RFID tracking of sublethal effects of two neonicotinoid insecticides on the foraging behavior of Apis mellifera

The development of insecticides requires valid risk assessment procedures to avoid causing harm to beneficial insects and especially to pollinators such as the honeybee Apis mellifera. In addition to testing according to current guidelines designed to detect bee mortality, tests are needed to determine possible sublethal effects interfering with the animal's vitality and behavioral performance. Several methods have been used to detect sublethal effects of different insecticides under laboratory conditions using olfactory conditioning. Furthermore, studies have been conducted on the influence insecticides have on foraging activity and homing ability which require time-consuming visual observation. We tested an experimental design using the radiofrequency identification (RFID) method to monitor the influence of sublethal doses of insecticides on individual honeybee foragers on an automated basis. With electronic readers positioned at the hive entrance and at an artificial food source, we obtained quantifiable data on honeybee foraging behavior. This enabled us to efficiently retrieve detailed information on flight parameters. We compared several groups of bees, fed simultaneously with different dosages of a tested substance. With this experimental approach we monitored the acute effects of sublethal doses of the neonicotinoids imidacloprid (0.15-6 ng/bee) and clothianidin (0.05-2 ng/bee) under field-like circumstances. At field-relevant doses for nectar and pollen no adverse effects were observed for either substance. Both substances led to a significant reduction of foraging activity and to longer foraging flights at doses of ≥0.5 ng/bee (clothianidin) and ≥1.5 ng/bee (imidacloprid) during the first three hours after treatment. This study demonstrates that the RFID-method is an effective way to record short-term alterations in foraging activity after insecticides have been administered once, orally, to individual bees. We contribute further information on the understanding of how honeybees are affected by sublethal doses of insecticides.     

Exposure to sublethal doses of fipronil and thiacloprid highly increases mortality of honeybees previously infected by Nosema ceranae

BACKGROUND: The honeybee, Apis mellifera, is undergoing a worldwide decline whose origin is still in debate. Studies performed for twenty years suggest that this decline may involve both infectious diseases and exposure to pesticides. Joint action of pathogens and chemicals are known to threaten several organisms but the combined effects of these stressors were poorly investigated in honeybees. Our study was designed to explore the effect of Nosema ceranae infection on honeybee sensitivity to sublethal doses of the insecticides fipronil and thiacloprid. METHODOLOGY/FINDING: Five days after their emergence, honeybees were divided in 6 experimental groups: (i) uninfected controls, (ii) infected with N. ceranae, (iii) uninfected and exposed to fipronil, (iv) uninfected and exposed to thiacloprid, (v) infected with N. ceranae and exposed 10 days post-infection (p.i.) to fipronil, and (vi) infected with N. ceranae and exposed 10 days p.i. to thiacloprid. Honeybee mortality and insecticide consumption were analyzed daily and the intestinal spore content was evaluated 20 days after infection. A significant increase in honeybee mortality was observed when N. ceranae-infected honeybees were exposed to sublethal doses of insecticides. Surprisingly, exposures to fipronil and thiacloprid had opposite effects on microsporidian spore production. Analysis of the honeybee detoxification system 10 days p.i. showed that N. ceranae infection induced an increase in glutathione-S-transferase activity in midgut and fat body but not in 7-ethoxycoumarin-O-deethylase activity. CONCLUSIONS/SIGNIFICANCE: After exposure to sublethal doses of fipronil or thiacloprid a higher mortality was observed in N. ceranae-infected honeybees than in uninfected ones. The synergistic effect of N. ceranae and insecticide on honeybee mortality, however, did not appear strongly linked to a decrease of the insect detoxification system. These data support the hypothesis that the combination of the increasing prevalence of N. ceranae with high pesticide content in beehives may contribute to colony depopulation.     

