Aim To determine whether expression of a cyanobacterial flavodoxin in soil bacteria of agronomic interest confers protection against the widely used herbicides paraquat and atrazine. Methods and Results The model bacterium Escherichia coli, the symbiotic nitrogen-fixing bacterium Ensifer meliloti and the plant growth-promoting rhizobacterium Pseudomonas fluorescens Aur6 were transformed with expression vectors containing the flavodoxin gene of Anabaena variabilis. Expression of the cyanobacterial protein was confirmed by Western blot. Bacterial tolerance to oxidative stress was tested in solid medium supplemented with hydrogen peroxide, paraquat or atrazine. In all three bacterial strains, flavodoxin expression enhanced tolerance to the oxidative stress provoked by hydrogen peroxide and by the reactive oxygen species-inducing herbicides, witnessed by the enhanced survival of the transformed bacteria in the presence of these oxidizing agents. Conclusions Flavodoxin overexpression in beneficial soil bacteria confers tolerance to oxidative stress and improves their survival in the presence of the herbicides paraquat and atrazine. Flavodoxin could be considered as a general antioxidant resource to face oxidative challenges in different micro-organisms. Significance and Impact of the study The use of plant growth-promoting rhizobacteria or nitrogen-fixing bacteria with enhanced tolerance to oxidative stress in contaminated soils is of significant agronomic interest. The enhanced tolerance of flavodoxin-expressing bacteria to atrazine and paraquat points to potential applications in herbicide-treated soils.