Hongfang Wang

Comparison of the nutrient composition of royal jelly and worker jelly of honey bees ( Apis mellifera )

In this study, the chemical and mineral composition and trace elements in royal jelly (RJ) and worker jelly (WJ) and in royal jelly on particular days (only-2-day RJ [O2d], only-3-day RJ [O3d] and only-4-day RJ [O4d]) were determined. Significant differences in levels of moisture, protein, 10-hydroxy-2-decenoic acid (10-HDA), fructose (F) and glucose (G) were found between the RJ and WJ samples. The nutrient content was significantly higher in samples on O2d than the O3d and O4d samples. The results of this study add to the current knowledge of the nutritional value of RJ and WJ. These results also imply a strong relationship between nutritional effects and polyphenism in honey bees.

Functional and mutational analyses of an omega-class glutathione S-transferase (GSTO2) that is required for reducing oxidative damage in Apis cerana cerana

Glutathione S‐transferases perform a variety of vital functions, particularly in reducing oxidative damage. Here, we investigated the expression patterns of Apis cerana cerana omega‐class glutathione S‐transferase 2 (AccGSTO2) under various stresses and explored its connection with antioxidant defences. We found that AccGSTO2 knockdown by RNA interference triggered increased mortality in Ap. cerana cerana, and immunohistochemistry revealed significantly decreased AccGSTO2 expression, particularly in the midgut and fat body. Further analyses indicated that AccGSTO2 knockdown resulted in decreases in catalase and glutathione reductase activities, ascorbate content and the ratio of reduced to oxidized glutathione, and increases in H2O2, malondialdehyde and carbonyl contents. We also analysed the transcripts of other antioxidant genes and found that many genes were down‐regulated in the AccGSTO2 knockdown samples, revealing that AccGSTO2 may be indispensable for attaining a normal lifespan by enhancing cellular oxidative resistance. In addition, the roles of cysteine residues in AccGSTO2 were explored using site‐directed mutagenesis. Mutants of Cys28 and Cys124 significantly affected the enzyme and antioxidant activities of AccGSTO2, which may be attributed to the changes in the spatial structures of mutants as determined by homology modelling. In summary, these observations provide novel insight into the structural and functional characteristics of GSTOs.

A glutathione S-transferase gene associated with antioxidant properties isolated from Apis cerana cerana

Glutathione S-transferases (GSTs) are an important family of multifunctional enzymes in aerobic organisms. They play a crucial role in the detoxification of exogenous compounds, especially insecticides, and protection against oxidative stress. Most previous studies of GSTs in insects have largely focused on their role in insecticide resistance. Here, we isolated a theta class GST gene designated AccGSTT1 from Apis cerana cerana and aimed to explore its antioxidant and antibacterial attributes. Analyses of homology and phylogenetic relationships suggested that the predicted amino acid sequence of AccGSTT1 shares a high level of identity with the other hymenopteran GSTs and that it was conserved during evolution. Quantitative real-time PCR showed that AccGSTT1 is most highly expressed in adult stages and that the expression profile of this gene is significantly altered in response to various abiotic stresses. These results were confirmed using western blot analysis. Additionally, a disc diffusion assay showed that a recombinant AccGSTT1 protein may be roughly capable of inhibiting bacterial growth and that it reduces the resistance of Escherichia coli cells to multiple adverse stresses. Taken together, these data indicate that AccGSTT1 may play an important role in antioxidant processes under adverse stress conditions.

A typical RNA-binding protein gene (AccRBM11) in Apis cerana cerana: characterization of AccRBM11 and its possible involvement in development and stress responses.