拟除虫菊酯类杀虫剂对蜜蜂的毒性和影响

蜜蜂是最重要的农业授粉昆虫之一,蜜蜂在授粉过程中极有可能接触到广泛使用的广谱杀虫剂-拟除虫菊酯,大多数拟除虫菊酯对蜜蜂等农业授粉昆虫有较高的毒性.本文对拟除虫菊酯类杀虫剂的作用机理进行了综述;总结了蜂群及蜂产品中拟除虫菊酯类杀虫剂的残留现状、拟除虫菊酯对蜜蜂的急性毒性以及亚致死效应,讨论了拟除虫菊酯类杀虫剂复配农药对蜜...     

Effects of a neonicotinoid pesticide on thermoregulation of African honey bees (Apis mellifera scutellata)

Thiamethoxam is a widely used neonicotinoid pesticide that, as agonist of the nicotinic acetylcholine receptors, has been shown to elicit a variety of sublethal effects in honey bees. However, information concerning neonicotinoid effects on honey bee thermoregulation is lacking. Thermoregulation is an essential ability for the honey bee that guarantees the success of foraging and many in-hive tasks, especially brood rearing. We tested the effects of acute exposure to thiamethoxam (0.2, 1, 2 ng/bee) on the thorax temperatures of foragers exposed to low (22 °C) and high (33 °C) temperature environments. Thiamethoxam significantly altered honey bee thorax temperature at all doses tested; the effects elicited varied depending on the environmental temperature and pesticide dose to which individuals were exposed. When bees were exposed to the high temperature environment, the high dose of thiamethoxam increased their thorax temperature 1–2 h after exposure. When bees were exposed to the low temperature, the higher doses of the neonicotinoid reduced bee thorax temperatures 60–90 min after treatment. In both experiments, the neonicotinoid decreased the temperature of bees the day following the exposure. After a cold shock (5 min at 4 °C), the two higher doses elicited a decrease of the thorax temperature, while the lower dose caused an increase, compared to the control. These alterations in thermoregulation caused by thiamethoxam may affect bee foraging activity and a variety of in-hive tasks, likely leading to negative consequences at the colony level. Our results shed light on sublethal effect of pesticides which our bees have to deal with.     

Bees exposed to climate change are more sensitive to pesticides

Bee populations are exposed to multiple stressors, including land-use change, biological invasions, climate change, and pesticide exposure, that may interact synergistically. We analyze the combined effects of climate warming and sublethal insecticide exposure in the solitary bee Osmia cornuta. Previous Osmia studies show that warm wintering temperatures cause body weight loss, lipid consumption, and fat body depletion. Because the fat body plays a key role in xenobiotic detoxification, we expected that bees exposed to climate warming scenarios would be more sensitive to pesticides. We exposed O. cornuta females to three wintering treatments: current scenario (2007–2012 temperatures), near-future (2021–2050 projected temperatures), and distant-future (2051–2080). Upon emergence in spring, bees were orally exposed to three sublethal doses of an insecticide (Closer, a.i. sulfoxaflor; 0, 4.55 and 11.64 ng a.i./bee). We measured the combined effects of wintering and insecticide exposure on phototactic response, syrup consumption, and longevity. Wintering treatment by itself did not affect winter mortality, but body weight loss increased with increasing wintering temperatures. Similarly, wintering treatment by itself hardly influenced phototactic response or syrup consumption. However, bees wintered at the warmest temperatures had shorter longevity, a strong fecundity predictor in Osmia. Insecticide exposure, especially at the high dose, impaired the ability of bees to respond to light, and resulted in reduced syrup consumption and longevity. The combination of the warmest winter and the high insecticide dose resulted in a 70% longevity decrease. Smaller bees, resulting from smaller pollen–nectar provisions, had shorter longevity suggesting nutritional stress may further compromise fecundity in O. cornuta. Our results show a synergistic interaction between two major drivers of bee declines, and indicate that bees will become more sensitive to pesticides under the current global warming scenario. Our findings have important implications for pesticide regulation and underscore the need to consider multiple stressors to understand bee declines.     

