Mortality and Cholinesterase Inhibition in Butterflies Following Aerial Naled Applications for Mosquito Control on the National Key Deer Refuge.

Natural resource managers are concerned about the impacts of aerial ultra-low volume spray (ULV) of insecticides for mosquito control (i.e., mosquito adulticides) and seek science-driven management recommendations that reduce risk but allow vector control for nearby human populations. Managers at the National Key Deer Refuge (Florida Keys, FL) are concerned for ULV effects upon conservation efforts for imperiled butterflies (Florida leafwing [Anaea troglodyta floridalis] and Bartram's hairstreak [Strymon acis bartrami] butterflies). No-spray zones were designated for protection of those butterflies, but their effectiveness for mitigation is unclear. To address this uncertainty, cholinesterase activity (ChE) and mortality were monitored for caged butterflies gulf fritillary [Agraulis vanilla] and great southern white [Ascia monuste]) deployed on the Refuge during three aerial ULV applications of the insecticide naled. Residue samplers also were deployed to estimate butterfly exposure. Spray efficacy against mosquitoes was assessed by deploying caged mosquitoes at the same locations as the butterflies. Average naled residue levels on filter paper samplers in the target area (1882-2898 µg/m2) was significantly greater than in the no-spray zone (9-1562 µg/m2). Differences between the no-spray zone and target area for butterfly mortality and ChE were inconsistent. Average mortality was significantly lower, and average ChE was significantly higher in the no-spray zone for larvae of one species but not for larvae of the other species. Mosquito mortality did not differ significantly between the two areas. Data from the present study reflect the inconsistent effectiveness of no-spray zones on the Refuge using standard methods employed at the time by the vector control agency in the Florida Keys and possibly by other vector control agencies in similar coastal environments. Furthermore, these findings helped to guide the design and to improve the conservation value of future no-spray zone delineations while allowing for treatment in areas where mosquito control is necessary for vector-borne disease reduction.

Elemental imaging (LA-ICP-MS) of zebrafish embryos to study the toxicokinetics of the acetylcholinesterase inhibitor naled.

The zebrafish embryo is an important model in ecotoxicology but the spatial distribution of chemicals and the relation to observed effects is not well understood. Quantitative imaging can help to gain insights into the distribution of chemicals in the zebrafish embryo. Laser ablation inductively coupled plasma-mass spectrometry (LA-ICP-MS) is used to quantify the uptake and the uptake kinetics of the bromine (Br) containing organophosphate naled (Dibrom®, dimethyl-1,2-dibromo-2,2-dichloroethylphosphate) and its distribution in zebrafish embryos using Br as the marker element. During exposure, the Br amounts increase in the embryos parallel to the irreversible inhibition of the acetylcholinesterase (AChE). The final amount of Br in the embryo (545 pmol/embryo) corresponds to a 280-fold enrichment of naled from the exposure solution. However, LC-MS/MS analyses showed that the internal concentration of naled remained below the LOD (7.8 fmol/embryo); also the concentration of its known transformation product dichlorvos remained low (0.85 to 2.8 pmol/embryo). These findings indicate the high reactivity and high transformation rate of naled to other products than dichlorvos. 12C normalized intensity distributions of Br in the zebrafish embryo showed an enrichment of Br in its head region. Kernel density estimates of the LA-ICP-MS data were calculated and outline the high reproducibility between replicated and the shift in the Br distribution during exposure. The Br enrichment indicates a preferential debromination or direct covalent reaction of naled with AChE in this region. Graphical abstract ᅟ.

Mosquito control pesticides and sea surface temperatures have differential effects on the survival and oxidative stress response of coral larvae.

