Aims: This study was designed to examine the breakdown of trehalose by rhizobia and to characterize the trehalose-degrading enzyme isolated from Rhizobium sp. NGR234. Methods and Results: Rhizobium sp. NGR234, Rhizobium fredii USDA257, R. phaseoli RCR3622, R. tropici CIAT899 and R. etli CE3 showed good growth in the presence of carbohydrate. Validamycin A did not prevent the growth of NGR234 on trehalose. The expression of a trehalose-degrading enzyme by NGR234 was intracellular and inducible by trehalose. The isolated enzyme digested other disaccharides, p-nitrophenyl-alpha-D-glucopyranoside and the substrate. The enzyme showed optimum activities at pH 7.0 and 30degreesC. Its pI was 4.75 and the V-max of the enzyme occurred at 35.7 mumol s(-1) mg(-1) protein with the K-m of 23 mmol when trehalose was hydrolysed. Conclusions: An enzyme capable of breaking down trehalose was produced. Some of the properties of the trehalose-degrading enzyme are similar to those isolated from other organisms but, this enzyme was validamycin resistant. These rhizobia like other trehalose-degrading microbes use trehalose by enzymatic catabolic action. Significance of Impact of the Study: Trehalose which accumulates during legume-rhizobia symbiosis is toxic to plants. Detoxification by trehalose-degrading enzymes is important for the progress of symbiosis.