Figures
Abstract
It has been suggested that the widespread use of neonicotinoid insecticides threatens bees, but research on this topic has been surrounded by controversy. In order to synthesize which research approaches have been used to examine the effect of neonicotinoids on bees and to identify knowledge gaps, we systematically reviewed research on this subject that was available on the Web of Science and PubMed in June 2015. Most of the 216 primary research studies were conducted in Europe or North America (82%), involved the neonicotinoid imidacloprid (78%), and concerned the western honey bee Apis mellifera (75%). Thus, little seems to be known about neonicotinoids and bees in areas outside Europe and North America. Furthermore, because there is considerable variation in ecological traits among bee taxa, studies on honey bees are not likely to fully predict impacts of neonicotinoids on other species. Studies on crops were dominated by seed-treated maize, oilseed rape (canola) and sunflower, whereas less is known about potential side effects on bees from the use of other application methods on insect pollinated fruit and vegetable crops, or on lawns and ornamental plants. Laboratory approaches were most common, and we suggest that their capability to infer real-world consequences are improved when combined with information from field studies about realistic exposures to neonicotinoids. Studies using field approaches often examined only bee exposure to neonicotinoids and more field studies are needed that measure impacts of exposure. Most studies measured effects on individual bees. We suggest that effects on the individual bee should be linked to both mechanisms at the sub-individual level and also to the consequences for the colony and wider bee populations. As bees are increasingly facing multiple interacting pressures future research needs to clarify the role of neonicotinoids in relative to other drivers of bee declines.
Citation: Lundin O, Rundlöf M, Smith HG, Fries I, Bommarco R (2015) Neonicotinoid Insecticides and Their Impacts on Bees: A Systematic Review of Research Approaches and Identification of Knowledge Gaps. PLoS ONE 10(8): e0136928. https://doi.org/10.1371/journal.pone.0136928
Editor: Nigel E. Raine, University of Guelph, CANADA
Received: April 23, 2014; Accepted: August 11, 2015; Published: August 27, 2015
Copyright: © 2015 Lundin et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Data Availability: All relevant data are within the paper and its Supporting Information files.
Funding: This research was financed by a grant from the Swedish Civil Contingencies Agency. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing interests: The authors have declared that no competing interests exist.
Introduction
Animal pollination, mainly performed by bees, is an important ecosystem service with almost 90 percent of flowering plants and 75 percent of the world’s most common crops benefiting from animal flower visitation [1–2]. Habitat loss and fragmentation, pesticides, pathogens, climate change, invasive species, intense management of managed bees, and decreased interest in beekeeping have all been suggested as threats to bees and pollination services, but the relative importance of these drivers remains uncertain [3–4].
More recently, the use of neonicotinoid insecticides has been specifically pointed out as a factor that might contribute to declines of both managed and wild bees [5–6]. Neonicotinoid compounds are used in more than 120 countries with at least 140 different crop uses (e.g. soil and foliar applications of the same compound in the same crop are defined as two different crop uses) [7]. Since their commercial introduction in the early 1990s, neonicotinoids have quickly become the most commonly used class of insecticides in the world. Their market share grew rapidly from 16 percent in 2005 to 24 percent in 2008, valued at roughly €1.5 billion in 2008 [7–8]. Neonicotinoids have high selectivity towards invertebrate over vertebrate organisms [9]. They are taken up systemically and can be present in all plant tissues, which makes them efficient against a wide range of pests over a protracted time period and when applied in small quantities, e.g. as seed treatments [7–9]. At the same time, several of the neonicotinoid compounds have been shown to be highly toxic to bees in very small quantities [10]. However, with the exception of exposure to dust emission from pneumatic seeders during sowing of treated seeds [11], estimates of bees’ exposure to neonicotinoids generally are substantially lower than levels causing acute mortality. Neonicotinoids can be translocated into pollen and nectar, the principal food sources for bees [12]. Moreover, some of the compounds degrade slowly and are present in the environment, e.g. in soil and/or treated plants for months, or even years, after the application [6,13–14]. Concern for pollinators has led to a temporary restriction of three neonicotinoids (clothianidin, thiamethoxam and imidacloprid) as seed treatments for use on crops attractive to bees in the European Union [15] and a policy to reduce the use of the same three insecticides as seed treatments for maize and soy by 80 percent from 2014 levels in Ontario, Canada [16]. However, significant knowledge gaps and controversy remain as to whether such restrictions are justified [17–19].
Reviews on the effect of neonicotinoids on bees have so far often dealt either specifically with the role of neonicotinoids for honey bee declines or colony losses [20–24], or more broadly with the effects of neonicotinoid use on the wider environment [6]. The most comprehensive reviews on neonicotinoid effects on bees have focused on concentrations of neonicotinoids found in the environment that bees might be exposed to, effects of neonicotinoids on bees, and risk assessment [5,25–26]. Additionally, the status of the natural science evidence base for effects of neonicotinoids on bees has recently been summarized [27]. In this study we examine which research approaches have been used in this area by conducting a systematic review of the literature. More specifically, for each study reviewed we asked the following questions: (i) from which country did the study originate, (ii) which neonicotinoid compounds were studied, (iii) which crops were studied, (iv) which bee species were studied, (v) which methodological approaches were used, and (vi) what biological levels, from the sub-individual to the population level, were studied? By asking these questions we aimed to systematically characterize how our current knowledge about the effect of neonicotinoids on bees has been derived and to identify research gaps that can be addressed in future studies.
Methods
We searched in the Web of Science Core Collection and PubMed for studies that had examined the effects of neonicotinoids on bees (last access date: 20 June 2015). The search was limited to the Web of Science and PubMed because it contained research articles that were available in full text, written in English, and that had undergone peer-review by scientists. We used the following search string to locate potential studies on neonicotinoids and bees: (neonic* OR imidacloprid OR clothianidin OR thiamethoxam OR acetamiprid OR thiacloprid OR nitenpyram OR dinotefuran) AND (*bee OR *bees).
The primary database consisted of 543 publications, from which we removed duplicates. Conference proceedings and book chapters that appeared in the search were excluded from further review to ensure easy access to full text publications. Publications not written in English were also excluded. Remaining database records were retrieved in full text and inspected in detail. For primary research articles to be included in our review they had to contain either a measure of an effect of a neonicotinoid on bees, or a measure of neonicotinoid contamination of bees or plants, hive products or other material that bees come into contact with or digest. Article types other than primary research (see categories below) were also included if they dealt with these topics. Publications in analytical chemistry that focused on developing methods to analyze neonicotinoids were included only if they included determinations of neonicotinoids in the natural environment.
Each publication included in the study was reviewed using a standard review protocol, with information collected as described below. We collected full reference information and extracted information about whether the publication was primary research, a review, meta-analysis, or another type of article (e.g. comment, opinion, essay or editorial). As our review questions were designed for primary research publications, we further evaluated only these publications. For each primary research study we noted the country where the study was performed, focal neonicotinoid compound(s), crop species and application methods examined in each crop studied, and bee species. For studies that lacked information about where the research was performed, as was the case for some laboratory studies, we used the location of the first author’s institution. A focal crop species was only noted if measurements were made in that crop, or if bees were exposed to that crop during the study. Hence, a focal crop was noted for laboratory studies only when it was included in the experiment, but not when the experimental treatment mimicked the application details for a certain crop. Each study could include zero to several focal crops and bee species, and one to several focal neonicotinoid compounds.
The methods used in primary research studies were categorized into five different methodological approaches: laboratory, semi-field, field, in silico, or combined approach. Studies in which the treatments and data collection were conducted in the laboratory or greenhouse were jointly classified into “laboratory approach”. Designs that used cages or tunnels in the field were classified as “semi-field approach”. “In silico approach” included modeling or risk assessments. “Combined approach” used more than one approach for single endpoints. Examples include combined laboratory and field designs, where the treatment applied to the study subjects was performed in the field, and the effects were observed in the laboratory on the same study subjects, or vice versa. Most studies could be assigned to a single methodological approach, but some studies that used different approaches for different endpoints were classified into multiple categories. For example, publications in which some endpoints were measured on one set of study subjects in a laboratory experiment, and another set of endpoints were measured on different set of study subjects in the field.
