Joint Action Effects of Emamectin Benzoate and Cypermethrin on the Marine Copepod Tigriopus Californicus Lindsay Schultz

Ursidae: The Undergraduate Research Journal at the University of Northern Colorado

Volume 5 | Number 3 Article 7

January 2016 Joint Action Effects of Emamectin Benzoate and Cypermethrin on the Marine Copepod Tigriopus californicus Lindsay Schultz

Michael Kelly

Follow this and additional works at: http://digscholarship.unco.edu/urj Part of the Biology Commons

Recommended Citation Schultz, Lindsay and Kelly, Michael (2016) "Joint Action Effects of Emamectin Benzoate and Cypermethrin on the Marine Copepod Tigriopus californicus," Ursidae: The Undergraduate Research Journal at the University of Northern Colorado: Vol. 5 : No. 3 , Article 7. Available at: http://digscholarship.unco.edu/urj/vol5/iss3/7

This Article is brought to you for free and open access by Scholarship & Creative Works @ Digital UNC. It has been accepted for inclusion in Ursidae: The ndeU rgraduate Research Journal at the University of Northern Colorado by an authorized editor of Scholarship & Creative Works @ Digital UNC. For more information, please contact [email protected]. Schultz and Kelly: Joint Action Effects of Emamectin Benzoate and Cypermethrin on th PESTICIDE EFFECTS ON A MARINE COPEPOD

Joint Action Effects of Emamectin Benzoate and Cypermethrin on the Marine Copepod Tigriopus californicus

Lindsay Schultz and Michael Kelly Faculty Mentor: Dr. Ginger Fisher, Biological Sciences

This project examined the effect of two pesticides on the marine copepod Tigriopus californicus. The pesticides are emamectin benzoate and cypermethrin, which are commonly used in salmon aquaculture to treat the salmon for sea lice, a type of parasitic copepod. The results of this study indicate that these pesticides are toxic to the free-living T. californicus at very low dosage levels when used individually. When the pesticides are used in combination, the mortality rate is similar to when they are used individually, but there is an additional paralytic effect on the animals. All animals exposed to any of the pesticide mixtures used in this study were irreversibly paralyzed and therefore effectively killed. This indicates a strong need for more testing on the combined impacts of pesticides and increased regulatory action for salmon aquaculture.

Keywords: Pesticides, paralysis, pemamectin benzoate, cypermethrin, aquaculture, salmon, sea lice, Tigriopus californicus

ishery stocks worldwide continue to Salmon are farmed in 24 countries, decline due to increased fishery with Norway, Scotland, Chile and Canada F pressure and unsustainable leading the way as producers (Burridge et management practices (Naylor et al., 2000). al., 2010). In the United States there has One potential solution to the decline in fish been an increase in these farms along the species has been aquaculture, where fish and Pacific Northwest and the coast of Maine shellfish are farmed at high densities and (Shaw et al., 2008). Salmon farms are harvested in large numbers to be brought typically located just offshore and consist of directly to market. Landings from a number of large circular nets known as aquaculture have increased rapidly since the floating pens. Juvenile fish are placed into turn of the century and Bush et al. (2013) the floating pens to continue to grow to state that aquaculture now provides close to market size. Due to the high density of the 50% of the world’s seafood supply. One fish in a relatively small space, parasites and fishery in particular where aquaculture has diseases are a concern (Morales-Serna et al., increased dramatically is the salmon fishery, 2014). In response, salmon farmers have where over 1.8 million metric tons are increased their use of pesticides to combat produced annually (Burridge et al., 2010). this issue. Pesticide use in the marine

87 Published by Scholarship & Creative Works @ Digital UNC, 2016 1 Ursidae: The Undergraduate Research Journal at the University of Northern Colorado, Vol. 5, No. 3 [2016], Art. 7 PESTICIDE EFFECTS ON A MARINE COPEPOD

