Leptospira interrogans is the causative agent of leptospirosis, which is an emerging zoonotic disease. Resistance to stress conditions is largely uncharacterized in these bacteria. We therefore decided to analyze a clpB mutant that we obtained by random transposon mutagenesis. The mutant did not produce any of the two isoforms of ClpB. The clpB(-) exhibited growth defects at 30 degrees , 37 degrees C, and poor nutrient medium, and increased susceptibility to oxidative stress, whereas the genetically complemented strain was restored in ClpB expression and in vitro wild-type growth. We also showed that the clpB(-) is attenuated in virulence in the acute animal model of leptospirosis. Our findings demonstrate that ClpB is involved in the general stress response. The chaperone is also necessary, either directly or indirectly, for the virulence of the pathogen L. interrogans.