Reduced-risk insecticides in Neotropical stingless bee species: impact on survival and activity

Background: As honeybees are the main pollinator species subject to an intense research regarding effects of pesticides, other ecologically important native bee pollinators have received little attention in ecotoxicology and risk assessment of pesticides in general, and insecticides in particular, some of which are perceived as reduced-risk compounds. Here the impact of three reduced-risk insecticides – azadirachtin, spinosad, and chlorantraniliprole – was assessed in two species of stingless bees, Partamona helleri and Scaptotrigona xanthotrica, which are important native pollinators in Neotropical America. The neonicotinoid imidacloprid was used as a positive control.Results: Spinosad exhibited high oral and contact toxicities in adult workers of both species at the recommended label rates, with median survival times (LT50s) ranging from 1 to 4 h, whereas these estimates were below 15 min for imidacloprid. Azadirachtin and chlorantraniliprole exhibited low toxicity at the recommended label rates, with negligible mortality that did not allow LT50 estimation. Sublethal behavioral assessments of these two insecticides indicated that neither one of them affected the overall group activity of workers of the two species. However, both azadirachtin and chlorantraniliprole impaired individual flight take-off of P. helleri and S. xanthotrica worker bees, which may compromise foraging activity, potentially leading to reduced colony survival.Conclusion: These findings challenge the common perception of non-target safety of reduced-risk insecticides and bioinsecticides, particularly regarding native pollinator species.Keywords: behavioral impact; biopesticides; colony and individual level effects; native bee pollinators; sublethal effect

Imidacloprid-Induced Impairment of Mushroom Bodies and Behavior of the Native Stingless Bee Melipona quadrifasciata anthidioides

Declines in pollinator colonies represent a worldwide concern. The widespread use of agricultural pesticides is recognized as a potential cause of these declines. Previous studies have examined the effects of neonicotinoid insecticides such as imidacloprid on pollinator colonies, but these investigations have mainly focused on adult honey bees. Native stingless bees (Hymenoptera: Apidae: Meliponinae) are key pollinators in neotropical areas and are threatened with extinction due to deforestation and pesticide use. Few studies have directly investigated the effects of pesticides on these pollinators. Furthermore, the existing impact studies did not address the issue of larval ingestion of contaminated pollen and nectar, which could potentially have dire consequences for the colony. Here, we assessed the effects of imidacloprid ingestion by stingless bee larvae on their survival, development, neuromorphology and adult walking behavior. Increasing doses of imidacloprid were added to the diet provided to individual worker larvae of the stingless bee Melipona quadrifasciata anthidioides throughout their development. Survival rates above 50% were only observed at insecticide doses lower than 0.0056 µg active ingredient (a.i.)/bee. No sublethal effect on body mass or developmental time was observed in the surviving insects, but the pesticide treatment negatively affected the development of mushroom bodies in the brain and impaired the walking behavior of newly emerged adult workers. Therefore, stingless bee larvae are particularly susceptible to imidacloprid, as it caused both high mortality and sublethal effects that impaired brain development and compromised mobility at the young adult stage. These findings demonstrate the lethal effects of imidacloprid on native stingless bees and provide evidence of novel serious sublethal effects that may compromise colony survival. The ecological and economic importance of neotropical stingless bees as pollinators, their susceptibility to insecticides and the vulnerability of their larvae to insecticide exposure emphasize the importance of studying these species

Bti-based insecticide enhances the predatory abilities of the backswimmer Buenoa tarsalis (Hemiptera: Notonectidae)

