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Abstract Long non-coding RNAs (lncRNAs) are crucial modulators of post-transcriptional gene expression regulation, cell fate determination, and disease development. However, lncRNA functions during short-term heat stress in adult worker bees are poorly understood. Here, we performed deep sequencing and bioinformatic analyses of honeybee lncRNAs. RNA interference was performed by using siRNA targeting the most highly expressed lncRNA. The silencing effect on lncRNA and the relative expression levels of seven heat shock protein (HSP) genes, were subsequently examined. Overall, 7,842 lncRNAs and 115 differentially expressed lncRNAs (DELs) were identified in adult worker bees following heat stress exposure. Structural analysis revealed that the overall expression abundance, length of transcripts, exon number, and open reading frames of lncRNAs were lower than those of mRNAs. GO analysis revealed that the target genes were mainly involved in “metabolism,” “protein folding,” “response to stress,” and “signal transduction” pathways. KEGG analysis indicated that the “protein processing in endoplasmic reticulum” and “longevity regulating pathway-multiple species” pathways were most enriched. Quantitative real-time polymerase chain reaction (qRT-PCR) detection of the selected DELs confirmed the reliability of the sequencing data. Moreover, the siRNA experiment indicated that feeding siRNA yielded a silencing efficiency of 77.51% for lncRNA MSTRG.9645.5. Upon silencing this lncRNA, the expression levels of three HSP genes were significantly downregulated (p

Bumblebees are primary social insects and a vital class of pollinating insects. Their distinctive reproductive mode is characterized by the independent initiation and construction of the nest by the queen and the subsequent production of sufficient workers, males, and gynes following colony development. After successful mating, the queen transitions to the first phase of its annual life cycle. The reproductive processes are directly influenced by environmental factors, including floral resources and pesticides. Moreover, the reproductive level is regulated by biological factors, particularly the role of workers, who participate in egg laying and pass on their genetic material to the next generation of queens. Successful reproduction can only be achieved by maintaining colony development under natural or artificial breeding conditions. Consequently, understanding the known factors that influence bumblebee reproduction is essential for developing conservation strategies for wild bumblebees and for successfully breeding diverse bumblebee species. Breeding various bumblebee species is crucial for in-depth research into known factors and for further exploration of other potential factors, which will also help to meet the demand for pollination in agricultural facilities globally.

A large number of studies have shown that propolis has positive effects in the treatment of type 2 diabetes mellitus (T2DM). However, there are have only been a few reports that are based on an ultra-high performance liquid chromatography-quadrupole-time-of-flight-mass spectrometry (UPLC-Q-TOF-MS) analysis of the fecal metabolomics of ethanol extract of propolis (EEP) in the treatment of T2DM. The present investigation was designed to screen potential biomarkers of T2DM by the metabonomic method and to explain the possible anti-diabetes mechanism of EEP according to the changes in the biomarkers. The results showed that EEP improved the body weight (BW) of T2DM mice, lowered blood sugar levels, and significantly restored blood biochemical indicators related to T2DM, such as fasting insulin (FINS), homeostasis model assessment of insulin resistance (HOMA-IR), aspartate transaminase (AST), and alanine aminotransferase (ALT). Liver pathology showed that EEP reversed liver damage caused by T2DM. Metabolomics data identified 27 potential biomarkers in fecal samples. EEP effectively regulated the dysfunction in the metabolic pathways of glycerophospholipids, sphingolipids, riboflavins, and sterol lipids caused by T2DM. In summary, our research results revealed positive effects of EEP in the treatment of T2DM and provided potential candidate markers for further research and in the clinical treatment of T2DM.

Bee pollen is collected from flowering plants and added with nectar and salivary gland secretions by honeybees. Bee pollen is abundant in various nutrients and active ingredients, and exerts excellent medicinal efficacy. However, some susceptible population developed clinical allergic symptoms due to eating bee pollen, which caused serious influences to the food safety and further application of bee pollen. Currently, the research on the allergenic mechanism of bee pollen is insufficient, which limits the utilization of bee pollen. The research advances of bee pollen allergenicity and allergens, as well as the microbial fermentation techniques for allergen degradation are summarized and discussed, aiming to provide a reference for the development of bee pollen allergen removal technology in the future.

