detection of ecotoxicological effects at environmentally relevant concentrations of some antifouling...
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Detection of ecotoxicological effects at environmentally relevant concentrations of
some antifouling and personal care products
Hans Blanck
Plant & Environmental Sciences
Biology
University of Gothenburg
Sweden
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Communityecotoxicology
Mixture ecotoxicologyNatural &
experimental ecosystems
Ecotoxicogenomics &bioinformatics
Ecotoxicology
METALS METALOIDS ANTIFOULANTSHERBICIDES
ANTIFOULANTS PHARMACEUTICALS
PERSONAL CARE PRODUCTS NANOPARTICLES AND MATERIALS
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Mind the gap!!!
• Regulatory Ecotoxicology mostly describes the immediate physiological consequences of toxicity in simplified systems
• Ecological status of the environment is the consequence of the a long-term selection pressures (also from mixtures of toxicants) on complex and variable systems
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PICT will respond when the most sensitive of these species are replaced by more tolerant ones
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Pollution-Induced Community Tolerance
PICT• PICT works with natural communities sampled in a
gradient of toxicant exposure• Community tolerance increases when sensitive species
are eliminated by toxicants• Community tolerance is quantified in short-term tests for
suspected toxicants• PICT can detect small community effects and ”identify”
the cause of the damage
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PERIPHYTON
- attached microbial communities on submerged surfaces
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Microcosm studiesEstablishment during controlled exposure
Flow-through system
Field studiesEstablishment during
environmental exposure
Periphyton communities
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Area for antifouling PICT studies and metasequencing of marine periphyton
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Ferreira, K.N., Iverson, T.M., Maghlaoui, K., Barber, J., Iwata, S., 2004. Architecture of the photosynthetic oxygen-evolving center. Science 303, 1831-1838.
IRGAROL INHIBITSPHOTOSYSTEM II IN PLANTS AND ALGAE
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Field study of irgarol tolerance
Blanck et al 2009 Marine Pollution Bulletin
• Irgarol levels stable over the years• Irgarol and Cu dominated the pleasure craft market in Sweden from 1994 -2004
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Irgarol-induced succesionIrgarol-induced succesion
1994 2001
Pre-exposure concentration (nM)
0 5 10 15 20Spe
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0 5 10 15 200
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No PICT detected in 1994 PICT readily detected 2001
DIFFERENTIAL SENSITIVITY IS A REQUIREMENT
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The distributions of psbA were very dissimilar between control and irgarol-exposed communities
The diversity in psbA was much greaterthan expected. • 72 haplotypes were found• these translated into 40 different D1-proteins • haplotypes came from various taxa
Community-wide sequencing of the psbA gene Community-wide sequencing of the psbA gene encoding the irgarol target D1encoding the irgarol target D1
Eriksson, K.M., Clarke, A.K., Franzen, L.G., Kuylenstierna, M.,Martinez, K., Blanck, H., 2009. Community-Level Analysis of psbA Gene Sequences and Irgarol Tolerance in Marine Periphyton. Applied and Environmental Microbiology 75, 897-906.
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PEST region
Herbicide binding niche
Ser264→Gly
Picture from from Erickson et al (1989) The Plant Cell (1) 361-371
D1 protein
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What about mixture toxicity with other PSII inhibitors ???
• Irgarol• Atrazine• Diuron• Isoproturon• Terbacil• etc.
TRIAZINES, PHENYL UREAS, URACILS ETC
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Congeneric mixtures of PSII inhibitorsPeriphyton photosynthesis
Observed April 1998
CA model provides good prediction
CA model provides good prediction
Arrhenius, A., Gronvall, F., Scholze, M., Backhaus, T., Blanck, H., 2004. Predictability of the mixture toxicity of 12 similarly acting congeneric inhibitors of photosystem II in marine periphyton and epipsammon communities. Aquatic Toxicology 68, 351-367.
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Congeneric and non-congeneric mixtures of PSII inhibitors
Periphyton community structure
Porsbring, T., Backhaus, T., Johansson, P., Kuylenstierna, M., Blanck, H., 2010. Mixture toxicity from photosystem II inhibitors on microalgal community succession is predictable by Concentration Addition. Environmental Toxicology and Chemistry 29, 2806-2813.
IrgarolDiuronTerbacil
DiuronIsoproturonFluometuron
DiuronIsoproturonFluometuron
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Clotrimazole
• Very toxic to aquatic organisms• OSPAR Commission 2005: ”currently no clear environ-mental risk from clotrimazole”• RQ > 2 for low tide marine scenarios• OSPAR Commission 2006: listed clotrimazole for priority action
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Clotrimazole inhibits 14 α-demethylase in plant-sterol metabolism
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Precursors increase and products decrease
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Sterol metablism in periphyton affected at environmental concentrations (50 pM)
MEC: 100 pM
PNEC: 300 pM
PEC: 600 pM
Porsbring, T., Blanck, H., Tjellstrom, H., Backhaus, T., 2009. Toxicity of the pharmaceutical clotrimazole to marine microalgal communities. Aquatic Toxicology 91, 203-211.
Porsbring, T., Blanck, H., Tjellstrom, H., Backhaus, T., 2009. Toxicity of the pharmaceutical clotrimazole to marine microalgal communities. Aquatic Toxicology 91, 203-211.
