Bee Pathogens Research Articles

Honey as a Source of Environmental DNA for the Detection and Monitoring of Honey Bee Pathogens and Parasites.

Environmental DNA (eDNA) has been proposed as a powerful tool to detect and monitor cryptic, elusive, or invasive organisms. We recently demonstrated that honey constitutes an easily accessible source of eDNA. In this study, we extracted DNA from 102 honey samples (74 from Italy and 28 from 17 other countries of all continents) and tested the presence of DNA of nine honey bee pathogens and parasites (Paenibacillus larvae, Melissococcus plutonius, Nosema apis, Nosema ceranae, Ascosphaera apis, Lotmaria passim, Acarapis woodi, Varroa destructor, and Tropilaelaps spp.) using qualitative PCR assays. All honey samples contained DNA from V. destructor, confirming the widespread diffusion of this mite. None of the samples gave positive amplifications for N. apis, A. woodi, and Tropilaelaps spp. M. plutonius was detected in 87% of the samples, whereas the other pathogens were detected in 43% to 57% of all samples. The frequency of Italian samples positive for P. larvae was significantly lower (49%) than in all other countries (79%). The co-occurrence of positive samples for L. passim and A. apis with N. ceranae was significant. This study demonstrated that honey eDNA can be useful to establish monitoring tools to evaluate the sanitary status of honey bee populations.

Propolis Envelope Promotes Beneficial Bacteria in the Honey Bee (Apis mellifera) Mouthpart Microbiome.

Honey bees collect and apply plant resins to the interior of their nest cavity, in order to form a layer around the nest cavity called a propolis envelope. Propolis displays antimicrobial activity against honey bee pathogens, but the effect of propolis on the honey bee microbiome is unknown. Honey bees do not intentionally consume propolis, but they do manipulate propolis with their mouthparts. Because honey bee mouthparts are used for collecting and storing nectar and pollen, grooming and trophallaxis between adults, feeding larvae, and cleaning the colony, they are an important interface between the bees’ external and internal environments and serve as a transmission route for core gut bacteria and pathogens alike. We hypothesized that the antimicrobial activity of an experimentally applied propolis envelope would influence the bacterial diversity and abundance of the worker mouthpart microbiome. The results revealed that the mouthparts of worker bees in colonies with a propolis envelope exhibited a significantly lower bacterial diversity and significantly higher bacterial abundance compared to the mouthparts of bees in colonies without a propolis envelope. Based on the taxonomic results, the propolis envelope appeared to reduce pathogenic or opportunistic microbes and promote the proliferation of putatively beneficial microbes on the honey bee mouthparts, thus reinforcing the core microbiome of the mouthpart niche.

The Pathogen Profile of a Honey Bee Queen Does Not Reflect That of Her Workers.

Throughout a honey bee queen’s lifetime, she is tended to by her worker daughters, who feed and groom her. Such interactions provide possible horizontal transmission routes for pathogens from the workers to the queen, and as such a queen’s pathogen profile may be representative of the workers within a colony. To explore this further, we investigated known honey bee pathogen co-occurrence, as well as pathogen transmission from workers to queens. Queens from 42 colonies were removed from their source hives and exchanged into a second, unrelated foster colony. Worker samples were taken from the source colony on the day of queen exchange and the queens were collected 24 days after introduction. All samples were screened for Nosema spp., Trypanosome spp., acute bee paralysis virus (ABPV), black queen cell virus (BQCV), chronic bee paralysis virus (CBPV), Israeli acute paralysis virus (IAPV), Lake Sinai virus (LSV), and deformed wing virus master variants (DWV-A, B, and C) using RT-qPCR. The data show that LSV, Nosema, and DWV-B were the most abundant pathogens in colonies. All workers (n = 42) were LSV-positive, 88% were Nosema-positive, whilst pathogen loads were low (<1 × 106 genome equivalents per pooled worker sample). All queens (n = 39) were negative for both LSV and Nosema. We found no evidence of DWV transmission occurring from worker to queen when comparing queens to foster colonies, despite DWV being present in both queens and workers. Honey bee pathogen presence and diversity in queens cannot be revealed from screening workers, nor were pathogens successfully transmitted to the queen.

