Biology subjects

Gourlay, C.

Publications and source records attributed to Gourlay, C..

6 recordsLinked to original sources

bioRxiv2025

Monitoring Candida albicans Biofilm Formation by Impedance Using Passive RFID

Medical implants are routinely colonized by microbial biofilms especially yeast biofilms such as those formed by Candida albicans. As medical implants move towards smart technology and IoT, it is more important than ever to understand the electrical properties of biofilm formation. Here we present an attempt at understanding how C. albicans biofilm formation effects sensor technology. We propose that as the biofilm matures, it becomes increasingly hydrophobic and undergoes a reduction in effective dielectric constant and ionic conductivity. This results in an impedance mismatch that disrupts capacitive coupling between the sensor and surrounding media, this effect that can be reliably detected. We show that a C. albicans biofilm changes its dielectric properties during maturation and, once mature, can be equated to a thin dielectric insulator. We demonstrate that these properties can be used to enable constant monitoring of a C. albicans biofilm using passive RFID, offering a new detection methodology that may be applied to surfaces known to be readily colonise.

microbiology↗

bioRxiv2025

Engineered yeast biofilms deliver plastic degrading enzymes to PET substrates

Enzymes that degrade PET and other plastics are being discovered with increasing pace, and engineering approaches further increase their activity. Most proposed applications for such enzymes require their purification, enabling the regeneration of monomeric subunits suitable for the production of new plastic. However, large amounts of plastics exist as environmental contaminants, and are unlikely to be suitable for standard recycling. Such waste may still be suitable for bioremediation using enzymatic activities, albeit without the economic incentives of a full recycling process. Here we develop a delivery system for PET hydrolases, based on engineered yeast biofilms that grow and secrete enzymes directly on the substrate to be degraded. The system does not require any enzyme purification steps, and the biofilms at least partially assimilate the degradation products into biomass. Our work provides proof-of-principle that demonstrates the potential of biofilm-based approaches.

bioengineering↗

bioRxiv2025

A novel C. elegans respirometry assay using low-cost optical oxygen sensors

Measurement of the oxygen consumption rate (OCR), or respirometry, is a powerful and comprehensive method for assessing mitochondrial function both in vitro and in vivo. Respirometry at the whole-organism level has been repeatedly performed in the model organism Caenorhabditis elegans, typically using high throughput microplate-based systems over traditional Clark-type respirometers. However, these systems are highly specialised, costly to purchase and operate, and inaccessible to many researchers. Here, we develop a respirometry assay using low-cost commercially available optical oxygen sensors (PreSens OxoPlates(R)) and fluorescence plate readers (the BMG FLUOstar), as an alternative to more costly standard respirometry systems. This assay uses standard BMG FLUOstar protocols and a set of custom scripts to perform repeated measurements of the C. elegans OCR, with the optional use of respiratory inhibitors or other interventions. We validate this assay by demonstrating the linearity of basal OCRs in samples with highly variable numbers of animals, and by examining the impact of respiratory inhibitors with previously demonstrated efficacy in C. elegans: carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone (a mitochondrial uncoupler) and sodium azide (a Complex IV inhibitor). Using this assay, we demonstrate that the sequential use of FCCP and sodium azide leads to an increase in the sodium azide-treated (non-mitochondrial) OCR, indicating that the sequential use of respiratory inhibitors, as standard in intact cell respirometry, may produce erroneous estimates of non-mitochondrial respiration in C. elegans and thus should be avoided.

cell biology↗

bioRxiv2024

Phenotypic plasticity in a novel set of EGFR tyrosine kinase inhibitor-adapted non-small cell lung cancer cell lines

