Ecology and Management of African Honey Bees (Apis mellifera L.).

In Africa, humans evolved as honey hunters of honey bee subspecies adapted to diverse geographical regions. Beekeeping today is practiced much as it was when Africans moved from honey hunting to beekeeping nearly 5,000 years ago, with beekeepers relying on seasonally available wild bees. Research suggests that populations are resilient, able to resist diseases and novel parasites. Distinct biomes, as well as environmental pressures, shaped the behavior and biology of these bees and in turn influenced how indigenous beekeeping developed. It appears that passive beekeeping practices that enabled free-living populations contributed to the overall resilience and health of the bee. There is clearly a need for research aimed at a deeper understanding of bee biology and the ecosystems from which they benefit and on which humans depend, as well as a growing realization that the management of these bees requires an indigenous approach that reflects a broader knowledge base and the economics of local communities and markets.

Colony Size, Rather Than Geographic Origin of Stocks, Predicts Overwintering Success in Honey Bees (Hymenoptera: Apidae) in the Northeastern United States.

Honey bees (Apis mellifera L.) are key pollinators of agricultural crops. However, approximately 30% of managed colonies die each winter in the United States. There has been great interest in breeding for 'locally adapted stocks' which survive winter conditions in a particular region. Here, we evaluate the impact of geographic origin of stock on colony weight, population size, and overwintering survival. Comparing four different U.S. honey bee stocks (two bred in southern and two bred in northern regions) under standard beekeeping practices in three different apiary locations in central Pennsylvania, we examined possible adaptation of these stocks to temperate conditions. We confirmed the genotypic difference among the stocks from different geographic origins via microsatellite analysis. We found that stock or region of origin was not correlated with weight, population size, or overwintering success. However, overwintering success was influenced by the weight and population size the colonies reached prior to winter where higher colony weight is a strong predictor of overwintering survival. Although the number of locations used in this study was limited, the difference in average colony sizes from different locations may be attributable to the abundance and diversity of floral resources near the honey bee colonies. Our results suggest that 1) honey bees may use similar strategies to cope with environmental conditions in both southern and northern regions, 2) colonies must reach a population size threshold to survive adverse conditions (an example of the Allee effect), and 3) landscape nutrition is a key component to colony survival.

Investigating the viral ecology of global bee communities with high-throughput metagenomics.

Bee viral ecology is a fascinating emerging area of research: viruses exert a range of effects on their hosts, exacerbate impacts of other environmental stressors, and, importantly, are readily shared across multiple bee species in a community. However, our understanding of bee viral communities is limited, as it is primarily derived from studies of North American and European Apis mellifera populations. Here, we examined viruses in populations of A. mellifera and 11 other bee species from 9 countries, across 4 continents and Oceania. We developed a novel pipeline to rapidly and inexpensively screen for bee viruses. This pipeline includes purification of encapsulated RNA/DNA viruses, sequence-independent amplification, high throughput sequencing, integrated assembly of contigs, and filtering to identify contigs specifically corresponding to viral sequences. We identified sequences for (+)ssRNA, (-)ssRNA, dsRNA, and ssDNA viruses. Overall, we found 127 contigs corresponding to novel viruses (i.e. previously not observed in bees), with 27 represented by >0.1% of the reads in a given sample, and 7 contained an RdRp or replicase sequence which could be used for robust phylogenetic analysis. This study provides a sequence-independent pipeline for viral metagenomics analysis, and greatly expands our understanding of the diversity of viruses found in bee communities.

Field Residues and Effects of the Insect Growth Regulator Novaluron and Its Major Co-Formulant N-Methyl-2-Pyrrolidone on Honey Bee Reproduction and Development.

Owing to the recent declines in honey bee (Apis mellifera L.) populations, there is a need for field and laboratory studies to investigate threats to pollinator health. This study examines the hypothesis that the organophosphate alternative, Rimon 0.83EC, can have consequences to honey bee health by combining newly acquired field residue data, laboratory bioassays, and colony level feeding studies. Following label rate applications of Rimon 0.83EC to apple trees, average residue concentrations of the active ingredient, novaluron, were found to be 3.38 ppm in tree-collected pollen. Residues of the major co-formulant in Rimon 0.83EC, N-methyl-2-pyrrolidone (NMP), were below the limit of detection in the field, but a growth chamber study described here found that NMP can persist in pollen for up to 7 d with average concentrations of 69.3 ppm. Concurrent larval rearing studies found novaluron and NMP to be toxic to developing honey bees at doses as low as 100 ppb and 100 ppm, respectively. Nucleus colony feeding studies found that chronic exposure to Rimon 0.83EC at doses as low as 200 ppm (18.6 ppm novaluron) can result in interruptions to brood production that can last for up to 2 wk after exposure. Taken together, these data indicate the use of Rimon 0.83EC on blooming flowers is a significant threat to honey bee reproduction, and suggest the need for more strict and clear usage guidelines.

