Influence of growth rate and starvation on fluorescent in situ hybridization of Rhodopseudomonas palustris

In situ hybridization with a fluorescently labeled 16S rRNA-targeted probe was examined using Rhodopseudomonas palustris as a model organism, which ha d been grown at different rates and under different conditions of growth an d starvation. The specific growth rate did not affect the percentage of hyb ridized cells in aerobically grown R. palustris cultures. However, signific ant changes in the percentage of hybridized cells occurred during extended periods of starvation. These changes were observed both in batch cultures g rown and starved aerobically in the dark, and in cultures grown phototrophi cally and starved anaerobically in the dark. Aerobic growth in batch cultur e and subsequent starvation resulted in a complete lack of detectable hybri dization after 20 days of starvation. In contrast, even after 30 days of st arvation, 50% of all cells were still detectable in cultures grown aerobica lly at growth rates < 0.06 h(-1) and then starved aerobically in the dark. The same was true for phototrophically grown cells that were starved anaero bically in the light. During starvation there was a clear, though non-linea r, positive correlation between the percentage of hybridized cells and the RNA content. In contrast, no direct correlation was observed between the nu mber of hybridized cells in a culture and the viability of this culture. Th us, in habitats with growing, non-growing, and starving bacteria, data on q uantitative detection of populations based on 16S rRNA-targeted probing sho uld be used with extreme caution as the detectability of the individual cel ls is strongly influenced by their physiological history and current physio logical state. (C) 2000 Federation of European Microbiological Societies. P ublished by Elsevier Science B.V. All rights reserved.