Finally, we mapped at what level of biological organization each study measured effects. Endpoint measurements of each study were grouped into four classes: (i) sub-individual measure (e.g. gene expression, cell death, neurotransmission, or physiological measures), (ii) individual measure (e.g. learning, memory, movement, or mortality), (iii) colony measure (e.g. colony mass, colony reproduction, or colony survival), or (iv) population measure (e.g. bee population size).
Fig 1 depicts a flow diagram for the systematic review. A checklist for the systematic review can be found in the Supplementary Information (S1 PRISMA Checklist).
Results
Article types and publication years
A total of 268 publications matched our criteria; 216 were primary research, 18 were reviews, one was a meta-analysis, and 33 were other publication types. Full references for all publications and data for each primary research publication are presented in S1 Table. Approximately half of the studies were published within the last three years, demonstrating a rapid expansion of the research field (Fig 2).
The single meta-analysis study was published in 2011 [12] and is not included in this figure. Data for 2015 (not complete; see text) included 20 primary research publications, 2 reviews and 6 other publications (not included in figure).
Geographical distribution of studies.
Primary research studies were conducted in 27 countries. However, more than half of the studies were from four countries: France (n = 44), the United States (n = 35), the United Kingdom (n = 23) and Italy (n = 20). Overall, 82 percent of the studies were done in Europe or North America (Fig 3). Nine percent of the studies were from Asia (n = 19) and 8 percent were from South America (n = 17). We found three studies from Oceania and two studies from Africa.
The number of primary research studies from each country is indicated. Colours indicate countries with 1–5 studies (red), 6–10 studies (orange) or more than 10 studies (green).
Neonicotinoid compounds
Imidacloprid was the most commonly studied compound (included in 78 percent of studies, n = 168), followed by thiamethoxam (34 percent, n = 73), clothianidin (33 percent, n = 71), acetamiprid (19 percent, n = 40), thiacloprid (18 percent, n = 39), dinotefuran (7 percent, n = 15), and nitenpyram (6 percent, n = 13).
Crop species
Maize was the most commonly studied crop (28 studies), followed by oilseed rape (canola: 7 studies) and sunflower (7 studies). Between one and four studies were found for each of 13 other crops (Table 1).
Bee species
The western honey bee Apis mellifera was the most common bee species studied (162 studies) followed by the two bumble bee species Bombus terrestris (24 studies) and Bombus impatiens (10 studies). Between one and six studies were found for each of 15 other bee species or species groups (Table 2).
Methodological approaches
A total of 112 studies used laboratory approaches, 92 used field approaches, 14 used semi-field approaches, and 12 used in silico approaches. Twenty-five studies used combined approaches. They most often combined laboratory and field approaches where the treatment was applied in the field and the effects were observed in the laboratory or vice versa. There were more methodological approaches than primary research studies because 32 studies used more than one approach (see S1 Table).
Biological levels–from sub-individual to population effects
Most studies measured the effects of neonicotinoids on bees at the individual level (n = 109 studies), followed by the colony level (n = 60) and the sub-individual level (n = 48). None of the studies investigated effects on the population level according to our definitions.
Discussion
Geographical distribution of studies
We found that most studies originated from just a few countries in Europe and North America. The skewed geographical distribution of studies is potentially problematic, especially when it comes to the paucity of field studies from countries outside Europe and North America. The use of neonicotinoids is geographically widespread: for example, imidacloprid has been registered for use in at least 120 countries [7]. Although more detailed global records of neonicotinoid use around the world are lacking, insecticide use is generally more intense in upper-middle income countries compared to high income countries, primarily in Europe and North America [42]. Furthermore, pesticides are also generally more weakly regulated in countries outside Europe and North America [42]. Finally, areas outside Europe and North America host the majority of the global crop pollination value. It has been estimated that 58 percent of the global economic value of insect pollination originates from Asia, with another 8 percent originating from Africa and a further 10 percent from South and Central America [43]. These factors suggest the need to assess the impacts of neonicotinoid use on bees and crop pollination services in countries outside Europe and North America. It is, however, important to acknowledge that our method of searching for studies only in the Web of Science and PubMed might have skewed results towards finding more studies from Europe and North America, and that alternative searches using Google Scholar, for example, may reveal additional studies from other countries.
Neonicotinoid compounds
When the number of studies we found for each neonicotinoid compound is compared against global sales data from 2009 (available in [7]) it can be concluded that there is a positive relationship between these two variables. Typically neonicotinoid compounds with higher sales figures have also attracted a greater number of studies. Thiamethoxam deviates the most from this relationship and has been studied less than would be expected from sales data. Imidacloprid, thiamethoxam and clothianidin are all highly toxic to honey bees (acute oral LD50 for A. mellifera: 0.004–0.005 μg per bee, acute contact LD50 for A. mellifera: 0.02–0.08 μg per bee [44]), although these quantities still are appreciably higher than those typically encountered by bees in the environment. They also have similar application methods and crop uses [7]. It is thus unlikely that the greater attention that imidacloprid and clothianidin have received relative to their market shares are due to bees being less at risk of being exposed to, and affected by, thiamethoxam. We conclude that more studies on thiamethoxam are warranted. Similarly, there is a need for comparative studies that assess how well the large numbers of studies on imidacloprid reflect the effects of neonicotinoid compounds as a group.
On the other hand, acetamiprid and thiacloprid, are several orders of magnitude less toxic to honey bees compared to the other neonicotinoids (acute oral LD50 for A. mellifera: 15–17 μg per bee, acute contact LD50 for A. mellifera: 8.1–39 μg per bee) [44]. A similar story seems to be true for the persistence of these compounds as thiacloprid and acetamiprid have shorter half-lives in the environment (3–74 days in soil for thiacloprid, 31–450 for acetamiprid) compared to imidacloprid (28–1250 days), clothianidin (148–6931 days) and thiamethoxam (7–353 days, clothianidin is a primary metabolite) [6]. The comparatively more “bee-friendly” properties of these compounds have, however, led to more liberal usage criteria: for instance, these chemicals are sometimes permitted for use on flowering crops [27]. In the case of the insecticide Biscaya OD 240, containing the active ingredient thiacloprid, the Swedish product information states that it can be used during daylight in flowering crops [45] when bees will be actively foraging. To our knowledge there have been few studies assessing the risks associated with such applications for bees and crop pollination (but see [46]), and this is an area that deserves further attention.
Crop species
Neonicotinoids are used in a wide number of crops, including many fruit and vegetables [8] that often are dependent on pollinators for yield, and are highly attractive to bees [1]. Imidacloprid has at least 140 different crop uses registered [7]. Despite this, we found that research about neonicotinoid impacts on bees has mainly focused on only three, albeit large, crop uses, namely seed treatments in maize, oilseed rape and sunflower.
The most common crop studied was maize (28 studies). These studies primarily investigated potential effects on honey bees resulting from (i) dust from seeds coated with neonicotinoids during sowing (e.g. [28]), (ii) guttation drops formed on treated plants (e.g. [47]), and (iii) pollen collection by bees from seed treated maize (e.g. [48]). In the seven studies each on oilseed rape and sunflowers, the focus was on potential sublethal effects on bees resulting from movement/transfer of neonicotinoids from seed treatments into pollen and nectar [29–30].
