Systemic insecticides are applied to plants using a wide variety of methods, ranging from foliar sprays to seed treatments and soil drenches. Neonicotinoids and fipronil are among the most widely used pesticides in the world. Their popularity is largely due to their high toxicity to invertebrates, the ease and flexibility with which they can be applied, their long persistence, and their systemic nature, which ensures that they spread to all parts of the target crop. However, these properties also increase the probability of environmental contamination and exposure of nontarget organisms. Environmental contamination occurs via a number of routes including dust generated during drilling of dressed seeds, contamination and accumulation in arable soils and soil water, runoff into waterways, and uptake of pesticides by nontarget plants via their roots or dust deposition on leaves. Persistence in soils, waterways, and nontarget plants is variable but can be prolonged; for example, the half-lives of neonicotinoids in soils can exceed 1,000 days, so they can accumulate when used repeatedly. Similarly, they can persist in woody plants for periods exceeding 1 year. Breakdown results in toxic metabolites, though concentrations of these in the environment are rarely measured. Overall, there is strong evidence that soils, waterways, and plants in agricultural environments and neighboring areas are contaminated with variable levels of neonicotinoids or fipronil mixtures and their metabolites (soil, parts per billion (ppb)-parts per million (ppm) range; water, parts per trillion (ppt)-ppb range; and plants, ppb-ppm range). This provides multiple routes for chronic (and acute in some cases) exposure of nontarget animals. For example, pollinators are exposed through direct contact with dust during drilling; consumption of pollen, nectar, or guttation drops from seed-treated crops, water, and consumption of contaminated pollen and nectar from wild flowers and trees growing near-treated crops. Studies of food stores in honeybee colonies from across the globe demonstrate that colonies are routinely and chronically exposed to neonicotinoids, fipronil, and their metabolites (generally in the 1–100 ppb range), mixed with other pesticides some of which are known to act synergistically with neonicotinoids. Other nontarget organisms, particularly those inhabiting soils, aquatic habitats, or herbivorous insects feeding on noncrop plants in farmland, will also inevitably receive exposure, although data are generally lacking for these groups. We summarize the current state of knowledge regarding the environmental fate of these compounds by outlining what is known about the chemical properties of these compounds, and placing these properties in the context of modern agricultural practices.

The death of honey bees, Apis mellifera L., and the consequent colony collapse disorder causes major losses in agriculture and plant pollination worldwide. The phenomenon showed increasing rates in the past years, although its causes are still awaiting a clear answer. Although neonicotinoid systemic insecticides used for seed coating of agricultural crops were suspected as possible reason, studies so far have not shown the existence of unquestionable sources capable of delivering directly intoxicating doses in the fields. Guttation is a natural plant phenomenon causing the excretion of xylem fluid at leaf margins. Here, we show that leaf guttation drops of all the corn plants germinated from neonicotinoid-coated seeds contained amounts of insecticide constantly higher than 10 mg/l, with maxima up to 100 mg/l for thiamethoxam and clothianidin, and up to 200 mg/l for imidacloprid. The concentration of neonicotinoids in guttation drops can be near those of active ingredients commonly applied in field sprays for pest control, or even higher. When bees consume guttation drops, collected from plants grown from neonicotinoid-coated seeds, they encounter death within few minutes.

Since seed coating with neonicotinoid insecticides was introduced in the late 1990s, European beekeepers have reported severe colony losses in the period of corn sowing (spring). As a consequence, seed-coating neonicotinoid insecticides that are used worldwide on corn crops have been blamed for honeybee decline. In view of the currently increasing crop production, and also of corn as a renewable energy source, the correct use of these insecticides within sustainable agriculture is a cause of concern. In this paper, a probable--but so far underestimated--route of environmental exposure of honeybees to and intoxication with neonicotinoid insecticides, namely, the atmospheric emission of particulate matter containing the insecticide by drilling machines, has been quantitatively studied. Using optimized analytical procedures, quantitative measurements of both the emitted particulate and the consequent direct contamination of single bees approaching the drilling machine during the foraging activity have been determined. Experimental results show that the environmental release of particles containing neonicotinoids can produce high exposure levels for bees, with lethal effects compatible with colony losses phenomena observed by beekeepers.

Losses of honeybees have been reported in Italy concurrent with the sowing of corn coated with neonicotinoids using a pneumatic drilling machine. Being unconvinced that solid particles containing systemic insecticide, falling on the vegetation surrounding the sown area, could poison bees foraging on contaminated nectar and pollen, the effect of direct aerial powdering was tested on foragers in free flight near the drilling machine. Bees were conditioned to visit a dispenser of sugar solution whilst a drilling machine was sowing corn along the flight path. Samples of bees were captured on the dispenser, caged and held in the laboratory. Chemical analysis showed some hundred nanograms of insecticide per bee. Nevertheless, caged bees, previously contaminated in flight, died only if kept in conditions of high humidity. After the sowing, an increase in bee mortality in front of the hives was also observed. Spring bee losses, which corresponded with the sowing of corn-coated seed, seemed to be related to the casual encountering of drilling machine during foraging flight across the ploughed fields

SummaryModern agriculture often involves the use of pesticides to protect crops. These substances are harmful to target organisms (pests and pathogens). Nevertheless, they can also damage non-target animals, such as pollinators and entomophagous arthropods. It is obvious that the undesirable side effects of pesticides on the environment should be reduced to a minimum. Western honey bees (Apis mellifera) are very important organisms from an agricultural perspective and are vulnerable to pesticide-induced impacts. They contribute actively to the pollination of cultivated crops and wild vegetation, making food production possible. Of course, since Apis mellifera occupies the same ecological niche as many other species of pollinators, the loss of honey bees caused by environmental pollutants suggests that other insects may experience a similar outcome. Because pesticides can harm honey bees and other pollinators, it is important to register pesticides that are as selective as possible. In this manuscript, we describe a selection of methods used for studying pesticide toxicity/selectiveness towards Apis mellifera. These methods may be used in risk assessment schemes and in scientific research aimed to explain acute and chronic effects of any target compound on Apis mellifera. Métodos estándar para la investigación toxicológica en Apis mellifera ResumenLa agricultura moderna a menudo implica el uso de plaguicidas para proteger los cultivos. Estas sustancias son dañinas para los organismos objetivo (plagas y patógenos). Sin embargo, también pueden dañar a animales que no son objetivo, como artrópodos polinizadores y entomófagos. Obviamente los efectos secundarios indeseables de los plaguicidas sobre el medio ambiente deben ser reducidos al mínimo. Las abejas occidentales (Apis mellifera) son organismos muy importantes desde el punto de vista agrícola y son vulnerables a los impactos inducidos por los plaguicidas. Contribuyen activamente a la polinización de los cultivos y de la vegetación silvestre, lo que hace posible la producción de alimentos. Como Apis mellifera ocupa el mismo nicho ecológico que muchas otras especies de polinizadores, la pérdida de las abejas melíferas causada por contaminantes ambientales sugiere que otros insectos pueden experimentar un resultado similar. Ya que los plaguicidas pueden dañar a las abejas y a otros polinizadores, es importante registrar los plaguicidas que sean lo más selectivos posible. En este artículo, se describe una selección de los métodos utilizados para el estudio de la toxicidad y el efecto selectivo de los plaguicidas hacia Apis mellifera. Estos métodos se pueden utilizar en sistemas de evaluación de riesgo y en la investigación científica para explicar los efectos agudos y crónicos en Apis mellifera de cualquier compuesto objetivo.