environment has been increasing copepods occur. Emamectin benzoate significantly in the past few decades due to a increases the permeability of muscle and concomitant increase in these aquaculture nerve membranes to chlorine (Durkin, activities, but the research to examine the 2010), thus interfering with effects and determine safe levels of these neurotransmission, causing irreversible chemicals has been lagging (Morales-Serna paralysis (Anderson, et al. 2009). et al., 2014). Emamectin benzoate has also been shown to The purpose of this study is to change life cycle patterns, and egg examine two pesticides commonly used in production in aquatic invertebrates (Durkin, salmon farming (cypermethrin and 2010). Although T. californicus is not the emamectin benzoate) to first determine their target organism for this pesticide, it is likely individual impact and then their impact to be exposed to the chemical if it is used when used together. The organism used to widely to treat sea lice in salmon farms examine the toxicity of these pesticides is along the western US coastline. the marine zooplanktonic copepod Tigriopus Cypermethrin, another pesticide californicus. Copepods such as T. commonly used in salmon farming, is a californicus are primary consumers and are synthetic pyrethroid that has traditionally considered the most common metazoans in been used in terrestrial agriculture to control the ocean (Feinberg and Dam, 1998). They pests such as moths in cotton farms, fruit are often used in toxicity studies due to their and vegetable crops. It has also been used as short life cycle and amenability to laboratory spot treatments for pests in stores, schools, conditions. Tigriopus californicus is found and office buildings (Meister, 1992). in marine tidal pools throughout the Western However, cypermethrin has recently been coast of the United States, from Baja used to control sea lice in salmon farming in California to Alaska (Peterson, et al., 2013), a formulation called Excis (Gowland et al., and is common in areas where salmon 2002). The pesticide is added directly to the farming is already occurring or proposed. water in and around the sea cage where the We anticipate that this increase in salmon salmon are held. Although cypermethrin is aquaculture will lead to an increase in the not readily absorbed into water, it has a high amount of pesticides discharged into marine tendency to bind to particulate matter in the tidal pools. water (Gowland et al., 2002) and remain Both pesticides examined in this suspended for some time before settling into study, emamectin benzoate and the sediments. (Muir, 1985; Agnihotri, 1986; cypermethrin, are typically used to treat sea Ayad, 2011). In invertebrates, cypermethrin lice, which are parasitic copepods in the is a neurostimulant that causes prolonged family Calgidae (Morales-Serna et al., contraction of muscles and interrupts normal 2015). Emamectin benzoate is often mobility (Willis, 2004). It has been found to administered orally, which allows the fish to damage the sodium channels, causing them have a direct source of the pesticide, without to remain open for extended periods of time releasing it directly into the water. This form (Jones, 1990). This is why it has been used of administration causes no ill effects on the as an effective pesticide for sea lice, and fish (Stone et al, 2002); however, it will be why there is concern about its impact on excreted in their feces (Sevatdal et al, 2005). non-target invertebrate species. This allows the pesticide to enter the marine The Environmental Protection environment, and can potentially move into Agency, under the Federal Environmental tidal pools where species of non-parasitic Pesticide Control Act (FEPCA), requires

88 http://digscholarship.unco.edu/urj/vol5/iss3/7 2 Schultz and Kelly: Joint Action Effects of Emamectin Benzoate and Cypermethrin on th PESTICIDE EFFECTS ON A MARINE COPEPOD

