Pathogenic Human Coronaviruses Research Articles

Unveiling the Multifaceted Capabilities of Endophytic Aspergillus flavus Isolated from Annona squamosa Fruit Peels against Staphylococcus Isolates and HCoV 229E-In Vitro and In Silico Investigations.

Recently, there has been a surge towards searching for primitive treatment strategies to discover novel therapeutic approaches against multi-drug-resistant pathogens. Endophytes are considered unexplored yet perpetual sources of several secondary metabolites with therapeutic significance. This study aims to isolate and identify the endophytic fungi from Annona squamosa L. fruit peels using morphological, microscopical, and transcribed spacer (ITS-rDNA) sequence analysis; extract the fungus's secondary metabolites by ethyl acetate; investigate the chemical profile using UPLC/MS; and evaluate the potential antibacterial, antibiofilm, and antiviral activities. An endophytic fungus was isolated and identified as Aspergillus flavus L. from the fruit peels. The UPLC/MS revealed seven compounds with various chemical classes. The antimicrobial activity of the fungal ethyl acetate extract (FEA) was investigated against different Gram-positive and Gram-negative standard strains, in addition to resistant clinical isolates using the agar diffusion method. The CPE-inhibition assay was used to identify the potential antiviral activity of the crude fungal extract against low pathogenic human coronavirus (HCoV 229E). Selective Gram-positive antibacterial and antibiofilm activities were evident, demonstrating pronounced efficacy against both methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive Staphylococcus aureus (MSSA). However, the extract exhibited very weak activity against Gram-negative bacterial strains. The ethyl acetate extract of Aspergillus flavus L exhibited an interesting antiviral activity with a half maximal inhibitory concentration (IC50) value of 27.2 µg/mL against HCoV 229E. Furthermore, in silico virtual molecular docking-coupled dynamics simulation highlighted the promising affinity of the identified metabolite, orienting towards three MRSA biotargets and HCoV 229E main protease as compared to reported reference inhibitors/substrates. Finally, ADME analysis was conducted to evaluate the potential oral bioavailability of the identified metabolites.

Low pathogenic human coronaviruses during the first waves of COVID-19 in Italy.

Low-pathogenic human coronaviruses (HCoVs) infect the upper respiratory tract and cause mild, cold-like respiratory illness. Although several studies have shown evidence of the global distribution of HCoVs, information about their distribution in Italy are often focused only on hospitalized children and elderly with respiratory symptoms. In this study, a total of 916 swab samples collected during the first two SARS-CoV-2 pandemic waves in Abruzzo region (central Italy) was selected for molecular screening of low pathogenic HCoVs by real-time RT-PCR. We identified low-pathogenic HCoV in nine samples. Positive samples underwent whole genome sequencing for genome characterization; indeed, we also report the whole genome sequence of a HCoV-229E strain.

Tubeimosides are pan-coronavirus and filovirus inhibitors that can block their fusion protein binding to Niemann-Pick C1.

SARS-CoV-2 and filovirus enter cells via the cell surface angiotensin-converting enzyme 2 (ACE2) or the late-endosome Niemann-Pick C1 (NPC1) as a receptor. Here, we screened 974 natural compounds and identified Tubeimosides I, II, and III as pan-coronavirus and filovirus entry inhibitors that target NPC1. Using in-silico, biochemical, and genomic approaches, we provide evidence that NPC1 also binds SARS-CoV-2 spike (S) protein on the receptor-binding domain (RBD), which is blocked by Tubeimosides. Importantly, NPC1 strongly promotes productive SARS-CoV-2 entry, which we propose is due to its influence on fusion in late endosomes. The Tubeimosides' antiviral activity and NPC1 function are further confirmed by infection with SARS-CoV-2 variants of concern (VOC), SARS-CoV, and MERS-CoV. Thus, NPC1 is a critical entry co-factor for highly pathogenic human coronaviruses (HCoVs) in the late endosomes, and Tubeimosides hold promise as a new countermeasure for these HCoVs and filoviruses.

