Autoradiographic methods have been developed to detect metabolic activity of viable but nonculturable cells of Helicobacter pylori in water. Four strains of H. pylori were studied by using microcosms containing suspensions of 72-h cultures in water. The suspensions of aged, nonculturable cells of H. pylori were incubated with ((-3)H)thymidine for 24 to 72 h, after which the cell suspensions were exposed to Kodak NTB2 emulsion for 3 to 28 days. Each sample was processed with three separate controls to rule out false-positive reactions. The organism remains viable and culturable under these conditions for up to 48 h and, in some cases, 20 to 30 days, depending on physical conditions of the environment. The authors found that temperature was a significant (P < or = 0.01) environmental factor associated with the viability of H. pylori cells in water. Autoradiographs of tritium-labeled cells of H. pylori revealed aggregations of silver grains associated with uptake by H. pylori of radiolabelled substrate. Findings based on the autoradiographic approach give strong evidence supporting the hypothesis that there is a waterborne route of infection for H. pylori. The possibility that H. pylori may persist in water in a metabolically active stage but not actively growing and dividing is intriguing and relevant to public health concerns. (Copyright (c) 1993, American Society for Microbiology.)