The citrus red mite, Panonychus citri (McGregor) (Acari: Tetranychidae), has a world-wide distribution and is the most important citrus (Citrus spp.) pest in many countries, including Japan, Spain, and China. To characterize the population genetic structure of the citrus red mite, the genetic variation in 15 populations from three citrus-planting regions of China (citrus belt of upper and middle reaches of Yangtze River [UMYR], citrus base of Yungui Plateau [YGP], and citrus belt of South China [SC]) was investigated using the ribosomal internal transcribed spacer (ITS) 1 region. These populations were characterized by high genetic diversity. Neutrality tests, mismatch analysis, and star-like network strongly supported the population expansion. Analysis of molecular variance (AMOVA) showed that no significant population genetic structure existed among UMYR, YGP, and SC. However, a significant global φ;ST and the genetic structure were found between (UMYR YGP) and SC. Pairwise φ;ST values between most populations were not significant, and a high gene flow was found among some geographically distant populations. The significant influence of host plants on the genetic structure of P. citri populations was found by the AMOVA for five host-related groups. In addition, the Mantel test revealed that geographical isolation was a factor responsible for the genetic differentiation. Therefore, our current observed population structure of P. citri can be contribute to several factors together, i.e., ongoing gene flow, the retention of ancestral polymorphisms, and natural selection (e.g., host plants, acaricides).