Professor Andrew Currie

Introduction. Polymicrobial bacteraemia, the simultaneous presence of multiple bacterial species in the bloodstream, complicates diagnosis and treatment and is linked to poorer patient outcomes. Accurate detection is essential for effective clinical management. Gap Statement. Despite being the diagnostic gold standard, conventional blood culture may fail to detect all pathogens in polymicrobial infections, particularly when species differ in growth rate or abundance. The relative performance of culture versus molecular methods under these conditions remains poorly characterized. Aim. To compare the performance of conventional blood culture and quantitative PCR (qPCR) for detecting pathogens in simulated polymicrobial blood cultures containing fast- and slow-growing bacteria at varying ratios and concentrations.

Methodology. Four clinically relevant polymicrobial mixtures were prepared using seven bacterial species. Human blood was spiked with bacterial suspensions at varying ratios and inoculated into BACTEC blood culture bottles. After incubation, samples were analysed using standard culture techniques and an in-house multiplex qPCR assay.

Results. Routine culture detected both organisms in only 42% of samples, while qPCR identified both pathogens in 83%. Differences in bacterial growth rates significantly influenced culture outcomes, with slower-growing or less abundant species frequently missed. Notably, Staphylococcus aureus was not detected when co-cultured with Escherichia coli at ratios of 1:1 to 25:1, and only visible on Gram stain when S. aureus was initially 50 times more abundant.

Conclusion. These findings highlight a key limitation of conventional methods in detecting polymicrobial infections and underscore the need for more sensitive diagnostics. qPCR offers improved detection, particularly when organism abundance and growth rates vary. Incorporating molecular tools alongside routine culture may enhance diagnostic accuracy and provide a more complete understanding of polymicrobial bacteraemia.

Sexual assault against women is prevalent, prompting initiatives such as Australia's National Plan To End Violence Against Women. Forensic DNA analysis is key to achieving justice by linking offenders to crime scenes. Differential extraction remains the gold standard for separating victim and perpetrator DNA through selective lysis of cells. Despite widespread adoption of the original differential lysis protocol, results can have mixed success, prompting consideration of inter-operator variability. This study assessed the reproducibility of the 1980s method by having three forensic researchers independently conduct the published method using identical sperm samples (n = 3 replicates each). Their protocol interpretation was recorded, revealing numerous procedural differences. Operator 1 recovered 1.95 ng (range 1.92 - 1.99 ng) of DNA, operator 2 obtained no detectable DNA, and operator 3 resulted in 0.58 ng (range 0.56 - 0.60 ng) of DNA. These findings suggest that the original published protocol contains sections with insufficient detail to reproduce with precision. Forensic research manuscripts must ensure that methods are comprehensive to avoid similar circumstances.

Sunshinevirus was originally isolated in 2008 from a collection of Australian pythons suffering an outbreak of neuro-respiratory disease. Sunshinevirus infection cases, including chronic asymptomatic infections, have subsequently been detected throughout Australia by PCR testing. To investigate the association between sunshinevirus infection and disease, controlled experimental infection studies in carpet pythons (Morelia spilota) were undertaken. The study aims were to determine if there was a causal relationship between sunshinevirus and disease, as well as collecting information on potential virus transmission routes, infectious doses, target organs for viral replication and pathology, and incubation period. Between August 2016 and February 2018, six pythons were inoculated with sunshinevirus, two were sham inoculated as control animals, and two were co-housed with infected animals. All animals were sampled weekly and sunshinevirus was detected in virus-inoculated animals by PCR 6–14 days post-inoculation. Five of the six virus-inoculated carpet pythons developed neuro-respiratory disease, and sunshinevirus could be re-isolated from liver and kidney tissues as well as blood. One of the co-housed pythons became infected, consistent with natural transmission. The other co-housed python remained uninfected. Clinical signs commenced 3–7 months after viral exposure. They were slow to develop, often subtle, and predominantly neurological. Histologically, there was a non-suppurative encephalomyelitis, with the most consistent finding being vacuolation and gliosis in the brain. Cytoplasmic inclusions were found in a range of predominantly epithelial tissues. Infected pythons shed infectious virus 2.5–7 months before they showed obvious clinical signs of disease and remained viremic (for several months) until euthanasia.IMPORTANCEAustralian pythons are extensively kept in captivity. Since the 1990s, native Australian snakes with neuro-respiratory signs of disease, sometimes fatal, had been presenting to veterinarians in Australia. Research conducted at Murdoch University on some of these snakes resulted in the discovery of a novel virus, now named sunshinevirus (Sunviridae). Widespread follow-up PCR testing of pythons throughout Australia linked sunshinevirus infection to both asymptomatic virus-carriers and pythons showing signs of disease. Experimental infections were conducted to determine whether sunshinevirus infection caused disease in Australian pythons. The experiments demonstrated a causal relationship by fulfilling Koch’s postulates, and 5/6 infected animals developed neurological disease. Infected pythons shed virus persistently for months, but control animals remained uninfected. Our findings provide veterinarians with valuable information for the management of sunshinevirus in zoos and private collections.

