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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).

The kynurenine (KYN) pathway is the major catabolic pathway for tryptophan in humans, producing several metabolites that influence health. In clinical settings, KYN levels serve as a valuable biomarker for the diagnosis and prognosis of inflammatory and neurological diseases. Nevertheless, KYN detection relies on mass spectrometry analysis, which requires specialized knowledge and expertise with high operational costs. The bacterial biosensor presents as a promising tool for rapid and cost-effective targeted substance detection due to its ease of genetic modification. Therefore, this study aimed to develop an engineered bacterial biosensor by integrating a genetic module in a plasmid designed for KYN detection harboured in an Escherichia coli chassis. The KYN biosensing component in the genetic module encodes a KYN pathway regulator (KynR) from Pseudomonas aeruginosa, driven by the P_BAD arabinose-inducible promoter. Upon expression, KynR would bind to the exogenous KYN and the bacterial responding kyn promoter to express the downstream green fluorescent protein gene to emit a fluorescence signal. However, despite successful induction by arabinose and the presence of KYN, biosensors with different gene orientations and genetic components failed to produce a significant fluorescence signal. These findings suggest that the sensitivity of P. aeruginosa KynR is insufficient to detect physiological levels of KYN. Further exploration of alternative biological sensing components is warranted.

Introduction. Despite the introduction 40 years ago of effective and low-cost treatment for tuberculosis (TB), morbidity and mortality from this disease remain substantial worldwide. According to the WHO, TB is once again the leading cause of death worldwide from a single infectious agent. In 2023, TB caused ~1.25 million deaths, surpassing COVID-19. In Morocco, the number of new TB cases rose from 30,897 in 2017 to 35,000 in 2019, highlighting a concerning upward trend that underscores the persistent challenge TB poses to the country's public health system. The incidence of multidrug-resistant (MDR) or rifampicin (RIF)-resistant TB was estimated at 1.7 per 100,000 inhabitants. Isoniazid (INH) is a cornerstone of first-line TB treatment, and resistance to it, even in the absence of RIF resistance, is associated with delayed treatment response, higher rates of treatment failure or relapse and increased risk of progression to MDR-TB if not promptly identified and appropriately managed. Moreover, current diagnostic algorithms in many settings, including Morocco, may miss INH monoresistance due to their reliance on rapid molecular tests that primarily detect RIF resistance, further emphasizing the emerging threat of drug-resistant TB. Despite this, national data on INH monoresistance remain scarce. Given the increasing burden of TB and the critical importance of early detection of drug resistance, it is essential to better understand patterns of resistance beyond RIF. It is within this context that we conducted the present study, which aims to investigate INH resistance in TB cases (pulmonary or extrapulmonary, new or previously treated) over a period of 3 years. Materials and methods. This is a retrospective study conducted at the Bacteriology Department of Mohammed V Military Instruction Hospital over a period of 3 years. Data were collected via the laboratory information system. Clinical samples underwent treatment using both conventional bacteriological methods and molecular techniques. The study of resistance to major anti-TB drugs was performed using the reverse hybridization technique, specifically the HAIN method (GenoType^® MTBDR plus by Hain Lifescience). Statistical analysis was performed using IBM SPSS Statistics 19 and Microsoft Excel 2019. Results. The study involved 464 patients treated for pulmonary and extrapulmonary TB, including both new cases and those previously treated with positive cultures. The mean age of the patients was 42.2 years, with a range from 8 to 88 years. There was a predominance of males at 74%, with a sex ratio of 2.8. Pulmonary sputum samples accounted for 84.8% of the cases, whereas extrapulmonary samples represented only 15.2%, and the positivity rates for direct examination and culture across all samples were 74% and 100%, respectively. INH resistance had a prevalence of 9% (43 out of 464). Genetic mutations observed indicated that 63% of the clinical isolates resistant to

