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Item type:Item, Immortalization of primary lamb kidney cells towards a continuous cell line for propagation of foot and mouth disease virus(Botswana International University of Science and Technology (BIUST), 2025-07) Gaboiphiwe, Kitsiso; Lebani, Kebaneilwe; Masisi, Kabo; Fana, E.MFoot and mouth disease (FMD) is a highly contagious disease of great economic importance. The disease is caused by foot and mouth disease virus (FMDV). Diagnosis of FMD is done through tests such as virus isolation (VI), enzyme linked immunosorbent assay (ELISA), virus neutralisation tests (VNT) and polymerase chain reaction (PCR) platforms. Tests such as VI and VNT require the use of cells that are susceptible to infection with FMDV. Cells that can be used for FMDV culture include primary cells and cell lines developed for diagnostic applications. Cells such as primary lamb kidney cells have been shown to be very sensitive to infection with FMDV, but the cells are short lived, laborious to prepare, and give variable results due to the use of different batches. Notwithstanding, the sensitivity of primary lamb kidney cells to infection by FMDV can be taken advantage of and developed into a cell line. Primary lamb kidneys cells’ lifespan can be increased through exogenous introduction of telomerase reverse transcriptase (TERT). In this study, transduction was conducted on primary lamb kidney cells at 80% confluency using recombinant lentivirus to over express TERT. The lentivirus construct included puromycin antibiotic resistance, thus, the cells were subjected to antibiotic selection, and surviving cells were expanded further. The over expression of TERT was confirmed by reverse transcription quantitative polymerase chain reaction (RT-qPCR) using a comparative ΔΔct method. The differences in the cycle thresholds for detection of TERT between primary cells and transduced cells showed increased expression of TERT in transduced cells. This increased expression however waned with increased cell passages. Encouragingly, the transduced cells were able to maintain similar morphology to primary cells over successive passages. The growth rate results also showed that primary cells’ growth regressed with progressive passaging as that of transduced cells increased. Analysis of TCID50/mL results for SAT and O serotypes of FMDV demonstrated no significant differences between primary and transduced cells. Virus isolation of SAT and O serotypes of FMDV gave similar time intervals to reach 100% cytopathic effect. These results demonstrate promise that the transduced cells could be used as a possible replacement for primary lamb kidney cells. Future work however must determine the extent of passaging that can be undertaken on transduced cells before there are notable changes to properties of the transduced cells. The transduced cells would also need to be validated using a broader range of virus serotypes and strain.Item type:Item, Antioxidant and antiproliferative activities of Azanza garckeana and Portulaca oleracea on HeLa cells(Botswana International University of Science and Technology (BIUST), 2024-07) Mosotho, Olorato Tefo; Gaobotse, Goabaone; Masisi, Kabo; Koobotse, MosesMortality and incidence rates of cervical cancer are on the rise in sub-Saharan Africa, particularly in Botswana. Current cervical cancer management strategies such as chemotherapy and radiation are not fully effective as they bring about major side effects and issues such as drug resistance. Indigenous plants of Botswana have been used in ethnomedicine to treat conditions related to cancer. Two known indigenous plants; Azanza garckeana and Portulaca oleracea have been found to contain phytochemicals which contribute to their antioxidant and anticancer properties. The aim of this study was to evaluate the antioxidant and anti-proliferation potential of extracts derived from Portulaca oleracea and Azanza garckeana. The study started with the derivation of extracts from the leaves, stems and roots of these two plants using six different solvents; methanol, ethanol, chloroform, distilled water, hexane and isopropanol in order of their polarity. Phytochemical screening of the extracts was then done to determine different secondary metabolites present. Total phenolic and flavonoid content of the different extracts was determined followed by the determination of the antioxidant activity of the extracts. GC-MS analysis was then conducted to detect different bioactive compounds present in the extracts. Anti-proliferation activity was determined by the cell viability assay; MTT to determine the IC50 values of the different extracts. It was found out that methanol effectively extracted antioxidant compounds. P. oleracea