High temperature affects behavior, physiology, survival, and the expression of related genes in adult honeybees. Apis mellifera is the common pollinator in greenhouse and is susceptible to high temperature stress. To further explore the molecular basis related to heat stress, we compared the transcriptome profiles of adult worker bees at 25 and 45°C, and detected the expression patterns of some differentially expressed genes (DEGs) in different tissues by q RT-PCR. Differential expression analysis showed that 277 DEGs were identified, including 167 genes upregulated and 110 genes downregulated after heat stress exposure in adult worker bees. In GO enrichment analysis, DEGs were mostly enriched for protein folding, unfold protein binding, and heme binding terms. Protein processing in endoplasmic reticulum and longevity regulating pathway-multiple species were significantly enriched in KEGG. The expression levels of 16 DEGs were consistent with the transcriptome results. The expression patterns of 9 DEGs in different tissues revealed high levels in the thorax, which was supposed that the thorax may be the most important part in the response to heat stress. This study provided valuable data for exploring the function of heat resistance-related genes.

Bees rely on their sensitive olfactory system to perform foraging activities in the surrounding environment. This ability is associated with the existence of olfactory receptors (ORs). In this study, we identified the AcerOr2 (ortholog to the Orco) protein in Apis cerana cerana, which contains a conserved, putative calmodulin (CaM)-binding site (CBS) indicating that CaM is involved in its function. We used immunofluorescence, Western blot, and Ca2 + imaging to monitor changes in the expression and activation of the signaling pathway associated with Ca2 + and Ca2 +/CaM-dependent protein kinase II (CaMKII) in Sf9 cells heterologously expressing AcerOr2 and a CaM-binding mutant. We used the synthetic Orco agonist VUAA1 to stimulate the cells or the antagonist W7 to inhibit CaM activity. The AcerOr2 CaM-binding mutant has a point mutation in the putative CBS (K331N). When heterologously expressed in Sf9 cells, the mutant should have less CaM activity. When the cells expressing AcerOr2 were treated with W7, the Ca2 + response of AceOr2 was similar to that of the mutant stimulated by VUAA1, and the expression of the CaM, CaMKII, and p-CaMKII has similar effects. Our results suggest that CaM activity affects the function of AceOr2 in vitro and can be used to further study the interaction between the AcerOr2 and calcium/CaM signaling pathway in the pollen collection behavior of bees.

Bee pollen is consumed for its nutritional and pharmacological benefits, but it also contains hazardous allergens which have not been identified. Here, we identified two potential allergens, glutaredoxin and oleosin-B2, in Brassica napus bee pollen using mass spectrometry-based proteomics analyses, and used bioinformatics to predict their antigenic epitopes. Comparison of fermented (by Saccharomyces cerevisiae) and unfermented bee pollen samples indicated that glutaredoxin and oleosin-B2 contents were significantly decreased following fermentation, while the contents of their major constituent oligopeptides and amino acids were significantly increased based on metabolomics analyses. Immunoblot analysis indicated that the IgE-binding affinity with extracted bee pollen proteins was also significantly decreased after fermentation, suggesting a reduction in the allergenicity of fermented bee pollen. Furthermore, fermentation apparently promoted the biosynthesis of L-valine, L-isoleucine, L-tryptophan, and L-phenylalanine, as well as their precursors or intermediates. Thus, fermentation could potentially alleviate allergenicity, while also positively affecting nutritional properties of B. napus bee pollen. Our findings might provide a scientific foundation for improving the safety of bee pollen products to facilitate its wider application.

