Development of Toxicity Indicators Scientific Steering Committee Meeting July 26, 2005 Sediment Quality Objectives For California Enclosed Bays and Estuaries

Presentation Overview Objectives Selection of candidate test methods Evaluation of candidates Recommendations Application and integration issues

Toxicity Indicators Many available data and methods Several challenges to effective use –Differential sensitivity/ reliability of methods –Confounding factors –Ecological relevance

Objectives Select a suite of recommended acute and chronic toxicity test methods Describe sensitivity, reliability, and ecological relevance for each method Develop thresholds for use in MLOE framework

Attributes of Toxicity Indicator Suite Protective of benthos and ecologically relevant –Sensitive at the scale of benthic impacts Dependable –Results are reproducible and comparable among labs Diverse endpoints –Mortality and sublethal response Different taxa –Greater representation of benthos Diverse exposure conditions –Matrix and duration

Approach for Test Selection Establish a list of candidate methods –Potential to meet desired attributes Compile and synthesize information about tests –Relate to desired test characteristics –SQO database, literature, lab studies, other scientists Select recommended tests –Match indicator attributes –Best combination of desired characteristics

California Test Data

Candidate Test Characteristics Well-documented and feasible for use in California Appropriate exposure method Good sensitivity and precision

Candidate Toxicity Indicators Short-term survival –Multiple species of amphipods –Direct sediment exposure –Widely used in California Short-term/embryo development and fertilization –Sea urchins and mussels –Frequently used in California –Pore water or sediment-water interface exposure Chronic/sublethal response –Usually species with limited use in California –Diverse endpoints and exposure methods

Growth Tests Polychaete –28 day exposure, fed –Dry weight –Frequently used in California Seed clam –7 day exposure, fed –Dry weight –Not used in California

Life Cycle Tests Amphipod –28 day exposure, fed –Dry weight, offspring –Occasionally used in California Copepod –14 day exposure, fed –Offspring –Not used in California

Cell Stability Test Oyster –4 day exposure, fed –Digestive gland cell stability –Not used in California

Candidate Tests Amphipod survival (10 day sediment exposure) –Ampelisca abdita –Eohaustorius estuarius –Leptocheirus plumulosus –Rhepoxynius abronius Growth/Reproduction (28 day sediment exposure) –L. plumulosus –Neanthes arenaceodentata (polychaete) Sea urchin fertilization (1 day pore water exposure) –Purple sea urchin (Strongylocentrotus purpuratus)

Candidate Tests Continued Embryo development (2-3 day pore water or sediment-water interface exposure) –Purple sea urchin –Mussel (Mytilus galloprovincialis) Copepod life cycle (14 day sediment exposure) –Amphiascus tenuiremus Clam growth (7 day sediment exposure) –Mercenaria mercenaria Oyster lysosomal stability (4 day sediment exposure) –Crassostrea virginica

Presentation Overview Objectives Selection of candidate test methods Evaluation of candidates Recommendations Application and integration issues

Evaluation Process Separate evaluation for short-term survival and sublethal test methods Short-term survival –10-day amphipod tests are accepted Species selection is primary issue Sublethal tests (many issues) –Feasibility –Consistency –Confounding factors –Sensitivity –Relevance –Cost

Evaluation Process Short-term survival tests –Compared attributes of four species typically used in 10-day amphipod tests Technical feasibility and supply Sensitivity Confounding factors

Amphipod Survival Test Characteristics SpeciesAvailability (months) SuppliersTechnical Difficulty Confounding Factors Ampelisca8Field/few Moderate ** Ammonia Eohaustorius12Field/fewLowAmmonia Leptocheirus12Lab/fewLowAmmonia Rhepoxynius12 Field/few * LowAmmonia Grain size * Special permit needed ** Higher test failure rate

Amphipod Survival Ampelisca is markedly less sensitive to CA sediment samples

Amphipod Species Recommendations Recommended –Eohaustorius estuarius –Leptocheirus plumulosus Not recommended –Rhepoxynius abronius Limited availability Grain size sensitivity –Ampelisca abdita Low sensitivity Low test success rate

