Botany

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    Determination of bacterial resistance profile of chronic wound pathogens and genetic characterisation of staphylococcus aureus from patients at meru teaching and referral hospital, Meru county, Kenya
    (Chuka University, 2025) Silas Rael Gacheri
    Chronic wound infections remain a major public health concern, particularly in low- and middle-income countries, where they are often complicated by polymicrobial colonization and the emergence of multidrug-resistant (MDR) pathogens. Effective management is challenged by incomplete pathogen identification, which hampers accurate diagnosis and treatment. This study aimed to determine the prevalence, bacterial profile, and antimicrobial resistance patterns of pathogens associated with chronic wound infections at Meru Teaching and Referral Hospital (MeTRH) in Kenya, and to provide genomic insights into their resistance and virulence determinants. A cross-sectional survey design was employed, in which 68 wound swab samples were collected from a stratified random sample of 293 patients over eight months, following consent. Data were analyzed using SPSS for epidemiological trends, R for visualization, and bioinformatics pipelines for sequencing data processing, assembly, annotation, and phylogenetic reconstruction to characterize antimicrobial resistance and virulence profiles. The findings indicate that the highest burden of chronic wounds occurred among patients aged 21–30 years (22%) and 51–60 years (19.2%). The predominant pathogens were Staphylococcus aureus (26.6%), Pseudomonas aeruginosa (13.2%), and Escherichia coli (10.3%). Clinically significant MDR organisms included Klebsiella pneumoniae, Proteus hauseri, Morganella morganii, Acinetobacter baumannii, and Enterococcus faecalis, while 20.5% of wounds showed no bacterial growth despite clinical evidence of infection. Notably, 50% Staphylococcus aureus isolates exhibited resistance to multiple antibiotics. Distinct susceptibility patterns were observed for Morganella morganii and Proteus spp, whereas coagulase-negative staphylococci displayed variable meropenem resistance reaching up to 100% in some cases. Klebsiella and Pseudomonas spp. were largely sensitive to meropenem, although resistance ranged from 0% to 11%, depending on the species. Whole-genome sequencing revealed frequent resistance determinants, including MecA (60%), blaZ (60%), aadD/aphA-3 (60%), mrsA (70%), ermB (30%), ermC (10%), ermA (20%), rpob_H481N (20%), and grlA_S80F; GrA_S84L (20%), reflecting co-occurrence of beta-lactam, MLS, sulfonamide/trimethoprim, tetracycline, aminoglycoside and quinolone resistance genes. Virulence markers such as aureolysin and toxin/exoenzyme genes (edinB, hlgA/B/C, lukPV, sak, scn, splA/B/E) were identified, alongside diverse spa types (t355-centered clusters), mosaic SCCmec elements, and varied MLST lineages (MLST/spa) were observed across Staphylococcus aureus. Plasmid replicons detected included Rep5a (n = 5), Rep16 (n = 4), Rep7a, Rep10, Rep9a, Rep9b, RepUS70, Rep21, Rep13, and Rep15, suggesting circulation of MRSA and MRSA-like lineages with mobile genetic elements. Clonal complexes CC30, CC121, CC45, and MLST 152 reflected both community- and hospitalassociated strains, indicating dynamic Staphylococcus aureus populations shaped by clonal expansion and horizontal gene transfer. The findings highlight the substantial antimicrobial resistance burden in chronic wound infections, the importance of integrating advanced molecular diagnostics with traditional surveillance, and the need for antibiotic stewardship programs. Moreover, the genomic diversity and virulence architecture revealed here highlight the necessity for localized surveillance strategies and virulence-informed risk stratification to guide infection control and therapeutic interventions.
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    Characterization of ochratoxigenic fungi in dry coffee cherry, effect of post-harvest handling practices and factors affecting its prevalence in Tharaka Nithi county, Kenya
    (Chuka University, 2024) Gitonga Tabitha Kanini
    Coffee is an economically significant cash crop valued for its unique flavour, aroma, and potential health benefits. In Kenya, most research efforts have focused on wet-processed coffee beans, with little attention directed towards the dry-processed cherry, known as dry coffee cherry, which is roasted and consumed locally. There remains a glaring gap in our understanding of ochratoxin A (OTA) occurrence in dry coffee cherry, particularly in major coffee-producing areas like Tharaka Nithi County. The pathogenic fungal species responsible for OTA production in coffee have not been adequately characterized. The Objectiveof this study was to investigate the prevalence of Ochratoxin A in dry coffee cherry in Tharaka Nithi County and its relationship with socio-economic factors and post-harvest handling practices for improved control and management. A cross-sectional design was used and the target population of coffee farmers drawn from three sub-counties. A cluster random sampling was applied to collect dry coffee cherry samples and information from farmers through a semi-structured questionnaire. A total of 143 farmers were sampled from 44 villages. Pathogens were isolated on Potato Dextrose Agar (PDA) media and OTA production assessed according to the International Commission on Microbiological Safety for Foods. Morphological characterization of fungal isolates was done using cultural characteristics while molecular identification was done using Sanger sequencing. Data was analysed using the SPSS version 28 and SAS version 9.4. Cluster analysis was carried out using Eucleadian distances. The fungal isolates obtained were aligned and phylogenetic tree was constructed using neighbor-joining clustering as implemented in MEGA X version 11. The results of the study revealed a high prevalence of Ochratoxigenic fungi in coffee samples across the different AEZs, with Aspergillus niger being the predominant fungal species. Ochratoxin A was detected by Enzyme-Linked Immunosorbent Assay (ELISA). The analysis of colony-forming units (CFU) across agroecological zones showed significant differences, with the highest CFU recorded in UM1 (941.14), followed by UM2 (852.06), and the lowest in UM3 (716.56). Genetic variability was measured using Principal Component Analysis (PCA) and Cluster Analysis. The Eigenvalues and proportion explained variance from the PCA provide insights into the genetic variability, with the first three components accounting for 77.82% of the total variation. Coefficient of Variation (CV) was used, with a CV of 27.59% indicating moderate variability in colony-forming units across samples. R-squared values were also used to indicate the extent of variability explained by agroecological zones and other factors. The levels of OTA contamination were influenced by several factors which includes; inadequate drying practices and improper storage conditions were identified as major contributors to the growth of Ochratoxin A fungi, Agro-ecological zones with higher humidity and warmer temperatures, socioeconomic factors such as farmers’ level of education, awareness about mycotoxins, and access to proper post-harvest facilities played a crucial role in determining the levels of contamination. Coffee beans stored for longer periods in humid, poorly ventilated conditions showed higher contamination levels. The results highlight the urgent need for interventions aiming at improving post-harvest handling practices to reduce OTA contamination. The study emphasizes the importance of educating farmers about the risks of mycotoxins. The findings also provide a basis for developing policies and interventions that can mitigate the risk of mycotoxin contamination and improve the overall quality and safety of Kenyan coffee product.