Biology subjects

Rawlinson, W.

Publications and source records attributed to Rawlinson, W..

4 recordsLinked to original sources

bioRxiv2026

Proteolytic control of the SARS-CoV-2 furin cleavage site defines the phenotypic evolution from the pandemic to endemic state

The successive emergence of SARS-CoV-2 variants with altered tissue tropism has progressively decoupled transmissibility from lower respiratory tract pathogenicity. Omicron lineages transmit exceptionally well whilst limiting severe lung disease. Here, we demonstrate that this phenotype has evolved through two temporally distinct tissue-specific proteolytic controls at the Spike (S) furin cleavage site (FCS). At the virion level, cell-type-dependent furin-mediated FCS hyper-cleavage depletes S in lung but not nasal epithelial cells. At the infected cell membrane during cell-cell spread, TMPRSS2 mediates S FCS cleavage but is negatively regulated in the presence of ACE2. Tissue-specific solute carriers SLC6A19 and SLC6A20 sequester ACE2, thereby relieving inhibition of TMPRSS2 and enabling S FCS cleavage during cell-cell spread. Critically, whilst all SARS-CoV-2 lineages benefit from this latter pathway, Omicron variants have evolved exclusive dependence on it, a strategic consolidation that focuses S proteolytic activation to TMPRSS2 alone. The combined outcomes of S protein regulation across viral and cell membranes reveal how tissue-specific proteolytic optimisation drives Omicron's transmission fitness advantage in the upper respiratory tract but at the cost of heavy attenuation in the lower respiratory tract.

microbiology↗

bioRxiv2026

Antiviral efficacy versus host recovery: contrasting transcriptional footprints of four antivirals in human cytomegalovirus-infected brain organoids

Infection with human cytomegalovirus (HCMV) is common and usually asymptomatic in healthy individuals, but can cause severe neurological injury, particularly following congenital transmission. For symptomatic congenital infection, standard antiviral treatment is ganciclovir, with maribavir and letermovir as alternative direct-acting agents. However, their relative antiviral activity in reducing HCMV RNA abundance and restoring the host transcriptome towards an uninfected state has not been directly assessed in a neural model. We infected human cerebral organoids with Merlin-strain HCMV and treated them for 14 days with aciclovir, ganciclovir, letermovir, or maribavir, comparing each with untreated infected organoids (NO). Relative to NO, antiviral treatment reduced viral load 3.3-fold with aciclovir, 20.1-fold with ganciclovir, 65.4-fold with letermovir, and 6.9-fold with maribavir. We identified >2,500 differentially expressed host genes in infected versus uninfected organoids, with enrichment of neurodevelopmental and metabolic stress pathways. Aciclovir, ganciclovir, and maribavir produced few differentially expressed host genes relative to NO and no significant GO or KEGG enrichment. In contrast, letermovir altered 312 genes enriched for glycolysis and related metabolic processes. An MSigDB Hallmark pathway analysis showed minimal perturbation with aciclovir and letermovir, whereas ganciclovir and maribavir produced small, coordinated pathway-level shifts. This was partly in the same direction as control uninfected organoids but also with additional perturbations not seen in controls. Collectively, antiviral choice influenced both the magnitude of reduction in HCMV RNA abundance and the gene expression profile of infected neural tissue. The results support further evaluation of ganciclovir and letermovir in therapy of neural damage resulting from HCMV infection.

microbiology↗

bioRxiv2021

Longitudinal characterisation of phagocytic and neutralisation functions of anti-Spike antibodies in plasma of patients after SARS-CoV-2 infection.

