Here, the expression profiles and differentially expressed miRNAs (DEmiRNAs) in the midguts of Apis cerana cerana workers at 7 d and 10 d post-inoculation (dpi) with N. ceranae were investigated via small RNA sequencing and bioinformatics. Five hundred and twenty nine (529) known miRNAs and 25 novel miRNAs were identified in this study, and the expression of 16 predicted miRNAs was confirmed by Stem-loop RT-PCR. A total of 14 DEmiRNAs were detected in the midgut at 7 dpi, including eight up-regulated and six down-regulated miRNAs, while 12 DEmiRNAs were observed in the midgut at 10 dpi, including nine up-regulated and three down-regulated ones. Additionally, five DEmiRNAs were shared, while nine and seven DEmiRNAs were specifically expressed in midguts at 7 dpi and 10 dpi. Gene ontology analysis suggested some DEmiRNAs and corresponding target mRNAs were involved in various functions including immune system processes and response to stimulus. KEGG pathway analysis shed light on the potential functions of some DEmiRNAs in regulating target mRNAs engaged in material and energy metabolisms, cellular immunity and the humoral immune system. Further investigation demonstrated a complex regulation network between DEmiRNAs and their target mRNAs, with miR-598-y, miR-252-y, miR-92-x and miR-3654-y at the center. Our results can facilitate future exploration of the regulatory roles of miRNAs in host responses to N. ceranae, and provide potential candidates for further investigation of the molecular mechanisms underlying eastern honeybee-microsporidian interactions.

Apis cerana is the original host for Nosema ceranae, a widespread fungal parasite resulting in honey bee nosemosis, which leads to severe losses to the apiculture industry throughout the world. However, knowledge of N. ceranae infecting eastern honey bees is extremely limited. Currently, the mechanism underlying N. ceranae infection is still largely unknown. Based on our previously gained high-quality transcriptome datasets derived from N. ceranae spores (NcCK group), N. ceranae infecting Apis cerana cerana workers at seven days post inoculation (dpi) and 10 dpi (NcT1 and NcT2 groups), comparative transcriptomic investigation was conducted in this work, with a focus on virulence factor-associated differentially expressed genes (DEGs). Microscopic observation showed that the midguts of A. c. cerana workers were effectively infected after inoculation with clean spores of N. ceranae. In total, 1411, 604, and 38 DEGs were identified from NcCK vs. NcT1, NcCK vs. NcT2, and NcT1 vs. NcT2 comparison groups. Venn analysis showed that 10 upregulated genes and nine downregulated ones were shared by the aforementioned comparison groups. The GO category indicated that these DEGs were involved in a series of functional terms relevant to biological process, cellular component, and molecular function such as metabolic process, cell part, and catalytic activity. Additionally, KEGG pathway analysis suggested that the DEGs were engaged in an array of pathways of great importance such as metabolic pathway, glycolysis, and the biosynthesis of secondary metabolites. Furthermore, expression clustering analysis demonstrated that the majority of genes encoding virulence factors such as ricin B lectins and polar tube proteins displayed apparent upregulation, whereas a few virulence factor-associated genes such as hexokinase gene and 6-phosphofructokinase gene presented downregulation during the fungal infection. Finally, the expression trend of 14 DEGs was confirmed by RT-qPCR, validating the reliability of our transcriptome datasets. These results together demonstrated that an overall alteration of the transcriptome of N. ceranae occurred during the infection of A. c. cerana workers, and most of the virulence factor-related genes were induced to activation to promote the fungal invasion. Our findings not only lay a foundation for clarifying the molecular mechanism underlying N. ceranae infection of eastern honey bee workers and microsporidian–host interaction.

Temperature is a pivotal ecological factor in the regulation of insect survival and reproduction

Honeybees are vital pollinators with functional differentiation as a key survival strategy. The Chinese honeybee (Apis cerana cerana) exhibits exceptional nectar foraging in complex terrains, yet how alternative splicing (AS) and polyadenylation (APA) regulate its labor division remains unclear. Here, we applied PacBio full-length transcriptome sequencing to annotate worker bee transcriptomes across three developmental stages (Ac3d, Ac10d, Ac21d), calibrating the third-generation sequencing data with second-generation sequencing to enhance transcriptome annotation accuracy. We identified 17,961 isoforms and 1922 APA genes, finding that alternative first exon was the major type of AS, while APA enhances transcriptomic diversity via dual polyadenylation sites in most genes. Functional analyses revealed AS enrichment in growth signaling (Vg6, CYP15A1) and immune pathways (PTPRR), whereas APA regulated growth signaling (Vg6), energy metabolism (Acsl1, AcceFE), and oxidative stress (PTPRR, PPO2). Validation by PCR and 3′RACE confirmed stage-specific AS/APA events in key genes. These findings significantly enhance the A. cerana cerana reference genome annotation and provide valuable insights into the mechanisms of AS and APA regulation underlying honeybee development and functional transitions.