Effects of Sublethal Doses of Imidacloprid on Young Adult Honeybee Behaviour

Imidacloprid (IMI), a neonicotinoid used for its high selective toxicity to insects, is one of the most commonly used pesticides. However, its effect on beneficial insects such as the honeybee Apis mellifera L is still controversial. As young adult workers perform in-hive duties that are crucial for colony maintenance and survival, we aimed to assess the effect of sublethal IMI doses on honeybee behaviour during this period. Also, because this insecticide acts as a cholinergic-nicotinic agonist and these pathways take part in insect learning and memory processes; we used IMI to assess their role and the changes they suffer along early adulthood. We focused on appetitive behaviours based on the proboscis extension response. Laboratory reared adults of 2 to 10 days of age were exposed to sublethal IMI doses (0.25 or 0.50ng) administered orally or topically prior to behavioural assessment. Modification of gustatory responsiveness and impairment of learning and memory were found as a result of IMI exposure. These outcomes differed depending on age of evaluation, type of exposure and IMI dose, being the youngest bees more sensitive and the highest oral dose more toxic. Altogether, these results imply that IMI administered at levels found in agroecosystems can reduce sensitivity to reward and impair associative learning in young honeybees. Therefore, once a nectar inflow with IMI traces is distributed within the hive, it could impair in-door duties with negative consequences on colony performance.     

Multiple routes of pesticide exposure for honey bees living near agricultural fields

Populations of honey bees and other pollinators have declined worldwide in recent years. A variety of stressors have been implicated as potential causes, including agricultural pesticides. Neonicotinoid insecticides, which are widely used and highly toxic to honey bees, have been found in previous analyses of honey bee pollen and comb material. However, the routes of exposure have remained largely undefined. We used LC/MS-MS to analyze samples of honey bees, pollen stored in the hive and several potential exposure routes associated with plantings of neonicotinoid treated maize. Our results demonstrate that bees are exposed to these compounds and several other agricultural pesticides in several ways throughout the foraging period. During spring, extremely high levels of clothianidin and thiamethoxam were found in planter exhaust material produced during the planting of treated maize seed. We also found neonicotinoids in the soil of each field we sampled, including unplanted fields. Plants visited by foraging bees (dandelions) growing near these fields were found to contain neonicotinoids as well. This indicates deposition of neonicotinoids on the flowers, uptake by the root system, or both. Dead bees collected near hive entrances during the spring sampling period were found to contain clothianidin as well, although whether exposure was oral (consuming pollen) or by contact (soil/planter dust) is unclear. We also detected the insecticide clothianidin in pollen collected by bees and stored in the hive. When maize plants in our field reached anthesis, maize pollen from treated seed was found to contain clothianidin and other pesticides; and honey bees in our study readily collected maize pollen. These findings clarify some of the mechanisms by which honey bees may be exposed to agricultural pesticides throughout the growing season. These results have implications for a wide range of large-scale annual cropping systems that utilize neonicotinoid seed treatments.     

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.     

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.     

A Four-Year Field Program Investigating Long-Term Effects of Repeated Exposure of Honey Bee Colonies to Flowering Crops Treated with Thiamethoxam