that toxicity testing be conducted before common response in the target species. pesticides can be released into the However, in this model, it is more difficult environment (FEPCA, 1972). These tests are to predict how the interaction of the typically done by examining the effects of chemicals will affect the target species, and one pesticide at a time by measuring the what concentrations are needed to reach this dose of the pesticide that is lethal to 50% of threshold. the test animals (LD50) in a defined time One mode of testing the joint action period, often 48 or 96 hours. While this type model is to use the toxic unit approach, of assay provides useful information about where the toxic unit (TU), in this case the the overall impact of the pesticide, it does LD50, is first determined and then the not reflect the reality of pesticide exposure chemicals are mixed in ratios of their toxic in the natural environment. Organisms are units to assess their impacts on the target rarely exposed to only one chemical at a species. This can be best conceptualized by time, and in fact are often exposed to the equation XTUA + YTUB = 1 TU(A-B), multiple chemicals simultaneously. The where A and B are the two pesticides, x and standard LD50 tests do not reveal how these y are the proportional toxic units of each mixtures of chemicals will impact the pesticide, and TU is the LD50 for each species exposed to them. Because of this, it (Forget et al., 1999). If x + y < 1, the is crucial to determine how the interaction of chemicals would be considered synergistic, chemicals can affect organisms. There are thus, you have a lower TU when the mathematical models that have been develop chemicals are added in a mixture. If the two to attempt to predict the effects of the pesticides are shown to be synergistic in interactions of various chemicals (DeMarch their effect, their overall impact is more than 1987), but there have been few experiments simply the addition of the two. If x + y = 1, conducted. then the chemicals are considered additive, Because this study focuses on the which is indicative of the concentration mixture of the two pesticides, it was addition model. In this case, it can be determined that we would be examining assumed that the toxicity of each pesticide aspects of the joint action model (Bliss compound can be summed to determine the 1939). The joint action model examines maximum acceptable dosage; i.e. one can how a mixture of chemicals can impact a combine the LD50 of each to arrive at an particular species. Typically, if the mode of acceptable level. If x + y >1, the chemicals action of the chemicals is similar the are considered antagonistic, in that you have concentration addition model applies to add more to get the same response as (Anderson &Weber, 1975). In this case, the 1TU, so the chemicals must be in some way effect of the mixture would be the same as counteracting one another. The goals of this simply adding the same amount of each study are to (1) to determine the toxicity individually. However, if the mode of level of cypermethrin and emamectin action of the chemicals is different, it is benzoate individually, and (2) to investigate likely that the effect on the organism will whether combined exposure more closely match the response addition model matches the concentration addition model or (Anderson &Weber, 1975). Here, it response addition model and if the latter, are assumed that each toxicant will have a the pesticides synergistic or antagonistic. threshold concentration before it causes a

89 Published by Scholarship & Creative Works @ Digital UNC, 2016 3 Ursidae: The Undergraduate Research Journal at the University of Northern Colorado, Vol. 5, No. 3 [2016], Art. 7 PESTICIDE EFFECTS ON A MARINE COPEPOD

Methods dissolve the pesticides, the impact of DMSO alone was also examined.

Test Chemicals Joint Action Effects A 95% pure formulation of cypermethrin (Sigma Aldrich) was dissolved To determine if the effects of the in dimethyl sulfoxide (DMSO) to produce a pesticides were additive, the two pesticides stock solution that was diluted to final were combined at 50% of their respective concentrations of 5 µg/L, 2.5 µg/L, .5 µg/L, LD50 values (50TUemamectin benzoate + and .25 µg/L. These concentrations were 50TUcypermethrin). Final concentrations were based on those of Willis and Ling (2004), 1.28µg/L emamectin benzoate and 1.3µg/L who examined the effect of cypermethrin on cypermethrin. Thus, if the pesticides were other marine copepods. This process was additive, this should result in 50% mortality repeated with a 95% pure formulation of 48 hours after exposure. Again, 12 emamectin benzoate (Sigma Aldrich), which ovigerous females were individually placed was diluted to the same concentrations. into the wells of a 24-well plate. At the end Both chemicals will persist in solution for a of the 48 hour exposure, animals were time period longer than the 48 hour toxicity examined under a stereomicroscope for their tests that were conducted in the study response to gentle prodding. These (Anderson et al., 2009; Muir et al,, 1995) experiments were repeated twice. To further elucidate the impact of the combined toxicants, and determine if there was a threshold response, copepods were Acute Toxicity Testing exposed to varying levels of the pesticides. Tigriopus californicus (Carolina The first set of experiments exposed Biology Supply) cultures were maintained in copepods to 75% of the LD50 concentration 35ppt salinity at 20oC (± 1.5) in a 12:12 light for emamectin benzoate and 25% of the dark cycle and fed Tetraselmis sp. (Carolina LD50 concentration of cypermethrin Biological Supply) to excess. To determine (75TUemamectin benzoate + 25TUcypermethrin), for the LD50 values at 48h, 12 ovigerous final concentrations of 1.92µg/L of females were placed individually into the emamectin benzoate and 0.065µg/L of wells of a 24-well plate and fed ad libitum cypermethrin. Additionally, experiments for the duration of the exposure. For each were conducted 25% of the LD50 toxicant concentration, at least three trials concentration of emamectin benzoate and were conducted for a total of 36 ovigerous 75% of the LD50 concentration of females per concentration. During the cypermethrin (25TUemamectin benzoate + toxicity testing, animals were maintained at 75TUcypermethrin) for final concentrations of the same temperature, light cycle and 0.64µg/L of emamectin benzoate and feeding regimen as the stock cultures. 1.95µg/L of cypermethrin. Three trials with Animals were examined at 48 hours and 12 ovigerous females (for a total of 36 tested for their ability to move in response to animals for each mixture) were conducted gentle prodding with a probe. Any animal for each mixture and were examined for that exhibited a discernable response was mortality after 48 hours using the methods considered to be alive at the end of the described above. exposure. Due to the need to use DMSO to

