Gammadelta T lymphocytes respond to different bacterial antigens and transformed cells. The antigenic molecules responsible for this activity have been studied extensively in antigenic preparations from Mycobacterium. We describe here the in vitro effect of Pseudomonas aeruginosa on gammadelta T lymphocytes and the properties of the implicated compounds. We found a preferential gammadelta T-cell expansion when we used heat-treated P. aeruginosa preparations and a dose-dependent inhibition of proliferation of peripheral blood mononuclear cells (PBMC) when non-heat-treated antigens were studied. This expansion corresponded to a Vgamma9-positive subpopulation. In contrast to alphabeta T lymphocytes, the highest stimulatory activity was restricted to very small cytosolic compounds. This activity was protease resistant and phosphatase sensitive and always dependent on interleukin (IL)-2 or alphabeta T-cell activation. We concluded that the antigenic molecules from P. aeruginosa that activated gammadelta T lymphocytes were small, non-peptidic, phosphorylated compounds, similar to those previously described from Mycobacterium.