Assessment of activated sludge viability with flow cytometry

The aim of the study was to evaluate the applicability of fluorescent dyes and multiparameter flow cytometry for the rapid and direct viability/activity assessment of activated sludge samples taken from wastewater treatment plants. Viability and activity of the biomass were estimated respectively through cellular membrane integrity, staining with SYBR Green I and Propidium Iodide, and through fluorogenic dyes capable of detecting enzymatic activity, as FDA and BCECF-AM. A procedure has been developed to disaggregate sludge flocs before dyes staining and cytometric analysis. The developed procedure allows a high recovery of bacteria with good accuracy and repeatability, and minimize the damage of the cells suspension obtained from the disaggregation of the flocs. These measurements were applied to estimate the two main parameters required to define the biological activated sludge process: the endogenous decay rate and the specific growth rate in exponential phase with high F/M ratio. Oxygen utilization rate measurements (OUR) were conducted to conventionally monitor the activity of the biomass. The preliminary data are encouraging and support the possibility to investigate bacteria dynamics on wastewater treatment plants.