Evaluation of acute and chronic toxicity of selected compounds in higher plants using cell culture

Summary The feasibility of using plant cell culture to measure toxicity was determined by investigating the toxicological effects of three chemical compounds, allyl alcohol, propargylglycine, and cadmium chloride, on cell cultures ofCatharanthus roseus G. Don (Madagascar periwinkle). Suspension cultures ofC. roseus were maintained in modified B5 medium and transferred every 5 d. Five-day-old cell cultures were exposed to various concentrations (10,3,1,0.3,0.1,0.03,0.01,0.003,0.001,0.0003,0.0001, 0.00003, and 0.0 mM) of the toxicants in both acute and chronic toxicity tests. In the acute test, cells were exposed to the toxicant for 24 h, washed three times with sterile medium, and plated in petri plates with an equal volume of 1.4% agar medium. Cells in the chronic test were plated with an equal volume of 1.4% agar medium containing various concentrations of the toxicant. Cells were incubated 28 d at 30°C in the dark. The colonies were counted and the results plotted as percent survival versus toxicant concentration. The results indicate, at the concentrations tested, thatC. roseus assay may be feasible in that it fulfills the criteria for a practical assay (e.g., rapid, simple, quantifiable, and reproducible). This work was submitted to the faculty of Miami University in partial fulfillment of the requirements for the degree of Master of Environmental Science, Institute of Environmental Sciences.     

Toxicity evaluation of 2-hydroxybiphenyl and other compounds involved in studies of fossil fuels biodesulphurisation

The acute toxicity of some compounds used in fossil fuels biodesulphurisation studies, on the respiration activity, was evaluated by Gordonia alkanivorans and Rhodococcus erythropolis. Moreover, the effect of 2-hydroxybiphenyl on cell growth of both strains was also determined, using batch (chronic bioassays) and continuous cultures. The IC₅₀ values obtained showed the toxicity of all the compounds tested to both strains, specially the high toxicity of 2-HBP. These results were confirmed by the chronic toxicity data. The toxicity data sets highlight for a higher sensitivity to the toxicant by the strain presenting a lower growth rate, due to a lower cells number in contact with the toxicant. Thus, microorganisms exhibiting faster generation times could be more resistant to 2-HBP accumulation during a BDS process. The physiological response of both strains to 2-HBP pulse in a steady-state continuous culture shows their potential to be used in a future fossil fuel BDS process.     

Acaricidal activity of spinosad and abamectin against two-spotted spider mites

Spinosad is a bioinsecticide with a high degree of selective toxicity towards insects of different orders, but its toxicity towards the two-spotted spider mite (TSSM), Tetranychus urticae Koch (Acari: Tetranychidae) is under debate. In this study, we compared the acaricidal properties of spinosad with the commercial bioacaricide abamectin on the life stages of TSSM. Adulticide and ovicide bioassays were performed on a susceptible laboratory strain using direct spraying of leaf disks with five rates of spinosad (20, 25, 30, 35 and 40 mg/l), five rates of abamectin (0.125, 0.25, 0.5, 1 and 2.5 mg/l), sublethal concentrations or a combination of spinosad and abamectin. Both adulticidal and ovicidal effects of spinosad against T. urticae in the laboratory were apparent, based on morality rates of the adults, reduction of female fecundity and death of offspring. Abamectin was also found to significantly reduce female fecundity and killed offspring when applied directly on the eggs. Interestingly, sublethal concentrations of spinosad reduced female fecundity stronger than abamectin. When a mixture of spinosad and abamectin was applied at LC_50, mortality was 74%, fecundity reduction was comparable to abamectin alone and egg hatching rate was lower than by either compound alone. In conclusion, spinosad was more harmful than abamectin for TSSM life stages and the combined application is recommended.     

