In-depth investigation of microRNA-mediated cross-kingdom regulation between Asian honey bee and microsporidian.

Asian honey bee is the original host for , a unicellular fungal parasite that causes bee nosemosis throughout the world. Currently, interaction between and is largely unknown. Our group previously prepared workers' midguts at 7 days post inoculation (dpi) and 10 dpi with spores as well as corresponding un-inoculated workers' midguts, followed by cDNA library construction and a combination of RNAs-seq and small RNA-seq. Meanwhile, we previously prepared clean spores of , which were then subjected to cDNA library construction and deep sequencing. Here, based on the gained high-quality transcriptome datasets, differentially expressed mRNAs (DEmiRNAs) targeted by host DEmiRNAs, and DEmRNAs targeted by microsporidian DEmiRNAs were deeply investigated, with a focus on targets involved in glycolysis/glyconeogenesis as well as virulence factors, and energy metabolism and immune response. In worker's midguts at 7 (10) dpi (days post inoculation), eight (seven) up-regulated and six (two) down-regulated miRNAs were observed to target 97 (44) down-regulated and 60 (15) up-regulated mRNAs, respectively. Additionally, two up-regulated miRNAs (miR-60-y and miR-676-y) in host midgut at 7 dpi could target genes engaged in spore wall protein and glycolysis/gluconeogenesis, indicating potential host miRNA-mediated regulation of microsporidian virulence factor and energy metabolism. Meanwhile, in at 7 (10) dpi, 121 (110) up-regulated and 112 (104) down-regulated miRNAs were found to, respectively, target 343 (247) down-regulated and 138 (110) down-regulated mRNAs in workers' midguts. These targets in host were relevant to several crucial cellular and humoral immune pathways, such as phagasome, endocytosis, lysosomes, regulation of autophagy, and Jak-STAT signaling pathway, indicative of the involvement of DEmiRNAs in regulating these cellular and humoral immune pathways. In addition, miR-21-x was up-regulated at 7 dpi and had a target relative to oxidative phosphorylation, suggesting that miR-21-x may be used as a weapon to modulate this pivotal energy metabolism pathway. Furthermore, potential targeting relationships between two pairs of host DEmiRNAs-microsporidian DEmRNAs and two pairs of microsporidian DEmiRNAs-host DEmRNAs were validated using RT-qPCR. Our findings not only lay a foundation for exploring the molecular mechanism underlying cross-kingdom regulation between workers and , but also offer valuable insights into Asian honey bee-microsporidian interaction.