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Grant #1R01ES024358-01

Project Period8/18/2014* to 5/31/2018

PIsW Clements, J. Meyer, J. Ranville, C. Vulpe

National Institute of Environmental Health

Science

Possible approaches for assessing and predicting ecological effects of metals

Ecological Effects

Microcoms -Benthic communities-Organism level tests

Field-scale Exps.-Community surveys-Caged organism experiments

Possible approaches for assessing and predicting ecological effects of metals

Approaches to Assess/Predict BioavailabilityEcological Effects

Tissue Metal Levels-MSMM Biotic Ligand Models-Stable Isotopes

Bioavailable Fraction-Passive samplers-Sensors

Bioassays-Genomics-Single-species tox. tests

Microcoms -Benthic communities-Organism level tests

Field-scale Exps.-Community surveys-Caged organism experiments

North Fork of Clear Creek, CO Black Hawk/Central City Superfund Site

(COD980717557)

Future AMD Treatment Plant

Current Ecological Conditions

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Project ObjectivesProject 1: Linking Metal Bioavailability to Ecological Effects

Conduct a before-after control-impact (BACI) field experiment to quantify the relative influence of metal bioavailability and substrate composition on ecosystem recovery

Project 2: Linking Metal Bioavailability to Genomic Effects

Utilize gene expression in aquatic bioindicator species as molecular biosensors to assess bioavailability of metal mixtures (Cd, Cu, Ni, Zn) in the water column

Project 3: Linking Metal Bioavailability to Tissue Concentrations and Metal Speciation

Conduct experiments to determine the bioavailable pool of metal mixtures released from contaminated sediments exposed to variable water chemistries, and directly measure metal accumulation by bioindicator species.

Project 1 will test the hypothesis that despite significant improvements in water quality, ecological effects will persist in NFCC because of residual, high metal bioavailability in sediments (i.e., a biogeochemical hysteresis) and poor substrate quality

Project 2 and 3 will test the hypothesis that despite significant improvements in water quality, low concentrations of bioavailable metals will persist in NFCC (i.e., a biogeochemical hysteresis effect)

7Metal Mixture Effects Must be Considered

Example: Laboratory tests with D. magna using constant Cd and variable Ni

Increasing Ni reduces Cd toxicity until the point at which Ni becomes toxic

1E-4 1E-3 0.01 0.1 10

20

40

60

80

100

0.025 mg/L Cd

0.075 mg/L Cd

0.100 mg/L Cd

0.175 mg/L Cd Ni Only, 95% Uncertainty

Mor

talit

y %

[Ni] mg/L

Field Measurements of Sediment Contamination and Recovery

DepositionRecovery

Chemical Hysteresis is Observed Longer studies needed

Expose to Clear Creek

Equilibrate organism with stable isotopes

Microcosm colonization

Tissue metals

Multi-site Multi-metal BLM

Sub-cellular distribution of isotopes

Mineral phase analysis

sequential extraction

Sediment metals

Donnan Membrane: free ion activity

Dissolved Metals

Gene sequencing

Research approach integrating metal bioavailability assessment with ecological effects

Biologic response to toxicant

Response reflected in transcriptional changes

Transcriptional response

“profile” indicative of particular toxicant

pollutant

AAAAA

AAAAA

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Gene C

Gene B

Gene A

• Contaminant-specific “fingerprint” of toxicant exposure and effect =greater specificity• Direct assessment of organism response (no chemical manipulation) = increased relevance• Sub-lethal endpoints provides dose response information = increased sensitivity

Metal

Metal

Metal

Metal

Microarrays or RNAseqmeasure genome wide

response to metal

Molecular approaches to assessMetals in Aquatic Ecosystems

addis Fly Molecular Bioindicators The dream…

Identify genes (mRNA) whose expression levels change in response to metal exposure AAAAA

AAAAA

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Isolate mRNA

RNAseq – the next generation technology

metals

control

Sequence it All! Let compbio sortthem out

To develop a QPCR based panel of genes which can distinguish between metal exposures

To test QPCR panel bioassay in simulated field situations

To test QPCR panel bioassay in field situation

More biological relevance to aquatic ecosystems of interestMetal bioavailability in habitat may be different than water column

ACKNOWLEDGEMENTS:

# 1RO1ES020917-01 1R01ES024358-01

Students:Jacob Williamson, Elizabeth Traudt, Pete Cadmus, Katherine Ebeling

Samuel Fritkin, Ryan Mason, Thomas Gately, Kristina Lucas, Jason Loving

Postdocs and CollaboratorsRamiro Pastorinho, Zaho Dong

Additional Funding:

Jake

Katherine

Sam

EllieRamiro

Pete

Problem:•Two AMD sources introduce dissolved metals (Cu, Zn, Cd)•Stream sediments have accumulated Fe, Al, Mn oxide precipitates and associated metals

Opportunity:•Examine the recovery of the stream following restoration

Research Approach•laboratory/microcosm/in-stream experiments & field observations

Golden, CO

Blackhawk, CO

sport fishingwater supplyrecreation

Water Chemistry

Geochemistry

Biological Effects/Toxicity

Predict Recovery

Effects of Metals on Aquatic Systems: Clear Creek CO