Biology subjects

Watterson, D.

Publications and source records attributed to Watterson, D..

17 recordsLinked to original sources

bioRxiv2026

Live attenuated YFV-17D vaccine induces robust B and T cell immunity in vaccine-draining lymph nodes

The yellow fever virus-17D (YFV-17D) vaccine is widely regarded as one of the most effective vaccines in existence, with protection linked to robust elicitation of neutralising antibodies1. The features of the germinal centre (GC) reaction driving this durable and potent antibody responses remain unclear. Here we examined immune dynamics in blood and lymph nodes (LNs) after primary YFV-17D immunisation in humans. Using fine needle biopsy (FNB), we identified robust T and B lymphocyte responses within draining LNs, underpinning maturation of antibodies targeting the viral envelope (E). Early antibody responses were largely IgM, with gradual accumulation of E-specific memory B cells and anti-viral IgG in the subsequent months. Monoclonal antibodies derived from GC B cells had specificity for E, with diversification in the LN capable of driving increases in binding and neutralising potency. While circulating vaccine-specific CD4+ T cells peaked two weeks post-vaccination, clonally related GC and IL-10+ T follicular helper populations progressively accumulated in the LN. Comparison of FNB transcriptional datasets following YFV-17D, SARS-CoV-2 mRNA/LNP, and seasonal influenza vaccination identified a conserved GC program involving similar B and T lymphocyte subpopulations. Collectively, we find that while YFV-17D induces robust GC activity, this trait is shared with other vaccine platforms and the notably durable protection against infection may reflect an incapacity of the virus to escape host immunity.

immunology↗

bioRxiv2026

The gut-liver axis and the role of bacteria in severe dengue virus infection in mice

Patients with severe dengue virus (DENV) infection can experience hemorrhage, shock and organ damage that coincides with defervescence and declining viremia. In addition to vascular leak, published work has shown that gut barrier permeability and elevated serum lipopolysaccharide (LPS) levels correlate with DENV disease severity. We previously described profound gut pathology in a mouse model of infection. This led to the hypothesis that a loss of gut epithelial barrier integrity and the influx of microbial products contribute to exacerbated inflammation and severe dengue disease. Consistent with this, here we demonstrated that DENV2 infection of AG129 mice compromised the gut epithelial barrier, promoted bacterial translocation, and induced microbial changes indicative of dysbiosis, including a reduced abundance of putatively beneficial microbial taxa and disrupted microbial circadian oscillations. Depleting gut bacteria with antibiotics from 1-day post-infection sharply reduced gut pathology as well as viral RNA in the proximal colon and liver, and the number of infected Kupffer cells in the liver. There was no evidence for systemic suppression of viral replication. We propose that viral infection of intestinal macrophages leads to inflammatory damage and mucosal barrier breakdown, dysbiosis and gut leak, with gut bacteria-dependent products promoting the susceptibility of Kupffer cells to DENV infection. Given the importance of liver damage in severe and fatal dengue disease, therapies that protect the gut barrier may help reduce disease severity.

microbiology↗

bioRxiv2026

Light Chain Dominant Quaternary Epitope Recognition and IgG3 Switching Drive Broad Dengue and Zika Virus Neutralization

Defining how antibodies achieve broad neutralization across antigenically related flaviviruses is essential for rational therapeutics and vaccine design. Here, we describe 3A06, a human monoclonal antibody that neutralizes dengue virus (DENV) serotypes 1-3 and zika virus (ZIKV). Notably, subclass switching to IgG3 exhibits expanded neutralizing activity against all four DENV serotypes. High-resolution cryo-electron microscopy (cryo-EM) analysis of 3A06 Fab bound to chimeric ZIKV virions shows that 3A06 engages a quaternary epitope spanning three E protomers. Functional analyses identify a major functional contribution from the light-chain, where somatic hypermutations are critical for neutralization potency, and reversion of improbable mutations substantially reduces neutralization and increases antibody dependent enhancement (ADE) at sub-neutralizing concentrations. Cryo-EM analyses of intact IgG3 in complex with chimeric DENV2 and DENV4 show that the conformational clexibility and extended IgG3 Fc-hinge enable bivalent engagement across E-protein rafts in multiple concigurations, overcoming geometrical constraints that limit binding in the IgG1 format. Together, these results define a distinct mode of epitope recognition relative to previously characterized E-dimer epitope (EDE)-specific broadly neutralizing antibodies (bnAbs), and demonstrate how IgG3 subclass architecture can expand virus neutralization breadth. These insights inform the rational engineering of antibodies and vaccines targeting quaternary epitopes on flaviviruses to overcome viral diversity.

