Detecting Pathogens in Water by Ultrafiltration and Microarray AnalysisEPA Grant Number: R833004
Title: Detecting Pathogens in Water by Ultrafiltration and Microarray Analysis
Investigators: Lee, Anthea K. , DeLeon, Ricardo , Rochelle, Paul A.
Institution: Metropolitan Water District of Southern California
EPA Project Officer: Page, Angela
Project Period: July 21, 2006 through July 20, 2009 (Extended to July 20, 2011)
Project Amount: $594,106
RFA: Development and Evaluation of Innovative Approaches for the Quantitative Assessment of Pathogens in Drinking Water (2005) RFA Text | Recipients Lists
Research Category: Drinking Water , Water
This project addresses critical issues in rapid pathogen detection methods by advancing sample processing, concentration, and nucleic acid extraction techniques so that high throughput sample interrogation tools, like microarrays, can be used to their full potential.
The ultimate objective of this project is to develop an innovative approach for detecting multiple waterborne bacterial, protozoan, and viral pathogens utilizing large volume (100 – 1,000 L) ultrafiltration (UF) as a universal pathogen concentration technique, direct extraction of nucleic acids, whole sample genome amplification (WSGA), and hybridization to a multi-pathogen, water quality microarray.
Cryptosporidium parvum, Salmonella typhimurium, and human adenovirus 2 are used as model pathogens for UF method development, infectivity assays, and real-time quantitative PCR to assess the efficiency of DNA extraction and WSGA procedures. Microarrays will be used for various aspects of the project including: (1) capturing of specific target-pathogen sequences on an array-based solid phase substrate to improve amplification sensitivity; (2) assessing the efficiency of WSGA techniques using an Escherichia coli genomic array; (3) measuring host cell response to pathogens as a rapid and sensitive infectivity detection assay; and (4) developing a multi-pathogen, multi-target, water quality microarray. The final detection array will target organisms on the Environmental Protection Agency’s candidate contaminant list, other potential waterborne pathogens, and traditional microbial indicators.
We anticipate that these experiments will clearly establish that microarray technology can be useful for the water industry and provide direction for future methods development. The ability to detect pathogens using several different methods, including cell culture infectivity, host response microarrays, and pathogen-specific microarrays will improve the water industry’s ability to protect consumers from potential microbiological threats.