School of Biomedical Sciences - Research Publications
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ItemHost-dependent resistance of Group A Streptococcus to sulfamethoxazole mediated by a horizontally-acquired reduced folate transporter(NATURE PORTFOLIO, 2022-11-30)Described antimicrobial resistance mechanisms enable bacteria to avoid the direct effects of antibiotics and can be monitored by in vitro susceptibility testing and genetic methods. Here we describe a mechanism of sulfamethoxazole resistance that requires a host metabolite for activity. Using a combination of in vitro evolution and metabolic rescue experiments, we identify an energy-coupling factor (ECF) transporter S component gene (thfT) that enables Group A Streptococcus to acquire extracellular reduced folate compounds. ThfT likely expands the substrate specificity of an endogenous ECF transporter to acquire reduced folate compounds directly from the host, thereby bypassing the inhibition of folate biosynthesis by sulfamethoxazole. As such, ThfT is a functional equivalent of eukaryotic folate uptake pathways that confers very high levels of resistance to sulfamethoxazole, yet remains undetectable when Group A Streptococcus is grown in the absence of reduced folates. Our study highlights the need to understand how antibiotic susceptibility of pathogens might function during infections to identify additional mechanisms of resistance and reduce ineffective antibiotic use and treatment failures, which in turn further contribute to the spread of antimicrobial resistance genes amongst bacterial pathogens.
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ItemThe effect of substance P and its common in vivo-formed metabolites on MRGPRX2 and human mast cell activation(JOHN WILEY & SONS LTD, 2022-08)The tachykinin neuropeptide substance P (SP) is the canonical agonist peptide for the neurokinin 1 receptor (NK1 R). More recently, it has also been shown to activate the Mas-related G protein-coupled receptor X2 (MRGPRX2) receptor on mast cells (MCs), triggering degranulation and release of inflammatory mediators. SP undergoes rapid C-terminal truncation in vivo by a number of proteases to generate the metabolites SP(1-9)-COOH and in particular SP(1-7)-COOH. While the C terminus of SP is critical for NK1 R activation, studies have shown that the peptide polycationic N terminus is key for MRGPRX2 and mast cell activation. The study thus aimed to determine if the C-terminally truncated metabolites of SP, SP(1-9)-COOH, and SP(1-7)-COOH retained stimulatory activity at MRGPRX2. SP, SP(1-9)-COOH, and SP(1-7)-COOH were synthesized and tested on HEK293 cells expressing NK1 R or MRGPRX2, and LAD2 human mast cells, to determine the activity of SP and its metabolites in Ca2+ mobilization, degranulation, and cytokine assays. As expected from prior studies, both C-terminally truncated SP metabolites had essentially no activity at NK1 R, even at very high concentrations. In contrast, the in vivo metabolite of SP, SP(1-9)-COOH retained ability to activate MRGPRX2 across all parameters tested, albeit with reduced potency compared to intact SP. SP(1-7)-COOH did not produce any significant MRGRPX2 activation. Our results suggest that the SP metabolite, SP(1-9)-COOH, may play a regulatory role through the activation of MRGPRX2. However, given the relatively low potency of both SP and SP(1-9)-COOH at MRGPRX2, additional work is needed to better understand the biological importance of this expanded SP/MRGPRX2 pathway.
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ItemTetraspanin CD82 restrains phagocyte migration but supports macrophage activation(CELL PRESS, 2022-07-15)Phagocytes migrate into tissues to combat infection and maintain tissue homeostasis. As dysregulated phagocyte migration and function can lead to inflammation or susceptibility to infection, identifying molecules that control these processes is critical. Here, we show that the tetraspanin CD82 restrains the migration of neutrophils and macrophages into tissues. Cd82 -/- phagocytes exhibited excessive migration during in vivo models of peritoneal inflammation, superfusion of CXCL1, retinopathy of prematurity, and infection with the protozoan parasite L. mexicana. However, with the latter, while Cd82 -/- macrophages infiltrated infection sites at higher proportions, cutaneous L. mexicana lesions were larger and persisted, indicating a failure to control infection. Analyses of in vitro bone-marrow-derived macrophages showed CD82 deficiency altered cellular morphology, and impaired gene expression and metabolism in response to anti-inflammatory activation. Altogether, this work reveals an important role for CD82 in restraining phagocyte infiltration and mediating their differentiation in response to stimulatory cues.
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ItemThe eyes have it: dim-light activity is associated with the morphology of eyes but not antennae across insect orders(OXFORD UNIV PRESS, 2021-10)Abstract The perception of cues and signals in visual, olfactory and auditory modalities underpins all animal interactions and provides crucial fitness-related information. Sensory organ morphology is under strong selection to optimize detection of salient cues and signals in a given signalling environment, the most well-studied example being selection on eye design in different photic environments. Many dim-light active species have larger compound eyes relative to body size, but little is known about differences in non-visual sensory organ morphology between diurnal and dim-light active insects. Here, we compare the micromorphology of the compound eyes (visual receptors) and antennae (olfactory and mechanical receptors) in representative pairs of day active and dim-light active species spanning multiple taxonomic orders of insects. We find that dim-light activity is associated with larger compound eye ommatidia and larger overall eye surface area across taxonomic orders but find no evidence that morphological adaptations that enhance the sensitivity of the eye in dim-light active insects are accompanied by morphological traits of the antennae that may increase sensitivity to olfactory, chemical or physical stimuli. This suggests that the ecology and natural history of species is a stronger driver of sensory organ morphology than is selection for complementary investment between sensory modalities.