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Complicated response pattern:some algae stimulated some inhibited
Pigment profiles
Xanthophyll cycle
Chlorophyll a
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Clotrimazole summary
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MIXTURES of pharmaceuticals & personal care products
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MIXTURES ofpharmaceuticals and personal care products
• Good predictions by both IA and CA
• Strong hormesis at low concentrations of mixture
• Effects based on total pigment content
• Periphyton EC50 for triclosan is 1-2 nM depending on endpoint
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Something from ”Nothing”....Periphyton communities
5-component mixture at the NOECs of the individual components yield a 28.3% inhibition, predicted to be 30% according to IA.Prediction based on previously generated data gave 23 % (IA) and 30% (CA).
Backhaus, T., Porsbring, T., Arrhenius, A., Brosche, S., Johansson, P., Blanck, H., 2011. SINGLE-SUBSTANCE AND MIXTURE TOXICITY OF FIVE PHARMACEUTICALS AND PERSONAL CARE PRODUCTS TO MARINE PERIPHYTON COMMUNITIES. Environmental Toxicology and Chemistry 30, 2030-2040.
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Antibiotics and periphyton- tetracycline
• Environmentally relevant tetracyline concentrations (< 1 – 100 microgram/l) in US
• 0.5 ug tetracycline per litre affected:
• Antibiotic resistance• Abundance and productivity of
bacteria• Algal and cyanobacterial biomass• Nematode abundance• At higher doses bacterial
productivity recovered, although the food web did not
US EPA´s Stream Mesocosm Facility
Quinlan EL et al. 2011.Temporal dynamics of periphyton exposed to tetracycline in stream mesocosms. Env Sci Technol. In press
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Ecological considerations•Ecological Energy Flow: 100 % through primary producer and decomposer compartments•Huge diversity and many phyla•Huge diversity implies broad set of potential target molecules and species•Potential for cascading effects in food webs (e.g. deficiency in nitrogen, sterols, energy, or food preferencies for species ,sizes, shapes, nutrient content etc )•Selected Test Organisms are Examples not Models...
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SCENIHRScientific Committee on Emerging and Newly Identified Health Risks
Assessment of theAntibiotic Resistance Effects of Biocides
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Triclosan favours antibiotic resistance in periphyton biofilms
Dosing Period (d)
0 7 14 21 28 35 42 49 56
% R
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Cu Exposure under Field Conditions Coselects for Antibiotic Resistance as Determined by a Novel Cultivation-Independent
Bacterial Community Tolerance Assay
Berg et al 2010. Env Sci Technol 44: 8724-28
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Chemical Hazard and Risk Assessement
•Present procedures tend to ignore the role of biodiversity and genetic diversity
•Present procedures tend to ignore the role of chemodiversity and mixture complexity
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Emergent comments...
•Emerging pollutants suggest that present control of chemicals fails•Why are we surprised that pharmaceuticals and preservatives are causing effects in the environment
• WFD is ambitious but its present approach to chemical status is disappointing. We tend to monitor those that we can handle.•Monitor ALL biocides, pharmaceuticals, PBT, vPvB, metals, HVCs at selected sites.•WFD requires good understanding of environmental mixtures
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The NICE projectNovel instruments for effect-based assessment
of chemical pollution in coastal ecosystems• Use present chemical monitoring data to provide
predictions of mixture toxicity in Swedish coastal waters
• Select sites and extend chemical analysis• Use biological monitoring tools (e.g. PICT, fish
biomarkers) to confirm suspected ecotoxicological effects or detect new ones
• Explore the sensitivity and specificity of ecotoxicogenomics in selected biota (periphyton,fish, invertebrates)
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Thank YOU !!!!!!
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PICT vision
• PICT detects ecological effects from toxicants in the environment
• PICT indicates the toxicant (group) that caused the effect.
• PICT is used systematically in Europe e.g. under the Water Framework Directive
• PICT is used together with community ecotoxico-genomics for mechanistic understanding, for higher specificity – and for mutual validation...
A EUROPEAN BIOFILM NETWORK IS BEING ESTABLISHED
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Irgarol, PICT and the psbA gene
• Irgarol induced community tolerance in periphyton, but only slowly.
• No differential sensitivity in 1994• Mutations in PEST region favoured by irgarol
selection pressure in diatoms (not cyanobacteria)
• A tolerance mechanisms for PS-II inhibitors proposed in the 1970´s, was discovered in nature.
• A mechanistic understanding of irgarol PICT
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Antibiotic contaminated sediment increase gene frequencies of resistance genes and genes involved in
horizontal gene transfer
Kristiansson, E., J. Fick, et al. (2011). "Pyrosequencing of Antibiotic-Contaminated River Sediments Reveals High Levels of Resistance and Gene Transfer Elements." Plos One 6(2).
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• Antibiotic resistance # biocides # overlapping resistance mechanims #
• mobile genetic elements (plasmids) # biocides may help maintaining plasmids for cross-resistance # dissemination of plasmids of cross-resistance in biofilms creates a potential risk of AR development
• Antibiotic resistance # biocides # overlapping resistance mechanims #
• mobile genetic elements (plasmids) # biocides may help maintaining plasmids for cross-resistance # dissemination of plasmids of cross-resistance in biofilms creates a potential risk of AR development