Strawberries are popular fruits around the world, and their yield and fruit quality rely on pollination by honey bees and bumblebee colonies. Both bee species have their own advantages in strawberry pollination. This study investigates the characteristic of strawberry (Fragaria × ananassa ‘Red Face’) flower volatiles and their effects on bee pollinators by (1) detecting the volatile compounds of strawberry flowers by polydimethylsiloxane (PDMS) combined gas chromatography-mass spectrometry (GC-MS), (2) determining whether Bombus terrestris or Apis mellifera showed antennae responses to certain compounds of strawberry flower volatiles by an electroantennography test (EAG), and (3) testing whether these compounds could elicit a corresponding behavioral response in bees. The results showed that (1) there were 38 chemical compounds in ‘Red Face’ volatiles with 7 types, most of which were known to be generally emitted by flowers but also have some compounds that have not been reported in strawberry flowers; (2) B. terrestris and A. mellifera had strong EAG responses to several compounds, respectively, especially to ethyl benzoate, (Z)-3-hexenyl propionate, (Z)-3-hexenyl acetate, benzeneacetaldehyde and melonal; and (3) both bee species showed significant avoidance behaviors to four tested compounds, especially the B. terrestris. Flower volatiles of strawberry ‘Red face’ were different from other strawberry varieties that have been reported; some of these electrophysiologically active compounds could cause antennal potential responses in bees, as well as behavioral responses. Our study supports the idea that strawberry flower volatiles are one of the factors influencing bee foraging decisions and provides a reference for formulating more reasonable bee pollination to improve strawberry fruit quality.

As important pollinators, honeybees and bumblebees present a pollination behavior that is influenced by flower volatiles through the olfactory system. In this study, volatile compounds from melon flowers were isolated and identified by headspace solid-phase microextraction (HS-SPME) and gas chromatography–mass spectrometry (GC-MS), and their effects on Apis mellifera and Bombus terrestris were investigated by electroantennogram (EAG) and behavior tests (Y-tube olfactometer). The results showed that 77 volatile compounds were detected in melon flowers, among which the relative content of aldehydes was the highest (61.34%; 82.09%). A. mellifera showed a strong EAG response to e-2-hexenal, e-2-octenal, and 1-nonanal. B. terrestris showed a strong EAG response to e-2-hexenal, e-2-octenal, 2,5-dimethyl-benzaldehyde, benzaldehyde and benzenepropanal. In behavior tests, the volatiles with the highest attractive rate to A. mellifera were e-2-hexenal (200 μg/μL, 33.33%) and e-2-octenal (300 μg/μL, 33.33%), and those to B. terrestris were e-2-hexenal (10 μg/μL, 53.33%) and 2,5-dimethyl-benzaldehyde (100 μg/μL, 43.33%). E-2-hexenal and e-2-octenal were more attractive to A. mellifera than B. terrestris, respectively (10 μg/μL, 10 μg/μL, 200 μg/μL). In conclusion, the volatiles of melon flowers in facilities have certain effects on the electrophysiology and behavior of bees, which is expected to provide theoretical and technical support for the pollination of A. mellifera and B. terrestris in facilities.

Bumble bees and honey bees are of vital importance for tomato pollination, although honey bees are less attracted to tomato flowers than bumble bees. Little is known about how tomato flower volatile compounds influence the foraging behaviors of honey bees and bumble bees. In this study, compounds of tomato flower volatiles were detected by gas chromatography–mass spectrometry. Electroantennography (EAG) and a dynamic two-choice olfactometer were used, respectively, to compare the differences of antennal and behavioral responses between Apis mellifera and Bombus terrestris towards selected volatile compounds. A total of 46 compounds were detected from the tomato flower volatiles. Of the 16 compounds tested, A. mellifera showed strong antennal responses to 3 compounds (1-nonanal, (+)-dihydrocarvone, and toluene) when compared with a mineral oil control, and B. terrestris showed 7 pronounced EAG responses (1,3-xylene, (+)-dihydrocarvone, toluene, piperitone, eucarvone, 1-nonanal, and β-ocimene). Additionally, 1-nonanal and (+)-dihydrocarvone elicited significant avoidance behavior of A. mellifera, but not of B. terrestris. In conclusion, bumble bees are more sensitive to the compounds of tomato flower volatiles compared to honey bees, and honey bees showed aversion to some compounds of tomato flower volatiles. The findings indicated that compounds of flower volatiles significantly influenced bee foraging preference for tomato.

