In honey bees, reproductive division of labor is maintained by social suppression of worker fertility, yet queen loss can trigger ovary activation in workers. Here, we tested whether endogenous dopaminergic signaling is progressively remodeled across successive stages of ovarian activation and how these changes relate to key hormone pathways. Newly emerged Apis mellifera workers were introduced into queenright or queenless colonies, collected after 14 days, and classified as having inactive, partially activated, or fully activated ovaries. We quantified brain dopamine and measured expression levels of genes involved in dopamine synthesis, transport, metabolism, and reception in both brain and ovary tissues, together with transcriptional markers of juvenile hormone (JH) and 20-hydroxyecdysone (20E) signaling. Brain dopamine increased with ovary activation and peaked in fully activated workers, coincident with elevated transcripts of tyrosine hydroxylase, dopa decarboxylase, dopamine transporter, and arylalkylamine N-acetyltransferase in the brain. Dopamine receptor genes were stable in the brain but were remodeled in the ovary, with Amdop1 increasing and Amdop3 decreasing during activation. Markers of JH signaling and ovarian 20E pathway activity also rose with ovarian development, consistent with early endocrine priming following queen loss. Collectively, these results support an integrated neuroendocrine framework in which dopaminergic remodeling and hormone pathway activation jointly accompany worker reproductive activation under queenless conditions.

Temporal reproductive plasticity is a central life-history trait for most iteroparous animals, yet its underlying molecular mechanisms remain poorly understood. We utilize the reversible reproductive arrest experienced by honey bee queens during colony swarming to investigate how these reproductive specialists dynamically adapt their physiology. We demonstrate that pre-swarming nutritional restriction triggers a resource reallocation, whereby ovarian mass declines as flight muscle function is enhanced, providing empirical support for a flight-fecundity trade-off. This transition is accompanied by elevated juvenile hormone and suppressed ecdysteroid and vitellogenin levels, together with the engagement of three spatially distinct oogenesis checkpoints. These checkpoints engage programmed cell death pathways: autophagy predominates in germarial stem cells and follicle cells, while apoptosis is the primary mechanism in vitellarium oocytes and nurse cells, collectively orchestrating oogenesis arrest. Integrated whole-transcriptomic and metabolomic analyses revealed broad molecular remodeling, including changes in FoxO-, Notch-, Wnt-, and Hippo-related signatures that accompanied oogenesis suppression. Our results indicate a "nutrition-hormone-checkpoint-programmed cell death" model that links colony-level swarming to individual ovary suppression. This work provides a systematic framework for understanding the molecular regulation of the reversible reproductive plasticity of honey bee queens.

The ectoparasitic honeybee (Apis mellifera) mite Tropilaelaps mercedesae represents a serious threat to Asian apiculture and a growing concern for global beekeeping due to its high reproductive capacity and host adaptability. However, the regulatory mechanisms underlying its host adaptation across life stages remain poorly characterized. Here, we performed integrated transcriptomic, proteomic, and metabolomic analyses of female mites at 4 key postembryonic developmental stages: protonymphs, deutonymphs, mature adults, and reproductive adults. Our multi-omics approach reveals distinct stage-specific regulatory programs. Nymphal stages exhibit coordinated activation of chitin metabolism pathways for structural growth, neuroactive ligand-receptor interactions for neural maturation, and ATP-binding cassette (ABC) transporters for detoxification. Adult maturation involves enhanced membrane dynamics, improved energy metabolism, and increased antioxidant defenses to support reproductive capacity and environmental resilience. Reproductive adults prioritize lipid metabolism and vitellogenin synthesis to fuel embryonic development. Crucially, host adaptation emerges as a critical driver of this stage-specific regulation. Additionally, multi-omics integration highlights a developmental transition from transcriptional dominance in early stages to post-transcriptional regulation in later phases. These insights provide a systems-level understanding of the physiological orchestration enabling this parasite's ecological success, while identifying valuable molecular targets for formulating more effective strategies to control this formidable pest.