Cryptosporidium, a protozoan parasite of significant public health concern, is responsible for severe diarrhoeal disease, particularly in immunocompromised individuals and young children in resource-limited settings. Analysis of whole-genome next-generation sequencing (NGS) data is critical in improving our understanding of Cryptosporidium epidemiology, transmission and diversity. However, effective analysis of NGS data in a public health context necessitates the development of robust, validated computational tools. We present Parapipe, an ISO-accreditable bioinformatic pipeline for high-throughput analysis of NGS data from Cryptosporidium and related taxa. Built using Nextflow DSL2 and containerized with Singularity, Parapipe is modular, portable, scalable and designed for use by public health laboratories. Using both simulated and real Cryptosporidium datasets, we demonstrate the power of Parapipe's genomic analysis for generating epidemiological insights. We highlight how whole-genome analysis yields substantially greater phylogenetic resolution than conventional gp60 molecular typing in Cryptosporidium parvum. Uniquely, Parapipe facilitates the integration of mixed infection analysis and phylogenomic clustering with epidemiological metadata, representing a powerful tool in the investigation of complex transmission pathways and identification of outbreak sources. Parapipe significantly advances genomic surveillance of Cryptosporidium, offering a streamlined, reproducible analytical framework. By automating a complex workflow and delivering detailed genomic characterization, Parapipe provides a valuable tool for public health agencies and researchers, supporting efforts to mitigate the global burden of cryptosporidiosis.

Large-scale skin microbiome studies are often restricted due to the need for participants to visit a research centre to have their skin swabbed by a trained individual. If samples taken by participants at home returned high-quality data, similar to that generated from samples taken by trained experts under controlled conditions, it would provide the potential for studies to have larger cohorts, include participants from multiple locations and facilitate longitudinal sample collection. Here, we describe the development of a novel unsupervised skin microbiome sample collection method and compare the data quality with that of supervised, in-lab sample collection. We enrolled 57 participants to collect skin swabs from their axillae, forearms, cheeks and scalps. Initially, samples were collected in our research centre under strict supervision by a trained expert. Participants then collected swabs from the same body sites 24 h later, unsupervised, at home, which they returned to the research centre within 3-5 days. All samples then underwent bacterial DNA extraction and 16S rRNA gene sequencing. Yield of extracted bacterial DNA was different depending on body site, with the dry swabs from the forearm producing the lowest amount. There were no significant differences in alpha and beta-diversities between supervised and unsupervised sampling methods, regardless of body site. Taxonomic analysis of bacterial genera also did not differ for axilla, cheek or scalp. Our data suggest that self-sampling skin microbiome methods can produce data that are comparable to samples collected under the supervision of a trained expert in lab settings. These findings should encourage the scalability of future research and allow for greater representative population diversity in genomic and microbiome research.

Rapid and accurate detection of vancomycin-resistant enterococci (VREs) can aid in the early application of appropriate antimicrobial treatment and the implementation of infection prevention measures. The VIASURE real-time PCR assay (Certest Biotec) is a multiplex nucleic acid-based in vitro diagnostic test intended for the detection of vanA and vanB, and geneLEAD VIII (Precision System Science) is a customizable, fully automated molecular detection platform. We evaluated the performance of the VIASURE assay on the geneLEAD VIII platform against 200 clinical enterococcal isolates consisting of 151 VREs and 49 vancomycin-susceptible enterococci collected in Japan, primarily in the Kinki region, and compared it to that of the in-house reference multiplex PCR assay. The performance of the VIASURE assay for the detection of vanA and vanB was comparable to that of the reference multiplex PCR assay, with both a sensitivity and specificity of 100%. Compared with the reference PCR assay, the VIASURE assay reduced the turn-around time by ~3 h 40 min (5 h 34 min vs. 1 h 54 min) and the hands-on time by 46 min per four samples. Fully automated molecular detection of vanA and vanB using the VIASURE assay and geneLEAD VIII for bacterial isolates can enable fast and reliable testing while reducing labour and is a promising tool for clinical laboratories and nosocomial infection control.

Background. Human immunodeficiency virus (HIV) is the major cause of failure to reach targets of tuberculosis (TB) control in settings with high HIV loads. TB, on the other hand, enhances the progression of HIV infection to AIDS. This study was done to understand the epidemiological and clinical profile of HIV-TB co-infected patients and to study the impact of TB on the recovery of CD4 counts. Methodology. An observational study was conducted in which of the 573 patients newly diagnosed with HIV infection and enrolled at the antiretroviral therapy (ART) centre, King George's Medical University, Lucknow, between May 2021 and June 2022, 80 patients who also had newly diagnosed TB were included. These HIV-TB co-infected patients were analysed for demographic factors. Also, clusters of differentiation 4 (CD4) counts were done at the time of enrolment on ART and then later, ~6 to 8 months of recieving ART and anti-tubercular treatment (ATT) initiation. For comparison, of the 493 HIV-only patients, 50 age- and gender-matched consecutive patients for whom baseline and follow-up CD4 counts were available were enrolled as controls. The change from baseline CD4 count was calculated using a paired t-test and Wilcoxon signed rank test. Results. In the present study, among HIV-TB co-infected patients, baseline CD4 levels were 194.52±162.27, and follow-up CD4 levels were 285.09±170.33. A statistically significant increment of 90.57±165.60 in mean CD4 levels was observed (t=4.019; P<0.001). Likewise, in only HIV-positive patients, a statistically significant increment of 125.26±191.48 (35.75%) cells in mean CD4 levels was observed (t=4.626; P<0.001). The increase in CD4 counts in HIV only population was significantly higher than that observed in HIV-TB co0infected patients. Conclusion. Though significant rise in CD4 counts was observed in both HIV-TB co-infected patients and HIV-only patients after 6 to 8 months of appropriate therapy, the rise was significantly higher among the HIV-only group as compared to the HIV-TB co-infected group.

