Xinle Duan

Colony merging is one of the very important operations in beekeeping. For a safe merger, nestmate recognition is a prerequisite, which is caused by nestmate odors produced by certain volatile chemicals. Some of these chemicals are now known; however, knowledge about the entire spectrum of these chemicals is still lacking. In this article, we have attempted at exploring such chemicals. We investigated the recognition behavior of bees of different ages. Volatile cuticular hydrocarbons (CHCs) in bee groups before and after merging were analyzed via gas chromatography-mass spectrometry (GC-MS). The results indicated that newly emerged bees, which possess the lowest nestmate recognition ability, with less than 5% of observing time showing the defense behavior, develop recognition behaviors rapidly, reaching peak levels of over 35% of observing time showing the behavior at 8 days old and maintaining them throughout their lives. Each bee group has a unique CHCs profile. Merging significantly altered the original CHC profiles of bee groups. Four CHCs were up-regulated in the merged bee group. Ethyl oleate, hexadecanoic acid ethyl ester, and octadecanoic acid ethyl ester were further confirmed to significantly promote bee group merging by reducing 50% of the nestmate recognition ability (p < 0.05). These results highlight the importance of nestmate recognition in the social lives of honey bees, establish the chemical basis for honey bee nestmate recognition, and have potential practical applications in honey bee colony merging.

Propolis contains abundant flavonoid and phenolic compounds, whose composition and concentration vary significantly in different geographical origins, thereby affecting its bioactive properties including anti-inflammatory, antioxidant, and antimicrobial activities. In this study, the flavonoid and phenolic content in the ethanol extract of propolis (EEP) from Henan (HN) and Shandong (SD) provinces was quantitatively analyzed, and the results showed that concentrations of both bioactive components in HN were slightly higher than those in SD. The non-targeted metabolomics technology was further employed to analyze the components of EEP, and a total of 10683 metabolites were detected. In the comparison between the samples of HN and SD, there were a total of 1436 differential metabolites, with 553 decreased and 883 increased in the HN sample. Among them, there were 205 differential metabolites related to flavonoids and phenols, with 108 decreased and 97 increased in the HN sample. However, a greater number of carboxylic acids and derivatives, fatty derivatives and organooxygen metabolites were found at higher relative levels in the HN sample. As a result, the EEP of the HN sample was selected to investigate its inhibitory effect on inflammation in lipopolysaccharide (LPS)-induced BV2 microglia cells. The results showed that LPS promoted the M1 polarization of BV2 microglia. However, treatment with EEP at concentrations of 10 µg/mL, 5 µg/mL, and 2.5 µg/mL could partially restore the cell morphology to its non-activated state. Meanwhile, LPS stimulation increased the protein levels of IL-1β, IL-6 and TNF-α significantly, as well as the relative gene expression levels of IL-1β, IL-6, TNF-α, COX-2, iNOS and TLR4. After treatment with the EEP, the expression levels of these three proteins and six genes were significantly decreased. These findings revealed that EEP effectively inhibited the M1 polarization of LPS-induced BV2 cells and decreased the expression of inflammatory factors, indicating its potential as a therapeutic agent for neuroinflammation.

Honey bees (Apis mellifera ligustica) are essential pollinators in both ecosystems and agricultural production. However, their populations are declining due to various factors, including pesticide exposure. Despite their importance, the reproductive castes, particularly drones, remain understudied in terms of pesticide effects. To investigate the effects of azoxystrobin and picoxystrobin on honey bee drones, the drones were exposed to different concentrations of azoxystrobin and picoxystrobin for 14 days; the drone survival, body weight, nutrient content, reproductive organs, and sperm concentration were assessed. Results showed that exposure to both fungicides caused a significant reduction in drone survival rates, with survival rates decreasing progressively as the duration of exposure increased. Compared to the control group, the body weights of drones in all treatment groups were significantly lower on days 7 and 14. Nutrient analysis revealed that low concentrations of azoxystrobin and picoxystrobin increased protein levels, while free fatty acid content decreased significantly in all treatment groups. No significant changes were observed in the total carbohydrate content. Morphological examination of reproductive organs showed that the lengths of the mucus glands and seminal vesicles in drones were significantly shorter in the treatment groups compared to the control group. Furthermore, exposure to azoxystrobin and picoxystrobin resulted in a significant decline in sperm concentration in the drones. These findings indicate that azoxystrobin and picoxystrobin have adverse effects on the health and reproductive capacity of honey bee drones. The present study highlights the need to reassess the risks posed by these fungicides to pollinators, particularly given the critical role of drones in maintaining the genetic diversity and resilience of honey bee colonies. Further research is warranted to elucidate the underlying mechanisms of these effects and explore potential mitigation strategies.