Neonicotinoid residues in nectar and pollen from crop plants have been implicated as one of the potential factors causing the declines of honey bee populations. Median residues of thiamethoxam in pollen collected from honey bees after foraging on flowering seed treated maize were found to be between 1 and 7 µg/kg, median residues of the metabolite CGA322704 (clothianidin) in the pollen were between 1 and 4 µg/kg. In oilseed rape, median residues of thiamethoxam found in pollen collected from bees were between <1 and 3.5 µg/kg and in nectar from foraging bees were between 0.65 and 2.4 µg/kg. Median residues of CGA322704 in pollen and nectar in the oilseed rape trials were all below the limit of quantification (1 µg/kg). Residues in the hive were even lower in both the maize and oilseed rape trials, being at or below the level of detection of 1 µg/kg for bee bread in the hive and at or below the level of detection of 0.5 µg/kg for hive nectar, honey and royal jelly samples. The long-term risk to honey bee colonies in the field was also investigated, including the sensitive overwintering stage, from four years consecutive single treatment crop exposures to flowering maize and oilseed rape grown from thiamethoxam treated seeds at rates recommended for insect control. Throughout the study, mortality, foraging behavior, colony strength, colony weight, brood development and food storage levels were similar between treatment and control colonies. Detailed examination of brood development throughout the year demonstrated that colonies exposed to the treated crop were able to successfully overwinter and had a similar health status to the control colonies in the following spring. We conclude that these data demonstrate there is a low risk to honey bees from systemic residues in nectar and pollen following the use of thiamethoxam as a seed treatment on oilseed rape and maize.     

High Levels of Miticides and Agrochemicals in North American Apiaries: Implications for Honey Bee Health

Background

Recent declines in honey bees for crop pollination threaten fruit, nut, vegetable and seed production in the United States. A broad survey of pesticide residues was conducted on samples from migratory and other beekeepers across 23 states, one Canadian province and several agricultural cropping systems during the 2007–08 growing seasons.

Methodology/Principal Findings

We have used LC/MS-MS and GC/MS to analyze bees and hive matrices for pesticide residues utilizing a modified QuEChERS method. We have found 121 different pesticides and metabolites within 887 wax, pollen, bee and associated hive samples. Almost 60% of the 259 wax and 350 pollen samples contained at least one systemic pesticide, and over 47% had both in-hive acaricides fluvalinate and coumaphos, and chlorothalonil, a widely-used fungicide. In bee pollen were found chlorothalonil at levels up to 99 ppm and the insecticides aldicarb, carbaryl, chlorpyrifos and imidacloprid, fungicides boscalid, captan and myclobutanil, and herbicide pendimethalin at 1 ppm levels. Almost all comb and foundation wax samples (98%) were contaminated with up to 204 and 94 ppm, respectively, of fluvalinate and coumaphos, and lower amounts of amitraz degradates and chlorothalonil, with an average of 6 pesticide detections per sample and a high of 39. There were fewer pesticides found in adults and brood except for those linked with bee kills by permethrin (20 ppm) and fipronil (3.1 ppm).

Conclusions/Significance

The 98 pesticides and metabolites detected in mixtures up to 214 ppm in bee pollen alone represents a remarkably high level for toxicants in the brood and adult food of this primary pollinator. This represents over half of the maximum individual pesticide incidences ever reported for apiaries. While exposure to many of these neurotoxicants elicits acute and sublethal reductions in honey bee fitness, the effects of these materials in combinations and their direct association with CCD or declining bee health remains to be determined.     

Field-Level Sublethal Effects of Approved Bee Hive Chemicals on Honey Bees (Apis mellifera L)

In a study replicated across two states and two years, we tested the sublethal effects on honey bees of the miticides Apistan (tau fluvalinate) and Check Mite+ (coumaphos) and the wood preservative copper naphthenate applied at label rates in field conditions. A continuous covariate, a colony Varroa mite index, helped us disambiguate the effects of the chemicals on bees while adjusting for a presumed benefit of controlling mites. Mite levels in colonies treated with Apistan or Check Mite+ were not different from levels in non-treated controls. Experimental chemicals significantly decreased 3-day brood survivorship and increased construction of queen supercedure cells compared to non-treated controls. Bees exposed to Check Mite+ as immatures had higher legacy mortality as adults relative to non-treated controls, whereas bees exposed to Apistan had improved legacy mortality relative to non-treated controls. Relative to non-treated controls, Check Mite+ increased adult emergence weight. Although there was a treatment effect on a test of associative learning, it was not possible to statistically separate the treatment means, but bees treated with Apistan performed comparatively well. And finally, there were no detected effects of bee hive chemical on colony bee population, amount of brood, amount of honey, foraging rate, time required for marked released bees to return to their nest, percentage of released bees that return to the nest, and colony Nosema spore loads. To our knowledge, this is the first study to examine sublethal effects of bee hive chemicals applied at label rates under field conditions while disambiguating the results from mite control benefits realized from the chemicals. Given the poor performance of the miticides at reducing mites and their inconsistent effects on the host, these results defend the use of bee health management practices that minimize use of exotic hive chemicals.     

