Non-target Aquatic Insects Research Articles

Non-target effects of methoprene and larvicidal surface films on invertebrate predators of mosquito larvae.

Mosquito larvicides are used across a variety of aquatic habitats, although when applied they likely affect other aquatic organisms. The removal or impairment of top insect predators via larvicides could be beneficial to mosquitoes by allowing their populations to rebound once pesticide levels dissipate. Our goal was to determine if two larvicide types, growth regulators (IGRs) and surface films (SFs), harm non-target aquatic insect communities, and if these chemicals influence the ability of predatory aquatic insects to regulate mosquitoes. We surveyed aquatic sites before and after IGR and SF-application, then compared changes in insect community structure. Evenness was lower in SF treated habitats, and when analyzing prey/controphic taxa only, evenness and diversity changed in untreated reference areas, suggesting that differences measured were due to other environmental factors, not larvicide presence. A field experiment was then conducted by exposing specific predatory aquatic insects to varying doses of IGRs and SFs and then placing them in mesocosms containing mosquito larvae. Surface films were directly lethal to adult dytiscids at recommended and high concentrations. Although we found no significant differences in mosquito emergence among all treatment levels, there was a trend of negative controls (no predator mesocosms) and SF-treated predators allowing the most mosquitoes to emerge compared to positive controls (predators not exposed to larvicides) and IGR-treated predators. Thus, these larvicides may have minimal effects on mosquito larvae predators, but the direct effects of surface films on insects that interact with the water's surface require further investigation.

A temporal perspective on aquatic subsidy: Bti affects emergence of Chironomidae

Emerging aquatic insects serve as one link between aquatic and adjacent riparian food webs via the flux of energy and nutrients. These insects provide high-quality subsidy to terrestrial predators. Thus, any disturbance of emergence processes may cascade to higher trophic levels and lead to effects across ecosystem boundaries. One stressor with potential impact on non-target aquatic insects, especially on non-biting midges (Diptera: Chironomidae), is the widely used mosquito control agent Bacillus thuringiensis var. israelensis (Bti). In a field experiment, we investigated emerging insect communities from Bti-treated (three applications, maximum field rate) and control floodplain pond mesocosms (FPMs) over 3.5 months for changes in their composition, diversity as well as the emergence dynamics and the individual weight of emerged aquatic insects over time. Bti treatments altered community compositions over the entire study duration – an effect mainly attributed to an earlier (∼10 days) and reduced (∼26%) peak in the emergence of Chironomidae, the dominant family (88% of collected individuals). The most reasonable explanation for this significant alteration is less resource competition caused by a decrease in chironomid larval density due to lethal effects of Bti. This is supported by the higher individual weight of Chironomidae emerging from treated FPMs (∼21%) during Bti application (April – May). A temporal shift in the emergence dynamics can cause changes in the availability of prey in linked terrestrial ecosystems. Consequently, terrestrial predators may be affected by a lack of appropriate prey leading to bottom-up and top-down effects in terrestrial food webs. This study indicates the importance of a responsible and elaborated use of Bti and additionally, highlights the need to include a temporal perspective in evaluations of stressors in aquatic-terrestrial meta-ecosystems.

The effect of diflubenzuron (Dimilin® 25 WP) on some non-target aquatic insect and crustacean species

The study is aimed at evaluating, under laboratory conditions, the side effects of a commercial formulation of diflubenzuron (Dimilin® 25 WP), which is an insecticide considered a chitin synthesis inhibitor, on some non-targeted aquatic species. The effect of two lethal doses of Dimilin LC10 (3.9 ng/L) and LC50 (16 ng/L) previously determined on fourth instar larvae of Culex pipiens L. 1758 were tested on some non-target aquatic species, abundant in the Lake of Birds (Northeast Algeria), a site classified under Ramsar Convention. The tested species belonged to two classes of invertebrates (Insects and Crustaceans). The insects included four species of Heteroptera: Corixa punctata Illiger, 1807, Notonecta glauca Latreille, 1802, Anisops sardea Latreille, 1802 and Plea minutissima Leach, 1817, and one species of Coleoptera Berosus signaticollis Charpentier,1825. For benthic crustaceans Daphnia magna Straus, 1820 (Cladocera) was retained. The results showed significant mortality recorded in C. punctata and medium mortality for B. signaticollis, A. sardea, N. glauca, P. minutissima and D. magna. Three-way ANOVA indicated highly significant effects of species, dose, and time. According to the sensitivity to Dimilin, the pairwise comparison of Tukey's test indicates that the most sensitive species was C. punctata followed by B. signaticollis and the least sensitive was N. glauca, followed by D. magna; then A. sardea and finally P. minutissima.

