Abstract: 【Aim】The purpose of this study is to detect the expression patterns of ass-milR0037-3p and its key target genes in worker larvae of Apis cerana cerana and A. mellifera ligustica in the process of Ascosphaera apis infection, so as to offer a foundation for further exploring the mechanism of ass-milR0037-3p regulating the As. apis infection. 【Methods】 The target genes of ass-milR0037-3p of As. apis were predicted using related software and then annotated to GO and KEGG databases. Stem-loop RT-PCR and RT-qPCR were employed to verify the expression of ass-milR0037-3p in the guts of the 6-day-old worker larvae of Ap. c. cerana and Ap. m. ligustica and to detect the expression levels of ass-milR0037-3p and its two key target genes (genomic protein acetyltransferase gene ESA1 and flavin containing amine oxidase gene FAO) in the guts of the 4-6-day-old worker larvae after feeding the 3-day-old worker larvae of Ap. c. cerana and Ap. m. ligustica with diets containing 1×10⁷ spores/mL As. apis, respectively.【Results】 ass-milR0037-3p could target 225 genes annotated to 28 GO terms such as reproduction, binding and cells, as well as 105 KEGG pathways such as splicing, biosynthesis of secondary metabolites, and amino sugar and nucleotide sugar metabolism. The target fragments of ass-milR0037-3p were amplified in the guts of the 6-day-old worker larvae of both Ap. c. cerana and Ap. m. ligustica infected with As. apis. The expression levels of ass-milR0037-3p in the guts of the 5- and 6-day-old worker larvae of Ap. c. cerana infected with As. apis were significantly up-regulated and the expression levels of ESA1 and FAO in the guts of the 5- and 6-day-old worker larvae of Ap. c. cerana infected with As. apis were significantly down-regulated, as compared with those of the 4-day-old worker larvae of Ap. c. cerana infected with As. apis. The expression level of ass-milR0037-3p in the gut of the 5-day-old worker larvae of Ap. m. ligustica infected with As. apis was up-regulated as compared with that of the 4-day-old worker larvae of Ap. m. ligustica infected with As. apis and that of the 6-day-old worker larvae of Ap. c. cerana infected with As. apis was significantly up-regulated as compared with that of the 4-day-old worker larvae of Ap. m. ligustica infected with As. apis. The expression levels of the target genes ESA1 and FAO in the gut of the 5-day-old worker larvae of Ap. m. ligustica infected with As. apis were up-regulated and those of the 6-day-old worker larvae of Ap. m. ligustica infected with As. apis were significantly up-regulated as compared with those of the 4-day-old worker larvae of Ap. m. ligustica infected with As. apis.【Conclusion】 ass-milR0037-3p potentially modulates the process of As. apis infecting worker larvae of Ap. c. cerana by negatively regulating the FAO expression, while potentially affects the process of As. apis infecting worker larvae of Ap. m. ligustica through positively regulating the ESA1 expression. The results provide a basis for elucidating the mechanism by which milRNAs respond to the As. apis infection of honeybee larval guts via regulating the expression of target genes.

Key words: Apis cerana cerana, Apis mellifera ligustica, microRNA, microRNA-like RNA, histone acetyltransferase gene, flavin-containing amine oxidase gene