ACE2-Fc and DPP4-Fc decoy receptors against SARS-CoV-2 and MERS-CoV variants: a quick therapeutic option for current and future coronaviruses outbreaks.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) and the Middle East respiratory syndrome coronavirus (MERS-CoV) are highly pathogenic human coronaviruses (CoVs). Anti-CoVs mAbs and vaccines may be effective, but the emergence of neutralization escape variants is inevitable. Angiotensin-converting enzyme 2 and dipeptidyl peptidase 4 enzyme are the getaway receptors for SARS-CoV-2 and MERS-CoV, respectively. Thus, we reformatted these receptors as Fc-fusion decoy receptors. Then, we tested them in parallel with anti-SARS-CoV (ab1-IgG) and anti-MERS-CoV (M336-IgG) mAbs against several variants using pseudovirus neutralization assay. The generated Fc-based decoy receptors exhibited a strong inhibitory effect against all pseudotyped CoVs. Results showed that although mAbs can be effective antiviral drugs, they might rapidly lose their efficacy against highly mutated viruses. We suggest that receptor traps can be engineered as Fc-fusion proteins for highly mutating viruses with known entry receptors, for a faster and effective therapeutic response even against virus harboring antibodies escape mutations.

Comparative transcriptome analysis of SARS-CoV-2, SARS-CoV, MERS-CoV, and HCoV-229E identifying potential IFN/ISGs targets for inhibiting virus replication.

Since its outbreak in December 2019, SARS-CoV-2 has spread rapidly across the world, posing significant threats and challenges to global public health. SARS-CoV-2, together with SARS-CoV and MERS-CoV, is a highly pathogenic coronavirus that contributes to fatal pneumonia. Understanding the similarities and differences at the transcriptome level between SARS-CoV-2, SARS-CoV, as well as MERS-CoV is critical for developing effective strategies against these viruses. In this article, we comparatively analyzed publicly available transcriptome data of human cell lines infected with highly pathogenic SARS-CoV-2, SARS-CoV, MERS-CoV, and lowly pathogenic HCoV-229E. The host gene expression profiles during human coronavirus (HCoV) infections were generated, and the pathways and biological functions involved in immune responses, antiviral efficacy, and organ damage were intensively elucidated. Our results indicated that SARS-CoV-2 induced a stronger immune response versus the other two highly pathogenic HCoVs. Specifically, SARS-CoV-2 induced robust type I and type III IFN responses, marked by higher upregulation of type I and type III IFNs, as well as numerous interferon-stimulated genes (ISGs). Further Ingenuity Pathway Analysis (IPA) revealed the important role of ISGs for impeding SARS-CoV-2 infection, and the interferon/ISGs could be potential targets for therapeutic interventions. Moreover, our results uncovered that SARS-CoV-2 infection was linked to an enhanced risk of multi-organ toxicity in contrast to the other two highly pathogenic HCoVs. These findings provided valuable insights into the pathogenic mechanism of SARS-CoV-2, which showed a similar pathological feature but a lower fatality rate compared to SARS-CoV and MERS-CoV.

Broad-spectrum antiviral activity of two structurally analogous CYP3A inhibitors against pathogenic human coronaviruses in vitro

Coronaviruses pose a permanent risk of outbreaks, with three highly pathogenic species and strains (SARS-CoV, MERS-CoV, SARS-CoV-2) having emerged in the last twenty years. Limited antiviral therapies are currently available and their efficacy in randomized clinical trials enrolling SARS-CoV-2 patients has not been consistent, highlighting the need for more potent treatments. We previously showed that cobicistat, a clinically approved inhibitor of Cytochrome P450-3A (CYP3A), has direct antiviral activity against early circulating SARS-CoV-2 strains in vitro and in Syrian hamsters. Cobicistat is a derivative of ritonavir, which is co-administered as pharmacoenhancer with the SARS-CoV-2 protease inhibitor nirmatrelvir, to inhibit its metabolization by CPY3A and preserve its antiviral efficacy. Here, we used automated image analysis for a screening and parallel comparison of the anti-coronavirus effects of cobicistat and ritonavir. Our data show that both drugs display antiviral activity at low micromolar concentrations against multiple SARS-CoV-2 variants in vitro, including epidemiologically relevant Omicron subvariants. Despite their close structural similarity, we found that cobicistat is more potent than ritonavir, as shown by significantly lower EC50 values in monotherapy and higher levels of viral suppression when used in combination with nirmatrelvir. Finally, we show that the antiviral activity of both cobicistat and ritonavir is maintained against other human coronaviruses, including HCoV-229E and the highly pathogenic MERS-CoV. Overall, our results demonstrate that cobicistat has more potent anti-coronavirus activity than ritonavir and suggest that dose adjustments could pave the way to the use of both drugs as broad-spectrum antivirals against highly pathogenic human coronaviruses.

