Morgyn Sylvia Warner

SUMMARY: DNA sequencing has revolutionized microbial surveillance in cystic fibrosis (CF), transforming pathogen identification from culture-dependent to total microbial community identification using molecular-based approaches. Techniques such as 16S rRNA gene sequencing have uncovered the complexity of the CF airway microbiome, while shotgun metagenomics, metatranscriptomics, and viromics now provide strain-level, functional, and viral insights beyond bacterial identification. Despite these advances, key technical and logistical challenges remain, including the processing of high-viscosity sputum samples, overwhelming host DNA contamination, managing large data sets, and the integration of complex bioinformatic outputs into clinical workflows. Emerging innovations such as host DNA depletion protocols, targeted enrichment panels, and adaptive sampling on Oxford Nanopore platforms are helping to overcome these barriers, improving microbial recovery and sequencing efficiency. As cystic fibrosis transmembrane conductance regulator (CFTR) modulator therapies are changing the lives of people with cystic fibrosis (pwCF), sequencing offers an unprecedented opportunity to track potential microbial adaptation in response. This review investigates current advances, limitations, and translational opportunities in DNA sequencing for CF airway microbiome surveillance, highlighting how these technologies can help reshape research and clinical microbiology in the post-modulator era.

Infection is a major cause of morbidity and mortality in myelodysplastic syndromes (MDS), yet infection risk remains incompletely defined in the contemporary treatment era. We conducted a retrospective study of 708 patients with MDS to characterize the incidence, microbiology, temporal dynamics, and predictors of infection-related hospitalization over two decades. Overall, 78.8% (n=558) of patients required hospitalization, of which 69.9% were infection related. Infection-related hospitalization was independently associated with inferior overall survival. In multivariable Cox proportional hazards model, comorbidity burden, red blood cell transfusion dependence, higher IPSS-R risk, exposure to chemotherapy or stem cell transplantation, and severe neutropenia independently predicted infection. Among azacitidine-treated patients, 71.4% experienced infection-related hospitalization, with nearly three-quarters occurring within the first six cycles, identifying a critical early vulnerability window. Neutropenia remained the dominant driver of infection risk; however, immune dysfunction independently increased susceptibility. Low monocyte counts, cytokine dysregulation, and TP53 mutations identified high-risk patients despite preserved neutrophil counts, reflecting impaired myeloid reserve. Consistent with this dynamic vulnerability, recent infection or severe neutropenia (<0.5×10⁹/L) increased subsequent infection risk by 2.47-fold. We observed a concerning shift in antimicrobial resistance, with 14% of Gram-negative infections producing extended-spectrum beta-lactamases, 21% of Pseudomonas aeruginosa isolates resistant to piperacillin-tazobactam, and vancomycin-resistant enterococci prevalence increasing from 14% to 46%. Together, these findings identify infection in MDS as a dynamic, prognostically important complication driven by cytopenia, treatment, and immune dysfunction. This supports time-adapted risk stratification, targeted prevention, and antimicrobial stewardship, particularly during early treatment and high-risk disease phase.

Objective: As antimicrobial resistance (AMR) continues to threaten global public health, bacteriophage therapy products (BTPs) offer a promising alternative to conventional antimicrobials. However, translation into routine clinical practice requires best practice standards for manufacturing and quality control to ensure the consistent safety, quality, and reliability of personalised BTPs produced for individual patients or small cohorts. Design: A modified Delphi methodology was used to develop consensus statements, engaging experts from Australia’s National Bacteriophage Therapy Regulatory Working Group across the fields of clinical microbiology, phage biology, good manufacturing practice (GMP), regulatory science, and government. The process comprised three iterative phases: (1) structured statement development, (2) an anonymous REDCap survey, and (3) a hybrid consensus meeting. The strength of evidence and recommendations was assessed using the GRADE (Grading of Recommendations Assessment, Development and Evaluation) framework. Results: Consensus was reached on 35 statements to provide best practice manufacture and quality control guidance for BTPs. These statements address requirements for phage identification and characterisation; define the point at which GMP-aligned processes commence for ubiquitous phages; outline quality control expectations for phage active pharmaceutical ingredient (pAPI) production and maintenance of BTP and host cell repositories. Additional guidance covers quality management systems, including documentation, traceability, and governance. Conclusion: These consensus statements provide comprehensive best practice recommendations for the manufacture and quality control of BTPs in Australia. By promoting consistent, safe, and quality-assured approaches to personalised BTPs, they aim to facilitate clinical implementation while remaining aligned with existing international pharmacopoeial standards and regulatory frameworks. Significance of this study: What is already known on this subject?: Bacteriophage Therapy Products (BTPs) have re-emerged as a potential solution to antimicrobial resistance (AMR), supported by growing clinical evidence and global research activity. Data available to the regulatory working party includes more than 70 high-risk patients treated with dozens of different bacteriophages under a national open-label protocol (unpubl.). Published guidance from the European Pharmacopoeia, European Medicines Agency (EMA), UK Medicines & Healthcare products Regulatory Agency (MHRA), and World Health Oragnization (WHO) exists, yet no practical standards for personalised BTP manufacture have been established. What are the new findings?: Recommendations have been made to define minimum quality and manufacturing stasndards for the safe, consistent production of BTPs. This is the first expert endorsed, risk-based framework to support the translation of personliased BTPs into routine clinical practice, cobering manufacture, repository management, quality control and supply. How might it impact on clinical practice in the foreseeable future?: These consensus statements, developed by Australia’s National Bacteriophage Therapy Regulatory Working Group, can assist researchers, clinicians and regulatory authorities to develop BTP production protocols.