Cadmium (Cd), a pervasive heavy metal and potent environmental contaminant, threatens ecosystem integrity, yet its impacts on pollinating insects remain insufficiently characterized. In this study, we systematically investigated the consequences of larval-stage Cd exposure on honeybee (Apis mellifera) queens through integrated physiological, metabolomic, and proteomic analyses. Sublethal Cd exposure disrupted hormone balance, induced midgut epithelial degeneration, and impaired hemolymph lipid homeostasis, collectively retarding queen developmental progression and physiological maturation. Ovarian proteomic profiling further revealed the Cd-mediated suppression of key proteins involved in muscle contraction, tracheal function, neural conduction, and antioxidant defense, ultimately leading to diminished reproductive vitality. Critically, high-dose Cd exposure elicited transgenerational impairments: F1 workers showed reduced body size, lowered sucrose responsiveness, and impaired olfactory learning, while F1 queens developed smaller ovaries with fewer ovarioles. Together, these results demonstrate that Cd imposes substantial multigenerational fitness costs on honeybee colonies via both direct physiological impairment and transgenerational toxicity. Our findings underscore the urgency of mitigating environmental Cd pollution to protect pollinator health and ensure sustainable ecosystems.

Fenazaquin, a potent insecticide widely used to control phytophagous mites, has recently emerged as a potential solution for managing Varroa destructor mites in honeybees. However, the comprehensive impact of fenazaquin on honeybee health remains insufficiently understood. Our current study investigated the acute and chronic toxicity of fenazaquin to honeybee larvae, along with its influence on larval hemolymph metabolism and gut microbiota. Results showed that the acute median lethal dose (LD50) of fenazaquin for honeybee larvae was 1.786 μg/larva, and the chronic LD50 was 1.213 μg/larva. Although chronic exposure to low doses of fenazaquin exhibited no significant effect on larval development, increasing doses of fenazaquin resulted in significant increases in larval mortality, developmental time, and deformity rates. At the metabolic level, high doses of fenazaquin inhibited nucleotide, purine, and lipid metabolism pathways in the larval hemolymph, leading to energy metabolism disorders and physiological dysfunction. Furthermore, high doses of fenazaquin reduced gut microbial diversity and abundance, characterized by decreased relative abundance of functional gut bacterium Lactobacillus kunkeei and increased pathogenic bacterium Melissococcus plutonius. The disrupted gut microbiota, combined with the observed gut tissue damage, could potentially impair food digestion and nutrient absorption in the larvae. Our results provide valuable insights into the complex and diverse effects of fenazaquin on honeybee larvae, establishing an important theoretical basis for applying fenazaquin in beekeeping.

Ectoparasitic mites of the genera Varroa and Tropilaelaps have evolved to exclusively exploit honey bees as food sources during alternating dispersal and reproductive life history stages. Here we show that the primary food source utilized by Varroa destructor depends on the host life history stage. While feeding on adult bees, dispersing V. destructor feed on the abdominal membranes to access to the fat body as reported previously. However, when V. destructor feed on honey bee pupae during their reproductive stage, they primarily consume hemolymph, indicated by wound analysis, preferential transfer of biostains, and a proteomic comparison between parasite and host tissues. Biostaining and proteomic results were paralleled by corresponding findings in Tropilaelaps mercedesae , a mite that only feeds on brood and has a strongly reduced dispersal stage. Metabolomic profiling of V. destructor corroborates differences between the diet of the dispersing adults and reproductive foundresses. The proteome and metabolome differences between reproductive and dispersing V. destructor suggest that the hemolymph diet coincides with amino acid metabolism and protein synthesis in the foundresses while the metabolism of non-reproductive adults is tuned to lipid metabolism. Thus, we demonstrate within-host dietary specialization of ectoparasitic mites that coincides with life history of hosts and parasites.

