RP220 (AAA-1117-094737) The use of a mode-of-action based test battery as an innovative approach to monitor surface water quality.
Start time: 8:00 AM
Chèvre, N¹, Behra, R¹, Eggen, R¹, Rutishauer, S¹, Schweigert, N¹, Escher, B¹, ¹ EAWAG, Duebendorf, Switzerland
Today, monitoring surface water quality is typically done by comparing chemical analytical measurements with water quality criteria, which are often based on the ecotoxicological effects of these substances. The analytically measured substances, however, only partly represent the substances in the environment leading to an incomplete evaluation of the surface water pollution. Furthermore, this approach does not consider the joint toxic action of the substances present in the environment. An alternative would therefore be to measure the toxicity of surface water samples with ecotoxicological test systems to evaluate the hazard of the pollutants for the environment. In our project, we developed a mechanism-based test battery, based on in-vitro and low complexity test systems, to monitor surface water quality. The different tests of the battery are based on different modes of toxic action that are frequently encountered in the environment, e.g. photosystem II inhibition or estrogenic effects. This test battery is proposed as early warning system to detect potentially dangerous substances in the environment and have preliminarily been tested to evaluate the quality of a river nearby Zürich, Switzerland. The results show that the test battery can successfully highlight the effects of photosystem II inhibitors and androgenic substances present in the river. The effects of estrogenic substances were more difficult to identify in this specific river, probably because of interactions of estrogenic substances with compounds present in the river samples′ extracts. No genotoxicity was detected in the river water samples. The results of a more complete study on an other Swiss river will also be presented. These studies should serve as basis to implement the test battery as a tool to assess the quality of the Swiss surface water.

RP221 (MEI-1117-568773) DNA damage and external lesions in Brown Bullheads from contaminated habitats.
Start time: 8:00 AM
Meier, J¹, Yang, X², Chang, L¹, Baumann, Paul², ¹ U.S. Environmental Protection Agency, Cincinnati, OH, U.S.A.² U.S. Geological Survey, Columbus, OH, U.S.A.
The single cell gel electrophoresis (Comet) assay was used to compare levels of DNA damage in brown bullheads (Ameiurus nebulosus) collected from three known contaminated locations, the Cuyahoga River, Ashtabula River, and Ashumet Pond (Cape Cod), with brown bullheads collected from three paired reference sites, Old Woman Creek, Conneaut River, and Great Herring Pond, respectively. Blood was sampled from the caudal vein of each fish and the Comet assay was conducted on erythrocytes. Raised external lesions and barbel abnormalities were also recorded. The results demonstrated that fish from the three contaminated sites each suffered significantly higher DNA damage (measured as % tail DNA, tail length and tail moment) than fish from their respective reference sites. The gender and age of the fish did not affect the Comet parameters. Positive relationships were observed between levels of DNA damage and presence of external abnormalities. This study provides evidence that the Comet assay is sufficiently sensitive to detect exposure of natural fish populations to environmental levels of genotoxic contaminants, and suggests this method may be useful as a recent or predictive indicator of carcinogenic exposure in aquatic ecosystems. Although this work was reviewed by EPA and approved for publication, it may not necessarily reflect Agency policy.

RP222 (HAL-1117-215761) Evaluation of macroinvertebrate bioassessment metrics in four pulp and paper mill effluent receiving waters.
Start time: 8:00 AM
Hall, T¹, Ikoma, J¹, Ragsdale, R¹, Arthurs, W¹, ¹ National Council for Air and Stream Improvement (NCASI), Anacortes, WA, US
An evaluation of data for 14 benthic macroinvertebrate bioassessment metrics collected over a six to seven year period for four U.S. rivers is reported. This evaluation is of interest because some of these metrics have been incorporated into assessment programs some of which have been developed and implemented by the USEPA (RBP), Canada (EEM) as well as in other countries. The current review provides an opportunity to evaluate the utility and sensitivity of individual metrics as influenced by season to season and year to year variation as well as from stream gradient changes. Also evaluated is the ability of these metrics to provide a meaningful signal from the influences of both point and non-point discharges as measured through a suite of concurrently measured water quality parameters. Metrics evaluated include those based on richness, diversity, similarity, tolerance, and functional feeding groups. Rivers in the study included the McKenzie and Willamette Rivers in Oregon, the Leaf River in Mississippi, and Codorus Creek in Pennsylvania. Each study site included five to six monitoring stations which extended over an area of from 28 to 52 km. All four watersheds included a pulp and paper mill effluent discharge, one or more tributary streams, and varying degrees of human influences in the riparian zone and surrounding landscape. Information utility from these individual bioassessment metrics is also compared to a multimetric assessment approach carried out previously with these same data.