Objective. Hepatitis B virus (HBV) spontaneous mutations may impact the severity of liver disease. This study aimed to assess the mutations in the pre-core (PC) region in HBV-HIV (human immunodeficiency virus) co-infected patients. Additionally, we explored its association with genotypes and examined the clinical implications. Methods. A total of 100 HBV-HIV co-infected patients and 50 HBV mono-infected patients were included in the study. We focused on the PC region of the HBV genome, sequencing it to identify PC mutant variants. PCR products were quantified via spectrophotometry and sequenced using the Sanger method. The resulting sequences were assembled, annotated and aligned in a single reading frame. Subsequent mutational and phylogenetic analyses were performed using UGENE software to determine the genotypes of the isolates. Results. The PC region was successfully amplified and sequenced in 27 samples, comprising 16 from HBV-HIV co-infected patients and 11 from HBV mono-infected patients. Phylogenetic analysis identified two HBV genotypes: genotype D, which was predominant and found in 24 samples (88.9%), and genotype A, present in 3 samples (11.1%). A T-to-C mutation at nucleotide position 1912 was detected in 48.1% of the patients. Furthermore, several additional PC mutations were observed, including A1850T, C1858T, G1899A, G1862T, G1951T, T1812C and T1809G, along with novel mutations such as C1936T, A2011G, T2020A and C2044T. Notably, the prevalence of these PC mutations did not significantly differ between the HBV mono-infected and HBV-HIV co-infected groups. Conclusion. This study underscored the prevalence of PC mutations in HBV-HIV co-infected patients. Although several of these mutations have been previously reported, our findings also revealed novel variants. Further research is needed to elucidate the clinical significance of these new mutations.

Diarrhoeal diseases remain a significant global health challenge, particularly in developing regions such as Africa, Asia and Latin America, where they are a leading cause of child mortality. Contaminated food, including raw or undercooked vegetables, is a major transmission route for diarrhoeal pathogens such as norovirus, Campylobacter, non-typhoid Salmonella and pathogenic Escherichia coli. This study aimed to assess the prevalence of enterotoxigenic E. coli (ETEC), a key diarrhoeal pathogen, in fresh produce and prepared salads in Mexico City. A total of 128 samples, including prepared salads (lettuce, carrots and tomatoes) and unprocessed coriander and lettuce, were analysed over 2 years using protocols from the Bacteriological Analytical Manual and the Official Mexican Standard (NOM) SSA 210. Genotyping was performed to detect ETEC-specific virulence genes encoding heat-stable and heat-labile enterotoxins (st and lt), respectively. ETEC was identified in 9.9% of the total samples, representing 51.56% of the confirmed E. coli isolates. Contamination rates varied by food type, with coriander showing the highest prevalence (78.78%), followed by lettuce (9.09%) and prepared salads from La Vicentina Market (9.09%) and La Purísima Market (3.03%). Genotyping revealed that 12.12% of the ETEC-positive samples carried both st and lt genes, while 33.3 and 54.6% carried only the lt or st gene, respectively. In lettuce samples, 9.09% were positive for ETEC, with 3.03% carrying the lt gene, 3.03% the st gene and 3.03% both genes. Similarly, in coriander, 21.21% were positive for the lt gene, 51.51% for the st gene and 6.06% for both genes. These findings highlight the widespread presence of ETEC in fresh produce sold in Mexico City, posing a significant public health risk, particularly given the increasing consumption of raw vegetables. The study provides the first reported data on ETEC contamination ratios in Mexico City, emphasizing the urgent need for improved food safety measures, including better hygiene practices during production, handling and preparation of fresh produce. This research underscores the importance of ongoing surveillance and preventive strategies to mitigate the risk of foodborne diarrhoeal diseases in urban populations.