Download Microbial Source Tracking • Pathogen Contamination –What is it?

Microbial Source Tracking
• Pathogen Contamination
–What is it?
–Where is it coming from?
–Who is making it?
–How can we address it?
Tiered Approach
Weight
of Evidence
Microbial Source
Tracking
Intensive sampling using
standard FIB measurements;
Infrared thermography
Sanitary Survey
Identification of impaired areas based on long-term
monitoring
Traditional Culturing Methods
EPA Rapid Methods
• Quantitative Polymerase Chain Reaction
(qPCR)
• 16S ribosomal RNA (16S rRNA) markers
Method A targets Enterococcus
Method B targets Bacteroidales
qPCR in Microbial Source Tracking
• 16S rRNA gene found in nearly all bacteria
and Archaea
• Small changes in genes allow for
identification of hosts
• qPCR allows for quantification of specific host
inputs
qPCR Basics
• Low detection limit
– Pro: very sensitive
– Con: low levels are not likely to be health risks
• Detects living and dead cells
– Pro: conveys source information
– Con: might be outdated source information
• Can detect lots of sources
– Pro: potentially unlimited host tracking ability
– Con: each host detection requires a separate assay
• Provides quantitative information
– Pro: enables evaluation of relative contribution of host animals
– Con: requires innovative calibration work
Year OneMethod Validation
Year Two Pilot Project: Withers Swash
Source Tracking Strategies
• Space and Time
– Targeted wet and dry weather sampling
• Tracers
– Bacteria concentrations
– Chemical concentrations
• Water mass indicators
• Optical brighteners
• Caffeine
– Host animals
• Multiple antibiotic resistance of fecal bacteria
• Genetic markers of fecal bacteria
Illicit Discharge Detection and Elimination: A Guidance Manual for
Program Development and Technical Assessments (CWP and Pitt,
2004)
Sampling
• 2 dry
• 3 wet
Nalgene
stormwater
sampler
– First flush sampling
Hobo
Site-Specific Storm Hydrographs
Hobo water level
loggers
Figure 3.3.1-1: Hydrograph showing the response to a rain event at Site 1 as recorded by an in12
situ Hobo™ water level logger.
Figure 3.4.1-1: Concentration of Enterococcus (MPN/100 mL) as a function of sampling
site. Black circles represent dry samples and white circles represent wet samples.
Figure 3.4.1-4:
Enterococcus
concentrations
(MPN/100 mL) in Withers
Basin sub-watersheds.
Figure 3.4.1-2: Enterococcus and E. coli concentrations
Figure 3.4.2.2-1: BacHum
concentrations (genome
copies/100 mL) in Withers
Basin sub-watersheds.
Results below the
reporting limit are in the
dark green-shaded
grouping.
Figure 3.4.2.3-1: BacCan
concentrations (genome
copies/100 mL) in
Withers Swash subwatersheds.
Concentrations grouped
into quartiles.
Weight-of-evidence approach using indices
• By Parameter
– Data grouped into quartiles with the bottom bin set at regulatory threshold
– Each data group assigned an integer value aka a “grade”
Table 3.4.1-1: Concentration ranges used to assign rank orders to E. coli and
Enterococcus concentrations. These values were used to create Table 3.4.1-2.
Weight-of-evidence approach using indices
•
•
By Parameter
By Site
– Each parameter assigned a “grade”
– Parameter grades summed to determine site “grade”
• Averaged Wet vs Dry
• Averaged Overall
Table 3.4.1-2. Fecal Indicator Bacteria (E. coli + Enterococcus) “Grades”
Weight-of-evidence approach using indices
•
•
•
By Parameter
By Site
Visualizing sites to prioritize for remediation
– Color coded matrixes
– Qualitative rankings
Table 3.4.3-3: Qualitative descriptors
Weight-of-evidence approach using indices
•
•
•
By Parameter
By Site
Visualizing which sites to prioritize for remediation
– Color coded matrixes
– Qualitative rankings
– Every site with a “very strong” detection of FIB had a “strong” or “very strong” detection for at least
one of the qPCR assays
Table 3.4.3-4: Summary of the results of the qualitative descriptors for regulatory FIB and qPCR
markers. Sites with very strong qualitative rankings are in red font.
Table 3.5.1-1: Sites with “strong” and “very strong” levels of BacHum concentrations.
Percentages determined by dividing the average rank order by the maximum rank
order for each tracer. Percentages greater than 50% are shaded red.
Table 3.5.1-2: Sites with “very strong” levels of BacCan. Percentages determined by
dividing the average rank order by the maximum rank order for each tracer.
Table 3.6.4-1: Sites with strong evidence for either human or canine sourced
Bacteroides.
Citation: Wood, J., J.M. Trapp, S.M. Libes, and E.J. Burge. 2013. Watershed Assessment Report:
Stormwater Management Planning: Development of a Pilot Investigative Approach to Remediate Bacterial
Source Impairments along the Grand Strand. Final Report, prepared under the authority of Section 22 of
the Water Resources Development Act of 1974 for the US Army Corps of Engineers, Charleston District;
Horry County, SC; Georgetown County, SC; City of Myrtle Beach, SC; and City of North Myrtle Beach, SC.
100 pgs + Appendices.
http://www.coastal.edu/envsci/projects/pollution/documents.html
Thank you to our
Partners
Figure 3.4.2.1-1: GenBac
concentrations (genome
copies/100 mL) in Withers
Basin sub-watersheds.
Concentrations are grouped
into quartiles.
What we want our
MST data to be
Sources of bacterial pollution
in Withers Swash
“Universal (GenBac)
Wildlife
Human (BacHum)
Dog (BacCan)
Goose (CGO-1F)
What our MST data actually are
Goose
(CGO-1F)
Wildlife
Dog
(BacCan)
“Universal”
(GenBac)
Human
(BacHum)
Sources of bacterial pollution
in Withers Swash