Facile green approach for solar energy assisted biogenic nanoparticles synthesis mediated by seed kernel aqueous extract of Trichosanthes tricupsidata and its potential biomedical applications

In this study, an eco-friendly approach is developed to green synthesize AgNPs using seed kernel aqueous extract of the bitter snake gourd, Trichosanthes tricupsidata (Tt). The biophysical properties of Tt-AgNPs formed under ideal conditions were characterized using UV–visible spectrophotometer, X-ray diffraction, Fourier transform infrared, and high-resolution transmission electron microscopy analyses. Tt-AgNPs showed substantial antibacterial activity against eight Gram-negative and five Gram-positive bacterial pathogens tested. Hemolytic assay performed against human-A erythrocytes ensured the biocompatibility of Tt-AgNPs at 2.8% of hemolysis that showed the standard requirements for nanoparticles hemolytic properties. Tt-AgNPs showed dose-dependent larval anti-mosquito activity against fourth instar larvae Aedes aegypti. Larval mortality of 100% was observed with highest concentration and the 24 h LC50 and LC90 values were found to be 233.18 and 446.22 µg/mL, respectively. No lethal effect was observed on non-target aquatic insect larvae Chironomus costatus. The overall findings of the study revealed the Tt-AgNPs are suitable for its potential biomedical applications.

Lethal and sub-lethal activity of Brevibacillus laterosporus on the mosquito Aedes albopictus and side effects on non-target water-dwelling invertebrates

The biocidal potential of Brevibacillus laterosporus against mosquitoes of major medical importance has been widely documented, but its effects on non-target invertebrates are still poorly known. In this study, we determined the lethal and sub-lethal effects of B. laterosporus strain UNISS 18, an entomopathogenic bacterium known for its effectiveness against synanthropic Diptera, on the larvae of the Asian tiger mosquito Aedes albopictus, a vector of several pathogens to humans. Moreover, we compared the larvicidal activity with the lethal action on the invasive snail Physella acuta and on two non-target water-dwelling species: the mayfly Cloeon dipterum, and the harlequin fly Chironomus riparius. B. laterosporus exhibited significant lethal effects on all the tested species with a concentration-dependent activity. However, the susceptibility varied among species, with a higher susceptibility of Ae. albopictus (LC50 = 0.16 × 107 spores mL−1) than the other species (LC50 = 0.31, 0.33, and 0.30 × 107 spores mL−1 for C. dipterum, C. riparius, and P. acuta, respectively). While 1st instar mosquito larvae were very susceptible to the bacterial infection, no effects on preimaginal development stages and adult emergence were observed at sub-lethal spores’ concentrations. Even if the efficacy of B. laterosporus against Ae. albopictus and the invasive freshwater snail P. acuta is promising for their control, the susceptibility of non-target beneficial aquatic insects, highlights the need of accurate evaluations before applying B. laterosporus for pest management in water environments.

