Evaluation of Trichoderma spp., Pseudomonas fluorescens and Bacillus subtilis for biological control of Ralstonia wilt of tomato [version 2; referees: 2 approved]

Abstract

Background: Ralstonia spp. is a major pathogenic microbe for tomato, which invades the roots of diverse plant hosts and colonizes xylem vessels causing wilt, especially in tropical, subtropical and warm-temperate regions. Ralstonia spp. produces several virulence factors helping it to invade the plant’s natural defense mechanism. Native isolates of Trichoderma spp., Pseudomonas fluorescens and Bacillus subtilis can be used as biocontrol agents to control the bacterial wilt and combined application of these beneficial microbes can give better results. Methods: Bacterial wilt infection in the field was identified by field experts and the infected plant part was used to isolate Ralstonia spp. in CPG media and was positively identified. Subsequently, the efficacy of the biocontrol agents was tested and documented using agar well diffusion technique and digital microscopy. 2ml of the microbial concentrate (109 cells/ml) was mixed in one liter of water and was applied in the plant root at the rate of 100 ml per plant as a treatment method. Results: It was observed that the isolated Trichoderma spp. AA2 and Pseudomonas fluorescens PFS were most potent in inhibiting the growth of Ralstonia spp., showing ZOI 20.67 mm and 22.33 mm, respectively. Digital microscopy showed distinct inhibitory effect on the growth and survival of Ralstonia spp. The results from the field data indicated that Trichoderma spp. and Pseudomonas fluorescens alone were able to prevent 92% and 96% of the infection and combination of both were more effective, preventing 97% of infection. Chemical control methods prevented 94% of infection. Bacillus subtilis could only prevent 84 % of the infection.   Conclusions: Antagonistic effect against Ralstonia spp. shown by native isolates of Trichoderma spp. and P. fluorescens manifested the promising potential as biocontrol agents. Combined application gave better results. Results shown by Bacillus subtilis were not significant