Following heightened colony losses, we screened 132 colonies from 23 commercial beekeeping operations in the states of Florida and California. We assessed chemical residues, finding beekeeper-applied miticides, followed by fungicides, were in the highest prevalence and concentration in all colony matrices. Insecticides comprised a minority of detections and concentration. Due to their heightened toxicity, insecticides contributed substantially to hazard quotients with one residue, the neonicotinoid imidacloprid, contributing 99.9% to overall HQ. We found both high prevalence and high levels of imidacloprid in adult bee bodies. Additionally, we preserved a small subsample of dying bees. Our findings highlight a survivorship bias, where dying bees had active ingredients known to have acute toxicity to bees. This result mirrors recent evidence for viral drivers of bee declines that were missed in whole-colony surveys. While bees die from multiple, often interacting, stressors, here we show single contributors at levels capable of causing acute harm.

Commercial beekeepers in the US reported severe colony losses early in 2025, as colonies were being staged for their critical role in the almond pollination season in California. Average reported losses since the preceding spring exceeded 60%, with substantial variation among operations. Many colonies were still actively collapsing in January 2025, at which time pooled and individual samples were collected and then screened for levels of 13 known honey bee pathogens and parasites. Acute bee paralysis virus and other known viral pathogens were found at high levels in pooled bee samples from all collapsing apiaries. Nevertheless, viral loads did not differ between healthy colonies and colonies in active collapse. However, individual bees exhibiting shaking behaviors and morbidity showed distinctly higher loads of two strains of deformed wing virus. Differences between these two analyses suggest that direct collections of morbid bees provide a complementary diagnostic for causal viruses, a suggestion supported by inoculation experiments that successfully replicated observed pathologies. Since these viruses are known to be vectored by the parasitic mite Varroa destructor, mites from collapsed colonies were in turn screened for resistance to amitraz, a critical miticide used widely by beekeepers, including all beekeepers surveyed in this study. A genetic trait linked with miticide resistance was found in all collected mites, underscoring the urgent need for new control strategies for this parasite. While viruses are a likely end-stage cause of colony death, other stressors such as nutritional stress and agrochemicals may have also played significant roles.

DNA methylation is an important epigenomic modification that significantly influences various cellular and organismal functions. In this study, we investigate the methylome of the small hive beetle, Aethina tumida. Our analysis reveals an average of 58,306 CpG methylation marks per beetle, representing approximately 0.99% of the genome's total CpGs. Notably, 85.4% of these methylation marks are located within genic regions on autosomes, with similar rates observed in both male and female beetles. However, male beetles exhibit a lower number of methylation marks and upregulated genes on Chromosome X when compared to female beetles. To evaluate the impact of epialleles on methylation, we identified 5828 associations between SNPs and methylation, with genotypes accounting for 39.2% of the variation observed at highly methylated sites. Interestingly, unfertilised eggs display slightly higher levels of DNA methylation compared to adult beetles, whereas embryos show methylation levels that are only about half of those in adults. This suggests that DNA methylation is dynamic during early development.

Complex host-vector-virus interfaces can impose serious health challenges. Western honeybees have experienced high colony losses globally, mainly driven by the host-shifted, virus-vectoring ectoparasitic mites Varroa destructor and Tropilaelaps mercedesae. Host populations can survive mite infestations through natural selection, offering a long-term strategy for colony health. However, host-vector-virus coevolution requires local adaptations of this triad, which is poorly understood. We propose harnessing natural selection through a global approach focused on standardized monitoring of colony survival, mite infestation levels, and control of reproductives. Studying native and adapted mite hosts, host shifts, and comparing susceptible to surviving hosts will enhance understanding of this host-vector-virus system. This strategy promotes colony health in both managed and wild host populations and provides insights into other host-vector-virus interfaces.

Honey bee fungal diseases, especially Nosema disease caused by the spore-forming intracellular parasite, Nosema, are listed with the Office International des Epizooties (OIE) and impact the health and performance of honey bees in multiple ways and are often associated with colony losses worldwide. The chapter Fungus Diseases presents an overview of causative agents, biology, pathology, and epidemiology of fungal diseases, and discusses the range of intervention options for prevention and treatment of the various fungal diseases in honeybees. The chapter aims to assist veterinarians in identifying diseases caused by fungal pathogens and to provide some basic information on fungal disease prevention and treatment strategies.