Exploring the Fatty Acid Binding Pocket in the SARS-CoV-2 Spike Protein - Confirmed and Potential Ligands.

Severe Acute Respiratory syndrome 2 (SARS-CoV-2) is a respiratory virus responsible for coronavirus disease 19 (COVID-19) and the still ongoing and unprecedented global pandemic. The key viral protein for cell infection is the spike glycoprotein, a surface-exposed fusion protein that both recognizes and mediates entry into host cells. Within the spike glycoprotein, a fatty acid binding pocket (FABP) was confirmed, with the crystallization of linoleic acid (LA) occupying a well-defined site. Importantly, when the pocket is occupied by a fatty acid, an inactive conformation is stabilized, and cell recognition is hindered. In this review, we discuss ligands reported so far for this site, correlating their activity predicted through in silico studies with antispike experimental activity, assessed by either binding assays or cell-infection assays. LA was the first confirmed ligand, cocrystallized in a cryo-EM structure of the spike protein, resulting in increased stability of the inactive conformation of the spike protein. The next identified ligand, lifitegrast, was also experimentally confirmed as a ligand with antiviral activity, suggesting the potential for diverse chemical scaffolds to bind this site. Finally, SPC-14 was also confirmed as a ligand, although no inhibition assays were performed. In this review, we identified 20 studies describing small-molecule compounds predicted to bind the pocket in in silico studies and with confirmed binding or in vitro activity, either inhibitory activity against the spike-ACE2 interaction or antiviral activity in cell-based assays. When considering all ligands confirmed with in vitro assays, a good overall occupation of the pocket should be complemented with the ability to make direct interactions, both hydrophilic and hydrophobic, with key amino acid residues defining the pocket surface. Among the active compounds, long flexible carbon chains are recurrent, with retinoids capable of binding the FABP, although bulkier systems are also capable of affecting viral fitness. Compounds able to bind this site with high affinity have the potential to stabilize the inactive conformation of the SARS-CoV-2 spike protein and therefore reduce the virus's ability to infect new cells. Since this pocket is conserved in highly pathogenic human coronaviruses, including MERS-CoV and SARS-CoV, this effect could be exploited for the development of new antiviral agents, with broad-spectrum anticoronavirus activity.

Inactivation of human coronaviruses using an automated room disinfection device

The emergence of more virulent and epidemic strains of viruses, especially in the context of COVID-19, makes it more important than ever to improve methods of decontamination. The objective of this study was to evaluate the potential of on-demand production of chlorine species to inactivate human coronaviruses. The commercial prototype disinfection unit was provided by Unipolar Water Technologies. The Unipolar device generates active chlorine species using an electrochemical reaction and dispenses the disinfectant vapour onto surfaces with an aspirator. The minimum effective concentration and exposure time of disinfectant were evaluated on human hepatoma (Huh7) cells using 50% tissue culture infectious dose (TCID50) assay and human coronavirus 229E (HCoV-229E), a surrogate for pathogenic human coronaviruses. We showed that chlorine species generated in the Unipolar device inactivate HCoV-229E on glass surfaces at ≥ 400 parts per million active chlorine concentration with a 5 min exposure time. Here, inactivation refers to the inability of the virus to infect the Huh7 cells. Importantly, no toxic effect was observed on Huh7 cells for any of the active chlorine concentrations and contact times tested.

