Resolution of SARS-CoV-2 infection in human lung tissues is driven by extravascular CD163+ monocytes (preprint)

The lung-resident immune mechanisms driving resolution of SARS-CoV-2 infection in humans remain elusive. Using mice co-engrafted with a genetically matched human immune system and fetal lung xenograft (fLX), we mapped the immunological events defining resolution of SARS-CoV-2 infection in human lung tissues. Viral infection is rapidly cleared from fLX following a peak of viral replication. Acute replication results in the emergence of cell subsets enriched in viral RNA, including extravascular inflammatory monocytes (iMO) and macrophage-like T-cells, which dissipate upon infection resolution. iMO display robust antiviral responses, are transcriptomically unique among myeloid lineages, and their emergence associates with the recruitment of circulating CD4+ monocytes. Consistently, mice depleted for human CD4+ cells but not CD3+ T-cells failed to robustly clear infectious viruses and displayed signatures of chronic infection. Our findings uncover the transient differentiation of extravascular iMO from CD4+ monocytes as a major hallmark of SARS-CoV-2 infection resolution and open avenues for unravelling viral and host adaptations defining persistently active SARS-CoV-2 infection.

A Drug-Free Pathogen Capture and Neutralizing Nasal Spray to Prevent Emerging Respiratory Infections (preprint)

Respiratory infections pose a global health crisis. Vaccines are pathogen specific, and new vaccines are needed for mutants and emerging pathogens. Here, we report a drug free prophylactic platform - a Pathogen Capture and Neutralizing Spray (PCANS) that acts via a multi-pronged approach to prevent a broad spectrum of respiratory infections. PCANS forms a protective coating in the nasal cavity that enhances the capture of large respiratory droplets. The coating acts as a physical barrier against a broad spectrum of viruses and bacteria, and rapidly neutralizes them, reducing the pathogen load by >99.99%. In mice, PCANS showed nasal retention for at least 8 h and was safe for daily administration. A single prophylactic dose of PCANS protected mice against supra-lethal dosages of a mouse-adapted H1N1 Influenza virus (PR8), reduced lung viral titer by >99.99%, improved survival, and suppressed pathological manifestations. Together, our data suggest PCANS as a promising daily-use prophylactic approach against current and emerging respiratory infections.

Complete substitution with modified nucleotides suppresses the early interferon response and increases the potency of self-amplifying RNA (preprint)

Self-amplifying RNA (saRNA) will revolutionize vaccines and in situ therapeutics by enabling protein expression for longer duration at lower doses. However, a major barrier to saRNA efficacy is the potent early interferon response triggered upon cellular entry, resulting in saRNA degradation and translational inhibition. Substitution of mRNA with modified nucleotides (modNTPs), such as N1-methylpseudouridine (N1m{Psi}), reduce the interferon response and enhance expression levels. Multiple attempts to use modNTPs in saRNA have been unsuccessful, leading to the conclusion that modNTPs are incompatible with saRNA, thus hindering further development. Here, contrary to the common dogma in the field, we identify multiple modNTPs that when incorporated into saRNA at 100% substitution confer immune evasion and enhance expression potency. Transfection efficiency enhances by roughly an order of magnitude in difficult to transfect cell types compared to unmodified saRNA, and interferon production reduces by >8 fold compared to unmodified saRNA in human peripheral blood mononuclear cells (PBMCs). Furthermore, we demonstrate expression of viral antigens in vitro and observe significant protection against lethal challenge with a mouse-adapted SARS-CoV-2 strain in vivo. A modified saRNA vaccine, at 100-fold lower dose than a modified mRNA vaccine, results in a statistically improved performance to unmodified saRNA and statistically equivalent performance to modified mRNA. This discovery considerably broadens the potential scope of self-amplifying RNA, enabling entry into previously impossible cell types, as well as the potential to apply saRNA technology to non-vaccine modalities such as cell therapy and protein replacement.