Only a few studies investigated fruit or vegetable crops, and most of them examined only the potential for neonicotinoid exposure to bees. For example, soil applications result in markedly higher neonicotinoid residue levels in the pollen and nectar of pumpkin and squash compared with seed treated maize, oilseed rape and sunflower [36,49]. This is an important finding because decisions of what constitutes field-realistic exposure for bees in experiments (e.g. [50–53]) and risk assessment studies ([54–55] but see [56]) are often based on information from seed treated maize, oilseed rape and sunflower. If it turns out that residue levels in the pollen and nectar of these crops (maize, oilseed rape and sunflower) are not representative of other crops, the hazard that neonicotinoids pose for bees might be incorrectly estimated.
Little attention has been given to neonicotinoid use on plants other than field crops, such as lawns with flowering weeds (but see [31,57–59]), fruit trees, home garden plants, ornamental plants, bushes and trees. Information available from studies not included on either Web of Science or PubMed [13], as well as from a recent study on pesticide residues in pollen and nectar from citrus trees treated with imidacloprid [35], suggests that neonicotinoids can be present at higher levels for an extended period of time in woody plants compared to non-woody plants. It was therefore surprising that exposure to bees from neonicotinoid treated woody plants, such as pome fruit, stone fruit and citrus fruit trees, and on ornamental and garden plants [7], has been largely unstudied. Future research should be conducted on a wider array of neonicotinoid treated agricultural crops, including methods of application other than seed treatment, and should more comprehensively assess the consequences for bees of neonicotinoid use on other plants.
Bee species
Most studies (75 percent) included measures of neonicotinoid exposure or effects on A. mellifera. Although the second and third most commonly studied species, the bumble bees B. terrestris and B. impatiens, were included in only eleven and five percent of studies respectively, they were still studied substantially more than any other bee species. Very few studies have examined the effect of neonicotinoids on solitary bees or social bees other than honey bees or bumble bees (e.g. stingless bees [60]).
An interesting question is whether this knowledge gap for bee species other than A. mellifera may be overcome by extrapolating information from honey bees to other bees. Comparative studies in which the same experimental protocols are applied to A. mellifera and other bee taxa can help answer this question. The few studies that have done this for neonicotinoids have found different responses among bee species [61–64]. However, results from a meta-analysis suggest there is a general positive correlation between the lethal toxicity of pesticides for A. mellifera and other bee species [65]. Part of the variation in pesticide toxicity among species might be explained by differences in body size, with larger bee species generally showing less sensitivity to a certain pesticide treatment [65–66]. However, this scope for predictability across bee species considers only lethal pesticide toxicity at the level of the individual bee. These trends may not hold true for the sublethal effects of neonicotinoid exposure. For example, B. terrestris has been found to be more sensitive than A. mellifera for some endpoints, even though the former has a larger body size [62,64]. In addition, there is a substantial variation in ecological, phenological and life-history traits among bee species, such as in preferred nesting habitats, flight seasons and degree of sociality. This implies that both the exposure to neonicotinoids and other pesticides, and their impacts at the colony and population levels, will vary significantly depending on the traits of the bee species [50,67–69]. For these reasons, it is unlikely that studies on A. mellifera can fully explain and/or predict the effects of neonicotinoids on other bee species.
A. mellifera is an important pollinator of many crops [1]. However, wild bees provide a complement to honey bees as pollinators, and the contribution of wild bees to crop pollination has likely been underestimated [70–71]. The reliance on the pollination services from wild bees might also be increasing as crop pollination demands are growing much faster than honey bee stocks [72–73], and wild bees may provide insurance against honey bee declines [74–75]. Future studies should therefore verify that neonicotinoid use does not jeopardize pollination across the wide range of bee species that deliver these essential ecosystem services.
Methodological approaches
Laboratory studies were most common. They are valuable for detecting the mechanisms of neonicotinoid impacts on bees [27]. Many laboratory studies have so far investigated the effects of neonicotinoids at concentrations above the field-realistic range [12]. This is a reasonable first step to verify if any effect is detectable. However, subsequent laboratory studies to investigate an effect that is already known to occur can be significantly improved by testing more realistic exposure scenarios. Increased coverage, especially within the potentially sublethal range of exposures to which bees might be exposed in the field, is needed to increase the predictive power of laboratory studies. Attention should be given not only to field-realistic concentrations, but also to feeding regimes, i.e. how the exposure is likely to occur over time in the field. For example, bumble bee colonies that are exposed to a two-week pulse of neonicotinoids simulating oilseed rape flowering, might be able to compensate for this exposure at the colony level through increased brood production if the exposure period is followed by a period of no neonicotinoid exposure [53]. Another way to increase realistic conditions in laboratory studies is to allow bees to forage for their food in the laboratory [76] or to use combined approaches with field realistic exposures of neonicotinoids in the laboratory and free foraging of bees in the field (e.g. [50–51,77–78]).
Field studies were the second most common methodological approach. Several studies measured only pesticide residues in bees or sources in the environment that bees come into contact with and/or ingest. These studies provide important information about field realistic exposure for bees, but the potential to draw further conclusions about the impacts on bees from such data are limited for several reasons. First, concentrations of neonicotinoids that were found are typically well below levels known to cause acute mortality [25], but they might be at levels that cause sublethal effects [27,79]. This complicates the interpretation of residue studies as sublethal effects are not fully characterised, and their consequences more difficult to assess. Second, metabolism of neonicotinoids within hours after ingestion by bees [64,80–82] may lead to an underestimation of exposure based on data from studies that only sample pesticide residues in bees. Third, for studies that measure neonicotinoids in the environment (e.g. in pollen and nectar, and in guttation fluid [41,83]), it often remains uncertain to what degree individual bees and colonies will be exposed to these sources. Finally, residue studies typically provide only a measure of exposure at one point in time (e.g. [84]). The exposure profile over time, i.e. how long bees are affected by varying concentrations of neonicotinoids in the field, typically remains unknown.
In another similar type of field studies, measures of bee exposure to neonicotinoids were related to effects by observational or correlational approaches. For example, measuring residues in bees suspected to be poisoned by pesticides (e.g. [85]) or in weak versus strong honey bee hives (e.g. [86–87]). Twelve field studies used designs in which bee colonies were placed in or near crop fields that were either treated with a neonicotinoid, or not, and then followed different aspects of bee or colony performance [29,48,88–97]. Recently, two such studies have demonstrated negative effects of seed treatments in oilseed rape on B. terrestris colony performance [29,96]. The study by Rundlöf et al. [29] also showed a negative effect on Osmia bicornis nesting success next to seed treated oilseed rape. No clear effects of neonicotinoids have been found in any of the studies that measured effects on honey bee colony performance, leading to the conclusion that neonicotinoid seed treatments of these crops are safe for honey bees. However, at least two reservations about this conclusion have been raised. First, given the low number of replicates (fields) and high level of intrinsic variability in performances of honey bee colonies, these field studies have low statistical power to detect any sublethal colony effects [12,29,98]. Second, as Goulson [18] rather skeptically pointed out, it is “essentially impossible to conduct a controlled experiment with free-flying bees.” The bee species studied in these field experiments, A. mellifera or B. terrestris, can forage over several kilometres in the landscape [99–100] (for A. mellifera in extreme cases up to 15 km from the hive [101]). This means that in a typical study landscape, bee colonies in control fields may be exposed to neonicotinoids from other fields that have been treated (see e.g. [102]), while bee colonies in treated fields are expected to also visit non-treated fields. A higher level of replication of fields, larger treated areas of crops per field, efforts to standardize bee colonies, and carefully paired treated and untreated fields in similar landscapes may be able to at least partially overcome these problems for honey bee studies. The current restriction on usage of neonicotinoid seed treatments on crops attractive to bees in the EU provides further opportunities to improve this type of field study, because contamination of control fields is less likely. Such trials should be facilitated by policy and authorities in the EU and member states during the current moratorium period.