PULMONARY REHABILITATION IN POST COVID SEQUELAE IN INDIA

An outbreak of pneumonia was reported in Wuhan, Hubei Province, China in December 2019 and quickly spread to all Chinese provinces. 1, 2 The coronavirus disease (COVID-19) pandemic spread through 190 countries within 20 weeks from Wuhan, China, affecting 334,000 populations. This caused more than 14,500 deaths by mid-March 2020. The number of infected people has risen to 1,210,956 in the month of April 6, 2020, with 67,594 associated expirations. 4 India recorded its first COVID-19 infection case on January 30, 2020 in a student from China's capital, Wuhan. In 2020, India reported 360 positive COVID-19 cases from 23 states across the country. 4 in India, the number of asymptomatic cases was higher. On April 4, 2019, a novel coronavirus pathogen was proved to be a novel betacoronavirus. 2 Sub-micron sized viruses, narrated as "organisms at the edge of life", are liable for some of the worst human pandemics in history, like influenza (1918), smallpox (1978), and COVID-19, as of September 17, 2020. 2 The SARS-CoV-2 virus is causative for COVID-19. Nearly 29 million people have been affected by this virus, which caused 936,521 deaths worldwide (WHO, 2020). 2 Corona viruses are single-stranded positive-strand RNA viruses that are known to contain some of the largest viral genomes, up to around 32 kbp in length. Following an augmentation in the number of coronavirus genome sequences available and following efforts to investigate the diversity in the wild, the family Corona viridae now contains four genera. The structure of the coronavirus as given in figure 1 Those species that belong to the genera Alpha corona virus and Beta corona virus can infect mammalian hosts, while those of Gamma corona virus and the recently defined Delta corona virus mainly infect avian species. 3 The highly pathogenic human coronaviruses, Severe Acute Respiratory Syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome coronavirus (MERS-CoV), belong to lineage B (sub-genus Sarbecovirus) and lineage C (sub-genus Merbecovirus) of Betacoronavirus. 3

Contrasting roles of MERS-CoV and SARS-CoV-2 internal proteins in pathogenesis in mice.

Betacoronaviruses encode an internal (I) gene via an alternative reading frame within the nucleocapsid gene, called ORF8b for Middle-East respiratory syndrome coronavirus (MERS-CoV) and ORF9b for severe acute respiratory syndrome coronavirus (SARS-CoV) and SARS-CoV-2. Previous reports suggested that proteins 8b and 9b are involved in evading multiple innate immune signaling pathways. However, their roles in mediating pathogenesis in infected animals have not been determined. In this study, we abrogated the expression of protein 8b in MERS-CoV and protein 9b in SARS-CoV-2. Using mouse models of MERS-CoV and SARS-CoV-2 infection, we found that MERS-CoV lacking protein 8b expression was more virulent, while SARS-CoV-2 lacking protein 9b expression was attenuated compared with the respective wild-type viruses. Upon further analysis, we detected increased levels of type I interferon and enhanced infiltration of immune cells to the lungs of mice infected with MERS-CoV lacking protein 8b expression. These data suggest that the I protein of MERS-CoV plays a role in limiting pathogenesis while that of SARS-CoV-2 enhances disease severity. IMPORTANCE The function of betacoronavirus internal protein has been relatively understudied. The earliest report on the internal protein of mouse hepatitis virus suggested that the internal protein is a structural protein without significant functions in virus replication and virulence. However, the internal proteins of severe acute respiratory syndrome coronavirus (SARS-CoV), Middle-East respiratory syndrome coronavirus, and SARS-CoV-2 have been shown to evade immune responses. Despite the reported functions of the internal protein in these highly pathogenic human coronaviruses, its role in mediating pathogenesis in experimentally infected animals has not been characterized. Our data indicated that despite the similar genomic location and expression strategy of these internal proteins, their effects on virulence are vastly different and virus specific, highlighting the complexity between host-virus interaction and disease outcome.

Vaccine-mediated protection against Merbecovirus and Sarbecovirus challenge in mice

The emergence of three highly pathogenic human coronaviruses-severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003, Middle Eastern respiratory syndrome (MERS)-CoV in 2012, and SARS-CoV-2 in 2019-underlines the need to develop broadly active vaccines against the Merbecovirus and Sarbecovirus betacoronavirus subgenera. While SARS-CoV-2 vaccines protect against severe COVID-19, they do not protect against other sarbecoviruses or merbecoviruses. Here, we vaccinate mice with a trivalent sortase-conjugate nanoparticle (scNP) vaccine containing the SARS-CoV-2, RsSHC014, and MERS-CoV receptor-binding domains (RBDs), which elicited live-virus neutralizing antibody responses. The trivalent RBD scNP elicited serum neutralizing antibodies against bat zoonotic Wuhan Institute of Virology-1 (WIV-1)-CoV, SARS-CoV, SARS-CoV-2 BA.1, SARS-CoV-2 XBB.1.5, and MERS-CoV live viruses. The monovalent SARS-CoV-2 RBD scNP vaccine only protected against Sarbecovirus challenge, whereas the trivalent RBD scNP vaccine protected against both Merbecovirus and Sarbecovirus challenge in highly pathogenic and lethal mouse models. This study demonstrates proof of concept for a single pan-sarbecovirus/pan-merbecovirus vaccine that protects against three highly pathogenic human coronaviruses spanning two betacoronavirus subgenera.

