School of Biomedical Sciences - Research Publications
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ItemEarly Depletion of Neutrophils Reduces Retinal Inflammation and Neovascularization in Mice with Oxygen-Induced Retinopathy(MDPI, 2023-11)Retinal inflammation is a central feature of ocular neovascular diseases such as diabetic retinopathy and retinopathy of prematurity, but the contribution of neutrophils to this process is not fully understood. We studied oxygen-induced retinopathy (OIR) which develops in two phases, featuring hyperoxia-induced retinal vaso-obliteration in phase I, followed by retinal neovascularization in phase II. As neutrophils are acute responders to tissue damage, we evaluated whether neutrophil depletion with an anti-Ly6G mAb administered in phase I OIR influenced retinal inflammation and vascular injury. Neutrophils were measured in blood and spleen via flow cytometry, and myeloperoxidase, an indicator of neutrophil activity, was evaluated in the retina using Western blotting. Retinal vasculopathy was assessed by quantitating vaso-obliteration, neovascularization, vascular leakage, and VEGF levels. The inflammatory factors, TNF, MCP-1, and ICAM-1 were measured in retina. In the OIR controls, neutrophils were increased in the blood and spleen in phase I but not phase II OIR. In OIR, the anti-Ly6G mAb reduced neutrophils in the blood and spleen, and myeloperoxidase, inflammation, and vasculopathy in the retina. Our findings revealed that the early rise in neutrophils in OIR primes the retina for an inflammatory and angiogenic response that promotes severe damage to the retinal vasculature.
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ItemNo Preview AvailableThe evolutionary mechanism of non-carbapenemase carbapenem-resistant phenotypes in Klebsiella spp(eLIFE SCIENCES PUBL LTD, 2023-07-06)Antibiotic resistance is driven by selection, but the degree to which a bacterial strain's evolutionary history shapes the mechanism and strength of resistance remains an open question. Here, we reconstruct the genetic and evolutionary mechanisms of carbapenem resistance in a clinical isolate of Klebsiella quasipneumoniae. A combination of short- and long-read sequencing, machine learning, and genetic and enzymatic analyses established that this carbapenem-resistant strain carries no carbapenemase-encoding genes. Genetic reconstruction of the resistance phenotype confirmed that two distinct genetic loci are necessary in order for the strain to acquire carbapenem resistance. Experimental evolution of the carbapenem-resistant strains in growth conditions without the antibiotic revealed that both loci confer a significant cost and are readily lost by de novo mutations resulting in the rapid evolution of a carbapenem-sensitive phenotype. To explain how carbapenem resistance evolves via multiple, low-fitness single-locus intermediates, we hypothesised that one of these loci had previously conferred adaptation to another antibiotic. Fitness assays in a range of drug concentrations show how selection in the antibiotic ceftazidime can select for one gene (blaDHA-1) potentiating the evolution of carbapenem resistance by a single mutation in a second gene (ompK36). These results show how a patient's treatment history might shape the evolution of antibiotic resistance and could explain the genetic basis of carbapenem-resistance found in many enteric-pathogens.
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ItemNo Preview AvailableDetection of Streptococcus pyogenes M1UK in Australia and characterization of the mutation driving enhanced expression of superantigen SpeA(NATURE PORTFOLIO, 2023-02-24)A new variant of Streptococcus pyogenes serotype M1 (designated 'M1UK') has been reported in the United Kingdom, linked with seasonal scarlet fever surges, marked increase in invasive infections, and exhibiting enhanced expression of the superantigen SpeA. The progenitor S. pyogenes 'M1global' and M1UK clones can be differentiated by 27 SNPs and 4 indels, yet the mechanism for speA upregulation is unknown. Here we investigate the previously unappreciated expansion of M1UK in Australia, now isolated from the majority of serious infections caused by serotype M1 S. pyogenes. M1UK sub-lineages circulating in Australia also contain a novel toxin repertoire associated with epidemic scarlet fever causing S. pyogenes in Asia. A single SNP in the 5' transcriptional leader sequence of the transfer-messenger RNA gene ssrA drives enhanced SpeA superantigen expression as a result of ssrA terminator read-through in the M1UK lineage. This represents a previously unappreciated mechanism of toxin expression and urges enhanced international surveillance.
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ItemNo Preview AvailableEvaluating the role of asymptomatic throat carriage of Streptococcus pyogenes in impetigo transmission in remote Aboriginal communities in Northern Territory, Australia: a retrospective genomic analysis(ELSEVIER, 2023-07)BACKGROUND: Streptococcus pyogenes, or group A Streptococcus (GAS), infections contribute to a high burden of disease in Aboriginal Australians, causing skin infections and immune sequelae such as rheumatic heart disease. Controlling skin infections in these populations has proven difficult, with transmission dynamics being poorly understood. We aimed to identify the relative contributions of impetigo and asymptomatic throat carriage to GAS transmission. METHODS: In this genomic analysis, we retrospectively applied whole genome sequencing to GAS isolates that were collected as part of an impetigo surveillance longitudinal household survey conducted in three remote Aboriginal communities in the Northern Territory of Australia between Aug 6, 2003, and June 22, 2005. We included GAS isolates from all throats and impetigo lesions of people living in two of the previously studied communities. We classified isolates into genomic lineages based on pairwise shared core genomes of more than 99% with five or fewer single nucleotide polymorphisms. We used a household network analysis of epidemiologically and genomically linked lineages to quantify the transmission of GAS within and between households. FINDINGS: We included 320 GAS isolates in our analysis: 203 (63%) from asymptomatic throat swabs and 117 (37%) from impetigo lesions. Among 64 genomic lineages (encompassing 39 emm types) we identified 264 transmission links (involving 93% of isolates), for which the probable source was asymptomatic throat carriage in 166 (63%) and impetigo lesions in 98 (37%). Links originating from impetigo cases were more frequent between households than within households. Households were infected with GAS for a mean of 57 days (SD 39 days), and once cleared, reinfected 62 days (SD 40 days) later. Increased household size and community presence of GAS and scabies were associated with slower clearance of GAS. INTERPRETATION: In communities with high prevalence of endemic GAS-associated skin infection, asymptomatic throat carriage is a GAS reservoir. Public health interventions such as vaccination or community infection control programmes aimed at interrupting transmission of GAS might need to include consideration of asymptomatic throat carriage. FUNDING: Australian National Health and Medical Research Council.
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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.