What s In Your Water? Microbial Source Tracking

Transcription

1 What s In Your Water? Microbial Source Tracking Valerie J. arwood, Ph.D. University of South Florida FSA Winter Conference 12/07/16

2 Agriculture Pets Sewage & Runoff Wildlife Fecal contamination affects water quality Water quality affects all of us!

3 Pathogens Associated with Sewage and Fecal Contamination of Water Are Numerous and Diverse Viral pathogens Bacterial pathogens O157:7 ID agar Norovirus Protozoan pathogens Salmonella E. coli O157:7 Cryptosporidium Giardia

4 Yet We Rely Almost Exclusively on Fecal Indicator Bacteria to Warn of Contamination Act as a warning signal of fecal pollution and increased risk of associated pathogens in water

5 Drawbacks of FIBs FIBs may be absent when pathogens are present FIBs are frequently present when pathogens are absent FIBs have many possible sources

6 Consequences of Error FIB Absent FIB Present Pathogen Absent Pathogen Present

7 Drawback: FIBs ave Many Possible Sources! Sources vary from highest risk (human sewage) to definite risk (cattle, poultry) to unknown risk (wildlife, pets, sediments, soils, stormwater) Can t remediate or prevent pollution effectively without knowledge of dominant source(s) of contamination

8 Who Poos There?

9 The Central ypothesis of MST Certain microbial species or types are associated with the gastrointestinal tract of specific animals hosts. This association can be used to track the fecal microorganism back to its host. S S S S S S S S S S S

10 ciudadswcd.org Library-Dependent MST Approach (Largely Outdated) Library????

11 The Library-Independent MST Approach Based on Genes of ost- Associated Microbes ( Markers ) Bov Polymerase chain reaction targets a specific gene & copies it Forensic-type evidence for contamination source

12 Strategy for Developing MST Markers 1. Identify microorganism that is confined to a particular host. 2. Identify gene that will discriminate this organism from all others. 3. Develop PCR method to selectively amplify the gene. 4. Test the PCR method for sensitivity, specificity, limit of detection and other performance characteristics. Bacteroides (microbewicki@kenyon.edu)

13 Sounds Simple ow ard Could MST Be? A. Specificity if we detect a given signature (marker), how sure are we that it came from a particular source? (poor sensitivity falsepositives) X X X X X X X

14 Non-Specific Markers Produce False Positives

15 Sounds Simple - ow ard B. Sensitivity Could It Be? In what percent of fecal samples from the target source can the marker be detected? And what is the concentration of the marker?

16 Non-Sensitive Markers Can Produce False Negatives????????

17 And Many Possible Sources Require Multiple MST Assays

18 Design-A-Study 1. What is study goal? Source apportionment TMDL? Remediation look for a major source and deal with it? Risk assessment how likely is human health risk? Legal whose animals or sewage is fouling the water?

19 Design-A-Study 2. Reconnaissance (old-fashioned sanitary survey) Define the watershed(s). Use google maps. Walk and drive around and look for animals, pipes, low-lying areas where water stagnates Talk to the health department, utilities, neighbors encourage squealing! Do some low-cost mapping with FIB to find trends and high/low areas

20 Design-A-Study 3. Choose markers based on (1) performance and (2) availability No marker is better than a non-specific marker. If you need to develop and validate a marker, count on it taking at least a year. Look for at least 85% specificity and 30% sensitivity in the literature. Test the marker(s) on fecal/sewage samples from your area assume nothing!

21 Bird (Gull2 and GFD) and uman- Associated Markers

22 Our Current Reality 100% Sensitive & Low LOD, BUT specificity issues uman Bacteroides Quantitative? Most methods target single source uman Viruses 100% Specific BUT low sensitivity

23 Questions?