Improved Culture Methods for Human Coronaviruses HCoV-OC43, HCoV-229E, and HCoV-NL63.

HCoV-OC43, HCoV-229E, HCoV-NL63, and HCoV-HKU1 are four of the seven known human coronaviruses (HCoVs) and, unlike the highly pathogenic SARS-CoV, MERS-CoV, and SARS-CoV-2, these four so-called seasonal HCoVs generally cause mild upper-respiratory-tract illness. As Biosafety Level 2 (BSL-2) pathogens, the seasonal HCoVs are more accessible and can be used as surrogates for studying the highly pathogenic HCoVs. However, scientists have for many years found these difficult to study because of the lack of a universal culture system and the inability of typical culture methods to yield high-titer infectious stocks. We have developed assays to grow and quantify infectious virus and viral RNA for HCoV-OC43, -229E, and -NL63. We identified which immortalized cell lines should be used to optimize the replication of HCoV-OC43, -229E, and -NL63 in order to generate high titers (Vero E6, Huh-7, and LLC-MK2 cells, respectively). Here we present protocols for improved propagation and quantification of each seasonal HCoV. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Growth of HCoVs Basic Protocol 2: Quantification of HCoV by plaque assay Basic Protocol 3: Quantification of HCoV RNA products of replication Basic Protocol 4: Concentrating HCoVs via ultracentrifugation.

Infection of Primary Nasal Epithelial Cells Grown at an Air-Liquid Interface to Characterize Human Coronavirus-Host Interactions.

Three highly pathogenic human coronaviruses (HCoVs) - SARS-CoV (2002), MERS-CoV (2012), and SARS-CoV-2 (2019) - have emerged and caused significant public health crises in the past 20 years. Four additional HCoVs cause a significant portion of common cold cases each year (HCoV-NL63, -229E, -OC43, and -HKU1), highlighting the importance of studying these viruses in physiologically relevant systems. HCoVs enter the respiratory tract and establish infection in the nasal epithelium, the primary site encountered by all respiratory pathogens. We use a primary nasal epithelial culture system in which patient-derived nasal samples are grown at an air-liquid interface (ALI) to study host-pathogen interactions at this important sentinel site. These cultures recapitulate many features of the in vivo airway, includingthe cell types present, ciliary function, and mucus production. We describe methods to characterize viral replication, host cell tropism, virus-induced cytotoxicity, and innate immune induction in nasal ALI cultures following HCoV infection, using recent work comparing lethal and seasonal HCoVs as an example1. An increased understanding of host-pathogen interactions in the nose has the potential to provide novel targets for antiviral therapeutics against HCoVs and other respiratory viruses that will likely emerge in the future.

Non-Structural Proteins (Nsp): A Marker for Detection of Human Coronavirus Families.

SARS-CoV-2 was the cause of the global pandemic that caused a total of 14.9 million deaths during the years 2020 and 2021, according to the WHO. The virus presents a mutation rate between 10-5 and 10-3 substitutions per nucleotide site per cell infection (s/n/c). Due to this, studies aimed at knowing the evolution of this virus could help us to foresee (through the future development of new detection strategies and vaccines that prevent the infection of this virus in human hosts) that a pandemic caused by this virus will be generated again. In this research, we performed a functional annotation and identification of changes in Nsp (non-structural proteins) domains in the coronavirus genome. The comparison of the 13 selected coronavirus pangenomes demonstrated a total of 69 protein families and 57 functions associated with the structural domain's differentials between genomes. A marked evolutionary conservation of non-structural proteins was observed. This allowed us to identify and classify highly pathogenic human coronaviruses into alpha, beta, gamma, and delta groups. The designed Nsp cluster provides insight into the trajectory of SARS-CoV-2, demonstrating that it continues to evolve rapidly. An evolutionary marker allows us to discriminate between phylogenetically divergent groups, viral genotypes, and variants between the alpha and betacoronavirus genera. These types of evolutionary studies provide a window of opportunity to use these Nsp as targets of viral therapies.

Regulation of coronavirus nsp15 cleavage specificity by RNA structure.