90 http://digscholarship.unco.edu/urj/vol5/iss3/7 4 Schultz and Kelly: Joint Action Effects of Emamectin Benzoate and Cypermethrin on th PESTICIDE EFFECTS ON A MARINE COPEPOD

Recovery within 30 minutes of exposure. An animal was considered paralyzed when it was only To determine if the effects of the capable of small twitching movements and combined pesticides were reversible, exhibited no swimming or locomotory recovery experiments were conducted. ability, even when prodded with a probe. At Following exposure to the 50:50 mixture, all the end of the 48 hour experiment, an copepods that were still alive were placed average of 47% of the animals were dead individually into wells containing only and all animals that remained alive were 35ppt seawater. After 2 hours and then again paralyzed (Table 1). Similar results were at 24 hours, animals were examined under a found with the other mixtures, in that all stereomicroscope to determine if they swam copepods were either paralyzed or dead at away in response to prodding with a probe. the end of the 48 hour exposure; however, the overall mortality decreased significantly Probit Analysis in comparison to the mortality of the 50:50 To calculate LD50 values for each exposure (Table 1 and Figure 2). concentration, Probit Analysis was used. Recovery Probit analysis transforms a curved dose- response curve to a straight line that can be There was no evidence of recovery used to determine the concentration that is from paralysis for any animals exposed to lethal to 50% of the population. Abbot’s the mixture of pesticides. This was true for correction was used to correct for the both the 2 hour and 24 hour recovery times. mortality that was seen in the DMSO control It appears that the paralysis caused by the (Rosenheim and Hoy, 1989) interaction of these two pesticides is not recoverable.

Results Discussion LD50 The data presented in this study Results from the acute toxicity tests indicated that both chemicals exhibit high of individual toxicants can be seen in Figure toxicity at relatively low concentrations, 1. From these data, Probit Analysis, with even when copepods are exposed to a single Abbott’s correction for the effect of the pesticide. These data have important DMSO, was used to determine the LD50 implications for the regulations of these values for each pesticide. The LD50 for chemicals. Presently, the recommended emamectin was 2.56 µg/L and the LD50 acute exposure limit for cypermethrin in value for cypermethrin was 2.6µg/L. These drinking water is 940 µg/L (US EPA, 2006), values appear slightly different than those in and the LD50 value for cypermethrin Figure 1 due to correction that was needed exposure to T. californicus found in this to account for the low levels of mortality study is 2.6 µg/L, a far lower level. There that occurred in the DMSO controls. are no comparable drinking water data Joint Action Effects available for emamectin benzoate, as this pesticide is still under review by the EPA When copepods were placed into the and is not considered an impairment of 50:50 mixture of emamectin benzoate and bodies of water under the Clean Water Act cypermethrin, all animals exhibited paralysis (US EPA 2011). However, it is added

91 Published by Scholarship & Creative Works @ Digital UNC, 2016 5 Ursidae: The Undergraduate Research Journal at the University of Northern Colorado, Vol. 5, No. 3 [2016], Art. 7 PESTICIDE EFFECTS ON A MARINE COPEPOD

100 90 80

60 Cypermethrin 50 40 30 Emmamectin Percent Survival Percent 20 Benzoate 10 0 20 2 0.4 0.04 0

Concentration (µg/L)

Figure 1 Survival of Tigriopus californicus after 48 hours exposure to either cypermethrin or emamectin benzoate (n = 36 for each toxicant at each concentration).

Figure 2 The effect of pesticide mixtures on mortality of T. californicus. Ratios refer to the proportion of the LD50 value for each pesticide. Asterisk indicates a significant difference (p<0.05) in mortality rate of the 50:50 ratio as compared to the others.

