Paulownia witches' broom (PaWB) caused by a phytoplasma, has caused extensive losses in the yields of paulownia timber and resulted in significant economic losses. However, the molecular mechanisms in Paulownia that underlie the phytoplasma stress are poorly characterized. In this study, we use an Illumina platform to sequence four small RNA libraries and four degradome sequencing libraries derived from healthy, PaWB-infected, and PaWB-infected 15 mg·L-1 and 30 mg·L-1 methyl methane sulfonate (MMS)-treated plants. In total, 125 conserved and 118 novel microRNAs (miRNAs) were identified and 33 miRNAs responsive to PaWB disease were discovered. Furthermore, 166 target genes for 18 PaWB disease-related miRNAs were obtained, and found to be involved in plant-pathogen interaction and plant hormone signal transduction metabolic pathways. Eleven miRNAs and target genes responsive to PaWB disease were examined by a quantitative real-time PCR approach. Our findings will contribute to studies on miRNAs and their targets in Paulownia, and provide new insights to further understand plant-phytoplasma interactions.