The declining health of coral reefs is intensifying worldwide at an alarming rate due to the combined effects of land-based sources of pollution and climate change. Despite the persistent use of mosquito control pesticides in populated coastal areas, studies examining the survival and physiological impacts of early life-history stages of non-targeted marine organisms are limited. In order to better understand the combined effects of mosquito pesticides and rising sea surface temperatures, we exposed larvae from the coral Porites astreoides to selected concentrations of two major mosquito pesticide ingredients, naled and permethrin, and seawater elevated +3.5 °C. Following 18-20 h of exposure, larvae exposed to naled concentrations of 2.96 µg L(-1) or greater had significantly reduced survivorship compared to controls. These effects were not detected in the presence of permethrin or elevated temperature. Furthermore, larval settlement, post-settlement survival and zooxanthellae density were not impacted by any treatment. To evaluate the sub-lethal stress response of larvae, several oxidative stress endpoints were utilized. Biomarker responses to pesticide exposure were variable and contingent upon pesticide type as well as the specific biomarker being employed. In some cases, such as with protein carbonylation and catalase gene expression, the effects of naled exposure and temperature were interactive. In other cases pesticide exposure failed to induce any sub-lethal stress response. Overall, these results demonstrate that P. astreoides larvae have a moderate degree of resistance against short-term exposure to ecologically relevant concentrations of pesticides even in the presence of elevated temperature. In addition, this work highlights the importance of considering the complexity and differential responses encountered when examining the impacts of combined stressors that occur on varying spatial scales.

Toxicity and risk of permethrin and naled to non-target insects after adult mosquito management.

We derived laboratory LC50 values, assessed non-target insect risks, and conducted a field bioassay for ultra-low-volume (ULV) aerosol applications of insecticides used to manage adult mosquitoes. The house cricket, Acheta domesticus (L.), was used as an indicator species for medium- to large-bodied ground dwelling insects. The 24-h LC(50) values for Permanone (formulated product of permethrin), Permanone + piperonyl butoxide (PBO), technical grade permethrin, and technical grade permethrin + PBO ranged from 0.052 to 0.9 microg/cm(2). The 24 h LC(50) for technical grade naled and Trumpet((R)) (formulated product of naled) were 0.038 and 0.44 microg/cm(2), respectively. The synergist ratio was 2.65 for Permanone + PBO and 1.57 for technical grade permethrin + PBO. The toxicity of technical grade permethrin was about 10-fold greater than Permanone. A risk assessment using modeled estimated environmental concentrations resulted in risk quotients (RQ) that exceeded regulatory levels of concern, but when compared to field-derived actual environmental concentrations RQs did not exceed a regulatory level of concern, except in the case of technical grade naled. These results were expected because higher tiered risk assessments using field-verified data generally lead to lower risk estimates. Field bioassays using caged crickets showed no significant mortality for permethrin or naled after a single truck-mounted ULV application. The results of the risk assessment using actual environmental concentrations are supported by the field bioassays and suggest that a single ULV application of synergized or unsynergized permethrin and naled most likely will not result in population impacts on medium- to large-bodied insects.

Prenatal naled and chlorpyrifos exposure is associated with deficits in infant motor function in a cohort of Chinese infants.

BACKGROUND: Organophosphate insecticides (OPs) are used worldwide, yet despite nearly ubiquitous exposure in the general population, few have been studied outside the laboratory. Fetal brains undergo rapid growth and development, leaving them susceptible to long-term effects of neurotoxic OPs. The objective here was to investigate the extent to which prenatal exposure to OPs affects infant motor development. METHODS: 30 OPs were measured in umbilical cord blood using gas chromatography tandem mass spectrometry in a cohort of Chinese infants. Motor function was assessed at 6-weeks and 9-months using Peabody Developmental Motor Scales 2nd edition (PDMS-2) (n=199). Outcomes included subtest scores: reflexes, stationary, locomotion, grasping, visual-motor integration (V-M), composite scores: gross (GM), fine (FM), total motor (TM), and standardized motor quotients: gross (GMQ), fine (FMQ), total motor (TMQ). RESULTS: Naled, methamidophos, trichlorfon, chlorpyrifos, and phorate were detected in ≥10% of samples. Prenatal naled and chlorpyrifos were associated with decreased 9-month motor function. Scores were 0.55, 0.85, and 0.90 points lower per 1ng/mL increase in log-naled, for V-M (p=0.04), FM (p=0.04), and FMQ (p=0.08), respectively. For chlorpyrifos, scores were 0.50, 1.98, 0.80, 1.91, 3.49, 2.71, 6.29, 2.56, 2.04, and 2.59 points lower for exposed versus unexposed infants, for reflexes (p=0.04), locomotion (p=0.02), grasping (p=0.05), V-M (p<0.001), GM (p=0.007), FM (p=0.002), TM (p<0.001), GMQ (p=0.01), FMQ (p=0.07), and TMQ (p=0.008), respectively. Girls appeared to be more sensitive to the negative effects of OPs on 9-month motor function than boys. CONCLUSIONS: We found deficits in 9-month motor function in infants with prenatal exposure to naled and chlorpyrifos. Naled is being aerially sprayed to combat mosquitoes carrying Zika virus, yet this is the first non-occupational human study of its health effects. Delays in early-motor skill acquisition may be detrimental for downstream development and cognition.

