Multilevel responses to copper toxicity – linking field community impacts with laboratory-bred organism responses Katy Jeppe, Jianghua Yang, Sara Long, Melissa Carew, Xioawei Zhang, Vin Pettigrove, Ary Hoffmann Research Summit 2016

Measuring toxicity Levels of ecological toxicity assessment More specific / causal Molecular, Biochemical Community Population Individual Physiological Ecosystem More ecologically relevant

Measuring toxicity Levels of ecological toxicity assessment More specific / causal Molecular, Biochemical Community Population Individual Physiological Ecosystem More ecologically relevant Field-based assessment More ecologically relevant Multiple factors Difficult to control Lab-based tests Less environmentally relevant One or a few factors More specific/causal Better control

Measuring toxicity Levels of ecological toxicity assessment More specific / causal Molecular, Biochemical Community Population Individual Physiological Ecosystem More ecologically relevant Field-based assessment More ecologically relevant Multiple factors Difficult to control Lab-based tests Less environmentally relevant One or a few factors More specific/causal Better control

How do these assessment techniques compare? Experiment aims: Are common lab-bred species representing field communities? Compare different ecological levels used to assess toxicity Assess the sensitivity of different levels used in lab-bred species and field community responses.

Copper causes toxicity in sediment Essential metal Agriculture Urban 5 concentrations of copper spiked sediment (5 reps) Unspiked control (15 reps)

Field-based microcosm approach Colonized by aerial invertebrates Field-based community responses Investigates sediment toxicity while controlling other factors Environmentally relevant field conditions

Field-based microcosm approach Experimental design Field-based microcosm approach Field-based community and abundance:

Field-based microcosm approach Experimental design Field-based microcosm approach Field-based community and abundance: Lab-bred individual and population:

Field-based microcosm approach Experimental design Field-based microcosm approach Field-based community and abundance: Lab-bred individual and population: Lab-bred sub-individual: Metabolomics and gene expression

Experimental design Field-based community (<7 week exposure) Aerial macroinvertebrates Lab-bred species Snails (6 week exposure) Potamopyrgus antipodarum (n=50) Physa acuta (n=5) week exposure) Insects (5 day exposure) Chironomus tepperi (n=40)

Endpoints at different ecological levels Field-based community Community structure and abundance Lab-bred individual and population Snail survival, reproduction and juvenile survival Chironomus survival and growth Lab-bred sub-individual Chironomus gene expression and metabolite abundance

Endpoints at different ecological levels Field-based community Community structure and abundance Lab-bred individual and population Snail survival, reproduction and juvenile survival Chironomus survival and growth Laboratory-bred sub-individual Chironomus gene expression and metabolite abundance

Endpoints at different ecological levels Field-based community Community structure and abundance Lab-bred individual and population Snail survival, reproduction and juvenile survival Chironomus survival and growth Lab-bred sub-individual Chironomus gene expression and metabolite abundance

Results Field-based community Rank Taxon Family Number of individuals % total abundance Number of microcosms 1 Paratanytarsus Chironomidae 5940 35.1 46 2 Procladius 3707 21.9 3 Chironomus 3635 21.5 33 4 Kiefferulus 2356 13.9 25 5 Cladopelma 279 1.7 11 6 Chaoboridae 211 1.2 7 Parachironomus 194 1.1 8 Berosus Hydrophilidae 182 9 Polypedilum 104 0.6 29 10 Oribatida Not identified 99 27 Microvelia Veliidae 48 0.3 12 Ceratopogonidae 45 13 Ephydridae 44 18 No difference in community structure (MRPP) but abundance did change

Results Field-based community To compare responses EC50 ranked by ecological guidelines Sensitive: Below PEC <149 mg/kg Moderate: Between PEC and ISQG-high 149-270 mg/kg Tolerant: Above ISQG-high >270 mg/kg ©entomart

Results Field-based community To compare responses EC50 ranked by ecological guidelines Sensitive: Below PEC <149 mg/kg Moderate: Between PEC and ISQG-high 149-270 mg/kg Tolerant: Above ISQG-high >270 mg/kg ©entomart

Results Field-based community To compare responses EC50 ranked by ecological guidelines Sensitive: Below PEC <149 mg/kg Moderate: Between PEC and ISQG-high 149-270 mg/kg Tolerant: Above ISQG-high >270 mg/kg ©entomart

Results Field-based community To compare responses EC50 ranked by ecological guidelines Sensitive: Below PEC <149 mg/kg Moderate: Between PEC and ISQG-high 149-270 mg/kg Tolerant: Above ISQG-high >270 mg/kg ©entomart

Results lab-bred individual and population

Results lab-bred individual and population P. antipodarum juvenile survival was the only sensitive lab-bred individual/population response Sensitive: Below PEC <149 mg/kg Moderate: Between PEC and ISQG-high 149-270 mg/kg Tolerant: Above ISQG-high >270 mg/kg

Results lab-bred individual and population P. antipodarum embryo production was a moderate lab-bred response Sensitive: Below PEC <149 mg/kg Moderate: Between PEC and ISQG-high 149-270 mg/kg Tolerant: Above ISQG-high >270 mg/kg

Results lab-bred individual and population All other lab-bred individual/population responses were tolerant Sensitive: Below PEC <149 mg/kg Moderate: Between PEC and ISQG-high 149-270 mg/kg Tolerant: Above ISQG-high >270 mg/kg

Results lab-bred sub-individual Cysteine metabolism in C. tepperi Genes measured are indicated in italic Metabolites measured are indicated in bold Metabolites not involved in the cysteine metabolism were also measured with LC-MS

Results Cysteine metabolism in C. tepperi Gene expression Metabolites

Results Other metabolites in C. tepperi Changes in: Ammonia metabolism, Melanin production and Copper complexing metabolites Opportunity for further investigation

Conclusions The most sensitive individual/population endpoints in lab-bred species was juvenile survival of P. antipodarum This endpoint was more tolerant than survival of Chironomus and Paratanytarsus field populations Sub-individual responses were the most sensitive lab-bred endpoints These were the only lab-bred endpoints to respond at concentrations where sensitive field-based populations were declining These responses also provided information on copper toxicity, demonstrating direct exposure through sequestration responses

Further work Metabolomic analysis revealed pathways of interest for investigating toxicity responses Melanin production and ammonia metabolism could provide useful biomarkers of metal exposure Mixtures – individual contaminant exposure was investigated here Are similar rankings observed with field sediments containing complex mixtures?

Thanks to Rebecca Reid, Cameron Amos, Patrick Bonny, Harry Eason, Rhianna Boyle, Georgia Sinclair, Tyler Mehler Daniel MacMahon, Dave Sharley, Steve Marshall, And all CAPIM staff and students