immunology↗

bioRxiv2025

Cloning, expression and characterisation of antigen-specific recombinant bat immunoglobulin from the black flying fox (Pteropus alecto)

Bats are natural reservoirs of viruses that cause severe disease in livestock and humans. Recent high-profile spillover events have directed significant attention towards the interplay between zoonotic viruses and antiviral immunity inherent to bats. Studies have highlighted that bats could harbour some deadly viruses without exhibiting outward symptoms. Various hypotheses have been proposed on how bats coexist with viruses; this includes dampened inflammation and altered innate immunity. However, there is limited literature on the humoral immune response in bats due to the scarcity of bat-specific reagents. To address this knowledge gap, we employed recombinant antibody design techniques to generate antigen-specific recombinant bat antibodies. This strategy involves combining the paratope of well-characterised antiviral antibodies with the IgG1 constant region of the black flying fox (Pteropus alecto). Characterisation of recombinant bat antibodies have revealed that they display canonical features of mammalian IgG. Additionally, recombinant bat antibodies display a binding and neutralising profile akin to human antibody counterparts. This approach provides much needed diagnostic tools and novel reagents to accelerate research into bat antibody immunity.

immunology↗

bioRxiv2025

A protective bispecific antibody targets both Nipah virus surface glycoproteins and limits viral escape

Nipah virus (NiV) and Hendra virus (HeV) are highly pathogenic henipaviruses without approved human vaccines or therapies. Here, we report on a highly potent bispecific therapeutic that combines an anti-fusion (F) nanobody with an anti-receptor binding protein (RBP) antibody to deliver a dual-targeting biologic that is resistant to viral escape. We show that the nanobody, DS90, engages a unique, conserved site within prefusion F of NiV and HeV, and provides neutralization and complete protection from NiV disease. Bispecific engineering of DS90 with the anti-RBP mAb m102.4 results in neutralization, elimination of viral escape and superior protection from NiV disease compared to leading monovalent approaches. These findings carry implications for the development of cross-neutralizing immunotherapies that limit the emergence of henipaviral escape mutants.

microbiology↗

bioRxiv2024

Vaccine-induced and hybrid immunity against SARS-CoV-2 variants of concern in two cohorts in Queensland, Australia (2021-2022).

The spike glycoprotein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the main target for vaccine development, as antibodies generated against the spike protein are the most immunodominant and neutralizing against the virus. However, variants of concern (VOC), often containing multiple mutations within neutralizing epitopes, confer immune evasion of the response generated by current SARS-CoV-2 vaccines. To assess the immunogenicity and virus-neutralisation ability of antibodies induced by individual or combination of Vaxzevria (AstraZeneca: AZ) and Comirnaty (Pfizer: PZ) vaccines and of a hybrid immunity induced in people who were also infected with SARS-CoV-2, longitudinal sera samples of participants recruited through the David Serisier Research biobank (Mater Research Hospital) or at the University of Queensland were collected. ELISA with a panel of purified Spike proteins from ancestral, alpha, delta and omicron BA.1 and BA.2 VOCs showed significantly (by [~]2 to 6-fold) reduced IgG antibody titres against Spike proteins from Omicron BA.1 and BA.2 compared to ancestral strain regardless of the number of vaccinations or presence of infection. Neutralisation assays showed reduced activity against delta and omicron BA.1, BA.5 and BA.5 VOCs. However, the differences were in general less pronounced than in the ELISA assay and some were not statistically significant, particularly after four (two AZ and two PZ) vaccinations. We also generated by circular polymerase extension reaction an attenuated SARS-CoV-2 strain with deletion of all accessory genes, ORF 3, 6, 7 and 8, based on the ancestral (QLD02) virus backbone (QLD02{Delta}3678) and validated it in virus-neutralization assays with our panel of sera samples. We showed the attenuation of the QLD02{Delta}3678 virus in Vero E6 and human Caco2 cells. We demonstrated that neutralization assays with the wild-type QLD02 virus and QLD02{Delta}3678 virus were concordant, providing a safe platform for neutralisation assays in BSL2/PC2 settings.