RP223 (RAT-1117-058161) Terrestrial Vertebrate Contaminant Exposure and Effects Data for Mid-Atlantic National Park Service Lands.
Start time: 8:00 AM
Ackerson, Betty¹, Rattner, Barnett¹, McKernan, Moira¹, ¹ USGS-Patuxent Wildlife Research Center, Beltsville, MD, USA
The USGS-Biomonitoring of Environmental Status and Trends project determines the effects of contaminants on biota and lands under the protection of the Department of the Interior. As part of this program the Contaminant Exposure and Effects-Terrestrial Vertebrates (CEE-TV) database (www.pwrc.usgs.gov/contaminants-online) was created in 1996 to compile ecotoxicological information and to help identify taxonomic, temporal and spatial data gaps in the Atlantic, Gulf and Pacific Coasts, Hawaii, Alaska and the Great Lakes Region. Previous analyses of CEE-TV records revealed that 60 of 126 National Park Service (NPS) units in these regions lacked recent (since 1990) contaminant exposure and effects data for terrestrial vertebrates. Currently we are conducting a more in-depth evaluation of contaminant threats and ecotoxicological data gaps for terrestrial vertebrates within or near (10 km) 23 NPS units in National Capital Region and Mid-Atlantic Networks. Contaminant exposure and effects data were compiled from each park unit and 10 km buffer. To identify contaminant threats, data layers from the Environmental Protection Agency's Toxic Release Inventory Sites, Fish Consumption Advisories, Section 303(d) Impaired Waterbodies, and USGS Land Use/Land Covers were overlaid onto NPS unit boundaries and their buffers in ArcGIS. Of the 23 park units reviewed, 20 units contained ecotoxicological data for terrestrial vertebrates and 13 units have recent (since 1990) information. Harper's Ferry National Historical Park, Antietam National Battlefield, and Hopewell Furnace National Historic Site and their respective buffers, are in regions with known water quality problems (e.g., exceed Total Maximum Daily Load for priority organics and metals) and vulnerable to pollution (e.g., >50% agricultural and urban land use). The NPS units that lack recent CEE-TV records and fall within areas of concern should be given priority for future ecotoxicological research and monitoring.

RP224 (NAL-1117-827001) Low Salinity Acclimation and Salinity Tolerance Of Americamysis bahia in Short-Term Chronic Toxicity Tests.
Start time: 8:00 AM
Nally, C¹, MacGillivray, A², Pallop, T¹, Koch, K¹, Fikslin, T², ¹ American Aquatic Testing, Inc., Allentown, PA, USA² Delaware River Basin Commission, West Trenton, NJ, USA
Efforts to study and characterize the nature and extent of chronic toxicity in the Delaware Estuary have been hampered by the limited number of EPA approved test species that are tolerant of low salinity levels present in the estuary. The purpose of this work was to develop supporting data for test species and test methods selected by the Delaware River Basin Commission Chronic Toxicity Workgroup for use in monitoring ambient water with salinities between 1 to 10 ppt. The mysid, Americamysis bahia, has been selected as a test species because mysids play an important role in the estuarine environment and have a history of use in standardized tests. However, based on data from long-term chronic toxicity tests, current EPA guidance is to conduct A. bahia tests at salinities between 20 to 30 ppt. This requires salinity adjustment in many samples from the estuary that may potentially alter the toxicity profile of the samples. This study evaluated the effects of salinities at 25, 20, 15, 10 and 5 ppt on the test species, A. bahia in short-term chronic toxicity tests after acclimation of the test organisms. Mysids tested at all salinities, with the exception of 5 ppt, exhibited survival of >80% and survival was not statistically significant different from organisms tested at 25 ppt. The growth endpoint exhibited a statistically significant difference in salinities <20 ppt. However, the average weights of the organisms were within the upper and lower control limits for the laboratory culture in all salinities except 5 ppt. The two different test lots of A. bahia that were acclimated to 10 ppt and used to conduct a series of standard reference toxicity tests met all test acceptability criteria for short-term chronic toxicity tests and produced results comparable to A. bahia testing performed at the recommended 25 ppt.