The abundant skin commensal, Staphylococcus epidermidis, is the leading cause of late-onset sepsis (LOS) in preterm infants but rarely causes infections in term infants and adults. Staphylococcal virulence mechanisms and the role of the preterm immune responses in driving these life-threatening infections remain poorly understood. Using an ex vivo sepsis model, we challenged whole blood from very preterm infants (30-32 weeks gestational age, GA; n = 8), term infants (> 37 weeks GA; n = 8), and young adults (18-25 years; n = 8) with either live S. epidermidis or S. aureus (~ 10
colony-forming units, CFU/ml) for 90 min. Dual RNA-sequencing (RNA-seq) was performed to simultaneously assess host and pathogen gene expression profiles, identifying common and pathogen-specific responses across cohorts. We found shared immune processes induced in all age groups upon bacterial challenge, including cytokine (IL1A, IL1B, IL6, IFNB1) and chemokine (CCL20, CCL3, CCL7, CXCL2) signalling. Preterm infants also exhibited unique responses, such as increased platelet activation and fibrin clot formation, Wnt signalling, and hypoxia pathways in response to S. epidermidis challenge. Our findings suggest that bacterial gene co-expression, including iron acquisition and heme biosynthesis genes, are also influenced by the hosts developmental age, highlighting the complexity of host-bacterial interactions in the early stages of neonatal sepsis.

Introduction: Eczema is a common, chronic inflammatory skin condition characterised by recurrent flares that may be exacerbated by increased Staphylococcus aureus colonisation in the setting of impaired skin barrier. In preterm infants, topical coconut oil improves skin integrity and demonstrates antimicrobial and anti-inflammatory properties, which are appealing characteristics of a treatment for eczema. However, there are concerns that transcutaneous sensitisation may lead to the development of food allergy in children with eczema.

Method: CocoKids was a single centre (Perth Children's Hospital), open label, pilot randomised controlled trial to investigate the safety, feasibility, clinical effects and skin microbial changes associated with topical coconut oil in children with eczema. Children <6 years old were randomised to standard eczema care with (intervention) or without (control) addition of daily topical coconut oil for 12 weeks. Children with coconut allergy or positive skin prick test (SPT) to coconut extract or coconut oil at screening were excluded. Outcomes were adverse events (including any new sensitisation to coconut on SPT or coconut specific IgE), adherence, changes in eczema severity and quality of life, and skin microbial changes associated with use of coconut oil.

Results: 58 participants were randomised (n = 27 intervention, n = 31 control; median age 1.55 years, range 0.2–5.9). Application of coconut oil was feasible, with 25/27 (93%) reporting daily application. No children in the coconut oil group developed a new positive coconut SPT or specific IgE following completion of the intervention period. Parent-reported eczema severity and quality of life improved significantly in the coconut oil group compared to controls, while eczema lesions showed reduced S. aureus load in those treated with coconut oil.

Conclusion: Daily application of coconut oil for 12 weeks was safe, with no evidence for de novo sensitisation to coconut in this cohort. The intervention was highly acceptable to parents and was associated with improvements in some clinical and microbiological outcomes. Results from this pilot study support the feasibility and rationale for larger trials of topical coconut oil in eczema.

The neonatal skin is central to early survival and immune development. Far from being a passive mechanical barrier, it integrates physical, chemical, and microbial defences that together protect the infant in the immediate postnatal period. In preterm infants, structural immaturity, reduced antimicrobial capacity, and altered microbial colonisation confer heightened vulnerability to infection and inflammation. At the same time, the neonatal period represents a critical window during which skin–microbe interactions shape tolerance and long-term immune trajectories. This review summarises recent advances in understanding the development of the skin barrier, antimicrobial and innate immune defences, and the role of commensals in immune programming. Translational opportunities for neonatal care are discussed, including skin protective practices, antisepsis, and emollient use that may reduce infection risk in the neonatal intensive care unit. Finally, we consider future directions in microbiome-informed and skin-centred strategies.

