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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.

Liver cancer is the fourth most deadly cancer, and early detection and timely treatment apparently play a crucial role in it. Intestinal bacteria affect the development of liver cancer through various pathways. In this study, the gut bacteria of liver cancer patients are analysed in detail by using metagenomic sequencing technology, and some of the bacterial species and metabolic pathways that may affect the development of liver cancer have been identified. Additionally, we identified bacterial factors that may impact key clinical indicators of the tumour. The findings of this study provide a scientific foundation for understanding the mechanisms underlying liver cancer development. This study freshened insights into clinical treatment strategies for liver cancer.

Background. The novel coronavirus disease 2019 (COVID-19) has highlighted vulnerabilities in healthcare systems and has brought the world to a standstill single-handedly. Diagnostic testing for COVID-19 is critical for understanding epidemiology, contact tracing, case management and controlling the transmission of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Real-time reverse transcription PCR test, the gold standard test, involves fairly complex steps and takes nearly 24-48 h to generate the results. GeneXpert is a rapid nucleic-acid-detection-based test approved by the Indian Council of Medical Research, which can shorten the turnaround time significantly. Aim. The study aimed to compare the performance of GeneXpert against the gold standard real-time reverse transcription PCR for SARS-CoV-2. Materials and methods. This retrospective study was conducted at a tertiary care centre from 25 March 2020 to 8 December 2020. The nasopharyngeal/oropharyngeal swabs that were sent to the Viral Research and Diagnostic Laboratory for testing of SARS-CoV-2 by GeneXpert [cartridge-based nucleic acid amplification test (CBNAAT)] were included for the study. A total of 270 samples (220 samples positive and 50 samples negative for SARS-CoV-2 by GeneXpert) were simultaneously tested for real-time reverse transcription PCR. Real-time reverse transcription PCR was considered as gold standard test (reference) for calculating the sensitivity and specificity of the GeneXpert (CBNAAT) test. Results. Out of the total samples tested (n=270) for SARS-CoV-2, 220 were positive, and 50 were negative for SARS-CoV-2 by GeneXpert. Among 220 GeneXpert SARS-CoV-2-positive samples, 118 (53.64%) were also positive by real-time reverse transcription PCR, while 102 (46.36%) showed negative results by real-time reverse transcription PCR. However, 50 GeneXpert negative samples showed 100% agreement with real-time reverse transcription PCR, i.e. they were also negative by real-time reverse transcription PCR. The GeneXpert sensitivity and specificity for COVID-19 were seen to be 100% (95% CI: 96.92-100%) and 32.89% (95% CI: 25.50-40.97%), respectively, as compared to the gold standard real-time reverse transcription PCR test. The positive predictive value and negative predictive value of GeneXpert for COVID-19 were found to be 53.64% and 100%, respectively. Conclusion. This study highlights that the GeneXpert test is highly sensitive and found to be useful in emergency and challenging situations for rapidly ruling out negative cases. However, due to its relatively low specificity, positive results should be confirmed with real-time reverse transcription PCR. Therefore, it can serve as a valuable tool for patients requiring urgent care by facilitating early diagnosis and management of COVID-19, ultimately contributing to preventing morbidity and mortality.

The increasing complexity and collaborative nature of scientific research projects underscore the need to implement project management practices to manage resources and funding, ensure data quality and prevent delays in project progress. Here, we introduce three major project management methodologies, including agile, waterfall and hybrid approaches, and explore their suitability for biological and microbiological research laboratories. Variables that may influence choosing an appropriate strategy for managing projects are considered, including the size and experience of a research group. In the following article, we provide an overview of the five major stages of project planning and execution, focusing on implementing each of the discussed strategies in the research laboratory. Furthermore, we discuss the composition of the research team and outline the responsibilities assigned to each team member based on their role in the project. This paper highlights potential risks and challenges that may negatively impact research progress, underscoring the need for proper project planning. Applying proper project management methodologies is often neglected in academic research, leading to serious delays and waste of valuable resources.

Blastomycosis is a serious fungal disease affecting humans and animals. It is typically caused by the thermally dimorphic fungus, Blastomyces dermatitidis. In this report, we describe an infection caused by the cryptic fungal species, Blastomyces gilchristii, in a tiger (Panthera tigris).