A final category of field studies consisted of experiments performed in the field with bees that were artificially fed with neonicotinoids after which bee or colony performances were measured (e.g. [103–105]). This type of field study provides the highest level of control. However, these studies have many similar disadvantages as laboratory studies. Although it may not be straightforward to assess, it is often concluded that exposures are at the higher end, or indeed above, those likely to be experienced in the field [27].
Semi-field studies, with cages in field conditions, were only occasionally used. It is a methodological approach that provides an intermediate level of control between field and laboratory studies. Semi-field studies provide a promising opportunity for further research on the side-effects of pesticides on non-target insects [106]. For example, in a semi-field study entire honey bee colonies were enclosed in 200 m² tunnels on flowering seed treated crops throughout the flowering period [92]. This simulates a realistic worst-case scenario in an intensively managed farmland landscape where an entire colony collects all of its food resources from a seed treated crop during its flowering period. Unfortunately, only residues of neonicotinoids in bees and plants were reported in that study, but it is a promising and evidently tractable approach for future studies if they also include cages on control fields and measure behavioural and life history impacts on the bees and their colonies.
Modelling and risk assessment in silico are useful tools for understanding the ecotoxicology of pesticides, especially at higher levels of biological organization [107], but there were few such approaches in the scientific literature on neonicotinoids and bees. Significant regulatory efforts are underway, at least in the EU, to improve risk assessment for neonicotinoid effects on bees [108]. Priorities for the development of risk assessments are to cover the full range of exposure routes and effects that are important for population development and to include bee species other than A. mellifera. Modelling efforts to predict the effects of neonicotinoids on bees at the colony level are still scarce (but see [109–112]), but represent a promising avenue for future expansion. Bee colony models offer a fast and cost effective alternative to large scale field studies, and they are useful tools to synthesize available knowledge and guide further empirical work [110,113]. Population modelling approaches are informative for evaluating the responses of beneficial insects to pesticide disturbance (e.g. [114]), but we are not aware of any models that have been developed to evaluate the effects of neonicotinoids on bees.
Biological levels–from sub-individual to population effects
Studies at the sub-individual level can reveal the mode of action of neonicotinoids on bees [115], provide insights on whether bees can detect and regulate their dietary intake of neonicotinoids [116] and give a better mechanistic understanding about how and when neonicotinoids are expected to interact with other pesticides [117] and bee pathogens [118–119]. However, only a minority of studies at this level made direct links to higher levels of organisation by measuring and reporting consequences for individual bees and the colony (S1 Table, but see e.g. [120]). In general, there is a limited understanding of how sub-individual effects of pesticides translate into consequences for the individual, population and community levels [107].
Studies that measure the effects on the individual were most common. They provide insights on the lethal toxicity of neonicotinoids on bees (e.g. [10]), as well as sublethal effects on factors such as longevity, foraging behaviour, feeding, learning and memory (e.g. [62,78,121–123]). Most studies measured the effects on the adult life stage whereas the effects on brood, larval or pupal stages received much less attention. Exposure to neonicotinoids in early life stages of bees can have both direct lethal effects and delayed sublethal effects on the adults [124,125]. Furthermore, chronic toxicity for bees at time points beyond a standard 10 days test time was rarely considered, even though neonicotinoids might cause delayed and time-cumulative toxicity [50,78,126]. A limitation of measuring only the effects at the individual level is that reinforcement or compensatory responses to pesticide exposure at the colony level are not accounted for. In honey bees, bumble bees and other social bees, the colony is the unit of reproduction, and losses of individual bees can be tolerated up to a certain level without consequences for survival or reproduction of the colony [127]. While most bumble bee studies included measurements of colony effects, few of the honey bee studies did (see S1 Table). Nevertheless, the colony level is of primary interest in managed honey bees, because reduced colony performance and higher rates of colony loss directly affect beekeepers. One reason for the relative paucity of colony level studies may be methodological difficulties of performing replicated studies on honey bees, which have large and variable colony sizes and strengths. Future research should therefore try to overcome these difficulties, for example by increasing sample sizes (see e.g. [98]) and employing increased levels of standardization among colonies in order to determine how neonicotinoids affect honey bee colony performance.
We defined population level effects as a change in the population size across at least one completed life cycle of the study organism. According to this definition we found no studies that measured population effects of neonicotinoid exposure on bees, although several studies indicated that such effects might be prevalent. For example, two studies recently showed significantly reduced queen production in bumble bee (B. terrestris) colonies exposed to neonicotinoids in field settings [29,96]. Further, in one of these studies O. bicornis did not colonize and reproduce in trap nests placed next to treated fields as compared with control fields [29], again suggesting population level effects. Whether such effects on reproductive success ultimately translate into impacts on the population size of wild bees depend on several factors. These include how reproduction is affected by exposure to neonicotinoids across the bee population on a landscape scale, and not just for bees nesting next to treated fields. It also depends on how negative effects on reproduction, caused by exposure to neonicotinoids, in turn affect density-dependent population regulation, for example through decreased competition for nesting and floral resources or through increased risk for Allee effects [27,128]. Possible ways of overcoming the lack of information on the population effects on wild bees would be to monitor bee populations over several seasons in landscapes with different patterns of neonicotinoid use and/or to use population modelling approaches that are informed by experimental data.
Conclusions
Based on our systematic literature review we conclude that despite considerable research efforts, there are still significant knowledge gaps concerning the impacts of neonicotinoids on bees. We found that studies were not representative of the diverse and global use of neonicotinoids that are applied in a multitude of insect pollinated crops and non-crop plants that are often visited by multiple bee species. Additionally, we found opportunities for methodological improvements. Laboratory approaches were most common, and we suggest that their capability to infer real-world consequences can be improved by using information from field studies to inform more field realistic exposures to neonicotinoids. Many of the studies with field approaches examined only potential exposure to neonicotinoids, and we suggest that more field studies should measure bee responses to these chemicals. While the majority of studies measured pesticide effects on individual bees, there is a need for more studies that link effects at the individual to mechanisms at the sub-individual level, and also to consequences for colonies and populations. Current research momentum, and recent rapid increases in the number of studies being published on this topic, provide opportunities to provide a more comprehensive understanding of how neonicotinoids are affecting bees. It is important to keep in mind that our review was based only on studies available on Web of Science and PubMed, and that additional information on the effect of neonicotinoids on bees is available in journals not listed in those databases, reports and regulatory pesticide risk assessments. We expect that additional studies from developing countries, individual lethal bee toxicity and risk assessment data might be available from these additional sources. Increased access and use of such data from regulatory pesticide risk assessments by the scientific community has the potential to improve our evidence base concerning the effects of neonicotinoids on bees [27].
Both managed and wild bees are subject to multiple, interacting environmental pressures [3–4,129]. Bees in modern agricultural landscapes are typically exposed to several classes of pesticides [84], creating opportunities for multiple combined effects of pesticide exposure. For example, the effect of neonicotinoids might be additive for bees in combination with other classes of insecticides such as pyrethroids [50,78] or in-hive pesticides [117], or perhaps even interact synergistically with fungicides [10,63], although the field relevance of some of these interactions has been questioned [46,130]. There is also evidence that neonicotinoids can promote additive or synergistic effects when combined with pathogens or parasites of A. mellifera [118–119,131–134] and B. terrestris [135]: but the importance of neonicotinoid-pathogen interactions under field-realistic conditions might have been overemphasized [136]. Future research must both disentangle how important neonicotinoid use is relative to other potential drivers of bee declines [3,4], as well as determine the identity and magnitude of interactive effects among these drivers in the field.
Author Contributions
Conceived and designed the experiments: OL MR HGS IF RB. Performed the experiments: OL. Analyzed the data: OL. Wrote the paper: OL MR HGS IF RB.
References
- 1. Klein AM, Vaissière BE, Cane JH, Steffan-Dewenter I, Cunningham SA, Kremen C, et al. Importance of pollinators in changing landscapes for world crops. Proc R Soc B. 2007;274: 303–313. pmid:17164193
- 2. Ollerton J, Winfree R, Tarrant S. How many flowering plants are pollinated by animals? Oikos. 2011;120: 321–326.