Using a Hazard Quotient to Evaluate Pesticide Residues Detected in Pollen Trapped from Honey Bees (Apis mellifera) in Connecticut

Analysis of pollen trapped from honey bees as they return to their hives provides a method of monitoring fluctuations in one route of pesticide exposure over location and time. We collected pollen from apiaries in five locations in Connecticut, including urban, rural, and mixed agricultural sites, for periods from two to five years. Pollen was analyzed for pesticide residues using a standard extraction method widely used for pesticides (QuEChERS) and liquid chromatography/mass spectrometric analysis. Sixty pesticides or metabolites were detected. Because the dose lethal to 50% of adult worker honey bees (LD₅₀) is the only toxicity parameter available for a wide range of pesticides, and among our pesticides there were contact LD₅₀ values ranging from 0.006 to >1000 μg per bee (range 166,000X), and even among insecticides LD₅₀ values ranged from 0.006 to 59.8 μg/bee (10,000X); therefore we propose that in studies of honey bee exposure to pesticides that concentrations be reported as Hazard Quotients as well as in standard concentrations such as parts per billion. We used both contact and oral LD₅₀ values to calculate Pollen Hazard Quotients (PHQ = concentration in ppb ÷ LD₅₀ as μg/bee) when both were available. In this study, pesticide Pollen Hazard Quotients ranged from over 75,000 to 0.01. The pesticides with the greatest Pollen Hazard Quotients at the maximum concentrations found in our study were (in descending order): phosmet, Imidacloprid, indoxacarb, chlorpyrifos, fipronil, thiamethoxam, azinphos-methyl, and fenthion, all with at least one Pollen Hazard Quotient (using contact or oral LD₅₀) over 500. At the maximum rate of pollen consumption by nurse bees, a Pollen Hazard Quotient of 500 would be approximately equivalent to consuming 0.5% of the LD₅₀ per day. We also present an example of a Nectar Hazard Quotient and the percentage of LD₅₀ per day at the maximum nectar consumption rate.     

High Concentrations of the Alarm Pheromone Component,Isopentyl Acetate,Reduces Foraging and Dancing in <Emphasis Type="Italic">Apis mellifera</Emphasis> Ligustica and <Emphasis Type="Italic">Apis cerana</Emphasis> Cerana

A honeybee colony is a superorganism that has evolved precise communication systems, which allow the colony to gather information from numerous individuals and coordinate its behavior. Alarm pheromones, such as isopentyl acetate (IPA), the main component of sting alarm pheromone, play a critical role in the coordination of individual behaviors as well as colony communication in honeybee colonies. In this study, honeybees (Apis mellifera ligustica and Apis cerana cerana) were exposed to relatively high levels of IPA at a foraging site (6–8 bee equivalents) and inside their colony (28–58 bee equivalents) to investigate the influence of alarm pheromones on foraging activity and hive flight activity. IPA reduced the number of bees that flew out the hive, foraged, and waggle danced. Under both contexts in the hive and at the food source, IPA can therefore inhibit honey bee foraging and foraging communication.     