High temperature is normally harmful to an organism. However, honey bees evolve, maintaining a relatively higher colony temperature of 34.5 °C in the long reproduction period. To determine the effect of such a higher colony temperature on adult bees and its biological significance, newly emerged bees were reared in cages at 34.5 °C and room temperature of 25.0 °C, respectively. Their survival rate, head weight, royal jelly-secreting gene expression, and morphology of the hypopharyngeal gland were investigated. Moreover, 40-day-old bees with significant differences in survival rate between the two temperature groups were subject to transcriptome and lipidome analysis. The result showed that the higher colony temperature was overall negative for the bees’ longevity. Transcriptome analysis showed that fatty acid metabolism-related items were enriched and the involved genes were upregulated in honey bees reared at 34.5 °C compared with the honey bees reared at 25.0 °C. Lipidomic analysis further validated that fatty acid metabolism, especially sphingolipid metabolism, was significantly altered. Such upregulation of fatty acid metabolism-related genes was also detected in young adult bees of 5 days old reared at 34.5 °C. These bees had heavier head weights, higher expression of royal jelly-secreting-related genes, and more developed hypopharyngeal glands. Such results showed that the colony temperature of 34.5 °C could accelerate the development process of newly emerged bees to be nurse bees, significantly increasing the colony nurturing capability, which in turn increased the development speed, size, and survivability of the colony. Thereby, the colony temperature of 34.5 °C shortened the lifespan of individual bees, but obtained huge returns at the colony level, with remarkable biological significance.

Fungal diseases of agricultural crops cause severe economic losses to the growers. For the control of these diseases, azoxystrobin is one of the recommended fungicides. This fungicide is systemic in action and is expected to reach the floral part of the treated crop and its residue in the pollen and nectar, the natural food sources of honey bees, which could be collected and fed on by honey bees, thus affecting their health. The purpose of this study was to determine the physiological and chemical changes caused by this fungicide in honey bee workers (Apis mellifera L). Workers of this honey bee at 1, 8, and 21 days old were treated with 125, 167, and 250 mg/L concentrations of azoxystrobin for seven days; their survival rates, activities of carboxylesterase (CarE), glutathione S-transferases (GSTs), cytochrome P450 enzyme (CYP450), catalase (CAT), and superoxide dismutase (SOD) enzymes, and the expression levels of immune (Aba, Api, Def1, and Hym) and nutrition genes (Ilp1, Ilp2, and Vg) were detected. Our findings revealed that azoxystrobin affected the survival of workers, particularly 1- and 21-day-old workers, who responded to azoxystrobin stress with increased activities of detoxification and protective enzymes, which might have physiological costs. Additionally, azoxystrobin affected the expression of immune and nutrition genes, with a decreased expression trend in 21-day-old workers compared to the 1- and 8-day-old workers, leading to reduced resistance to external stressors and increased mortality rates. These findings provide important insights into the adverse effects of azoxystrobin on workers of different ages and emphasize the potential risks of this chemical to colony stability and individual health. This study recommends an urgent ban on such a harmful fungicide being used for fungi control in agriculture, especially during plant flowering.

Honeybee larvae and pupae form the cornerstone of colony survival, development, and reproduction. Azoxystrobin is an effective strobilurin fungicide that is applied during the flowering stage for controlling plant pathogens. The contaminated nectar and pollen resulting from its application are collected by forager bees and impact the health of honeybee larvae and pupae. The current study evaluated the survival, development, and physiological effects of azoxystrobin exposure on the larvae and pupae of Apis mellifera worker bees. The field-recommended concentrations of azoxystrobin were found to suppress the survival indices and lifespan in the larval as well as pupal stages; moreover, the rates of the survival and pupation of larvae as well as the body weights of the pupae and newly-emerged adult bees were significantly reduced upon long-term exposure to azoxystrobin. In addition, azoxystrobin ingestion induced changes in the expression of genes critical for the development, immunity, and nutrient metabolism of larvae and pupae, although the expression profile of these genes differed between the larval and pupal stages. Results indicated the chronic toxicity of azoxystrobin on the growth and development of honeybee larvae and pupae, which would affect their sensitivity to pathogens and other external stresses during the development stage and the study will provide vital information regarding the pollination safety and rational use of pesticides.