Managed honey bee colonies (Apis mellifera) in the US continue to experience high overwinter loss rates driven by parasites, pathogens, poor nutrition, and pesticides. To mitigate these losses, inspection and monitoring are critical for identifying traits of colonies in decline and potential causal factors. In this study, we apply molecular methods to associate potential causative agents with colonies in various stages of decline. Initially, we investigated in-hive bee metagenomic RNA isolated from 15 colonies across seven managed operations in California whose adult bee and brood populations were classified as Strong, Medium, or Weak in strength. We discovered that Weak colonies harbored 2.2- and 3.6- fold more viral species than Medium and Strong colonies, respectively, as well as larger viral read pools despite similar library sizes. They also displayed higher nucleotide variation in Varroa-vectored viruses, indicating associations with high mite populations. When investigating differences in host gene expression, we discovered an upregulation of immune-related pathways in Weak colonies relative to Strong. Specifically, Weak colonies upregulated genes related to wound healing, phagocytosis, oxidative stress resistance, apoptosis, and RNA interference. Most antimicrobial peptides were upregulated in Weak colonies, although defensin1 was significantly higher in Strong colonies, along with several detoxification enzymes and the royal jelly peptide apisimin. Weak colonies also showed an upregulation of transcripts tied to abnormal protein digestion. The low levels of viral replication and fewer species of mite-vectored viruses in Strong colonies may be due to successful Varroa management. Strong colonies also displayed upregulated levels of nine different ubiquinone transcripts, arguably reflecting increasing longevity or a younger in-hive population compared to Weak colonies. Overall, these results provide a detailed account of viral metagenomics and associated host responses, providing new insights into the mechanisms underlying honey bee colony decline under comparable management conditions.

In 2025, a swarm of giant honey bees, Apis dorsata dorsata, was detected on board a cargo vessel prior to arrival at the port of Elizabeth, New Jersey, USA. Apis dorsata is a quarantine species not native to the United States, and further inspection of the bees uncovered 28 mites that were identified as Tropilaelaps mercedesae by morphological and DNA barcoding analysis. Tropilaelaps mercedesae is an ectoparasite of honey bees that is not yet present in the USA. An additional mite species, Kuzinia morsei, was also observed. Molecular screening of common honey bee pathogens in 42 intercepted bees showed a 5% prevalence of trypanosomatid infections and a 2% prevalence of American foulbrood. Additionally, Black queen cell virus (BQCV) and Deformed wing virus B (DWV-B) were detected in 38% and 7% of the samples, respectively. To our knowledge, this is the first detection of T. mercedesae and K. morsei on a vessel bound for the USA, as well as the first interception of a whole swarm of A. dorsata dorsata. Phylogenetic analysis of the CO1 sequence data indicated that the intercepted bee swarm originated from a southern India lineage. Our findings indicate that exotic bee swarms can harbor parasitic mites and multiple bee pathogens, both critical to the apiculture industry. Additionally, our findings reveal that Tropilaelaps mites can survive extended periods associated with adult bees and in the absence of brood, suggesting greater potential for long-distance movement and introduction risk. This study provides new insights into Tropilaelaps dispersal and swarm-mediated pathogen movement, thereby improving our ability to predict establishment and outbreak risk crucial for countering invasive agricultural threats. The findings also emphasize that early detection, through coordinated port and shipboard surveillance, and strong interagency collaboration, is vital for reducing the risk of agricultural pest and pathogen introductions into the USA.

Biological invasions may induce adaptive shifts in traits that increase individual reproductive success, thereby accelerating population growth and amplifying ecological impacts. One such trait is polyandry, i.e. multiple mating by females, which may vary between endemic and invasive populations due to variation in demographic and environmental factors that alter the costs and benefits of multiple mating. Despite polyandry being widespread across taxa, comparative studies among multiple endemic and invasive populations of invasive species remain scarce. Here, we estimated the prevalence of polyandry in two endemic and five invasive populations of small hive beetle (Aethina tumida), a globally invasive parasite affecting bee health. Using DNA microsatellite genotyping of field-sampled females and their offspring to estimate mating frequencies, we show that polyandry varies in SHB populations and females mate with 1-13 males. However, the polyandry levels were not significantly different across the endemic and invasive populations compared. These results suggest that polyandry in SHB may be a stable reproductive trait rather than a plastic response to invasion-related factors. Our work supports further research on how multiple mating may improve both individual reproductive success and population viability in the context of biological invasions.