stem and leaf methanolic extract had high total phenolic and flavonoid content of 16.25 ± 0.48 TE mg/100 g, 16.39 ± 0.51 TE mg/100 g, 45.81 ± 1.02 QE mg/100 g and 41.70 ± 3.69 QE mg/100 g respectively whereas A. garckeana leaf methanolic had high total phenolic and flavonoid content of 14.00 ± 1.00 TE mg/100 g and 52.03 ± 2.44 QE mg/100 g respectively. xii | P a g e It was also found out that chloroform extracts had good antioxidant activity as the P. oleracea stem chloroform extracts gave a high antioxidant compounds concentration of 687.50 ± 0.88 TE Mmol/g and radical scavenging activity of 97.83 % ± 1.59 due to the presence of antioxidant compounds in the extracts. Methanolic and ethanolic extracts displayed anti-proliferative activity towards HeLa cells used in study and the extracts affected cell viability in a dose dependent fashion. A. garckeana leaf ethanolic extracts reduced cell viability by 92.00 % at 400 µg/ml extract concentration, P. oleracea leaf methanolic extracts reduced cell viability by 89.00 % at 400 µg/ml extract concentration, A. garckeana leaf methanolic extracts reduced cell viability by 91.00 % and P. oleracea stem methanolic extracts reduced cell viability by 69.00 % at 400 µg/ml extract concentration. The IC50 values of the extracts which measured the cytotoxicity degree of the extracts were found to be 329.31 µg/mL ± 54.69, 176.44 µg/mL ± 25.49 µg/mL, 190.47 µg/mL ± 46.10, 242.09 µg/mL ± 23.60, 22.78 µg/mL ± 10.37 for P. oleracea leaf methanolic extract, A. garckeana leaf methanolic extract, A. garckeana leaves ethanolic extract, P. oleracea stem methanolic extract and cisplatin (anti cancer drug) respectively. The anti-proliferative and antioxidant activity of the extracts of P. oleracea and A. garckeana was speculated to be due to the presence of antioxidant and anticancer compounds in the extract such as; 10-acetoxy-1,4a-dimethyl-9-oxo-1,2,3,4,4a,9-hexahydrophenanthrene-1-carboxylic acid, methyl ester ,octasiloxane,1,1,3,3,5,5,7,7,9,9,11,11,13,13,15,15-hexadecamethyl-, methoxy-2-[(3-methoxyphenyl)methylene]-,(E), cholestan-26-oic 3(2H)-benzofuranone, acid, 6- 3,7,12-trihydroxy-, (3α,5β,7α,12α)-, 3-O-Methyl-D-glucose, cyclononasiloxane and octadecamethyl.Item type:Item, Investigating the contribution of germline and somatic variants in the fatty acid metabolism pathway to prostate cancer aggressiveness in Southern African men(Botswana International University of Science and Technology (BIUST), 2025-04) Maendo, Umuna; Lebani, Kebaneilwe; Zhou, Nerve; Hayes, VanessaProstate cancer (PCa) is a significant global health issue, ranking second in cancer diagnoses and is the fifth leading cause of cancer-related deaths among men worldwide. Men of African ancestry have disproportionately higher PCa-associated mortality rates compared to other racial groups, underscoring a critical health disparity that necessitates further research. Despite this, most PCa genome studies focus predominantly on populations of European ancestry, excluding African men from tailored diagnosis and treatment options. PCa progression is facilitated by altering various signalling and metabolic pathways including fatty acid metabolism (FAM). To support their rapid growth, PCa cells meet their higher energy demands primarily through the oxidative degradation of fatty acids. While genes involved in FAM have been extensively studied, it is not well understood how both germline and somatic mutations within FAM influence patient predisposition and PCa progression. This project sought to address this gap by investigating the impact of germline and somatic variants within the FAM pathway on PCa aggressiveness in men of African and European ancestry. Data generated from the Southern African Prostate Cancer Study (SAPCS) was used in this study. A total of 743 Black South African men participating in the SAPCS underwent whole-exome genotyping. Common variants (MAF > 0.01) in 158 FAM genes were tested for association with PCa (451 cases vs 292 controls) and PCa aggressiveness (203 high risk, ISUP 3-5, vs 461 low risk, ISUP 1-2 and controls) using logistic regression. The significant variants were assessed for deleterious or damaging effects using SIFT and PolyPhen. Rare germline and somatic variant analysis were performed on whole-genome sequencing data from 158 treatment-naive individuals with PCa (Australia: n=53; South Africa: n=105). Rare variants (MAF < 0.05) were annotated with ANNOVAR, ClinVar, InterVar, and cancer genome interpreter as well as functional impact prediction tools (SIFT, PolyPhen, CADD, LOFTEE). The somatic variants were assessed for deleteriousness using SIFT and PolyPhen and tumour mutational burden (TMB) of the somatic variants was calculated. Nine common SNPs in FAM-related genes were significantly associated with PCa, including three deleterious variants in . Eight SNPs were linked to aggressive PCa, with deleterious variants in INMT and AOC3. These common SNPs had not been previously associated with PCa. Rare germline analysis revealed 15 pathogenic variants, four oncogenic variants and 15 variants with predicted loss of function. 