RNA-binding motif proteins (RBMs) belong to RNA-binding proteins that display extraordinary posttranscriptional gene regulation roles in various cellular processes, including development, growth, and stress responses. Nevertheless, only a few examples of the roles of RBMs are known in insects, particularly in Apis cerana cerana. In the present study, we characterized the novel RNA-binding motif protein 11 from Apis cerana cerana, which was named AccRBM11 and whose promoter sequence included abundant potential transcription factor binding sites that are connected to responses to adverse stress and early development. Quantitative PCR results suggested that AccRBM11 was expressed at highest levels in 1-day postemergence worker bees. AccRBM11 mRNA and protein levels were higher in the poison gland and the epidermis than in other tissues. Moreover, levels of AccRBM11 transcription were upregulated upon all the simulation of abiotic stresses. Furthermore, Western blot analysis indicated that AccRBM11 protein expression levels could be induced under some abiotic stressors, a result that did not completely in agree with the qRT-PCR results. It is also noteworthy that the expression of some genes that connected with development or stress responses were remarkably suppressed when AccRBM11 was silenced, which suggested that AccRBM11 might play a similar role in development or stress reactions with the above genes. Taken together, the data presented here provide evidence that AccRBM11 is potentially involved in the regulation of development and some abiotic stress responses. We expect that this study will promote future research on the function of RNA-binding proteins.

A novel 1-Cys thioredoxin peroxidase gene in Apis cerana cerana: characterization of AccTpx4 and its role in oxidative stresses

Thioredoxin peroxidase (Tpx), also named peroxiredoxin (Prx), is an important peroxidase that can protect organisms against stressful environments. AccTpx4, a 1-Cys thioredoxin peroxidase gene from the Chinese honey bee Apis cerana cerana, was cloned and characterized. The AccTpx4 gene encodes a protein that is predicted to contain the conserved PVCTTE motif from 1-Cys peroxiredoxin. Quantitative real-time PCR (Q-PCR) and Western blotting revealed that AccTpx4 was induced by various oxidative stresses, such as cold, heat, insecticides, H2O2, and HgCl2. The in vivo peroxidase activity assay showed that recombinant AccTpx4 protein could efficiently degrade H2O2 in the presence of DL-dithiothreitol (DTT). In addition, disc fusion assays revealed that AccTpx4 could function to protect cells against oxidative stresses. These results indicate that AccTpx4 plays an important role in oxidative stress responses and may contribute to the conservation of honeybees.

Identification and Characterization of a Novel Methionine Sulfoxide Reductase B Gene (AccMsrB) from Apis cerana cerana (Hymenoptera: Apidae)

ABSTRACT Methionine sulfoxide reductase B genes (MsrBs) play a crucial role protecting cells from oxidative damage. In this study, we isolated and characterized an MsrB gene from Apis cerana cerana F., designated AccMsrB. The full cDNA of AccMsrB was 757 bp with an open reading frame (ORF) of 414 bp, and the predicted translation product was a 137 amino acid polypeptide with an estimated molecular mass of 15.5 kDa and an isoelectric point of 7.77. Multiple sequence alignment revealed that AccMsrB shares high identity with other known MsrBs (Apis florea F., Apis mellifera L., Apis dorsata F., and Bombus terrestris L.) and contains conservative Cys residues. Quantitative real-time polymerase chain reaction showed AccMsrB to be highly expressed in the epidermis of adult workers, reaching high levels in first-instar larvae, prepupae, and 15-d-old adults. Furthermore, the expression of AccMsrB was upregulated by various oxidative stresses, including 4, 16, 25, and 42°C, ultraviolet light (30mj/cm2), H2O2, CdCl2, HgCl2, paraquat, imidacloprid, and cyhalothrin. However, AccMsrB was downregulated when exposed to phoxime. These results indicate that AccMsrB might respond to various environmental stresses and protect against reactive oxygen species.

Effects of Astragalus membranaceus on growth performance, carcass characteristics, and antioxidant status of broiler chickens

Abstract An experiment was conducted using 720 one-day-old Arbor Acres broiler chickens to evaluate the effects of Astragalus membranaceus root powder (AMP), Astragalus polysaccharides (APS) on growth performance, carcass characteristics, and antioxidant enzymes, and metabolites of blood and liver in broiler chickens. The six dietary treatments included a control that was a corn–soybean meal basal diet; basal diet supplemented with 120 mg/kg antibiotic; 350 mg/kg APS; or 5000, 10,000, or 15,000 mg/kg diet of AMP. Supplementation of 10,000 mg/kg AMP increased average daily gain and feed conversion rate of broilers in grower period (22–42 d) and over the entire experiment compared to basic diet. Responses of antioxidant status to dietary APS and AMP varied depending on the age of the broilers. Beneficial effects of AMP on growth and feed utilization of broilers could be obtained by inclusion of AMP at the rate of 10,000 mg/kg diet.