Aerial and tidal transport of mosquito control pesticides into the Florida Keys National Marine Sanctuary.

This project was undertaken as the initial monitoring program to determine if mosquito adulticides applied along the Florida Keys cause adverse ecological effects in the Florida Keys National Marine Sanctuary (FKNMS). The study monitored the distribution and persistence of two mosquito adulticides, permethrin and dibrom (naled), during three separate routine applications by the Florida Keys Mosquito Control District. The approach was to determine if toxic concentrations of the pesticides entered the FKNMS by aerial drift or tidal transport. The amount of pesticide entering the FKNMS by way of aerial drift was monitored by collection on glass fiber filter pads, set on floats in a grid pattern on either side of the FKNMS. Permethrin was recovered from filter pads on the leeward side for each of the three applications, ranging from 0.5 to 50.1 microg/m(2) throughout the study. Tidal current transport was monitored by collection of surface and subsurface water samples at each grid site. Tidal transport of naled and dichlorvos (naled degradation product) was apparent in the adjacent waters of the FKNMS. These compounds were detected in subsurface, offshore water at 0.1 to 0.6 microg/1, 14 hr after application. Permethrin was not detected in offshore water samples; however, concentrations ranging from 5.1 to 9.4 microg/l were found in surface water from the canal system adjacent to the application route. Comparison of the observed environmental concentrations with toxicity data (permethrin LC-50, 96 hr for Mysidopsis bahia = 0.02 microg/l) indicated a potential hazard to marine invertebrates in the canals with possible tidal transport to other areas.

Mosquito control insecticides: a probabilistic ecological risk assessment on drift exposures of naled, dichlorvos (naled metabolite) and permethrin to adult butterflies.

A comprehensive probabilistic terrestrial ecological risk assessment (ERA) was conducted to characterize the potential risk of mosquito control insecticide (i.e., naled, it's metabolite dichlorvos, and permethrin) usage to adult butterflies in south Florida by comparing the probability distributions of environmental exposure concentrations following actual mosquito control applications at labeled rates from ten field monitoring studies with the probability distributions of butterfly species response (effects) data from our laboratory acute toxicity studies. The overlap of these distributions was used as a measure of risk to butterflies. The long-term viability (survival) of adult butterflies, following topical (thorax/wings) exposures was the environmental value we wanted to protect. Laboratory acute toxicity studies (24-h LD50) included topical exposures (thorax and wings) to five adult butterfly species and preparation of species sensitivity distributions (SSDs). The ERA indicated that the assessment endpoint of protection, of at least 90% of the species, 90% of the time (or the 10th percentile from the acute SSDs) from acute naled and permethrin exposures, is most likely not occurring when considering topical exposures to adults. Although the surface areas for adulticide exposures are greater for the wings, exposures to the thorax provide the highest potential for risk (i.e., SSD 10th percentile is lowest) for adult butterflies. Dichlorvos appeared to present no risk. The results of this ERA can be applied to other areas of the world, where these insecticides are used and where butterflies may be exposed. Since there are other sources (e.g., agriculture) of pesticides in the environment, where butterfly exposures will occur, the ERA may under-estimate the potential risks under real-world conditions.

