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More Information about PATHSCAN and PATHCAST
Background
Information about the effects of anthropogenic influences and environmental degradation on the distribution, abundance, and virulence of waterborne pathogens is rapidly increasing (Henrickson et al., 2001). Human enteric pathogens are known to cause widespread waterborne disease in the United States both through ingestion of contaminated seafood and direct water contact, costing an estimated $20 billion per year in lost productivity (CDC). In 1999 alone, for example, 342 cases of confirmed non-cholera Vibrio infections were reported to the Center for Disease Control through their Vibrio Surveillance System. Fourty six percent of these cases were hospitalized and 11% resulted in patient death. While Vibrio parahaemolyticus was most prevalent, 94% of the mortalities were associated with V. vulnificus. While monitoring human health and disease outcomes is important, protecting human health and the economic value of coastal and estuarine environments and resources requires the development of rapid pathogen detection and forecasting tools.
Traditional identification techniques for pathogenic bacteria are labor-intensive and time-consuming because long incubation periods and selective media are required to culture these organisms (Lyon, 2001). With the advent of Polymerase Chain Reaction (PCR) technology, however, the ability to screen for pathogens in environmental samples has been greatly enhanced. Unfortunately, simply detecting these pathogens in the environment is often of limited value, as many are a normal component of healthy marine⁄estuarine ecosystems. The need to rapidly quantify pathogens, and virulent strains, has begun to be addressed by the development of real-time or quantitative PCR which utilizes a fluorescent reporter to quantify the exponential increase in PCR product with each cycle. In addition, coupling real-time PCR with multiplex PCR, or the use of several primers and fluorescent reporters simultaneously, offers the ability to screen and quantify multiple species or multiple genes within a species rapidly, within hours (Blackstone et. al, 2003). This is particularly beneficial in the quantification of slow growing and difficult to culture organisms such as many Mycobacterium spp. (weeks to months) and viable but non-culturable states. While molecular techniques offer a mechanism for rapid screening and reporting of pathogens in water, a further understanding of conditions which favor growth of specific pathogen species and virulent strains is important for the development of predictive capabilities.
Literature Cited
Bruijnesteijn van Coppenraet, E.S, J.A. Lindeboom, J.M. Prins, M.F. Peeters, E.C.J. Claas, and E.J. Kuijper. 2004. Real-time pcr assay using fine-needle aspirates and tissue biopsy specimens for the rapid diagnosis of mycobacterial lymphadenitis in children. Journal of Clinical Microbiology, 42(6): 2644-2650.
Blackstone, G.M., J.L. Nordstrom, M.C.L. Vickery, M. Bowen, R. Meyer, and A.DePaola. 2003. Detection of pathogenic Vibrio parahaemolyticus in oyster enrichments by real time PCR. The Journal of Microbiological Methods, 53(2): 149-155.
Henrickson, S.E., T. Wong, P. Allen, T. Ford, and P.R. Epstein. 2001. Marine swimming related illness: Implications for monitoring and environmental policy. Environmental Health Perspectives, 109 (7): 645- 650.
Lyon, W.J. 2001. TaqMan PCR for detection of Vibrio cholerae 01, 0139, Non-01, and Non-0139 in pure cultures, raw oysters, and synthetic seawater. Applied and Environmental Microbiology, 67(10):4685-4693.