Project description:Apis mellifera and Apis cerana are two sibling species of Apidae. Apis cerana is adept at collecting sporadic nectar in mountain and forest region and exhibits stiffer hardiness and acarid resistance as a result of natural selection, whereas Apis mellifera has the advantage of producing royal jelly. To identify differentially expressed genes (DEGs) that affect the development of hypopharyngeal gland (HG) and/or the secretion of royal jelly between these two honeybee species, we performed a digital gene expression (DGE) analysis of the HGs of these two species at three developmental stages (newly emerged worker, nurse and forager).Twelve DGE-tag libraries were constructed and sequenced using the total RNA extracted from the HGs of newly emerged workers, nurses, and foragers of Apis mellifera and Apis cerana. Finally, a total of 1482 genes in Apis mellifera and 1313 in Apis cerana were found to exhibit an expression difference among the three developmental stages. A total of 1417 DEGs were identified between these two species. Of these, 623, 1072, and 462 genes showed an expression difference at the newly emerged worker, nurse, and forager stages, respectively. The nurse stage exhibited the highest number of DEGs between these two species and most of these were found to be up-regulated in Apis mellifera. These results suggest that the higher yield of royal jelly in Apis mellifera may be due to the higher expression level of these DEGs.In this study, we investigated the DEGs between the HGs of two sibling honeybee species (Apis mellifera and Apis cerana). Our results indicated that the gene expression difference was associated with the difference in the royal jelly yield between these two species. These results provide an important clue for clarifying the mechanisms underlying hypopharyngeal gland development and the production of royal jelly.
Project description:Various environmental stresses, such as heat shock, heavy metals, ultraviolet (UV) radiation and different pesticides, induce a cellular oxidative stress response. The cellular oxidative stress response is usually regulated by heat shock proteins (Hsps) acting as molecular chaperones. Stress-induced phosphoprotein 1 (STIP1), one of the most widely studied co-chaperones, functions as an adaptor that directs Hsp90 to Hsp70-client protein complexes. However, the biological functions of STIP1 remain poorly understood in honeybee (Apis cerana cerana). In this study, AccSTIP1 was identified in Apis cerana cerana. AccSTIP1 transcription was found to be induced by heat (42 °C), HgCl2, H2O2 and different pesticides (emamectin benzoate, thiamethoxam, hexythiazox and paraquat) and inhibited by CdCl2, UV and kresoxim-methyl. Moreover, western blot analysis indicated that the expression profiles of AccSTIP1 were consistent with its transcriptional expression levels. The disc diffusion assay showed that chemically competent transetta (DE3) bacteria expressing a recombinant AccSTIP1 protein displayed the smaller death zones than did control bacteria after exposure to paraquat and HgCl2. The DNA nicking assay suggested that recombinant purified AccSTIP1 protected supercoiled pUC19 plasmid DNA from damage caused by a thiol-dependent mixed-function oxidation (MFO) system. After knocking down AccSTIP1 gene expression via RNA interference (RNAi), the transcript levels of antioxidation-related genes were obviously lower in dsAccSTIP1 honeybees compared with those in the uninjected honeybees. Collectively, these results demonstrated that AccSTIP1 plays an important role in counteracting oxidative stress. This study lays a foundation for revealing the mechanism of AccSTIP1 in the Apis cerana cerana antioxidant system.
Project description:Varroa destructor is an ectoparasitic mite of Asian or Eastern honeybees Apis cerana (A. cerana) which has become a serious threat to European subspecies of Western honeybees Apis mellifera (A. mellifera) within the last century. V. destructor and its vectored honeybee viruses became serious threats for colony survival. This is a short period for pathogen- and host-populations to adapt. To look for possible variation in the composition of viral populations we performed RNA metagenomic analysis of the Western honeybee subspecies A. m. ligustica, A. m. syriaca, A. m. intermissa, and A. cerana and their respective V. destructor mites. The analysis revealed two novel viruses: Varroa orthomyxovirus-1 (VOV-1) in A. mellifera and V. destructor and a Hubei like-virga virus-14 homolog in V. destructor. VOV-1 was more prevalent in V. destructor than in A. mellifera and we found evidence for viral replication in both hosts. Interestingly, we found differences in viral loads of A. cerana and their V. destructor, A. m. intermissa, and its V. destructor showed partial similarity, while A. m. ligustica and A. m. syriaca and their varroa where very similar. Deformed wing virus exhibited 82.20%, 99.20%, 97.90%, and 0.76% of total viral reads in A. m. ligustica, A. m. syriaca, A. m. intermissa, and A. cerana, respectively. This is the first report of a complete segmented-single-stranded negative-sense RNA virus genome in honeybees and V. destructor mites.
Project description:BACKGROUND: The Eastern hive honey bee, Apis cerana cerana is a native and widely bred honey bee species in China. Molecular biology research about this honey bee species is scarce, and genomic information for A. c. cerana is not currently available. Transcriptome and expression profiling data for this species are therefore important resources needed to better understand the biological mechanisms of A. c. cerana. In this study, we obtained the transcriptome information of A. c. cerana by RNA-sequencing and compared gene expression differences between queens and workers of A. c. cerana by digital gene expression (DGE) analysis. RESULTS: Using high-throughput Illumina RNA sequencing we obtained 51,581,510 clean reads corresponding to 4.64 Gb total nucleotides from a single run. These reads were assembled into 46,999 unigenes with a mean length of 676 bp. Based on a sequence similarity search against the five public databases (NR, Swissport, GO, COG, KEGG) with a cut-off E-value of 10(-5) using BLASTX, a total of 24,630 unigenes were annotated with gene descriptions, gene ontology terms, or metabolic pathways. Using these transcriptome data as references we analyzed the gene expression differences between the queens and workers of A. c. cerana using a tag-based digital gene expression method. We obtained 5.96 and 5.66 million clean tags from the queen and worker samples, respectively. A total of 414 genes were differentially expressed between them, with 189 up-regulated and 225 down-regulated in queens. CONCLUSIONS: Our transcriptome data provide a comprehensive sequence resource for future A. c. cerana study, establishing an important public information platform for functional genomic studies in A. c. cerana. Furthermore, the DGE data provide comprehensive gene expression information for the queens and workers, which will facilitate our understanding of the molecular mechanisms of the different physiological aspects of the two castes.