Achromobacter species are emerging multidrug-resistant (MDR) pathogens in people with cystic fibrosis. Their increasing resistance has grown an interest in phage therapy as an alternative treatment strategy. However, the factors governing phage susceptibility remain poorly understood, thereby limiting the rational selection of phage candidates. Using 15 strictly lytic Achromobacter phages and 7 clinical cystic fibrosis isolates representing Achromobacter insolitus and Achromobacter xylosoxidans, we demonstrate substantial variation in infection efficiency across all 105 phage-host combinations, variation that could not be discerned from qualitative plaque assays alone. We integrated complete bacterial and phage genomes with quantitative efficiency-of-plating (EOP) assays and lineage-aware Bayesian mixed-effects modelling to show that phage infectivity in Achromobacter is governed predominantly by bacterial lineage and strain identity, accounting for 90% of total variance in log-normalised EOP, with individual strains varying substantially in permissiveness irrespective of species membership. After accounting for this lineage structure, no individual defence system, antimicrobial resistance gene class, or phage tail cluster retained a statistically significant independent or interaction association with infectivity. Together, these findings demonstrate that bacterial strain identity is the primary driver of infection outcome. Host defence systems and phage tail-associated genes remain biologically plausible contributors; their independent effect could not be resolved after accounting for lineage structure, indicating that infection outcomes are largely strain-dependent. This work shifts the question from which individual traits predict infection to how strain lineage and specific host-phage combinations jointly determine infectivity and argues that quantitative phenotyping of individual phage-host pairs is essential for guiding phage candidate selection and supporting rational cocktail design against multidrug-resistant Achromobacter infections in cystic fibrosis.

Chronic infections in cystic fibrosis (CF) emerge from gradual ecological transitions in the airway microbiome, yet early predictive markers remain poorly defined. We developed a new autoencoder-based framework that outperforms read-based or metagenome-assembled genome-based analyses at capturing the continuum from health-associated commensals to pathogen-dominated, antibiotic-tolerant communities. This improvement is achieved by integrating taxonomic and functional data from 127 sputum and bronchoalveolar lavage metagenomes from 64 people with CF into latent “Clusters of Phylogeny and Functions” (COPFs). Coupled with gradient-boosted random forests, COPFs predicted Pseudomonas aeruginosa colonisation, multidrug resistance, and impending infection up to a year before clinical detection. The multidrug-resistant P. aeruginosa signature showed the same resistance-mechanism evolution as found in laboratory experiments. The inclusion of eukaryotic markers revealed persistent Aspergillus fumigatus signatures even during culture-negative intervals. Applying our South Australian-trained model to over 1,000 global metagenomes from 22 independent CF datasets, we achieved 94% accuracy in predicting P. aeruginosa status across platforms and geographies, validating the model’s universal utility. Our results demonstrate that combining datasets with deep learning reveals conserved ecological and metabolic mechanisms in disease progression, transforming metagenomics into a predictive framework for managing chronic infections.