Role of spike in the pathogenic and antigenic behavior of SARS-CoV-2 BA.1Omicron (preprint)

The recently identified, globally predominant SARS-CoV-2 Omicron variant (BA.1) is highly transmissible, even in fully vaccinated individuals, and causes attenuated disease compared with other major viral variants recognized to date1-7. The Omicron spike (S) protein, with an unusually large number of mutations, is considered the major driver of these phenotypes3,8. We generated chimeric recombinant SARS-CoV-2 encoding the S gene of Omicron in the backbone of an ancestral SARS-CoV-2 isolate and compared this virus with the naturally circulating Omicron variant. The Omicron S-bearing virus robustly escapes vaccine-induced humoral immunity, mainly due to mutations in the receptor-binding motif (RBM), yet unlike naturally occurring Omicron, efficiently replicates in cell lines and primary-like distal lung cells. In K18-hACE2 mice, while Omicron causes mild, non-fatal infection, the Omicron S-carrying virus inflicts severe disease with a mortality rate of 80%. This indicates that while the vaccine escape of Omicron is defined by mutations in S, major determinants of viral pathogenicity reside outside of S.

Platelet proteome analysis reveals an early hyperactive phenotype in SARS-CoV-2-infected humanized ACE2 mice (preprint)

Coronavirus disease-2019 (COVID-19) provokes a hypercoagulable state with increased incidence of thromboembolism and mortality. Platelets are major effectors of thrombosis and hemostasis. Suitable animal models are needed to better understand COVID-19-associated coagulopathy (CAC) and underlying platelet phenotypes. Here, we assessed K18-hACE2 mice undergoing a standardized SARS-CoV-2 infection protocol to study dynamic platelet responses via mass spectrometry-based proteomics. In total, we found significant changes in >1,200 proteins. Strikingly, protein alterations occurred rapidly by 2 days post-infection (dpi) and preceded outward clinical signs of severe disease. Pathway enrichment analysis of 2dpi platelet proteomes revealed that SARS-CoV-2 infection upregulated complement-coagulation networks (F2, F12, CFH, CD55/CD59), platelet activation-adhesion-degranulation proteins (PF4, SELP, PECAM1, HRG, PLG, vWF), and chemokines (CCL8, CXCL5, CXCL12). When mice started to lose weight at 4dpi, pattern recognition receptor signaling (RIG-I/MDA5, CASP8, MAPK3), and interferon pathways (IFIT1/IFIT3, STAT1) were predominant. Interestingly, SARS-CoV-2 spike protein in the lungs was observed by immunohistochemistry, but in platelets was undetected by proteomics. Similar to patients, K18-hACE2 mice during SARS-CoV-2 infection developed progressive lymphohistiocytic interstitial pneumonia with platelet aggregates in the lungs and kidneys. In conclusion, this model recapitulates activation of coagulation, complement, and interferon responses in circulating platelets, providing valuable insight into platelet pathology during COVID-19. Key PointsO_LISARS-CoV-2-infected humanized ACE2 mice recapitulate platelet reprogramming towards activation-degranulation-aggregation. C_LIO_LIComplement/coagulation pathways are dominant in platelets at 2 days post-infection (dpi), while interferon signaling is dominant at 4dpi. C_LI

Macrophages govern antiviral responses in human lung tissues protected from SARS-CoV-2 infection (preprint)

The majority of SARS-CoV-2 infections among healthy individuals result in asymptomatic to mild disease. However, the immunological mechanisms defining effective lung tissue protection from SARS-CoV-2 infection remain elusive. Unlike mice solely engrafted with human fetal lung xenograft (fLX), mice co-engrafted with fLX and a myeloid-enhanced human immune system (HNFL mice) are protected against SARS-CoV-2 infection, severe inflammation, and histopathology. Effective control of viral infection in HNFL mice associated with significant macrophage infiltration, and the induction of a potent macrophage-mediated interferon response. The pronounced upregulation of the USP18-ISG15 axis (a negative regulator of IFN responses), by macrophages was unique to HNFL mice and represented a prominent correlate of reduced inflammation and histopathology. Altogether, our work shed light on unique cellular and molecular correlates of lung tissue protection during SARS-CoV-2 infection, and underscores macrophage IFN responses as prime targets for developing immunotherapies against coronavirus respiratory diseases.

