Phillyrin (KD-1) exerts anti-viral and anti-inflammatory activities against novel coronavirus (SARS-CoV-2) and human coronavirus 229E (HCoV-229E) by suppressing the nuclear factor kappa B (NF-κB) signaling pathway.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has extensively and rapidly spread in the world, causing an outbreak of acute infectious pneumonia. However, no specific antiviral drugs or vaccines can be used. Phillyrin (KD-1), a representative ingredient of Forsythia suspensa, possesses anti-inflammatory, anti-oxidant, and antiviral activities. However, little is known about the antiviral abilities and mechanism of KD-1 against SARS-CoV-2 and human coronavirus 229E (HCoV-229E). PURPOSE: The study was designed to investigate the antiviral and anti-inflammatory activities of KD-1 against the novel SARS-CoV-2 and HCoV-229E and its potential effect in regulating host immune response in vitro. METHODS: The antiviral activities of KD-1 against SARS-CoV-2 and HCoV-229E were assessed in Vero E6 cells using cytopathic effect and plaque-reduction assay. Proinflammatory cytokine expression levels upon infection with SARS-CoV-2 and HCoV-229E infection in Huh-7 cells were measured by real-time quantitative PCR assays. Western blot assay was used to determine the protein expression of nuclear factor kappa B (NF-κB) p65, p-NF-κB p65, IκBα, and p-IκBα in Huh-7 cells, which are the key targets of the NF-κB pathway. RESULTS: KD-1 could significantly inhibit SARS-CoV-2 and HCoV-229E replication in vitro. KD-1 could also markedly reduce the production of proinflammatory cytokines (TNF-α, IL-6, IL-1ß, MCP-1, and IP-10) at the mRNA levels. Moreover, KD-1 could significantly reduce the protein expression of p-NF-κB p65, NF-κB p65, and p-IκBα, while increasing the expression of IκBα in Huh-7 cells. CONCLUSIONS: KD-1 could significantly inhibit virus proliferation in vitro, the up-regulated expression of proinflammatory cytokines induced by SARS-CoV-2 and HCoV-229E by regulating the activity of the NF-кB signaling pathway. Our findings indicated that KD-1 protected against virus attack and can thus be used as a novel strategy for controlling the coronavirus disease 2019.

Coronaviruses and Endocrine System: A Systematic Review on Evidence and Shadows.

Coronaviruses are a big family of viruses that can infect mammalians and birds. In humans they mainly cause respiratory tract infections, with a large spectrum of severity, from mild, self-limited infections to highly lethal forms as severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV) and Coronavirus Disease 2019 (COVID-19). Scanty data are reported for the involvement of endocrine glands in human coronaviruses, in particular SARS-CoV-2. In this review, we summarize endocrinological involvement in human coronaviruses, including data on animal coronaviruses. Avians, ferrets and bovine are affected by specific coronavirus syndromes, with variable involvement of endocrine glands. SARS-CoV and SARS-CoV-2 use angiotensin-converting enzyme 2 (ACE2) as a target receptor, so ACE2 plays a central role in viral transmission and initial organ involvement. Autoptic studies on SARS patients revealed that thyroid, parathyroid, pituitary gland, endocrine pancreas and especially adrenals and testis could be impaired by different mechanisms (direct damage by SARS-CoV, inflammation, vascular derangement and autoimmune reactions) and few clinical studies have evidenced functional endocrine impairment. Only few data are available for COVID-19 and gonads and endocrine pancreas seem to be involved. International endocrinological societies have brought some recommendations for the COVID-19 pandemic, but further studies need to be performed, especially to detect long-term hormonal sequelae.

Serological profiles of pan-coronavirus-specific responses in COVID-19 patients using a multiplexed electro-chemiluminescence-based testing platform.