Recovery Patterns of Moina Macrocopa Exposed Previously to Different Concentrations of Cadmium and Methyl Parathion: Life-Table Demography and Population Growth Studies

In most toxicity studies using Cladocera, bioassays are routinely done to determine median lethal concentration (LC₅₀) or the responses to sublethal exposure. However, information on the patterns of recovery of cladocerans exposed to different concentrations of toxicants is scarce. This is important because cladocerans exposed to toxicants for a short duration may later recuperate under favourable conditions. Using the life table demographic and population growth, the present study was conducted to evaluate the recovery patterns of Monia macrocopa exposed to five different concentrations (0, 25, 50, 75 and 100% of 24 h LC₅₀ for CdCl₂ or methyl parathion) and then returned to toxicant-free medium containing alga (Chlorella vulgaris) at low (0.25 × 10⁶cells ml^–1), medium (0.5 × 10⁶cells ml^–1) or high (1 × 10⁶cells ml^–1) levels. We measured selected life history variables such as average lifespan, life expectancy at birth, gross and net reproductive rates, generation time and the rate of population increase. Results indicated that regardless of food concentration, surviving individuals of M. macrocopa exposed to a median lethal concentration did not recover. The effect of food level was significant at 25 and 50% of the median lethal concentration for cadmium or methyl parathion. Age-specific fecundity curves showed that exposure to either toxicant for a duration as short as 24 h at one-fourth of the LC₅₀ showed reduced output of offspring, especially at a lower food level. At and above exposures of 0.037 mgl^–1 of methyl parathion, no reproduction occurred. The highest gross and net reproductive rates (127 and 55 offspring female^–1) were obtained in controls at the high (1 × 10⁶ cells ml^–1) algal food level. The rate of population increase obtained from life table data was around 0.7 per day in controls but decreased when exposed to toxicant concentrations. The rates of population increase per day derived from population growth data varied from 0.22 to 0.33 per day for the controls, depending on the food levels.     

Effect of food concentration on the chronic toxicity of sodium dodecyl sulphate to Daphnia magna

Food concentration supplied during subchronic and chronic toxicity tests, is one of the most important factors that might influence the response of test organisms to toxicants. The green microalga Scenedesmus incrassatulus was used as food for the cladoceran Daphnia magna, in a chronic toxicity test with the toxicant sodium dodecyl sulphate (SDS). Test concentrations were 0.625, 1.25, and 2.5 mg l^-1 of SDS, equivalent to 1/40, 1/20, and 1/10 of the average 48-h LC50 previously determined. Food concentrations were 9.5, 19, and 38 mg l^-1 (dry wt.). Survival and reproduction were recorded, and the data were analyzed using a life-table approach. After 55 days, the main findings were as follows:

Hypothesis formulation and testing in aquatic bioassays: a deterministic model approach

The paper examines the significance of toxicant kinetics information obtained from aquatic toxicity bioassays and bioconcentration tests. The data, bioconcentration kinetics and acute mortality versus exposure-duration information for juvenile American flagfish (Jordanella foridae) exposed to 1,4-dichlorobenzene, are interpreted in terms of a one-compartment, first-order kinetics model. The output of the model is used to formulate a testable hypothesis regarding the comparison of toxicant kinetics derived from both bioconcentration test exposures and toxicity bioassays. The model's estimates of the toxicant body burden attained at mortality are compared with theoretical and observed body burdens from literature sources. The use of a simple, deterministic residue-based, one-compartment, first-order kinetics model to evaluate existing data, as well as to formulate hypotheses to direct experimental designs, is examined.     

Cultivation of the heart urchin Echinocardium cordatum and validation of its use in marine toxicity testing for environmental risk assessment

To study environmental risk assessment, echinoderms provide a useful model for ecotoxicological testing. However, limited knowledge of the life history of field collected heart urchins is a problem and the use of cultured urchins has been investigated here. The present study describes a culture method for the heart urchin Echinocardium cordatum under controlled laboratory conditions, providing organisms with a low biological variation. Based on our optimized growth protocol both larvae and juveniles have a growth rate comparable to E. cordatum in the wild. The toxicological response of cultured and field-collected E. cordatum was compared in standard saltwater toxicity bioassays. Using ammonium chloride as a water-soluble reference toxicant, mean 96 h LC₅₀ values for cultured heart urchins versus field collected animals were 37.4 ± 7.6 mg NH₄+/l (n = 5) versus 22.5 ± 4.9 mg NH₄+/l (n = 19), respectively. Additional toxicity experiments with tributyl tin (TBT) spiked sediments revealed 14d LC₅₀ values of 1,242 (95% confidence interval 986–1,564) and 964 (95% confidence interval 843–1,102) µg Sn/kg dw respectively in cultured and field collected E. cordatum. From this it was concluded that cultured heart urchins are less sensitive to TBT than field collected E. cordatum. Furthermore in whole sediment toxicity tests, survival of cultured sea urchins was higher or at least similar to that of field collected E. cordatum. The increased sensitivity of field urchins compared to cultured urchins in various toxicity tests may be due to multiple environmental stressors reducing their overall performance. Overall it was demonstrated that the use of cultured E. cordatum provides a significant advance for urchin-based bioassays for marine environmental toxicity testing, resulting in a more homogeneous, vital population with experimental data displaying reduced variability.     