Fatal neuroinvasion of SARS-CoV-2 in K18-hACE2 mice is partially dependent on hACE2 expression (preprint)

Animal models recapitulating the distinctive features of severe COVID-19 are critical to enhance our understanding of SARS-CoV-2 pathogenesis. Transgenic mice expressing human angiotensin-converting enzyme 2 (hACE2) under the cytokeratin 18 promoter (K18-hACE2) represent a lethal model of SARS-CoV-2 infection. However, the cause(s) and mechanisms of lethality in this mouse model remain unclear. Here, we evaluated the spatiotemporal dynamics of SARS-CoV-2 infection for up to 14 days post-infection. Despite infection and moderate inflammation in the lungs, lethality was invariably associated with viral neuroinvasion and neuronal damage (including spinal motor neurons). Neuroinvasion occurred following virus transport through the olfactory neuroepithelium in a manner that was only partially dependent on hACE2. Interestingly, SARS-CoV-2 tropism was overall neither widespread among nor restricted to only ACE2-expressing cells. Although our work incites caution in the utility of the K18-hACE2 model to study global aspects of SARS-CoV-2 pathogenesis, it underscores this model as a unique platform for exploring the mechanisms of SARS-CoV-2 neuropathogenesis. SUMMARYCOVID-19 is a respiratory disease caused by SARS-CoV-2, a betacoronavirus. Here, we show that in a widely used transgenic mouse model of COVID-19, lethality is invariably associated with viral neuroinvasion and the ensuing neuronal disease, while lung inflammation remains moderate.

Using image-based haplotype alignments to map global adaptation of SARS-CoV-2 (preprint)

Quantifying evolutionary change among viral genomes is an important clinical device to track critical adaptations geographically and temporally. We built image-based haplotype-guided evolutionary inference (ImHapE) to quantify adaptations in expanding populations of non-recombining SARS-CoV-2 genomes. By combining classic population genetic summaries with image-based deep learning methods, we show that different rates of positive selection are driving evolutionary fitness and dispersal of SARS-CoV-2 globally. A 1.35-fold increase in evolutionary fitness is observed within the UK, associated with expansion of both the B.1.177 and B.1.1.7 SARS-CoV-2 lineages.

Targeting conserved viral virulence determinants by single domain antibodies to block SARS-CoV2 infectivity (preprint)

We selected SARS-CoV2 specific single domain antibodies (sdAbs) from a previously constructed phage display library using synthetic immunogenic peptides of the virus spike (S) protein as bait. The sdAbs targeting the cleavage site (CS) and the receptor binding domain (RBD) in S protein efficiently neutralised the infectivity of a pseudovirus expressing SARS-CoV2 S protein. Anti-CS sdAb blocked the virus infectivity by inhibiting proteolytic processing of SARS-CoV2 S protein. Both the sdAbs retained characteristic structure within the pH range of 2 to 12 and remained stable upto 65{degrees}C. Furthermore, structural disruptions induced by a high temperature in both the sdAbs were largely reversed upon their gradual cooling and the resulting products neutralised the reporter virus. Our results therefore suggest that targeting CS in addition to the RBD of S protein by sdAbs could serve as a viable option to reduce SARS-CoV2 infectivity and that proteolytic processing of the viral S protein is critical for infection.

In vitro screening of anti-viral and virucidal effects against SARS-CoV-2 by Hypericum perforatum and Echinacea. (preprint)

