Bioaccumulation Pattern of the SARS-CoV-2 Spike Proteins in Pacific Oyster Tissues.

There is mounting evidence of the contamination of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the sewage, surface water, and even marine environment. Various studies have confirmed that bivalve mollusks can bioaccumulate SARS-CoV-2 RNA to detectable levels. However, these results do not provide sufficient evidence for the presence of infectious viral particles. To verify whether oysters can bind the viral capsid and bioaccumulate the viral particles, Pacific oysters were artificially contaminated with the recombinant SARS-CoV-2 spike protein S1 subunit (rS1). The bioaccumulation pattern of the rS1 in different tissues was investigated by immunohistological assays. The results revealed that the rS1 was bioaccumulated predominately in the digestive diverticula. The rS1 was also present in the epithelium of the nondigestive tract tissues, including the gills, mantle, and heart. In addition, three potential binding ligands, including angiotensin-converting enzyme 2 (ACE 2)-like substances, A-type histo-blood group antigen (HBGA)-like substances, and oyster heat shock protein 70 (oHSP 70), were confirmed to bind rS1 and were distributed in tissues with various patterns. The colocalization analysis of rS1 and those potential ligands indicated that the distributions of rS1 are highly consistent with those of ACE 2-like substances and oHSP 70. Both ligands are distributed predominantly in the secretory absorptive cells of the digestive diverticula and may serve as the primary ligands to bind rS1. Therefore, oysters are capable of bioaccumulating the SARS-CoV-2 capsid readily by filter-feeding behavior assisted by specific binding ligands, especially in digestive diverticula. IMPORTANCE This is the first article to investigate the SARS-CoV-2 spike protein bioaccumulation pattern and mechanism in Pacific oysters by the histochemical method. Oysters can bioaccumulate SARS-CoV-2 capsid readily by filter-feeding behavior assisted by specific binding ligands. The new possible foodborne transmission route may change the epidemic prevention strategies and reveal some outbreaks that current conventional epidemic transmission routes cannot explain. This original and interdisciplinary paper advances a mechanistic understanding of the bioaccumulation of SARS-CoV-2 in oysters inhabiting contaminated surface water.

Small molecule therapeutics for COVID-19: repurposing of inhaled furosemide.

The novel coronavirus SARS-CoV-2 has become a global health concern. The morbidity and mortality of the potentially lethal infection caused by this virus arise from the initial viral infection and the subsequent host inflammatory response. The latter may lead to excessive release of pro-inflammatory cytokines, IL-6 and IL-8, as well as TNF-α ultimately culminating in hypercytokinemia ("cytokine storm"). To address this immuno-inflammatory pathogenesis, multiple clinical trials have been proposed to evaluate anti-inflammatory biologic therapies targeting specific cytokines. However, despite the obvious clinical utility of such biologics, their specific applicability to COVID-19 has multiple drawbacks, including they target only one of the multiple cytokines involved in COVID-19's immunopathy. Therefore, we set out to identify a small molecule with broad-spectrum anti-inflammatory mechanism of action targeting multiple cytokines of innate immunity. In this study, a library of small molecules endogenous to the human body was assembled, subjected to in silico molecular docking simulations and a focused in vitro screen to identify anti-pro-inflammatory activity via interleukin inhibition. This has enabled us to identify the loop diuretic furosemide as a candidate molecule. To pre-clinically evaluate furosemide as a putative COVID-19 therapeutic, we studied its anti-inflammatory activity on RAW264.7, THP-1 and SIM-A9 cell lines stimulated by lipopolysaccharide (LPS). Upon treatment with furosemide, LPS-induced production of pro-inflammatory cytokines was reduced, indicating that furosemide suppresses the M1 polarization, including IL-6 and TNF-α release. In addition, we found that furosemide promotes the production of anti-inflammatory cytokine products (IL-1RA, arginase), indicating M2 polarization. Accordingly, we conclude that furosemide is a reasonably potent inhibitor of IL-6 and TNF-α that is also safe, inexpensive and well-studied. Our pre-clinical data suggest that it may be a candidate for repurposing as an inhaled therapy against COVID-19.