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Applied and Environmental Microbiology, February 2007, p. 890-896, Vol. 73, No. 3
0099-2240/07/$08.00+0     doi:10.1128/AEM.01395-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

High-Throughput and Quantitative Procedure for Determining Sources of Escherichia coli in Waterways by Using Host-Specific DNA Marker Genes{triangledown}

Tao Yan,1 Matthew J. Hamilton,1,2 and Michael J. Sadowsky1,2,3*

BioTechnology Institute,1 Department of Microbiology,2 Department of Soil, Water, and Climate, University of Minnesota, St. Paul, Minnesota 551083

Received 16 June 2006/ Accepted 1 December 2006

Escherichia coli is currently used as an indicator of fecal pollution and to assess water quality. While several genotypic techniques have been used to determine potential sources of fecal bacteria impacting waterways and beaches, they do not allow for the rapid analysis of a large number of samples in a relatively short period of time. Here we report that gene probes identified by Hamilton and colleagues (M. J. Hamilton, T. Yan, and M. J. Sadowsky, Appl. Environ. Microbiol. 72:4012-4019, 2006) were useful for the development of a high-throughput and quantitative macroarray hybridization system to determine numbers of E. coli bacteria originating from geese/ducks. The procedure we developed, using a QBot robot for picking and arraying of colonies, allowed us to simultaneously analyze up to 20,736 E. coli colonies from water samples, with minimal time and human input. Statistically significant results were obtained by analyzing 700 E. coli colonies per water sample, allowing for the analysis of approximately 30 sites per macroarray. Macroarray hybridization studies done on E. coli collected from water samples obtained from two urban Minnesota lakes and one rural South Carolina lake indicated that geese/ducks contributed up to 51% of the fecal bacteria in the urban lake water samples, and the level was below the detection limit in the rural lake water sample. This technique, coupled with the use of other host source-specific gene probes, holds great promise as a new quantitative microbial source tracking tool to rapidly determine the origins of E. coli in waterways and on beaches.

* Corresponding author. Mailing address: University of Minnesota, Department of Soil, Water, and Climate, 1991 Upper Buford Circle, 439 Borlaug Hall, St. Paul, MN 55108. Phone: (612) 624-2706. Fax: (612) 625-2208. E-mail: sadowsky{at}umn.edu.

{triangledown} Published ahead of print on 8 December 2006.

Applied and Environmental Microbiology, February 2007, p. 890-896, Vol. 73, No. 3
0099-2240/07/$08.00+0     doi:10.1128/AEM.01395-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.





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