Determination of in situ bacterial growth rates in aquifers and aquifer sediments

Laboratory and field-scale studies with stained cells were performed to monitor cell growth in groundwater systems. During cell division, the fluorescence intensity of the protein stain 5-(and 6-)-carboxyfluorescein diacetate succinimidyl ester (CFDA/SE) for each cell is halved, and the intensity can be tracked with a flow cytometer. Two strains of bacteria, Comamonas sp. strain DA001 and Acidovorax sp. strain OY-107, both isolated from a shallow aquifer, were utilized in this study. The change in the average generation or the average fluorescence intensity of the CFDA/SE-stained cells could be used to obtain estimates of doubling times. In microcosm experiments, the CFDA/SE-based doubling times were similar to the values calculated by total cell counting and were independent of cell concentration. Intact and repacked sediment core experiments with the same bacteria indicated that changes in groundwater chemistry were just as important as growth rates in determining planktonic cell concentrations. The growth rates within the sediment cores were similar to those calculated in microcosm experiments, and preferential transport of the daughter cells was not observed. The experiments indicated that the growth rates could be determined in systems with cell losses due to other phenomena, such as attachment to sediment or predation. Application of this growth rate estimation method to data from a field-scale bacterial transport experiment indicated that the doubling time was approximately 15 days, which is the first known direct determination of an in situ growth rate for bacteria in an aquifer.