In-hive Pesticide Exposome: Assessing risks to migratory honey bees from in-hive pesticide contamination in the Eastern United States.

This study measured part of the in-hive pesticide exposome by analyzing residues from live in-hive bees, stored pollen, and wax in migratory colonies over time and compared exposure to colony health. We summarized the pesticide burden using three different additive methods: (1) the hazard quotient (HQ), an estimate of pesticide exposure risk, (2) the total number of pesticide residues, and (3) the number of relevant residues. Despite being simplistic, these models attempt to summarize potential risk from multiple contaminations in real-world contexts. Colonies performing pollination services were subject to increased pesticide exposure compared to honey-production and holding yards. We found clear links between an increase in the total number of products in wax and colony mortality. In particular, we found that fungicides with particular modes of action increased disproportionally in wax within colonies that died. The occurrence of queen events, a significant risk factor for colony health and productivity, was positively associated with all three proxies of pesticide exposure. While our exposome summation models do not fully capture the complexities of pesticide exposure, they nonetheless help elucidate their risks to colony health. Implementing and improving such models can help identify potential pesticide risks, permitting preventative actions to improve pollinator health.

Toxicological Risks of Agrochemical Spray Adjuvants: Organosilicone Surfactants May Not Be Safe.

Agrochemical risk assessment that takes into account only pesticide active ingredients without the spray adjuvants commonly used in their application will miss important toxicity outcomes detrimental to non-target species, including humans. Lack of disclosure of adjuvant and formulation ingredients coupled with a lack of adequate analytical methods constrains the assessment of total chemical load on beneficial organisms and the environment. Adjuvants generally enhance the pesticidal efficacy and inadvertently the non-target effects of the active ingredient. Spray adjuvants are largely assumed to be biologically inert and are not registered by the USA EPA, leaving their regulation and monitoring to individual states. Organosilicone surfactants are the most potent adjuvants and super-penetrants available to growers. Based on the data for agrochemical applications to almonds from California Department of Pesticide Regulation, there has been increasing use of adjuvants, particularly organosilicone surfactants, during bloom when two-thirds of USA honey bee colonies are present. Increased tank mixing of these with ergosterol biosynthesis inhibitors and other fungicides and with insect growth regulator insecticides may be associated with recent USA honey bee declines. This database archives every application of a spray tank adjuvant with detail that is unprecedented globally. Organosilicone surfactants are good stand alone pesticides, toxic to bees, and are also present in drug and personal care products, particularly shampoos, and thus represent an important component of the chemical landscape to which pollinators and humans are exposed. This mini review is the first to possibly link spray adjuvant use with declining health of honey bee populations.

Early-life experience affects honey bee aggression and resilience to immune challenge.

Early-life social experiences cause lasting changes in behavior and health for a variety of animals including humans, but it is not well understood how social information ''gets under the skin'' resulting in these effects. Adult honey bees (Apis mellifera) exhibit socially coordinated collective nest defense, providing a model for social modulation of aggressive behavior. Here we report for the first time that a honey bee's early-life social environment has lasting effects on individual aggression: bees that experienced high-aggression environments during pre-adult stages showed increased aggression when they reached adulthood relative to siblings that experienced low-aggression environments, even though all bees were kept in a common environment during adulthood. Unlike other animals including humans however, high-aggression honey bees were more, rather than less, resilient to immune challenge, assessed as neonicotinoid pesticide susceptibility. Moreover, aggression was negatively correlated with ectoparasitic mite presence. In honey bees, early-life social experience has broad effects, but increased aggression is decoupled from negative health outcomes. Because honey bees and humans share aspects of their physiological response to aggressive social encounters, our findings represent a step towards identifying ways to improve individual resiliency. Pre-adult social experience may be crucial to the health of the ecologically threatened honey bee.

Assessing Honey Bee (Hymenoptera: Apidae) Foraging Populations and the Potential Impact of Pesticides on Eight U.S. Crops.

Beekeepers who use honey bees (Apis mellifera L.) for crop pollination services, or have colonies making honey on or in close proximity to agricultural crops, are concerned about the reductions of colony foragers and ultimate weakening of their colonies. Pesticide exposure is a potential factor in the loss of foragers. During 2009-2010, we assessed changes in the field force populations of 9-10 colonies at one location per crop on each of the eight crops by counting departing foragers leaving colonies at regular intervals during the respective crop blooming periods. The number of frames of adult bees was counted before and after bloom period. For pesticide analysis, we collected dead and dying bees near the hives, returning foragers, crop flowers, trapped pollen, and corn-flowers associated with the cotton crop. The number of departing foragers changed over time in all crops except almonds; general patterns in foraging activity included declines (cotton), noticeable peaks and declines (alfalfa, blueberries, cotton, corn, and pumpkins), and increases (apples and cantaloupes). The number of adult bee frames increased or remained stable in all crops except alfalfa and cotton. A total of 53 different pesticide residues were identified in samples collected across eight crops. Hazard quotients (HQ) were calculated for the combined residues for all crop-associated samples and separately for samples of dead and dying bees. A decrease in the number of departing foragers in cotton was one of the most substantial crop-associated impacts and presented the highest pesticide risk estimated by a summed pesticide residue HQ.