Serological assessment of SARS-CoV-2 specific responses are an essential tool for determining the prevalence of past SARS-CoV-2 infections in the population especially when testing occurs after symptoms have developed and limited contact tracing is in place. The goal of our study was to test a new 10-plex electro-chemiluminescence-based assay to measure IgM and IgG responses to the spike proteins from multiple human coronaviruses including SARS-CoV-2, assess the epitope specificity of the SARS-CoV-2 antibody response against full-length spike protein, receptor-binding domain and N-terminal domain of the spike protein, and the nucleocapsid protein. We carried out the assay on samples collected from three sample groups: subjects diagnosed with COVID-19 from the U.S. Army hospital at Camp Humphreys in Pyeongtaek, South Korea; healthcare administrators from the same hospital but with no reported diagnosis of COVID-19; and pre-pandemic samples. We found that the new CoV-specific multiplex assay was highly sensitive allowing plasma samples to be diluted 1:30,000 with a robust signal. The reactivity of IgG responses to SARS-CoV-2 nucleocapsid protein and IgM responses to SARS-CoV-2 spike protein could distinguish COVID-19 samples from non-COVID-19 and pre-pandemic samples. The data from the three sample groups also revealed a unique pattern of cross-reactivity between SARS-CoV-2 and SARS-CoV-1, MERS-CoV, and seasonal coronaviruses HKU1 and OC43. Our findings show that the CoV-2 IgM response is highly specific while the CoV-2 IgG response is more cross-reactive across a range of human CoVs and also showed that IgM and IgG responses show distinct patterns of epitope specificity. In summary, this multiplex assay was able to distinguish samples by COVID-19 status and characterize distinct trends in terms of cross-reactivity and fine-specificity in antibody responses, underscoring its potential value in diagnostic or serosurveillance efforts.

Molecular Evolution of Human Coronavirus 229E in Hong Kong and a Fatal COVID-19 Case Involving Coinfection with a Novel Human Coronavirus 229E Genogroup.

Compared to other human coronaviruses, the genetic diversity and evolution of human coronavirus 229E (HCoV-229E) are relatively understudied. We report a fatal case of COVID-19 pneumonia coinfected with HCoV-229E in Hong Kong. Genome sequencing of SARS-CoV-2 and HCoV-229E from a nasopharyngeal sample of the patient showed that the SARS-CoV-2 strain HK13 was most closely related to SARS-CoV-2 type strain Wuhan-Hu-1 (99.99% nucleotide identity), compatible with his recent history of travel to Wuhan. The HCoV-229E strain HK20-42 was most closely related to HCoV-229E strain SC0865 from the United States (99.86% nucleotide identity). To investigate if it may represent a newly emerged HCoV-229E genotype in Hong Kong, we retrieved 41 archived respiratory samples that tested positive for HCoV-229E from 2004 to 2019. Pneumonia and exacerbations of chronic airway diseases were common among infected patients. Complete RdRp, S, and N gene sequencing of the 41 HCoV-229E strains revealed that our contemporary HCoV-229E strains have undergone significant genetic drift with clustering of strains in chronological order. Two novel genogroups were identified, in addition to previously described genogroups 1 to 4, with recent circulating strains including strain HK20-42 belonging to novel genogroup 6. Positive selection was detected in the spike protein and receptor-binding domain, which may be important for viral evolution at the receptor-binding interphase. Molecular dating analysis showed that HCoV-229E shared the most recent common ancestor with bat and camel/alpaca 229E-related viruses at ∼1884, while camel/alpaca viruses had a relatively recent common ancestor at ∼1999. Further studies are required to ascertain the evolutionary origin and path of HCoV-229E.IMPORTANCE Since its first appearance in the 1960s, the genetic diversity and evolution of human coronavirus 229E (HCoV-229E) have been relatively understudied. In this study, we report a fatal case of COVID-19 coinfected with HCoV-229E in Hong Kong. Genome sequencing revealed that our SARS-CoV-2 strain is highly identical to the SARS-CoV-2 strain from Wuhan, compatible with the patient's recent travel history, whereas our HCoV-229E strain in this study is highly identical to a recent strain in the United States. We also retrieved 41 archived HCoV-229E strains from 2004 to 2019 in Hong Kong for sequence analysis. Pneumonia and exacerbations of chronic airway diseases were common diagnoses among the 41 patients. The results showed that HCoV-229E was evolving in chronological order. Two novel genogroups were identified in addition to the four preexisting HCoV-229E genogroups, with recent circulating strains belonging to novel genogroup 6. Molecular clock analysis dated bat-to-human and bat-to-camelid transmission to as early as 1884.

