The whitefly Bemisia tabaci (Gennadius) is one of the most important pests causing economic losses in a variety of cropping systems around the world. This species was recently found in a coastal region of Colombia and has now spread inland. To investigate this invasive process, the genetic structure of B. tabaci was examined in 8 sampling locations from 2 infested regions (coastal, inland) using 9 microsatellite markers and the mitochondrial COI gene. The mitochondrial analysis indicated that only the invasive species of the B. tabaci complex Middle East-Asia Minor 1 (MEAM 1 known previously as biotype B) was present. The microsatellite data pointed to genetic differences among the regions and no isolation by distance within regions. The coastal region in the Caribbean appears to have been the initial point of invasion, while the inland region in the Southwest showed genetic variation among populations most likely reflecting founder events and ongoing changes associated with climatic and topographical heterogeneity. These findings have implications for tracking and managing B. tabaci.