Monoclonal Human Antibodies That Recognise the Exposed N and C Terminal Regions of the Often-Overlooked SARS-CoV-2 ORF3a Transmembrane Protein.

ORF3a has been identified as a viroporin of SARS-CoV-2 and is known to be involved in various pathophysiological activities including disturbance of cellular calcium homeostasis, inflammasome activation, apoptosis induction and disruption of autophagy. ORF3a-targeting antibodies may specifically and favorably modulate these viroporin-dependent pathological activities. However, suitable viroporin-targeting antibodies are difficult to generate because of the well-recognized technical challenge associated with isolating antibodies to complex transmembrane proteins. Here we exploited a naïve human single chain antibody phage display library, to isolate binders against carefully chosen ORF3a recombinant epitopes located towards the extracellular N terminal and cytosolic C terminal domains of the protein using peptide antigens. These binders were subjected to further characterization using enzyme-linked immunosorbent assays and surface plasmon resonance analysis to assess their binding affinities to the target epitopes. Binding to full-length ORF3a protein was evaluated by western blot and fluorescent microscopy using ORF3a transfected cells and SARS-CoV-2 infected cells. Co-localization analysis was also performed to evaluate the "pairing potential" of the selected binders as possible alternative diagnostic or prognostic biomarkers for COVID-19 infections. Both ORF3a N and C termini, epitope-specific monoclonal antibodies were identified in our study. Whilst the linear nature of peptides might not always represent their native conformations in the context of full protein, with carefully designed selection protocols, we have been successful in isolating anti-ORF3a binders capable of recognising regions of the transmembrane protein that are exposed either on the "inside" or "outside" of the infected cell. Their therapeutic potential will be discussed.

COVID-19-associated mucormycosis, diabetes and steroid therapy: Experience in a single centre in Western Mexico.

BACKGROUND: COVID-19-associated mucormycosis (CAM) has emerged as a challenging complication as the current pandemic has increased the population requiring treatment with corticosteroids. CAM has caused a massive outbreak in India, reported to be causing cases in Iran, Egypt and The Netherlands. OBJECTIVES: To describe CAM cases occurring in a single centre in Western Mexico. METHODS: Our group carried out a retrospective study from May 2020 to May 2021 to identify CAM cases in patients with previous COVID-19 diagnosis. RESULTS: Six CAM cases occurred in a single centre in Western Mexico during the study period, most of them with diabetes (n = 5/6) and all received corticosteroid therapy even when only three had severe COVID-19. After analysing local COVID-19 burden, it was estimated that in this region, CAM was 300 times more frequent among COVID individuals than the estimates for general population. CONCLUSION: Similar to large reports in India and other countries, CAM cases reported in this study were diagnosed in individuals with diabetes, hyperglycaemic status and with history of previous use of corticosteroids. Identifying these individuals at risk can help the early identification of CAM. In addition, strict glycaemic control and avoidance of unnecessary corticosteroid in non-severe COVID-19 cases could help in preventing this complicated fungal infection.

Neutralisation of SARS-CoV-2 by anatomical embalming solutions.

Teaching and learning anatomy by using human cadaveric specimens has been a foundation of medical and biomedical teaching for hundreds of years. Therefore, the majority of institutions that teach topographical anatomy rely on body donation programmes to provide specimens for both undergraduate and postgraduate teaching of gross anatomy. The COVID-19 pandemic has posed an unprecedented challenge to anatomy teaching because of the suspension of donor acceptance at most institutions. This was largely due to concerns about the potential transmissibility of the SARS-CoV-2 virus and the absence of data about the ability of embalming solutions to neutralise the virus. Twenty embalming solutions commonly used in institutions in the United Kingdom and Ireland were tested for their ability to neutralise SARS-CoV-2, using an established cytotoxicity assay. All embalming solutions tested neutralised SARS-CoV-2, with the majority of solutions being effective at high-working dilutions. These results suggest that successful embalming with the tested solutions can neutralise the SARS-CoV-2 virus, thereby facilitating the safe resumption of body donation programmes and cadaveric anatomy teaching.