Abstract: Background: Tobacco bacterial wilt caused by Ralstonia solanacearum is a devastating disease in Solanaceae crops and the pathogen is a notorious pathogen worldwide. Methodology: The phenotypic characterization of R. solanacearum under various osmolytes and pH environments were analyzed by using biolog Phenotype Microarray system (PMs). Using PM plates 9 and 10, 192 different assays were tested, including 96 different osmolytes and 96 pH environments. Results: Result presented that all four isolates of R. solanacearum exhibited similar phenotypic fingerprints. They had wide range adaptabilities in osmolytes with up to 4% sodium chloride, up to 20% ethylene glycol, up to 4% urea, up to 2% sodium lactate, up to 200 mM sodium phosphate (pH 7), up to 50 mM ammonium sulfate (pH 8), up to 100 mM sodium nitrate and up to 80 mM sodium nitrite. The bacterium did not grow at other osmolytes, including sodium chloride ranging from 5.5-10%, sodium formate ranging from 3-6%, urea ranging from 5-7%, sodium lactate ranging 7-12% and sodium benzoate (pH 5.2) ranging from 100-200 mM. They also exhibited active metabolism under pH values between 5 and 10, with an optimal pH value of around 6. The R. solanacearum showed active deaminase activity, while no decarboxylase activity in the presence of various amino acids. Conclusion: This study increased understanding on the bacterium, especially in survivability of the bacterium in the environment and would provide valuable profiles in developing practical ideas and methods for the disease control.