The interactions between plants and pollinators are complex. Flower volatiles as special olfactory cues could influence the foraging choices of pollinators. Here, we conducted bioassays to evaluate the role of flower volatiles on the attraction of honey bees (native Apis cerana and exotic Apis mellifera) to pears (native Pyrus bretschneideri and exotic Pyrus communis). Chemical and electrophysiological approaches were used to determine flower volatiles and evaluate the antennal responses of honey bees to volatiles from pear flowers. Bioassays demonstrated that flower volatiles were crucial for the attraction of honey bees to pear flowers; honey bees preferred to forage on P. communis flowers (p > 0.05), with approximately 64.37 ± 0.02% (A. mellifera) and 62.10 ± 0.02% (A. cerana) foraging on P. communis. Flowers of P. communis and P. bretschneideri yielded 27 and 31 compounds, respectively, with 17 of them being common. Honey bee antennae responded to 16 chemicals, including 5 contained in both pear species: 1-nonanol, linalool, methyl 2-hydroxy-3-methylpentanoate, methyl L-isoleucinate, and α-farnesene. In addition, there were 8 electrophysiologically active compounds in P. bretschneideri: methyl L-valine ester, benzaldehyde, 6-methyl-5-hepten-2-one, isophorone, 2-methyl octane, longicyclene, longifolene, and caryophyllene; and 3 electrophysiologically active compounds in P. communis: β-ocimene, 4-oxoisophorone and lilac alcohol D. In conclusion, our study demonstrated the significant impact of pear flower volatiles on honey bee foraging choices. This knowledge provides a basis for the selection of honey bees for pear pollination and lays a foundation for further study of the chemical communication of pear attractiveness to honey bees.

The decreasing number of bees is a global ecological problem. With the advancement of agricultural modernisation, the large-scale use of neonicotinoid insecticides is one of the main factors leading to the decline of bees. The aim of the present study was to investigate the effect and the mechanisms underlying bees impaired by dinotefuran. Acute (48 h) oral toxicity tests showed that a 5% lethal concentration (LC5) was 0.220 mg/L, and a 20% lethal concentration (LC20) was 0.458 mg/L. The gene expression profile shows that when compared with the control group, the LC5 group induced 206 significantly upregulated, differentially expressed genes (DEGs) and 363 significantly downregulated DEGs, while the LC20 group induced 180 significantly upregulated DEGs and 419 significantly downregulated DEGs. Significantly, transcriptomic analysis revealed DEGs involved in immunity, detoxification, and the nervous system, such as antimicrobial peptides, vitellogenin, synaptotagmin-10, AChE-2, and nAChRa9. Furthermore, Gene Ontology (GO) annotation and Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway analysis revealed that DEGs were enriched in amino acid and fatty acid biosynthesis and metabolism pathways. Collectively, our findings will help clarify the deleterious physiological and behavioural impacts of dinotefuran on bees and provide a basis for future research on the mechanisms underlying bees impaired by dinotefuran.

With the availability of various plants in bloom simultaneously, honey bees prefer to collect some pollen types over others. To better understand pollen’s role as a reward for workers, we compared the digestibility and nutritional value of two pollen diets, namely, pear (Pyrus bretschneideri Rehd.) and apricot (Armeniaca sibirica L.). We investigated the visits, pollen consumption, and pollen extraction efficiency of caged Apis mellifera workers. Newly emerged workers were reared, and the effects of two pollen diets on their physiological status (the development of hypopharyngeal glands and ovaries) were compared. The choice-test experiments indicated a significant preference of A. mellifera workers for apricot pollen diets over pear pollen diets (number of bees landing, 29.5 ± 8.11 and 9.25 ± 5.10, p < 0.001 and pollen consumption, 0.052 ± 0.026 g/day and 0.033 ± 0.013 g/day, p < 0.05). Both pollen diets had comparable extraction efficiencies (67.63% for pear pollen and 67.73% for apricot pollen). Caged workers fed different pollen diets also exhibited similar ovarian development (p > 0.05). However, workers fed apricot pollen had significantly larger hypopharyngeal glands than those fed pear pollen (p < 0.001). Our results indicated that the benefits conferred to honey bees by different pollen diets may influence their foraging preference.