Special Infectious Agent Unit in King Fahd Medical Research Center at King Abdulaziz University, Jeddah, Saudi Arabia, has pursed the anti-viral project field to optimize the group of medicinal plants for human-infectious diseases. We have begun virtually in this field since COVID-19 pandemic, besides our divergence in the infectious agents. In this study and based on the previous review, Hypericum perforatum (St. Johns Wort) and Echinacea (gaia HERBS(R)) were tested in vitro using Vero E6 cells for their anti-viral effects against the newly identified Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) through its infectious cycle from 0 to 48 hours post infection. The hypericin (0.9 mg) of H. perforatum and the different parts (roots, seeds, aerial) of two types of Echinacea species (Echinacea purpurea and Echinacea angustifolia) were examined their efficacy in certain concentration and under light-dependent anti-viral activities to measure the inhibition of the SARS-CoV-2 mRNA expression of RNA-dependent RNA polymerase (RdRP) gene and the viral load with quantitative real-time polymerase chain reaction (qRT-PCR), and to assess the neutralization of the SARS-CoV-2 spike receptor binding on cell culture assay. Interestingly, the mixture (H.E.) of 100 mg/mL of H. perforatum and Echinacea was tested too on SARS-CoV-2 and showed crucial anti-viral activity competing H. perforatum then Echinacea effects as anti-viral treatment. Therefore, the results of gaia HERBS(R) products, H. perforatum and Echinacea species, applied in this study showed significant anti-viral and virucidal effects in the following order of potency: H. perforatum, H.E., and Echinacea on SARS-CoV-2 infectious cycle; and will definitely required a set up of clinical trial with specific therapeutic protocol based on the outcome of this study. Author SummaryAfter an outbreak of Rift Valley Fever in the Southern region of Saudi Arabia, particularly in May 2003, Special Infectious Agents Unit (SIAU) was established and founded by Prof. Esam Ibraheem Azhar. This unit contains a full range of facilities including Biosafety Level 3, allows him and his research groups to ambulate and culture risk group 3 viruses in Saudi Arabia & Gulf States for the first time. Since that time, SIAU and our international collaboration have been extended to implement a standard protocols in the infectious agents diagnostics procedure through different mode of collaboration including exchange of expertise, joint research program and more recently a technology transfer agreements with number of international institute sharing same interests. Furthermore, we have been engaged in number of researches related to Hajj & Umrah plus number of national services with the Ministry of Health (MOH) through which, we utilize our Mobile biosafety level 3 Lab to enhance the diagnostics of MERS CoV in the Holly sites during Hajj since 2014. In our SIAU and with a powerful team, we have excellent researches made valuable contributions through in vivo and in vitro animal and human studies, and several human viral pathogens which are a threat to global health security due to millions of pilgrims visiting Saudi Arabia every year from 182 countries: with particular areas of interests in: Alkhurma Viral Hemorrhagic Fever, Dengue Hemorrhagic Fever Viruses, Rift Valley Fever Virus, MERS-CoV and more recently the new global infectious diseases threat, Sever Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2).

Decomposing the sources of SARS-CoV-2 fitness variation in the United States (preprint)

The fitness of a pathogen is composite phenotype determined by many different factors influencing growth rates both within and between hosts. Determining what factors shape fitness at the host population-level is especially challenging because both intrinsic factors like pathogen genetics and extrinsic factors such as host behaviour influence between-host transmission potential. These challenges have been highlighted by controversy surrounding the population-level fitness effects of mutations in the SARS-CoV-2 genome and their relative importance when compared against non-genetic factors shaping transmission dynamics. Building upon phylodynamic birth-death models, we develop a new framework to learn how hundreds of genetic and non-genetic factors have shaped the fitness of SARS-CoV-2. We estimate the fitness effects of all amino acid variants and several structural variants that have circulated in the United States between February and September 2020 from viral phylogenies. We also estimate how much fitness variation among pathogen lineages is attributable to genetic versus non-genetic factors such as spatial heterogeneity in transmission rates. Up to September 2020, most fitness variation between lineages can be explained by background spatial heterogeneity in transmission rates across geographic regions. Furthermore, no genetic variant including the Spike D614G mutation has had a significant effect on population-level fitness. Instead, the rapid increase in the frequency of the Spike D614G can be explained by the variant having a spatial transmission advantage due to first establishing in regions with higher transmission rates during the earliest stages of the pandemic.

