SEMINAR: MBF Student Seminar Series Presents David Wanless Friday 10/22 4p

[Sidney Hartley <shartley@rsmas.miami.edu>]

Improved monitoring of recreational waters by

incorporating molecular techniques

 

David Wanless

 

Abstract

 

The monitoring of recreational waters is important to gauge risks to human health. Currently, risks to human health within recreational waters are uncovered through classic microbiological testing techniques that measure fecal indicators. Methods currently in use are time-intensive (18 hours to test results) [1], test for a small # of indicators, and by the time results are known, actions such as closures or warnings may be irrelevant, as contamination concerns may have dissipated through natural currents or tides. [2]  There are new molecular techniques capable of delivering faster results (within 2-4 hours of collection) for indicator organisms along with alternative targets.  Additional targets of molecular assays could focus on source specific markers.  This would provide more data to water managers and a possible start to remediation of contaminants.  These molecular techniques rely on genetic discrimination instead of bacterial growth and metabolic activity.  However, these techniques cannot be utilized until extraction efficiencies are known.  By using known quantities of Lactococci to mimic the characteristics of Enterococci, it is possible to determine this efficiency.  This was accomplished by adding known quantities of Lactococci cells and Enterococci cells to filters and extracting those filters. After extraction, the Lactococci and Enterococci DNA was quantified using qPCR.  The percentage of Lactococci remaining in the elution was then compared to the results from the Enterococci assay. The cells correlated by .94 and slope and intercept were close to a 1:1 linear relationship (y = .9436x +.011) and a R² value of .89.  Assays were also developed to organisms associated with dog and gull fecal pollution.  The Dog assay was present in 90% of individual dog fecal samples (26/29), and 69% (20/29) of the fecal samples were highly positive (>E4 copies) for the marker. The gull assay was also tested for prevalence with our smaller library of gull and non-gull fecal samples.  All gull samples were positive for the marker (3/3) with the only cross reactivity being pelican.  By adding extraction controls and source tracking markers, results of assays can be fast, accurate, and provide information on the source of pollution in recreational waters.

 

B.S.  University of Wisconsin, 1998

Entered Masters Program August 2006

Advisor: Dr. Mike Schmale

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[1] U.S. EPA Method 1600  http://www.epa.gov/nerlcwww/1600sp02.pdf

[2] Leecaster, M., Weisberg,  S.B., 2001.  Effect of sampling frequency on shoreline microbiology assessments.  Mar. Pollut. Bull. 42, 1150-1154

-- 
Sidney L.S. Hartley
University of Miami
Rosenstiel School of Marine and 
Atmospheric Science
Division of Marine Biology and Fisheries
4600 Rickenbacker Causeway
S Grosvenor  315
Miami, Fl 33149
p: 305.421.4176
f: 305.421.4600
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