The threat of neonicotinoids to insect pollinators, particularly honeybees (Apis mellifera), is a global concern, but the risk of chiral neonicotinoids to insect larvae remains poorly understood. In the current study, we evaluated the acute and chronic toxicity of dinotefuran enantiomers to honeybee larvae in vitro and explored the mechanism of toxicity. The results showed that the acute median lethal dose (LD50) of S-dinotefuran to honeybee larvae was 30.0 μg/larva after oral exposure for 72 h, which was more toxic than rac-dinotefuran (92.7 μg/larva) and R-dinotefuran (183.6 μg/larva). Although the acute toxicity of the three forms of dinotefuran to larvae was lower than that to adults, chronic exposure significantly reduced larval survival, larval weight, and weight of newly emerged adults. Analysis of gene expression and hormone titer indicated that dinotefuran affects larval growth and development by interfering with nutrient digestion and absorption and the molting system. Analysis of hemolymph metabolome further revealed that disturbances in the neuroactive ligand-receptor interaction pathway and energy metabolism are the key mechanisms of dinotefuran toxicity to bee larvae. In addition, melatonin and vitellogenin are used by larvae to cope with dinotefuran-induced oxidative stress. Our results contribute to a comprehensive understanding of dinotefuran damage to bees and provide new insights into the mechanism of enantioselective toxicity of insecticides to insect larvae.

Tropilaelaps mercedesae, an ectoparasitic mite of honeybees, is currently a severe health risk to Apis mellifera colonies in Asia and a potential threat to the global apiculture industry. However, our understanding of the physiological and developmental regulation of this pest remains significantly insufficient. Using ultra-high resolution mass spectrometry, we provide the first comprehensive proteomic profile of T. mercedesae spanning its entire post-embryonic ontogeny, including protonymphs, deutonymphs, mature adults, and reproductive mites. Consequently, a total of 4,422 T. mercedesae proteins were identified, of which 2,189 proteins were significantly differentially expressed (FDR < 0.05) throughout development and maturation. Our proteomic data provide an important resource for understanding the biology of T. mercedesae, and will contribute to further research and effective control of this devastating honeybee pest.

In this article, we report the pathogenicity of a new strain of fungus, Rhizopus oryzae to honeybee larvae, isolated from the chalkbrood-diseased mummies of honeybee larvae and pupae collected from apiaries in China. Based on morphological observation and internal transcribed spacer (ITS) region analyses, the isolated pathogenic fungus was identified as R. oryzae. Koch’s postulates were performed to determine the cause-and-effect pathogenicity of this isolate fungus. The in vitro pathogenicity of this virulent fungus in honeybees was tested by artificially inoculating worker larvae in the lab. The pathogenicity of this new fungus for honeybee larvae was both conidial-concentration and exposure-time dependent; its highly infectious and virulent effect against the larvae was observed at 1 × 105 conidia/larva in vitro after 96 h of challenge. Using probit regression analysis, the LT50 value against the larvae was 26.8 h at a conidial concentration of 1 × 105 conidia/larva, and the LC50 was 6.2 × 103 conidia/larva. These results indicate that the new isolate of R. oryzae has considerable pathogenicity in honeybee larvae. Additionally, this report suggests that pathogenic phytofungi may harm their associated pollinators. We recommend further research to quantify the levels, mechanisms, and pathways of the pathogenicity of this novel isolated pathogen for honeybee larvae at the colony level.

Environmental variation selects for the adaptive plasticity of maternal provisioning. Even though developing honeybees find themselves in a protected colony environment, their reproductively specialized queens actively adjust their maternal investment, even among worker-destined eggs. However, the potentially adaptive consequences of this flexible provisioning strategy and their mechanistic basis are unknown. Under natural conditions, we find that the body size of larvae hatching from small eggs in large colonies converges with that of initially larger larvae hatching from large eggs typically produced in small colonies. However, large eggs confer a persistent body size advantage when small and large eggs are cross-fostered in small and large colonies, respectively. We substantiate the increased maternal investment by identifying growth-promoting metabolomes and proteomes in large eggs compared to small eggs, which are primarily enriched in amino acid metabolism and cell maturation. Thus, our study provides a comprehensive adaptive explanation for the worker egg size plasticity of honeybees.