SARS-CoV-2 Requires Cholesterol for Viral Entry and Pathological Syncytia Formation (preprint)

Summary Many enveloped viruses induce multinucleated cells (syncytia), reflective of membrane fusion events caused by the same machinery that underlies viral entry. These syncytia are thought to facilitate replication and evasion of the host immune response. Here, we report that co-culture of human cells expressing the receptor ACE2 with cells expressing SARS-CoV-2 spike, results in synapse-like intercellular contacts that initiate cell-cell fusion, producing syncytia resembling those we identify in lungs of COVID-19 patients. To assess the mechanism of spike/ACE2-driven membrane fusion, we developed a microscopy-based, cell-cell fusion assay to screen ∼6000 drugs and >30 spike variants. Together with cell biological and biophysical approaches, the screen reveals an essential role for membrane cholesterol in spike-mediated fusion, which extends to replication-competent SARS-CoV-2 isolates. Our findings provide a molecular basis for positive outcomes reported in COVID-19 patients taking statins, and suggest new strategies for therapeutics targeting the membrane of SARS-CoV-2 and other fusogenic viruses. Highlights Cell-cell fusion at ACE2-spike clusters cause pathological syncytia in COVID-19 Drug screen reveals critical role for membrane lipid composition in fusion Spike’s unusual membrane-proximal cysteines and aromatics are essential for fusion Cholesterol tunes relative infectivity of SARS-CoV-2 viral particles

Evaluation of in vitro activity of copper gluconate against SARS-CoV-2 using confocal microscopy-based high content screening (preprint)

ContextSevere Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) that emerged late in 2019 is the etiologic agent of coronavirus disease 2019 (Covid-19). There is an urgent need to develop curative and preventive therapeutics to limit the current pandemic and to prevent the re-emergence of Covid-19. This study aimed to assess the in vitro activity of copper gluconate against SRAS-CoV-2. MethodsVero E6 cells were treated with copper gluconate 18 hours before infection. Cells were infected with a recombinant GFP expressing SARS-CoV-2. Infected cells were maintained in fresh medium containing copper gluconate for an additional 48-hour period. The infection level was measured by the confocal microscopy-based high content screening method. The cell viability in presence of copper gluconate was assessed by XTT assay. ResultsThe viability of Vero E6 cells treated with copper gluconate up to 200 M was found to be similar to that of untreated cells, but it dropped below 40% with 400 M of this agent. The infection rate was 23.8%, 18.9%, 20.6%, 6.9%, 5.3%,5.2% in cells treated with 0, 2, 10, 25, 50 and 100 M of copper gluconate respectively. As compared to untreated cells, the number of infected cells was reduced by 71%, 77%, and 78% with 25, 50, and 100 M of copper gluconate respectively (p < 0.05). ConclusionCopper gluconate was found to mitigate SARS-CoV-2 infection in Vero E6 cells. Furthers studies are needed to determine whether copper homeostasis could play a role in SARS-CoV-2 infection. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=76 SRC="FIGDIR/small/422548v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@1f99949org.highwire.dtl.DTLVardef@1bebae6org.highwire.dtl.DTLVardef@e05e37org.highwire.dtl.DTLVardef@49a3b2_HPS_FORMAT_FIGEXP M_FIG C_FIG

A novel cell culture system modeling the SARS-CoV-2 life cycle (preprint)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes the global pandemic of COVID-19, and no effective antiviral agents and vaccines are available. SARS-CoV-2 is classified as a biosafety level-3 (BLS-3) agent, impeding the basic research into its biology and the development of effective antivirals. Here, we developed a biosafety level-2 (BSL-2) cell culture system for production of transcription and replication-competent SARS-CoV-2 virus-like-particles (trVLP). This trVLP expresses a reporter gene (GFP) replacing viral nucleocapsid gene (N), which is required for viral genome packaging and virion assembly (SARS-CoV-2-GFP/{Delta}N trVLP). The complete viral life cycle can be achieved and exclusively confined in the cells ectopically expressing SARS-CoV or SARS-CoV-2 N proteins, but not MERS-CoV N. Genetic recombination of N supplied in trans into viral genome was not detected, as evidenced by sequence analysis after one-month serial passages in the N-expressing cells. Moreover, intein-mediated protein trans-splicing approach was utilized to split the viral N gene into two independent vectors, and the ligated viral N protein could function in trans to recapitulate entire viral life cycle, further securing the biosafety of this cell culture model. Based on this BSL-2 SARS-CoV-2 cell culture model, we developed a 96-well format high throughput screening for antivirals discovery. We identified salinomycin, tubeimoside I, monensin sodium, lycorine chloride and nigericin sodium as potent antivirals against SARS-CoV-2 infection. Collectively, we developed a convenient and efficient SARS-CoV-2 reverse genetics tool to dissect the virus life cycle under a BSL-2 condition. This powerful tool should accelerate our understanding of SARS-CoV-2 biology and its antiviral development.