microbiology↗

bioRxiv2023

Alternative cell entry mechanisms for SARS-CoV-2 and multiple animal viruses

The cell entry mechanism of SARS-CoV-2, the causative agent of the COVID-19 pandemic, is not fully understood. Most animal viruses hijack cellular endocytic pathways as an entry route into the cell. Here, we show that in cells that do not express serine proteases such as TMPRSS2, genetic depletion of all dynamin isoforms blocked the uptake and strongly reduced infection with SARS-CoV-2 and its variant Delta. However, increasing the viral loads partially and dose-dependently restored infection via a thus far uncharacterized entry mechanism. Ultrastructural analysis by electron microscopy showed that this dynamin-independent endocytic processes appeared as 150-200 nm non-coated invaginations and was efficiently used by numerous mammalian viruses, including alphaviruses, influenza, vesicular stomatitis, bunya, adeno, vaccinia, and rhinovirus. Both the dynamin-dependent and dynamin-independent infection of SARS-CoV-2 required a functional actin cytoskeleton. In contrast, the alphavirus Semliki Forest virus, which is smaller in diameter, required actin only for the dynamin-independent entry. The presence of TMPRSS2 protease rescued SARS-CoV-2 infection in the absence of dynamins. Collectively, these results indicate that some viruses such as canine parvovirus and SARS-CoV-2 mainly rely on dynamin for endocytosis-dependent infection, while other viruses can efficiently bypass this requirement harnessing an alternative infection entry route dependent on actin.

microbiology↗

bioRxiv2023

The role of N-glycosylation in spike antigenicity for the SARS-CoV-2 Gamma variant

The emergence of SARS-CoV-2 variants alters the efficacy of existing immunity towards the viral spike protein, whether acquired from infection or vaccination. Mutations that impact N-glycosylation of spike may be particularly important in influencing antigenicity, but their consequences are difficult to predict. Here, we compare the glycosylation profiles and antigenicity of recombinant viral spike of ancestral Wu-1 and the Gamma strain, which has two additional N-glycosylation sites due to amino acid substitutions in the N-terminal domain (NTD). We found that a mutation at residue 20 from threonine to asparagine within the NTD caused the loss of NTD-specific antibody binding. Glycan site-occupancy analyses revealed that the mutation resulted in N-glycosylation switching to the new sequon at N20 from the native N17 site. Site-specific glycosylation profiles demonstrated distinct glycoform differences between Wu-1, Gamma, and selected NTD variant spike proteins, but these did not affect antibody binding. Finally, we evaluated the specificity of spike proteins against convalescent COVID-19 sera and found reduced cross-reactivity against some mutants, but not Gamma spike compared to Wuhan spike. Our results illustrate the impact of viral divergence on spike glycosylation and SARS-CoV-2 antibody binding profiles.

biochemistry↗

bioRxiv2023

Senolytic therapy alleviates physiological human brain aging and COVID-19 neuropathology.

