Anti-microbial Effect of Cocoa Leaf Extracts on Botryodiplodia theobromae; Leading Causative Organism of Crown Rot Disease of Banana (Musa acuminata)

Abstract

An investigation was carried to assess in-vitro management of crown rot disease on Banana using extracts of cocoa leaves. The causative organism (Botryodiplodia theobromae) was sub cultured and the concentration determined through microbial count. Different cocoa leaves extracts were prepared using two different solvents. A conventional fungicide known as Mancozeb was used as a positive control and the nutrient broth only served as negative controls to help in assessment of the antimicrobial effect of the cocoa leave extracts. Phenolic Methanol Extracts (PME), Crude Cocoa Leaves Aqueous Extracts (CCAE) and Mancozeb were employed to prepare aqueous solutions through serial dilution with varied concentration. The nutrient broth (100 µL of double strength), 100 µL each of the plant extract concentrations, and 20 µL of inoculums were dispensed in a sterile micro-titre plate containing 96 wells. Incubation of the micro-titre plate was carried out at a temperature of 37°C for 1 day (24 hours). Susceptibility or otherwise of the microorganism (B. theobromae) from four sample stations was analyzed with 20 µL of a 5% solution of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide MTT. The presence of blue-black wells indicated the growth of B. theobromae. The experiment was conducted in triplicates for PME (refined), CCAE and Mancozeb. The minimum inhibition concentrations (MICs) of PME (refined) that inhibited the growth of B. theobromae collected from four different locations were 5mg/ml, 2.5 mg/ml, 10 mg/ml and 2.5 mg/ml respectively for B. theobromae (B1)(Offinso), B. theobromae (B2)(Effiduase), B. theobromae (B4)(Kejetia) and B. theobromae (B5)(Ejisu) which was lower compared to the MICs of CCAE which was 30 mg/ml for the four sample stations. Comparatively, the MIC of the PME (5 mg/ml, 2.5 m g/ml, 10 mg/ml and 2.5mg/ml) was almost equivalent to the MIC of the conventional fungicide (Mancozeb) (2.5 mg/ml) for all the four sample stations of B. theobromae.