RP225 (SAR-1117-798363) Towards the development of a new automated sediment toxicity test with Lumbriculus variegatus.
Start time: 8:00 AM
Sardo, A.M.¹, Gerhardt, A.^{1, 2}, Soares, A.M.V.M.¹, ¹ Department of Biology, University of Aveiro, Aveiro, Portugal² LimCo International, Ibbenbüren, Germany
Lumbriculus variegatus is a very common freshwater oligochaeta, that is used in a large number of toxicity tests and that presents a remarkable ability to regenerate. The Multispecies Freshwater Biomonitor® (MFB), measures online different behaviors of aquatic species based on the registration of changes in a high frequency alternating current, caused by movements of the organisms in their test chambers and it is based on quadropole impedance technology. The aim of this short term experiment was to develop a new automated sediment toxicity test using L. variegatus. Lead was chosen in this test as model substance due to the affinity to sediment particles. The following concentrations were used, in order to contaminate the sediment: 25.0, 50.0, 100.0, 150.0 and 300.0 mgPb/L, and a control with uncontaminated ASTM water. In each concentration level, 6 worms (about 3 cm each) cut into equally long two pieces per replicate were used. An experiment with clean sediment and contaminated water was also performed. Survival, fitness (color), regeneration ability and speed as well as and position (sediment/water) were monitored daily. Some tests with the MFB were performed in order to obtain behaviour signals.

RP226 (CUR-1117-769666) Evaluation of the Asiatic Clam (Corbicula fluminea) as a Standard Toxicity Test Organism.
Start time: 8:00 AM
Currie, R¹, Baxter, R¹, Alexander, A², Echols, B², Cherry, D², ¹ Roanoke College, Salem, VA, USA² VA Tech, Blacksburg, VA, USA
Acute and chronic toxicity tests rely upon commonly accepted organisms such as Ceriodaphnia dubia, Daphnia magna and Pimephales promelas, to indicate various levels of pollution and to set protective limits for industrial discharges. These species are easily cultured and widely researched relative to toxicological responses which have resulted in well-established toxicity databases. However, due to the lentic nature of both C. dubia and D. magna and the wide distribution of P. promelas, these organisms may not be protective of more lotic, sedimentary organisms such as freshwater mussels. The goal of this research was to provide an additional test organism, Corbicula fluminea, for use in standard toxicity tests. Corbicula has the potential to be utilized in both acute and chronic water column tests and chronic sediment tests. The first objective of this research focused on the laboratory culturing of recently released pediveligers in an effort to effectively maintain pediveliger populations through maturity into umbonal juveniles. Attempts to culture pediveligers through the juvenile stage were unsuccessful. A secondary objective was to use developing juveniles to establish LC₅₀ values for comparison to currently used test organisms. Acute toxicity test results showed C. fluminea (≤24 hrs) to be more sensitive to sodium chloride (NaCl) than C. dubia or D. magna with a mean LC₅₀ value of 1.41 g/L NaCl in 48-hr toxicity tests compared to 2.26 g/L (C. dubia) and 4.56 g/L (D. magna). Corbicula were even more sensitive to NaCl after 7-days old with a mean LC₅₀ value of 0.96 g/L.

RP227 (PET-1117-750052) Element concentrations and fluctuating asymmetry in a terrestrial isopod (Armadillidium vulgare) in metropolitan Chicago parklands.
Start time: 8:00 AM
Peters, E¹, Kaufmann-Daszczuk, B¹, Brant, H², Jagoe, C², Richter, R¹, ¹ Chicago State University, Chicago, IL, USA² Savannah River Ecology Lab, Aiken, SC, USA
Terrestrial isopods have long been known to bioaccumulate high levels of Cu, Fe, Zn, Cd, and Pb, and are established biological indicators of these metals. Southern metropolitan Chicago, Illinois has a long history of contamination from industrial activities (e.g., metal smelting, electroplating, and paint manufacture) and associated waste production (e.g., steel mill slag, coal and coke ash). Many of the sources of these contaminants were located within or near residential areas. We sampled populations of the terrestrial isopod Armadillidium vulgare collected neighborhood parks and other natural areas in metropolitan Chicago and its surrounding suburbs, including NW Lake County, Indiana (total area > 600 km²). We sampled isopods (n > 60) from 21 sites in late June-early July 2004, and again from 12 of these sites in September. We analyzed these isopods for 20 elements by ICP-MS. Only Ag was consistently below method detection limits: most populations were above the MDLs for Sb and Hg, and all populations had detectable concentrations of the other elements. As expected from previous studies, isopod populations within the city accumulated higher levels of Cu, Fe, Zn, Cd, and Pb than did isopods in suburban areas. We also found elevated concentrations of other less common but potentially hazardous elements, e.g., V, Cr, Ni, As, and Ba. We noted apparent effects of these contaminants on postnatal development: concentrations of toxic metals and metalloids (and combinations of these elements) were positively correlated with the degree of fluctuating asymmetry in compound eye lens (ommatidia) number. As an abundant (and invasive) species in habitats ranging from woodlands, residential neighborhoods, and parks to industrialized areas throughout the Northern Hemisphere, A. vulgare may be an ideal sentinel animal for monitoring the presence and effects of metal and metalloid pollutants in urban environments.

