus, which caused Severe Acute Respiratory Coronavirus-2 Syndrome (SARS-CoV-2), a major respiratory outbreak in Wuhan, China in December 2019, is now prevalent in many countries around the world Identifying PCR-based viruses is a well-known and relatively stable protocol Unfortunately, the high mutation rates may lead to widespread changes in viral nucleic acid sequences, and so using specific primers for PCR can be recom-mended In this study, we evaluated the power of a conventional RT-PCR to detect SARS-CoV-2 RNA among the five set primer sets Materials and Methods: The five genomic regions of the Coronavirus SARS-2 virus including Nucleocapsids (N), Enve-lope (E), RNA depended RNA Polymerase (RdRp), ORF1ab and Spike (S) were selected for primer designing A conventional RT-PCR was performed to compare sensitivity, specificity and other analytical characteristics of primers designed against two Real Time PCR commercial kits Results: The result of the comparative analysis showed that the ORF1ab, N and RdRp primers had a sensitivity, specificity and positive predictive value higher than other primers A significant difference in the analytical sensitivity between the studied primer sets in RT-PCR kits was observed Conclusion: In this study, the ORF1ab, Nucleocapsid and RdRp regions have the best primers for identifying the SARS-CoV-2 RNA between different genes that have been suggested


Ratcheting down the virulence of SARS-CoV-2 in the COVID-19 pandemic


Overwhelming mutations or SNPs of SARS-CoV-2: A point of caution

Abstract: The morbidity of SARS-CoV-2 (COVID-19) is reaching 3 Million landmark causing and a serious public health concern globally and it is enigmatic how several antiviral and antibody treatments were not effective in the different period across the globe. With the drastic increasing number of positive cases around the world WHO raised the importance in the assessment of the risk of spread and understanding genetic modifications that could have occurred in the SARS-CoV-2. Using all available deep sequencing data of complete genome from all over the world (NCBI repository), we identified several hundreds of point mutations or SNPs in SARS-CoV-2 all across the genome. This could be the cause for the constant change and differed virulence with an increase in mortality and morbidity. Among the 12 different countries (one sequence from each country) with complete genome sequencing data, we noted the 47 key point mutations or SNPs located along the entire genome that might have impact in the virulence and response to different antivirals against SARS-CoV-2. In this regard, key viral proteins of spike glycoprotein, Nsp1, RdRp and the ORF8 region got heavily mutated within these 3 months via person-to-person passage. We also discuss what could be the possible cause of this rapid mutation in the SARS-CoV-2.