Epitope-resolved profiling of the SARS-CoV-2 antibody response identifies cross-reactivity with endemic human coronaviruses.

The SARS-CoV-2 proteome shares regions of conservation with endemic human coronaviruses (CoVs), but it remains unknown to what extent these may be cross-recognized by the antibody response. Here, we study cross-reactivity using a highly multiplexed peptide assay (PepSeq) to generate an epitope-resolved view of IgG reactivity across all human CoVs in both COVID-19 convalescent and negative donors. PepSeq resolves epitopes across the SARS-CoV-2 Spike and Nucleocapsid proteins that are commonly targeted in convalescent donors, including several sites also recognized in some uninfected controls. By comparing patterns of homologous reactivity between CoVs and using targeted antibody-depletion experiments, we demonstrate that SARS-CoV-2 elicits antibodies that cross-recognize pandemic and endemic CoV antigens at two Spike S2 subunit epitopes. We further show that these cross-reactive antibodies preferentially bind endemic homologs. Our findings highlight sites at which the SARS-CoV-2 response appears to be shaped by previous CoV exposures and which have the potential to raise broadly neutralizing responses.

High prevalence of pre-existing serological cross-reactivity against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in sub-Saharan Africa.

OBJECTIVE: Significant morbidity and mortality have occurred in the USA, Europe, and Asia due to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), whereas the numbers of infections and deaths in sub-Saharan Africa (SSA) have remained comparatively low. It has been hypothesized that exposure of the population in SSA to other coronaviruses prior to the COVID-19 pandemic resulted in some degree of cross-protection against SARS-CoV-2 infection and pathogenesis. We evaluated this hypothesis by comparing SARS-CoV-2 cross-reactive antibodies in pre-pandemic plasma samples collected from SSA and the USA. METHOD: Pre-COVID-19 pandemic plasma samples from SSA and the USA were collected and tested by immunofluorescence assay against the spike and nucleocapsid proteins of all known human coronaviruses (HCoVs). RESULTS: The prevalence of SARS-CoV-2 serological cross-reactivity was significantly higher in samples from SSA compared with the USA. Most of these cross-reactive samples cross-recognized the SARS-CoV-2 nucleocapsid protein and the spike proteins of other HCoVs. Nucleocapsid proteins from HCoV-NL63 and HCoV-229E were detected in most samples, thereby implicating prior exposure to these two HCoVs as the likely source of cross-reactive antibodies against SARS-CoV-2. CONCLUSION: The low incidences of SARS-CoV-2 infection and disease in SSA appear to be correlated with the pre-pandemic serological cross-recognition of HCoVs, which are substantially more prevalent in SSA than the USA.

Immunoinformatic Analysis of SARS-CoV-2 Nucleocapsid Protein and Identification of COVID-19 Vaccine Targets.

COVID-19 is a worldwide emergency; therefore, there is a critical need for foundational knowledge about B and T cell responses to SARS-CoV-2 essential for vaccine development. However, little information is available defining which determinants of SARS-CoV-2 other than the spike glycoprotein are recognized by the host immune system. In this study, we focus on the SARS-CoV-2 nucleocapsid protein as a suitable candidate target for vaccine formulations. Major B and T cell epitopes of the SARS-CoV-2 N protein are predicted and resulting sequences compared with the homolog immunological domains of other coronaviruses that infect human beings. The most dominant of B cell epitope is located between 176-206 amino acids in the SRGGSQASSRSSSRSRNSSRNSTPGSSRGTS sequence. Further, we identify sequences which are predicted to bind multiple common MHC I and MHC II alleles. Most notably there is a region of potential T cell cross-reactivity within the SARS-CoV-2 N protein position 102-110 amino acids that traverses multiple human alpha and betacoronaviruses. Vaccination strategies designed to target these conserved epitope regions could generate immune responses that are cross-reactive across human coronaviruses, with potential to protect or modulate disease. Finally, these predictions can facilitate effective vaccine design against this high priority virus.