Identification of developmental toxicants using the Frog Embryo Teratogenesis Assay-Xenopus (FETAX)

Because growth and development are processes sensitive to the action of many chemicals, bioassays that screen for developmental toxicants may be more indicative of chronic effects than acute toxicity assays. FETAX is a 96 h whole embryo static renewal test employing the embryos of the frog Xenopus laevis. Endpoints are mortality, malformation and growth. Because of the frog's fecundity, its extensive use in basic research and the ability to obtain embryos year-round, it is an ideal organism to use in screening for developmental toxicants. By validating using known mammalian teratogens and the use of rat liver microsomes to stimulate mammalian metabolism, we have extended the use of the system for the prescreening of human developmental toxicants. In past validation work, we have correctly identified the teratogenicity of 15 to 17 compounds used in validation for a predictive accuracy of approximately 88%. In the present study, the ability of FETAX to detect developmental toxicants in groundwater samples taken from an industrial waste dump was evaluated. FETAX showed that it was sensitive enough to detect developmental toxicants in samples without prior concentration. In some samples, less than half the LC50 concentration was required to cause significant malformation. In some cases, a dose-response curve was not obtainable but the test results nonetheless indicated some developmental toxicity. The results of this study indicate that it is necessary to routinely screen for developmental toxicants when establishing water quality criteria for the preservation of species and for human health.     

Acute and chronic toxicity of the pesticide methyl parathion to the rotifer Brachionus angularis (Rotifera) at different algal (Chlorella vulgaris) food densities

Methyl parathion is a commonly used insecticide in Mexico to eradicate insect pests. We evaluated the effects of this insecticide on rotifer B. angularis using both acute and chronic toxicity tests. Median lethal concentration (LC₅₀) of methyl parathion for B. angularis for a 24-h bioassay in the presence and absence of an algal diet was derived. Elevated LC₅₀ due to the survival of a greater number of test individuals in the presence of food was observed. Regardless of the toxicant concentration, population growth curves of the animals maintained at the low food level (0.75×10⁶ cells ml^-1) had a longer lag phase than those at the high food level (1.5×10⁶ cells ml^-1). Regardless of food level, an increase in the toxicant concentration in the medium resulted in decreased population growth. The lowest peak population density (50 ind. ml^-1) was observed at the highest toxicant concentration and the lower food level. The highest population density (200 ind. ml^-1) was observed in the controls at high food level. The rates of population increase per day (r) in the controls were higher (from 0.14 to 0.37 depending on the food level). Irrespective of food level, there was a decrease in the r values with increasing pesticide concentration in the medium. In order to detect the effect of population density on the growth rates in relation to the toxicant stress, we plotted the daily growth rate against initial density for the entire duration of the experiment. We observed the existence of a significantly inverse relation at all treatments except at the low food level and high toxicant concentrations (0.625 and 1.25 mg l^-1). We discuss the role of algae in the toxicity of methyl parathion to zooplankton.     

<Emphasis Type="Italic">Dunaliella salina</Emphasis> (Chlorophyta) as a Test-Object for Assessment of Detergent Pollution of a Marine Environment

The dynamics of the number of the microalga Dunaliella salina, depending on the age of the matrix culture, the number of cells, and the time of toxicant administration in the culture medium and on oxygen production as a parameter of the functional condition of the dunaliella, was studied in solutions of sodium dodecylsulfate (SDS) containing 0.1, 1, and 10 mg/l of the detergent. The parameters at which the application of the considered test-object allowed determination of the most exact information on the environment quality and toxicity of substances were determined. It was shown that the SDS in concentrations of 0.1 and 1 mg/l did not affect significantly the growth of the microalga, and an inhibiting effect was recorded at a toxicant content of 10 mg/l.Original Russian Text Copyright ¢ 2005 by Biologiya Morya, Markina, Aizdaicher.     