Dual RNA-sequencing (dual RNA-seq) holds significant promise for deciphering bacterial virulence mechanisms during systemic infections. However, its application in sepsis research is hindered by technical challenges, including a low bacterial burden in blood and limited sample volumes and RNA yield from vulnerable populations, such as neonates. We developed an optimized protocol [dual RNA isolation from blood (DRIB)] for simultaneous stabilization, isolation and purification of high-quality host leukocyte and bacterial RNA from low-volume whole blood samples (0.5 ml). This protocol is compatible with clinical sample collection workflows and high-throughput RNA sequencing. The feasibility of DRIB for dual RNA-seq was validated using a pilot cohort of clinical adult sepsis samples, enabling the investigation of host–bacterial gene expression during sepsis. The DRIB protocol yielded 2.10–6.91 µg of total RNA per clinical sample in our pilot cohort. Dual-species ribosomal RNA (rRNA) depletion and RNA-seq generated 16.6–24.8 million filtered reads per sample, with 63±7% of reads uniquely mapped to host or bacterial sequences. Host genes accounted for 51–68% (8.4–10.9 million) reads, while 0.5–6.7% (79,496–789,808 reads) mapped to bacterial genomes. Bioinformatic analysis revealed that both shared and individual transcriptional patterns were identified in host and bacterial responses, including pathways related to immune metabolism and metal-ion binding. Our optimized DRIB protocol and RNA-seq pipeline effectively captured both host and bacterial RNA transcription in clinical sepsis samples. Expanding this approach to larger cohorts and varying disease timepoints will provide crucial new insights into host–bacterial gene co-expression dynamics in sepsis progression and outcomes.

Understanding the distribution and variation in NMR-based inflammatory markers is crucial to the evaluation of their clinical utility in disease prognosis and diagnosis. We applied high-resolution 1H NMR spectroscopy of blood plasma and serum to measure the acute phase reactive glycoprotein signals (GlycA and GlycB) and the subregions of the lipoprotein-based Supramolecular Phospholipid Composite signals (SPC1, SPC2, and SPC3) in a large multicohort population study. A total of 5702 samples were studied to determine the signal variations in a range of chronic and acute inflammatory conditions. We found that while GlycA and GlycB were increased in inflammation, the SPC regions behaved independently of Glyc signals, with SPC2 and SPC3 being reduced in chronic inflammation in comparison to healthy controls (p-value SPC2 = 2.9 × 10–10, p-value SPC3 = 2.2 × 10–3) and SPC1 (p-value = 0.29) being unchanged. SPC1 was decreased in acute inflammation, indicating a link to the immune response (p-value = 2.5 × 10–11). These findings confirm the independent biological relevance of all three SPC subregions and contraindicate the use of aggregate SPC values as general inflammatory markers.

Introduction: Skin care for very and extremely preterm infant is an important and previously underappreciated topic. Coconut oil skin care for preterm infants is a promising option, but several important questions remain including the theoretical potential for allergic sensitization.

Methods: This prospective study conducted skin prick testing and allergy questionnaires in a cohort of very preterm infants who received routine skin care with virgin coconut oil during their neonatal admission.

Results: Ninety infants (median GA 28.2 weeks, median BW 1,048 g) were assessed at corrected age 15 months. A total of 8 children had positive skin prick tests to 1 or more of the most common allergens (egg, peanut, cashew, dust mite). No child was sensitized to coconut oil or extract.

Conclusions: We did not find evidence of allergic sensitization following neonatal skin care with coconut oil in children born preterm.

Tick bites and tick-related diseases are on the rise. Diagnostic tests that identify well-characterised tick-borne pathogens (TBPs) possess limited capacity to address the causation of symptoms associated with poorly characterised tick-related illnesses, such as debilitating symptom complexes attributed to ticks (DSCATT) in Australia. Identification of local signals in tick-bitten skin that can be detected systemically in blood would have both clinical (diagnostic or prognostic) and research (mechanistic insight) utility, as a blood sample is more readily obtainable than tissue biopsies. We hypothesised that blood samples may reveal signals which reflect relevant local (tissue) events and that the time course of these signals may align with local pathophysiology. As a first step towards testing this hypothesis, we compared molecular signatures in skin biopsies taken from the tick-bite location of human participants, as published in our previous study, together with peripheral blood signatures obtained concurrently. This approach captures differentially expressed molecules across multiple omics datasets derived from peripheral blood (including cellular and cell-free transcriptomics, proteomics, metabolomics, and DNA methylation), and skin biopsies (spatial transcriptomics). Our original data revealed that extracellular matrix organisation and platelet degranulation pathways were upregulated in the skin within 72 h of a tick bite. The same signals appeared in blood, where they then remained elevated for three months, displaying longitudinally consistent alterations of biological functions. Despite the limited sample size, these data represent proof-of-concept that molecular events in the skin following a tick bite can be detectable systemically. This underscores the potential value of blood samples, akin to a liquid biopsy, to capture biomarkers reflecting local tissue processes.