Apis cerana cerana is a subspecies of eastern honeybee, Apis cerana , and it plays a vital role in ecological maintenance in China. However, A. c. cerana is threatened by many pathogenic microorganisms including Nosema ceranae , a widespread fungal parasite that infected worldwide colonies. In this article, un-challenged (AcCK1, AcCK2) and N. ceranae -challenged midguts of A. c. cerana workers (AcT1, AcT2) were sequenced utilizing Nanopore long-read sequencing technology. Totally, 11,727,628, 6,996,395, 14,383,735 and 11,580,154 raw reads were yielded from AcCK1, AcCK2, AcT1 and AcT2; the average lengths were 1147 bp, 908 bp, 992 bp and 1077 bp, and the average N50 were 1308 bp, 911 bp, 1079 bp and 1192 bp. The length distribution of was ranged 1 kb to more than 10 kb. Additionally, the quality (Q) score distribution of raw reads was among Q7~Q17. Further, 11,617,144, 6,940,895, 14,277,240 and 11,501,562 clean reads were respectively obtained from AcCK1, AcCK2, AcT1 and AcT2, and among them 78.40%, 82.50%, 79.05% and 80.20% were identified as full-length clean reads. In addition, full-length clean reads from AcCK1, AcT1, AcT2 and AcCK2 were ranged from 1 kb to more than 10 kb in length. Finally, the length distribution of redundant reads-removed full-length transcripts was among 1 kb~5 kb. Value of the data: ♦ This dataset enables better understanding the complexity of A. c. cerana transcriptome. ♦ Current dataset contributes to identification of genes and transcripts engaged in response of eastern honeybee to N. ceranae stress. ♦ The data provides a valuable genetic resource for deciphering alternative splicing and polyadenylation of A. c. cerana mRNAs involved in host response to N. ceranae challenge. ♦ The reported data is beneficial for uncovering the molecular mechanism regulating interaction between eastern honeybee and microsporidian.

Apis cerana cerana , a subspecies of eastern honey, Apis cerana , plays a specific role in beekeeping industry and ecosystem in China and other Asian countries. Larvae of A. c. cerana can be infected by Ascosphaera apis , the fungal pathogen of chalkbrood. In this article, normal 4-, 5-, and 6-day-old larval guts (AcCK1, AcCK2, AcCK3) and A. apis -infected 4-, 5- and 6-day-old larval guts (AcT1, AcT2, AcT3) of A. c. cerana workers were respectively harvested followed by DNA isolation, bisulfite conversion, cDNA library construction and Illumina sequencing. Based on genome-wide bisulfite sequencing, 79167210, 82175052, 79331489, 81051009, 74742842 and 74849091 raw reads were generated from AcCK1, AcCK2, AcCK3, AcT1, AcT2 and AcT3, and after quality control, 73417030 (92.73%), 76660370 (93.27%), 71804727 (90.44%), 75046507 (92.82%), 67487782 (90.30%) and 67367023 (90.04%) clean reads were obtained, respectively. Additionally, 73333333, 76533333, 71466667, 75066667, 67590965 and 67200000 clean reads were mapped to the reference genome of A. cerana , including 54656767, 58583415, 54127407, 57943220, 52547867 and 51295824 unique mapped clean reads, and 8624392, 8789458, 7531333, 7747337, 6249679 and 5394174 multiple mapped clean reads. The genome-wide bisulfite sequencing data reported here can be used for genome-wide identification of 5mC methylation sites in eastern honeybee larval guts and systematic investigation of DNA methylation-mediated host response to A. apis infection. Value of the data: The current dataset contributes to genome-wide identification of 5mC methylation sites in normal and A. apis -infected larval guts of eastern honeybee. The reported data could be used for systematic investigation of DNA methylation-mediated response of eastern honeybee larvae to A. apis infection. Our data offers a valuable genetic resource for better understanding epigenetic regulation mechanism involved in eastern honeybee larvae- A. apis interaction.