Simultaneous measurement of metabolic activity and membrane integrity in marine bacterioplankton determined by confocal laser-scanning microscopy

We simultaneously assessed the metabolic activity and viability of individual bacterioplankton cells in the coastal and open North Sea. Three different techniques were applied to determine cell features related to the physiological status of the cell. SYBR Green I was used to estimate the nucleic acid content of the cell. Propidium iodide (PI) stains cells with a compromised cell membrane, commonly interpreted as indicative of dead cells. Microautoradiography (MA) with radiolabeled glucose and leucine was applied to indicate metabolically active cells. The relative abundance of metabolically active cells determined by MA was usually <20% of the total abundance of bacteria. In contrast, the percentage of PI-positive cells in the total bacterial community was generally high (∼80 %). However, the overwhelming majority (97 %) of cells taking up glucose and leucine were also PI-positive. Apparently, the uptake of radiolabeled substrate is related to PI accumulation in cells, indicating that PI is not a reliable stain to indicate non-active or dead bacteria. We suggest that several methods should be combined to assess the physiological status of individual cells in natural bacterioplankton communities.