The Potential for SARS-CoV-2 to Evade Both Natural and Vaccine-induced Immunity (preprint)

SARS-CoV-2 attaches to the surface of susceptible cells through extensive interactions between the receptor binding domain (RBD) of its spike protein and angiotensin converting enzyme type 2 (ACE2) anchored in cell membranes. To investigate whether naturally occurring mutations in the spike protein are able to prevent antibody binding, yet while maintaining the ability to bind ACE2 and viral infectivity, mutations in the spike protein identified in cases of human infection were mapped to the crystallographically-determined interfaces between the spike protein and ACE2 (PDB entry 6M0J), antibody CC12.1 (PDB entry 6XC2), and antibody P2B-2F6 (PDB entry 7BWJ). Both antibody binding interfaces partially overlap with the ACE2 binding interface. Among 16 mutations that map to the RBD:CC12.1 interface, 11 are likely to disrupt CC12.1 binding but not ACE2 binding. Among 12 mutations that map to the RBD:P2B-2F6 interface, 8 are likely to disrupt P2B-2F6 binding but not ACE2 binding. As expected, none of the mutations observed to date appear likely to disrupt the RBD:ACE2 interface. We conclude that SARS-CoV-2 with mutated forms of the spike protein may retain the ability to bind ACE2 while evading recognition by antibodies that arise in response to the original wild-type form of the spike protein. It seems likely that immune evasion will be possible regardless of whether the spike protein was encountered in the form of infectious virus, or as the immunogen in a vaccine. Therefore, it also seems likely that reinfection with a variant strain of SARS-CoV-2 may occur among people who recover from Covid-19, and that vaccines with the ability to generate antibodies against multiple variant forms of the spike protein will be necessary to protect against variant forms of SARS-CoV-2 that are already circulating in the human population.

Generation of a SARS-CoV-2 Replicon as a Model System to Dissect Virus Replication and Antiviral Inhibition (preprint)

SARS-CoV-2 research and antiviral discovery are hampered by the lack of a cell-based virus replication system that can be readily adopted without biosafety level 3 (BSL-3) restrictions. Here, the construction of a non-infectious SARS-CoV-2 reporter replicon and its application in deciphering viral replication mechanisms and evaluating SARS-CoV-2 inhibitors are presented. The replicon genome is replication competent but does not produce progeny virions. Its replication can be inhibited by RdRp mutations or by known SARS-CoV-2 antiviral compounds. Using this system, a high-throughput antiviral assay has also been developed. Significant differences in potencies of several SARS-CoV-2 inhibitors in different cell lines were observed, which highlights the challenges of discovering antivirals capable of inhibiting viral replication in vivo and the importance of testing compounds in multiple cell culture models. The generation of a SARS-CoV-2 replicon provides a powerful platform to expand the global research effort to combat COVID-19.