Project description:Nosema ceranae is a widespread fungal pathogen of honeybees, which is infective to all castes in the colony, including queens, drones and workers. Nosemosis caused by N. ceranae poses a big challenge for apiculture all over the world. Here, midguts of normal and N. ceranae-infected Apis cerana cerana workers at 7 and 10 days post infection were sequenced utilizing small RNA sequencing (sRNA-seq) technology. Totally, more than 150.54 Mb raw reads were produced in this article, and over 144.26 Mb high-quality clean reads with a mean ratio of 95.83% were obtained after strict filtering and quality control. For more insight please see "Comparative identification of microRNAs in Apis cerana cerana workers' midguts responding to Nosema ceranae invasion" (Chen et al., 2019). Raw data are available in NCBI Sequence Read Archive (SRA) database under the BioProject number PRJNA487111. Our data can be used for investigating differentially expressed microRNAs (miRNAs) and piRNAs and their regulatory roles engaged in A. c. cerana response to N. ceranae infection, and for offering potential candidates for uncovering the molecular mechanisms regulating eastern honeybee-microsporidian interactions.
Project description:Cytochrome P450 monooxygenases (P450s) are widely distributed multifunctional enzymes that play crucial roles in insecticide detoxification or activation. In this study, to ascertain the molecular mechanisms of P450s in the detoxification of Chinese honeybees, Apis cerana cerana Fabricius (A. c. cerana), we isolated and characterized four new P450 genes (Acc301A1, Acc303A1, Acc306A1, and Acc315A1). The open reading frames of the four genes are 1263 to 1608 bp in length and encode four predicted polypeptides of 499 to 517 amino acids in length. Real-time quantitative PCR (RT-qPCR) results showed that expression of all four genes was observed in all developmental stages. In addition, Western blot assays further indicated the RT-qPCR results that showed that the four genes were induced by pesticide (thiamethoxam, deltamethrin, dichlorovos, and paraquat) treatments. Furthermore, we also used double-stranded RNA-mediated RNA interference to investigate the functions of Acc301A1, Acc303A1,and Acc306A1 in the antioxidant defense of honeybees. RNA interference targeting Acc301A1, Acc303A1, and Acc306A1 significantly increased the mortality rate of A. c. cerana upon pesticide treatment. These results provide important evidence about the role of the four P450 genes involved in detoxification.
Project description:DNA methylation is an epigenetic modification primarily responsible for individual phenotypic variation. This modification has been reported to play an important role in caste, brain plasticity, and body development in honeybees (Apis mellifera). Here, we report the DNA methylation profile of honeybee hypopharyngeal glands, from atrophy in winter to arousal in the following spring, through the use of whole-genome bisulfite sequencing. Consistent with previous studies in other Apis species, we found low methylation levels of the hypopharyngeal gland genome that were mostly of the CG type. Notably, we observed a strong preference for CpG methylation, which was localized in promoters and exon regions. This result further indicated that, in honeybees, DNA methylation may regulate gene expression by mediating alternative splicing, in addition to silencing gene in the promoter regions. After assessment by correlation analysis, we identified seven candidate proteins encoded by differentially methylated genes, including aristaless-related homeobox, forkhead box protein O, headcase, alpha-amylase, neural-cadherin, epidermal growth factor receptor, and aquaporin, which are reported to be involved in cell growth, proliferation, and differentiation. Hypomethylation followed by upregulated expression of these candidates suggested that DNA methylation may play significant roles in the activation of hypopharyngeal glands in overwintering honeybees. Overall, this study elucidates epigenetic modification differences in honeybee hypopharyngeal glands by comparing an inactive winter state to an aroused state in the following spring, which could provide further insight into the evolution of insect sociality and regulatory plasticity.
Project description:Mitogen-activated protein kinase kinases (MKKs) are important components of the MAPK signaling pathways, which play a key role in responding to stress and inflammatory stimuli. Here, a new MKK gene, AccMKK6, was identified and functionally analyzed in Apis cerana cerana Real-time quantitative PCR (qPCR) and Western blot analysis demonstrated that the AccMKK6 expression level was up-regulated by several environmental stresses. Moreover, the knockdown of AccMKK6 by RNA interference technology altered the expression levels of some antioxidant genes. In addition, the knockdown of AccMKK6 resulted in increased malonyldialdehyde (MDA) concentration and decreased antioxidant-related enzymes activity in honeybees. To explore the MAPK signaling pathways involved in AccMKK6, we identified the transcription factor kayak in A. cerana cerana We analyzed the interactions of AccMKK6, Accp38b, and Acckayak using the yeast two-hybrid system. AccMKK6 and Acckayak showed similar expression profiles after several stress treatments. In addition, the expression level of Acckayak was significantly increased when AccMKK6 was silenced. Therefore, we speculate that AccMKK6 may be involved in the MAPK cascades, which play a crucial role in counteracting oxidative stress caused by external stimuli.