Identification and Tracking of the
Sources of Microbial Pollution in
Watersheds and Distribution Systems

Mansour Samadpour
Department of Environmental Health
School of Public Health and Community Medicine
University of Washington, Seattle, WA

The lack of appropriate methodology for tracing bacterial contamination in the environment is a major impediment identification and control of the sources of these pollutants and adversely affects the decision-making process in water quality management, and management of fisheries resources. Several methods for tracking genetically engineered microorganisms have been used, but their utility is limited to the detection of organisms carrying reporter genes or their products. Limited efforts to track sources of natural bacterial populations have been made, the approach used was based on quantitation of indicator organisms at various sites. These studies invariably have resulted in raising more questions rather than providing an answer. I have developed and tested a tracking system for identification of sources of microbial pollution. The methodology can be used to asses the impact and contribution of non-point sources and point-sources of fecal pollution (animals, septic tanks, farms, sewage treatment plants, etc.) on the fecal coliform level in watersheds and mapping the distribution, transportation, and movement of indicator organisms and microbial pathogens in watersheds and distribution systems.

The Ribo-Tracking method was used to determine the impact of the human source on the main source of water supply for the City of Seattle (Cedar River Watershed).

1268 water and environmental Escherichia coli isolates were collected over a two-year period and their ribotypes were compared to determine the origin of non-point source contamination of the Cedar River. A collection of E.coli isolates was established from Cedar River water (701 isolates), humans and septic tanks in the watershed (415 isolates), and from fecal samples of animals that reside in the watershed (12 isolates). Water samples were taken at the intake point for Seattle's drinking water supply and various sampling stations on the Cedar River. Samples were plated on selective differentiative media for isolation of fecal coliforms. A collection of E. coli isolates were established. Genomic DNA was extracted from each isolate, digested with EcoR1 restriction enzyme electrophoretically separated, and analyzed by Southern blot hybridization with radiolabeled cDNA probe for the ribosomal RNA operon. The autoradiograms were then assigned a ribotype pattern according to band grouping and size, and were entered into a database and compared to all other isolates which had been treated in the same manner. If a strain from a given source was found to have the exact same ribotype as isolates from the Cedar River water, the source was identified as the contributor of the polluting strain.

Results indicate that the fecal coliforms found in the Cedar River are not of human origin. While several matches were found between animal (avian, cat, coyote, deer, dog, and rodents) and water isolates, there were not matches between human and water isolates.

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