Throughout their life cycle, plants are inevitably confronted with various challenges imposed by adverse environmental conditions, including both biotic and abiotic stresses. To adapt to these environmental fluctuations, plants have evolved a highly efficient regulatory mechanism, in which microRNAs (miRNAs) play pivotal roles. miRNAs are a class of 20-24 nucleotide non-coding RNAs generated from MIR genes, which regulate gene expression at the post-transcriptional level through mRNA degradation or translational repression. Over the past decades, accumulating evidence has demonstrated that miRNAs serve as master regulators in plant responses to biotic stresses, such as those caused by bacteria, fungi, oomycetes, viruses, nematodes, and insects. In this review, we summarize recent advances in miRNA biogenesis and highlight the regulatory roles of plant miRNAs in biotic stress tolerance. Additionally, we discuss future directions of miRNA research.