Identification of groundwater microbes that decrease the concentration of the conservative tracer uranine

Fluorescent dyes are commonly used as conservative groundwater tracers to track the migration of water. Over- or underestimation of important parameters such as the water flow rate can occur if the concentration of a dye is changed by unexpected reactions. Because such errors may seriously affect the results of experiments, the reactions and processes that change fluorescent dye concentrations need to be understood. In this study, we focused on the widely used fluorescent dye uranine (UR) and aimed to identify microbes contributing to decreases in UR concentrations in groundwater. First, we identified the conditions (water temperature, pH, and salinity) under which significant decreases in UR concentrations occurred to show that the decrease in UR concentrations were caused by the effects of microbes in the groundwater. Next, we obtained information about the metabolism of organic matter by potential contributing microbes. These results were used to narrow down possible microbes that could decrease the UR concentration. Analysis of the microbial community in groundwater using 16S rRNA gene sequencing was then used to further identify contributing microbes. Finally, a verification experiment was conducted using a strain of one of the identified microbes (Parapontixanthobacter aurantiacus). Our results showed that conservation of the concentration of fluorescent dye solutions prepared with on-site groundwater was affected by several microbes with different metabolic characteristics, including P. aurantiacus. When fluorescent dye solutions prepared with on-site groundwater are used in field investigations or tracer tests, the pros and cons of using fluorescent dyes should be carefully evaluated because of the potential effects of microbes in the groundwater.