WP107 (LON-1117-812969) Expression and Induction of a Cytochrome P450 family 4 gene identified from Chironomus tentans Exposed to atrazine.
Start time: 8:00 AM
Londono, D.¹, Siegfried, B.¹, Sarath, G.², Lydy, M.³, ¹ University of Nebraska, Lincoln, NE, U.S.² USDA-ARS, Lincoln, NE, U.S.³ Southern Illinois University, Carbondale, IL, U.S.
Cytochrome P450s represent the single most important enzyme family involved with detoxification of xenobiotics. Induction of specific P450 isozymes in insects is often observe after treatment with a variety of xenobiotics and induction of the P450 system can have important consequences to the ability of insects to tolerate exposure to pesticides. Studies performed in our lab have measured the effect of atrazine exposure on cytochrome P450-dependent monooxygenase activity and have found increased activity as a result of atrazine exposure (10 ppm) in midge larvae (Chironomus tentans). A 45 kDa protein of increased intensity was observed after SDS-PAGE of microsomal protein that is similar in size to cytochrome P450 enzymes reported by other insects. Immunochemical studies using a Drosophila melanogaster anti-P450 polyclonal antiserum, further support the cytochrome P450 nature of this inducible 45-KDa protein. Identification of specific atrazine-inducible genes could enhance sensitivity of detection and provide insight into potential consequences of exposure. The present research identified an atrazine-inducible CYP4 gene with an open reading frame of 1,680 bp (accession number AY880065) which was amplified and sequenced from C. tentans. Alignments of inferred amino acid sequences with other insect CYP4 gene homologues indicate a high degree of similarity. Northern blot analysis employing a fragment of 1,200 bp form the CYP4 gene as a probe was used to examine expression of CYP4 among different developmental stages and among atrazine exposed and unexposed larvae. The gene was expressed in all life stages although was most abundant in larval instars. Exposure to atrazine at 10 ppm caused approximately 2-fold increase in signal intensity confirming the inducible nature of gene.

WP108 (COU-1117-830688) Pesticide concentrations in critical zones of the Saint Lawrence Estuary and their toxicity to fish.
Start time: 8:00 AM
Couillard, C.¹, Lebeuf, M.¹, Roy, R.¹, DeBlois, C.², ¹ Fisheries and Oceans Canada, Mont-Joli, Quebec, Canada² Centre d'expertise en analyse environnementale du Québec, Sainte-Foy, Quebec, Canada
Coastal marshes of the St. Lawrence Estuary (SLE) are critical habitats for several fish species and may be contaminated by pesticides from urban or agricultural runoffs. The objectives of this study were: (1) to measure concentrations of pesticides in SLE tributaries irrigating four coastal marshes, (2) to evaluate fish health and biomarker responses in resident three-spine stickleback (Gasterosteus aculeatus), and (3) to determine toxicity thresholds of selected pesticides for marine fish larvae. From May to September 2003, water was sampled weekly at each site. Concentrations of pesticides were generally low (e.g., atrazine ≤ 200 ng/L; diazinon ≤ 10 ng/L) but were higher at the two agricultural sites than either the urban or the reference sites. In 2003 and 2004, sticklebacks were collected from the four sites during the months of June and July. The activity of acetylcholinesterase (AChEase), measured in muscle and brain tissues of captured sticklebacks, was not different among sites. In the laboratory, mummichog (Fundulus heteroclitus) larvae were exposed to concentrations of the organophosphorous insecticide diazinon for 96-h. Mummichog growth was inhibited at the lowest exposure concentration (124 ng/L), while inhibition of AChEase was observed only at concentrations ≥ 361 ng/L. These results indicate that inputs of pesticides from the SLE tributaries to the coastal marshes are relatively low and that there are no obvious toxic impacts in resident fish. Further studies are underway to evaluate pesticide inputs from the marine water of the SLE and to investigate potential endocrine disruption in these fish populations.