Shufeng Jiedu, a promising herbal therapy for moderate COVID-19:Antiviral and anti-inflammatory properties, pathways of bioactive compounds, and a clinical real-world pragmatic study.

BACKGROUND: Shufeng Jiedu capsules (SFJDC), a patented herbal drug composed of eight medicinal plants, is used for the treatment of different viral respiratory tract infectious diseases. Based on its antiviral, anti-inflammatory and immunoregulatory activity in acute lung injury, SFJDC might be a promising candidate for the treatment of COVID-19. PURPOSE: To evaluate the antiviral and anti-inflammatory properties and to discover the mechanism of action of SFJDC as a potential drug for the treatment of COVID-19. Furthermore, the study should determine the clinical effectiveness of SFJDC for the treatment of COVID-19. DESIGN: We analyzed the antiviral and anti-inflammatory effects of SFJDC in a HCoV-229E mouse model on lung index, virus load in the lung, the release of cytokines, and on T- and B-lymphocytes. The mechanism of action was further investigated by network analysis. Additionally, we investigated data from a clinical pragmatic real-world study for patients with confirmed COVID-19, to evaluate the clinical effect of SFJDC and to determine the best time to start the treatment. RESULTS: SFJDC significantly reduced the virus load in the lung of HCoV-229E mice (from 1109.29 ± 696.75 to 0 ± 0 copies/ml), decreased inflammatory factors IL-6, IL-10, TNF-α, and IFN-γ in the lung, and increased the amount of CD4+ and CD8+ cells in the blood compared to the model group. Network analysis revealed that SFJDC reduces the activity of NFκB via several signaling pathways. Quercetin, wogonin, and polydatin bind directly to the main protease (Mpro) of SARS-CoV-2. Clinical data showed that SFJDC, added to standard antiviral therapy (AVD), significantly reduced the clinical recovery time of COVID-19 and fatigue (from 3.55 ± 4.09 to 1.19 ± 2.28 days) as well as cough (from 5.67 ± 5.64 to 3.47 ± 3.75) days compared to AVD alone. SFJDC therapy was significantly more effective when used within the first 8 days after the onset of symptoms. CONCLUSION: SFJDC might be a promising drug for the treatment of COVID-19, but large-scale randomized, double-blinded, placebo-controlled clinical trials are needed to complement the real-world evidence. It might be beneficial to start SFJDC treatment as early as possible in suspected cases of COVID-19.

The receptor binding domain of the viral spike protein is an immunodominant and highly specific target of antibodies in SARS-CoV-2 patients.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that first emerged in late 2019 is responsible for a pandemic of severe respiratory illness. People infected with this highly contagious virus can present with clinically inapparent, mild, or severe disease. Currently, the virus infection in individuals and at the population level is being monitored by PCR testing of symptomatic patients for the presence of viral RNA. There is an urgent need for SARS-CoV-2 serologic tests to identify all infected individuals, irrespective of clinical symptoms, to conduct surveillance and implement strategies to contain spread. As the receptor binding domain (RBD) of the spike protein is poorly conserved between SARS-CoVs and other pathogenic human coronaviruses, the RBD represents a promising antigen for detecting CoV-specific antibodies in people. Here we use a large panel of human sera (63 SARS-CoV-2 patients and 71 control subjects) and hyperimmune sera from animals exposed to zoonotic CoVs to evaluate RBD's performance as an antigen for reliable detection of SARS-CoV-2-specific antibodies. By day 9 after the onset of symptoms, the recombinant SARS-CoV-2 RBD antigen was highly sensitive (98%) and specific (100%) for antibodies induced by SARS-CoVs. We observed a strong correlation between levels of RBD binding antibodies and SARS-CoV-2 neutralizing antibodies in patients. Our results, which reveal the early kinetics of SARS-CoV-2 antibody responses, support using the RBD antigen in serological diagnostic assays and RBD-specific antibody levels as a correlate of SARS-CoV-2 neutralizing antibodies in people.