RP228 (CON-1117-820857) Rhythmic Motions of Invertebrates as Assay Systems for Environmental Toxins.
Start time: 8:00 AM
Nath, N.¹, Stockero, J.¹, Klaper, R¹, Consi, T¹, ¹ UWM Great Lakes WATER Institute, Milwaukee, WI, USA
Many organs and structures in the transparent bodies of small freshwater invertebrates exhibit rhythmic motion. The frequencies of these motions may provide useful information about the effect of environmental toxins on these animals at sub-lethal but still physiologically relevant concentrations. Conversely, changes in the frequency of a specific organ may provide a qualitative and/or quantitative assay for the presence of a specific toxin or class of toxins. A great advantage of using frequency as the indicating parameter is the ease with which it can be measured. This could lead to the development of automated whole-animal assay systems that could be deployed in the field. Another advantage is that the different rhythmically moving organs of these animals will permit measurement of the effects of different classes of toxic compounds (e.g. neuro-toxins, compounds that affect cilliary activity). We are using two freshwater invertebrates to explore this idea, Daphnia (various species) and the embryos of the ramshorn snail (Planorbis sp.). The Daphnia has a pulsing heart, rhythmically oscillating mouth parts and thoracic legs, and a large mobile eye. Ramshorn snail embryos are deposited in a 2-dimensional mass of several eggs thus permitting the examination and analysis of multiple animals. These embryos exhibit muscular motions, ciliary movement, and heart pulsations. We have developed a simple computerized system to extract the frequency spectrum from video recordings of these animals. Our analysis system will be presented along with preliminary results on the effects of exposure to several compounds including: caffeine, fluoxetine, and clofibrinic acid.

RP229 (GOL-1117-818085) Characterization of Esterases in Different Organs of the White Grunt (Haemulon plumieri) for their Use as Biomarkers in the Mesoamerican Barrier Reef System.
Start time: 8:00 AM
Alpuche-Gual, L¹, Gold-Bouchot, G¹, ¹ CINVESTAV Unidad Merida, Merida, Yucatan, Mexico
The Mesoamerican Barrier Reef System is considered the second longest barrier reef in the world. It is an area with a high biodiversity, both in terms of species and habitats, which is threatened by development of coastal areas. The white grunt (Haemulon plumieri; Lacepede, 1801) has been chosen as a bioindicator species for the regional monitoring program; it is a fish of tropical and warm-temperate waters that is distributed in the Atlantic coast from Virginia to Florida, along the Caribbean, and to the south of Brazil. The biological, physiological and ecological information that exists on this organism is scarce, and neither has it been studied from the ecotoxicological point of view. The objective of this work is to characterize the activities of cholinesterases and carboxylesterases in brain, liver, muscle gills, eye and spleen, and to determine if there any differences in the activities with respect to size. Twenty five fish were collected in a place near a fishing village, without agricultural activity, and were taken alive to the laboratory where they were sacrificed and brain, liver, muscle, gills, spleen and eye were excised; the gonads were preserved to determine sex and maturity. Cholinesterase activity was determined by the Ellman modified technique, carboxylesterase activity by the method of Mastropaolo and Yurno, and protein according to Lowry. Results indicate that cholinesterase activity is highest in the liver, followed by the brain and muscle; the gills, eye and spleen had a very low activity, and are inadequate to determine variations caused by pollutants. Only the liver presented a measurable carboxylesterase activity, and the other organs had very low activities of this enzyme. There were no significant correlations between correlation weight or total length with esterase activity, and thus size is not a factor that affects esterase activity in the of the white grunt.

