EXTRACTION OF CHITINASE ENZYME FROM CHITINOLYTIC FUNGI AND EVALUATION OF ITS ROLE IN PLANT DEFENSE AGAINST PHYTOPATHOGENIC FUNGI

Abstract

Phytopathogenic fungi result in a significant loss in agricultural production. The use of bio fungicides to manage these fungal pathogens has become a more appealing option than using conventional synthetic fungicides since they are relatively environmentally safe. Chitinolytic enzymes are environmentally friendly antifungal agents used to control fungal phytopathogens. Therefore, this study was conducted to extract chitinase enzyme from fungi and determine the fungicidal effect of the crude extracts against Aspergillus niger and Fusarium oxysporum. The chitinolytic fungi were isolated from soil using the dilution plate method and the best chitnolytic fungi with the highest colloidal chitin degrading potentials were selected by plate screening method and identified based on cultural and morphological features. The selected isolates were used in chitinase extraction. Solid state fermentation was used for chitinase production using colloidal chitin. The growth inhibitory effects of the crude chitinase were evaluated in vitro against Aspergillus niger and Fusarium oxysporum by applying different concentrations of crude chitinase using the agar dilution method. The experiment was done in a completely randomized design with three replications. In this study 8 morphologically different fungal isolates were identified and screened on colloidal chitin agar media. Based on the results of the chitinolytic index, CPF5 (0.97), CPF3 (0.89) and CPF6 (0.63) isolates were selected and identified as Trichoderma sp, Penicillium sp and Rhizopus sp, respectively. The highest chitinase activity was observed in Trichoderma sp (33.67U/ml) followed by Penicillium sp (26.76U/ml) and Rhizopus sp (14.97U/ml). Crude chitinase of the isolates significantly (p≤0.05) inhibited the mycelial growth of A. niger and F. oxysporum. Crude chitinase of all isolates exhibited greater inhibition at high concentrations. Therefore, it can be concluded that crude chitinase extracted from these fungal isolates effectively inhibited mycelial growth of the test pathogens. The effective mycelial growth inhibitory activity of crude chitinase extracts suggests that chitinase enzyme need to be produced in large quantity and applied in the field to manage phytopathogenic fungi.