Browsing by Author "Shaw, Megan L."

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    The development of a sars-cov-2 pseudoparticle assay for the detection of neutralizing antibodies in sera from covid-19 patients
    (University of the Western Cape, 2024) Daniels, Ame-Leigh; Shaw, Megan L.
    The COVID-19 pandemic has caused devastating effects on the global socio-economic landscape. Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is the causative agent of COVID-19, and infections have led to approximately 7 million fatalities worldwide. Previous findings have confirmed that neutralizing antibodies are a highly predictive measure of immune protection. Thus, measuring SARS-CoV-2 neutralizing responses after infection or vaccination remains a priority, especially in the event of newly emerging SARS-CoV-2 variants. Due to the limitations of working with live virus in a BSL3 facility, pseudoparticles are an alternative tool used to study viral surface proteins. This research aims to develop a SARSCoV-2 pseudoparticle system to detect neutralizing antibodies in sera from previously infected or vaccinated individuals. The use of vesicular stomatitis virus (VSV) as a vector for the pseudoparticle system, provides several advantages over the lentivirus system including faster assay time, and lack of interference from antiretroviral drugs present in sera from HIVpositive patients on antiretroviral therapy (ART). The latter is particularly relevant in South Africa where there are more than 7.8 million HIV people living with HIV. After generating VSV pseudoparticles bearing the SARS-CoV-2 spike from the Wuhan/D614G, Beta, Delta and Omicron variants, a neutralizing assay was optimized using characterized human monoclonal antibodies. The assay was then applied to a sample set of patient sera and the ID50 values were compared to those obtained using a lentivirus-based SARS-CoV-2 neutralization assay. The comparison highlighted a strong concordance between the VSV and lentivirus neutralization assays particularly for the Wuhan/D614G and Omicron variants. The findings indicate that the development of a VSV-based neutralization assay is a valuable contribution to our ongoing efforts to characterize protective immune responses arising from SARS-CoV-2 infection or vaccination.
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    Functional landscape of SARS-CoV-2 cellular restriction
    (Cell Press, 2021) Martin-Sancho, Laura; Lewinski, Mary K.; Shaw, Megan L.
    A deficient interferon (IFN) response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has been implicated as a determinant of severe coronavirus disease 2019 (COVID-19). To identify the molecular effectors that govern IFN control of SARS-CoV-2 infection, we conducted a large-scale gain-of-function analysis that evaluated the impact of human IFN-stimulated genes (ISGs) on viral replication. A limited subset of ISGs were found to control viral infection, including endosomal factors inhibiting viral entry, RNA binding proteins suppressing viral RNA synthesis, and a highly enriched cluster of endoplasmic reticulum (ER)/Golgi-resident ISGs inhibiting viral assembly/egress.
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    A plant-produced SARS-CoV-2 spike protein elicits heterologous immunity in hamsters
    (Frontiers Media, 2023) Margolin, Emmanuel; Schäfer, Georgia; Shaw, Megan L.
    Molecular farming of vaccines has been heralded as a cheap, safe and scalable production platform. In reality, however, differences in the plant biosynthetic machinery, compared to mammalian cells, can complicate the production of viral glycoproteins. Remodelling the secretory pathway presents an opportunity to support key post-translational modifications, and to tailor aspects of glycosylation and glycosylation-directed folding. In this study, we applied an integrated host and glyco-engineering approach, NXS/T Generation™, to produce a SARS-CoV-2 prefusion spike trimer in Nicotiana benthamiana as a model antigen from an emerging virus. The size exclusion-purified protein exhibited a characteristic prefusion structure when viewed by transmission electron microscopy, and this was indistinguishable from the equivalent mammalian cell-produced antigen.
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    Protective efficacy of a plant-produced beta variant rsars-cov-2 vlp vaccine in golden syrian hamsters
    (Elsevier Ltd, 2024) Lemmer, Yolandy; Chapman, Ros; Shaw, Megan L.
    In the quest for heightened protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants, we engineered a prototype vaccine utilizing the plant expression system of Nicotiana benthamiana, to produce a recombinant SARS-CoV-2 virus-like particle (VLP) vaccine presenting the S-protein from the Beta (B.1.351) variant of concern (VOC). This innovative vaccine, formulated with either a squalene oil-in-water emulsion or a synthetic CpG oligodeoxynucleotide adjuvant, demonstrated efficacy in a golden Syrian Hamster challenge model. The Beta VLP vaccine induced a robust humoral immune response, with serum exhibiting neutralization not only against SARS-CoV-2 Beta but also cross-neutralizing Delta and Omicron pseudoviruses. Protective efficacy was demonstrated, evidenced by reduced viral RNA copies and mitigated weight loss and lung damage compared to controls. This compelling data instills confidence in the creation of a versatile platform for the local manufacturing of potential pan-sarbecovirus vaccines, against evolving viral threats.