MicroRNAs (miRNAs) are a class of noncoding RNA molecules and play important roles in a wide spectrum of biological processes, including in immune response. Recent years have witnessed considerable amount of research interest in studies on miRNA-mediated modulation gene function during neuroinflammation. Here, we evaluated Streptococcus agalactiae infected tilapia (Oreochromis niloticus) brain for the expression profile of miRNAs, potential functions and their correlation with genes involved in inflammatory pathways. A total of 1981 miRNAs were identified, including in 486 miRNAs which have homologues in the currently available databases and 1945 novel miRNAs. The expression levels of 547 miRNAs were significantly altered at 6 h–48 h post-bacterial infection, and these miRNAs were therefore classified as differentially expressed tilapia miRNAs. Real-time PCR were implemented for 14 miRNAs co-expressed in five samples, and agreement was confirmed between the high-throughput sequencing and real-time PCR data. For the 486 differentially expressed miRNAs target 41,820 genes. GO and KEGG enrichment analysis revealed that some target genes of miRNAs were grouped mainly into the categories of apoptotic, signal pathwayand immune response. This is the first report of comprehensive identification of teleost miRNAs being differentially regulated in brain in normal conditions relating to bacterial infection.