Laboratory analysis of Au\u2013Pd bimetallic nanoparticles synthesized with Citrus limon leaf extract and its efficacy on mosquito larvae and non-target organisms

The current study provides novel results on the synthesis of bimetallic nanoparticles (BNPs) of gold and palladium (Au–Pd) with an eco-friendly and non-toxic aqueous leaf extract of plant Citrus limon. The BNPs were characterized and toxicity bioassay was examined on the larvae of the pathogen vectors such as Anopheles stephensi and Aedes aegypti mosquitoes. The predation efficiency test was evaluated on the invertebrate non-target organisms such as natural predatory nymphs of dragonfly and damselfly. The results of material characterization using UV VIS spectroscopy confirmed the synthesis of Au–Pd BNPs with the appearance of the SPR bands. FT-IR spectroscopy indicates the presence of functional groups containing high amounts of nitro compounds and amines on the surface of BNPs. TEM result shows the presence of spherical polydisperse Au–Pd BNPs in the sample. The XRD pattern displayed the semi-crystalline nature and the changes in the hydrodynamic size and surface potential was determined for the sample at 0 h, 24 h, 48 h, and 72 h of synthesis through DLS and ZP analysis. Au–Pd BNPs Bioassay provided the effective lethal concentrations (LC50) against the I–IV instar larvae of An. stephensi and Ae. aegypti after 24 h, 48 h, and 72 h of exposure. The LC50 obtained from the larvicidal bioassay was used to test its effect on the predation efficiency of the selected nymphs which showed increased predation from 40 to 48 h of exposure as compared to the negative control. Hereby, we conclude that Au–Pd BNPs bioassay shows toxic mosquito larvicidal activity at the selected concentration with no lethal effect on the predation efficiency of the selected stage of the predatory non-target aquatic invertebrate insects.

Binding properties to nicotinic acetylcholine receptors can explain differential toxicity of neonicotinoid insecticides in Chironomidae

Neonicotinoids are neuroactive insecticides commonly detected in freshwater ecosystems. Recent studies have indicated that these compounds are markedly toxic to Chironomidae, a widespread family of ecologically important aquatic insects. However, despite their sensitivity, the pharmacological mechanisms driving neonicotinoid toxicity have yet to be characterized in these insect species. Here, we used a combination of saturation and competition binding studies to characterize neonicotinoid binding properties to nicotinic acetylcholine receptors (nAChR) in two different Chironomidae (Chironomus riparius and Chironomus dilutus) at two different life stages (larval and adult). Using radiolabeled imidacloprid ([3H]-IMI), we characterized and compared receptor density (Bmax), imidacloprid binding affinity (KD), and receptor binding affinity (Ki) to three different neonicotinoid competitors (imidacloprid, clothianidin, and thiamethoxam). We then compared receptor density and binding affinity parameters derived for Chironomidae to data previously generated for other dipterans and agricultural pests. We found that there were limited differences in neonicotinoid binding between C. riparius and C. dilutus, with both organisms demonstrating high affinities for imidacloprid (KD = 0.22−0.87 nM) and high receptor densities (Bmax = 0.92–6.53 pmol/mg). However, there were significant differences between life-stages, with larvae expressing higher densities of nicotinic acetylcholine receptors and higher imidacloprid affinities than adults. Moreover, there were compound-specific differences in receptor affinity, with larval stages displaying relative affinities (Ki) that generally correlated with acute neonicotinoid toxicity (e.g. clothianidin ≥ imidacloprid >>> thiamethoxam). Finally, compared to other dipterans and agricultural pests, Chironomidae display very high densities of high affinity nicotinic acetylcholine receptors, which likely contribute to their sensitivity. Results indicated that receptor-level differences in neonicotinoid binding may be responsible for ecotoxicological differences amongst distinct insect species, and they likely play a role in life stage-, and compound-level toxicity differences previously observed for Chironomidae. Overall, this study highlights the value of understanding the toxicological mechanisms of action of neonicotinoids in sensitive, non-target aquatic insects, to better predict adverse effects associated with unintentional neonicotinoid exposure.