SARS-CoV-2, the etiologic agent of the COVID-19 pandemic, has had an enduring impact on global public health. However, SARS-CoV-2 is only one of multiple pathogenic human coronaviruses (CoVs) to have emerged since the turn of the century. CoVs encode for several nonstructural proteins (nsps) that are essential for viral replication and pathogenesis. Among them is nsp15, a uridine-specific viral endonuclease that is important in evading the host immune response and promoting viral replication. Despite the established endonuclease function of nsp15, little is known about other determinants of its cleavage specificity. In this study we investigate the role of RNA secondary structure in SARS-CoV-2 nsp15 endonuclease activity. Using a series of in vitro endonuclease assays, we observed that thermodynamically stable RNA structures were protected from nsp15 cleavage relative to RNAs lacking stable structure. We leveraged the s2m RNA from the SARS-CoV-1 3'UTR as a model for our structural studies as it adopts a well-defined structure with several uridines, two of which are unpaired and thus highly probable targets for nsp15 cleavage. We found that SARS-CoV-2 nsp15 specifically cleaves s2m at the unpaired uridine within the GNRNA pentaloop of the RNA. Further investigation revealed that the position of uridine within the pentaloop also impacted nsp15 cleavage efficiency suggesting that positioning within the pentaloop is necessary for optimal presentation of the scissile uridine and alignment within the nsp15 catalytic pocket. Our findings indicate that RNA secondary structure is an important determinant of nsp15 cleavage and provides insight into the molecular mechanisms of RNA recognition by nsp15.

IgG Fc-Binding Peptide-Conjugated Pan-CoV Fusion Inhibitor Exhibits Extended In Vivo Half-Life and Synergistic Antiviral Effect When Combined with Neutralizing Antibodies.

The peptide-based pan-coronavirus fusion inhibitor EK1 is in phase III clinical trials, and it has, thus far, shown good clinical application prospects against SARS-CoV-2 and its variants. To further improve its in vivo long-acting property, we herein developed an Fc-binding strategy by conjugating EK1 with human immunoglobulin G Fc-binding peptide (IBP), which can exploit the long half-life advantage of IgG in vivo. The newly engineered peptide IBP-EK1 showed potent and broad-spectrum inhibitory activity against SARS-CoV-2 and its variants, including various Omicron sublineages and other human coronaviruses (HCoVs) with low cytotoxicity. In mouse models, IBP-EK1 possessed potent prophylactic and therapeutic efficacy against lethal HCoV-OC43 challenge, and it showed good safety profile and low immunogenicity. More importantly, IBP-EK1 exhibited a significantly extended in vivo half-life in rhesus monkeys of up to 37.7 h, which is about 20-fold longer than that reported for EK1. Strikingly, IBP-EK1 displayed strong in vitro or ex vivo synergistic anti-HCoV effect when combined with monoclonal neutralizing antibodies, including REGN10933 or S309, suggesting that IBP-conjugated EK1 can be further developed as a long-acting, broad-spectrum anti-HCoV agent, either alone or in combination with neutralizing antibodies, to combat the current COVID-19 pandemic or future outbreaks caused by emerging and re-emerging highly pathogenic HCoVs.

An engineered recombinant protein containing three structural domains in SARS-CoV-2 S2 protein has potential to act as a pan-human coronavirus entry inhibitor or vaccine antigen

ABSTRACT The threat to global health caused by three highly pathogenic human coronaviruses (HCoV), SARS-CoV-2, MERS-CoV and SARS-CoV, calls for the development of pan-HCoV therapeutics and vaccines. This study reports the design and engineering of a recombinant protein designated HR1LS. It contains three linked molecules, each consisting of three structural domains, including a heptad repeat 1 (HR1), a central helix (CH), and a stem helix (SH) region, in the S2 subunit of SARS-CoV-2 spike (S) protein. It was found that HR1LS protein automatically formed a trimer able to bind with heptad repeat 2 (HR2) region in the SARS-CoV-2 S2 subunit, thus potently inhibiting HCoV fusion and entry into host cells. Furthermore, immunization of mice with HR1LS, when combined with CF501 adjuvant, resulted in the production of neutralizing antibodies against infection of SARS-CoV-2 and its variants, as well as SARS-CoV, MERS-CoV, HCoV-229E, HCoV-NL63 and MjHKU4r-CoV-1. These results suggest that HR1LS is a promising candidate for further development as a novel HR1-trimer-based pan-HCoV entry inhibitor or vaccine for the treatment and prevention of infection by SARS-CoV-2 and its variants, but also other HCoVs with the potential to cause future emerging and re-emerging infectious coronavirus diseases.