SARS-CoV-2 RNA reverse-transcribed and integrated into the human genome (preprint)

Prolonged SARS-CoV-2 RNA shedding and recurrence of PCR-positive tests have been widely reported in patients after recovery, yet these patients most commonly are non-infectious1-14. Here we investigated the possibility that SARS-CoV-2 RNAs can be reverse-transcribed and integrated into the human genome and that transcription of the integrated sequences might account for PCR-positive tests. In support of this hypothesis, we found chimeric transcripts consisting of viral fused to cellular sequences in published data sets of SARS-CoV-2 infected cultured cells and primary cells of patients, consistent with the transcription of viral sequences integrated into the genome. To experimentally corroborate the possibility of viral retro-integration, we describe evidence that SARS-CoV-2 RNAs can be reverse transcribed in human cells by reverse transcriptase (RT) from LINE-1 elements or by HIV-1 RT, and that these DNA sequences can be integrated into the cell genome and subsequently be transcribed. Human endogenous LINE-1 expression was induced upon SARS-CoV-2 infection or by cytokine exposure in cultured cells, suggesting a molecular mechanism for SARS-CoV-2 retro-integration in patients. This novel feature of SARS-CoV-2 infection may explain why patients can continue to produce viral RNA after recovery and suggests a new aspect of RNA virus replication.

SARS-CoV-2 desensitizes host cells to interferon through inhibition of the JAK-STAT pathway (preprint)

SARS-CoV-2 can infect multiple organs, including lung, intestine, kidney, heart, liver, and brain. The molecular details of how the virus navigates through diverse cellular environments and establishes replication are poorly defined. Here, we performed global proteomic analysis of the virus-host interface in a newly established panel of phenotypically diverse, SARS-CoV-2-infectable human cell lines representing different body organs. This revealed universal inhibition of interferon signaling across cell types following SARS-CoV-2 infection. We performed systematic analyses of the JAK-STAT pathway in a broad range of cellular systems, including immortalized cell lines and primary-like cardiomyocytes, and found that several pathway components were targeted by SARS-CoV-2 leading to cellular desensitization to interferon. These findings indicate that the suppression of interferon signaling is a mechanism widely used by SARS-CoV-2 in diverse tissues to evade antiviral innate immunity, and that targeting the viral mediators of immune evasion may help block virus replication in patients with COVID-19.

In vitro assessment of the virucidal activity of four mouthwashes containing Cetylpyridinium Chloride, ethanol, zinc and a mix of enzyme and proteins against a human coronavirus (preprint)

Background: saliva is established to contain high counts SARS-CoV-2 virus and contact with saliva droplets, contaminated surfaces or airborne particles are sources of viral transmission. The generation of infective aerosols during clinical procedures is of particular concern. Therefore, a fuller understanding of the potential of mouthwash to reduce viral counts and modulate the risk of transmission in medical professional and public context is an important research topic. Method: we determined the virucidal activity of four anti-bacterial mouthwashes against a surrogate for SARS-CoV-2, Human CoV-SARS 229E, using a standard ASTM suspension test, with dilution and contact times applicable to recommended mouthwash use. Results: the mouthwash formulated with 0.07% Cetylpyridinium Chloride exhibited virucidal effects providing a [≥]3.0 log reduction HCoV-229E viral count. Mouthwashes containing 15.7% ethanol, 0.2% zinc sulphate heptahydrate and a mix of enzymes and proteins did not demonstrate substantive virucidal activity in this test. Conclusion: mouthwash containing 0.07% Cetylpyridinium Chloride warrants further laboratory and clinical assessment to determine their potential benefit in reducing the risk of SARS-CoV-2.

Ethacridine inhibits SARS-CoV-2 by inactivating viral particles in cellular models (preprint)

More than a million people have now died from COVID-19, because of infection with the SARS-CoV-2 coronavirus. Currently, the FDA has approved remdesivir, an inhibitor of SARS-CoV-2 replication, to treat COVID-19, though very recent data from WHO showed little if any COVID19 protective effect. Here we report that ethacridine, a safe and potent antiseptic use in humans, effectively inhibits SARS-CoV-2, at very low concentrations (EC50 ~ 0.08 M). Ethacridine was identified through a high-throughput screening of an FDA-approved drug library in living cells using a fluorescent assay. Interestingly, the main mode of action of ethacridine is to inactivate virus particles, preventing binding to the host cells. Thus, our work has identified a potent drug with a distinct mode of action against SARS-CoV-2.