Toxicity of the insecticides spinosad and indoxacarb to the non-target aquatic midge Chironomus riparius

Spinosad and indoxacarb are two relatively new insecticides mainly used in agriculture to control insect pests. However, at their current application rates, non-target aquatic insect species may also be impacted. In this study, larvae of the non-biting midge Chironomus riparius were exposed in the laboratory to both insecticides and their effects evaluated at the organismal level, using standard ecotoxicological tests, and at the biochemical level, by monitoring specific oxidative stress, neuronal, and energy metabolism biomarkers. Chronic exposure to both insecticides compromised growth and emergence of C. riparius. Short-term exposures revealed alterations at biochemical level that might be related to the toxicological targets of both insecticides. Growth and development time were the most sensitive endpoints at individual level for both pesticides, while at the biochemical level, the electron transport system activity was the most sensitive biomarker for spinosad exposure, suggesting an increase in energy demands associated with the activation of defense mechanisms. Glutathione-S-transferase was the most sensitive biomarker for indoxacarb exposure, underlining the role of this enzyme in the detoxification of indoxacarb. Additionally, changes in lactate dehydrogenase and glutathione peroxidase activities were observed for both insecticides, and evidences of oxidative damage were found for spinosad. This study contributes to the growing knowledge on sublethal effects of novel insecticides on non-target aquatic invertebrates and strengthens the usefulness of biochemical biomarkers to support the interpretation of their potentially deleterious effects on aquatic insects near agricultural fields.

Lethal and sublethal toxicity of neonicotinoid and butenolide insecticides to the mayfly, Hexagenia spp.

Neonicotinoid insecticides are environmentally persistent and highly water-soluble, and thus are prone to leaching into surface waters where they may negatively affect non-target aquatic insects. Most of the research to date has focused on imidacloprid, and few data are available regarding the effects of other neonicotinoids or their proposed replacements (butenolide insecticides). The objective of this study was to assess the toxicity of six neonicotinoids (imidacloprid, thiamethoxam, acetamiprid, clothianidin, thiacloprid, and dinotefuran) and one butenolide (flupyradifurone) to Hexagenia spp. (mayfly larvae). Acute (96-h), water-only tests were conducted, and survival and behaviour (number of surviving mayflies inhabiting artificial burrows) were assessed. Acute sublethal tests were also conducted with imidacloprid, acetamiprid, and thiacloprid, and in addition to survival and behaviour, mobility (ability to burrow into sediment) and recovery (survival and growth following 21 d in clean sediment) were measured. Sublethal effects occurred at much lower concentrations than survival: 96-h LC50s ranged from 780 μg/L (acetamiprid) to >10,000 μg/L (dinotefuran), whereas 96-h EC50s ranged from 4.0 μg/L (acetamiprid) to 630 μg/L (thiamethoxam). Flupyradifurone was intermediate in toxicity, with a 96-h LC50 of 2000 μg/L and a 96-h EC50 of 81 μg/L. Behaviour and mobility were impaired significantly and to a similar degree in sublethal exposures to 10 μg/L imidacloprid, acetamiprid, and thiacloprid, and survival and growth following the recovery period were significantly lower in mayflies exposed to 10 μg/L acetamiprid and thiacloprid, respectively. A suite of effects on mayfly swimming behaviour/ability and respiration were also observed, but not quantified, following exposures to imidacloprid, acetamiprid, and thiacloprid at 1 μg/L and higher. Imidacloprid concentrations measured in North American surface waters have been found to meet or exceed those causing toxicity to Hexagenia, indicating that environmental concentrations may adversely affect Hexagenia and similarly sensitive non-target aquatic species.

Exposure to chlorantraniliprole affects the energy metabolism of the caddisfly Sericostoma vittatum.

Caddisflies have been included in ecotoxicological studies because of their sensitivity and ecological relevance. The present study aimed to assess the sublethal effects of an anthranilic diamide insecticide, chlorantraniliprole (CAP), to Sericostoma vittatum. Used worldwide, CAP is a persistent compound that has been found in surface waters at concentrations from 0.1 μg/L to 9.7 μg/L. It targets the ryanodine receptors, and the present ecotoxicological assessment focused on biomarkers related to neurotransmission, biotransformation, oxidative stress damage, and endpoints related to energy processing (feeding, energy reserves, and cellular metabolism). Six days of exposure trials revealed that feeding activity was significantly decreased in S. vittatum larvae exposed to 0.9 μg/L CAP. Concomitantly, a reduction in cellular metabolism and a significant decrease in protein content were also observed in caddisfly larvae exposed to CAP, suggesting metabolic depression. The results show that sublethal concentrations of CAP can cause detrimental sublethal effects on S. vittatum total glutathione content at concentrations as low as 0.2 μg/L. Bioenergetics can be used to assess physiological effects of contaminants, and the present results show that exposure to low, environmentally relevant, concentrations of CAP alter energy acquisition and metabolism in nontarget aquatic insects with potential population level effects. Environ Toxicol Chem 2017;36:1584-1591. © 2016 SETAC.

