球囊菌胁迫中华蜜蜂幼虫肠道过程中病原的转录组学研究

Transcriptome analysis of Ascosphaera apis stressing larval gut of Apis cerana cerana

摘要:

[目的] 本研究利用RNA-seq技术对球囊菌胁迫的中华蜜蜂（中蜂）幼虫肠道进行深度测序，经趋势分析得到差异表达基因（DEGs）的显著表达模式，进而对胁迫过程中的球囊菌进行转录组学分析。[方法] 利用Illumina HiSeq 2500平台对球囊菌胁迫的中蜂幼虫肠道进行深度测序，并利用相关软件进行了深入分析。最后，通过RT-qPCR对RNA-seq数据进行了验证。[结果] 本研究共得到球囊菌的41133932条高质量clean reads。22865个DEGs共聚类为8个基因表达模式，其中，16769个DEGs聚类为2个显著上调趋势与2个显著下调趋势。GO富集分析结果显示，显著上调与显著下调趋势中的DEGs分别富集于40与37个GO term，基因富集数最多的为细胞进程（2486 unigenes）。KEGG代谢通路（pathway）富集分析结果显示，显著上调与显著下调趋势中的DEGs分别富集于119和112个pathway，基因富集数最多的分别是氨基酸生物合成（127 unigenes）与核糖体（98 unigenes）。进一步分析表明球囊菌在胁迫中蜂幼虫肠道的过程中通过提高物质合成促进其增殖，而宿主通过抑制球囊菌的蛋白合成抵御病原入侵。富集在MAPK信号通路的11个DEGs的表达水平随着胁迫时间的延长而逐渐下降，推测中蜂幼虫通过抑制该通路而阻遏球囊菌增殖。[结论] 本研究不仅为揭示白垩病过程中的球囊菌-中蜂幼虫互作提供了重要信息，也为阐明不同抗性蜂种的球囊菌抗性差异奠定了基础。

Abstract:

[Objective] RNA-seq technology was used to sequence the larval guts of Apis cerana cerana under stress of Ascosphaera apis. Subsequently, trend was analyzed for differentially expressed genes (DEGs) to obtain significant gene expression patterns, followed by transcriptome analysis of A. apis stressing the larval gut.[Methods] Infected honeybee larval guts were sequenced at Illumina HiSeq 2500 platform and in-depth analyses were done using corresponding biological software. Finally, RT-qPCR was conducted to validate RNA-seq data.[Results] A total of 41133932 high-quality clean reads were obtained. Trend analysis result showed that 22865 DEGs were grouped into 8 gene expression patterns, among them 16769 DEGs were assigned to 4 significant expression patterns including 2 up-regulated trends and 2 down-regulated trends. GO enrichment analysis result showed that all DEGs within significant up-and down-regulated patterns were enriched in 40 and 37 GO terms, respectively, and the mostly enriched one is cellular process (2486 unigenes). KEGG enrichment analysis result displayed that the DEGs within significant up-and down-regulated trends were enriched in 119 and 112 pathways, respectively, and biosynthesis of amino acids (127 unigenes) and ribosome (98 unigenes) were mostly enriched. A. apis facilitated its proliferation through enhancing the biosynthesis and the host could fight A. apis by inhibiting the protein synthesis of the fungal pathogen during the stress process. Furthermore, expression levels of 11 DEGs enriched in the pathogen's MAPK signaling pathway decreased when the stressing time of A. apis was prolonged, suggesting that A. c. cerana larvae could constrain the pathogen's replication by disturbing this pathway.[Conclusion] This is the first report of transcriptome investigation of A. apis infecting A. c. cerana larvae. Our data provide gene expression profiles of A. apis stressing the larval gut of A. c. cerana, as well lay the foundation of unraveling molecular mechanisms regulating the pathogenesis of A. apis.