Algal bioassays to determine toxicity of metal mixtures

The shortcomings of many established toxicity criteria for metals have resulted from a simplification of bioassays performed with single metals. A more comprehensive approach was needed to diagnose the effects of metal mixtures on aquatic organisms. Using Chlorella, Ankistrodesmus and Scenedesmus as test organisms, we experimented on a number of factors which affected metal toxicity bioassays. These included metal interactions, algal competitions, species sensitivities, the ratio of an excess metal to other metals and nutrient levels. An alternative technique was finally established which involved an evaluation of individual assays of Chlorella and Ankistrodesmus in separate media and a tissue-metal analysis on Chlorella. Chlorella fusca, commonly found in lakes with high metal concentrations, showed high tolerance to mixed-metal solutions, while Ankistrodesmus proved to be very sensitive. By determining the maximum yield ratio between Ankistrodesmus and Chlorella (i.e. the A/Ch ratio) it was possible to compare the toxic strength of harmful metals according to an established standard curve of A/Ch ratio versus mixed-metal concentrations. The levels of tissue-metal analysed in Chlorella also gave some indication as to which metals were responsible for the toxicity.     

The role of bivalve molluscs as tools in estuarine sediment toxicity testing: a review

Estuarine sediments frequently are repositories and therefore potential sources of anthropogenic contaminants. Many organic and metallic chemical compounds released into aquatic systems bind to particulates and so accumulate in the sediments, thus, sediments become repositories of contaminants in estuaries. These may also cause contamination through diffusion of porewater, resuspension of particulates and dispersal of benthic fauna. There is a need to assess the biological affects of these anthropogenic contaminants because they may be toxic to infauna and bottomfish. Sediment toxicity bioassays are a means for carrying out such an assessment and primarily provide data on toxicity by measuring the effects on the test organism. Existing sediment toxicity bioassays rely on a battery of aquatic toxicity tests, which are based on the extraction of pore water, and elutriate from sediments and then subjecting these sediment phases to toxicity testing regimes. Two estuarine bivalve molluscs, Scrobicularia plana and Tapes semidecussatus were used to assess the ecotoxicity of field-collected sediments from estuarine and coastal areas around the Irish and English Coast over a 3-year study period. A variety of endpoints were measured during the study including survival in air, behaviour, animal condition, biochemistry, soft tissue metal concentrations, lysosomal membrane integrity and histopathology. Of these endpoints, the most sensitive were survival, survival in air, lysosomal membrane integrity, behaviour and histopathology.     

Toxicity of Santander Bay Sediments to the Euryhaline Freshwater Oligochaete Limnodrilus hoffmeisteri

The freshwater euryhaline oligochaete Limnodrilus hoffmeisteri was selected for use in bioassays with polluted sediment from Santander Bay. It is easy to culture; is tolerant of low to moderate, up to 15‰, salinity; and is common in oligohaline conditions in European and North American estuaries. Worms were collected from an estuarine population and kept in unpolluted sediment for between 2 and 4 weeks, under laboratory conditions, at 7–8.5‰ salinity and 22.5 °C. Sediment from different sites in Santander Bay were sieved through 250 μm mesh and adjusted to a salinity of approx. 7‰ prior to the bioassays, either by adding sea water or dechlorinated tap water to the overlying water. High levels of ammonia in some sediment, which may confound results in ecotoxicity bioassays, were reduced by oxidation of the sediments in shallow trays. Sediment bioassays were performed with sexually mature Limnodrilus hoffmeisteri worms in 250-ml beakers, with a 1:3 ratio of sediment:water and 4 worms per baker. Endpoints in the 14-day bioassay were % mortality, adult final biomass, % adults that have shown resorption of the clitellum, number of cocoons, and burrowing behaviour. It was possible to rank the sites according to their toxicity using both mortality rates and sublethal effects. The control site had the following values for the endpoints: 5% mortality, ( ) 2.40 ± 1.52 cocoons per beaker and 1.271 ± 0.470 mg dw adult final biomass. The most toxic sediment resulted in 65% mortality, resorption of the clitellum in 67% of the adults, no production of cocoons and a low final biomass ( =0.681 ± 0.489 mg dw per adult). A second site had high mortality (60%) and no reproduction, although resorption of the clitellum did not occur in surviving animals. The remaining sites showed similar mortality (35–42%), and at only one of them was low reproduction observed (0.8 ± 0.447 cocoons per beaker). Behavioural effects, measured as length of galleries in a fixed area of the test-vessel at the end of the bioassay, were significant compared with control at only one site. Multivariate analysis showed the mortality gradient to be the strongest, with a second unassociated gradient representing clitellum resorption. The mortality gradient was associated with Cu and Zn concentration, and PAH and Pb possibly with resorption.     

