Beneficial endophytes for the management of brown Spot of rice by Bipolaris oryzae (breda de haan)

Abstract

The research study entitled “Beneficial endophytes for the management of brown spot of rice incited by Bipolaris oryzae (Breda de Haan)” was undertaken at College of Agriculture, Vellayani and Integrated Farming System Research Station (IFSRS), Karamana during 2023-2025 with the objective to develop an ecofriendly management practice for brown spot of rice incited by Bipolaris oryzae using beneficial endophytes. The three bacterial endophytes viz., Bacillus subtilis PAL-11, Bacillus altitudinis TSL-4 and Bacillus altitudinis KAM-11 as well as the fungal endophyte viz., Xylaria sp. KTD-2 maintained at IFSRS, Karamana were used for the study. Rice plants exhibiting brown leaf spot symptoms were collected from the fields of IFSRS, Karamana. Typical symptoms on leaves included oval shaped brown lesions with grey centre and yellow margin. Several such spots coalesced leading to drying of leaves. Minute brown spots appeared on grains, resulting in grain discolouration. B. oryzae was isolated and sub cultured on potato dextrose agar medium following single hyphal tip method. Mycelium was grey to black in colour with its growth appearing as zonations from centre to periphery. Conidiophores were multi-septate, brown in colour, bearing slightly curved and fusiform conidia, initially hyaline which later turned brown. Pathogenicity study was done by artificially inoculating leaves of healthy rice plants (var. Uma) with seven day old mycelial agar discs of B. oryzae. Cultural, morphological and molecular characterization confirmed the identity of the pathogen to be Bipolaris oryzae (Teleomorph: Cochliobolus miyabeanus) with 98.73 per cent homology. In vitro screening of bacterial and fungal endophytes against B. oryzae was undertaken by dual culture technique and disc diffusion assay. Xylaria sp. KTD-2 and B. subtilis PAL-11 recorded the highest inhibition (73.11 and 68.88 % respectively) of mycelial growth of the pathogen when tested by dual culture technique. B. subtilis PAL- 11 recorded the highest antifungal potential, with 68.32 per cent inhibition of mycelial growth and an inhibition zone of 0.51 mm, when tested by disc diffusion assay. Compatibility assay by cross streak assay demonstrated that the endophytic Bacillus isolates were compatible with each other. Compatibility study of endophytes with the 143 recommended fungicide, trifloxystrobin 25% + tebuconazole 50% (0.04%) revealed that all bacterial endophytes were compatible with the fungicide. However, the fungal antagonist, Xylaria sp. showed complete incompatibility and high fungicide sensitivity. Evaluation of seedling growth promotion by promising endophytes using roll paper towel method revealed that both Xylaria sp. KTD-2 and B. subtilis PAL-11 significantly enhanced seedling vigour and growth. Pot culture study to evaluate the antifungal potential of the two promising treatments selected from in vitro study against B. oryzae revealed that, seed treatment with Xylaria sp. KTD -2 followed by its foliar application @ 30, 45 and 60 days after transplanting (DAT) recorded the lowest disease incidence (46.65%) and disease severity (15.60%), with the highest grain yield (31.73 g plant -1). To further understand the interaction between Xylaria sp. KTD-2 and B. oryzae on rice leaves, a scanning electron microscopy analysis was conducted. The images revealed that the thin hyphae of Xylaria sp. KTD-2 growing into the mesophyll cells, the primary entry point of B. oryzae and subsequently invaded and disintegrated the vesicles produced by the pathogen. Biochemical assays were undertaken to assess the changes in the activities of peroxidase, polyphenol oxidase, phenylalanine ammonia lyase, catalase, superoxide dismutase at 0, 24, 48, and 72 h after foliar inoculation of the pathogen in the KTD-2 and PAL-11 treated plants in pot culture. The results revealed that, seed treatment with Xylaria sp. KTD-2 and B. subtilis PAL-11 activated defense mechanisms in rice plants against B. oryzae by significantly enhancing production of different antioxidant enzymes activity in rice leaves. Thus, the present study revealed that, the fungal endophyte, Xylaria sp. KTD-2 and the bacterial endophyte, Bacillus subtilis PAL-11, possessed strong potential for disease management and plant growth promotion in rice. Thus, these endophytes can serve as suitable alternatives or supplements to chemical fungicides in rice disease management programs. Thus, it can be concluded that, seed treatment with Xylaria sp. KTD-2 followed by its foliar application at 30, 45 and 60 DAT can reduce the incidence of brown spot disease of rice along with the additional benefit of plant growth promotion.