Impact of Terminalia chebula Retz. against Aedes aegypti L. and non-target aquatic predatory insects

Aedes aegypti Linn is one of the most important mosquito species. The vectors are responsible for causing deadly diseases like dengue and dengue hemorrhagic fever. Several chemical pesticides used to control these dengue vectors caused severe toxic significances on human health and other non-target beneficial insects. Therefore the current investigation has been made to access the bio-efficacy of the crude seed extracts of T. chebula against the dengue vector Ae. aegypti. The GC-MS analysis of crude seed extracts of T. chebula identified nine chemical compounds with major peak area in the 1,2,3-Benzenetriol (61.96%), followed by Tridecanoic acid (09.55%). Ae. aegypti larvae showed dose dependent mortality rate was observed between the treatments. Prominent protection rate at greater concentrations of 100ppm and moderate protection at 75 and 50ppm was observed in the repellent assay. Lethal concentration (LC50 and LC90) of fourth instar larvae of Ae. aegypti was observed in 138 and 220ppm concentration respectively. Similarly, the seed extracts showed 100% adulticidal activity at the concentration of 400ppm at 30min of exposure time. Phytochemicals present in the seed extracts of T. chebula significantly affects the major portions of the midgut tissues of Ae. aegypti at the concentration of 100ppm. The toxicological evaluation of seed extracts also proved non-toxic towards the A. bouvieri and Tx. splendens aquatic predatory insects. Hence, the present result suggest that bio-rational plant derived T. chebula could be incorporated in the dengue vector control and have no adverse effects on non-target beneficial insects.

Effect of seed kernel aqueous extract from Annona squamosa against three mosquito vectors and its impact on non-target aquatic organisms

ObjectiveTo evaluate the toxicity of Annona squamosa (A. squamosa) aqueous (physiological saline) seed soluble extract and its control of mosquito population. MethodsOvicidal, larvicidal and pupicidal activity of A. squamosa crude soluble seed kernel extract was determined according to World Health Organization. The mortality of each mosquito stage was recorded after 24 h exposured to plant material. Toxicity assay was used to assess the non-target organisms with different concentrations according to Organisation for Economic Co-operation and Development. ResultsThe aqueous solubilized extracts of A. squamosa elicit the toxicity against all stages of Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus, and the LC50 values against stages of egg, 1st-4th larvae were (1.45 and 1.26–2.5 mg/mL), (1.12 and 1.19–2.81 mg/mL) and (1.80 and 2.12–3.41 mg/mL) respectively. The pupicidal activity also brought forth amended activity against all three mosquitoes species, and the LC50 values were consider to be 3.19, 2.42 and 4.47 mg/mL. Ultimately there was no mortality observed from non-target organism of Chironomus costatus. ConclusionsBased on the findings of the study, it suggests that the use of A. squamosa plant extract can act as an alternate insecticidal agents for controlling target mosquitoes without affecting the non-target aquatic insect. Further investigation to identify the active compounds and their mechanisms of action is recommended.

Comparative chronic toxicity of imidacloprid, clothianidin, and thiamethoxam to Chironomus dilutus and estimation of toxic equivalency factors.

