Multidrug-resistant (MDR) Klebsiella pneumoniae poses a significant global health concern, particularly in hospital setting where it causes severe and hard-to-treat infections. In this study, 329 fungal-derived compounds were screened for their potential to inhibit MrkD1P, a key fimbrial adhesin protein (PDB ID: 3U4K) involved in host tissue adhesion. Molecular docking analysis identified ochratoxin A (− 9.1 kcal/mol), bromadiolone (− 8.6 kcal/mol), and permethrin (− 8.2 kcal/mol) as top-performing candidates, exhibiting strong binding affinities and stable molecular interactions, including hydrogen bonding and hydrophobic contacts. These findings were reinforced by 100-nanosecond molecular dynamics (MD) simulations, which showed sustained ligand–protein stability, particularly for ochratoxin A. Free energy estimations using the MM/PBSA method further suggested the thermodynamic favourability of these interactions. Pharmacokinetic profiling (ADMET) indicated favourable absorption and distribution properties for all three compounds, with low toxicity predictions, though some hepatotoxicity was noted. Principal component analysis (PCA) demonstrated that ochratoxin A and permethrin induced substantial alterations in protein dynamics, suggesting ligand-specific structural effects. Experimental validation confirmed the antibacterial activity of ochratoxin A against K. pneumoniae , producing a 34 ± 0.67 mm inhibition zone at 100 µg/disc, surpassing ciprofloxacin (33 mm) with a MIC of 18.33 ± 0.72 µg/mL and MBC of 39.33 ± 1.36 µg/mL ( p < 0.05). Collectively, these in silico and in vitro results highlight fungal metabolites, particularly ochratoxin A, as promising therapeutic leads against MDR K. pneumoniae . However, further in vivo investigations are required to establish their safety and clinical potential. Graphical abstract

Diagnostic inaccuracies are a major yet often overlooked threat to global health, leading to delayed treatment, preventable harm and systemic gaps in disease control. Among the most affected domains are Orthoflavivirus infections, which pose ongoing diagnostic challenges due to antigenic cross-reactivity, overlapping clinical symptoms and the narrow temporal sensitivity of standard tools such as serology and reverse transcription polymerase chain reaction. These constraints have led to widespread misdiagnoses and underreporting, ultimately hampering both effective clinical management and public health response. Recent advances in metagenomic and metatranscriptomic sequencing offer a transformative solution by enabling unbiased, simultaneous pathogen detection and real-time profiling of viral and host transcriptomics. In this review, we assess the diagnostic performance and translational value of these approaches in resolving Orthoflavivirus infections, with case examples from clinical settings in countries like the USA, UK, China and Germany which have already implemented these approaches into routine diagnosis in some settings. We examine key methodological considerations, including optimal sample timing, sample types and processing, sequencing strategy selection and the diagnostic performance of various platforms. We highlight the growing use of metatranscriptomics for detecting active infections, profiling viral and host responses, identifying coinfections and supporting real-time surveillance. We also discuss the key challenges such as technical expertise, lack of standardization, cost, turnaround time and regulatory approval that currently limit global implementation. Finally, we highlight emerging international efforts to integrate sequencing-based diagnostics into routine hospital workflows. Together, these innovations mark a critical shift toward precision diagnostics for Orthoflavivirus infections, with broad implications for clinical settings.

Soil salinity is a major constraint to crop establishment and yield, particularly during germination and early seedling development of Brassica napus. Here

Wildlife-origin viral diseases are an increasing One Health challenge in Australia, with climate change emerging as a key driver of disease emergence, transmission, and spillover. Australian wildlife, particularly marsupials, birds, and bats, serve as important reservoirs of zoonotic and vector-borne viruses. However, the combined influence of wildlife reservoirs and climate change on viral disease ecology in Australia has not been comprehensively synthesised. This review examines the epidemiology, significance, and climate sensitivity of wildlife-associated viral diseases and evaluates how climate change and variability affect reservoir host ecology, vector biology, viral persistence, and host–pathogen–vector interactions. Thirteen major wildlife-associated viruses of One Health importance were identified, with transmission predominantly involving wildlife reservoirs and mosquito vectors. Temperature and rainfall were the principal climatic drivers influencing the transmission of Ross River virus (RRV), Barmah Forest virus (BFV), Murray Valley encephalitis virus (MVEV), Japanese encephalitis virus (JEV), West Nile virus/Kunjin virus (WNV/KUNV), and avian influenza A virus (AIV). For most mosquito-borne viruses (MBVs), temperature exhibited a non-linear relationship with transmission, while above-average rainfall generally increased outbreak risk by promoting mosquito abundance, habitat availability, and reservoir host activity. Significant knowledge gaps remain in wildlife virome surveillance, climate–disease pathways, and integrated One Health surveillance. Strengthening climate-informed surveillance that integrates wildlife, vectors, genomics, epidemiology, and predictive modelling will improve outbreak preparedness and reduce the risk of viral emergence and spillover in Australia.

Clostridioides difficile infection (CDI) remains a significant healthcare challenge, primarily due to its toxin‐mediated pathogenesis that results in severe gastrointestinal complications. This study investigates the inhibitory potential of bioactive compounds derived from Brassica oleracea L., specifically targeting the CDTa subunit of the binary toxin, using both computational and experimental approaches. Molecular docking analysis identified indolylmethyl glucosinolate (−9.1 kcal/mol) and indole‐3‐carbinol (−8.75 kcal/mol) as top candidates, demonstrating high binding affinities to the CDTa protein. Stability and dynamic behavior of the ligand–protein complexes were further assessed through 100‐ns molecular dynamics simulations, which confirmed their stable interactions. Thermodynamic evaluations using MM‐GBSA calculations revealed favorable binding free energies, supporting their potential as effective inhibitors. In parallel, the antibacterial efficacy of these compounds was validated through in vitro antibacterial assays, where indolylmethyl glucosinolate and indole‐3‐carbinol exhibited maximum inhibition zones of 23.33 ± 0.67 mm and 22.67 ± 0.33 mm, respectively, at a concentration of 100 μg/mL. In MIC and MBC assays, both compounds showed significant antibacterial activity, with indolylmethyl glucosinolate demonstrating slightly higher potency (MIC: 10.33 ± 0.72 μg/mL; MBC: 23.33 ± 1.36 μg/mL) than indole‐3‐carbinol (MIC: 11.33 ± 0.27 μg/mL; MBC: 26.67 ± 3.60 μg/mL). Additionally, PCA and DCCM revealed distinct conformational stabilization patterns and correlated motion of CDTa upon ligand binding, supporting a potential antivirulence interaction. Moreover, ADMET analysis revealed differences in their pharmacokinetic and toxicity profiles, providing further insights into their therapeutic potential. Overall, these findings suggest that B. oleracea –derived compounds exhibit a direct antibacterial activity against C. difficile while also demonstrating computationally supported inhibition of CDTa, indicating a complementary antivirulence mechanism that may contribute to their therapeutic potential in CDI.