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Background. Pseudomonas aeruginosa is a key pathogen in cystic fibrosis (CF), driving pulmonary decline and exhibiting resistance through virulence factors and adaptive mutations. Cefiderocol (FDC) is a novel siderophore cephalosporin with activity against Gram-negative bacteria. We aimed to assess the in vitro efficacy of FDC against P. aeruginosa isolates in a CF population. Methods. The study was conducted in a tertiary hospital with a specialist adult CF service. All first isolates of significant respiratory pathogens among this cohort are cryopreserved at -80 °C. Antimicrobial susceptibility testing to FDC was performed as per European Committee on Antimicrobial Susceptibility Testing Disk-Diffusion (version 10) for all stored isolates of P. aeruginosa from 2017 to 2022 inclusive. Results. Eighty-five isolates from seventy-one patients were included. Resistance phenotypes comprised 19% (n=16) multidrug-resistant (MDR), 16% (n=14) extensively drug-resistant (XDR) and 24% (n=20) pandrug-resistant (PDR), with 24 % (n=20) exhibiting the mucoid phenotype. Overall, 85% of isolates were susceptible to FDC, with a mean inhibition zone of 25.2 mm. Antimicrobial activity was retained in 81% of MDR, 86% of XDR, 60% of PDR and 90% of mucoid isolates. Seventy-four per cent of meropenem-non-susceptible isolates remained susceptible to FDC, compared with lower susceptibility to ceftolozane-tazobactam (42%), tobramycin (36%) and ciprofloxacin (22%). Conclusion. FDC exhibited excellent in vitro activity against P. aeruginosa from adults with CF, including highly resistant and mucoid phenotypes. These findings highlight its potential as a salvage option in this high-risk population and provide the first Irish surveillance data to inform antimicrobial stewardship and future clinical use.

Azoles inhibit the cytochrome P450-dependent enzyme lanosterol 14α-demethylase (CYP51) that is encoded by the ERG11 gene. Azole resistance in Candida species arises through different mechanisms, like mutations in the ERG11 gene, ERG11 overexpression, CDR1,2 (Candida drug resistance) overexpression that actively efflux azole drugs, reducing their intracellular concentration and therapeutic effectiveness, and biofilm formation. We sequenced the ERG11 gene to determine mutations in the coding and non-coding regions of ERG11 in clinical isolates of Candida glabrata (Nakaseomyces glabratus) from Pakistan. Eight C. glabrata (N. glabratus) strains from our fungal strain bank (five fluconazole-resistant and three susceptible dose-dependent) were revived and used. The ERG11 gene was amplified by PCR, sequenced using the Sanger methodology and analysed using bioinformatic tools. We identified a change in nucleotide at c. -66 T/G upstream of the start codon ATG in the promoter region of the ERG11 gene in fluconazole-resistant C. glabrata (N. glabratus). Within the downstream (coding region), where numbering begins at the ATG start codon as position +1, two novel synonymous mutations at positions T300C and T834C and previously reported synonymous mutations T768C, A1023G, T1557A and A1581G were also observed. This is the first study evaluating ERG11 mutations in C. glabrata (N. glabratus) from Pakistan. The clinical significance of such uncommon ERG11 gene mutations, such as c. -66 T/G, should be explored further through correlation with treatment outcome data.

Extra-colonic infections caused by Clostridioides difficile are exceptionally rare, with prosthetic joint infections (PJIs) comprising only a small fraction of the reported cases. Moreover, there is limited guidance on the optimal management of such infections. We present the case of a 76-year-old man who developed a left hip PJI due to C. difficile 6 weeks after undergoing surgical revision for a periprosthetic fracture. Given the complexity of the case, curative surgical intervention was not considered feasible. The patient was treated with repeated debridement, intravenous vancomycin and oral (PO) metronidazole, followed by successful suppression with PO doxycycline - a novel therapeutic approach not previously documented. To date, only seven cases of C. difficile-associated PJI have been reported in the literature; this is the first known instance in which suppression of a C. difficile PJI has been achieved and the first to utilize whole-genome sequencing for further analysis of the isolate.

