Because glutathione (GSH) is an important antioxidant and metal chelator, this study was undertaken to determine whether the role of this tripeptide in cadmium tolerance of Rhizobium leguminosarum is linked to its antioxidant properties. Two strains expressing different degrees of tolerance to cadmium (Cd) stress were used and the influence of the addition of extracellular GSH to the growth media was determined. Oxidative stress indexes and reactive oxygen species (ROS) scavenging enzymes were evaluated, as well as changes in GSH and oxidized GSH (GSSG) levels. Results confirmed that CdCl2 imposes oxidative stress in R. leguminosarum, which was characterized by an increase in GSSG formation and an induction of the activities of superoxide dismutase (SOD), GSH peroxidase (GPX), GSH reductase (GR) and catalase (CAT). Addition of reduced GSH to the growth media had a protective effect, particularly in the sensitive strain. Our findings show that increased tolerance in R. leguminosarum is not related to a higher efficiency in the oxygen scavengers per se, but to ratios of intracellular reduced GSH. Furthermore, we demonstrated that the supply of GSH to the sensitive strain enhanced survival, concluding that GSH plays a crucial role in Cd tolerance in this species. However, this role consists in preventing the oxidative stress to the cell, rather than reducing its effects. (c) 2006 Elsevier Inc. All rights reserved.