Shotgun sequencing of honey DNA can describe honey bee derived environmental signatures and the honey bee hologenome complexity

Honey bees are large-scale monitoring tools due to their extensive environmental exploration. In their activities and from the hive ecosystem complex, they get in close contact with many organisms whose traces can be transferred into the honey, which can represent an interesting reservoir of environmental DNA (eDNA) signatures and information useful to analyse the honey bee hologenome complexity. In this study, we tested a deep shotgun sequencing approach of honey DNA coupled with a specifically adapted bioinformatic pipeline. This methodology was applied to a few honey samples pointing out DNA sequences from 191 organisms spanning different kingdoms or phyla (viruses, bacteria, plants, fungi, protozoans, arthropods, mammals). Bacteria included the largest number of species. These multi-kingdom signatures listed common hive and honey bee gut microorganisms, honey bee pathogens, parasites and pests, which resembled a complex interplay that might provide a general picture of the honey bee pathosphere. Based on the Apis mellifera filamentous virus genome diversity (the most abundant detected DNA source) we obtained information that could define the origin of the honey at the apiary level. Mining Apis mellifera sequences made it possible to identify the honey bee subspecies both at the mitochondrial and nuclear genome levels.

Sudden and Prevalent Deaths of Foraging Honey Bees in Early Spring During Sowing of Clothianidin Coated Maize Seeds Between 2013 and 2018 in Turkey

Abstract Between 2013 and 2018, there was an outbreak of sudden foraging honey bee deaths in Turkey’s Çukurova Region from the beginning of February to the middle of March, a period of time when the sowing of maize seeds occurs in the region. Upon returning, the affected bees were unable to enter the hive because of unbalanced flying and walking. Deaths were seen at apiaries located on plains or near plains where maize seeds were sowed. When winds blew from the direction of the plains towards the hives, honey bee deaths increased, but when the weather was rainy, honey bee deaths decreased or stopped. Honey bee losses were mainly between 30% and 80%. Investigations related to the honey bee deaths were carried out in the affected apiaries using on-site inspection and laboratory tests. The main reason of deaths would be linked to the clothianidin coated maize seeds used in the plain during sowing period. No remarkable correlation with any bee pathogens was detected related to the sudden and prevalent honey bee deaths.

Interactions Between Thiamethoxam and Deformed Wing Virus Can Drastically Impair Flight Behavior of Honey Bees.

Exposure to multiple stress factors is believed to contribute to honey bee colony decline. However, little is known about how co-exposure to stress factors can alter the survival and behavior of free-living honey bees in colony conditions. We therefore studied the potential interaction between a neonicotinoid pesticide, thiamethoxam, and a highly prevalent honey bee pathogen, Deformed wing virus (DWV). For this purpose, tagged bees were exposed to DWV by feeding or injection, and/or to field-relevant doses of thiamethoxam, then left in colonies equipped with optical bee counters to monitor flight activity. DWV loads and the expression of immune genes were quantified. A reduction in vitellogenin expression level was observed in DWV-injected bees and was associated with precocious onset of foraging. Combined exposure to DWV and thiamethoxam did not result in higher DWV loads compared to bees only exposed to DWV, but induced precocious foraging, increased the risk of not returning to the hive after the first flight, and decreased survival when compared to single stress exposures. We therefore provided the first evidence for deleterious interactions between DWV and thiamethoxam in natural conditions.