92 http://digscholarship.unco.edu/urj/vol5/iss3/7 6 Schultz and Kelly: Joint Action Effects of Emamectin Benzoate and Cypermethrin on th PESTICIDE EFFECTS ON A MARINE COPEPOD

Table 1 The effect of pesticide mixtures on T. californicus, including rates of mortality and paralysis. Ratios refer to the proportion of the LD50 value for each pesticide. Pesticide Ratio (Emamectin Average % Average % Total % Benzoate: Cypermethrin) Mortality Paralyzed Affected

50:50 42.7 55.3 100

75:25 16.67 83.33 100

25:75 11.11 88.89 100

directly to water during treatments of recoverable. Any copepod that is paralyzed salmon in salmon farms and levels of and unable to swim or feed will not be 4.16X10-6 µg/L have been recorded as long capable of survival for more than a few as 8 days after treatment (Willis and Ling, days, and therefore the pesticide has 2004). effectively killed it, but will require a longer While the data from the single time to do so. Thus is appears that these exposure experiments indicate a need for pesticides act synergistically when presented more studies on the impact of these in a mixture. pesticides in the marine environment, the To further examine the response results from the mixture studies are even addition model and look for threshold more concerning. When examining the joint concentrations, mixtures of 75% of the action of these two toxicants, it appears that emamectin benzoate LD50 to 25% of the they do not fit well into either the cypermethrin LD50 and 25% of the concentration addition or the response emamectin benzoate LD50 to 75% of the addition model. Because both pesticides cypermethrin LD50 were tested. It appears affect the nervous system of the target that the mortality rate declines in these organism, and therefore have a similar mode mixtures, but the overall paralysis effects of action, one could predict that they would remain the same, resulting in an eventual follow the concentration addition model. If 100% mortality. So, while there appears to that were the case, the 50:50 mixture of be a threshold effect of these toxicants in pesticides should have resulted in a 50% their ability to cause mortality, there is no mortality rate at the end of the 48h exposure. threshold effect in their paralytic actions. The data indicate that the mortality rate was We propose that this is evidence of the 47.2%, which is very close to the expected response addition model, in that these value, but mortality rate alone does not paint toxicants have slightly different effects on the full picture of the effect of these the neuromuscular system of the target toxicants. At the end of the 48h exposure, species, and therefore the result is paralysis all copepods were dead or were completely rather than mortality. The response addition paralyzed and this paralysis was not model predicts that the interaction of two

93 Published by Scholarship & Creative Works @ Digital UNC, 2016 7 Ursidae: The Undergraduate Research Journal at the University of Northern Colorado, Vol. 5, No. 3 [2016], Art. 7 PESTICIDE EFFECTS ON A MARINE COPEPOD