80% of individuals with these rare variants had aggressive PCa. Somatic analysis showed African tumours had a higher (though not statistically significant) mean TMB than European tumours ( ). Several PDVs were exclusive to African-derived tumours, including mutations in FASN, ACADL, ACOT8, and ADH1C. Although limited by a small sample size, this study reveals novel germline and somatic variants in FAM pathway genes, highlighting their contribution to PCa risk and aggressiveness. Future research should prioritise African population inclusion, increased sample size, and include phenotypic studies in PCa genomics to improve diagnostics, treatment, and health equity.Item type:Item, An ontology-driven framework for animal traceability(Botswana International University of Science and Technology (BIUST), 2025-07) Mokgetse, Tshepiso Larona; Hlomani, Hlomani; Sigwele, TshiamoTraceability systems are crucial for regulatory compliance and consumer trust in food supply integrity and animal protection. Animal traceability systems ensure food safety and disease outbreak control. However, the current systems lack interoperability and are badly integrated across multiple stakeholders. The non-interoperable animal traceability systems pose challenges for farmers and the government in tracking stray animals, diseases, feeding, vaccinations, and livestock export regulations. The main cattle identification and traceability system being used in Botswana is the non-interoperable Botswana Livestock Identification and Traceability System. The six stakeholders concentrate on cow but lack a single system for interoperability, integration, standardization, and domain understanding among them. This research proposes the development of an interoperable and ontology-driven animal traceability framework to solve the gap of poor interoperability in current frameworks for animal traceability, with Botswana as a case study. Limitations of current traceability frameworks were identified, and the components for the design and development of the proposed interoperability framework were generated using the mixed method design methodology - qualitative and quantitative research. Involving robust data analysis. Quantitative identified system gaps and patterns in traceability practices though descriptive statistics. Qualitative research carried out a thematic analysis using NVivo tool on stakeholder interviews and documents, for in-depth understanding of socio-technical aspects of traceability systems. Together the designs reinforced the validity and reliability of the findings. The analysis identified cattle, farmers, system, and records as the key concepts. 39% of Botswana participants find BAITS ineffective, making it the main challenge of animal traceability. An interoperable ontology-driven animal traceability framework is developed. In addition, a new and novel animal traceability ontology was developed using the METHONTOLOGY ontology development methodology, covering knowledge acquisition, specification, conceptualization, formalization, and evaluation. The process ensures a transparent and systematic ontology development of the animal traceability domain to model the key traceability components. The ontology was evaluated for quality based on consistency, clarity, adaptability, and coverage. The Ontometric evaluation tool was used to evaluate revealing a high level of inheritance richness 0.692308, relationship richness 0.678571, and average depth 0.9, indicating detailed knowledge ontology, strong class connections, and seamless data integration and exchange. The evaluation of reference and candidate ontologies revealed 66.67% of animal traceability ontology is more interoperable than animal health ontology at 33.33%. Altogether, results indicate that the framework achieved interoperability and standardization, producing seamless integrated systems. Overall, this study contributes both as a template that is methodological for agricultural research in semantic systems. Also, providing a foundation for adaptable solutions to support worldwide supply chain compliance and strategies in animal health. The ontology-driven framework is designed as a guiding principle for system developers, a blueprint to implement semantically correct integrated traceability. The ontology is developed to contribute semantic interoperability and can be integrated with data infrastructures already existing.