Department of Biological Sciences

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    EVALUATION OF EFFECTS OF Trichoderma asperellum STRAINS IN BIOCONTROL OF CHARCOAL-ROT CAUSING Macrophomina phaseolina (Tassi) Goidanich IN COWPEA
    (Covenant University Ota, 2025-09) OYESOLA, Olushola Luke; Covenant University Thesis
    Macrophomina phaseolina, a soil-borne fungal pathogen, is responsible for loss in cowpea at both seedling and adult stages. Synthetic fungicides, although proven effective in M. phaseolina control, lead to severe cowpea poisoning and pose significant health risks to humans and animals. Trichoderma are efficient bioagents for managing plant pathogens, representing a promising strategy for managing M. phaseolina. This study evaluated the antifungal efficacy of some Trichoderma asperellum strains against M. phaseolina. The Trichoderma isolates obtained from soil were macroscopically, microscopically and genotypically identified based on the Internal Transcribed Spacer genes. Bioactive compounds were isolated from selected Trichoderma species and evaluated using Gas Chromatography-Mass Spectrometry (GC-MS). The inhibitory potential of the suspensions of selected Trichoderma species against M. phaseolina was tested singly in vitro and in combination in vivo. The in vivo experiments were carried out in the screenhouse across pre-planting, post-germination and seed treatment for eighty days, and plant growth indicators such as plant height, stem girth and leaf number were assessed at ten-day intervals. Disease incidence and severity were also determined following standard methods. After the termination of the in vivo experiment, yield assessment was carried out to determine the cowpea pod number, pod weight, and seed number, as well as the fresh and dry weights of leaves, stems and roots. A histopathological assessment was carried out on the lower stems of the cowpea plants to determine the effect of M. phaseolina on the cowpea tissues. Phenotypic, microscopic and genotypic characterisation identified all the Trichoderma isolates as Trichoderma asperellum. Among the Trichoderma species isolated and tested, T. asperellum (Tric13), T. asperellum (Tric4) and T. asperellum (Tric12) exhibited significant inhibitory potential (p<0.05) against M. phaseolina in vitro, with inhibition values of 82.51%, 82.41%, and 81.95%, respectively. The production of volatile organic compounds varied among the Trichoderma species. Specifically, Tric13 yielded terpenoids, ketones, sesquiterpenes, cycloalkanes, and alcohols. In contrast, Tric4 produced fatty acids, aldehydes, alkanes, terpenoids, and aromatic and bicyclic compounds, while Tric12 generated carboxylic acids, terpenoids, phenolics, bicyclic and aromatic compounds. The in vivo results showed that Trt7 (plant height = 49.8000 cm; stem girth = 0.53625 cm; leaf number = 21.028), followed by Trt3 (plant height = 49.8250 cm; stem girth = 0.49986 cm; leaf number = 18.611), recorded higher cowpea biomass across the Trichoderma formulations than the negative control. Zero disease incidence was observed in Trt3 and Trt7 (0%) upon treatment with the Trichoderma formulations. Also, zero disease severity was noted in Trt3 and Trt7 (0%), compared to the negative controls, which displayed 100% incidence and severity. The cowpea yield assessment showed that the highest pod weight (11.3 g) and pod number (8) were recorded in Trt7 (pre-planting), and seed number (32) was recorded in Trt5 (seed treatment). The histopathological assessment of the cowpea showed the efficacy of the treatments in inhibiting the growth of M. phaseolina, minimising its tissue entry, and xylem vessel occlusion. This study established the efficacy of Trichoderma in M. phaseolina biomanagement and the synergistic potential of different Trichoderma asperellum strains in its biocontrol.
