Social disruption: Sublethal pesticides in pollen lead to Apis mellifera queen events and brood loss

Eusocial Apis mellifera colonies depend on queen longevity and brood viability to survive, as the queen is the sole reproductive individual and the maturing brood replenishes the shorter-lived worker bees. Production of many crops rely on both pesticides and bee pollination to improve crop quantity and quality, yet sublethal impacts of this pesticide exposure is often poorly understood. We investigated the resiliency of queens and their brood after one month of sublethal exposure to field relevant doses of pesticides that mimic exposure during commercial pollination contracts. We exposed full size colonies to pollen contaminated with field-relevant doses of the fungicides (chlorothalonil and propicanizole), insecticides (chlorypyrifos and fenpropathrin) or both, noting a significant reduction in pollen consumption in colonies exposed to fungicides compared to control. While we found no difference in the total amount of pollen collected per colony, a higher proportion of pollen to nonpollen foragers was detected in all pesticide exposed colonies. After ceasing treatments, we measured brood development, discovering a significant increase in brood loss and/or cannibalism across all pesticide exposed groups. Sublethal pesticide exposure in general was linked to reduced production of replacement workers and a change in protein acquisition (pollen vs. non-pollen foraging). Fungicide exposure also resulted in increased loss of the reproductive queen

Prioritizing changes in management practices associated with reduced winter honey bee colony losses for US beekeepers.

peer reviewedBeekeepers attempt to manage their honey bee colonies in ways that optimize colony health. Disentangling the impact of management from other variables affecting colony health is complicated by the diversity of practices used and difficulties handling typically complex and incomplete observational datasets. We propose a method to 1) compress multi-factored management data into a single index, to holistically investigate the real world impact of management on colony mortality, and 2) simplify said index to identify the core practices for which a change in behavior is associated with the greatest improvement in survivorship. Experts scored the practices of US beekeepers (n = 18,971) documented using four years of retrospective surveys (2012-2015). Management Index scores significantly correlated with loss rates, with beekeepers most in line with recommendations suffering lower losses. The highest ranked practices varied by operation type, as recommendations accounted for the current prevalence of practices. These results validate experts' opinion using empirical data, and can help prioritize extension messages. Improving management will not prevent all losses; however, we show that few behavioral changes (in particular related to comb management, sources of new colonies and Varroa management) can lead to a non-negligible reduction in risk

Pesticide exposure in honey bees results in increased levels of the gut pathogen Nosema

Global pollinator declines have been attributed to habitat destruction, pesticide use, and climate change or some combination of these factors, and managed honey bees, Apis mellifera, are part of worldwide pollinator declines. Here we exposed honey bee colonies during three brood generations to sub-lethal doses of a widely used pesticide, imidacloprid, and then subsequently challenged newly emerged bees with the gut parasite, Nosema spp. The pesticide dosages used were below levels demonstrated to cause effects on longevity or foraging in adult honey bees. Nosema infections increased significantly in the bees from pesticide-treated hives when compared to bees from control hives demonstrating an indirect effect of pesticides on pathogen growth in honey bees. We clearly demonstrate an increase in pathogen growth within individual bees reared in colonies exposed to one of the most widely used pesticides worldwide, imidacloprid, at below levels considered harmful to bees. The finding that individual bees with undetectable levels of the target pesticide, after being reared in a sub-lethal pesticide environment within the colony, had higher Nosema is significant. Interactions between pesticides and pathogens could be a major contributor to increased mortality of honey bee colonies, including colony collapse disorder, and other pollinator declines worldwide

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

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

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

A Survey of Honey Bee Colony Losses in the U.S., Fall 2007 to Spring 2008

Honey bees are an essential component of modern agriculture. A recently recognized ailment, Colony Collapse Disorder (CCD), devastates colonies, leaving hives with a complete lack of bees, dead or alive. Up to now, estimates of honey bee population decline have not included losses occurring during the wintering period, thus underestimating actual colony mortality. Our survey quantifies the extent of colony losses in the United States over the winter of 2007–2008.Surveys were conducted to quantify and identify management factors (e.g. operation size, hive migration) that contribute to high colony losses in general and CCD symptoms in particular. Over 19% of the country's estimated 2.44 million colonies were surveyed. A total loss of 35.8% of colonies was recorded; an increase of 11.4% compared to last year. Operations that pollinated almonds lost, on average, the same number of colonies as those that did not. The 37.9% of operations that reported having at least some of their colonies die with a complete lack of bees had a total loss of 40.8% of colonies compared to the 17.1% loss reported by beekeepers without this symptom. Large operations were more likely to have this symptom suggesting that a contagious condition may be a causal factor. Sixty percent of all colonies that were reported dead in this survey died without dead bees, and thus possibly suffered from CCD. In PA, losses varied with region, indicating that ambient temperature over winter may be an important factor.Of utmost importance to understanding the recent losses and CCD is keeping track of losses over time and on a large geographic scale. Given that our surveys are representative of the losses across all beekeeping operations, between 0.75 and 1.00 million honey bee colonies are estimated to have died in the United States over the winter of 2007–2008. This article is an extensive survey of U.S. beekeepers across the continent, serving as a reference for comparison with future losses as well as providing guidance to future hypothesis-driven research on the causes of colony mortality