Honey bee virus causes context-dependent changes in host social behavior

Anthropogenic changes create evolutionarily novel environments that present opportunities for emerging diseases, potentially changing the balance between host and pathogen. Honey bees provide essential pollination services, but intensification and globalization of honey bee management has coincided with increased pathogen pressure, primarily due to a parasitic mite/virus complex. Here, we investigated how honey bee individual and group phenotypes are altered by a virus of concern, Israeli acute paralysis virus (IAPV). Using automated and manual behavioral monitoring of IAPV-inoculated individuals, we find evidence for pathogen manipulation of worker behavior by IAPV, and reveal that this effect depends on social context; that is, within versus between colony interactions. Experimental inoculation reduced social contacts between honey bee colony members, suggesting an adaptive host social immune response to diminish transmission. Parallel analyses with double-stranded RNA (dsRNA)-immunostimulated bees revealed these behaviors are part of a generalized social immune defensive response. Conversely, inoculated bees presented to groups of bees from other colonies experienced reduced aggression compared with dsRNA-immunostimulated bees, facilitating entry into susceptible colonies. This reduction was associated with a shift in cuticular hydrocarbons, the chemical signatures used by bees to discriminate colony members from intruders. These responses were specific to IAPV infection, suggestive of pathogen manipulation of the host. Emerging bee pathogens may thus shape host phenotypes to increase transmission, a strategy especially well-suited to the unnaturally high colony densities of modern apiculture. These findings demonstrate how anthropogenic changes could affect arms races between human-managed hosts and their pathogens to potentially affect global food security.

Competition and Identification of Anaerobic Organisms Isolated from Apis mellifera Gut Microbiota

Apis mellifera, the most common species of honeybee, has been suffering the epidemic of colony collapse disorder (CCD). This is defined by the sudden and unexplainable death of an entire colony of bees, and populations worldwide dwindle further every year. Although research continues, there exists no lasting or effective treatment. One underdeveloped area of study has been the option to treat bees with probiotics ( Audisio, 2017; Baffoni et al., 2015). There have been numerous studies within other organisms and honeybees that show that a well‐maintained gut microbiome is a key component of health, especially when the pathogen is endemic to the intestines – as in the case of Nosema pathogen, which has been implicated in CCD ( Raymann, 2018; Shreiner, 2015). The hypothesis being investigated is that microbes cultivated from the gut of normal bees will suppress the growth of known honey bee pathogens. Identification of such normal flora may serve as a probiotic for treatment of bees experiencing CCD due to infectious agents.Support or Funding InformationResearch Grant from the University of Wisconsin ‐ Stout

Occurrence of honey bee-associated pathogens in Varroa-free pollinator communities

Australia remains the last significant land mass free of Varroa, a parasitic mite which has caused dramatic honey bee (Apis mellifera) colony losses across the globe, due to its association with the pathogenic deformed wing virus (DWV). As such, Australia continues to maintain relatively healthy honey bee populations, despite recent work showing apiaries harbor a surprisingly high prevalence of microbial pathogens. We sought to determine the prevalence of these microbial pathogens in honey bees and native pollinators actively co-foraging on mass flowering crops and to understand the extent to which they may be shared between taxa. We found high prevalences of black queen cell virus (BQCV) and sacbrood virus (SBV) in the honey bees (88% and 41% respectively), and correspondingly, these were the most common honey bee pathogens detected in native pollinator taxa, albeit at much lower prevalence; the maximum prevalence for any pathogen in a native pollinator group was 24% (BQCV in Halictidae spp.). The viral pathogens Israeli acute paralysis virus and Lake Sinai viruses 1 and 2, and the fungal parasites Nosema apis and Nosema ceranae, were only rarely detected. Phylogenetic analyses of the most common pathogens revealed similar genotypes circulating between species. Our data suggest that, in Australian orchards, pathogen prevalence in honey bees is a good predictor of pathogen prevalence in native pollinators, which raises concerns about how the viral landscape may change in native taxa if, or when, Varroa arrives.

Engineered symbionts activate honey bee immunity and limit pathogens.

Honey bees are essential pollinators threatened by colony losses linked to the spread of parasites and pathogens. Here, we report a new approach for manipulating bee gene expression and protecting bee health. We engineered a symbiotic bee gut bacterium, Snodgrassella alvi, to induce eukaryotic RNA interference (RNAi) immune responses. We show that engineered S. alvi can stably recolonize bees and produce double-stranded RNA to activate RNAi and repress host gene expression, thereby altering bee physiology, behavior, and growth. We used this approach to improve bee survival after a viral challenge, and we show that engineered S. alvi can kill parasitic Varroa mites by triggering the mite RNAi response. This symbiont-mediated RNAi approach is a tool for studying bee functional genomics and potentially for safeguarding bee health.

