Abstract: 【Objective】So far, no study on application of next-generation sequencing technology for the research of chalkbrood disease was reported. In the present research, RNA-seq technology was utilized to deep sequence of normal and Ascosphaera apis-treated 4th instar Apis cerana cerana larval gut to mine larvae’s responses to A. apis challenge.【Method】AcCK (un-treated group) and AcT (A. apis-treated group) were sequenced on Illumina HiSeq 2500 platform. After evaluation and filtration of raw data from RNA-seq, differentially expressed gene (DEG) analysis was performed using edgeR software, further, Gene Ontology (GO) and KEGG pathway enrichment analyses were carried out, and finally, real-time quantitative PCR (qRT-PCR) was conducted to validate the RNA-seq data.【Result】 In total, RNA-seq produced 188 457 338 raw reads, and after filtration, 182 088 448 clean reads were obtained, Q20 and Q30 of each sample were above 97.96% and 94.97%, respectively, indicating that RNA-seq data in this research were with high quality. principle component analysis (PCA) was performed on all genes level and the result showed PC1 and PC2 were able to account for 75.8% and 10.7% of the expressed genes’ overall differences, respectively. DEG analysis result displayed that there were 344 up-regulated genes and 239 down-regulated genes in AcCK VS AcT. GO enrichment analysis result showed that the DEGs were enriched in 36 GO terms, among them, the mostly enriched ones were cell (106 unigenes), cell part (106 unigenes) and metabolic process (104 unigenes). KEGG pathway enrichment analysis result suggested that up- and down-regulated genes were enriched in 72 and 45 pathways, respectively, and the mostly enriched pathways for up-regulated genes were ribosome (72 unigenes), carbon metabolism (16 unigenes) and glycolysis/gluconeogenesis (14 unigenes), while the mostly enriched pathways for down-regulated genes were carbon metabolism (9 unigenes), glyoxylate and dicarboxylate metabolism (8 unigenes) and amino acids biosynthesis (7 unigenes). further analysis demonstrated that the immune-related genes in A. c. cerana larval gut were activated, while the metabolism-related genes were greatly inhibited.【Conclusion】The findings of the study not only uncovered the A. c. cerana larval gut’s responses to A. apis during the early stage of invasion, but also provided key information for clarifying the mechanism underlying the host’s responses to A. apis, thus laying a foundation for functional investigation of key responding genes.

Key words: Apis cerana cerana, larval gut, Ascosphaera apis, transcriptome, RNA-seq