Assessing viability of Escherichia coli in mining-impacted surface waters

Multiple threats to surface water quality, including both fecal pollution and acid mine drainage (AMD) are frequently coincident in mining regions, such as parts of the Appalachian region of the U.S. While it has been established that AMD can have toxic effects to fecal bacteria in surface waters, there is also evidence that AMD may induce a viable but non-culturable (VBNC) state, leaving fecal bacteria undetectable by common standard methods. This study aims to better understand the occurrence of VBNC Escherichia coli in mining-impacted waters (MIW) through three objectives: (1) assess the relationship between MIWs and concentrations of culturable fecal bacteria, (2) compare standard methods for quantification of E. coli in MIW, and (3) investigate whether MIW can induce a VBNC state in E. coli. An analysis of historic data from the Deckers Creek and West Run Watersheds of West Virginia revealed a moderate correlation between pH and E. coli abundance determined via culture and enzyme-based methods. In samples collected from sites within the two watersheds impacted by historic mining activities, the E. coli uidA gene was measured via quantitative polymerase chain reaction (qPCR) and found to be significantly more abundant than E. coli concentrations measured via culture or enzyme substrate coliform test. However, this discrepancy between methods was greatest in samples with a low pH. Further, experiments in which an environmental strain of E. coli was dosed into samples of MIW diluted with saline at different concentrations confirmed that the culturability of E. coli significantly decreased with increasing MIW concentration, while the total number of intact cells, determined via "live/dead” staining and microscopy, remained high. This finding suggests that culture and enzyme methods may underestimate viable E. coli levels in AMD-impacted waters.