toxicants can produce unexpected effects in References the target species when combined, rather than simply an increase in mortality, which Agnihotri, N.P. Jain, HK. Gajbhiye, VT. (1986). Persistence of some synthetic pyrethroid is what our data show. insecticides in soil, water, and sediment, If use of these pesticides becomes Part I. Journal of Entomological Research. widespread, it could potentially devastate 10:147-151 the native marine copepod community, Anderson, B., Doelling, P., & Hetrick, J. (2009). which in turn will have a significant impact Section 3 Request for a New Use of the on the ecosystem as a whole due to the role Insecticide Emamectin Benzoate. United that these organisms play in the marine food States. Environmental Protection Agency. chain. A variety of tidepool fish, several Office of Pesticide Programs. Washington, genera of anemones, and some grapsid and DC. pagurid crabs are known predators of T. Anderson, P.D., & Weber, L.J. (1975). The toxicity californicus specifically (Dethier, 1980), and to aquatic pollutants of mixtures of heavy copepods in general are a major component metals. Proceedings, Heavy Metals in the of their ecosystem. They are significant Environment, Toronto, ON, Canada, grazers of the phytoplankton community, are October 27-31, pp 933-954. important prey for pelagic fish species, and Ayad, M., Fdil, M., & Mouabad, A. 2011. Effects of play a critical role in the microbial loop of cypermethrin (pyrethroid insecticide) on the the world’s oceans (Turner 2004). Because valve activity behavior, byssal thread of their role is prey items, bioaccumulation formation, and survival in air of the marine mussel Mytilus galloprovincialis. Archives is a potential threat, but neither emamectin of Environmental Contamination and benzoate nor cypermethrin appear to Toxicology 60:462- 470. bioaccumulate in the food chain (Chukwudebe et al., 1996). However, there Bliss, C.I. 1939. The toxicity of poisons applied jointly. Ann. Appl. Biol. 26: 585-615. is preliminary evidence that cypermethrin can accumulate in fish (Corcellas et al., Boudin, M. 2003. US Pub. Int. Res. Group vs. 2015) and mussel (Gowland et al., 2002) Atlantic Salmon of ME, 257 F. Supp. 2d 407 tissue following exposure to cypermethrin in Dist.Court, D. Maine 2003 the water column. This has implications for Burridge, L., Weis, J.S., Cabello, F., Pizarro, J., & the combined use of these pesticides in Bostick, K. (2010). Chemical use in salmon salmon farming and their impact on non- aquaculture: A review of current practices target species. The toxicity of both of these and possible environmental effects. pesticides to copepods at the low levels Aquaculture 306: 7-23. found in the current study suggest that Bush, S.R., Belton, B., Hall, D., Vendergeest,P., copepods may act as an effective indicator Murray, J.F., Ponte, S., Oosterveer, P., species for the health of the ecosystem as a Islam, M.S., Mol, A.P.J., Hatanaka, M., whole. Based on the data presented here we Krujssen, F., Ha, T.T.T., Little, D.C., & Kusumawati,R. (2013). Certify sustainable argue that more testing needs to be done to aquaculture? Science 341: 1067-1068. determine the impact of these pesticides, especially their use in combination, on other Chukwudebe, A.C., Andrew, N., Drottar, K., Swigert, aquatic species that are likely to be prevalent J., & Wislocki, P.G. (1996). Bioaccumulation potential of 4 “-epi- in areas where salmon farming occurs. (Methylamino)-4”-deoxyavermectin B1a Benzoate (emmamectin benzoate) in bluegill sunfish. Journal of Agriculture and Food Chemistry. 44(9): 2894-2899.

94 http://digscholarship.unco.edu/urj/vol5/iss3/7 8 Schultz and Kelly: Joint Action Effects of Emamectin Benzoate and Cypermethrin on th PESTICIDE EFFECTS ON A MARINE COPEPOD