Inducing immune bee genes

Bee Health Honey bees are prone to parasitism by the Varroa mite, which is a vector for several bee pathogens. However, honey bees are also host to the symbiotic gut bacterium Snodgrassella alvi. Leonard et al. engineered S. alvi to produce double-stranded RNA (dsRNA)—a stimulus for insect RNA interference defense responses—from a plasmid containing two inverted promoters tagged with a fluorescent label (see the Perspective by Paxton). This dsRNA module can be targeted to interfere with specific bee genes as well as crucial viral and mite genes. The authors found that gene expression could be blocked for at least 15 days as the symbionts established in the bees' guts and continuously expressed the dsRNA constructs. S. alvi with specifically targeted plasmids not only suppressed infection with deformed wing virus but also effectively reduced Varroa mite survival. Science , this issue p. [573][1]; see also p. [504][2] [1]: /lookup/doi/10.1126/science.aax9039 [2]: /lookup/doi/10.1126/science.aba6135

Characterization of C3larvinA, a novel RhoA-targeting ADP-ribosyltransferase toxin produced by the honey bee pathogen, Paenibacillus larvae.

C3larvinA is a putative virulence factor produced by Paenibacillus larvae enterobacterial-repetitive-intergenic-consensus (ERIC) III/IV (strain 11-8051). Biochemical, functional and structural analyses of C3larvinA revealed that it belongs to the C3-like mono-ADP-ribosylating toxin subgroup. Mammalian RhoA was the target substrate for its transferase activity suggesting that it may be the biological target of C3larvinA. The kinetic parameters of the NAD+ substrate for the transferase (KM = 75 ± 10 µM) and glycohydrolase (GH) (KM = 107 ± 20 µM) reactions were typical for a C3-like bacterial toxin, including the Plx2A virulence factor from Paenibacillus larvae ERIC I. Upon cytoplasmic expression in yeast, C3larvinA caused a growth-defective phenotype indicating that it is an active C3-like toxin and is cytotoxic to eukaryotic cells. The catalytic variant of the Q187-X-E189 motif in C3larvinA showed no cytotoxicity toward yeast confirming that the cytotoxicity of this factor depends on its enzymatic activity. A homology consensus model of C3larvinA with NAD+ substrate was built on the structure of Plx2A, provided additional confirmation that C3larvinA is a member of the C3-like mono-ADP-ribosylating toxin subgroup. A homology model of C3larvinA with NADH and RhoA was built on the structure of the C3cer-NADH-RhoA complex which provided further evidence that C3larvinA is a C3-like toxin that shares an identical catalytic mechanism with C3cer from Bacillus cereus. C3larvinA induced actin cytoskeleton reorganization in murine macrophages, whereas in insect cells, vacuolization and bi-nucleated cells were observed. These cellular effects are consistent with C3larvinA disrupting RhoA function by covalent modification that is shared among C3-like bacterial toxins.

Age and Method of Inoculation Influence the Infection of Worker Honey Bees (Apis mellifera) by Nosema ceranae.

The microsporidian parasite Nosema ceranae is a highly prevalent, global honey bee pathogen. Apis mellifera is considered to be a relatively recent host for this microsporidia, which raises questions as to how it affects its host’s physiology, behavior and longevity, both at the individual and colony level. As such, honey bees were inoculated with fresh purified spores of this pathogen, both individually (Group A) or collectively (Group B) and they were studied from 0 to 15 days post-emergence (p.e.) to evaluate the effect of bee age and the method of inoculation at 7 days post-infection. The level of infection was analyzed individually by qPCR by measuring the relative amount of the N. ceranae polar tubule protein 3 (PTP3) gene. The results show that the bee’s age and the method of infection directly influence parasite load, and thus, early disease development. Significant differences were found regarding bee age at the time of infection, whereby the youngest bees (new-born and 1 day p.e.) developed the highest parasite load, with this load decreasing dramatically in bees infected at 2 days p.e. before increasing again in bees infected at 3–4 days p.e. The parasite load in bees infected when older than 4 days p.e. diminished as they aged. When the age cohort data was pooled and grouped according to the method of infection, a significantly higher mean concentration and lower variation in N. ceranae infection was evident in Group A, indicating greater variation in experimental infection when spores were administered collectively to bees through their food. In summary, these data indicate that both biological and experimental factors should be taken into consideration when comparing data published in the literature.