The ambient temperature and relative humidity affect the metabolic and physiological responses of bees, thus affecting their life activities. However, the physiological changes in bee due to high temperature and high humidity remain poorly understood. In this study, we explored the effects of higher temperature and humidity on the epiphysiology of bees by evaluating the survival, tolerance and body water loss in two bee species (Apis cerana and Apis mellifera). We also evaluated the changes in the activity of antioxidant and detoxification enzymes in their body. We observed that under higher temperature and humidity conditions, the survival rate of A. mellifera was higher than that of A. cerana. On the other hand, a comparison of water loss between the two species revealed that A. mellifera lost more water. However, under extremely high temperature conditions, A. cerana was more tolerant than A. mellifera. Moreover, under higher temperature and humidity conditions, the activity of antioxidant and detoxification enzymes in bees was significantly increased. Overall, these results suggest that high temperatures can adversely affect bees. They not only affect the survival and water loss, but also stimulate oxidative stress in bees. However, unlike our previous understanding, high humidity can also adversely affect bees, although its effects are lower than that of temperature.

Apis cerana cerana relies on its sensitive olfactory system to perform foraging activities in the surrounding environment. Olfactory receptors (ORs) are a primary requirement for odorant recognition and coding. However, the molecular recognition of volatile compounds with ORs in A. cerana cerana is still not clear. Hence, in the present study, we achieved transient transfection and cell surface expression of A. cerana cerana ORs (AcerOr1 and AcerOr2; AcerOr2 is orthologous to the co-receptor) in Spodoptera frugiperda (Sf9) cells. AcerOr2 narrowly responded to N-(4-ethylphenyl)-2-((4-ethyl-5-(3-pyridinyl)-4H-1,2,4-triazol-3-yl) thio) acetamide (VUAA1), whereas AcerOr1 was sensitive to eugenol, lauric acid, ocimene, 1-nonanol, linolenic acid, hexyl acetate, undecanoic acid, 1-octyl alcohol, and nerol. Of the compounds tested, AcerOr1 showed the highest sensitivity to these odorants with EC50 values of 10−7 and 10−8 M, and AcerOr2 recognized VUAA1 with higher sensitivity [EC50 = (6.621 ± 0.26) × 10−8]. These results indicate that AcerOr2 is an essential gene for olfactory signaling, and AcerOr1 is a broadly tuned receptor. We discovered ligands that were useful for probing receptor activity during odor stimulation and validated three of them by electroantennography. The response increased with concentration of the odorant. The present study provides insight into the mechanism of olfactory discrimination in A. cerana cerana.

Mutillid wasps are ectoparasitic insects that parasitize the enclosed developmental stages of their hosts. Adults are sexually dimorphic, with brilliantly colored and hardened cuticles. The biology of parasitic mutillid wasps has rarely been addressed. Here, we investigated the parasitization by Mutilla europaea on an important pollinator, Bombus breviceps. The parasitic biology and dispersal ability of M. europaea were observed and tested under experimental conditions. We provide the first record of M. europaea parasitizing B. breviceps in southwestern China. As is the case with other bumblebee species, M. europaea mainly parasitized the puparia of males. The dispersal and invasion ability of this parasite under experimental conditions indicates that it spreads rapidly, as far as 20 m in one week, and invades different hosts (B. breviceps and Bombus haemorrhoidalis). This report not only clarifies the parasitic relationship between M. europaea and B. breviceps, but also has important ecological implications for the conservation of bumblebees in China.