In January of 2025, U.S. commercial beekeepers reported unusually high honey bee colony losses as they prepared colonies for almond pollination. Two industry groups launched nationwide surveys to document colony losses between June 2024 and March 2025 across all scales of beekeeping. This study analyzes these survey data to assess colony losses, estimate financial impacts, and identify correlations with beekeeper management practices and geographical locations. Unlike past surveys, commercial beekeepers experienced more severe losses than smaller-scale beekeepers during this period. Respondents, managing over half of U.S. colonies, most frequently cited Varroa mites as the cause for their losses. Varroa mites were followed by pesticides and pathogens in the case of commercial beekeepers and by queen failure and weather in the case of smaller-scale beekeepers. Although Varroa was the most frequently cited cause, losses did not significantly differ between users and non-users of amitraz, suggesting that rising amitraz resistance alone does not explain observed trends. Differences in protein and carbohydrate feeding frequencies also played a role in net losses. While colony loss rates and financial concern varied widely among respondents, commercial beekeepers understandably showed higher sensitivity to financial impacts, with concerns increasing linearly with loss severity. This study highlights the value of beekeeper surveys which, alongside direct analyses of bee samples and longitudinal studies, help identify effective management strategies and environmental risks. Such insights are crucial for addressing the leading causes of colony losses on a national scale, and ultimately aid in safeguarding honey bee health, pollination services, and agricultural production.

High infestation levels of small hive beetle (SHB), Aethina tumida, can cause more damage to honeybee, Apis mellifera, host colonies. However, the spatiotemporal variation of SHB infestations is poorly understood. Here, we show that SHB infestations can be equally high in native and invasive ranges, suggesting that differences between host populations are the key criterion for damage. The data reveal that spatial variation within locations was not correlated with migratory beekeeping, SHB management strategies, nor the number of colonies at an apiary. Despite no annual changes in SHB infestations, the data confirm seasonal variation in infestations in two locations probably due to environmental factors affecting SHB. Infestations are lower in Italy than elsewhere, possibly due to strongly implemented management strategies. It is apparent that our understanding of varying SHB infestations is still limited. This suggests that further efforts are required to elucidate our knowledge of this important host-parasite system.

Honey bees create complex societies of self-organized individuals in intricate colonies. Studies of honey bees are carried out in both the field and the laboratory. However, field research is encumbered by the difficulties of making reliable observations and environmental confounders. Meanwhile, laboratory trials produce data that are not field realistic as they lack key characteristics of a natural colony. Additionally, advances in honey bee research have been hindered without reliable methodology to rear queens in the laboratory. Here we provide a new system to reliably produce queens and worker brood in the laboratory and describe how this system fits with artificial insemination of queens as a step towards a continuous self-contained source of bees. The process creates a bridge between field research and laboratory trials and provides a secure system for contagious or regulated elements while maintaining many of the intrinsic characteristics of a honey bee colony.

The passion flower bee, Protandrena (Anthemurgus) passiflorae (Robertson) is a monolectic, host-plant specialist of the passionflower plant Passiflora lutea L. Using a single adult male individual, we generated long-read PacBio HiFi, HiC, and short-read RNA sequencing data to build a well-annotated, chromosome-level genome assembly for this species. The final nuclear genome is 249 Mb with 150x coverage and with most of the genome scaffolding into 12 chromosomes. The scaffold N50 is 21.4 Mb and the genome has a Benchmarking Universal Single-Copy Ortholog (BUSCO) score of 97.2% for 5991 hymenopteran genes. BRAKER3 annotation of the genome identified 12,098 genes and 15,353 total transcripts and found that 20.27% of the genome is made up of repetitive elements. We resolved a mitochondrial genome of 12.7 kb. The P. passiflorae genome represents one of only a few published andrenid bee genomes and one of the first monolectic bees. This new high-quality genome will serve as a valuable resource for investigating the genomic basis of specialization and for providing a useful resource for studying pollinator health and conservation.

Bees are key species for pollination and apiculture. Within the multiple biotic threats, parasites are one of the main players involved in bee health. Among them, trypanosomatid parasites have been the focus of recent studies that have placed them as one of the most prevalent microorganisms in the digestive tract of bees around the world. Here, we review what is known in epidemiology, cell biology and genetics of these parasites in bees with a particular focus on the work made on honeybees. We also discuss the possible implications for honeybee health and describe research gaps to be explored both from the honeybee host and trypanosomatid parasites sides.

