Unraveling the mechanism underlying the immune responses of Apismelliferaligustica to Nosemaceranae stress based on small RNA omics anlyses

• 1、College of Animal Sciences(College of Bee Science),Fujian Agriculture and Forestry University, Fuzhou, 350002

Abstract：Nosema ceranae specifically parasites the cell of adult bee, leading to bee microsporidiosis. In this work, N. ceranae-stressed Apis mellifera ligustica workers\' midguts at 7 d post inoculation (dpi) and 10 dpi (Am7T and Am10T) un-stressed midguts (Am7CK, Am10CK) were deeply sequenced using small RNA-seq (sRNA-seq) technology, a total of 165 895 574 raw reads and 132 028 990 clean tags were yielded, and average Pearson correlation coefficients among different biological replicas in each group were above 87.92%. Here, 928 known miRNAs and 56 novel miRNAs were identified. Majority of these miRNAs were 18 nt and 22 nt in length, and the first base of most miRNAs has a U bias. The expression of five novel miRNAs was verified using Stem-loop RT-PCR. Moreover, 48 up-regulated miRNAs and 36 down-regulated miRNAs were identified in Am7CK vs Am7T comparison group, while 56 up-regulated miRNAs and 51 down-regulated miRNAs were identified in Am10CK vs Am10T. DEmiRNAs in these two comparison groups can target 9 827 and 1 0720 mRNAs, respectively. DEmiRNA-targeted mRNAs were respectively engaged in 50 and 47 functional terms, and 138 and 135 pathways, includingcellular immune pathways such as endocytosis and ubiquitin-mediated proteolysis, and five humoral immune pathways such as MAPK signaling pathway and FoxO signaling pathway. Further, regulatory network analysis showed 26 DEmiRNAs in Am7CK vs Am7T could target 10 DEmRNAs including endocytosis, ubiquitin-mediated proteolysis, phagosome, FoxO signaling pathway and MAPK signaling pathway; whereas 15 DEmiRNAs in Am10CK vs Am10T can target 10 immune-associated pathways. Finally, the reliability of sequencing data and differential expression trend of four DEmiRNA was validated using RT-qPCR. Our results revealed that host DEmiRNAs may response to N. ceranae and participate in immune defense through regulating material and energy metabolisms, apoptosis, pathogen identification, cellular and humoral immunity; however, host DEmiRNAs might not regulate the expression of antimicrobial peptide-encoded genes; miR-1-z was likely to be engaged in cell proliferation, apoptosis and immune processes during host response to N. ceranae stress; oxidative phosphorylation pathway may play a special role in host immune response and interaction between A. m. ligustica and N. ceranae.

Keywords： Apis mellifera ligustica Nosema ceranae microRNA immune response host-pathogen interaction molecular mechanism