Background <p>Patients with multidrug-resistant tuberculosis (MDR-TB) who are resistant to at least both rifampicin and isoniazid, lack effective treatment options in clinic. The gold standard for the diagnosis of MDR-TB is drug sensitivity test, which is time-consuming and has a relatively low positive detection rate. Screening early diagnostic biomarker for MDR-TB is urgent need in clinical practice.</p> Methods <p>A total of 33 patients with MDR-TB, healthy controls and drug-sensitive tuberculosis (DS-TB) were included in this study. Total plasma exosomal RNA was extracted from the subjects, and the MDR-TB plasma-specific exosomal miRNAs were obtained by Illumina sequencing.</p> Results <p>There were 644 and 647 differentially expressed miRNAs in the plasma exosomes of MDR-TB patients obtained by sequencing and biogenic analysis compared with DS-TB patients and healthy controls, respectively. Differential miRNAs are mainly involved in the biological function of regulation of transcription and protein binding, and enriched in the pathways in cancer and MAPK signaling pathway. Moreover, seven plasma exosomal miRNAs in MDR-TB patients were significantly different from those in DS-TB patients and healthy controls. Among them, three of the miRNAs (hsa-miR-122-5p_R-1, hsa-miR-23b-3p_R + 1, and hsa-miR-15a-5p_R-1) were found to be in target relationship with MDR-TB related genes (NTRK2, KIDINS220, NCKAP1, MAPK9, NFAT5, ATF6 and SLC11A2) by target gene prediction analysis. Further the bioinformatic analysis showed that hsa-miR-122-5p_R-1 targets the protein PGLYRP2, a diagnostic biomarker identified in our previous study.</p> Conclusions <p>We suggest that hsa-miR-122-5p_R-1, hsa-miR-23b-3p_R + 1, and hsa-miR-15a-5p_R-1 are closely related to MDR-TB.</p>