Rice striped stem borer (RSSB, Chilo suppressalis) is a serious lepidopteron pest occurring in rice ecosystems of Asia and Europe. Genetically modified (GM) insect-resistant Bt rice has been developed to deter lepidopteron pests including RSSB. The concern of resistance evolution to the Bt toxin by the pests under commercial cultivation of GM Bt rice and the need of effective management of the resistance encourage the studies of genetic variation and divergence, as well as gene flow of RSSB populations. We analyzed 13 RSSB populations fed on water-oats or rice plants, respectively, from southeast China applying the fluorescent amplified fragment length polymorphism fingerprints. A generally moderate level of genetic variation was detected in the populations, as estimated by Nei's genetic diversity (0.27) and Shannon's index (0.42). The FST and amova values indicated a low level (&amp;#8764;12%) of genetic divergence among the RSSB populations. A relatively frequent gene flow (an average Nm = 2.62) was detected among the 12 RSSB populations, which may explain the limited genetic divergence among the rice-feeding populations. This explanation gains support by the assignment test of the corresponding populations, suggesting that a considerable proportion of individuals was contributed from non-native populations. Our results revealed that the moderate level of genetic diversity combined with relatively frequent gene flow among RSSB populations across a large geographical range may slow down the resistance evolution of the RSSB populations, given that a proper measure of resistance management is taken.