A Mathematical Model of Forager Loss in Honeybee Colonies Infested with <Emphasis Type="Italic">Varroa destructor</Emphasis> and the Acute Bee Paralysis Virus

We incorporate a mathematical model of Varroa destructor and the Acute Bee Paralysis Virus with an existing model for a honeybee colony, in which the bee population is divided into hive bees and forager bees based on tasks performed in the colony. The model is a system of five ordinary differential equations with dependent variables: uninfected hive bees, uninfected forager bees, infected hive bees, virus-free mites and virus-carrying mites. The interplay between forager loss and disease infestation is studied. We study the stability of the disease-free equilibrium of the bee-mite-virus model and observe that the disease cannot be fought off in the absence of varroacide treatment. However, the disease-free equilibrium can be stable if the treatment is strong enough and also if the virus-carrying mites become virus-free at a rate faster than the mite birth rate. The critical forager loss due to homing failure, above which the colony fails, is calculated using simulation experiments for disease-free, treated and untreated mite-infested, and treated virus-infested colonies. A virus-infested colony without varroacide treatment fails regardless of the forager mortality rate.     

Effects of Imidacloprid and Varroa destructor on survival and health of European honey bees,Apis mellifera

There has been growing concern over declines in populations of honey bees and other pollinators which are a vital part to our food security. It is imperative to identify factors responsible for accelerated declines in bee populations and develop solutions for reversing bee losses. While exact causes of colony losses remain elusive, risk factors thought to play key roles are ectoparasitic mites Varroa destructor and neonicotinoid pesticides. The present study aims to investigate effects of a neonicotinoid pesticide Imidacloprid and Varroa mites individually on survivorship, growth, physiology, virus dynamics and immunity of honey bee workers. Our study provides clear evidence that the exposure to sublethal doses of Imidacloprid could exert a significantly negative effect on health and survival of honey bees. We observed a significant reduction in the titer of vitellogenin (Vg), an egg yolk precursor that regulates the honey bees development and behavior and often are linked to energy homeostasis, in bees exposed to Imidacloprid. This result indicates that sublethal exposure to neonicotinoid could lead to increased energy usage in honey bees as detoxification is a energy‐consuming metabolic process and suggests that Vg could be a useful biomarker for measuring levels of energy stress and sublethal effects of pesticides on honey bees. Measurement of the quantitative effects of different levels of Varroa mite infestation on the replication dynamic of Deformed wing virus (DWV), an RNA virus associated with Varroa infestation, and expression level of immune genes yields unique insights into how honey bees respond to stressors under laboratory conditions.     

Pesticide Residues and Bees – A Risk Assessment

Bees are essential pollinators of many plants in natural ecosystems and agricultural crops alike. In recent years the decline and disappearance of bee species in the wild and the collapse of honey bee colonies have concerned ecologists and apiculturalists, who search for causes and solutions to this problem. Whilst biological factors such as viral diseases, mite and parasite infections are undoubtedly involved, it is also evident that pesticides applied to agricultural crops have a negative impact on bees. Most risk assessments have focused on direct acute exposure of bees to agrochemicals from spray drift. However, the large number of pesticide residues found in pollen and honey demand a thorough evaluation of all residual compounds so as to identify those of highest risk to bees. Using data from recent residue surveys and toxicity of pesticides to honey and bumble bees, a comprehensive evaluation of risks under current exposure conditions is presented here. Standard risk assessments are complemented with new approaches that take into account time-cumulative effects over time, especially with dietary exposures. Whilst overall risks appear to be low, our analysis indicates that residues of pyrethroid and neonicotinoid insecticides pose the highest risk by contact exposure of bees with contaminated pollen. However, the synergism of ergosterol inhibiting fungicides with those two classes of insecticides results in much higher risks in spite of the low prevalence of their combined residues. Risks by ingestion of contaminated pollen and honey are of some concern for systemic insecticides, particularly imidacloprid and thiamethoxam, chlorpyrifos and the mixtures of cyhalothrin and ergosterol inhibiting fungicides. More attention should be paid to specific residue mixtures that may result in synergistic toxicity to bees.     