Exposure of Larvae of the Solitary Bee Osmia bicornis to the Honey Bee Pathogen Nosema ceranae Affects Life History.

Wild bees are important pollinators of wild plants and agricultural crops and they are threatened by several environmental stressors including emerging pathogens. Honey bees have been suggested as a potential source of pathogen spillover. One prevalent pathogen that has recently emerged as a honey bee disease is the microsporidian Nosema ceranae. While the impacts of N. ceranae in honey bees are well documented, virtually nothing is known about its effects in solitary wild bees. The solitary mason bee Osmia bicornis is a common pollinator in orchards and amenable to commercial management. Here, we experimentally exposed larvae of O. bicornis to food contaminated with N. ceranae and document spore presence during larval development. We measured mortality, growth parameters, and timing of pupation in a semi-field experiment. Hatched individuals were assessed for physiological state including fat body mass, wing muscle mass, and body size. We recorded higher mortality in the viable-spore-exposed group but could only detect a low number of spores among the individuals of this treatment. Viable-spore-treated individuals with higher head capsule width had a delayed pupation start. No impact on the physiological status could be detected in hatched imagines. Although we did not find overt evidence of O. bicornis infection, our findings indicate that exposure of larvae to viable N. ceranae spores could affect bee development.

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 outcomes 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 previous reports on the reduction of DWV diversity following the arrival of the parasitic mite Varroa 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 quasispecies. 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 complementation 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 genotypes were adapted to coexist. Mutational and recombination events were observed across clone progeny, providing new insights into the forces that drive and constrain virus diversification. Accordingly, our results suggest that Varroa influences DWV dynamics by causing an initial selective sweep, which is followed by virus diversification fueled by negative frequency-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.

Wild bumble bees (Hymenoptera: Apidae: Bombini) as a potential reservoir for bee pathogens in northeastern Argentina

The genus Bombus is represented by 38 subgenera around the world with nearly 250 described species. They are among the most efficient insect pollinators and therefore important to the ecology and the economy. In Argentina, this genus it is represented by eight native and two introduced species. Several bee pathogens related to colony losses have been found in wild pollinators around the world, including bumble bees. We studied the presence of these pathogens in bumble bee species from different locations in northeastern Argentina to determine their relationship with prevalent bee pathogens previously detected in the country. We collected 93 specimens of three bumble bee species and screened them for eleven honey bee pathogens. We detected Nosema ceranae, Ascosphaera apis, Melisococcus plutonius, and four viruses. Sixteen samples were pathogen-free and 77 samples contained one or more pathogens. If bumble bees are a potential reservoir for bee pathogens, this could lead to the development of Emerging Infection Diseases in wild bees. However, further studies are required to confirm this assumption and to determine the direction of the spillover between wild and managed bees.

Deformed wing virus type A, a major honey bee pathogen, is vectored by the mite Varroa destructor in a non-propagative manner

Honey bees, the primary managed insect pollinator, suffer considerable losses due to Deformed wing virus (DWV), an RNA virus vectored by the mite Varroa destructor. Mite vectoring has resulted in the emergence of virulent DWV variants. The basis for such changes in DWV is poorly understood. Most importantly, it remains unclear whether replication of DWV occurs in the mite. In this study, we exposed Varroa mites to DWV type A via feeding on artificially infected honey bees. A significant, 357-fold increase in DWV load was observed in these mites after 2 days. However, after 8 additional days of passage on honey bee pupae with low viral loads, the DWV load dropped by 29-fold. This decrease significantly reduced the mites’ ability to transmit DWV to honey bees. Notably, negative-strand DWV RNA, which could indicate viral replication, was detected only in mites collected from pupae with high DWV levels but not in the passaged mites. We also found that Varroa mites contain honey bee mRNAs, consistent with the acquisition of honey bee cells which would additionally contain DWV replication complexes with negative-strand DWV RNA. We propose that transmission of DWV type A by Varroa mites occurs in a non-propagative manner.