Phagocytic responses by effector cells to antibody or complement-opsonised viruses have been recognized to play a key role in anti-viral immunity. These include antibody dependent cellular phagocytosis mediated via Fc-receptors, phagocytosis mediated by classically activated complement-fixing IgM or IgG1 antibodies and antibody independent phagocytosis mediated via direct opsonisation of viruses by complement products activated via the mannose-binding lectin pathway. Limited data suggest these phagocytic responses by effector cells may contribute to the immunological and inflammatory responses in SARS-CoV-2 infection, however, their development and clinical significance remain to be fully elucidated. In this cohort of 62 patients, acutely ill individuals were shown to mount phagocytic responses to autologous plasma-opsonised SARS-CoV-2 Spike protein-coated microbeads as early as 10 days post symptom onset. Heat inactivation of the plasma prior to use as an opsonin caused 77-95% abrogation of the phagocytic response, and pre-blocking of Fc-receptors on the effector cells showed only 18-60% inhibition. These results suggest that SARS-CoV-2 can provoke early phagocytosis, which is primarily driven by heat labile components, likely activated complements, with variable contribution from anti-Spike antibodies. During convalescence, phagocytic responses correlated significantly with anti-Spike IgG titers. Older patients and patients with severe disease had significantly higher phagocytosis and neutralisation functions when compared to younger patients or patients with asymptomatic, mild, or moderate disease. A longitudinal study of a subset of these patients over 12 months showed preservation of phagocytic and neutralisation functions in all patients, despite a drop in the endpoint antibody titers by more than 90%. Interestingly, surface plasmon resonance showed a significant increase in the affinity of the anti-Spike antibodies over time correlating with the maintenance of both the phagocytic and neutralisation functions suggesting that improvement in the antibody quality over the 12 months contributed to the retention of effector functions. Author SummaryLimited data suggest antibody dependent effector functions including phagocytosis may contribute to the immunological and inflammatory responses in SARS CoV-2 infection, however, their development, maintenance, and clinical significance remain unknown. In this study we show: O_LIPatients with acute SARS CoV-2 infection can mount phagocytic responses as early as 10 days post symptom onset and these responses were primarily driven by heat labile components of the autologous plasma. These results indicate that the current approach of studying phagocytosis using purified or monoclonal antibodies does not recapitulate contribution by all components in the plasma. C_LIO_LIIn convalescent patients, high phagocytic responses significantly correlated with increasing age, increasing disease severity, high neutralisation functions and high anti-Spike antibody titers, particularly IgG1. C_LIO_LILongitudinal study of convalescent patients over a 12-month period showed maintenance of phagocytic and neutralisation functions, despite a drop in the anti-Spike endpoint antibody titers by more than 90%. However, we found significant increase in the affinity of the anti-Spike antibodies over the 12-month period and these correlated with the maintenance of functions suggesting that improvement in the antibody quality over time contributed to the retention of effector functions. Clinically, measuring antibody titers in sera but not the quality of antibodies is considered a gold standard indicator of immune protection following SARS-CoV 2 infection or vaccination. Our results challenge this notion and recommends change in the current clinical practice. C_LI

immunology↗

bioRxiv2020

Chimeric synthetic reference standards enable cross-validation of positive and negative controls in SARS-CoV-2 molecular tests.

DNA synthesis in vitro has enabled the rapid production of reference standards. These are used as controls, and allow measurement and improvement of the accuracy and quality of diagnostic tests. Current reference standards typically represent target genetic material, and act only as positive controls to assess test sensitivity. However, negative controls are also required to evaluate test specificity. Using a pair of chimeric A/B RNA standards, this allowed incorporation of positive and negative controls into diagnostic testing for the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). The chimeric standards constituted target regions for RT-PCR primer/probe sets that are joined in tandem across two separate synthetic molecules. Accordingly, a target region that is present in standard A provides a positive control, whilst being absent in standard B, thereby providing a negative control. This design enables cross-validation of positive and negative controls between the paired standards in the same reaction, with identical conditions. This enables control and test failures to be distinguished, increasing confidence in the accuracy of results. The chimeric A/B standards were assessed using the US Centers for Disease Control real-time RT-PCR protocol, and showed results congruent with other commercial controls in detecting SARS CoV-2 in patient samples. This chimeric reference standard design approach offers extensive flexibility, allowing representation of diverse genetic features and distantly related sequences, even from different organisms.

molecular biology↗