- 3. Potts SG, Biesmeijer JC, Kremen C, Neumann P, Schweiger O, Kunin WE. Global pollinator declines: trends, impacts and drivers. Trends Ecol Evol. 2010;25: 345–353. pmid:20188434
- 4. Vanbergen AJ, the Insect Pollinators Initiative. Threats to an ecosystem service: pressures on pollinators. Front Ecol Environ. 2013;11: 251–259.
- 5. van der Sluijs JP, Simon-Delso N, Goulson D, Maxim L, Bonmatin JM, Belzunces LP. Neonicotinoids, bee disorders and the sustainability of pollinator services. Curr Opin Environ Sustainability. 2013;5: 293–305.
- 6. Goulson D. An overview of the environmental risks posed by neonicotinoid insecticides. J Appl Ecol. 2013;50: 977–987.
- 7. Jeschke P, Nauen R, Schindler M, Elbert A. Overview of the status and global strategy for neonicotinoids. J Agr Food Chem. 2011;59: 2897–2908.
- 8. Elbert A, Haas M, Springer B, Thielert W, Nauen R. Applied aspects of neonicotinoid uses in crop protection. Pest Manag Sci. 2008;64: 1099–1105. pmid:18561166
- 9. Jeschke P, Nauen R. Neonicotinoids—from zero to hero in insecticide chemistry. Pest Manag Sci. 2008;64: 1084–1098.
- 10. Iwasa T, Motoyama N, Ambrose JT, Roe RM. Mechanism for the differential toxicity of neonicotinoid insecticides in the honey bee, Apis mellifera. Crop Prot. 2004;2: 371–378.
- 11. Nuyttens D, Devarrewaere W, Verboven P, Foqué D. Pesticide‐laden dust emission and drift from treated seeds during seed drilling: a review. Pest Manag Sci. 2013;69: 564–575. pmid:23456984
- 12. Cresswell JE. A meta-analysis of experiments testing the effects of a neonicotinoid insecticide (imidacloprid) on honey bees. Ecotoxicology. 2011;20: 149–157. pmid:21080222
- 13.
Hopwood J, Vaughan M, Shepherd M, Biddinger D, Mader E, Hoffman Black S, et al. Are neonicotinoids killing bees? A review of research into the effects of neonicotinoid insecticides on bees, with recommendations for action. Portland: The Xerces Society for Invertebrate Conservation; 2012.
- 14. Krupke CH, Hunt GJ, Eitzer BD, Andino G, Given K. Multiple routes of pesticide exposure for honey bees living near agricultural fields. PLoS One. 2012;7: e29268. pmid:22235278
- 15. EU. Regulation (EU) No 485/2013. Official Journal of the European Union. 2013;139: 12–26.
- 16.
Ministry of the Environment and Climate Change. Neonicotinoid regulations. 2015. Available: http://www.ontario.ca/environment-and-energy/neonicotinoid-regulations.
- 17. Dicks L. Bees, lies and evidence-based policy. Nature. 2013;494: 283. pmid:23426287
- 18. Goulson D. Neonicotinoids and bees: what's all the buzz? Significance. 2013;10: 6–11.
- 19. Gross M. EU ban puts spotlight on complex effects of neonicotinoids. Curr Biol. 2013;23: R462–R464. pmid:23894744
- 20. Cresswell JE, Desneux N, van Engelsdorp D. Dietary traces of neonicotinoid pesticides as a cause of population declines in honey bees: an evaluation by Hill's epidemiological criteria. Pest Manag Sci. 2012;68: 819–827. pmid:22488890
- 21. Maini S, Medrzycki P, Porrini C. The puzzle of honey bee losses: a brief review. B Insectol. 2010;63: 153–160.
- 22. Farooqui T. A potential link among biogenic amines-based pesticides, learning and memory, and colony collapse disorder: a unique hypothesis. Neurochem Int. 2013;62: 122–136. pmid:23059446
- 23. Fairbrother A, Purdy J, Anderson T, Fell R. Risks of neonicotinoid insecticides to honeybees. Environ Toxicol Chem. 2014;33: 719–731. pmid:24692231
- 24. Smith KM, Loh EH, Rostal MK, Zambrana-Torrelio CM, Mendiola L, Daszak P. Pathogens, pests, and economics: drivers of honey bee colony declines and losses. EcoHealth. 2013;10: 434–445. pmid:24496582
- 25. Blacquière T, Smagghe G, Van Gestel CA, Mommaerts V. Neonicotinoids in bees: a review on concentrations, side-effects and risk assessment. Ecotoxicology. 2012;21: 973–992. pmid:22350105
- 26. Pisa L, Amaral-Rogers V, Belzunces LP, Bonmatin J-M, Downs C, Goulson D, et al. Effects of neonicotinoids and fipronil on non-target invertebrates. Environ Sci Pollut Res. 2014;22: 68–102.
- 27. Godfray HCJ, Blacquière T, Field LM, Hails RS, Petrokofsky G, Potts SG, et al. A restatement of the natural science evidence base concerning neonicotinoid insecticides and insect pollinators. Proc R Soc B. 2014;281: 20140558. pmid:24850927
- 28. Girolami V, Marzaro M, Vivan L, Mazzon L, Giorio C, Marton D, et al. Aerial powdering of bees inside mobile cages and the extent of neonicotinoid cloud surrounding corn drillers. J Appl Entomol. 2013;137: 35–44.
- 29. Rundlöf M, Andersson GK, Bommarco R, Fries I, Hederström V, Herbertsson L, et al. Seed coating with a neonicotinoid insecticide negatively affects wild bees. Nature. 2015;521: 77–80. pmid:25901681
- 30. Schmuck R, Schöning R, Stork A, Schramel O. Risk posed to honeybees (Apis mellifera L, Hymenoptera) by an imidacloprid seed dressing of sunflowers. Pest Manag Sci. 2001;57: 225–238. pmid:11455652
- 31. Larson JL, Redmond CT, Potter DA. Mowing mitigates bioactivity of neonicotinoid insecticides in nectar of flowering lawn weeds and turfgrass guttation. Environ Toxicol Chem. 2015;34: 127–132. pmid:25319809
- 32. Hoffmann EJ, Castle SJ. Imidacloprid in melon guttation fluid: a potential mode of exposure for pest and beneficial organisms. J Econ Entomol. 2012;105: 67–71. pmid:22420257
- 33. Stewart SD, Lorenz GM, Catchot AL, Gore J, Cook D, Skinner J, et al. Potential exposure of pollinators to neonicotinoid insecticides from the use of insecticide seed treatments in the mid-southern United States. Environ Sci Tech. 2014;48: 9762–9769.
- 34. Sechser B, Freuler J. The impact of thiamethoxam on bumble bee broods (Bombus terrestris L.) following drip application in covered tomato crops. J Pest Sci. 2003;76: 74–77.
- 35. Byrne FJ, Visscher PK, Leimkuehler B, Fischer D, Grafton‐Cardwell EE, Morse JG. Determination of exposure levels of honey bees foraging on flowers of mature citrus trees previously treated with imidacloprid. Pest Manag Sci. 2014;70: 470–482. pmid:23788449
- 36. Stoner KA, Eitzer BD. Movement of soil-applied imidacloprid and thiamethoxam into nectar and pollen of squash (Cucurbita pepo). PLoS One. 2012;7: e39114. pmid:22761727
- 37. Skerl MIS, Bolta SV, Cesnik HB, Gregorc A. Residues of pesticides in honeybee (Apis mellifera carnica) bee bread and in pollen loads from treated apple orchards. B Environ Contam Tox. 2009;83: 374–377.