Genome-wide analysis of signatures of selection in populations of African honey bees (Apis mellifera) using new web-based tools.

BACKGROUND: With the development of inexpensive, high-throughput sequencing technologies, it has become feasible to examine questions related to population genetics and molecular evolution of non-model species in their ecological contexts on a genome-wide scale. Here, we employed a newly developed suite of integrated, web-based programs to examine population dynamics and signatures of selection across the genome using several well-established tests, including F ST, pN/pS, and McDonald-Kreitman. We applied these techniques to study populations of honey bees (Apis mellifera) in East Africa. In Kenya, there are several described A. mellifera subspecies, which are thought to be localized to distinct ecological regions. RESULTS: We performed whole genome sequencing of 11 worker honey bees from apiaries distributed throughout Kenya and identified 3.6 million putative single-nucleotide polymorphisms. The dense coverage allowed us to apply several computational procedures to study population structure and the evolutionary relationships among the populations, and to detect signs of adaptive evolution across the genome. While there is considerable gene flow among the sampled populations, there are clear distinctions between populations from the northern desert region and those from the temperate, savannah region. We identified several genes showing population genetic patterns consistent with positive selection within African bee populations, and between these populations and European A. mellifera or Asian Apis florea. CONCLUSIONS: These results lay the groundwork for future studies of adaptive ecological evolution in honey bees, and demonstrate the use of new, freely available web-based tools and workflows ( http://usegalaxy.org/r/kenyanbee ) that can be applied to any model system with genomic information.

The formulation makes the honey bee poison.

Dr. Fumio Matsumura's legacy embraced a passion for exploring environmental impacts of agrochemicals on non-target species such as bees. Why most formulations are more toxic to bees than respective active ingredients and how pesticides interact to cause pollinator decline cannot be answered without understanding the prevailing environmental chemical background to which bees are exposed. Modern pesticide formulations and seed treatments, particularly when multiple active ingredients are blended, require proprietary adjuvants and inert ingredients to achieve high efficacy for targeted pests. Although we have found over 130 different pesticides and metabolites in beehive samples, no individual pesticide or amount correlates with recent bee declines. Recently we have shown that honey bees are sensitive to organosilicone surfactants, nonylphenol polyethoxylates and the solvent N-methyl-2-pyrrolidone (NMP), widespread co-formulants used in agrochemicals and frequent pollutants within the beehive. Effects include learning impairment for adult bees and chronic toxicity in larval feeding bioassays. Multi-billion pounds of formulation ingredients like NMP are used and released into US environments. These synthetic organic chemicals are generally recognized as safe, have no mandated tolerances, and residues remain largely unmonitored. In contrast to finding about 70% of the pesticide active ingredients searched for in our pesticide analysis of beehive samples, we have found 100% of the other formulation ingredients targeted for analysis. These 'inerts' overwhelm the chemical burden from active pesticide, drug and personal care ingredients with which they are formulated. Honey bees serve as an optimal terrestrial bioindicator to determine if 'the formulation and not just the dose makes the poison'.

Overwintering honey bees: biology and management.

In temperate climates, honey bees (Apis mellifera) survive the winter by entering a distinct physiological and behavioral state. In recent years, beekeepers are reporting unsustainably high colony losses during the winter, which have been linked to parasitization by Varroa mites, virus infections, geographic location, and variation across honey bee genotypes. Here, we review literature on environmental, physiological, and social factors regulating entrance, maintenance, and exit from the overwintering state in honey bees in temperate regions and develop a testable model to explain how multiple factors may be acting synergistically to regulate this complex transition. We also review existing knowledge of the factors affecting overwintering survival in honey bees and providing suggestions to beekeepers aiming to improve their colonies' overwintering success.

Evaluation of the distribution and impacts of parasites, pathogens, and pesticides on honey bee (Apis mellifera) populations in East Africa.