A comparison of nutrient availability measured by chemical analysis and calculated from bioassay yields

The concentrations of soluble biologically available nitrogen and phosphorus were determined using Selenastrum capricornutum bioassays and compared with analytically measured soluble nitrate (NO₃-N) and soluble reactive phosphorus concentrations during enrichment studies in a South African impoundment. The NO₃-N analyses consistently underestimated the soluble biologically available nitrogen and the extent of the discrepancy decreased with increasing NO₃-N concentration. Biological availability of soluble organic nitrogen during the bioassays is suggested as a reason for the discrepancies. At low soluble reactive phosphorus concentrations the analytical measurements underestimated the soluble biologically available phosphorus while at high soluble reactive phosphorus concentrations the analytical measurements were considerable overestimates of soluble biologically available phosphorus. Possible reasons for the observed trend are discussed.     

Ecotoxicological studies with the freshwater rotifer Brachionus calyciflorus II. An assessment of the chronic toxicity of lindane and 3,4-dichloroaniline using life tables

The effects of chronic exposure of the freshwater rotifer Brachionus calyciflorus to the toxicants lindane and 3,4-dichloroaniline (DCA), were evaluated. The parameters used to determine the toxicity on these compounds were the age-specific and fertility, and the demographic parameters: intrinsic rate of natural increase (r), generation time (T), net reproductive rate (R _o), reproductive value (V _x) and life expectancy (e _o). All the demographic parameters studied decreased with increasing toxicant concentrations. The use of life tables techniques with B. calyciflorus as a test method for the determination of chronic toxicity of xenobiotics is discussed.     

Fecundity,reproduction, and growth of Moina macrocopa fed different algae

Three micro algae, Ankistrodesmus convolutus, Scenedesmus incrassatulus and Chlorella vulgaris, at three concentrations, were tested as diets for Moina macrocopa. Their effect on reproduction, fecundity, growth and survival was evaluated. All three algae satisfied the nutritional requirements of M. macrocopa, despite their difference in size. The best concentration, expressed in dry weight, was 5 mg l^–1 for all three. Time to first reproduction was 4 days with all diets. Average time between clutches was 33 hours, and maximum number of clutches was 12; the highest average number of offspring per brood was 27. The largest broods were the fourth to sixth, depending on the diet. Ephippium hatching depends on temperature, taking only 24 hours at 30 °C and 48 hours at 27 °C. This species could be an important test organism in aquatic bioassays, and a live food in aquaculture.     

The use of life-tables for evaluating the chronic toxicity of pollutants toMysidopsis bahia

This study examines the application of population statistics to laboratory-derived toxicological data for the purpose of developing a predictive model that assesses the population consequences of pollutant and environmentally-induced stress. Life tables are used to calculate age-specific survivorship, fecundity, intrinsic rate of population increase (r), and reproductive value (V_a), for populations ofMysidopsis bahia chronically exposed, in separate tests, to mercury and nickel. The population statistics, r, and V_a, and pollutant dose are defined quantitatively, and the critical value (r = 0) for the intrinsic rate of population increase is compared with traditional toxicological measures of acute and chronic toxicity.The effects of an environmental variable, predation, significantly reduce the critical value (r = 0) for the intrinsic rate of growth and demonstrate the interactions of multiple stressors. The use of the population statistic, r, is recommended for estimating the ecological significance of toxicological data and for facilitating the assessment of interactions between pollutant-stressed populations and dynamic environmental pressures.Contribution No. 258 of the U.S. Environmental Protection Agency.     

