Recently, the number of collection records of Pemphigus galls from Populus nigra has been increasing in Japan. To identify the galls on P. nigra, mitochondrial COI sequences were analyzed from galling aphid samples collected on P. nigra in Tokyo and Hokkaido. From the BLAST search and neighbor-joining (NJ) analysis, the aphid samples were identified as Pemphigus bursarius, which has not been recorded from Japan. Two samples from Tokyo and Hokkaido showed a genetic difference of 0.30%. This result suggests that different strains of P. bursarius might have been introduced into the Japanese islands at least twice.