Aging is the primary risk factor for most neurodegenerative diseases, and recently coronavirus disease 2019 (COVID-19) has been associated with severe neurological manifestations that can eventually impact neurodegenerative conditions in the long-term. The progressive accumulation of senescent cells in vivo strongly contributes to brain aging and neurodegenerative co-morbidities but the impact of virus-induced senescence in the aetiology of neuropathologies is unknown. Here, we show that senescent cells accumulate in physiologically aged brain organoids of human origin and that senolytic treatment reduces inflammation and cellular senescence; for which we found that combined treatment with the senolytic drugs dasatinib and quercetin rejuvenates transcriptomic human brain aging clocks. We further interrogated brain frontal cortex regions in postmortem patients who succumbed to severe COVID-19 and observed increased accumulation of senescent cells as compared to age-matched control brains from non-COVID-affected individuals. Moreover, we show that exposure of human brain organoids to SARS-CoV-2 evoked cellular senescence, and that spatial transcriptomic sequencing of virus-induced senescent cells identified a unique SARS-CoV-2 variant-specific inflammatory signature that is different from endogenous naturally-emerging senescent cells. Importantly, following SARS-CoV-2 infection of human brain organoids, treatment with senolytics blocked viral retention and prevented the emergence of senescent corticothalamic and GABAergic neurons. Furthermore, we demonstrate in human ACE2 overexpressing mice that senolytic treatment ameliorates COVID-19 brain pathology following infection with SARS-CoV-2. In vivo treatment with senolytics improved SARS-CoV-2 clinical phenotype and survival, alleviated brain senescence and reactive astrogliosis, promoted survival of dopaminergic neurons, and reduced viral and senescence-associated secretory phenotype gene expression in the brain. Collectively, our findings demonstrate SARS-CoV-2 can trigger cellular senescence in the brain, and that senolytic therapy mitigates senescence-driven brain aging and multiple neuropathological sequelae caused by neurotropic viruses, including SARS-CoV-2.

neuroscience↗

bioRxiv2022

An alpaca-derived nanobody neutralizes the SARS-CoV-2 omicron variant

The SARS-CoV2 Omicron variant sub-lineages spread rapidly through the world, mostly due to their immune-evasive properties. This has put a significant part of the population at risk for severe disease and underscores the need for anti-SARS-CoV-2 agents that are effective against emergent strains in vulnerable patients. Camelid nanobodies are attractive therapeutic candidates due to their high stability, ease of large-scale production and potential for delivery via inhalation. Here, we characterize the RBD-specific nanobody W25, which we previously isolated from an alpaca, and show superior neutralization activity towards Omicron lineage BA.1 in comparison to all other SARS-CoV2 variants. Structure analysis of W25 in complex with the SARS-CoV2 spike surface glycoprotein shows that W25 engages an RBD epitope not covered by any of the antibodies previously approved for emergency use. Furthermore, we show that W25 also binds the spike protein from the emerging, more infectious Omicron BA.2 lineage with picomolar affinity. In vivo evaluation of W25 prophylactic and therapeutic treatments across multiple SARS-CoV-2 variant infection models, together with W25 biodistribution analysis in mice, demonstrates favorable pre-clinical properties. Together, these data endorse prioritization of W25 for further clinical development.

microbiology↗

bioRxiv2022

SARS-CoV-2 drives NLRP3 inflammasome activation in human microglia through spike-ACE2 receptor interaction

Coronavirus disease-2019 (COVID-19) is primarily a respiratory disease, however, an increasing number of reports indicate that SARS-CoV-2 infection can also cause severe neurological manifestations, including precipitating cases of probable Parkinsons disease. As microglial NLRP3 inflammasome activation is a major driver of neurodegeneration, here we interrogated whether SARS-CoV-2 can promote microglial NLRP3 inflammasome activation utilising a model of human monocyte-derived microglia. We identified that SARS-CoV-2 isolates can bind and enter microglia, triggering inflammasome activation in the absence of viral replication. Mechanistically, microglial NLRP3 could be both primed and activated with SARS-CoV-2 spike glycoprotein in a NF-{kappa}B and ACE2-dependent manner. Notably, virus- and spike protein-mediated inflammasome activation in microglia was significantly enhanced in the presence of -synuclein fibrils, which was entirely ablated by NLRP3-inhibition. These results support a possible mechanism of microglia activation by SARS-CoV-2, which could explain the increased vulnerability to developing neurological symptoms akin to Parkinsons disease in certain COVID-19 infected individuals, and a potential therapeutic avenue for intervention. SIGNIFICANCE STATEMENTSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) principally affects the lungs, however there is evidence that the virus can also reach the brain and lead to chronic neurological symptoms. In this study, we examined the interaction SARS-CoV-2 with brain immune cells, by using an ex-vivo model of human monocyte-derived microglia. We identified robust activation of the innate immune sensor complex, NLRP3 inflammasome, in cells exposed to SARS-CoV-2. This was dependent on spike protein-ACE2 receptor interaction and was potentiated in the presence of -synuclein. We therefore identify a possible mechanism for SARS-CoV-2 and increased vulnerability to developing neurological dysfunction. These findings support a potential therapeutic avenue for treatment of SARS-CoV-2 driven neurological manifestations, through use of NLRP3 inflammasome or ACE2 inhibitors.