- 38. Choudhary A, Sharma DC. Dynamics of pesticide residues in nectar and pollen of mustard (Brassica juncea (L.) Czern.) grown in Himachal Pradesh (India). Environ Monit Assess. 2008;144: 143–150. pmid:17952621
- 39. Chen M, Tao L, McLean J, Lu C. Quantitative analysis of neonicotinoid insecticide residues in foods: implication for dietary exposures. J Agric Food Chem. 2014;62: 6082–6090. pmid:24933495
- 40. Vaikkinen A, Schmidt HS, Kiiski I, Rämö S, Hakala K, Haapala M, et al. Analysis of neonicotinoids from plant material by desorption atmospheric pressure photoionization‐mass spectrometry. Rapid Commun Mass Spectrom. 2015;29: 424–430.
- 41. Reetz JE, Zühlke S, Spiteller M, Wallner K. Neonicotinoid insecticides translocated in guttated droplets of seed-treated maize and wheat: a threat to honeybees? Apidologie. 2011;42: 596–606.
- 42. Schreinemachers P, Tipraqsa P. Agricultural pesticides and land use intensification in high, middle and low income countries. Food Policy. 2012;37: 616–626.
- 43. Gallai N, Salles JM, Settele J, Vaissière BE. Economic valuation of the vulnerability of world agriculture confronted with pollinator decline. Ecol Econ. 2009;68: 810–821.
- 44. European Food Safety Authority. Statement on the findings in recent studies investigating sub-lethal effects in bees of some neonicotinoids in consideration of the uses currently authorised in Europe. EFSA Journal. 2012;10: 2752.
- 45.
Bayer CropScience. Product information for Biscaya OD 240 (in Swedish). 2015. Available: http://www.cropscience.bayer.se/~/media/Bayer%20CropScience/Scandinavia/Sweden/Produkter/Produktsidor%202015/Biscaya%202015.ashx.
- 46. Schmuck R, Stadler T, Schmidt HW. Field relevance of a synergistic effect observed in the laboratory between an EBI fungicide and a chloronicotinyl insecticide in the honeybee (Apis mellifera L, Hymenoptera). Pest Manag Sci. 2003;59: 279–286. pmid:12639044
- 47. Girolami V, Mazzon L, Squartini A, Mori N, Marzaro M, Di Bernardo A, et al. Translocation of neonicotinoid insecticides from coated seeds to seedling guttation drops: a novel way of intoxication for bees. J Econ Entomol. 2009;102: 1808–1815. pmid:19886445
- 48. Nguyen BK, Saegerman C, Pirard C, Mignon J, Widart J, Thirionet B, et al. Does imidacloprid seed-treated maize have an impact on honey bee mortality? J Econ Entomol. 2009;102: 616–623. pmid:19449641
- 49. Dively GP, Kamel A. Insecticide residues in pollen and nectar of a cucurbit crop and their potential exposure to pollinators. J Agr Food Chem. 2012;60: 4449–4456.
- 50. Gill RJ, Ramos-Rodriguez O, Raine NE. Combined pesticide exposure severely affects individual-and colony-level traits in bees. Nature. 2012;491: 105–108. pmid:23086150
- 51. Whitehorn PR, O’Connor S, Wackers FL, Goulson D. Neonicotinoid pesticide reduces bumble bee colony growth and queen production. Science. 2012;336: 351–352. pmid:22461500
- 52. Elston C, Thompson HM, Walters KF. Sub-lethal effects of thiamethoxam, a neonicotinoid pesticide, and propiconazole, a DMI fungicide, on colony initiation in bumblebee (Bombus terrestris) micro-colonies. Apidologie. 2013;44: 563–574.
- 53. Laycock I, Cresswell JE. Repression and recuperation of brood production in Bombus terrestris bumble bees exposed to a pulse of the neonicotinoid pesticide imidacloprid. PLoS One. 2013;8: e79872. pmid:24224015
- 54. Rortais A, Arnold G, Halm MP, Touffet-Briens F. Modes of honeybees exposure to systemic insecticides: estimated amounts of contaminated pollen and nectar consumed by different categories of bees. Apidologie. 2005;36: 71–83.
- 55. Halm MP, Rortais A, Arnold G, Tasei JN, Rault S. New risk assessment approach for systemic insecticides: the case of honey bees and imidacloprid (Gaucho). Environ Sci Technol. 2006;40: 2448–2454. pmid:16646488
- 56. Sanchez-Bayo F, Goka K. Pesticide residues and bees–a risk assessment. PLoS One. 2014;9: e94482. pmid:24718419
- 57. Gels JA, Held DW, Potter DA. Hazards of insecticides to the bumble bees Bombus impatiens (Hymenoptera: Apidae) foraging on flowering white clover in turf. J Econ Entomol. 2002;95: 722–728. pmid:12216812
- 58. Larson JL, Redmond CT, Potter DA. Assessing insecticide hazard to bumble bees foraging on flowering weeds in treated lawns. PLoS One. 2013;8: e66375. pmid:23776667
- 59. Larson JL, Redmond CT, Potter DA. Impacts of a neonicotinoid, neonicotinoid–pyrethroid premix, and anthranilic diamide insecticide on four species of turf-inhabiting beneficial insects. Ecotoxicology. 2014;23: 252–259. pmid:24493235
- 60. Barbosa WF, Smagghe G, Guedes RNC. Pesticides and reduced‐risk insecticides, native bees and pantropical stingless bees: pitfalls and perspectives. Pest Manag Sci. 2015;71: 1049–1053. pmid:25892651
- 61. Stark JD, Jepson PC, Mayer DF. Limitation to the use of topical toxicity data for prediction of pesticide side-effect in the field. J Econ Entomol. 1995;88: 1081–1088.
- 62. Cresswell JE, Page CJ, Uygun MB, Holmbergh M, Li Y, Wheeler JG, et al. Differential sensitivity of honey bees and bumble bees to a dietary insecticide (imidacloprid). Zoology. 2012;115: 365–371. pmid:23044068
- 63. Biddinger DJ, Robertson JL, Mullin C, Frazier J, Ashcraft SA, Rajotte EG, et al. Comparative toxicities and synergism of apple orchard pesticides to Apis mellifera (L.) and Osmia cornifrons (Radoszkowski). PLoS One. 2013;8: e72587. pmid:24039783
- 64. Cresswell JE, Robert FXL, Florance H, Smirnoff N. Clearance of ingested neonicotinoid pesticide (imidacloprid) in honey bees (Apis mellifera) and bumblebees (Bombus terrestris). Pest Manag Sci. 2014;70: 332–337. pmid:23633150
- 65. Arena M, Sgolastra F. A meta-analysis comparing the sensitivity of bees to pesticides. Ecotoxicology. 2014;23: 324–334. pmid:24435220
- 66. Devillers J, Decourtye A, Budzinski H, Pham-Delègue MH, Cluzeau S, Maurin G. Comparative toxicity and hazards of pesticides to Apis and non-Apis bees. A chemometrical study. SAR QSAR Environ Res. 2003;14: 389–403. pmid:14758982
- 67. Thompson HM, Hunt LV. Extrapolating from honeybees to bumblebees in pesticide risk assessment. Ecotoxicology. 1999;8: 147–166.
- 68. Williams NM, Crone EE, Minckley RL, Packer L, Potts SG. Ecological and life-history traits predict bee species responses to environmental disturbances. Biol Conserv. 2010;143: 2280–2291.
- 69. Brittain C, Potts SG. The potential impacts of insecticides on the life-history traits of bees and the consequences for pollination. Basic Appl Ecol. 2011;12: 321–331.
- 70. Breeze TD, Bailey AP, Balcombe KG, Potts SG. Pollination services in the UK: how important are honeybees? Agr Ecosyst Environ. 2011;142: 137–143.