Nontarget aquatic insects are susceptible to chronic neonicotinoid insecticide exposure during the early stages of development from repeated runoff events and prolonged persistence of these chemicals. Investigations on the chronic toxicity of neonicotinoids to aquatic invertebrates have been limited to a few species and under different laboratory conditions that often preclude direct comparisons of the relative toxicity of different compounds. In the present study, full life-cycle toxicity tests using Chironomus dilutus were performed to compare the toxicity of 3 commonly used neonicotinoids: imidacloprid, clothianidin, and thiamethoxam. Test conditions followed a static-renewal exposure protocol in which lethal and sublethal endpoints were assessed on days 14 and 40. Reduced emergence success, advanced emergence timing, and male-biased sex ratios were sensitive responses to low-level neonicotinoid exposure. The 14-d median lethal concentrations for imidacloprid, clothianidin, and thiamethoxam were 1.52 μg/L, 2.41 μg/L, and 23.60 μg/L, respectively. The 40-d median effect concentrations (emergence) for imidacloprid, clothianidin, and thiamethoxam were 0.39 μg/L, 0.28 μg/L, and 4.13 μg/L, respectively. Toxic equivalence relative to imidacloprid was estimated through a 3-point response average of equivalencies calculated at 20%, 50%, and 90% lethal and effect concentrations. Relative to imidacloprid (toxic equivalency factor [TEF] = 1.0), chronic (lethality) 14-d TEFs for clothianidin and thiamethoxam were 1.05 and 0.14, respectively, and chronic (emergence inhibition) 40-d TEFs were 1.62 and 0.11, respectively. These population-relevant endpoints and TEFs suggest that imidacloprid and clothianidin exert comparable chronic toxicity to C. dilutus, whereas thiamethoxam induced comparable effects only at concentrations an order of magnitude higher. However, the authors caution that under field conditions, thiamethoxam readily degrades to clothianidin, thereby likely enhancing toxicity. Environ Toxicol Chem 2017;36:372-382. © 2016 SETAC.

Sensitivity of the early-life stages of freshwater mollusks to neonicotinoid and butenolide insecticides

Neonicotinoid insecticides can be transported from agricultural fields, where they are used as foliar sprays or seed treatments, to surface waters by surface or sub-surface runoff. Few studies have investigated the toxicity of neonicotinoid or the related butenolide insecticides to freshwater mollusk species. The current study examined the effect of neonicotinoid and butenolide exposures to the early-life stages of the ramshorn snail, Planorbella pilsbryi, and the wavy-rayed lampmussel, Lampsilis fasciola. Juvenile P. pilsbryi were exposed to imidacloprid, clothianidin, or thiamethoxam for 7 or 28 d and mortality, growth, and biomass production were measured. The viability of larval (glochidia) L. fasciola was monitored during a 48 h exposure to six neonicotinoids (imidacloprid, thiamethoxam, clothianidin, acetamiprid, thiacloprid, or dinotefuran), or a butenolide (flupyradifurone). The 7-d LC50s of P. pilsbryi for imidacloprid, clothianidin, and thiamethoxam were ≥4000 μg/L and the 28-d LC50s were ≥182 μg/L. Growth and biomass production were considerably more sensitive endpoints than mortality with EC50s ranging from 33.2 to 122.0 μg/L. The 48-h LC50s for the viability of glochidia were ≥456 μg/L for all seven insecticides tested. Our data indicate that neonicotinoid and butenolide insecticides pose less of a hazard with respect to mortality of the two species of mollusk compared to the potential hazard to other non-target aquatic insects.

Toxicological effects of pyrethroids on non-target aquatic insects.

The toxicological effects of pyrethroids on non-target aquatic insects are mediated by several modes of entry of pyrethroids into aquatic ecosystems, as well as the toxicological characteristics of particular pyrethroids under field conditions. Toxicokinetics, movement across the integument of aquatic insects, and the toxicodynamics of pyrethroids are discussed, and their physiological, symptomatic and ecological effects evaluated. The relationship between pyrethroid toxicity and insecticide uptake is not fully defined. Based on laboratory and field data, it is likely that the susceptibility of aquatic insects (vector and non-vector) is related to biochemical and physiological constraints associated with life in aquatic ecosystems. Understanding factors that influence aquatic insects susceptibility to pyrethroids is critical for the effective and safe use of these compounds in areas adjacent to aquatic environments.