Here, we introduce sublines of the EGFR-mutant non-small cell lung cancer (NSCLC) cell lines HCC827 and HCC4006 adapted to the EGFR kinase inhibitors gefitinib (HCC827rGEFI2{micro}M, HCC4006rGEFI1{micro}M), erlotinib (HCC827rERLO2{micro}M, HCC4006rERLO1{micro}M), and afatinib (HCC827rAFA50nM, HCC4006rAFA100nM). All sublines displayed resistance to gefitinib, erlotinib, afatinib, and the third-generation EGFR kinase inhibitor osimertinib that overcomes T790M-mediated resistance. HCC4006rERLO1{micro}M displayed a spindle-like morphology in agreement with previous findings that had detected epithelial-mesenchymal-transition (EMT) in its precursor cell line HCC4006rERLO0.5{micro}M. EMT had also been reported for the HCC4006rGEFI1{micro}M precursor cell line HCC4006rGEFI0.5{micro}M and for HCC4006rAFA100nM, but the morphologies of HCC4006rGEFI1{micro}M or HCC4006rAFA100nM did not support this, suggesting plasticity in EMT regulation during the drug adaptation process and in established resistant cell lines. Accordingly, HCC4006rERLO1{micro}M displayed resistance to MEK and AKT inhibitors in contrast to its precursor HCC4006rERLO0.5{micro}M. We also detected metabolic plasticity, i.e., a temporary Warburg metabolism, in HCC4006 and HCC827rGEFI2{micro}M. Response profiles to cytotoxic anti-cancer drugs, kinase inhibitors, and HDAC inhibitors resulted in complex patterns that were specific for each individual subline without obvious overlaps, indicating individual resistance phenotypes. All resistant sublines remained sensitive or displayed collateral sensitivity to at least one of the investigated drugs. In conclusion, the comparison of EGFR kinase-resistant NSCLC sublines with their precursor cell lines that had been previously characterised at a lower resistance level and metabolic investigations indicated phenotypic plasticity during the resistance formation process and in established cell lines. This plasticity may contribute to the well-known variability in cell line phenotypes observed between different laboratories and in intra-laboratory experiments.

cancer biology↗

bioRxiv2020

Localization and Functional Characterization of the Alternative Oxidase in Naegleria

The Alternative oxidase (AOX) is a protein involved in maintaining the Krebs cycle in instances where the respiratory chain has been inhibited, while allowing for the maintenance of cell growth and necessary metabolic processes for survival. Among eukaryotes, alternative oxidases have disperse distribution and are found in plants, fungi and a few protists, including Naegleria ssp. Naegleria species are free-living unicellular amoeboflagellates, and include the pathogenic species of N. fowleri, the so-called brain eating amoeba. Using a multidisciplinary approach, we aimed to understand the evolution, localization and function of AOX and the role that plays in Naeglerias biology. Our analyses suggest that the protein was present in last common ancestor of the genus and structure prediction showed that all functional residues are also present in Naegleria species. Using a combination of cellular and biochemical techniques, we also functionally characterize N. gruberis AOX in its mitochondria and we demonstrate that its inactivation affects its proliferation. Consequently, we discuss the benefits of the presence of this protein in Naegleria species, along with its potential pathogenicity role in N. fowleri. We predict that our findings will spearhead new explorations to understand the cell biology, metabolism and evolution of Naegleria and other free-living relatives.

microbiology↗

bioRxiv2018

Inhibition of classical and alternative modes of respiration in C. albicans leads to cell wall remodelling and increased macrophage recognition

The human fungal pathogen C. albicans requires respiratory function for normal growth, morphogenesis and virulence. As such the mitochondria represent an enticing target for the development of new antifungal strategies. This possibility is further bolstered by the presence of fungal specific characteristics. However, respiration in C. albicans, as is the case in many fungal organisms, is facilitated by redundant electron transport mechanisms that makes direct inhibition a challenge. In addition, many chemicals known to target the electron transport chain are highly toxic. Here we make use of chemicals with low toxicity in mammals to efficiently inhibit respiration in C. albicans. We find that use of the Nitric Oxide donor, Sodium Nitroprusside (SNP), and the alternative oxidase inhibitor, SHAM, prevent respiration, lead to a loss in viability and to cell wall rearrangements that increase the rate of uptake by macrophages in vitro and in vivo. We propose that SNP+SHAM treatment leads to transcriptional changes that drive cell wall re-arrangement but which also prime cells to activate transition to hyphal growth. In line with this we find that pre-treatment of C. albicans with SNP+SHAM leads to an increase in virulence. Our data reveals strong links between respiration, cell wall remodelling and activation of virulence factors. Our findings also demonstrate that respiration in C. albicans can be efficiently inhibited with chemicals which are not damaging to the mammalian host, but that we need to develop a deeper understanding of the roles of mitochondria in cellular signalling if they are to be developed successfully as a target for new antifungals.\n\nAuthor SummaryCurrent approaches to tackling fungal infections are limited and new targets must be identified to protect against the emergence of resistant strains. We investigate the potential of targeting mitochondria, organelles required for energy production, growth and virulence, in the yeast human fungal pathogen Candida albicans. Our findings suggest that mitochondria can be targeted using drugs that can be tolerated by humans and that this treatment enhances their recognition by immune cells. However release of C. albicans cells from mitochondrial inhibition appears to activate a stress response that increases traits associated with virulence. Our results make it clear that mitochondria are a valid target for the development of anti-fungal strategies but that we must determine the mechanisms by which they regulate stress signalling and virulence ahead of successful therapeutic advance.

microbiology↗