Acute toxicity and risk assessment of permethrin, naled, and dichlorvos to larval butterflies via ingestion of contaminated foliage.

Three Florida native larval butterflies (Junonia coenia, Anartia jatrophae, Eumaeus atala) were used in the present study to determine the acute toxicity, hazard, and risk of a 24h ingestion of leaves contaminated with the adult mosquito control insecticides permethrin, naled, and dichlorvos to late 4th and early 5th in-star caterpillars. Based on 24-h LD50s for ingestion, naled was more acutely toxic than permethrin and dichlorvos to caterpillars. Hazard quotients using the ratio of the highest doses and the 90th percentile doses from field measurements in host plant foliage following actual mosquito control applications to the toxicological benchmarks from laboratory toxicity tests indicate potential high acute hazard for naled compared to permethrin and dichlorvos. Based on probabilistic ecological risk methods, naled exposure doses in the environment also presented a higher acute risk to caterpillars than permethrin and dichlorvos. The acute toxicity laboratory results and ecological risk assessment are based only on dietary ingestion and single chemical doses. It does not include other typical exposure scenarios that may occur in the environment. It is thus plausible to state that the ecological risk assessment presented here underestimates the potential risks in the field to caterpillars. However, one assumption that is scientifically feasible and certainly real from the results - if the environmental exposure doses of mosquito control operations are similar or higher to those presented here in leaves from the field, after applications, there will likely be significant mortalities and other adverse effects on caterpillar populations.

Characterizing stress gene expression in reef-building corals exposed to the mosquitoside dibrom.

We characterize two genes expressed in Acropora cervicornis upon exposure to 0.5 microg/l of dibrom, a pesticide used for mosquito control in the Florida Keys. Fragments of these genes were isolated, sequenced, and developed into chemiluminescent probes for Northern slot blots. Expression of target transcripts was detected in corals exposed to a variety of stressors including organophosphates, organochlorines, heavy metals, naphthalene, and temperature. Within the context of stressors examined, the D25 probe demonstrates toxicant and concentration specificity for organophosphates, whereas the D50 probe had broader specificity, detecting transcripts in corals exposed to dibrom, naphthalene, and temperature stress. After characterizing specificity in the lab, these probes were used on field samples taken from the Florida Keys. Both probes detected their targets in samples taken from the upper Florida Keys in August 2000. Preliminary search of sequence databases suggest similarity exists between D25 and a thioesterase.

Minimizing the impact of the mosquito adulticide naled on honey bees, Apis mellifera (Hymenoptera: Apidae): aerial ultra-low-volume application using a high-pressure nozzle system.

The impact of the mosquito adulticide naled on honey bees, Apis mellifera L., was evaluated by exposing test beehives to nighttime aerial ultra-low-volume (ULV) applications using a high-pressure nozzle system. The tests were conducted during routine mosquito control missions at Manatee County, Florida, in summer 2000. Two treatment sites were sprayed a total of four times over a 10-wk period. Honey bees, which clustered outside of the hive entrances, were subjected to naled exposure during these mosquito control sprays. The highest average naled ground deposition was 2,688 microg/m2 at the Port Manatee site, which resulted in statistically significant bee mortality (118) compared with the controls. At the Terra Ceia Road site, an intermediate level of naled deposition was found (1,435 microg/m2). For this spray mission, the range of dead bees per hive at Terra Ceia was 2 to 9 before spraying and 5 to 36 after naled application. Means of all other naled ground depositions were < 850 microl/m2. We concluded that substantial bee mortality (> 100 dead bees) resulted when naled residue levels were > 2,000 kg/m2 and honey bees were clustered outside of the hive entrances during mosquito adulticide applications. Compared with the flat-fan nozzle systems currently used by most of Florida's mosquito control programs, the high-pressure nozzle system used in this experiment substantially reduced environmental insecticide contamination and lead to decreased bee mortality. Statistical analysis also showed that average honey yield at the end of the season was not significantly reduced for those hives that were exposed to the insecticide.