Prevalence and population genetics of the emerging honey bee pathogen DWV in Chinese apiculture

Honey bees are agriculturally important, both as pollinators and by providing products such as honey. The sustainability of beekeeping is at risk through factors of global change such as habitat loss, as well as through the spread of infectious diseases. In China and other parts of Asia, beekeepers rely both on native Apis cerana and non-native Apis mellifera, putting bee populations at particular risk of disease emergence from multi-host pathogens. Indeed, two important honey bee parasites have emerged from East Asian honey bees, the mite Varroa destructor and the microsporidian Nosema ceranae. As V. destructor vectors viral bee diseases, we investigated whether another key bee pathogen, Deformed Wing Virus (DWV), may also have originated in East Asian honey bee populations. We use a large-scale survey of apiaries across China to investigate the prevalence and seasonality of DWV in managed A. mellifera and A. cerana colonies, showing that DWV-A prevalence was higher in A. mellifera, with a seasonal spike in prevalence in autumn and winter. Using phylogenetic and population genetic approaches, we show that while China and East Asian DWV isolates show comparatively high levels of genetic diversity, these bee populations are not a source for the current global DWV epidemic.

Seasonal dynamics and co-occurrence patterns of honeybee pathogens revealed by high-throughput RT-qPCR analysis.

The health of the honey bee Apis mellifera is challenged by introduced parasites that interact with its inherent pathogens and cause elevated rates of colony losses. To elucidate co‐occurrence, population dynamics, and synergistic interactions of honey bee pathogens, we established an array of diagnostic assays for a high‐throughput qPCR platform. Assuming that interaction of pathogens requires co‐occurrence within the same individual, single worker bees were analyzed instead of collective samples. Eleven viruses, four parasites, and three pathogenic bacteria were quantified in more than one thousand single bees sampled from sixteen disease‐free apiaries in Southwest Germany. The most abundant viruses were black queen cell virus (84%), Lake Sinai virus 1 (42%), and deformed wing virus B (35%). Forager bees from asymptomatic colonies were infected with two different viruses in average, and simultaneous infection with four to six viruses was common (14%). Also, the intestinal parasites Nosema ceranae (96%) and Crithidia mellificae/Lotmaria passim (52%) occurred very frequently. These results indicate that low‐level infections in honey bees are more common than previously assumed. All viruses showed seasonal variation, while N. ceranae did not. The foulbrood bacteria Paenibacillus larvae and Melissococcus plutonius were regionally distributed. Spearman's correlations and multiple regression analysis indicated possible synergistic interactions between the common pathogens, particularly for black queen cell virus. Beyond its suitability for further studies on honeybees, this targeted approach may be, due to its precision, capacity, and flexibility, a viable alternative to more expensive, sequencing‐based approaches in nonmodel systems.

Impact of Biotic and Abiotic Stressors on Managed and Feral Bees.

Large-scale declines in bee abundance and species richness over the last decade have sounded an alarm, given the crucial pollination services that bees provide. Population dips have specifically been noted for both managed and feral bee species. The simultaneous increased cultivation of bee-dependent agricultural crops has given rise to additional concern. As a result, there has been a surge in scientific research investigating the potential stressors impacting bees. A group of environmental and anthropogenic stressors negatively impacting bees has been isolated. Habitat destruction has diminished the availability of bee floral resources and nest habitats, while massive monoculture plantings have limited bee access to a variety of pollens and nectars. The rapid spread and increased resistance buildup of various bee parasites, pathogens, and pests to current control methods are implicated in deteriorating bee health. Similarly, many pesticides that are widely applied on agricultural crops and within beehives are toxic to bees. The global distribution of honey bee colonies (including queens with attendant bees) and bumble bee colonies from crop to crop for pollination events has been linked with increased pathogen stress and increased competition with native bee species for limited resources. Climatic alterations have disrupted synchronous bee emergence with flower blooming and reduced the availability of diverse floral resources, leading to bee physiological adaptations. Interactions amongst multiple stressors have created colossal maladies hitting bees at one time, and in some cases delivering additive impacts. Initiatives including the development of wild flower plantings and assessment of pesticide toxicity to bees have been undertaken in efforts to ameliorate current bee declines. In this review, recent findings regarding the impact of these stressors on bees and strategies for mitigating them are discussed.