RP230 (BOW-1117-812785) Evaluation of WET testing as an indicator of aquatic health.
Start time: 8:00 AM
Bowersox, M¹, Latimer, H¹, Diamond, J¹, Lessard, J¹, Stribling, J¹, Bixler, C², ¹ Tetra Tech, Inc., Owings Mills, MD, USA² WERF, Alexandria, VA, USA
There is uncertainty regarding WET compliance limits as effective predictors of effluent effects on aquatic life resources. Other previous studies have examined the relationship between WET and instream biological integrity, most of these used outdated test methods with unspecified data quality. This study is a 2-year pilot project focusing on the relationship between WET testing results and instream bioassessment using standardized WET test methods and high quality bioassessment data. Standardized WET test methods will include three-brood chronic Ceriodaphnia dubia, 7-day chronic fathead minnow (Pimephales promelas), and 96-hour green algae (Selenastrum capricornutum) as described by USEPA, 2002. Six dischargers have been selected to participate in the study representing a range of influent and effluent conditions including industrial, municipal, effluent dominated and chronic toxicity. Quarterly WET tests using all three species and a full bioassessment of all three representative communities will be conducted. Macroinvertebrates will be assessed using a 20-jab dipnet method and, for select sites, using Hester-Dendy artificial substrate samplers. Fish communities will be assessed via electrofishing, and periphyton will be sampled for volume and diversity. The overall study plan uses a data quality objective approach (DQOs), which includes the type and quality of data needed to satisfy the study objectives. This pilot study will be used to test relationships between WET test results and biological conditions, as well as provide a framework for a broader, more definitive study.

RP231 (HIN-1117-804837) Contaminant concentrations and biomarker responses of fish from the Mobile, Apalachicola, Savannah, and Pee Dee River Basins.
Start time: 8:00 AM
Hinck, J¹, Blazer, V², Denslow, N³, Gross, T⁴, May, T¹, Orazio, C¹, Coyle, J⁵, Tillitt, D¹, ¹ USGS-CERC, Columbia, Missouri, USA² USGS-LSC, Kearneysville, West Virginia, USA³ University of Florida, Gainesville, Florida, USA⁴ USGS-FISC, Gainesville, Florida, USA⁵ USGS BEST Program, Fort Collins, Colorado, USA
Many chemical manufacturers, pulp and paper mills, textile operations, and power plants are located in the Mobile, Apalachicola, Savannah, and Pee Dee River Basins. Contaminants released from these industries and other sources may pose a risk to fish in nearby waters. To address these concerns, the Large River Monitoring Network (LRMN) of the Biomonitoring of Environmental Status and Trends (BEST) Program measured tissue concentrations of selected contaminants and evaluated biomarker responses in largemouth bass (Micropterus salmoides) and common carp (Cyprinus carpio) from four sites in the Mobile River Basin (MRB) and three sites each in the Apalachicola, Savannah, and Pee Dee River Basins. Composite samples of whole fish from each site were analyzed for organochlorine and elemental contaminants and for 2,3,7,8-tetrachloro dibenzo-p-dioxin-like activity (TCDD-EQ) using the H4IIE rat hepatoma cell bioassay. Microsomal ethoxyresorufin O-deethylase (EROD) activity was measured to determine exposure to AhR agonists in individual fish. Mean EROD activity was uniformly greater in carp (>20 pmol/min/mg) from sites in the MRB than those from other basins; mean activity in bass was generally >40 pmol/min/mg at all sites. Fish health indicators (condition factor, somatic indices, general histopathology) were examined in relation to chemical concentrations and reproductive biomarkers (steroid hormones, gonadal histology, vitellogenin) to identify potential effects of exposure. Mean health assessment index (HAI) values were >100 in bass from the MRB and were attributed to abnormalities of the gills, liver, kidney, and spleen, which were likely the result of parasitic infestations; HAI values in carp were generally low. In addition, concentrations of 17-estradiol and 11-ketotestosterone were relatively low in female carp from the Coosa and Tombigbee Rivers in the MRB compared to those from other sites. These data will aid in characterizing fish health and contaminant concerns in the Mobile, Apalachicola, Savannah, and Pee Dee River Basins.