Chlorous acid water and sodium hypochlorite solution are effective disinfectants against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the virus that caused the pandemic. Recent studies have shown that both compounds have equivalent inactivation effects when tested on purified viruses. However, in practical applications, the presence of organic matter is common and can significantly affect disinfectant performance. We conducted several experiments comparing these two disinfectants under different conditions to better understand their practical efficacy. When an infected cell culture medium (serum-free) was used as the test virus, chlorous acid water and sodium hypochlorite solution showed reduced efficacy. This decrease was attributed to the presence of aa in the medium. Notably, sodium hypochlorite solution showed a more pronounced reduction in potency compared with chlorous acid water. In addition, we evaluated the SARS-CoV-2 inactivation effects of chlorous acid water and sodium hypochlorite solution under various organic loading conditions simulating real-world contamination scenarios such as blood, vomit and saliva. The organic materials used included BSA, SRBCs, polypeptone, FBS and artificial saliva. The results showed that chlorous acid water demonstrated superior resilience to organic matter interference compared with sodium hypochlorite solution. These results suggest that chlorous acid water may be more effective than sodium hypochlorite solution in inactivating viruses on contaminated surfaces, particularly in healthcare settings where organic contamination is common. In summary, our research suggests that chlorous acid water may be a more effective disinfectant in practical settings.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection triggers complex host responses, including alterations in RNA transcription and modification. Understanding these changes is crucial for elucidating viral pathogenesis and identifying potential therapeutic targets. We used direct RNA sequencing to comprehensively profile the transcriptomic and epitranscriptomic landscapes of human HEK-AT cells infected with SARS-CoV-2 at 8 h post-infection, compared to mock controls. We analysed viral and host transcriptomes, focusing on gene and transcript expression, isoform usage and RNA m6A modifications. Viral RNA sequencing reads showed 3' end-biassed coverage indicative of subgenomic RNA synthesis, with high expression of N gene subgenomic RNA reads. Sixteen m6A modification sites were consistently identified in the viral genome, primarily within the ORF1ab and S genes. In the human transcriptome, we found 254 positions with significantly altered m6A modification rates, with 119 showing decreased modification and 135 showing increased modification in infected cells. Genes with decreased m6A modifications were enriched in the neurotrophin signalling pathway. Transcript-level analysis identified 19 upregulated and 12 downregulated transcripts. Notably, transcript discovery and quantification revealed a novel isoform of the HIST1H2BK gene, which was significantly more expressed in infected cells compared to mock controls. Isoform switching analysis revealed 24 significant switches involving 21 genes, implicating mitochondrial reprogramming and immune-related pathways. In conclusion, this study provides a detailed, direct RNA sequencing-based characterization of host-virus RNA interactions, revealing key insights into SARS-CoV-2 infection mechanisms and potential therapeutic targets.

Genitourinary tuberculosis is a severe form of extrapulmonary tuberculosis. The kidneys are the most commonly affected organs, followed by the epididymis, testicles, bladder, ureter and prostate. Notably, epididymal tuberculosis is one of the forms of genital tuberculosis presenting with specific clinical features, which may include epididymitis, orchid-epididymitis or hydrocele. We report the case of a patient with a hydrocele that revealed epididymal tuberculosis. Utilizing molecular biology techniques, a diagnostic test for epididymal tuberculosis was established. The patient was treated conservatively with tuberculosis medication for 6 months.

Among the five MLST clades that define the global population structure of the bacterial pathogen Clostridioides difficile, Clade 2 has received special attention due to the global spread, clinical severity and hospital prevalence of ST01 strains. To identify features potentially contributing to the historically attributed higher virulence and epidemic potential of ST01 strains, we compared a range of phenotypic traits across the infection cycle between clinical Clade 2 ST01 and non-ST01 strains from ST41, ST47, ST67, ST154 and ST638. We found no significant differences in canonical virulence-associated characteristics such as spore adherence, motility, biofilm formation and resistance to a disinfectant. However, ST01 strains exhibited distinct profiles in surface layer protein A (SlpA)-mediated immune activation and toxin B (TcdB)-induced cytotoxicity that were consistent with allelic divergence. These findings highlight the need to reconsider current paradigms of Clade 2 hypervirulence and underscore the importance of allele-specific phenotypic variation in developing targeted public health strategies.