Efficacy of Dibrom, Trumpet, and Scourge against four mosquito species in Louisiana.

Adult mortality of Anopheles quadrimaculatus, Culex quinquefasciatus, and the Aedes spp. complex (Aedes sollicitans and Aedes taeniorhynchus) was observed after aerial ultra-low volume (ULV) exposure to Dibrom, Trumpet, and Scourge. Dibrom was applied at 112 g active ingredient (AI)/ha, Trumpet at 112 g AI/ha, and Scourge at 1.96 g AI/ha. At all time intervals, Dibrom and Trumpet were significantly more effective against the Aedes spp. complex than against An. quadrimaculatus and Cx. quinquefasciatus. Scourge was significantly more effective against An. quadrimaculatus and Cx. quinquefasciatus than Dibrom or Trumpet. Trumpet was evaluated at lower labeled rates (28, 56, and 84 g AI/ha) against Cx. quinquefasciatus and the Aedes spp. complex. Adult mortality with Trumpet increased significantly at 1 and 24 h against Cx. quinquefasciatus. With the Aedes spp. complex, mortality increased with rate at 1 h, but at 12 and 24 h, the medium and high dosages were not significantly different from each other. Culex quinquefasciatus and the Aedes spp. complex were also subjected to ULV ground applications of Dibrom, Trumpet, and Scourge. Dibrom was applied at 22.4 g AI/ha, Trumpet at 22.4 g AI/ha, and Scourge as a 1:6 mineral oil mixture at 1.96 g AI/ha. Relative to Dibrom and Trumpet, mortality from Scourge differed greatly with mosquito species. Against Cx. quinquefasciatus, Scourge was significantly more effective than Dibrom and Trumpet at all times and distances, but against the Aedes spp. complex Scourge was significantly less effective.

Acute toxicity of mosquitocidal compounds to young mosquitofish, Gambusia affinis.

Toxicity of Florida mosquito larvicides and adulticides to 3-5 day old Gambusia affinis was determined in the laboratory. After 24-h exposure, the larvicides, temephos, fenoxycarb and petroleum distillates had LC50 values of 5.60, 1.05 and 593.4 ppm, respectively. After 24 h the adulticides resmethrin, fenthion, naled and malathion had LC50 values of 0.007, 2.94, 3.50 and 12.68 ppm, respectively. The only compound toxic to young mosquitofish at maximum field application rates was resmethrin. However, in the light of earlier tests, aerially applied adulticides generally reach the water surface at reduced concentrations and thus probably pose little or no risk to mosquitofish populations.

Impact of naled on the mosquito vectors of eastern equine encephalitis virus.

This letter questions the appropriateness of methodology used in a study by Howard and Oliver (J. Am. Mosq. Control Assoc. 13:315-325; 1988). Two independent data sets, collected for different purposes by 2 different groups, were subjected to statistical analysis to determine if the data sets differed. The experimental "design," as described by the authors, is an example of pseudoreplication, which arises when replicates are collected at a scale finer than the one for which conclusions of statistical testing are intended to be drawn. All of the components of a properly designed field experiment (control, replication, randomization, and interspersion) are missing from this study. The authors proceed to draw a series of conclusions from the data presented. Few, if any, of the conclusions can be supported by the evidence presented. The assertions put forward in this paper could have a severe negative impact on efforts to prevent transmission of arboviruses or other pathogens to humans and domestic animals.

Monitoring the aquatic toxicity of mosquito vector control spray pesticides to freshwater receiving waters.