- 71. Garibaldi LA, Steffan-Dewenter I, Winfree R, Aizen MA, Bommarco R, Cunningham SA, et al. Wild pollinators enhance fruit set of crops regardless of honey bee abundance. Science. 2013;339: 1608–1611. pmid:23449997
- 72. Aizen MA, Harder LD. The global stock of domesticated honey bees is growing slower than agricultural demand for pollination. Curr Biol. 2009;19: 915–918. pmid:19427214
- 73. Breeze TD, Vaissière BE, Bommarco R, Petanidou T, Seraphides N, Kozák L, et al. Agricultural policies exacerbate honeybee pollination service supply-demand mismatches across Europe. PLoS One. 2014;9: e82996. pmid:24421873
- 74. Kremen C, Williams NM, Thorp RW. Crop pollination from native bees at risk from agricultural intensification. Proc Natl Acad Sci USA. 2002;99: 16812–16816. pmid:12486221
- 75. Winfree R, Williams NM, Dushoff J, Kremen C. Native bees provide insurance against ongoing honey bee losses. Ecol Lett. 2007;10: 1105–1113. pmid:17877737
- 76. Mommaerts V, Reynders S, Boulet J, Besard L, Sterk G, Smagghe G. Risk assessment for side-effects of neonicotinoids against bumblebees with and without impairing foraging behavior. Ecotoxicology. 2010;19: 207–215. pmid:19757031
- 77. Feltham H, Park K, Goulson D. Field realistic doses of pesticide imidacloprid reduce bumblebee pollen foraging efficiency. Ecotoxicology. 2014;23: 317–323. pmid:24448674
- 78. Gill RJ, Raine NE. Chronic impairment of bumblebee natural foraging behaviour induced by sublethal pesticide exposure. Funct Ecol. 2014;28: 1459–1471.
- 79. Desneux N, Decourtye A, Delpuech JM. The sublethal effects of pesticides on beneficial arthropods. Annu Rev Entomol. 2007;52: 81–106. pmid:16842032
- 80. Suchail S, Debrauwer L, Belzunces LP. Metabolism of imidacloprid in Apis mellifera. Pest Manag Sci. 2004;60: 291–296. pmid:15025241
- 81. Suchail S, De Sousa G, Rahmani R, Belzunces LP. In vivo distribution and metabolisation of 14C‐imidacloprid in different compartments of Apis mellifera L. Pest Manag Sci. 2004;60: 1056–1062. pmid:15532678
- 82. Brunet JL, Badiou A, Belzunces LP. In vivo metabolic fate of (14C)‐acetamiprid in six biological compartments of the honeybee, Apis mellifera L. Pest Manag Sci. 2005;61: 742–748. pmid:15880574
- 83. Bonmatin JM, Moineau I, Charvet R, Fleche C, Colin ME, Bengsch ER. A LC/APCI-MS/MS Method for analysis of imidacloprid in soils, in plants, and in pollens. Anal Chem. 2003;75: 2027–2033. pmid:12720336
- 84. Mullin CA, Frazier M, Frazier JL, Ashcraft S, Simonds R, vanEngelsdorp D, et al. High levels of miticides and agrochemicals in North American apiaries: implications for honey bee health. PLoS One. 2010;5: e9754. pmid:20333298
- 85. Chauzat MP, Martel AC, Blanchard P, Clemént MC, Schurr F, Lair C, et al. A case report of a honey bee colony poisoning incident in France. J Apicult Res. 2010;49: 113–115.
- 86. Chauzat MP, Carpentier P, Martel AC, Bougeard S, Cougoule N, Porta P, et al. Influence of pesticide residues on honey bee (Hymenoptera: Apidae) colony health in France. Environ Entomol. 2009;38: 514–523. pmid:19508759
- 87. Pareja L, Colazzo M, Pérez-Parada A, Niell S, Carrasco-Letelier L, Besil N, et al. Detection of pesticides in active and depopulated beehives in Uruguay. Int J Environ Res Publ Health. 2011;8: 3844–3858.
- 88. Tasei JN, Ripault G, Rivault E. Hazards of imidacloprid seed coating to Bombus terrestris (Hymenoptera: Apidae) when applied to sunflower. J Econ Entomol. 2001;94: 623–627. pmid:11425015
- 89. Cutler GC, Scott-Dupree CD. Exposure to clothianidin seed-treated canola has no long-term impact on honey bees. J Econ Entomol. 2007;100: 765–772. pmid:17598537
- 90. Ondo Zue Abaga N, Alibert P, Dousset S, Savadogo PW, Savadogo M, Sedogo M. Insecticide residues in cotton soils of Burkina Faso and effects of insecticides on fluctuating asymmetry in honey bees (Apis mellifera Linnaeus). Chemosphere. 2011;83: 585–592. pmid:21190716
- 91. Boily M, Sarrasin B, DeBlois C, Aras P, Chagnon M. Acetylcholinesterase in honey bees (Apis mellifera) exposed to neonicotinoids, atrazine and glyphosate: laboratory and field experiments. Environ Sci Pollut Res. 2013;20: 5603–5614.
- 92. Pilling E, Campbell P, Coulson M, Ruddle N, Tornier I. A four-year field program investigating long-term effects of repeated exposure of honey bee colonies to flowering crops treated with thiamethoxam. PLoS One. 2013;8: e77193. pmid:24194871
- 93. Cutler GC, Scott-Dupree CD. A field study examining the effects of exposure to neonicotinoid seed-treated corn on commercial bumble bee colonies. Ecotoxicology. 2014;23: 1755–1763. pmid:25194943
- 94. Cutler GC, Scott-Dupree CD, Sultan M, McFarlane AD, Brewer L. A large-scale field study examining effects of exposure to clothianidin seed-treated canola on honey bee colony health, development, and overwintering success. PeerJ. 2014;2: e652. pmid:25374790
- 95. Pohorecka K, Skubida P, Miszczak A, Semkiw P, Sikorski P, Zagibajło K, et al. Residues of neonicotinoid insecticides in bee collected plant materials from oilseed rape crops and their effect on bee colonies. J Apic Sci. 2012;56: 115–134.
- 96. Goulson D. Neonicotinoids impact bumblebee colony fitness in the field; a reanalysis of the UK’s Food & Environment Research Agency 2012 experiment. PeerJ. 2015;3: e854. pmid:25825679
- 97. Alburaki M, Boutin S, Mercier PL, Loublier Y, Chagnon M, Derome N. Neonicotinoid-coated Zea mays seeds indirectly affect honeybee performance and pathogen susceptibility in field trials. PLoS One. 2015;10: e0125790. pmid:25993642
- 98. Pirk CW, de Miranda JR, Kramer M, Murray TE, Nazzi F, Shutler D, et al. Statistical guidelines for A. mellifera research. J Apicult Res. 2013;52: 52.4.13.
- 99. Steffan-Dewenter I, Kuhn A. Honeybee foraging in differentially structured landscapes. Proc R Soc B. 2003;270: 569–575. pmid:12769455
- 100. Walther‐Hellwig K, Frankl R. Foraging habitats and foraging distances of bumblebees, Bombus spp. (Hym., Apidae), in an agricultural landscape. J Appl Entomol. 2000;124: 299–306.
- 101. Beekman M, Ratnieks FLW. Long-range foraging by the honey-bee, Apis mellifera L. Funct Ecol. 2000;14: 490–496.
- 102.
The Food and Environment Research Agency. Effects of neonicotinoid seed treatments on bumble bee colonies under field conditions. 2013. Available: http://fera.co.uk/ccss/documents/defraBumbleBeeReportPS2371V4a.pdf.
- 103. Faucon JP, Aurières C, Drajnudel P, Mathieu L, Ribiere M, Martel AC, et al. Experimental study on the toxicity of imidacloprid given in syrup to honey bee (A. mellifera) colonies. Pest Manag Sci. 2005;61: 111–125. pmid:15619715
- 104. Lu C, Warchol KM, Callahan RA. Sub-lethal exposure to neonicotinoids impaired honey bees winterization before proceeding to colony collapse disorder. B Insectol. 2014;67: 125–130.