WP109 (CAR-1116-956681) Assessing persistence and aquatic toxicity of a xylene-emulsifier herbicide to salmonids and daphnids.
Start time: 8:00 AM
Cardwell, R.¹, van Genderen, E.¹, Buhler, G.², Caldwell, R.², Edgington, A.¹, Power, L.¹, Smith, C.¹, ¹ Parametrix Environmental Research Laboratory, Albany, OR, USA² Northwest Aquatic Sciences, Newport, OR, USA
A mixture of xylene and an emulsifier (the adjuvant) is used as an herbicide to control some of the aquatic plants that can clog irrigation fixtures and retard flow in eastern Washington irrigation canals. Because the treated water cannot be released downstream before it is considered acutely nontoxic to aquatic life, acute toxicity bioassays of rainbow trout fry and Daphnia magna were used to define the risks posed by the emulsifier, xylene, and herbicide mixture. Bioassays rather than chemical analysis were judged to be the most cost effective means for assessing net bioavailability and toxicity of the chemical mixtures. Because the emulsifier, a mixture of seven chemicals, was acutely toxic and affected mixture toxicity, its persistence was measured using bioassay, as was that of the herbicide mixture. Although the emulsifier constituted only 2% of the formulated herbicide (98% xylene), mixture toxicity was considered because definitive toxicity tests with both organisms suggested the emulsifier was up to seven times more toxic than xylene alone on a volume per volume basis. In fact, rainbow trout were equally sensitive to the emulsifier and the formulated herbicide while D. magna appeared to be most sensitive to the herbicide exposure. Whether the interaction was additive or more-than-additive could not be determined. Based on slow stirring in beakers to simulate mixing in flowing canals, the emulsifier was less persistent (half-life 3 hr) than the xylene (half-life 7.5 hr), but both dissipated rapidly via volatilization. There was evidence that some of the emulsifier's degradation intermediates were acutely toxic if the treated water was held long enough (≥75 hr @ 20 C) before release to natural streams. Toxicity bioassays proved to be a cost-effective and technologically effective means for assessing toxicity, persistence and toxicological interactions of this complex mixture.

WP111 (SUN-1117-767696) Responses of non-target species to two algaecides and an adjuvant used to control Lyngbya wollei.
Start time: 8:00 AM
Sundberg, S¹, Jones, R¹, Rodgers, Jr., J¹, ¹ Clemson University, Clemson, SC, USA
Two algaecides (Algimycin PWF® and PAK-27™) and an adjuvant (Cide-Kick II®) have been used to control the nuisance algal species, Lyngbya wollei, in Lay Lake and Lake Jordon, Alabama. Information regarding the comparative toxicities of these products is essential for assessing potential risks or adverse effects to non-target species. The objectives of this study were: 1) to measure the relative toxicities and potencies of Algimycin PWF, PAK-27, and Cide-Kick II to a vertebrate, Pimephales promelas Rafinesque, and two invertebrates, Hyalella azteca Saussure and Daphnia magna Straus, in static, non-renewal, short-term (96-hr) aqueous and sediment laboratory exposures with survival as the measured endpoint, and 2) to measure the relative toxicities and potencies of mixtures of the two algaecides and adjuvant as they were applied in the Alabama reservoirs. Three replicate test chambers (250-mL beakers), 10 juvenile organisms each, were used for each concentration tested and a control for all experiments using site water. Test chamber temperature was maintained at 25±1^oC in a temperature-controlled incubator with a light:dark photoperiod of 16hr:8hr. Water characteristics including dissolved oxygen, pH, hardness, alkalinity, and conductivity were measured throughout exposure experiments. Relative toxicities (NOECs and LOECs) of each algaecide, adjuvant, and mixture were calculated based on organism survival using analysis of variance (ANOVA, alpha = 0.05). Lethal concentrations (LC₅₀s) of algaecides and/or adjuvant were calculated using the Spearman-Karber method. Estimated aqueous LC₅₀s for PAK-27 were above the targeted concentration of 6.21 mg/L in Lay Lake for treatment of L. wollei, whereas estimated aqueous LC₅₀s for Cide-Kick II were much lower than the targeted concentration of 5 ml/L. However, the presence of sediment in the test chambers helped to alleviate toxicity to organisms. Results from this research provide essential information regarding the risks to non-target species involved in treating L. wollei with these products in these Alabama reservoirs.