Pesticides are applied to state and local waterways in California to control insects such as mosquitoes, which are known to serve as a vector for West Nile Virus infection of humans. The California State Water Resources Control Board adopted a National Pollutant Discharge Elimination System General Permit to address the discharge to waters of the United States of pesticides resulting from adult and larval mosquito control. Because pesticides used in spray activities have the potential to cause toxicity to nontarget organisms in receiving waters, the current study was designed to determine whether toxicity testing provides additional, useful environmental risk information beyond chemical analysis in monitoring spray pesticide applications. Monitoring included a combination of aquatic toxicity tests and chemical analyses of receiving waters from agricultural, urban, and wetland habitats. The active ingredients monitored included the organophosphate pesticides malathion and naled, the pyrethroid pesticides etofenprox, permethrin, and sumithrin, pyrethrins, and piperonyl butoxide (PBO). Approximately 15% of the postapplication water samples were significantly toxic. Toxicity of half of these samples was attributed to the naled breakdown product dichlorvos. Toxicity of 2 other water samples likely occurred when PBO synergized the effects of pyrethroid pesticides that were likely present in the receiving system. Four of 43 postapplication sediment samples were significantly more toxic than their corresponding pre-application samples, but none of the observed toxicity was attributed to the application events. These results indicate that many of the spray pesticides used for adult mosquito control do not pose significant acute toxicity risk to invertebrates in receiving systems. In the case of naled in water, analysis of only the active ingredient underestimated potential impacts to the receiving system, because toxicity was attributed to the breakdown product, dichlorvos. Toxicity testing can provide useful risk information about unidentified, unmeasured toxicants or mixtures of toxicants. In this case, toxicity testing provided information that could lead to the inclusion of dichlorvos monitoring as a permit requirement.

Effects of mosquito control pesticides on competent queen conch (Strombus gigas) larvae.

Pesticides are applied seasonally in the Florida Keys to control nuisance populations of mosquitoes that pose a health threat to humans. There is, however, a need to investigate the effects of these pesticides on non-target marine organisms. We tested naled and permethrin, two mosquito adulticides used in the Keys, on a critical early life-history stage of queen conch (Strombus gigas). We conducted 12-h exposure experiments on competent (i.e., capable of undergoing metamorphosis) queen conch larvae using environmentally relevant pesticide concentrations. We found that there was little to no mortality and that the pesticides did not induce or interfere with metamorphosis. However, after introduction of a natural metamorphic cue (extract of the red alga Laurencia potei), a significantly greater proportion of larvae underwent metamorphosis in the pesticide treatments than in those with the alga alone. In addition to the morphogenetic pathway that induces metamorphosis when stimulated, there thus appears to be a regulatory pathway that enhances the response to metamorphic triggers, as suggested by the increased sensitivity of the queen conch larvae to the algal cue after pesticide exposure (i.e., the pesticides stimulated the regulatory pathway). The regulatory pathway probably plays a role in the identification of high-quality habitat for metamorphosis, as the increased response to the algal cue suggests. Aerial drift and runoff can carry these pesticides into nearshore waters, where they may act as a false signal of favorable conditions and facilitate metamorphosis in suboptimal habitat, thus adversely affecting recruitment in nearshore queen conch populations.

The relationship between total cholinesterase activity and mortality in four butterfly species.

The relationship between total cholinesterase activity (TChE) and mortality in four butterfly species (great southern white [Ascia monuste], common buckeye [Junonia coenia], painted lady [Vanessa cardui], and julia butterflies [Dryas julia]) was investigated. Acute contact toxicity studies were conducted to evaluate the response (median lethal dose [LD50] and TChE) of the four species following exposure to the organophosphate insecticide naled. The LD50 for these butterflies ranged from 2.3 to 7.6 µg/g. The average level of TChE inhibition associated with significant mortality ranged from 26 to 67%, depending on the species. The lower bounds of normal TChE activity (2 standard deviations less than the average TChE for reference butterflies) ranged from 8.4 to 12.3 µM/min/g. As a percentage of the average reference TChE activity for the respective species, the lower bounds were similar to the inhibition levels associated with significant mortality, indicating there was little difference between the dose resulting in significant TChE inhibition and that resulting in mortality.

Risk assessment for adult butterflies exposed to the mosquito control pesticide naled.