The backswimmer Buenoa tarsalis (Hemiptera: Notonectidae) is a naturally occurring predator of immature stages of mosquitoes. These aquatic predators can suffer from non-targeted exposure to insecticides that are commonly used in aquatic environments to control mosquitoes. Here, we evaluated whether insecticide formulations containing the bacterium Bacillus thuringiensis var. israelensis (Bti) or the organophosphate pirimiphos-methyl would affect the survival and the predatory abilities of B. tarsalis. First, we conducted survival bioassays to estimate the median survival time (LT50) of B. tarsalis when exposed to Bti-based insecticide (at 0.25 and 25 mg a.i./L) and pirimiphos-methyl (at 1, 10 and 1000 mg a.i./L). The highest concentrations of the insecticides were equivalent to the label-recommended field rates. Second, the predatory abilities of B. tarsalis exposed to insecticides were evaluated at three prey densities (3, 6 and 9 mosquito larvae/100 mL water) just after insecticide exposure or after a 24 h recovery time. While the survival of B. tarsalis was significantly reduced with pirimiphos-methyl concentrations ≥10 mg a.i./L, the Bti-exposed predators exhibited similar survival as unexposed predators. Interestingly, after a recovery time of 24 h, B. tarsalis sublethally exposed to pirimiphos-methyl or Bti-based insecticide consistently killed more A. aegypti larvae (at the intermediate density) than unexposed predators. However, for the without-recovery bioassays, the pirimiphos-methyl-exposed predators exhibited reduced predatory abilities at the lowest prey density. Because they do not reduce the survival or the predatory abilities of B. tarsalis, Bti-based insecticides can be considered a safe insecticide to use in the presence of backswimmers

Survival and swimming behavior of insecticide-exposed larvae and pupae of the yellow fever mosquito Aedes aegypti

The yellow fever mosquito Aedes aegypti is essentially a container-inhabiting species that is closely associated with urban areas. This species is a vector of human pathogens, including dengue and yellow fever viruses, and its control is of paramount importance for disease prevention. Insecticide use against mosquito juvenile stages (i.e. larvae and pupae) is growing in importance, particularly due to the ever-growing problems of resistance to adult-targeted insecticides and human safety concerns regarding such use in human dwellings. However, insecticide effects on insects in general and mosquitoes in particular primarily focus on their lethal effects. Thus, sublethal effects of such compounds in mosquito juveniles may have important effects on their environmental prevalence. In this study, we assessed the survival and swimming behavior of A. aegypti 4th instar larvae (L4) and pupae exposed to increasing concentrations of insecticides. We also assessed cell death in the neuromuscular system of juveniles.Third instar larvae of A. aegypti were exposed to different concentrations of azadirachtin, deltamethrin, imidacloprid and spinosad. Insect survival was assessed for 10 days. The distance swam, the resting time and the time spent in slow swimming were assessed in 4th instar larvae (L4) and pupae. Muscular and nervous cells of L4 and pupae exposed to insecticides were marked with the TUNEL reaction. The results from the survival bioassays were subjected to survival analysis while the swimming behavioral data were subjected to analyses of covariance, complemented with a regression analysis. All insecticides exhibited concentration-dependent effects on survival of larvae and pupae of the yellow fever mosquito. The pyrethroid deltamethrin was the most toxic insecticide followed by spinosad, imidacloprid, and azadirachtin, which exhibited low potency against the juveniles. All insecticides except azadirachtin reduced L4 swimming speed and wriggling movements. A similar trend was also observed for swimming pupa, except for imidacloprid, which increased the swimming activity of pupa. Curiously, the insecticides did not affect cell damage in the neuromuscular system of larvae and pupae. Deltamethrin and spinosad were the main compounds to exhibit lethal effects, which allowed the control of A. aegypti larvae and pupae, and impair their swimming potentially compromising foraging and predation likelihood

Spinosad in the native stingless bee Melipona quadrifasciata: regrettable non-target toxicity of a bioinsecticide

The risks imposed by novel insecticides, mainly bioinsecticides, are largely unknown despite their increased use and their perceived environmental safety, which is based on their natural origin. Furthermore, unlike honeybees, native pollinator species have received little attention. In the present study, the lethal and sublethal effects of the neonicotinoid imidacloprid and the bioinsecticide spinosad were assessed in the stingless bee species Melipona quadrifasciata, an important native pollinator in the Neotropical region. The adult stingless bee workers exhibited high oral insecticide susceptibility, with LD50s of 23.54 and 12.07 ng a.i./bee for imidacloprid and spinosad, respectively. Imidacloprid also impaired worker respiration and overall group activity and flight, while spinosad significantly impaired only worker flight despite exhibiting higher oral toxicity to adult workers than imidacloprid. These findings indicate the hazardous nature not only of imidacloprid but also the bioinsecticide spinosad to adult workers of the native pollinator M. quadrifasciata. Therefore, bioinsecticides should not be exempted from risk assessment analysis due to their lethal and sublethal components