- 105. Sandrock C, Tanadini M, Tanadini LG, Fauser-Misslin A, Potts SG, Neumann P. Impact of chronic neonicotinoid exposure on honeybee colony performance and queen supersedure. PLoS One. 2014;9: e103592. pmid:25084279
- 106. Macfadyen S, Banks JE, Stark JD, Davies AP. Using semifield studies to examine the effects of pesticides on mobile terrestrial invertebrates. Annu Rev Entomol. 2014;59: 383–404. pmid:24160417
- 107. Köhler HR, Triebskorn R. Wildlife ecotoxicology of pesticides: can we track effects to the population level and beyond? Science. 2013;341: 759–765. pmid:23950533
- 108.
European Food Safety Authority. Bee health. 2015. Available: http://www.efsa.europa.eu/en/topics/topic/beehealth.
- 109. Henry M, Beguin M, Requier F, Rollin O, Odoux JF, Aupinel P, et al. A common pesticide decreases foraging success and survival in honey bees. Science. 2012;336: 348–350. pmid:22461498
- 110. Bryden J, Gill RJ, Mitton RA, Raine NE, Jansen VA. Chronic sublethal stress causes bee colony failure. Ecol Lett. 2013;16: 1463–1469. pmid:24112478
- 111. Henry M, Bertrand C, Le Féon V, Requier F, Odoux JF, Aupinel P, et al. Pesticide risk assessment in free-ranging bees is weather and landscape dependent. Nature Comm. 2014;5: 4359.
- 112. Cresswell JE, Thompson HM. Comment on “A common pesticide decreases foraging success and survival in honey bees”. Science. 2012;337: 1453. pmid:22997307
- 113. Becher MA, Grimm V, Thorbek P, Horn J, Kennedy PJ, Osborne JL. BEEHAVE: a systems model of honeybee colony dynamics and foraging to explore multifactorial causes of colony failure. J Appl Ecol. 2014;51: 470–482. pmid:25598549
- 114. Banks JE, Stark JD, Vargas RI, Ackleh AS. Deconstructing the surrogate species concept: a life history approach to the protection of ecosystem services. Ecol Appl. 2014;24: 770–778. pmid:24988775
- 115. Tomizawa M, Yamamoto I. Binding of nicotinoids and the related compounds to the insect nicotinic acetylcholine receptor. J Pestic Sci. 1992;17: 231–236.
- 116. Kessler SC, Tiedeken EJ, Simcock KL, Derveau S, Mitchell J, Softley S, et al. Bees prefer foods containing neonicotinoid pesticides. Nature. 2015;521: 74–76. pmid:25901684
- 117. Palmer MJ, Moffat C, Saranzewa N, Harvey J, Wright GA, Connolly CN. Cholinergic pesticides cause mushroom body neuronal inactivation in honeybees. Nat Commun. 2013;4: 1634. pmid:23535655
- 118. Alaux C, Brunet JL, Dussaubat C, Mondet F, Tchamitchan S, Cousin M, et al. Interactions between Nosema microspores and a neonicotinoid weaken honeybees (A. mellifera). Environ Microbiol. 2010;12: 774–782. pmid:20050872
- 119. Di Prisco G, Cavaliere V, Annoscia D, Varricchio P, Caprio E, Nazzi F, et al. Neonicotinoid clothianidin adversely affects insect immunity and promotes replication of a viral pathogen in honey bees. Proc Natl Acad Sci USA. 2013;110: 18466–18471. pmid:24145453
- 120. Moffat C, Pacheco JG, Sharp S, Samson AJ, Bollan KA, Huang J, et al. Chronic exposure to neonicotinoids increases neuronal vulnerability to mitochondrial dysfunction in the bumblebee (Bombus terrestris). FASEB J. 2015;29: 2112–2119. pmid:25634958
- 121. Decourtye A, Armengaud C, Renou M, Devillers J, Cluzeau S, Gauthier M, et al. Imidacloprid impairs memory and brain metabolism in the honeybee (A. mellifera L.). Pestic Biochem Phys. 2004;78: 83–92.
- 122. Decourtye A, Devillers J, Cluzeau S, Charreton M, Pham-Delègue MH. Effects of imidacloprid and deltamethrin on associative learning in honeybees under semi-field and laboratory conditions. Ecotox Environ Safe. 2004;57: 410–419.
- 123. Schneider CW, Tautz J, Grünewald B, Fuchs S. RFID tracking of sublethal effects of two neonicotinoid insecticides on the foraging behavior of A. mellifera. PLoS One. 2012;7: e30023. pmid:22253863
- 124. Yang EC, Chang HC, Wu WY, Chen YW. Impaired olfactory associative behavior of honeybee workers due to contamination of imidacloprid in the larval stage. PLoS One. 2012;7: e49472. pmid:23166680
- 125. Tan K, Chen W, Dong S, Liu X, Wang Y, Nieh JC. A neonicotinoid impairs olfactory learning in Asian honey bees (Apis cerana) exposed as larvae or as adults. Sci Rep. 2015;5 10989. pmid:26086769
- 126. Rondeau G, Sánchez-Bayo F, Tennekes HA, Decourtye A, Ramírez-Romero R, Desneux N. Delayed and time-cumulative toxicity of imidacloprid in bees, ants and termites. Sci Rep. 2014;4: 5566. pmid:24993452
- 127. Schmid-Hempel P, Heeb D. Worker mortality and colony development in bumblebees, Bombus lucorum (L.)(Hymenoptera, Apidae). Mitt Schweiz Entomol Ges. 1991;64: 93–108.
- 128. Roulston TAH, Goodell K. The role of resources and risks in regulating wild bee populations. Annu Rev Entomol. 2011;56: 293–312. pmid:20822447
- 129. González-Varo JP, Biesmeijer JC, Bommarco R, Potts SG, Schweiger O, Smith HG, et al. Combined effects of global change pressures on animal-mediated pollination. Trends Ecol Evol. 2013;28: 524–530. pmid:23746938
- 130. Thompson HM, Fryday SL, Harkin S, Milner S. Potential impacts of synergism in honeybees (A. mellifera) of exposure to neonicotinoids and sprayed fungicides in crops. Apidologie. 2014;45: 545–553.
- 131. Vidau C, Diogon M, Aufauvre J, Fontbonne R, Viguès B, Brunet JL, et al. Exposure to sublethal doses of fipronil and thiacloprid highly increases mortality of honeybees previously infected by Nosema ceranae. PLoS One. 2011;6: e21550. pmid:21738706
- 132. Pettis JS, Johnson J, Dively G. Pesticide exposure in honey bees results in increased levels of the gut pathogen Nosema. Naturwissenschaften. 2012;99: 153–158. pmid:22246149
- 133. Doublet V, Labarussias M, Miranda JR, Moritz RF, Paxton RJ. Bees under stress: sublethal doses of a neonicotinoid pesticide and pathogens interact to elevate honey bee mortality across the life cycle. Environ Microbiol. 2015;17: 969–983. pmid:25611325
- 134. Retschnig G, Neumann P, Williams GR. Thiacloprid-Nosema ceranae interactions in honey bees: host survivorship but not parasite reproduction is dependent on pesticide dose. J Invertebr Pathol. 2014;118: 18–19. pmid:24594300
- 135. Fauser‐Misslin A, Sadd BM, Neumann P, Sandrock C. Influence of combined pesticide and parasite exposure on bumblebee colony traits in the laboratory. J Appl Ecol. 2014;51: 450–459.
- 136.
Retschnig G, Williams GR, Odemer R, Boltin J, Di Poto C, Mehmann MM, et al. Effects, but no interactions, of ubiquitous pesticide and parasite stressors on honey bee (Apis mellifera) lifespan and behaviour in a colony environment. Environ Microbiol. 2015; In press.