Characterization of the CDK5 gene in Apis cerana cerana ( AccCDK5 ) and a preliminary identification of its activator gene, AccCDK5r1

Cyclin-dependent kinase 5 (CDK5) is an unusual CDK whose function has been implicated in protecting the central nervous system (CNS) from oxidative damage. However, there have been few studies of CDK5 in insects. In this study, we identified the AccCDK5 gene from Apis cerana cerana and investigated its role in oxidation resistance. We found that AccCDK5 is highly conserved across species and contains conserved features of the CDK5 family. The results of qPCR analysis indicated that AccCDK5 is highly expressed during the larval and pupal stages and in the adult head and muscle. We further observed that AccCDK5 is induced by several environmental oxidative stresses. Moreover, the overexpression of the AccCDK5 protein in E. coli enhances the resistance of the bacteria to oxidative stress. The activation of CDK5 requires binding to its activator. Therefore, we also identified and cloned cyclin-dependent kinase 5 regulatory subunit 1, which we named AccCDK5r1, from Apis cerana cerana. AccCDK5r1 contains a conserved cell localization targeting domain as well as binding and activation sites for CDK5. Yeast two-hybrid analysis demonstrated the interaction between AccCDK5 and AccCDK5r1. The expression patterns of the two genes were similar after stress treatment. Collectively, these results suggest that AccCDK5 plays a pivotal role in the response to oxidative stresses and that AccCDK5r1 is a potential activator of AccCDK5.

Characterization of a Decapentapletic Gene (AccDpp) from Apis cerana cerana and Its Possible Involvement in Development and Response to Oxidative Stress

To tolerate many acute and chronic oxidative stress-producing agents that exist in the environment, organisms have evolved many classes of signal transduction pathways, including the transforming growth factor β (TGFβ) signal pathway. Decapentapletic gene (Dpp) belongs to the TGFβ superfamily, and studies on Dpp have mainly focused on its role in the regulation of development. No study has investigated the response of Dpp to oxidative pressure in any organism, including Apis cerana cerana (A. cerana cerana). In this study, we identified a Dpp gene from A. cerana cerana named AccDpp. The 5΄ flanking region of AccDpp had many transcription factor binding sites that relevant to development and stress response. AccDpp was expressed at all stages of A. cerana cerana, with its highest expression in 15-day worker bees. The mRNA level of AccDpp was higher in the poison gland and midgut than other tissues. Furthermore, the transcription of AccDpp could be repressed by 4°C and UV, but induced by other treatments, according to our qRT-PCR analysis. It is worth noting that the expression level of AccDpp protein was increased after a certain time when A. cerana cerana was subjected to all simulative oxidative stresses, a finding that was not completely consistent with the result from qRT-PCR. It is interesting that recombinant AccDpp restrained the growth of Escherichia coli, a function that might account for the role of the antimicrobial peptides of AccDpp. In conclusion, these results provide evidence that AccDpp might be implicated in the regulation of development and the response of oxidative pressure. The findings may lay a theoretical foundation for further genetic studies of Dpp.

The Wisdom of Honeybee Defenses Against Environmental Stresses

As one of the predominant pollinator, honeybees provide important ecosystem service to crops and wild plants, and generate great economic benefit for humans. Unfortunately, there is clear evidence of recent catastrophic honeybee colony failure in some areas, resulting in markedly negative environmental and economic effects. It has been demonstrated that various environmental stresses, including both abiotic and biotic stresses, functioning singly or synergistically, are the potential drivers of colony collapse. Honeybees can use many defense mechanisms to decrease the damage from environmental stress to some extent. Here, we synthesize and summarize recent advances regarding the effects of environmental stress on honeybees and the wisdom of honeybees to respond to external environmental stress. Furthermore, we provide possible future research directions about the response of honeybees to various form of stressors.