Dynamic evolution in the key honey bee pathogen deformed wing virus: Novel insights into virulence and competition using reverse genetics

The impacts of invertebrate RNA virus population dynamics on virulence and infection out- comes are poorly understood. Deformed wing virus (DWV), the main viral pathogen of honey bees, negatively impacts bee health, which can lead to colony death. Despite previ- ous reports on the reduction of DWV diversity following the arrival of the parasitic mite Var- roa destructor, the key DWV vector, we found high genetic diversity of DWV in infested United States honey bee colonies. Phylogenetic analysis showed that divergent US DWV genotypes are of monophyletic origin and were likely generated as a result of diversification after a genetic bottleneck. To investigate the population dynamics of this divergent DWV, we designed a series of novel infectious cDNA clones corresponding to coexisting DWV genotypes, thereby devising a reverse-genetics system for an invertebrate RNA virus qua- sispecies. Equal replication rates were observed for all clone-derived DWV variants in single infections. Surprisingly, individual clones replicated to the same high levels as their mixtures and even the parental highly diverse natural DWV population, suggesting that complemen- tation between genotypes was not required to replicate to high levels. Mixed clone–derived infections showed a lack of strong competitive exclusion, suggesting that the DWV geno- types were adapted to coexist. Mutational and recombination events were observed across clone progeny, providing new insights into the forces that drive and constrain virus diversifi- cation. Accordingly, our results suggest that Varroa influences DWV dynamics by causing an initial selective sweep, which is followed by virus diversification fueled by negative fre- quency-dependent selection for new genotypes. We suggest that this selection might reflect the ability of rare lineages to evade host defenses, specifically antiviral RNA interference (RNAi). In support of this hypothesis, we show that RNAi induced against one DWV strain is less effective against an alternate strain from the same population

RNA viruses in hymenopteran pollinators : evidence of inter-taxa virus transmission via pollen and potential impact on non-Apis hymenopteran species

Although overall pollinator populations have declined over the last couple of decades, the honey bee (Apis mellifera) malady, colony collapse disorder (CCD), has caused major concern in the agricultural community. Among honey bee pathogens, RNA viruses are emerging as a serious threat and are suspected as major contributors to CCD. Recent detection of these viral species in bumble bees suggests a possible wider environmental spread of these viruses with potential broader impact. It is therefore vital to study the ecology and epidemiology of these viruses in the hymenopteran pollinator community as a whole. We studied the viral distribution in honey bees, in their pollen loads, and in other non-Apis hymenopteran pollinators collected from flowering plants in Pennsylvania, New York, and Illinois in the United States. Viruses in the samples were detected using reverse transcriptase-PCR and confirmed by sequencing. For the first time, we report the molecular detection of picorna-like RNA viruses (deformed wing virus, sacbrood virus and black queen cell virus) in pollen pellets collected directly from forager bees. Pollen pellets from several uninfected forager bees were detected with virus, indicating that pollen itself may harbor viruses. The viruses in the pollen and honey stored in the hive were demonstrated to be infective, with the queen becoming infected and laying infected eggs after these virus-contaminated foods were given to virus-free colonies. These viruses were detected in eleven other non-Apis hymenopteran species, ranging from many solitary bees to bumble bees and wasps. This finding further expands the viral host range and implies a possible deeper impact on the health of our ecosystem. Phylogenetic analyses support that these viruses are disseminating freely among the pollinators via the flower pollen itself. Notably, in cases where honey bee apiaries affected by CCD harbored honey bees with Israeli Acute Paralysis virus (IAPV), nearby non-Apis hymenopteran pollinators also had IAPV, while those near apiaries without IAPV did not. In containment greenhouse experiments, IAPV moved from infected honey bees to bumble bees and from infected bumble bees to honey bees within a week, demonstrating that the viruses could be transmitted from one species to another. This study adds to our present understanding of virus epidemiology and may help explain bee disease patterns and pollinator population decline in general

Large-scale field application of RNAi technology reducing Israeli acute paralysis virus disease in honey bees (Apis mellifera, Hymenoptera: Apidae).

The importance of honey bees to the world economy far surpasses their contribution in terms of honey production; they are responsible for up to 30% of the world's food production through pollination of crops. Since fall 2006, honey bees in the U.S. have faced a serious population decline, due in part to a phenomenon called Colony Collapse Disorder (CCD), which is a disease syndrome that is likely caused by several factors. Data from an initial study in which investigators compared pathogens in honey bees affected by CCD suggested a putative role for Israeli Acute Paralysis Virus, IAPV. This is a single stranded RNA virus with no DNA stage placed taxonomically within the family Dicistroviridae. Although subsequent studies have failed to find IAPV in all CCD diagnosed colonies, IAPV has been shown to cause honey bee mortality. RNA interference technology (RNAi) has been used successfully to silence endogenous insect (including honey bee) genes both by injection and feeding. Moreover, RNAi was shown to prevent bees from succumbing to infection from IAPV under laboratory conditions. In the current study IAPV specific homologous dsRNA was used in the field, under natural beekeeping conditions in order to prevent mortality and improve the overall health of bees infected with IAPV. This controlled study included a total of 160 honey bee hives in two discrete climates, seasons and geographical locations (Florida and Pennsylvania). To our knowledge, this is the first successful large-scale real world use of RNAi for disease control