Evaluation of Sublethal Tests Compared characteristics of candidate tests –Technical feasibility and supply –Relevance to program objectives –Reproducibility and precision –Documentation and history –Sensitivity –Cost Sequential process

Sublethal Test Characteristics TestAvailability (months) SuppliersTechnical Difficulty Method Guidance Amphipod growth12Lab/fewModerateStandard Polychaete growth12Lab/oneModerateStandard Pore water fertilization 8Field/manyModerateStandard Pore water development 6-12Field/manyLowStandard SWI development6-12Field/manyLowPublished Copepod life cycle12Lab/oneHighPublished Clam growth8Lab/manyLowPublished Oyster lysosome8Field/manyModerateReport

Sublethal Test Characteristics TestAmong Lab Variability Within Lab Variability More Sensitive Than Amphipod Survival Test Relative Precision of Response* Amphipod growthFairGoodSometimesLow Polychaete growthGood SometimesLow Pore water fertilization GoodFairSometimesHigh Pore water development Good SometimesHigh SWI developmentFairGoodSometimesLow Copepod life cycle?GoodOftenHigh Clam growthGoodFairSometimesSimilar Oyster lysosome??RarelyLow * Compared to 10 day amphipod test

Sublethal Tests: Relative Sensitivity

Sublethal Tests: Feasibility and Supply Not recommended –Copepod life cycle test No commercial test lab capability in California or U.S. Technically difficult No information on reproducibility No standard method –Oyster lysosome stability No commercial test lab capability in California or U.S. Technically difficult No information on reproducibility No standard method

Sublethal Tests: Exposure Method Not recommended –Interstitial water tests with sea urchins or mussels High sensitivity to confounding factors Less realistic exposure scenario Remaining candidates have differing combinations of desirable attributes for use in SQO program –Clam growth –Polychaete growth –Amphipod growth/reproduction –Embryo development at sediment-water interface

Sublethal Tests: Documentation and History Not recommended –Juvenile clam growth test No standard method Limited applied experience with method in California

Sublethal Tests: Sensitivity and Cost Not recommended –Leptocheirus reproduction and growth Higher relative cost Lower relative sensitivity

Recommendations Conduct both acute survival and sublethal test to evaluate sediment toxicity Acute survival test –Either E. estuarius L. plumulosus Sublethal response test –Either Polychaete growth test (N. arenaceodentata) Sediment-water interface test using mussel or sea urchin embryos

Application and Integration Issues Thresholds –How to interpret results of each test Integration of results –Multiple tests –Weighting Applications –New studies –Past data

Thresholds Several options for establishment –Statistical Significant difference from control Variability –Biological Magnitude of response Association with benthic community response A combined approach is recommended –Unaffected (good) vs. affected (bad) Desire confidence in choice Statistically-based threshold –Severity of effect Magnitude and ecological relevance of response is important Biological-based threshold

Proposed Thresholds Reference/no effect Low effect Moderate effect High effect –Response not statistically significant different from control –Response different from control, but less than test-specific minimum significant difference (msd) –Result may be within test variability –Clear effect, but below level associated with high probability of benthic effects –Up to 50% effect relative to control (if benthic association unknown) –Above level associated with likely benthic community impacts –> 50% response relative to control (if association unknown)

Integration of Results A combination of concordance and magnitude of response Sublethal test results receive less weight Four categories of effect

Integration of Results CategoryDescription ReferenceNo effect in any test Marginal DeviationLow effect in only one test, may not be reproducible or ecologically significant ModerateEffect measured in multiple tests or a severe effect in one test HighSevere effect in one test and concordance of effects among tests

Proposed Integration Framework LOE Category Amphipod Survival Result Sublethal Result RefLowModerateHigh Ref Marginal Deviation ModerateModerate? LowRefModerate High Moderate Marginal Deviation ModerateHigh Moderate High

Applications New studies –Should use both types of tests –If only one used Reject information? Only amphipod survival: use result as LOE classification? Only sublethal data: Use conditionally with modified weighting? –Other test methods used Ancillary information, not a substitute for specified tests

Applications Past data –Incomplete test suite? –Other amphipod species? –Other methods?