The biological activity and geographical distribution of honey bees is strongly temperature-dependent, due to their ectothermic physiology. In China, the endemic Apis cerana cerana exhibits stronger cold hardiness than Western honey bees, making the former species important pollinators of winter-flowering plants. Although studies have examined behavioral and physiological mechanisms underlying cold resistance in bees, data are scarce regarding the exact molecular mechanisms. Here, we investigated gene expression in A. c. cerana under two temperature treatments, using transcriptomic analysis to identify differentially expressed genes (DEGs) and relevant biological processes, respectively. Across the temperature treatments, 501 DEGs were identified. A gene ontology analysis showed that DEGs were enriched in pathways related to sugar and amino acid biosynthesis and metabolism, as well as calcium ion channel activity. Additionally, heat shock proteins, zinc finger proteins, and serine/threonine-protein kinases were differentially expressed between the two treatments. The results of this study provide a general digital expression profile of thermoregulation genes responding to cold hardiness in A. c. cerana. Our data should prove valuable for future research on cold tolerance mechanisms in insects, and may be beneficial in breeding efforts to improve bee hardiness.

[This corrects the article DOI: 10.1371/journal.pone.0179922.].

Chemosensory genes play a central role in sensing chemical signals and guiding insect behavior. The Chinese honeybee, Apis cerana cerana, is one of the most important insect species in China in terms of resource production, and providing high-quality products for human consumption, and also serves as an important pollinator. Communication and foraging behavior of worker bees is likely linked to a complex chemosensory system. Here, we used transcriptome sequencing on adult A. c. cerana workers of different ages to identify the major chemosensory gene families and the differentially expressed genes(DEGs), and to investigate their expression profiles. A total of 109 candidate chemosensory genes in five gene families were identified from the antennal transcriptome assemblies, including 17 OBPs, 6 CSPs, 74 ORs, 10 IRs, and 2SNMPs, in which nineteen DEGs were screened and their expression values at different developmental stages were determined in silico. No chemosensory transcript was specific to a certain developmental period. Thirteen DEGs were upregulated and 6were downregulated. We created extensive expression profiles in six major body tissues using qRT-PCR and found that most DEGs were exclusively or primarily expressed in antennae. Others were abundantly expressed in the other tissues, such as head, thorax, abdomen, legs, and wings. Interestingly, when a DEG was highly expressed in the thorax, it also had a high level of expression in legs, but showed a lowlevel in antennae. This study explored five chemoreceptor superfamily genes using RNA-Seq coupled with extensive expression profiling of DEGs. Our results provide new insights into the molecular mechanism of odorant detection in the Asian honeybee and also serve as an extensive novel resource for comparing and investigating olfactory functionality in hymenopterans.

The greater wax moth, Galleria mellonella (Linnaeus, 1758), is a notorious pest of honey bee colonies that has negatively affected the global apicultural industry. Olfactory cues influence the behavior of wax moth, where males attract females, making them an ideal candidate for pheromone studies. However, the molecular mechanism of chemoreception in G. mellonella pertaining to sex pheromone recognition has not been elucidated. In this study, transcriptome sequencing was conducted on the antennae of male and female greater wax moths to assess the differential expression patterns of chemosensory genes and better understand the underlying olfactory mechanism. In the results, a total of 121 chemosensory gene transcripts were identified, including 37 odorant-binding proteins, 35 chemosensory proteins, 33 olfactory receptors, 14 ionotropic receptors and 2 sensory neuron membrane proteins. The expression patterns of these genes were determined using the estimated fragments per kilobase of transcript per million fragments mapped. Among the 114 DEGs, 66 were expressed exclusively in the female antennae, whereas the remaining were expressed predominantly in the male antennae. Additionally, five chemosensory-related genes (OBP69alike, OBP72-like, CSP7, CSP10 and OR29) were differentially expressed between the two samples. In conclusion, the study lay a foundation for understanding the olfactory functions of chemosensory genes in G. mellonella, which can help to control and prevent the damage caused by this pest. [ABSTRACT FROM AUTHOR]

