Evaluation of bacillus and trichoderma species for Biological control of bacterial wilt caused by ralstonia solanacearum in tomato

Abstract

Tomato (Lycopersicum esculentum) has been rated second most important vegetable crop in Kenya. Bacterial wilt caused by Ralstonia solanacearum, is a major biotic constraint to tomato productivity with yield losses of up to 64%. The available management strategies such as cultural practices and use of chemicals are limited in effectiveness. This study therefore focused on biological control agents (BCAs) in the management of Ralstonia solanacearum. Bacillus and Trichoderma isolates that are antagonistic to Ralstonia solanacearum and other important bacteria pathogens were screened and identified in vitro. The study also evaluated the effect of the Bacillus and Trichoderma isolates in management of bacterial wilt disease under field conditions. Twenty-eight Trichoderma species isolated from local soils at Kabete and 19 Bacillus isolates retrieved from earlier screened isolates maintained at the Plant Science and Crop Protection Department, University of Nairobi were used as the antagonists against three pathogens. Paper disc method was used to test the antagonistic activity of the isolates against Ralstonia solanacearum, Xanthomonas campestris pv campestris and Pseudomonas spp in vitro. The experiment was conducted in a complete randomized design, with three replicates. Antagonistic activity was assessed by measuring the radius of zone of inhibition (ZOI) of the pathogen due to the antagonist. Field experiments were conducted in a randomized complete block design at Kabete and Mwea sites in Kenya. The treatments included; 3 Trichoderma isolates (T1, T2 and T4), 2 Bacillus isolates (CB64 and CA7), a mixture of T1, T2 and T4, chemical standard and distilled water as control. Trichoderma and Bacillus isolates were grown on sterilized sorghum grain and cow manure carriers, respectively. Antagonist’s inoculation was carried out by dipping tomato plants for 30 minutes in each treatment suspension. Each treatment was applied at a rate of 150ml/plant hole and this was repeated after 35 days. Soils were sampled prior to transplanting, 60 days and 112 days after transplanting for quantification of R. solanacearum population and at 126 days for determining the total microbial count in the soil. Bacterial wilt incidence was assessed every week by counting the number of wilted plants in each plot. Yield parameters were assessed at physiological maturity. In vitro studies showed that 10 Bacillus and 11 Trichoderma isolates had varied antagonistic activity against all the pathogens tested. Trichoderma isolate T1 was the most effective in inhibiting the growth of Ralstonia solanacearum with a mean ZOI measuring 13.5mm while Bacillus isolates CB64 was the best antagonist with a mean ZOI measuring 4.3mm. Trichoderma isolate T28 and Bacillus isolate CA5 showed the highest ZOI of 15.2mm and 6.6mm, respectively against Xanthomonas campestris. The antagonists screened gave lower activity against Pseudomonas sp. compared to other pathogens. Trichoderma isolate T28 showed the highest ZOI of 9.3mm and Bacillus isolate CB14 and CB22 gave similar ZOI of 5.3mm. Isolates of Trichoderma showed better activity by more than 56.67% compared to isolates of Bacillus. All the treatments evaluated under field conditions significantly reduced bacterial wilt incidence and severity at P≤ 0.05 than the control at Kabete and Mwea sites. Trichoderma isolate T1 followed by Bacillus isolate CB64 were the best in reducing the disease incidence by more than 61.66% and 53%, respectively at both sites. Treatment CB64 and T1 had the highest reduction of R. solanacearum population in the soil by 93.17% and 92.07%, respectively. However, control had a pathogen increase of 20.40%. The total microbial count was highest in Bacillus treated plots in both sites. Isolate CB64 had the highest count of 1.32×105 CFU/ml at Kabete and 1.21×105 CFU/ml at Mwea site. CB64 and T1 performed significantly better compared to the standard, while the mixture of isolates T1, T2 and T4 performed poorest in all parameters. The treatments also increased the yield of tomato. Results from this study showed that Trichoderma and Bacillus isolates are effective biological control agents for use in management of bacterial wilt. Keywords: Tomato, bacterial wilt, Ralstonia solanacearum, Trichoderma, Bacillus