Effects of extract of soapnut Sapindus emarginatus on esterases and phosphatases of the vector mosquito, Aedes aegypti (Diptera: Culicidae)

Our earlier investigations with kernels from the soapnut Sapindus emarginatus revealed it as a new source of botanical biocide with potent antimosquito activity, as evident from the proven unique ability of the aqueous kernel extract to kill all the developmental stages of three important vector mosquito species, Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus. This extract was also found to be safe for two non-target aquatic insects. As a sequel to these findings, we have further examined quantitative and qualitative changes in total proteins, esterases, and phosphatases in whole body homogenates of fourth instar larvae and pupae of A. aegypti exposed to this extract at an appropriate threshold time for its lethal effect to gain insights into the impact of the botanical biocide on biochemical characteristics of the target vector mosquito at two distinct developmental stages. The profiles of proteins, esterases (acetylcholinesterse, α- and β-carboxylesterases), and phosphatases (acid and alkaline) exhibited distinct patterns of variation during normal development of fourth instar larvae and pupae, indicating intrinsic difference in biochemical features between these two developmental stages of A. aegypti. Upon exposure of the larvae to the extract, significant reduction in the activities of acetylcholinesterse, β-carboxylesterase, and acid phosphatases were recorded, whereas the total proteins, α-carboxylesterase and alkaline phosphatase activities were unaffected. By contrast, only alkaline phosphatase activity was significantly affected in pupae exposed to the extract. Analysis of these enzymes in native PAGE revealed that they exist in isoforms in both the larvae and pupae. The alterations in the levels of enzymatic activities observed from the quantitative assays of various enzymes were reflected by the respective zymograms with perceptible differences in the intensity and the number of bands detected especially with β-carboxylesterase, acid and alkaline phosphatase activity between the control and exposed test organisms. Despite the fact that the soapnut kernel extract causes mortality of both the larvae and pupae of A. aegypti, the findings of this study demonstrate that the impact of this extract is most pronounced in various enzyme profiles of the larvae rather than the pupae. Such discrepancy implicates the presence of unique biochemical mechanisms in the pupae of mosquito for detoxification of botanical biocides.

Antimosquito activity of aqueous kernel extract of soapnut Sapindus emarginatus: impact on various developmental stages of three vector mosquito species and nontarget aquatic insects

Aqueous (physiological saline) extracts of seed kernel from seven indigenous plants were initially screened for their antimosquito activity against eggs, larvae of all instars, and pupae of Aedes aegypti. Among various seed kernels tested, the soapnut Sapindus emarginatus (Sapindaceae) extract was found to exhibit, for the first time, a strong antimosquito activity as evident from its ability to inflict 100% mortality of all the developmental stages of A. aegypti. Furthermore, the kernel extract of S. emarginatus also exerted ovicidal, larvicidal, and pupicidal activity against two other important vector mosquitoes, namely, Anopheles stephensi and Culex quinquefasciatus. Differential susceptibility of the various developmental stages of the three mosquito species exposed to soapnut extract was also noticed. The kernel extract was found to be safe for two nontarget aquatic insects tested: the larvae of Chironomus costatus and the nymphs of Diplonychus rusticus. Lethal concentration values of soapnut extract to these nontarget insects were always threefold or fivefold higher than those that produced 100% mortality of the larvae of the three mosquito species examined. The findings of this study clearly demonstrate that the aqueous kernel extract of S. emarginatus has potent antimosquito activity detectable against all the developmental stages of three important vector mosquito species as well as it is safe for nontarget aquatic organisms, and thus this new botanical resource could be used as an eco-friendly alternative biocidal agent in control of mosquitoes.