Objective. To evaluate the performance of a novel immunochromatographic (IC) card test (TRURAPID^® O.K.N.V.I. RESIST-5) for rapid detection of five carbapenemase enzymes in metallo-beta-lactamase (MBL)-producing organisms, compared to real-time PCR and an Advanced Expert System (AES). Methods. Clinically isolated 100 non-duplicates of multidrug-resistant Gram-negative bacilli expressing MBL production were tested using the novel IC card, real-time PCR and the Vitek-2 AES. Sensitivity, specificity and turnaround time were evaluated. Results. The novel IC card showed high sensitivity for detecting NDM (93%) and KPC (91.7%) carbapenemases, but lower sensitivity for OXA-48 (60%), VIM (67%) and IMP (33%) compared to PCR. It had a rapid turnaround time of 15-20 min versus 5-7 h for PCR and 18-22 h for AES. Conclusion. The novel IC card offers a rapid, cost-effective approach for detecting carbapenemases, particularly NDM and KPC, in clinical microbiology practice. It may be beneficial in resource-limited settings where these enzymes are prevalent. Considering the limited sensitivity for the IMP and VIM genes, this warrants confirmatory testing by PCR. Further evaluation is needed to assess its role as a screening or confirmatory test, especially during nosocomial outbreaks.

Introduction. Optimization of diagnostic testing is essential for the sustainable delivery of laboratory services. To date, little consideration has been given to the potential benefits of diagnostic stewardship to laboratories looking to reduce their environmental footprint. Hypothesis. Implementing a pre-analytical diagnostic stewardship intervention for the testing of superficial wound swabs would result in a measurable reduction in the environmental footprint of the microbiology laboratory. Aim. To assess the consequential impact of a diagnostic stewardship intervention on test volume, carbon footprint and quantity of non-recyclable plastic waste generated. Methodology. Superficial wound swabs received in the absence of clinical details suggestive of active skin and soft tissue infection were rejected by the laboratory. The carbon footprint of testing was estimated using Publicly Available Specification 2050:2011 methodology in a cradle-to-grave, attributional life-cycle assessment within a defined system boundary. The mass of laboratory plastic waste was calculated through the accurate weighing of associated laboratory consumables. Results. The intervention resulted in a reduction of 35.77 kg CO₂e and 9.06 kg of unrecyclable plastic waste over an 8-day period without measurable patient harm. Conclusion. This study demonstrates, in relation to specific testing pathways, that the optimization of microbiology laboratory diagnostic testing can result in a reduction in greenhouse gas emissions and non-recyclable plastic waste generation without negatively impacting patient care.

Introduction. Human metapneumovirus (hMPV), first identified in 2001, is one of the major respiratory pathogens causing acute respiratory tract infections (ARTIs). In Sri Lanka, data on epidemiology and clinical characteristics of hMPV infections are limited. In this study, we aimed to investigate the epidemiology and clinical characteristics of hMPV infection in adults and children with ARTIs in different locations in Sri Lanka from January 2021 to December 2023. Methods. A total of 1,582 respiratory samples from patients with ARTIs were enrolled from four tertiary care hospitals. Nasopharyngeal swab samples were subjected to real-time reverse transcriptase PCR to test for hMPV using a commercial multiplex assay. Demographic and clinical data were extracted from the patients' clinical records. A selected subset of positive samples was subjected to genomic sequencing using an amplicon-based approach with the Respiratory Pathogen ID/AMR Library Prep and Enrichment Kit using the Illumina platform. Results. hMPV infection was identified in 1.64% (26/1,582) of patients, with the majority being children under 5 years of age. The co-infection rate was 0.34% with other respiratory viruses. The most common clinical presentation in hMPV infection included acute upper respiratory tract infection with fever, cough and cold and sore throat. Conclusion. hMPV is an important respiratory pathogen in children, causing ARTIs. hMPV-infected patients showed a range of respiratory symptoms with varying severity ranging from common cold to life-threatening lower respiratory tract infections. Continuous surveillance on hMPV infection may help in monitoring the hMPV activity, which will help in tracking the emergence of hMPV infections.