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    MOLECULAR DOCKING, LIGAND QUALITY AND ANTIPLASMODIAL EVALUATION OF BENZAMIDE, COUMARIN AND BENZODIAZEPINE ANALOGS
    (Covenant University Ota, 2025-04) ADEBAYO GLORY PIPELOLUWA; Covenant University Thesis
    Malaria chemotherapy is an essential strategy for malaria elimination but resistance has challenged existing antimalarials, including frontline artemisinin combination therapy (ACT); hence, new antimalarial drugs must be discovered and developed. This study investigated the antiplasmodial efficacy and cytotoxicity through in vitro models while also testing the antiplasmodial efficacy, and the in vivo acute toxicity of benzamide, coumarin and benzodiazepine analogss. This study also evaluated the ligand quality of the molecules and their possible Plasmodium falciparum protein targets. Three molecules, 4- amino-N-hydroxybenzamide (AHB), ethyl 2-oxo-2H-chromene-3-carboxylate (ECC), and 2,2,4-trimethyl-2-3-dihyro-1H-benzo[b][1,4] diazepine (BDZ) were screened for their in vitro antiplasmodial activities tested against P. falciparum 3D7 standard strain using the SYBR Green Dye I measuring IC50 and their cytotoxicities against MCF-7 breast cancer cells using the [3(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] (MTT) assay. Their antiplasmodial efficacies were determined using Peter’s 4-day suppressive test against Plasmodium berghei in M. musculus while acute toxicities were investigated in the Mus musculus (mice). Ligand qualities were determined using ligand efficiency metrics, and molecular docking was conducted to determine the ligand interactions between ECC and the following enzymatic proteins, P. falciparum dihydroorotate dehydrogenase (PfDHOH) and P. falciparum purine nucleoside phosphorylase (PfPNP); and the molecular interaction between BDZ and PfDXR - Plasmodium falciparum 1-deoxy-D-xylulose-5-phosphate reductoisomerase, P. falciparum falcipain-2 and P. falciparum plasmepsin X (PfPMX). AHB showed no cytotoxicity against MCF-7 at (CC50) = 277.7 μM, while ECC showed inhibition with CC50= 3.930 μM, and BDZ showed no cytotoxicity CC50= 7373 μM. The in vitro antiplasmodial activity showed potency at (AHB)IC50 = 0.0020 ± 0.008 μM, (ECC) IC50= 0.0010 ± 0.002 μM, (BDZ) IC50= 0.0036 ± 0.003 μM respectively. BDZ showed the highest selectivity index at > 200,000, suggesting that it exhibited the best safety/efficacy among the three compounds. AHB displayed LD50 = >5000 mg/kg while ECC and BDZ displayed LD50 = 3162.28 mg/kg. Histopathological examinations showed non-toxicity by the three analogs on the liver and kidney of M. musculus. The percentage suppression of AHB (80.53 ± 3.26 %) at 400 mg/kg, was comparable to the standard chloroquine (81.71 ± 1.82 %) at 100 mg/kg where the mean survival time for both exceeded 30 days. ECC and BDZ showed excellent efficacies (70.98 ± 20.89 % and 83.66 ± 11.67 %) at 200 mg/kg, comparable to chloroquine 80.97 ± 5.82 %. The chemosuppression values for AHB and BDZ were significant at P value < 0.05. The ligand quality of ECC and BDZ displayed good Ligand Efficiency compared to chloroquine and artemisinin and higher enzyme affinities, and ligand efficiency dependent lipophilicity than the standard drugs. ECC and BDZ displayed good characteristics. The docking studies displayed strong hydrophobic interactions between ECC, PfDHODH, and PfPNP, suggesting good potency. BDZ’s binding with PfDXR, Pffalcipain-2, and PfPMX also displayed potency derived from hydrophobic and hydrogen interactions. Conclusively, this study showed AHB, ECC and BDZ were non-toxic to mammalian cells rodents’ liver and kidneys. These molecules exhibited good antiplasmodial inhibitory potential against both P. falciparum in vitro and P. berghei in vivo. ECC and BDZ displayed high ligand efficiency and strong molecular interactions with their protein targets. Therefore, all three analogs can be moved for further optimization in drug development.