In social insects, the foraging gene (for) regulates insect age- and task-based foraging behaviors. We studied the expression and localization of the for gene (Acfor) in Apis cerana cerana workers to explore whether the differential regulation of this gene is associated with the behaviors of nurses and foragers. The expression profiles of Acfor in different tissues and at different ages were examined using real-time quantitative reverse transcription polymerase chain reaction. Cellular localization in the brain was detected using in situ hybridization. Acfor transcripts in different ages workers showed that Acfor expression was detected in all the heads of 1- to 30-d-old worker bees. Acfor expression reached a peak at 25 d of age, and then declined with increasing age. The results showed that Acfor gene expression in five tissues was respectively significantly higher in foragers than in nurses. In nurses, the relative expression of Acfor was the highest in the antennae. There was a highly significant difference in expression between antennae, legs, and the other three tissues. In foragers, Acfor expression was the highest in the thorax, which was significantly different from all other tissues. In situ hybridization showed that Acfor was highly expressed in the lamina of the optic lobes, in a central column of Kenyon cells in the mushroom bodies of the brain of workers, and in the antennal lobes. This suggested that Acfor expression affects age-related foraging behavior in Apis cerana cerana, and that it may be related to flight activity. [ABSTRACT FROM AUTHOR]

Antifreeze proteins (AFPs) are biological cryoprotectants with unique properties that play a crucial role in regulating the molecular mechanisms governing cold resistance in insects. To identify and characterize Apis cerana cerana AFP (AcerAFP), we cloned the full-length cDNA of AcerAFP and examined its expression patterns in A. cerana cerana. A nucleotide alignment analysis showed that the entire coding region of AcerAFP is 1095 bp and encodes a polypeptide of 365 amino acids. The amino acid sequence of this protein exhibits 63-96% homology with AFP homologs from other hymenopterans. α-helices form the main secondary and tertiary structures of AcerAFP, which is similar to the molecular structure of fish AFP type-I. The expression profiles of AcerAFP revealed that expression was tissue, sex, and developmentally specific. In response to cold stress, the mRNA and protein expression of AcerAFP were both induced by low temperatures, and were also related to the concentrations of several cryoprotectants, including glucose, glycerin, glutamic acid, cysteine, histidine, alanine, and methionine. In addition, we found that the knockdown of AcerAFP by RNA interference remarkably increased the total freezing temperature of hemolymph in A. cerana cerana, where levels of AcerAFP mRNA were correlated with the expression of most antifreeze-related proteins. Taken together, these results suggest that AcerAFP plays an essential role as a biological cryoprotectant in honeybees, and is in turn regulated by small cryoprotectants and antifreeze-related proteins. [ABSTRACT FROM AUTHOR]

Insects use olfaction to recognize a wide range of volatile cues, to locate food sources, mates, hosts and oviposition sites. These chemical volatiles are perceived by odorant receptors (ORs) expressed on the dendritic membrane of olfactory neurons, most of which are housed within the chemosensilla of antennae. Most insect ORs are tandemly arrayed on chromosomes and some of them are formed by gene duplication. Here, we identified a pair of duplicated Or genes, AcerOr1 and AcerOr3, from the antennae of the Asian honeybee, Apis cerana cerana, and reported their molecular characterization and temporal expression profiles. The results showed that these two genes shared high similarity both in sequence and the gene structure. Quantitative real-time PCR analysis of temporal expression pattern indicated that in drones the expression pattern of these two genes were very similar. The transcripts expressed weakly in larvae and pupae, then increased gradually in adults. In workers, the expression level of AcerOr1 changed more drastically and expressed higher than that of AcerOr3. However, both reached their highest expression level in one-day-old adults. In addition, the expression profiles between different sexes revealed that AcerOr3 appear to be expressed biased in male antennae. These results suggest that AcerOr1 may perceive odours of floral scents, while AcerOr3 may detect odours critical to male behaviour, such as the queen substance cues. [ABSTRACT FROM AUTHOR]