Evaluation of Bacillus spp. as biocontrol agents against chili leaf spot caused by Xanthomonas vesicatoria

Abstract

In this study, we investigated native soil antagonists and the exotic Bacillus thuringiensis (Bt) in their effectiveness against chili bacterial pathogens, specifically Pseudomonas syringae and Xanthomonas vesicatoria. Putative native soil antagonists were isolated through plating heat-shocked (60 °C) serial dilutions on nutrient agar plates. Among these isolates, two (SK, CM) were further characterized based on phenotypic and biochemical traits, revealing gram-positive and catalase-positive characteristics, while being negative for urease, oxidase, and arginine dihydrolase. To evaluate the efficacy of SK, CM, and exotic Bt against P. syringae and X. vesicatoria, a dual culture confrontational assay was conducted. SK, CM, and Bt were found to restrict the growth of X. vesicatoria, not P. syringae. Consequently, further tests were exclusively performed with X. vesicatoria. Chemical compatibility with copper sulfate, topsin M, and sodium benzoate was assessed using the poisoned food technique, revealing inhibitory effects on microbial growth except for topsin M. Antibiotic sensitivity testing (erythromycin, streptomycin, tetracycline, penicillin) of X. vesicatoria, SK, CM, and Bt was carried out using the disk diffusion method. Results indicated a minimum zone of inhibition (0 mm) against penicillin and a maximum (29 mm) against streptomycin. The efficacy of antagonists was also tested in planta using detached leaf and seedling inoculation methods. Pre-inoculating leaves of a susceptible chili cultivar with antagonists and subsequently re-inoculating with X. vesicatoria demonstrated that, except for the positive control, leaves remained asymptomatic after 7 days post-inoculation. A pot experiment involving soil drenching and foliar spray inoculation further confirmed that plants initially treated with antagonists exhibited resistance against subsequent pathogen application compared to the positive control. In conclusion, this pilot study revealed that native putative Bacillus spp. isolates (SK, CM) and commercial exotic Bt have the potential to counteract X. vesicatoria in both local and distant tissues.