Use of a prou-gfp transcriptional fusion to quantify water stress on the leaf surface.
Catherine A. Axtell and Gwyn A. Beattie, Iowa State University, Ames, Iowa

Although water availability is thought to be a major factor influencing bacterial population dynamics on leaves, the quantitative impact of limited water availability on bacterial growth and survival has not been characterized. We introduced into the common leaf inhabitant Pantoea agglomerans a green fluorescent protein-based transcriptional fusion that was induced in a dose-dependent manner by both osmotic stress and matric (desiccation) stress. In this study, we evaluated the effectiveness of this P. agglomeransprou-gfp biosensor for quantifying bacterial water stress on leaves. After introducing cells onto bean plants, we recovered the cells after various periods of drying and determined using flow cytometry that the mean fluorescence of the recovered cells increased significantly after only 10 minutes of drying. After 30 minutes of drying, the average cell exhibited a 1.4-fold increase in fluorescence, and thus was exposed to a water potential equivalent to that imposed by 27mM NaCl. A small subpopulation of cells (19%) exhibited a much larger increase (equivalent to 127mm NaCl exposure). Based on our studies with surface-grown cells, sudden exposure to 27 or 127mm NaCl does not impact cell survival but does increase the lag time before growth by at least 20% (27mM) or 100% (127mM) relative to in the absence of water stress. These studies demonstrated that the P. agglomerans biosensor functions at ecophysiologically relevant levels of water stress, and that under the conditions tested, P. agglomerans were exposed to sufficiently high levels of water stress on leaves to affect their growth dynamics but not their survival.