Abstract: 【Aim】 To enrich the information of microRNA (miRNA) in Nosema ceranae and to provide theoretical and experimental bases for further exploration of the function of miRNAs in pathogen spore and pathogen infection. 【Methods】 Based on the gained data from small RNA-seq, miRNAs in the clean spores of N. ceranae were identified and analyzed using bioinformatics software. Stem-loop RT-PCR was performed to detect the expression of the identified miRNAs. Sequences of miRNAs were validated by molecular cloning and Sanger sequencing. Target genes of these miRNAs were predicted with TargetFinder software and then annotated to databases. The regulatory network was constructed based on the the targeting relationship between miRNAs and their target genes, followed by visualization with Cytoscape software. 【Results】 In total, 10 miRNAs were identified in N. ceranae spores. The length distribution of these miRNAs ranged from 21 to 25 nt. The first base of these miRNAs showed a U bias, and there was an obvious difference in the bias at each base. The result of stem-loop RT-PCR indicated that the 10 miRNAs were truly expressed, and Sanger sequencing confirmed the reliability of sequences of two miRNAs randomly selected from them. A total of 249 target genes were predicted, among which 249, 118, 136 and 3 target genes could be annotated to Nr, Swiss-Prot, KOG and eggNOG databases, respectively. In addition, 134 and 71 target genes could be respectively annotated to 30 functional terms in GO database and 54 pathways in KEGG database. 【Conclusion】 The findings reveal the presence and expression of miRNAs in N. ceranae spores. These miRNAs may participate in vital activities in spores via regulating the expression of potential target genes.

Key words: Honey bee, host, Nosema ceranae, small RNA sequencing, microRNA, regulation