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Lord, M.

Publications and source records attributed to Lord, M..

5 recordsLinked to original sources

A novel chitinase-like family of candidate effectors unique to aphids

Molecular interactions between aphids and plants include delivery of salivary effector proteins into host cells, acting as virulence factors to suppress host immunity, or as avirulence functions triggering immune activation. However, understanding of virulence and avirulence mechanisms in aphid-plant systems is currently limited. Here, we report discovery of an effector candidate family that is unique to aphids. Using functional genomics data on divergent pea aphid (Acyrthosiphon pisum) genotypes and their F1 progeny, we filtered for differentially expressed saliva proteins that co-segregated with virulence or avirulence phenotypes. LOC100575698 (ACPISUM_029930), annotated as an uncharacterized protein, was the sole candidate effector for which RNA-Seq and saliva proteomics data showed significantly different expression both between avirulent and virulent parents and between their segregating F1 progeny, with this gene upregulated in avirulent genotypes. BLASTP searches revealed multiple divergent homologs only in genomes of the Aphidomorpha infra-order, suggesting a hitherto undefined ancient aphid-specific gene family. AlphaFold models indicate strong structural similarities but weak sequence homology to chitinases. Because the aphid-specific clade all lack canonical DxxDxDxE motifs for catalytic activity, we designate the proteins as a novel CHitinase-Like (CHL) family. Association of ACPISUM_029930 (ApCHL1) with avirulence was further supported by co-segregating SNPs and a genotype-specific alternatively spliced isoform. We hypothesise that CHL proteins may function similarly to phylogenetically unrelated chitin-binding fungal effectors that sequester chitin, also present in aphid stylets, potentially preventing defence activation through plant chitin receptors and/or blocking chitin degradation by host-secreted chitinases.

molecular biology↗

The level of endothelial glycocalyx maturity modulates interactions with charged nano-materials

Nanomaterials have been extensively investigated for their potential in delivering therapeutics to target tissues, but few have advanced to clinical application. The luminal surface of endothelial cells that line blood vessels are covered by a glycocalyx, a complex extracellular matrix rich in anionic glycans. However, the role of this glycocalyx in governing nanomaterial-cell interactions is often overlooked. In this study, we demonstrate that gold nanoparticles functionalized with branched polyethyleneimine (AuNP+) bind to primary human endothelial cells expressing either a developing or mature glycocalyx, with the interaction involving hyaluronan and heparan sulfate. Notably, the mature glycocalyx decreases the toxicity of AuNP+. In contrast, lipoic acid-functionalized gold nanoparticles (AuNP-) bind to endothelial cells with a developing glycocalyx, but not a mature glycocalyx. To further investigate this phenomenon, we studied charged polymers, including poly(arginine) (polyR) and poly(glutamic acid) (polyE). PolyE does not associate with endothelial cells regardless of glycocalyx maturity, but when glycans are enzymatically degraded, it can bind to the cells. Conversely, polyR associates with endothelial cells irrespective of glycocalyx maturity or glycan degradation. These findings highlight the intricate relationship between nanomaterial charge and presentation in interactions with endothelial cells, offering insights for modulating nanomaterial interactions with the blood vessel wall.

bioengineering↗

Global siRNA Screen Reveals Critical Human Host Factors of SARS-CoV-2 Multicycle Replication

Defining the subset of cellular factors governing SARS-CoV-2 replication can provide critical insights into viral pathogenesis and identify targets for host-directed antiviral therapies. While a number of genetic screens have previously reported SARS-CoV-2 host dependency factors, these approaches relied on utilizing pooled genome-scale CRISPR libraries, which are biased towards the discovery of host proteins impacting early stages of viral replication. To identify host factors involved throughout the SARS-CoV-2 infectious cycle, we conducted an arrayed genome-scale siRNA screen. Resulting data were integrated with published datasets to reveal pathways supported by orthogonal datasets, including transcriptional regulation, epigenetic modifications, and MAPK signalling. The identified proviral host factors were mapped into the SARS-CoV-2 infectious cycle, including 27 proteins that were determined to impact assembly and release. Additionally, a subset of proteins were tested across other coronaviruses revealing 17 potential pan-coronavirus targets. Further studies illuminated a role for the heparan sulfate proteoglycan perlecan in SARS-CoV-2 viral entry, and found that inhibition of the non-canonical NF-kB pathway through targeting of BIRC2 restricts SARS-CoV-2 replication both in vitro and in vivo. These studies provide critical insight into the landscape of virus-host interactions driving SARS-CoV-2 replication as well as valuable targets for host-directed antivirals.

systems biology↗

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↗

Near-infrared Spectroscopy evaluations for rapid differentiation of carbapenem resistant Enterobacteriaceae from susceptible strains

Antimicrobial resistance (AMR) caused by Carbapenem-Resistant Enterobacteriaceae (CRE) is a global threat. Accurate identification of these bacterial species with associated AMR is critical for their management. While highly accurate methods to detect CRE are available, they are costly, timely and require expert skills making their application infeasible in low-resource settings. Here, we investigated the potential of Near-infrared Spectroscopy (NIRS) for a range of applications; i) the detection and differentiation of isolates of two pathogenic Enterobacteriaceae species, Klebsiella pneumoniae and Escherichia coli and, ii) the differentiation of carbapenem resistant and susceptible K. pneumoniae. NIRS has successfully differentiated between K. pneumoniae and E. coli isolates with a predictive accuracy of 89.04% (95% CI; 88.7-89.4%). K. pneumoniae isolates harbouring carbapenem resistance determinants were differentiated from susceptible K. pneumoniae strains with an accuracy of 85% (95% CI; 84.2-86.1%). To our knowledge, this is the largest demonstration of a proof of concept for the utility and feasibility of NIRS for rapidly differentiating between K. pneumoniae from E.coli as well as from carbapenem resistant K. pneumoniae from susceptible strains.

microbiology↗