PRODUCTION OPTIMIZATION OF BACILLUS ISOLATES WITH A COST-EFFECTIVE PROCESS AS A BIOLOGICAL CONTROL AGENT

Project no. 20159 · Status: Kapanmış · 2019

Summary

Soil is a dynamic habitat and is not only a critical resource in agriculture and food security, but also ensures the continuity of the entire life process. Pathogenic microorganisms, which adversely affect plant health, are a major and chronic threat to sustainable agriculture worldwide. Agrochemicals used to increase productivity in agricultural production and to kill pathogens, pests and weeds have great harmful effects on the ecosystem. Due to current public concerns about the side effects of these chemicals, understanding the cooperation between plants and the microbial population in the rhizosphere has become important. Therefore, the need to develop biological agents that are accepted worldwide has arisen. The use of rhizobacteria that promote plant growth is a good alternative to solve this problem. They play an important role in increasing soil fertility, promoting plant growth and suppressing phytopathogen for the development of environmentally friendly sustainable agriculture.The main purpose of this thesis study is to determine the antifungal effects of native and original Bacillus isolates against phytopathogenic fungi and to optimize the production parameters and an economical production medium to obtain maximum viable cell count by using the most effective isolate. For this purpose, 1574 Bacillus isolates were investigated in vitro for their antifungal effects against phytopathogenic fungi such as Botrytis cinerea, Fusarium solani and Rhizoctonia solani. Percent growth inhibition values were calculated for 118 Bacillus isolates, which were determined to form inhibition zones against all three phytopathogens. It was determined that the maximum percent growth inhibition values of the examined isolates were 65.6% against B.cinerea, 42.2% against F.solani and 55.6% against R.solani. For antifungal lipopeptide extracts obtained from isolates whose hemolytic activities were also examined, these values were determined as 60.0% against B.cinerea, 36.4% against F.solani and 37.9% against R.solani.The specific enzyme activity values for protease, chitinase and chitosanase which obtained from the examined 77 Bacillus isolates were determined respectively between 9.7-6652 U/mg, 0.22-46.64 U/mg and 8, 02-22.17 U/mg. In addition, 11 of these isolates were determined to be HCN producers, while 70 of them were determined to be siderophore producers. It has been shown that the siderophore index value for siderophore producing isolates varies between 1.22 and 3.11. Among the isolates whose coagulase activity was also examined, two of them were found to be coagulase positive. By examining the biocontrol mechanisms, 19 isolates were selected among those with the highest antifungal effect which can ne used as a biological control agent and for these isolates species determination was made by molecular identification.Bacillus velezensis 4-Ka 22 strain, which was determined to be suitable for mass production and to produce high lipopeptide antibiotics, was selected for production optimization. For this isolate, the production medium containing molasses, soy flour, CaCl2 and glycerol, and production parameters such as temperature, shaker speed (rpm), pH and inoculation rate were optimized using conventional methods and RSM. The optimized production medium contains 0.1% molasses, 1% soy flour, 0.5% CaCl2 and 1% glycerol. The production temperature was determined as 30°C. In addition, as a result of the optimization performed with RSM for the production parameters, the shaker speed was determined as 195 rpm, pH 5.0 and the inoculation rate was 4.3%. As a result of the optimization, the number of viable cells obtained for the Bacillus velezensis 4-Ka-22 strain was increased by 13.4 times and the cost of the production media components was reduced by 467 times.

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