A prospective risk assessment was conducted for adult butterflies potentially exposed to the mosquito control insecticide naled. Published acute mortality data, exposure data collected during field studies, and morphometric data (total surface area and fresh body weight) for adult butterflies were combined in a probabilistic estimate of the likelihood that adult butterfly exposure to naled following aerial applications would exceed levels associated with acute mortality. Adult butterfly exposure was estimated based on the product of (1) naled residues on samplers and (2) an exposure metric that normalized total surface area for adult butterflies to their fresh weight. The likelihood that the 10th percentile refined effect estimate for adult butterflies exposed to naled would be exceeded following aerial naled applications was 67 to 80%. The greatest risk would be for butterflies in the family Lycaenidae, and the lowest risk would be for those in the family Hesperidae, assuming equivalent sensitivity to naled. A range of potential guideline naled deposition levels is presented that, if not exceeded, would reduce the risk of adult butterfly mortality. The results for this risk assessment were compared with other risk estimates for butterflies, and the implications for adult butterflies in areas targeted by aerial naled applications are discussed.

Use of butterflies as nontarget insect test species and the acute toxicity and hazard of mosquito control insecticides.

Honeybees are the standard insect test species used for toxicity testing of pesticides on nontarget insects for the U.S. Environmental Protection Agency (U.S. EPA) under the Federal Insecticide Fungicide and Rodenticide Act (FIFRA). Butterflies are another important insect order and a valued ecological resource in pollination. The current study conducted acute toxicity tests with naled, permethrin, and dichlorvos on fifth larval instar (caterpillars) and adults of different native Florida, USA, butterfly species to determine median lethal doses (24-h LD50), because limited acute toxicity data are available with this major insect group. Thorax- and wing-only applications of each insecticide were conducted. Based on LD50s, thorax and wing application exposures were acutely toxic to both caterpillars and adults. Permethrin was the most acutely toxic insecticide after thorax exposure to fifth instars and adult butterflies. However, no generalization on acute toxicity (sensitivity) of the insecticides could be concluded based on exposures to fifth instars versus adult butterflies or on thorax versus wing exposures of adult butterflies. A comparison of LD50s of the butterflies from this study (caterpillars and adults) with honeybee LD50s for the adult mosquito insecticides on a µg/organism or µg/g basis indicates that several butterfly species are more sensitive to these insecticides than are honeybees. A comparison of species sensitivity distributions for all three insecticides shows that permethrin had the lowest 10th percentile. Using a hazard quotient approach indicates that both permethrin and naled applications in the field may present potential acute hazards to butterflies, whereas no acute hazard of dichlorvos is apparent in butterflies. Butterflies should be considered as potential test organisms when nontarget insect testing of pesticides is suggested under FIFRA.

Aerial ultra-low-volume application of naled: impact on nontarget imperiled butterfly larvae (Cyclargus thomasi bethunebakeri) and efficacy against adult mosquitoes (Aedes taeniorhynchus).

We assessed the exposure and acute toxicity of naled, applied aerially as an ultra-low-volume spray for mosquito control, on late instar larvae of the Miami blue (Cyclargus thomasi bethunebakeri) (Comstock and Huntington 1943) (Lepidoptera: Lycaenidae), an imperiled South Florida butterfly. We concurrently evaluated the control efficacy against caged adult female salt-marsh mosquitoes (Aedes taeniorhynchus) (Wiedemann 1821) (Diptera: Culicidae). This 3-yr study was conducted in north Key Largo (Monroe County, FL) beginning in 2006. The field trials incorporated 15 sampling stations: nine in the target spray zone, three in the spray drift zone at varying distances from the target zone, and three in the control zone not subjected to naled spray drift. A total of six field spray trials were completed, three at an altitude of 30.5 m (100 feet), and three at 45.7 m (150 feet). For all trials, the ultra-low-volume application of Trumpet EC insecticide (78% naled) at a rate of 54.8 ml/ha (0.75 fl. oz/acre) was effective in killing caged adult mosquitoes in the target zone. Butterfly larvae survival was significantly reduced in the spray zone compared with drift and control zones. Analysis of insecticide residue data revealed that the mortality of the late instar butterfly larvae was a result of exposure to excess residues of naled. Additional research is needed to determine mitigation strategies that can limit exposure of sensitive butterflies to naled while maintaining mosquito control efficacy.