Agrochemical synergism imposes higher risk to Neotropical bees than to honeybees

Bees are key pollinators whose population numbers are declining, in part, owing to the effects of different stressors such as insecticides and fungicides. We have analysed the susceptibility of the Africanized honeybee, Apis mellifera, and the stingless bee, Partamona helleri, to commercial formulations of the insecticides deltamethrin and imidacloprid. The toxicity of fungicides based on thiophanate-methyl and chlorothalonil were investigated individually and in combination, and with the insecticides. Results showed that stingless bees were more susceptible to insecticides than honeybees. The commercial fungicides thiophanate-methyl or chlorothalonil caused low mortality, regardless of concentration; however, their combination was as toxic as imidacloprid to both species, and over 400-fold more toxic than deltamethrin for A. mellifera. There were highly synergistic effects on mortality caused by interactions in the mixture of imidacloprid and the fungicides thiophanate-methyl, chlorothalonil and the combined fungicide formulation in A. mellifera, and also to a lesser extent in P. helleri. By contrast, mixtures of the deltamethrin and the combined fungicide formulation induced high synergy in P. helleri, but had little effect on the mortality of A. mellifera. Differences in physiology and modes of action of agrochemicals are discussed as key factors underlying the differences in susceptibility to agrochemicals

Detection of a new pyrethroid resistance mutation (V410L) in the sodium channel of Aedes aegypti: a potential challenge for mosquito control

The yellow fever mosquito, Aedes aegypti, particularly in Neotropical regions, is the principal vector of dengue, yellow fever, Zika and Chikungunya viruses. Pyrethroids remain one of the most used insecticides to control Aedes mosquitoes, despite the development of pyrethroid resistance in many mosquito populations worldwide. Here, we report a Brazilian strain of A. aegypti with high levels (approximately 100–60,000 fold) of resistance to both type I and type II pyrethroids. We detected two mutations (V410L and F1534C) in the sodium channel from this resistant strain. This study is the first report of the V410L mutation in mosquitoes. Alone or in combination with the F1534C mutation, the V410L mutation drastically reduced the sensitivity of mosquito sodium channels expressed in Xenopus oocytes to both type I and type II pyrethroids. The V410L mutation presents a serious challenge for the control of A. aegypti and will compromise the use of pyrethroids for the control of A. aegypti in Brazil; therefore, early monitoring of the frequency of the V410L mutation will be a key resistance management strategy to preserve the effectiveness of pyrethroid insecticides

Insecticide resistance and size assortative mating in females of the maize weevil (Sitophilus zeamais)

Random mating is a common assumption in studies of insecticide resistance evolution, but seldom tested despite its potential consequences. Therefore, the existing evidence of female choice and insecticide resistance in populations of the maize weevil (Sitophilus zeamais), a key pest of stored cereals, led to the assessment of mating preferences and their association with insecticide resistance in this species. Mixed lines of a maize weevil colony were established from field‐collected populations, which after 5 months of natural breeding were selected for deltamethrin resistance for five generations, reaching over 100‐fold resistance. Mating preference was significantly based on the partner size, measured as body mass (χ2 = 5.83, df = 1, P = 0.016). Susceptible females preferred heavier males for mating (χ2 = 5.83, df = 1, P = 0.015), a trait that was more frequently associated with deltamethrin resistance (χ2 = 7.38, df = 1, P = 0.007). Deltamethrin resistance compromised daily fertility, although the reduced offspring production observed in matings between susceptible females and resistant males was negligible. Susceptible female weevils prefer larger (and heavier) males to mate, a trait associated with deltamethrin resistance, favouring the maintenance and spread of the resistant phenotype in the population

Volume of the mushroom bodies in the brains of the stingless bee workers (<i>Melipona quadrifasciata anthidioides</i>) reared on imidacloprid-contaminated diets during the larval period.

<p>The symbols represent means and standard error.</p

Survival plots of stingless bee workers (<i>Melipona quadrifasciata anthidioides</i>) reared on imidacloprid-contaminated diets containing increasing doses of the insecticide.

<p>The survival curves of workers bees exposed to imidaclorpid doses between 0.28 e 28 µg a.i./bee were not significantly different and were therefore coded with the same color (i.e., blue).</p