In East Africa, honey bees (Apis mellifera) provide critical pollination services and income for small-holder farmers and rural families. While honey bee populations in North America and Europe are in decline, little is known about the status of honey bee populations in Africa. We initiated a nationwide survey encompassing 24 locations across Kenya in 2010 to evaluate the numbers and sizes of honey bee colonies, assess the presence of parasites (Varroa mites and Nosema microsporidia) and viruses, identify and quantify pesticide contaminants in hives, and assay for levels of hygienic behavior. Varroa mites were present throughout Kenya, except in the remote north. Levels of Varroa were positively correlated with elevation, suggesting that environmental factors may play a role in honey bee host-parasite interactions. Levels of Varroa were negatively correlated with levels of hygienic behavior: however, while Varroa infestation dramatically reduces honey bee colony survival in the US and Europe, in Kenya Varroa presence alone does not appear to impact colony size. Nosema apis was found at three sites along the coast and one interior site. Only a small number of pesticides at low concentrations were found. Of the seven common US/European honey bee viruses, only three were identified but, like Varroa, were absent from northern Kenya. The number of viruses present was positively correlated with Varroa levels, but was not correlated with colony size or hygienic behavior. Our results suggest that Varroa, the three viruses, and Nosema have been relatively recently introduced into Kenya, but these factors do not yet appear to be impacting Kenyan bee populations. Thus chemical control for Varroa and Nosema are not necessary for Kenyan bees at this time. This study provides baseline data for future analyses of the possible mechanisms underlying resistance to and the long-term impacts of these factors on African bee populations.

Weighing risk factors associated with bee colony collapse disorder by classification and regression tree analysis.

Colony collapse disorder (CCD), a syndrome whose defining trait is the rapid loss of adult worker honey bees, Apis mellifera L., is thought to be responsible for a minority of the large overwintering losses experienced by U.S. beekeepers since the winter 2006-2007. Using the same data set developed to perform a monofactorial analysis (PloS ONE 4: e6481, 2009), we conducted a classification and regression tree (CART) analysis in an attempt to better understand the relative importance and interrelations among different risk variables in explaining CCD. Fifty-five exploratory variables were used to construct two CART models: one model with and one model without a cost of misclassifying a CCD-diagnosed colony as a non-CCD colony. The resulting model tree that permitted for misclassification had a sensitivity and specificity of 85 and 74%, respectively. Although factors measuring colony stress (e.g., adult bee physiological measures, such as fluctuating asymmetry or mass of head) were important discriminating values, six of the 19 variables having the greatest discriminatory value were pesticide levels in different hive matrices. Notably, coumaphos levels in brood (a miticide commonly used by beekeepers) had the highest discriminatory value and were highest in control (healthy) colonies. Our CART analysis provides evidence that CCD is probably the result of several factors acting in concert, making afflicted colonies more susceptible to disease. This analysis highlights several areas that warrant further attention, including the effect of sublethal pesticide exposure on pathogen prevalence and the role of variability in bee tolerance to pesticides on colony survivorship.

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.

Colony collapse disorder: a descriptive study.

BACKGROUND: Over the last two winters, there have been large-scale, unexplained losses of managed honey bee (Apis mellifera L.) colonies in the United States. In the absence of a known cause, this syndrome was named Colony Collapse Disorder (CCD) because the main trait was a rapid loss of adult worker bees. We initiated a descriptive epizootiological study in order to better characterize CCD and compare risk factor exposure between populations afflicted by and not afflicted by CCD. METHODS AND PRINCIPAL FINDINGS: Of 61 quantified variables (including adult bee physiology, pathogen loads, and pesticide levels), no single measure emerged as a most-likely cause of CCD. Bees in CCD colonies had higher pathogen loads and were co-infected with a greater number of pathogens than control populations, suggesting either an increased exposure to pathogens or a reduced resistance of bees toward pathogens. Levels of the synthetic acaricide coumaphos (used by beekeepers to control the parasitic mite Varroa destructor) were higher in control colonies than CCD-affected colonies. CONCLUSIONS/SIGNIFICANCE: This is the first comprehensive survey of CCD-affected bee populations that suggests CCD involves an interaction between pathogens and other stress factors. We present evidence that this condition is contagious or the result of exposure to a common risk factor. Potentially important areas for future hypothesis-driven research, including the possible legacy effect of mite parasitism and the role of honey bee resistance to pesticides, are highlighted.

"Entombed Pollen": A new condition in honey bee colonies associated with increased risk of colony mortality.

Here we describe a new phenomenon, entombed pollen, which is highly associated with increased colony mortality. Entombed pollen is sunken, capped cells amidst "normal", uncapped cells of stored pollen, and some of the pollen contained within these cells is brick red in color. There appears to be a lack of microbial agents in the pollen, and larvae and adult bees do not have an increased rate of mortality when they are fed diets supplemented with entombed pollen in vitro, suggesting that the pollen itself is not directly responsible for increased colony mortality. However, the increased incidence of entombed pollen in reused wax comb suggests that there is a transmittable factor common to the phenomenon and colony mortality. In addition, there were elevated pesticide levels, notably of the fungicide chlorothalonil, in entombed pollen. Additional studies are needed to determine if there is a causal relationship between entombed pollen, chemical residues, and colony mortality.