Corcellas, C., Eljarrat, E., & Barcelo, D. (2015). First (Copepoda: Caigidae) parasitic on marine report of pyrethroid bioaccumulation in wild fishes with commercial and aquaculture river fish: A case study in Iberian river importance in Chamela Bay, Pacific coast of basins (Spain). Environmental International. Mexico by using morphology and DNA 75: 110-116. barcoding with description of a new species of Caligus. Parasitology International. 63: DeMarch, B.G.E. (1987). Simple similar action and 69-79. independent joint action: Two similar models for the joint effects of toxicants Muir, D.C.G. Rawn, G.P. Townsend, B.E. Lockhart, applied as mixtures. Aquat. Toxicol. 9: 291- & W. Greenhalgh, R. (1985). 304. Bioconcentration of cypermethrin, deltamethrin, fenvalerate, and permethrin by Dethier, M.N. (1980). Tidepools as refuges: Chironomus tentans larvae in sediment and Predation and the limits of the harpacticoid water. Environmental Toxicology and copepod Tigriopus californicus. Journal of Chemistry. 4, 51-61 Experimental Marine Biology and Ecology. 42: 99-111. Naylor, R.L., Goldburg, R.J., Primavera, J.H., Kautsky, N., Beveridge, M.C.M., Clay, J., Durkin, P. (2010). Emamectin benzoate: human Folke, C., Lubchenco, J., Mooney, H. & health and ecological risk assessment. Torell, M. (2000). Effect of aquaculture on United States. USDA Forest Service. world fish supplies. Nature 405: 1017-1024. Syracuse Environmental Research Associates. Atlanta, Georgia. Peterson, D., Kubow, K., Connolly, M., Kaplan, L., Wetkowski, M., Leong, W., Phillips, B., & Federal Environmental Pesticide Control Act of Edmands, S. (2013). Reproductive and 1972, 7. U.S.C. §§ 135 (1972). phylogenetic divergence of tidepool copepod populations across a narrow Feinberg, L.R. & Dam, H.G. (1998). Effects of diet geographical boundary in Baja California. on dimensions, density and sinking rates of Journal of Biogeography, 40, 1664-1675 fecal pellets of the copepod Acartia tonsa. Mar. Ecol. Prog. Ser., 175: 87–96. Rosenheim, J.A., & Hoy, M.A. (1989). Confidence Intervals for the Abbott’s Formula Forget, J., Pavillon, J, Beliaeff, B., & Bocquene, G. Correction of Bioassay Data for Control (1999). Joint action of pollutant Response. Journal of Economic combinations (pesticides and metals) on Entomology, 82(2), 331-335. survival (LC50 values) and acetylcholinesterase activity of Tigriopus Sevatdal, S., Magnusson, A., Ingebrigtsen, R., & brevicornis (Copepoda, Harpacticoida). Env. Horsberg, T.E. (2005). Distribution of Toxicol. Chem. 18: 912-918. emamectin benzoate in Atlantic salmon (Salmo salar L.). Journal of Veterinary Gowland, B., Webster, L. Fryer, R., Davies, I., Pharmacological Therapeutics, 28, 101-107. Moffat, C., & Stagg, R. (2002). Uptake and effects of the cypermethrin-containing sea Shaw, S.D., Berger, M.L., Brenner, D., Carpenter, lice treatment Excis in the marine mussel D.O., Tao, L. Hong, C., & Kannan, K., Mytilus edulis. Environmental Pollution (2008). Polybrominated diphenyl ethers 120: 805-811. (PBDEs) in farmed and wild salmon marketed in the Northeastern United States. Jones, D. (1990). Environmental fate of Chemosphere 71: 1422-1431. cypermethrin. Department Monitoring & Pest Management. Sacramento, CA. Stone, J., Roy, W. J., Sutherland, I. H., Ferguson, H.W., Sommerville, C., & Endris, R. (2002). Meister, R.T. (1992). Farm Chemicals Handbook ’92. Safety and efficacy of emamectin benzoate Meister Publishing Company, Wiloughby, administration in- feed to Atlantic salmon, OH. Salmo salar L., smolts in freshwater, as a preventative treatment against infestations of Morales-Serna, FN., CD Pinacho-Pinacho, S. Gomez, sea lice, Lepeophtheirus salmonis. & GP de Leon. 2014. Diversity of sea lice Aquaculture 210: 21-34.

95 Published by Scholarship & Creative Works @ Digital UNC, 2016 9 Ursidae: The Undergraduate Research Journal at the University of Northern Colorado, Vol. 5, No. 3 [2016], Art. 7 PESTICIDE EFFECTS ON A MARINE COPEPOD

Taylor, W.W., Schechter. M.G., & Wolfson, L.G. US Environmental Protection Agency, (2011). (2007) (eds). Globalization: Effects on Emamectin Benzoate Summary Document. Fisheries Resources. Cambridge University Office of Pesticide Programs, Washington, Press, Cambridge. DC.

Trask, J., Harbourt, C., Miller, P., Cox, M., Jones, R., US Environmental Protection Agency. (2006). Hendley, P., & Lam, C. (2014). Washoff of Reregistration Eligibility Decision for cypermethrin residues from slabs of external Cypermethrin. Office of Pesticide Programs, building material surfaces using simulated Washington, DC. rainfall. Environmental Toxicology and Chemistry 33: 302-307. US Environmental Protection Agency. Jan. 3, 1989. Pesticide Fact Sheet Number 199: Turner, J.T. (2004). The importance of small Cypermethrin. Us EPA, Office of Pesticide planktonic copepods and their roles in Programs, Registration Div., Washington, pelagic marine food webs. Zoological DC. Studies. 43(2): 255-266. Willis, K., & Ling, N. (2004). Toxicity of the US Department of Agriculture, Soil Conservation aquaculture pesticide cypermethrin to Service. 1990 (Nov.). SCS/ARS/CES planktonic marine copepods. Aquaculture Pesticide Properties Database: Version 2.0 Research 35: 263-270. (Summary). USDA- Soil Conservation Service, Syracuse, NY.

96 http://digscholarship.unco.edu/urj/vol5/iss3/7 10