Evaluating human fecal contamination sources in Kranji Reservoir Catchment, Singapore

Singapore government through its Public Utilities Board is interested in opening Kranji Reservoir to recreational use. However, water courses within the Kranji Reservoir catchment contain human fecal indicator bacteria above recreational water quality criteria; their sources and distribution under dry and wet weather are also unknown. The goal of this study was to evaluate the distribution of E. coli under dry and wet weather, to determine the sources of the human fecal contamination, and to validate the use of human-specific 16S rRNA Bacteroides marker for human fecal source tracking in Singapore and tropical regions. Environmental water and DNA water samples (332) collected in the Kranji catchment in January and July 2009, and January 2010 were analyzed for E. coli using Hach m-ColiBlue24@ and IDEXX Colilert Quanti-Tray*/2000. Touchdown PCR and Nested-PCR HF183F assays were used to assess the absence or presence of the HF marker in Kranji catchment. Selected positive HF marker samples were sequenced and mapped using a phylogenetic tree to confirm their similarity in base order to the human factor identified in the temperate climate. The indicator bacteria (E. coli) results showed consistently high E. coli concentrations (geometric mean 3240 CFU/100 ml) in dry and wet weather in residential, horticultural and animal farming areas. The DNA analysis results showed that 94% of the 34 environmental DNA water samples collected in residential, horticultural and animal farming areas were positive to the HF marker. Generally, 74% and 94% of DNA samples respectively collected in dry and wet weather in the Kranji catchment were positive. The sequence and phylogenetic tree analysis confirmed that the HF marker identified was similar to the HF marker identified in temperate climates. Based on the results we conclude that human fecal contamination sources are widespread in the animal farming, horticultural and residential areas of Kranji catchment. The HF marker analysis validated its applicability as 16S rRNA gene of human-specific Bacteroides for human fecal source tracking in Singapore and elsewhere in tropical climates. Thesis Supervisor: Peter Shanahan Title: Senior Lecturer of Civil and Environmental Engineering Thesis Supervisor: Janelle Thompson Title: Assistant Professor of Civil and Environmental Engineering