DYRK1A promotes viral entry of highly pathogenic human coronaviruses in a kinase-independent manner.

Identifying host genes essential for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has the potential to reveal novel drug targets and further our understanding of Coronavirus Disease 2019 (COVID-19). We previously performed a genome-wide CRISPR/Cas9 screen to identify proviral host factors for highly pathogenic human coronaviruses. Few host factors were required by diverse coronaviruses across multiple cell types, but DYRK1A was one such exception. Although its role in coronavirus infection was previously undescribed, DYRK1A encodes Dual Specificity Tyrosine Phosphorylation Regulated Kinase 1A and is known to regulate cell proliferation and neuronal development. Here, we demonstrate that DYRK1A regulates ACE2 and DPP4 transcription independent of its catalytic kinase function to support SARS-CoV, SARS-CoV-2, and Middle East Respiratory Syndrome Coronavirus (MERS-CoV) entry. We show that DYRK1A promotes DNA accessibility at the ACE2 promoter and a putative distal enhancer, facilitating transcription and gene expression. Finally, we validate that the proviral activity of DYRK1A is conserved across species using cells of nonhuman primate and human origin. In summary, we report that DYRK1A is a novel regulator of ACE2 and DPP4 expression that may dictate susceptibility to multiple highly pathogenic human coronaviruses.

Epidemiological studies of COVID-19 disease

The Chinese population having epidemiological link with wholesale market of seafood develop respiratory illness with pneumonia like clinical presentations in Hubei Province, Wuhan City, recently in December 2019. Laboratory results revealed a novel coronavirus that was named severe acute respiratory syndrome coronavirus-2 (SARS CoV-2) and disease it caused given name of coronavirus disease 2019 (COVID-19). COVID-19 turn out to be third outbreak of human pathogenic coronaviruses and first pandemic of 21st century. The previous coronaviruses proved merely as a tip of iceberg after emergence of recently identified SARS-CoV-2 with potential of pandemic status that significantly revealed the concealed capabilities of virulence and contagiousness of betacoronaviruses group. The aim of this review is to discuss the origin of COVID-19, transmission patterns, susceptible hosts, factors that increase disease severity, nucleic acid base diagnosis, mortality and morbidity rates and therapeutic option against COVID-19. The diagnostic methods like microarray and RNA-targeting CRISPR were adopted recently for COVID-19 diagnosis after advancing them further for rapid and accurate detection of COVID-19 and high throughput.

Antiviral activity of marine sulfated glycans against pathogenic human coronaviruses

Great interest exists towards the discovery and development of broad-spectrum antivirals. This occurs due to the frequent emergence of new viruses which can also eventually lead to pandemics. A reasonable and efficient strategy to develop new broad-spectrum antivirals relies on targeting a common molecular player of various viruses. Heparan sulfate is a sulfated glycosaminoglycan present on the surface of cells which plays a key role as co-receptor in many virus infections. In previous work, marine sulfated glycans (MSGs) were identified as having antiviral activities. Their mechanism of action relies primarily on competitive inhibition of virion binding to heparan sulfate, preventing virus attachment to the cell surface prior to entry. In the current work we used pseudotyped lentivirus particles to investigate in a comparative fashion the inhibitory properties of five structurally defined MSGs against SARS-CoV-1, SARS-CoV-2, MERS-CoV, and influenza A virus (IAV). MSGs include the disaccharide-repeating sulfated galactan from the red alga Botryocladia occidentalis, the tetrasaccharide-repeating sulfated fucans from the sea urchin Lytechinus variegatus and from the sea cucumber Isostichopus badionotus, and the two marine fucosylated chondroitin sulfates from the sea cucumbers I. badionotus and Pentacta pygmaea. Results indicate specificity of action against SARS-CoV-1 and SARS-CoV-2. Curiously, the MSGs showed decreased inhibitory potencies against MERS-CoV and negligible action against IAV. Among the five MSGs, the two sulfated fucans here studied deserve further attention since they have the lowest anticoagulant effects but still present potent and selective antiviral properties.