RP232 (PAR-1118-113484) The monitoring of heavy metals in human bloods of middle school students.
Start time: 8:00 AM
Park, Hee Ra¹, Kwun, Ki-Sung¹, Kim, Meehye¹, Kim, Soon Ki², Lee, Hyo Min¹, Kim, Soo Yeon¹, Choi, Kwang Sik¹, ¹ Korea Food & Drug Administration National Institute of Toxicological Research, South of Korea² Inha University Medical School, South of Korea
This study was conducted to estimate the contents of heavy metals including lead, cadmium, zinc, copper as well as iron status(serum iron, total iron binding capacity, ferritin etc)in blood samples of middle school students(n = 300). The contents of heavy metals were determined using the GF-AAS (Graphite Furnace Atomic Absorption Spectrophotometer). The microwave digestion method and dilution method were compared. The dilution method showed the better recovery and detection limit than microwave digestion method. The values of toxic metals in whloe blood of boys & girls were 3.46 & 3.05 for Pb, 0.063 & 0.065 for Cd respectively (ug/dL). Also the values of trace metals in serum of boys & girls were 105.9 & 92.6 for Zn, 98.3 & 99.0 for Cu respectively (ug/dL). The prevalence of iron deficiency was 7.5% in 146 boys and 14.3% in 156 girls. The mean values of lead in girls were higher in iron deficiency, iron deficiency anemia and anemia groups than normal group. The mean values of lead and zinc were higher in boys compared to those in girls(P<0.05), the mean values of cadmium and copper in boys were similar to those in girls. Our results of toxic metals such as Pb & Cd showed lower to CDC's(Centers for Disease Control) blood lead levels of concern for children, 10 ug/dL.

RP234 (PUL-1117-837846) Biomonitoring for the Georgia coast: Oysters (Crassostrea virginica) as sentinels of coastal water quality.
Start time: 8:00 AM
Pulster, E¹, Singleton, M¹, Danforth, J¹, Frischer, M¹, Maruya, K², ¹ Skidaway Institute of Oceanography, Savannah, Georgia, USA² Southern California Coastal Water Research Project, Westminster, California, USA
The eastern oyster (Crassostrea virginica) is one of the most abundant bivalve species inhabiting the eastern U.S. coastal zone. Because they accumulate chemical and microbial contaminants and thus integrate exposures via the water column, bivalves including C. virginica have been used as integrative water quality bioindicators for decades. The overall objective of this project is to initiate an environmental observation system for coastal Georgia (USA) water quality based on this ecologically relevant indicator species. Oysters were collected from viable beds from up to 16 sites in the six coastal GA counties and analyzed for a suite of heavy metal and organic contaminants, as well as bacterial indicators of water quality and oyster health (total and fecal coliforms, Enterococci, and bioluminescent strains). Site selection was based on differing land use (pristine, oyster harvesting beds, industrial, commercial, and residential waterfront). Collection is taking place in summer and winter for a three year period, beginning in May 2005. The data will be compiled to compare contaminant levels associated with different land uses, to assess for relationships among chemical contaminants and biological levels/indicators, and to ultimately create a map of coastal GA water quality.

RP235 (KIM-1117-821753) The development of six-channel biological early warning system and new methodology to detect the activity of Daphnia magna.
Start time: 8:00 AM
Jeon, J¹, Kim, S¹, Kim, K¹, ¹ Gwangju Institute of Science and Technology, Gwangju, Korea
Biological early warning system (BEWS) has been requested to reduce false alarms in application to real sites by the system operator. The false alarm is serious problem and may result in workloads at the water quality monitoring station. To achieve the accurate detection, a new methodology was designed to identify and to quantify the activity of Daphnia magna, it was equipped with a six-channel individual monitoring system including a software program, grid counter (GC) and several hardware for measuring water quality parameters. We also used the relative activity parameter (_a) to detect the increased activity of D. magna. It is defined as _a=(N_t−A_c)/A_c where N_t= counted number by GC at time t, A_c = cumulative average until time t. Using six _avalues calculated from 6 different channel monitoring systems that produced the activity data at every 5 min, the Student′s t-test was performed to verify the difference between the average value as a control and the exposure values at every measurement simultaneously. The exposure tests were conducted with different copper concentrations, 50, 100, 200, and 400 ppb. Each test chamber (total 6 chambers) contained one D. magna and the alarm was generated at p < 0.005 in the basis of _a value. The average time to reach two consecutive detections of significant differences was 1.2, 0.58, 0.60, and 0.29 hours for Cu concentration of 50, 100, 200, and 400 ppb, respectively. From the results, we suggest that p values from the t-test based on _a be more accurate, stable and predictable parameter to detect the chemical spike than other values like swimming speed, trajectory, and so on. Consequently, it is possible to reduce false alarms as well as to give the confidence for system to operators with highly accurate detection ability by using the six-channel monitoring system.