Mechanistic Approach to Understanding the Toxicity of the Azole Fungicide Triadimefon to a Nontarget Aquatic Insect and Implications for Exposure Assessment

Mechanistic and stereoselective based in vitro metabolism assays were utlilized to gain insight into the toxic mode of action of the 1,2,4-triazole fungicide, triadimefon, with black fly (Diptera: Simuliidae) larvae. Based on results from enzyme inhibitor studies, the metabolism of triadimefon in black fly larvae microsomes was found to occur predominantly via an oxidative P450-mediated pathway; triadimenol was formed via the stereoselective reduction of the prochiral carbonyl group of triadimefon. The relatively minor contribution of carbonyl reduction suggests that triadimefon may inhibit ecdysone 20-monooxygenase and disrupt insect molting hormone biosynthesis. 48-h LC50 tests for triadimefon and triadimenol with black fly larvae yielded median values (with 95% confidence intervals) of 6.1 (5.8-6.4) and 22.3 (20.3-24.1) mg/L respectively. The exposure of black fly larvae to sublethal concentrations of triadimefon resulted in increased microsomal P450 activity and affected the microsomal rates of both triadimefon depletion and triadimenol formation. In contrast to trout, black fly larvae produced a higher fraction of the more toxic triadimenol stereoisomers, which may explain in part why triadimefon exhibited a significantly greater toxicity with black fly larvae than trout. These results illustrate that while LC50 tests conducted with commercial triadimenol would presumably expose each organism to the same relative abundance of the four triadimenol stereoisomers, LC50 tests with triadimefon ultimately expose each organism to a unique set of triadimenol stereoisomers depending upon the organism's stereoselective metabolism.

Acute lethal and sublethal effects of a neem-based insecticide on nontarget aquatic insects in stream channels.

Extracts from seeds and leaves of the neem tree, Azadirachta indica, express insecticidal activity against a broad range of insect pests (Schmutterer and Singh 1995). The primary active ingredient in neem extracts is the triterpenoid, azadirachtin. Because several common forest pests are among the most susceptible insects to azadirachtin, the use of neem-based insecticides in Canadian forest pest management programs is being investigated (Isman et al. 1991; Helson et al. 1999). Insecticide applications to forests may result in some of the product entering nearby water bodies and it is therefore important to ensure that these insecticides do not post a risk of adverse effects on aquatic organisms. Small streams are particularly susceptible to contamination by runoff from adjacent sprayed areas because of low dilution potential, and from inadvertent overspray because they are difficult to avoid during aerial applications. We investigated the potential for adverse effects on stream insects by applying a neem-based insecticide to outdoor stream channels and measuring acute lethal and sublethal responses.

Effect of an Azadirachtin Formulation on Six Nontarget Aquatic Macroinvertebrates

Effects of a botanical pesticide, azadirachtin, were evaluated on nontarget aquatic macroinvertebrates found in SW Montana. A neem product formulated for commercial use and its petroleum-based, inert carrier alone were used to test the hypothesis that a commercial, U.S. Environmental Protection Agency-approved formulation of the pesticide, azadirachtin-A, derived fromextractsofneemtreeseeds, Azadirachtaindica A.Juss.(Family:Meliaceae),istoxictonontarget aquatic macroinvertebrates in Montana river systems. LC50s (24 h) for 5 insect species, Drunella grandis (McDunnough), Drunella doddsi (Needham), Skwala parallela (Frison), Brachycentrus occidentalis Banks, and Brachycentrus americanus (Banks), and 1 isopod species, Caecidotea inter- media (Forbes) varied from 1.8 to 9.2 ppm of the full formulation. The full formulation consisted of: azadirachtin (3% wt:wt); other neem-extracted material (32% wt:wt); petroleum solvents (65% wt:wt). LT50s for 4 ppm of this formulation were 9.4 to 748 hours for these field-collected macro- invertebrates. Without azadirachtin, but with the petroleum-based carrier, LT50s were 13.6-46.7 h. These bioassays conducted late in winter and early spring of 1994 and 1995 and late summer and early fall of 1994 indicated mortality from the neem formulation was similar to herbicides 2,4-D and picloram, but mortality could be accounted for by action of petroleum solvents in the inert materials. This suggests the registration process of new pesticides should also include documented potential toxicity of inert components in the formulation to non-target aquatic insects.