Evaluation and application of aquatic toxicity tests: use of the Microtox test for the prediction of toxicity based upon concentrations of contaminants in soil

The quality control for the reuse of cleaned soil from a contaminated site consisted in the determination of the main contaminants by analytical chemical methods such as GC and HPLC. Since it is not possible to analyze for all contaminants a toxicity test should be used to detect large concentrations of not routinely analyzed chemicals. The aim of the study was to develop a system for toxicity testing, which should be able to predict the toxicity of soil samples based on the concentration of chemicals in the soil and to detect toxic chemicals not analyzed by the routinely conducted soil analysis.Based upon the relative sensitivity to various contaminants as well as practical aspects such as test duration and costs the Microtox® test was favoured over the bioassays with Daphnia magna and Scenedesmus subspicatus. The Microtox® test was used to measure the toxicity of various pesticides and their major metabolites. The toxicity data of the pure compounds were used to predict the toxicity (EC₅₀ and % inhibition of the bioluminescence reaction) of defined mixtures of chemicals in water by applying two different mathematical appriaches which are based on the additivity of the effects of the single chemicals. The predicted values were compared with the experimental data and showed good agreement.In order to be able to predict the toxicity of soil samples using the Microtox® test the soil/water partition coefficient (K _d) was measured for the main contaminants. The toxicity of soil samples was predicted by calculating the concentration of the contaminants in the leachate by using the corresponding concentration in the soil and applying the K _d values determined. From the calculated composition of the leachate the expected toxicity was estimated. This value was compared with the toxicity experimentally determined in the Microtox® test.     

The influence of mineral salts on fecundity of the water flea (Daphnia magna) and the implications on toxicity testing of industrial wastewater

The effects of mineral salts constituting water hardness on fecundity ofDaphnia magna were assessed. Of the salts tested, increased concentrations of NaHCO₃ and MgSO₄ had no effect on fecundity, CaSO₄ significantly increased fecundity, and KCl significantly reduced fecundity. The number of offspring produced per daphnid was correlative to the CaSO₄ concentration at CaSO₄ concentrations between 91 and 2100 mg/ℓ. The effects of CaSO₄ on daphnid fecundity could influence the interpretive outcome of industrial wastewater toxicity tests using this species when the waste and dilution waters contain different concentrations of CaSO₄. It is recommended that when performing these tests, dilution water be sampled at the intake site of the industry's water source, thus assuring initial comparability of the waste and dilution waters. The CaSO₄ content of the water prior to and after industrial use should be determined to identify any alterations of CaSO₄ concentration during use. Identification of CaSO₄ concentration differences can aid in the interpretation of effects associated with the wastewater.     

Lethal and sublethal effects of abamectin, spinosad, methoxyfenozide and acetamiprid on the predaceous plant bug Deraeocoris brevis in the laboratory

Deraeocoris brevis (Uhler) (Hemiptera: Miridae), an important generalist predator in pome fruits in the western United States, was reared in the laboratory on frozen Ephestia kuehniella (Zeller) (Lepidoptera: Pyralidae) eggs and treated in a Potter spray tower to assess acute toxicity and chronic sublethal effects of abamectin, spinosad, methoxyfenozide, and acetamiprid. Acute toxicity was assessed using topical application. Sublethal effects were examined as the combined result of topical, residual and oral exposure. Two different dose rates, the full field rate and 10% rate of the full field rate, were compared to distilled water as the untreated check. Methoxyfenozide and spinosad had no acute toxicity to nymphs and adults at the 10% and full field rate, and no effect on egg hatch and nymph survival just after hatch. Acetamiprid and abamectin at the full field rate did not affect egg hatch, but the residue had moderate to high toxicity to hatched nymphs. Also, topically applied acetamiprid and abamectin had moderate to high acute toxicity to nymphs and adults at the full field rate, but moderate toxicity at the 10% rate. In sublethal bioassays, abamectin-treated adults (10% field rate) laid 80% fewer and less viable eggs compared with the untreated check. Spinosad-treated (full field rate) adults laid fewer and less viable eggs. Also, egg hatch in the subsequent generation was lower. Methoxyfenozide had no sublethal effects on adults at the full rate, but slowed development of 4th instars following treatment of 2nd instar nymphs, and lowered fecundity by 30% in the subsequent generation compared with the untreated check. Acetamiprid (10% rate) applied to nymphs or adults had no effects on development or reproduction. Results from this study suggest that the newer reduced risk insecticides, which have begun to replace organophosphate insecticides in pome fruits in the United States, are not as selective to natural enemies as initially thought. Their impact on D. brevis varied with chemistry and mode of action from primarily acute toxicity (i.e., acetamiprid) to reproductive and other sublethal effects (i.e., methoxyfenozide, spinosad) or a combination of both (i.e., abamectin). In addition to acute toxicity, sublethal effects need to be quantified in order to accurately predict the total impact of a pesticide on a natural enemy in the field.