Continuous Improvement and Evolution of Michigan’s

Water Quality Testing

Presented By Alexandra Strohm

Outline

•Background

•History of Water Quality Testing

•Current Testing Methods

•Applications

•Future Methods

Background

Alexandra Strohm

• Junior Michigan State University

• Biochemistry and Molecular Biology, Pharmacology and Toxicology

• MDEQ Student Assistant Water Toxic Unit

• Lyman Briggs Molecular Biology PCR Lab MSU

“Short History of Water Pollution Abatement in Michigan”

• Francis Frost wrote a short a summary of Michigan’s history of water pollution

• Frost was one of 3 member Michigan Water Resources Commission staff

• Documented the struggle of making Michigan’s water quality a priority

1923 1965 1968 1929 1935 1948 1952 1972 1990 2000 1997 1988 1995

“Short History of Water Pollution Abatement in Michigan”

• From 1923 to 1929 pollution control personnel were divided between several divisions with separate budgets Problem

• Act 245 Public Acts of 1929 created The Stream Control Commission Act • Towns that were ordered to build sewage plants tried to abolish commission

until about 1946

1923 1965 1968 1929 1935 1948 1952 1972 1990 2000 1997 1988 1995

“Short History of Water Pollution Abatement in Michigan”

1923 1965 1968 1929 1935 1948 1952 1972 1990 2000 1997 1988 1995

1972 Clean Water Act

• NPDES (National Pollutant Discharge Elimination System) demanded permits for discharge and waste

• Limits were set to meet standards to make water fishable and swimmable

• Grants awarded for wastewater treatment

1923 1965 1968 1929 1935 1948 1952 1972 1990 2000 1997 1988 1995

MDEQ BEACH Act

• 1995 MDEQ was created by Governor John Engler

• In 2000, BEACH (Beaches Environmental Assessment and Coastal Health) Act passed

• EPA provides grants for beach monitoring and notification programs

1923 1965 1968 1929 1935 1948 1952 1972 1990 2000 1997 1988 1995

Water Quality Standards

Michigan Public Health Code and Rule 323.1062(1) of the Part 4. Water Quality Standards (Promulgated pursuant to Part 31 of the Natural

Resources and Environmental Protection Act, 1997 PA 451, as amended)

MDEQ E. coli Testing

Idexx Colilert-18 Quanti Tray Method

• MPN determines E. coli density

• ‘Defined Substrate Technology’ (MUG and ONPG nutrient indicators)

• Synthesizes fluorescent E. Coli

• ONPG is substrate for B-galactosidase (enzyme) which turns the coliform yellow

• MUG is substrate for B-glucuronidase (enzyme) which causes E. coli to fluoresce

What is PCR?

Polymerase Chain Reaction

Amplify target segments of DNA Billions of copies!!!

Test amount or presence/absence of specific DNA markers

PCR: DNA Synthesis in Vitro

Cocktail ingredients:

Template DNA

Oligonucleotide Primers

dNTPs

Taq Polymerase

PCR Buffer

Mg2+

What is PCR?

• Denaturation (95 degrees Celsius)

• Annealing (~68 degrees Celsius)

• Elongation (72 egress Celsius)

The Problem

“In the United States alone, it is estimated that 39.2% of all rivers, lakes, and streams are unsafe for recreational use, with fecal pathogens as the number one cause of impairment”

How is qPCR useful in Water Testing?

The Problem

“The results showed…Enterococcus measured by QPCR can predict GI illness after swimming in fecally contaminated fresh water…Incorporation of rapid measurements such as these into a regulatory framework has the potential to improve beach management decisions and protect swimmers’ health.”

“Ultimately, the use of faster indicators of recreational water quality will result in the ability to make decisions about recreational water quality on the day of sample collection. This, in turn, could lower GI illnesses in communities, especially in those dependent on beach-related tourism.”

1

10

100

1000

10000

Singing Bridge

MPN_Singing GEC_Singing

y = 0.1505x + 20.386 R² = 0.1283

1

10

100

1000

10000

1 10 100 1000 10000

IDEX

X

EPA Method

Holland

y = 0.4688x + 30.961 R² = 0.5267

1

10

100

1000

10000

1 10 100 1000 10000

IDEX

X

EPA Method

Singing Bridge

Source Tracking: How do fix the problem?

• Identify the sources of E. coli present in a water sample

•Genetic markers in organisms

•Potential path to successful remediation

•qPCR

Chrysler

0

500

1000

1500

2000

2500

3000

2009 2010 2011 2012 2013 2014 2015 2016

E.co

li C

olo

ny

Form

ing

Un

its

pe

r 1

00

mL

Wat

er

Year

Chrysler Park: Individual Sample E.coli Concentrations (2009-2016)

Individual Sample of E.coliConcentration

TBC Maximum

Linear (Individual Sample ofE.coli Concentration)

Linear (TBC Maximum)

0

200

400

600

800

1000

1200

1400

2009 2010 2011 2012 2013 2014 2015 2016

E. c

oli

CFU

in p

er

10

0 m

L W

ate

r

Year

Chrysler Park: Individual Sample 50th and 95th Percentile E. coli Concentration (2009-2016)

50th Percentile

95th Percentile

Linear (50th Percentile)

Linear (95th Percentile)

Brimley State Park

Brimley State Park

0

500

1000

1500

2000

2500

2010 2011 2012 2013 2014 2015 2016

E. c

oli

Co

nce

ntr

atio

n in

MP

N/1

00

mL

Wat

er

Year

Brimley State Park: Individual Sample E. coli Concentrations (2010-2016)

Individual Sample E. coliConcentrations

TBC WQS

Linear (Individual Sample E. coliConcentrations)

Linear (TBC WQS)

0

500

1000

1500

2000

2500

2010 2011 2012 2013 2014 2016

E. c

oli

Co

nce

ntr

atio

n in

MP

N/1

00

mL

Wat

er

Year

Brimley State Park: Individual Sample 50th and 95th Percentile E. coli Concentrations (2010-2016)

50th Percentile

95th Percentile

Linear (50th Percentile)

Linear (95th Percentile)

Challenges with Source Tracking

•Fails near LLOQ

•Genetic markers must be highly specific

•Primers could bind to multiple organism DNA (Dogs and Human cross)

•Best markers and criteria still being developed

Source tracking: Human Vs. Dogs • “Study estimates that 39.1 percent of human pathogens also infect

domestic animals”

• Used genome fragment enrichment (GFE) to identify nonribosomal microbial genetic markers for PCR-based testing of dog fecal contamination

• DG3, DG37, DG72 markers

• 244 Fecal samples below LOD for noncanines (humans)

Digital PCR • Eliminates the need for a standard curve

• Plasson statistics/ corrections

• More sensitive smaller differences detected

• Less reagents needed (no standards, less wells used)

• No cross reactions

• Partitions sample into droplets or chambers

• Flow cytometer checks droplets

• Measures positive or negative/ accepted or not accepted

Thank You!