Management of bacterial wilt disease of tomato by the root endophytic fungus piriformospora indica, rhizobacteria and bacterial endophytes

Abstract

The study entitled “Management of bacterial wilt disease of tomato by the root endophytic fungus Piriformospora indica, rhizobacteria and bacterial endophytes” was conducted during 2016-2018 at Department of Agricultural Microbiology, College of Agriculture, Vellayani with the objective of assessing the potential of root endophytic fungus Piriformospora indica, plant growth-promoting rhizobacteria and bacterial endophytes in suppressing bacterial wilt incidence in tomato. The bacterial wilt pathogen, Ralstonia solanacearum was isolated from infected tomato plants on Semi selective medium from South Africa (SMSA). Koch’s postulates were proved by artificial inoculation of tomato seedlings. A functional relationship between population and optical density of the broth culture was worked out and was used for determining the inoculum density for challenge inoculation in the biocontrol experiment. In vitro antagonistic interaction between the bioagents and the pathogen was worked out both by direct and indirect assays. In all direct assays which involved cross streak plating, agar plug diffusion technique, disc diffusion and spot on lawn method, it was found that Bacillus amyloliquefaciens VLY24, Bacillus velezensis PCSE10 and Streptomyces leeuwenhoekii KBT004 exhibited antagonism against Ralstonia solanacearum. Indirect antagonism was checked by agar well diffusion and disc diffusion methods using culture filtrate. Out of the eight bacterial bioagents tested, only two i.e., Bacillus velezensis PCSE10 and Bacillus amyloliquefaciensVLY24 had inhibitory effect on the pathogen. Dual culture plate assay on PDA has shown that three bacterial bioagents, Rhizobium radiobacter PCRE10, Bacillus megaterium NAT001 and Streptomyces leeuwenhoekii KBT004 were compatible with Piriformospora indica. However, when the compatible bacteria were co-cultured in a single fermentor system along with Piriformospora indica it was observed that Bacillus megaterium NAT001 failed to grow along with the fungal endophyte. Suppression of bacterial wilt incidence by the individual and combined application of bacterial bioagents and fungal endophytes were tested with the wilt susceptible tomato variety Naveen (Indo-American hybrid seeds Pvt. Ltd, Bengaluru) and the moderately tolerant KAU variety Vellayani Vijay. Bioagents were applied during the nursery production of seedlings and the 21 days old seedlings were transplanted to pots filled with unsterile garden soil. Challenge inoculation with the pathogen was done five days after transplanting by drenching the pots with 10 ml each of the bacterial suspension (107cfu/ml). The disease incidence was scored at weekly intervals for 21 days. When the bioagents were tested individually for the suppression of bacterial wilt incidence in the hybrid variety Naveen, maximum disease suppression was observed in plants treated with Rhizobium radiobacter PCRE10 (15 percent) after 21 days. The disease suppressive ability of Bacillus velezensis PCSE10 (25 percent) was also significantly superior to all other treatments. Selection of bacterial bioagents for combined application with Piriformospora indica was done based on compatibility with the fungal endophyte and inhibitory action against Ralstonia solanacearum. Combined application of the fungal endophyte and Bacillus amyloliquefaciens VLY24 suppressed the wilt incidence to the highest extent (40 percent). When bacterial wilt suppression in Vellayani Vijay was tested by the individual application of bacterial bioagents, maximum disease suppression was recorded in plants treated with Streptomyces leeuwenhoekii KBT004 (30 percent), when observed 21 days after challenge inoculation. The combination of Piriformospora indica with Rhizobium radiobacter PCRE10 (50 percent) and Bacillus velezensis PCSE10 (50 percent) showed significantly lower disease incidence compared to the rest of the treatments. In the plant growth promotion experiment done with the variety Vellayani Vijay in the nursery stage, maximum plant height was observed in plants treated with Rhizobium radiobacter PCRE10 (14.49cm). However, those plants treated with Bacillus pumilus VLY17 had the highest number of leaves per plant (4.58). Combined inoculation of Piriformospora indica and Bacillus amyloliquefaciens VLY24 resulted in improved shoot fresh weight (2.08 g per plant) which was on a par with the plants treated with Bacillus velezensis PCSE10 (1.95 g plant-1). However, shoot weight on dry weight basis was the maximum for plants treated with Bacillus velezensis PCSE10 (122.55 mg plant-1) which was at par with those treated with combination of Piriformospora indica and Bacillus amyloliquefaciens VLY24 (104.78 mg plant-1). The present study revealed that biological management of bacterial wilt in tomato could be a feasible strategy under controlled conditions. The same has to be validated under field conditions before making any recommendations.