COVID-19: viral-host interactome analyzed by network based-approach model to study pathogenesis of SARS-CoV-2 infection.

BACKGROUND: Epidemiological, virological and pathogenetic characteristics of SARS-CoV-2 infection are under evaluation. A better understanding of the pathophysiology associated with COVID-19 is crucial to improve treatment modalities and to develop effective prevention strategies. Transcriptomic and proteomic data on the host response against SARS-CoV-2 still have anecdotic character; currently available data from other coronavirus infections are therefore a key source of information. METHODS: We investigated selected molecular aspects of three human coronavirus (HCoV) infections, namely SARS-CoV, MERS-CoV and HCoV-229E, through a network based-approach. A functional analysis of HCoV-host interactome was carried out in order to provide a theoretic host-pathogen interaction model for HCoV infections and in order to translate the results in prediction for SARS-CoV-2 pathogenesis. The 3D model of S-glycoprotein of SARS-CoV-2 was compared to the structure of the corresponding SARS-CoV, HCoV-229E and MERS-CoV S-glycoprotein. SARS-CoV, MERS-CoV, HCoV-229E and the host interactome were inferred through published protein-protein interactions (PPI) as well as gene co-expression, triggered by HCoV S-glycoprotein in host cells. RESULTS: Although the amino acid sequences of the S-glycoprotein were found to be different between the various HCoV, the structures showed high similarity, but the best 3D structural overlap shared by SARS-CoV and SARS-CoV-2, consistent with the shared ACE2 predicted receptor. The host interactome, linked to the S-glycoprotein of SARS-CoV and MERS-CoV, mainly highlighted innate immunity pathway components, such as Toll Like receptors, cytokines and chemokines. CONCLUSIONS: In this paper, we developed a network-based model with the aim to define molecular aspects of pathogenic phenotypes in HCoV infections. The resulting pattern may facilitate the process of structure-guided pharmaceutical and diagnostic research with the prospect to identify potential new biological targets.

Infectivity of human coronavirus in the brain.

A new strain of human coronaviruses (hCoVs), Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), has been identified to be responsible for the current outbreak of the coronavirus disease 2019 (COVID-19). Though major symptoms are primarily generated from the respiratory system, neurological symptoms are being reported in some of the confirmed cases, raising concerns of its potential for intracranial invasion and neurological manifestations, both in the acute phase and in the long-term. At present, it remains unclear the extent to which SARS-CoV-2 is present in the brain, and if so, its pathogenic role in the central nervous system (CNS). Evidence for neuroinvasion and neurovirulence of hCoVs has been recognised in animal and human studies. Given that SARS-CoV-2 belongs to the same family and shares characteristics in terms of receptor binding properties, it is worthwhile exploring its potential CNS manifestations. This review summarises previous findings from hCoVs in relation to the CNS, and compares these with the new strain, aiming to provide a better understanding of the effects of SARS-CoV-2 on the CNS.

Possible therapeutic role of a highly standardized mixture of active compounds derived from cultured Lentinula edodes mycelia (AHCC) in patients infected with 2019 novel coronavirus.

The outbreak of SARS-CoV-2 disease (COVID-19) is currently, March 2020, affecting more than 100,000 people worldwide and, according to the WHO (World Health Organization), a pandemic is shortly expected. The virus infects the lower respiratory tract and causes severe pneumonia and mortality in approximately 10% and 3-5%, respectively, of cases, mainly among the elderly and/or people affected by other diseases. AHCC is an α-glucan-based standardized mushroom extract that has been extensively investigated as an immunostimulant both in animals and/or in humans affected by West Nile virus, influenza virus, avian influenza virus, hepatitis C virus, papillomavirus, herpes virus, hepatitis B virus and HIV by promoting a regulated and protective immune response. Although the efficacy of AHCC has not yet been specifically evaluated with respect to SARS-CoV-2 disease, its action in promoting a protective response to a wide range of viral infections, and the current absence of effective vaccines, could support its use in the prevention of diseases provoked by human pathogenic coronavirus, including COVID-19.