The economic and environmental value of honeybees has been severely challenged in recent years by the collapse of their colonies worldwide, often caused by outbreaks of infectious diseases. However, our understanding of the diversity, prevalence, and transmission of honeybee viruses is largely obscure due to a lack of large-scale and longitudinal genomic surveillance on a global scale. We report the meta-transcriptomic sequencing of nearly 2000 samples of the two most important economic and widely maintained honeybee species, as well as an associated ectoparasite mite, collected across China during 2016–2019. We document the natural diversity and evolution of honeybee viruses in China, providing evidence that multiple viruses commonly co-circulate within individual bee colonies. We also expanded the genomic data for 12 important honeybee viruses and revealed novel genetic variants and lineages associated with China. We identified more than 23 novel viruses from the honeybee and mite viromes, with some exhibiting ongoing replication in their respective hosts. Together, these data provide additional support to the idea that mites are an important reservoir and spill-over host for honeybee viruses. Our data show that honeybee viruses are more widespread, prevalent, and genetically diverse than previously realized. The information provided is important in mitigating viral infectious diseases in honeybees, in turn helping to maintain sustainable productive agriculture on a global scale.

Tissue phenotypic plasticity facilitates rapid adaptation of organisms to biotic and/or abiotic pressure. The reproductive capacity of honey bee workers (Apis mellifera) is plastic and responsive to pheromones produced by broods and the queen. Egg laying workers (ELWs), which could reactivate their ovaries and lay haploid eggs upon queen lost, have been commonly discussed from many aspects. However, it remains unclear whether midgut homeostasis in ELWs is affected during plastic changes. Here, we found that the expression of nutrition- and autophagy-related genes was up-regulated in the midguts of ELWs, compared with that in nurse workers (NWs) by RNA-sequencing. Furthermore, the area and number of autophagosomes were increased, along with significantly increased cell death in the midguts of ELWs. Moreover, cell cycle progression in the midguts of ELWs was increased compared with that in NWs. Consistent with the up-regulation of nutrition-related genes, the body and midgut sizes, and the number of intestinal proliferation cells of larvae reared with royal jelly (RJ) obviously increased more than those reared without RJ in vitro. Finally, cell proliferation was dramatically suppressed in the midguts of ELWs when autophagy was inhibited. Altogether, our data suggested that autophagy was induced and required to sustain cell proliferation in ELWs’ midguts, thereby revealing the critical role of autophagy played in the intestines during phenotypic plasticity changes.

一、切叶蜂的分类地位及种类 切叶蜂科(Megachilidae)是蜜蜂总科中长口器类的一个大科.切叶蜂科形态多样,具有寄生和非寄生两个亚群.切叶蜂科其下包括两个亚科:切叶蜂亚科和毛腹蜂亚科.据不完全统计,目前全世界切叶蜂亚科共203属及亚属,近4000种.中国有记载的切叶蜂18属,47亚属,305种及亚种.

Egg size plasticity represents an adaptive reproductive strategy in numerous organisms, including the honey bee, Apis mellifera . However, the proximate causation of this plasticity and egg size in general is unknown. We show that honey bee queens predictably and reversibly adjust egg size in response to their colony size and that this plasticity is an active response to the queens’ perception of colony size instead of a consequence of egg laying rate. The egg size increase involves changes of 290 ovarian proteins, mostly related to increased energy metabolism, protein transport, and cytoskeleton functions. Spatio-temporal expression analysis of the small GTPase Rho1 indicates its central role in egg size regulation, which we confirm by RNAi-mediated gene knock-down and expression analyses. The molecular adjustments that promote maternal investment of honey bee queens in response to their social environment thus reveal a novel mechanism of egg size regulation.