neuroscience↗

bioRxiv2022

SARS-CoV-2 triggers complement activation through interactions with heparan sulfate

The complement system has been heavily implicated in severe COVID-19 with clinical studies revealing widespread gene induction, deposition, and activation. However, the mechanism by which complement is activated in this disease remains incompletely understood. Herein we examined the relationship between SARS-CoV-2 and complement by inoculating the virus in lepirudin-anticoagulated human blood. This caused progressive C5a production after 30 minutes and 24 hours, which was blocked entirely by inhibitors for factor B, C3, C5, and heparan sulfate. However, this phenomenon could not be replicated in cell-free plasma, highlighting the requirement for cell surface deposition of complement and interactions with heparan sulfate. Additional functional analysis revealed that complement-dependent granulocyte and monocyte activation was delayed. Indeed, C5aR1 internalisation and CD11b upregulation on these cells only occurred after 24 hours. Thus, SARS-CoV-2 is a non-canonical complement activator that triggers the alternative pathway through interactions with heparan sulfate.

immunology↗

bioRxiv2021

Complete protection by a single dose skin patch delivered SARS-CoV-2 spike vaccine

SARS-CoV-2 has infected over 160 million people and resulted in more than 3.3 million deaths, and we still face many challenges in the rollout of vaccines. Here, we use the high-density microarray patch to deliver a SARS-CoV-2 spike subunit vaccine directly to the skin. We show the vaccine, dry-coated on the patch is thermostable, and delivery of spike via HD-MAP induced greater cellular and antibody immune responses, with serum able to potently neutralize clinically relevant isolates including those from the B.1.1.7 and B.1.351 lineages. Finally, a single dose of HD-MAP-delivered spike provided complete protection from a lethal virus challenge, demonstrating that HD-MAP delivery of a SARS-CoV-2 vaccine is superior to traditional needle-and-syringe vaccination and has the potential to greatly impact the ongoing COVID-19 pandemic.

microbiology↗

bioRxiv2021

Human genome integration of SARS-CoV-2 contradicted by long-read sequencing

A recent study proposed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) hijacks the LINE-1 (L1) retrotransposition machinery to integrate into the DNA of infected cells. If confirmed, this finding could have significant clinical implications. Here, we applied deep (>50x) long-read Oxford Nanopore Technologies (ONT) sequencing to HEK293T cells infected with SARS-CoV-2, and did not find the virus integrated into the genome. By examining ONT data from separate HEK293T cultivars, we completely resolved 78 L1 insertions arising in vitro in the absence of L1 overexpression systems. ONT sequencing applied to hepatitis B virus (HBV) positive liver cancer tissues located a single HBV insertion. These experiments demonstrate reliable resolution of retrotransposon and exogenous virus insertions via ONT sequencing. That we found no evidence of SARS-CoV-2 integration suggests such events are, at most, extremely rare in vivo, and therefore are unlikely to drive oncogenesis or explain post-recovery detection of the virus.

genomics↗

bioRxiv2021

Zika virus noncoding RNA cooperates with the viral protein NS5 to inhibit STAT1 phosphorylation and facilitate viral pathogenesis