Background: Residential aged care facilities (RACFs) are potential reservoirs for methicillin‐resistant Staphylococcus aureus (MRSA), with residents at increased risk due to compromised immunity, comorbidities, frequent hospital visits, and close living quarters. Objectives: This study is aimed at assessing MRSA prevalence in RACFs and analyzing antimicrobial resistance mechanisms both phenotypically and genotypically. Methods: A total of 106 participants were recruited: 84 from three RACFs and 22 from retirement villages (as control samples). MRSA was isolated from nasal, wound, and urine samples using selective media. Identification was confirmed via MALDI‐TOF MS. Antimicrobial susceptibility testing and MRSA screening were conducted using ISO broth microdilution and EUCAST disc diffusion. Whole‐genome sequencing and bioinformatics were used to explore resistance mechanisms and clonal transmission. Results: Nasal carriage of MRSA was 8.2% ( n = 7/85) among RACF residents; none were found in retirement village participants. All isolates formed strong biofilms, and 66.6% (10/15) showed multidrug resistance. Genotypic analysis revealed ST22 as the predominant clone. Genotypic resistance largely matched phenotypic profiles, except for unexplained tetracycline resistance. No isolates carried Panton–Valentine leukocidin (PVL), consistent with previously reported data on MRSA in long‐term care facilities. SNP‐based phylogenetic analysis indicated persistent colonization and circulation of multidrug‐resistant clones within the facilities. Nasal carriage of MRSA was associated with UTIs and/or wound infections in some cases, whereas a UTI caused by the same clone in a resident within the same facility without nasal carriage indicates possible intrafacility clonal dissemination. Conclusion: This study highlights persistent colonization with hospital‐associated MRSA and potential clonal transmission among residents of aged care facilities despite several courses of antibiotics. The predominance of the epidemic ST22 clone and high rates of multidrug resistance underscore the urgent need for targeted infection control, routine genomic surveillance, and antimicrobial stewardship in long‐term care settings to prevent MRSA dissemination.

Background: Data on the health-related quality of life (HRQoL) for invasive meningococcal disease (IMD) survivors, particularly among adolescents and young adults (AYAs), are limited. This study aimed to investigate the in-depth experiences and impacts of IMD on AYAs. Methods: Participants were recruited from two Australian states, Victoria and South Australia. We conducted qualitative, semi-structured interviews with 30 patients diagnosed with IMD between 2016 and 2021. The interview transcripts were analyzed thematically. Results: Of the participants, 53% were aged 15–19 years old, and 47% were aged 20–24. The majority (70%) were female. Seven themes relating to the participants’ experience of IMD were identified: (1) underestimation of the initial symptoms and then rapid escalation of symptoms; (2) reliance on social support for emergency care access; (3) the symptoms prompting seeking medical care varied, with some key symptoms missed; (4) challenges in early medical diagnosis; (5) traumatic and life-changing experience; (6) a lingering impact on HRQoL; and (7) gaps in the continuity of care post-discharge. Conclusion: The themes raised by AYA IMD survivors identify multiple areas that can be addressed during their acute illness and recovery. Increasing awareness of meningococcal symptoms for AYAs may help reduce the time between the first symptoms and the first antibiotic dose, although this remains a challenging area for improvement. After the acute illness, conducting HRQoL assessments and providing multidisciplinary support will assist those who require more intensive and ongoing assistance during their recovery.

Antimicrobial-resistant Klebsiella pneumoniae is one of the predominant pathogens in healthcare settings. However, the prevalence and resistome of this organism within residential aged care facilities (RACFs), which are potential hotspots for antimicrobial resistance, remain unexplored. Here, we provide a phenotypic and molecular characterization of antimicrobial-resistant K. pneumoniae isolated from RACFs. K. pneumoniae was isolated from urine, faecal and wastewater samples and facility swabs. The antimicrobial susceptibility profiles of all the isolates were determined and the genomic basis for resistance was explored with whole-genome sequencing on a subset of isolates. A total of 147 K. pneumoniae were isolated, displaying resistance against multiple antimicrobials. Genotypic analysis revealed the presence of beta-lactamases and the ciprofloxacin-resistance determinant QnrB4 but failed to confirm the basis for the observed cephalosporin resistance. Clonal spread of the multidrug-resistant, widely disseminated sequence types 323 and 661 was observed. This study was the first to examine the resistome of K. pneumoniae isolates from RACFs and demonstrated a complexity between genotypic and phenotypic antimicrobial resistance. The intra-facility dissemination and persistence of multidrug-resistant clones is concerning, given that residents are particularly vulnerable to antimicrobial resistant infections, and it highlights the need for continued surveillance and interventions to reduce the risk of outbreaks.