Bacterial Source Tracking in Calleguas Creek Watershed

B.J. KILDARE; D. THOMPSON; S. TIWARI; V. RAJAL; S. WUERTZ

San Francisco, USA

Congreso; Joint Meeting of the 3 Divisions of the International Union of Microbiological Societies (IUMs); 2005

BackgroundSurface water quality is strongly influenced by increasing anthropogenic activities, as environmental waters receive a diversity of point and non-point source pollution. Specifically, microbial contamination of surface waters in the Calleguas Creek watershed has become a major concern, as many reaches within the watershed appear in the 2002 Clean Water Act Section 303(d) as impaired due to fecal coliform bacteria. Current efforts focus on the development of a Bacteria Total Maximum Daily Load (TMDL) to address this issue. For the protection of human and ecosystem health it is important to determine the abundance and diversity of human pathogens in these waters, as well as to identify the sources of the fecal contamination.MethodsThe study comprises a 12-month sampling and analysis plan for the Calleguas Creek watershed. In short, 100 L of water are collected monthly at each of 7 sites representing the receiving waters of the watershed. Hollow fiber ultrafiltration is used to reduce each water sample to approximately 100 ml. Performance of the concentration process is monitored for each sample with bacteriophage PP7 and E. coli spikes. Nucleic acids are extracted from the concentrated sample, and pathogen (adenovirus and enterovirus) enumeration is performed by quantitative polymerase chain reaction (qPCR). Additionally, a method of bacterial source tracking using Bacteroidales will be performed. Total Bacteroidales concentrations are determined using qPCR, and the relative contributions of human and non-human species-specific Bacteroides strains are approximated by conventional PCR coupled with a replicate limiting dilution method.ResultsThe combination of large volume ultrafiltration and molecular analysis led to highly reproducible environmental analysis of the watershed. Traces of Bacteroidales were found in every sample and both adenovirus and enterovirus qPCR assays successfully determined the presence of viruses. Detection limits varied from sample to sample and were strongly influenced by the extent of enzyme inhibition.ConclusionsThe project aims to provide data necessary to assist in the development of a new TMDL for the Calleguas Creek watershed. The results will help direct mitigation efforts to improve microbial water quality within the watershed and its receiving waters.