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ItemHypnotics with novel modes of action(WILEY, 2020-02)Insomnia and, more generally, lack of sleep are on the rise. Traditionally treated by classical hypnotics, such as benzodiazepines and Z drugs, which both act on the GABAA receptor, and other modalities, including nondrug therapies, such as cognitive behavioural therapy, there is a range of new hypnotics which are being developed or have recently received market approval. Suvorexant and the like target the orexin/hypocretin system: they should have less side effects in terms of drug-drug interactions with e.g. alcohol, less memory impairment and dependence potential compared to classical hypnotics.
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ItemLong-term captivity is associated with changes to sensory organ morphology in a critically endangered insect(WILEY, 2022-02)Abstract Captive breeding programmes are key to many threatened species reintroduction strategies but could potentially be associated with adaptations to captivity that are maladaptive in their natural habitat. Despite the importance of sensory ecology to biological fitness, few studies explore sensory system adaptations to captivity. Captive environments are devoid of predators and provide ready access to food sources and potential mates, thus reducing the need for individuals to use signals and cues to identify and locate resources or detect potential threats. With reduced complexity of the signalling environment, relaxation of selective pressures may favour reduced investment in sensory organs in captivity. We test this prediction in an iconic critically endangered invertebrate, the Lord Howe Island stick insect Dryococelus australis, which was extirpated from the island in the 1920s/30s and rediscovered on a nearby volcanic stack, Ball's Pyramid, in 2001. Using historical specimens from these populations and specimens from the 8–10th and 14th generations of a long‐term conservation captive breeding programme, we examine differences in behaviourally relevant morphological traits of the compound eyes (visual organs) and antennae (olfactory organs). We find that captivity is associated with smaller compound eye size, smaller eye ommatidia and reduced density of antennal odour receptors. These morphological changes are indicative of reduced sensitivity to visual and olfactory signals and cues, and therefore are likely to have fitness implications when reintroducing a captive population into the wild. Synthesis and applications. We observe differences in sensory organ morphology between wild and captive‐bred populations of the critically endangered Lord Howe Island stick insect. Our results emphasise the importance of incorporating evolutionary biology and sensory ecology into conservation programme design: to minimise the potential for captive breeding environments to compromise sensory systems that support appropriate behaviours upon reintroduction of populations into a natural habitat.
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ItemThe Salmonella Effector SseK3 Targets Small Rab GTPases(FRONTIERS MEDIA SA, 2020-08-19)During infection, Salmonella species inject multiple type III secretion system (T3SS) effector proteins into host cells that mediate invasion and subsequent intracellular replication. At early stages of infection, Salmonella exploits key regulators of host intracellular vesicle transport, including the small GTPases Rab5 and Rab7, to subvert host endocytic vesicle trafficking and establish the Salmonella-containing vacuole (SCV). At later stages of intracellular replication, interactions of the SCV with Rab GTPases are less well defined. Here we report that Rab1, Rab5, and Rab11 are modified at later stages of Salmonella infection by SseK3, an arginine N-acetylglucosamine (GlcNAc) transferase effector translocated via the Salmonella pathogenicity island 2 (SPI-2) type III secretion system. SseK3 modified arginines at positions 74, 82, and 111 within Rab1 and this modification occurred independently of Rab1 nucleotide binding. SseK3 exhibited Golgi localization that was independent of its glycosyltransferase activity but Arg-GlcNAc transferase activity was required for inhibition of alkaline phosphatase secretion in transfected cells. While SseK3 had a modest effect on SEAP secretion during infection of HeLa229 cells, inhibition of IL-1 and GM-CSF cytokine secretion was only observed upon over-expression of SseK3 during infection of RAW264.7 cells. Our results suggest that, in addition to targeting death receptor signaling, SseK3 may contribute to Salmonella infection by interfering with the activity of key Rab GTPases.
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ItemHigh antibody titres induced by protein subunit vaccines using Mycobacterium ulcerans antigens Hsp18 and MUL _3720 with a TLR-2 agonist fail to protect against Buruli ulcer in mice(PEERJ INC, 2020-08-07)BACKGROUND: Mycobacterium ulcerans is the causative agent of a debilitating skin and soft tissue infection known as Buruli ulcer (BU). There is no vaccine against BU. The purpose of this study was to investigate the vaccine potential of two previously described immunogenic M. ulcerans proteins, MUL_3720 and Hsp18, using a mouse tail infection model of BU. METHODS: Recombinant versions of the two proteins were each electrostatically coupled with a previously described lipopeptide adjuvant. Seven C57BL/6 and seven BALB/c mice were vaccinated and boosted with each of the formulations. Vaccinated mice were then challenged with M. ulcerans via subcutaneous tail inoculation. Vaccine performance was assessed by time-to-ulceration compared to unvaccinated mice. RESULTS: The MUL_3720 and Hsp18 vaccines induced high titres of antigen-specific antibodies that were predominately subtype IgG1. However, all mice developed ulcers by day-40 post-M. ulcerans challenge. No significant difference was observed in the time-to-onset of ulceration between the experimental vaccine groups and unvaccinated animals. CONCLUSIONS: These data align with previous vaccine experiments using Hsp18 and MUL_3720 that indicated these proteins may not be appropriate vaccine antigens. This work highlights the need to explore alternative vaccine targets and different approaches to understand the role antibodies might play in controlling BU.