Varroa destructor injects a salivary secretion into honeybees during their feeding process. The salivary secretion plays a vital role in mite–bee interactions and is the main cause of honeybee illness. To determine the biological function of cystatin-L2-like, one of the components of V. destructor salivary secretion, its gene expression in mites during the reproductive phase and dispersal phase was quantified using RT-qPCR, respectively. Moreover, the E. coli-expressed and -purified cystatin was injected into the white-eyed honeybee pupae, and its effects on the survival, the weight of the newly emerged bee, and the transcriptome were determined. The results showed that cystatin was significantly upregulated in mites during the reproductive phase. Cystatin significantly shortened the lifespan of pupae and decreased the weight of the newly emerged bees. Transcriptome sequencing showed that cystatin upregulated 1496 genes and downregulated 1483 genes in pupae. These genes were mainly enriched in ATP synthesis, the mitochondrial respiratory chain, and cuticle structure and function. Cystatin comprehensively downregulated the metabolism of carbohydrates, fatty acids, and amino acids, and energy production in the pupae. The downregulation of metabolic activity could save more nutrients and energy for V. destructor, helping it to maximize its reproduction potential, implying that the mite could manipulate the metabolism of host bees through the injected salivary secretion. The results provide new insights into mite–bee interactions, providing a basis for related studies and applications.

The strobilurin fungicide pyraclostrobin is widely used to prevent and control the fungal diseases of various nectar and pollen plants. Honeybees also directly or indirectly contact this fungicide with a long-term exposure period. However, the effects of pyraclostrobin on the development and physiology of Apis mellifera larvae and pupae during continuous exposure have been rarely known. To investigate the effects of field-realistic concentrations of pyraclostrobin on honeybee survival and development, the 2-day-old larvae were continuously fed with different pyraclostrobin solutions (100 mg/L and 83.3 mg/L), and the expression of development-, nutrient-, and immune-related genes in larvae and pupae were examined. The results showed that two field-realistic concentrations of pyraclostrobin (100 and 83.3 mg/L) significantly decreased the survival and capped rate of larvae, the weight of pupae and newly emerged adults, and such decrease was a positive correlation to the treatment concentrations. qPCR results showed that pyraclostrobin could induce the expression of Usp, ILP2, Vg, Defensin1, and Hymenoptaecin, decrease the expression of Hex100, Apidaecin, and Abaecin in larvae, could increase the expression of Ecr, Usp, Hex70b, Vg, Apidaecin, and Hymenoptaecin, and decreased the expression of ILP1, Hex100 and Defensin1in pupae. These results reflect pyraclostrobin could decrease nutrient metabolism, immune competence and seriously affect the development of honeybees. It should be used cautiously in agricultural practices, especially in the process of bee pollination.

Increasing ELF-EMF pollution in the surrounding environment could impair the cognition and learning ability of honeybees, posing a threat to the honeybee population and its pollination ability. In a social honeybee colony, the numbers of adult bees rely on the successful large-scale rearing of larvae and continuous eclosion of new adult bees. However, no studies exist on the influence of ELF-EMFs on honeybee larvae. Therefore, we investigated the survival rate, body weight, and developmental duration of first instar larvae continuously subjected to ELF-EMF exposure. Moreover, the transcriptome of fifth instar larvae were sequenced for analyzing the difference in expressed genes. The results showed that ELF-EMF exposure decreases the survival rate and body weight of both white-eye pupae and newly emerged adults, extends the duration of development time and seriously interferes with the process of metamorphosis and pupation. The transcriptome sequencing showed that ELF-EMF exposure decreases the nutrient and energy metabolism and impedes the degradation of larvae tissues and rebuilding of pupae tissues in the metamorphosis process. The results provide an experimental basis and a new perspective for the protection of honeybee populations from ELF-EMF pollution.