Ascosphaera apis is the causative agent of honey bee chalkbrood disease, and spores are the only known source of infections. Interference with sporulation is therefore a promising way to manage A. apis. The versicolorin reductase gene (StcU-2) is a ketoreductase protein related to sporulation and melanin biosynthesis. To study the StcU-2 gene in ascospore production of A. apis, CRISPR/Cas9 was used, and eight hygromycin B antibiotic-resistant transformants incorporating enhanced green fluorescent protein (EGFP) were made and analyzed. PCR amplification, gel electrophoresis, and sequence analysis were used for target gene editing analysis and verification. The CRISPR/Cas9 editing successfully knocked out the StcU-2 gene in A. apis. StcU-2 mutants had shown albino and non-functional spore-cyst development and lost effective sporulation. In conclusion, editing of StcU-2 gene has shown direct relation with sporulation and melanin biosynthesis of A. apis; this effective sporulation reduction would reduce the spread and pathogenicity of A. apis to managed honey bee. To the best of our knowledge, this is the first time CRISPR/Cas9-mediated gene editing has been efficiently performed in A. apis, a fungal honey bee brood pathogen, which offers a comprehensive set of procedural references that contributes to A. apis gene function studies and consequent control of chalkbrood disease.

Varroa destructor is a major pathogenic driver of the Western honeybee colony losses globally. Understanding the developmental regulation of V. destructor is critical to develop effective control measures. Development is a complex biological process regulated by numerous genes and long non-coding RNAs (lncRNAs); however, the underlying regulation of lncRNAs in the development of V. destructor remains unknown. In this study, we analyzed the RNA sequencing (RNA-Seq) data derived from the four stages of female V. destructor in the reproductive phase (i.e., egg, protonymph, deutonymph, and adult). The identified differentially expressed mRNAs and lncRNAs exhibited a stage-specific pattern during developmental transitions. Further functional enrichment established that fat digestion and absorption, ATP-binding cassette (ABC) transporters, mitogen-activated protein kinase (MAPK) signaling pathway, and ubiquitin-proteasome pathway play key roles in the maturation of female V. destructor. Moreover, the lncRNAs and mRNAs of some pivotal genes were significantly upregulated at the deutonymph stage, such as cuticle protein 65/6.4/63/38 and mucin 5AC, suggesting that deutonymph is the key stage of metamorphosis development and pathogen resistance acquisition for female V. destructor. Our study provides novel insights into a foundational understanding of V. destructor biology.

Insect pollinators provide major pollination services for wild plants and crops. Honeybee viruses can cause serious damage to honeybee colonies. However, viruses of other wild pollinating insects have yet to be fully explored. In the present study, we used RNA sequencing to investigate the viral diversity of 50 species of wild pollinating insects. A total of 3 pathogenic honeybee viruses, 8 previously reported viruses, and 26 novel viruses were identified in sequenced samples. Among these, 7 novel viruses were shown to be closely related to honeybee pathogenic viruses, and 4 were determined to have potential pathogenicity for their hosts. The viruses detected in wild insect pollinators were mainly from the order Picornavirales and the families Orthomyxoviridae, Sinhaliviridae, Rhabdoviridae, and Flaviviridae. Our study expanded the species range of known insect pollinator viruses, contributing to future efforts to protect economic honeybees and wild pollinating insects.

Reproduction involves the investment of resources into offspring. Although variation in reproductive effort often affects the number of offspring, adjustments of propagule size are also found in numerous species, including the Western honey bee, Apis mellifera . However, the proximate causes of these adjustments are insufficiently understood, especially in oviparous species with complex social organization in which adaptive evolution is shaped by kin selection. Here, we show in a series of experiments that queens predictably and reversibly increase egg size in small colonies and decrease egg size in large colonies, while their ovary size changes in the opposite direction. Additional results suggest that these effects cannot be solely explained by egg-laying rate and are due to the queens’ perception of colony size. Egg-size plasticity is associated with quantitative changes of 290 ovarian proteins, most of which relate to energy metabolism, protein transport, and cytoskeleton. Based on functional and network analyses, we further study the small GTPase Rho1 as a candidate regulator of egg size. Spatio-temporal expression analysis via RNAscope and qPCR supports an important role of Rho1 in egg-size determination, and subsequent RNAi-mediated gene knockdown confirmed that Rho1 has a major effect on egg size in honey bees. These results elucidate how the social environment of the honey bee colony may be translated into a specific cellular process to adjust maternal investment into eggs. It remains to be studied how widespread this mechanism is and whether it has consequences for population dynamics and epigenetic influences on offspring phenotype in honey bees and other species.