We present the first chromosome-level genome assembly for Bombus pensylvanicus, a historically widespread native pollinator species that was distributed across eastern North America but has subsequently undergone declines in range area and local relative abundance. This species has been of significant interest as a model for understanding both patterns and possible causes of bumble bee decline in the region, including the role of genetic variation. Here we present a chromosome-level reference genome assembled using Pacific Biosciences singe-molecule HiFi sequences and Hi-C data and annotated using evidence derived from RNA sequencing of multiple tissue types. The B. pensylvanicus genome has a total length of ∼352.6 Mb and was assembled into a total of 224 scaffolds, with 19 primary pseudomolecules representing putative chromosomes and an N50 = 14.872 Mb. Annotation with the Eukaryotic Genome Annotation Pipeline-External (EGAPx) identified 11,411 genes (10,263 protein coding), and BUSCO analysis of 5,991 Hymenoptera-specific BUSCO groups indicated a completeness for the proteins of 99.0% (98.6% single-copy, 0.5% duplicated) and for the genome of 98.5% (98.2% single-copy, 0.3% duplicated). We present synteny analyses with other recently assembled Bombus genomes representing different subgenera and examine the distribution of repetitive regions of the genome relative to the distribution of genes and noncoding RNAs.

BACKGROUND:The protist family Trypanosomatidae includes parasites of insects, vertebrates, plants, and even other unicellular eukaryotes. The genomes of these species harbor clues to the evolution of parasitism, adaptation to new hosts, and infection of mammals. We present an analysis of a chromosome-level genome assembly of Lotmaria passim, the most prevalent known trypanosomatid of honey bees, linking genome sequence and organization to gene expression and infection of bees. RESULTS:The genome showed a high degree of synteny with assemblies of other trypanosomatids and especially to the closely related Leptomonas pyrrhocoris. It included four copies of chromosomes that shared ancestry with the tetrasomic Leishmania Chromosome 31 and are consistently supernumerary throughout Trypanosomatidae. However, these chromosomes showed lower similarity to L. passim relatives than did the genome overall, with sufficient variation across haplotypes to distinguish two separate disomic chromosomes. Transcriptomic analyses showed that these chromosomes are enriched in genes upregulated during bee infection, and each include five paralogs of the GP63 gene implicated in infection of both insects and mammals. Patterns of expression in bees suggested decreased protein synthesis, a shift from carbohydrate- to amino acid-based metabolism, and reduced cell motility in bee guts versus cell culture. In contrast, genes involved in cell adhesion were upregulated, consistent with the importance of attachment to insect tissue in this species and the family overall. CONCLUSIONS:Our analysis links differentiation of a conserved supernumerary chromosome with infection of bees, parallel to this chromosome's role in Leishmania infection of mammals and linking chromosome-level changes with adaptation to new hosts.

The western honey bee, Apis mellifera, faces a new threat from the spread of parasitic Tropilaelaps (Tropi) mites, specifically T. mercedesae, which adds additional complexity to an apicultural landscape heavily impacted by Varroa destructor and its associated virus complex. In this study conducted in central Thailand, we investigated the efficacy of two methods of applying formic acid and a thermal remediation technique in controlling Tropi mites and Varroa. We focused our attention on the reproductive stage of the mites, which is restricted to capped brood cells. Results revealed that both formic acid treatments we tested (Formic Pro and liquid formic acid) demonstrated an immediate and substantial reduction in live Tropi and Varroa populations, maintaining near-zero levels for the 3-week duration of the study. In contrast, thermal remediation, employing heating pads, exhibited a more gradual decline, achieving an 85.42% reduction in Tropi mites and a 92.33% reduction in Varroa mites by week three. Both applications of formic acid maintained residual efficacy, continuing to result in dead mites being found in cells 14 days post exposure to the chemical. No dead mites were found inside the cells after the first week of the study in the thermal remediation treatment. Notably, heat-treated colonies experienced an unexpected resurgence in mite populations during week two. The findings contribute valuable insights into potential strategies for mitigating the threat of Tropi mites (via chemical and nonchemical treatment measures) and highlight the urgency of further research to safeguard global honey bee populations.