Effect of fipronil on side-specific antennal tactile learning in the honeybee

In the honeybee, the conditioning of the proboscis extension response using tactile antennal stimulations is well suited for studying the side-specificity of learning including the possible bilateral transfer of memory traces in the brain, and the role of inhibitory networks. A tactile stimulus was presented to one antenna in association with a sucrose reward to the proboscis. The other antenna was either not stimulated (A+/0 training), stimulated with a non-reinforced tactile stimulus B (A+/B− training) or stimulated with B reinforced with sucrose to the proboscis (A+/B+ training). Memory tests performed 3 and 24 h after training showed in all situations that a tactile stimulus learnt on one side was only retrieved ipsilaterally, indicating no bilateral transfer of information. In all these groups, we investigated the effect of the phenylpyrazole insecticide fipronil by applying a sublethal dose (0.5 ng/bee) on the thorax 15 min before training. This treatment decreased acquisition success and the subsequent memory performances were lowered but the distribution of responses to the tactile stimuli between sides was not affected. These results underline the role of the inhibitory networks targeted by fipronil on tactile learning and memory processes.     

Population statistics and biological traits of Hippodamia variegata (Goeze) (Coleoptera: Coccinellidae) affected by LC30 of thiamethoxam and pirimicarb

Although selective pesticides are recommended to use in integrated pest management (IPM) programmes, they could adversely affect biological agents. So, the aim of the current study was to investigate the LC₃₀ of pirimicarb and thiamethoxam on Hippodamia variegata. Two pesticides, pirimicarb and thiamethoxam, when applied at concentrations of 1522.8 and 251.3?mg (ai)/L, respectively, on the third instar larvae of the insect, produced 30% mortality. Also, results showed that neither pirimicarb nor thiamethoxam affect stage age distribution (c_x), fecundity (eggs/female) and adult development time. These two pesticides extended preadult duration significantly (p?<?0.0001, F?=?31.22, df?=?122). Also, survival rate (l_x) and age-specific reproductive value (v_x) decreased and some changes in age-specific life expectancy (e_x) happened. Generally, more adverse effects were found in the population treated by thiamethoxam. The results showed that pirimicarb and thiamethoxam have potential to harm the predatory ladybird in IPM programme, though in sublethal doses.     

Multi-Drug Resistance Transporters and a Mechanism-Based Strategy for Assessing Risks of Pesticide Combinations to Honey Bees

Annual losses of honey bee colonies remain high and pesticide exposure is one possible cause. Dangerous combinations of pesticides, plant-produced compounds and antibiotics added to hives may cause or contribute to losses, but it is very difficult to test the many combinations of those compounds that bees encounter. We propose a mechanism-based strategy for simplifying the assessment of combinations of compounds, focusing here on compounds that interact with xenobiotic handling ABC transporters. We evaluate the use of ivermectin as a model substrate for these transporters. Compounds that increase sensitivity of bees to ivermectin may be inhibiting key transporters. We show that several compounds commonly encountered by honey bees (fumagillin, Pristine, quercetin) significantly increased honey bee mortality due to ivermectin and significantly reduced the LC₅₀ of ivermectin suggesting that they may interfere with transporter function. These inhibitors also significantly increased honey bees sensitivity to the neonicotinoid insecticide acetamiprid. This mechanism-based strategy may dramatically reduce the number of tests needed to assess the possibility of adverse combinations among pesticides. We also demonstrate an in vivo transporter assay that provides physical evidence of transporter inhibition by tracking the dynamics of a fluorescent substrate of these transporters (Rhodamine B) in bee tissues. Significantly more Rhodamine B remains in the head and hemolymph of bees pretreated with higher concentrations of the transporter inhibitor verapamil. Mechanism-based strategies for simplifying the assessment of adverse chemical interactions such as described here could improve our ability to identify those combinations that pose significantly greater risk to bees and perhaps improve the risk assessment protocols for honey bees and similar sensitive species.