Use of Nitrogen-15-Enriched Escherichia coli as a Bacterial Tracer in Karst Aquifers

James W. Ward, John G. Warden, Ashley M. Bandy, Alan E. Fryar, Gail M. Brion, Stephen A. Macko, Christopher S. Romanek, Mark S. Coyne

Research output: Contribution to journalArticlepeer-review

4 Scopus citations


Karst aquifers are susceptible to contamination by microorganisms, but relatively few studies have used bacteria as tracers. We demonstrate the utility of Escherichia coli enriched in the stable isotope nitrogen-15 (15N) as a novel bacterial tracer. Nonpathogenic E. coli from two springs in central Kentucky were grown on 15N-enriched media. Survival of E. coli and persistence of the isotopic signal were assessed in two sets of laboratory experiments conducted with sterilized spring water in dark microcosms at 14 °C. First, isotopically labeled bacteria survived for 130 d at concentrations within one log unit of the average initial value, and there was no significant difference in δ15N values from Day 1 to Day 130. Second, water samples with E. coli were inoculated with either of two different species of protozoa (Tetrahymena pyriformis or Colpoda steinii). During 7 d, δ15N values increased in T. pyriformis while bacterial populations decreased. In a field test, following a 2.1-cm rainfall, 15N-labeled E. coli, solutes (rhodamine WT dye and bromide), and latex microspheres were injected into a sinkhole approximately 530 m upgradient of a spring. Breakthrough of all tracers coincided, but microspheres were remobilized by subsequent storms, unlike other tracers. Enriched E. coli exhibited more tailing than solute tracers during the initial storm-flow recession. These results indicate that 15N-enriched E. coli is a viable tracer of bacterial transport in karst aquifers, although predation may attenuate the isotopic signal in systems that are not rapidly flushed.

Original languageEnglish
Pages (from-to)830-839
Number of pages10
Issue number6
StatePublished - Nov 1 2016

Bibliographical note

Funding Information:
The authors acknowledge support from the University of Kentucky (UK) College of Agriculture SB-271 program, the U.S. Department of the Interior under U.S. Geological Survey Grant Number 06HQGR0087 to the UK Research Foundation, the Kentucky NSF-EPSCoR program, the UK Department of Earth and Environmental Sciences, and the Tracy Farmer Institute for Sustainability and the Environment. We thank the Kentucky Geological Survey, the UK Environmental Research and Training Laboratory staff, and the City of Versailles for access; Dr. Harry Rowe (Bureau of Economic Geology, University of Texas at Austin) for his suggestions during the project; and three anonymous reviewers for their substantive comments. The views and conclusions contained herein are those of the authors and do not necessarily represent the official policies, either expressed or implied, of the U.S. Government.

Publisher Copyright:
© 2016, National Ground Water Association.

ASJC Scopus subject areas

  • Water Science and Technology
  • Computers in Earth Sciences


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