Zika virus (ZIKV) is a re-emerging pathogenic flavivirus, which causes microcephaly in infants and poses a continuing threat to public health. ZIKV, like all other flaviviruses, produces highly abundant noncoding RNA known as subgenomic flaviviral RNA (sfRNA). Herein we utilized wild-type and mutant ZIKV defective in production of sfRNA to elucidate for the first time how production of sfRNA affects all aspects of ZIKV pathogenesis. We found that in mouse pregnancy model of infection sfRNA is required for trans-placental dissemination of ZIKV and subsequent infection of fetal brain. Using human brain organoids, we showed that sfRNA promotes apoptosis of neural progenitor cells leading to profound cytopathicity and disintegration of organoids. We also found by transcriptome profiling and gene network analysis that in infected human placental cells sfRNA inhibits multiple antiviral pathways and promotes apoptosis with STAT1 identified as a key shared factor linking these two interconnected sfRNA activities. We further showed for the first time that sfRNA inhibits phosphorylation and nuclear translocation of STAT1 by a novel mechanism which involves binding to and stabilizing viral protein NS5. This allows accumulation of NS5 at the levels required for efficient inhibition of STAT1 phosphorylation. Thus, we elucidated the molecular mechanism by which ZIKV sfRNA exerts its functions in vertebrate hosts and discovered a co-operation between viral noncoding RNA and a viral protein as a novel strategy employed by viruses to counteract antiviral responses.

microbiology↗

bioRxiv2021

Concerns regarding vaccination as a management strategy against koala retrovirus

Koalas are of great cultural, ecological and economic significance to Australia. However, they are in steep decline throughout much of their geographic range, with the recently endogenized koala retrovirus (KoRV) presumed to be a contributing factor. Olagoke, et al. 1 have proposed vaccination against KoRV as a suitable control mechanism to reduce the severity of KoRV infections and suggest that their recombinant vaccine is effective at invoking a neutralising immune response and significantly reducing viral loads. Here, we report the findings of our own attempts to detect antibodies specific to the KoRV envelope protein (Env) in infected animals. Antibodies against Env were undetectable in all animals, as would be expected in the case of an endogenous virus antigen which should proceed towards a pathway of self-tolerance. This finding calls into question the results of Olagoke, et al. 1, and prior work by this team 2,3. Following a critical review of this work 1-3, we have identified a number of assay inaccuracies and possible over interpretations of the data and conclude that further work to develop and test a vaccine against KoRV in koalas is not supported by evidence.

immunology↗

bioRxiv2020

Pixatimod (PG545), a clinical-stage heparan sulfate mimetic, is a potent inhibitor of the SARS-CoV-2 virus

Heparan sulfate (HS) is a cell surface polysaccharide recently identified as a co-receptor with the ACE2 protein for recognition of the S1 spike protein on SARS-CoV-2 virus, providing a tractable new target for therapeutic intervention. Clinically-used heparins demonstrate inhibitory activity, but world supplies are limited, necessitating alternative solutions. Synthetic HS mimetic pixatimod is a drug candidate for cancer with immunomodulatory and heparanase-inhibiting properties. Here we show that pixatimod binds to and destabilizes the SARS-CoV-2 spike protein receptor binding domain (S1-RBD), and directly inhibits its binding to human ACE2, consistent with molecular modelling identification of multiple molecular contacts and overlapping pixatimod and ACE2 binding sites. Assays with multiple clinical isolates of live SARS-CoV-2 virus show that pixatimod potently inhibits infection of monkey Vero E6 and human bronchial epithelial cells at concentrations within its safe therapeutic dose range. Furthermore, in a K18-hACE2 mouse model pixatimod demonstrates that pixatimod markedly attenuates SARS-CoV-2 viral titer and COVID-19-like symptoms. This demonstration of potent anti-SARS-CoV-2 activity establishes proof-of-concept for targeting the HS-Spike protein-ACE2 axis with synthetic HS mimetics. Together with other known activities of pixatimod our data provides a strong rationale for its clinical investigation as a potential multimodal therapeutic to address the COVID-19 pandemic.

microbiology↗