Spatial genetic variability of the fall armyworm, Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae), was studied by collecting samples from 31 locations in the United States, Argentina, Panama, and Puerto Rico, and then using amplified fragment length polymorphism to detect genetic variation. Analysis of molecular variance showed significant genetic variation in fall armyworm among all (28%) sample locations and individuals within (71%) sample locations; genetic variation of fall armyworm was minimal between sample locations grouped into regions. The pairwise fixation index (F_ST) comparisons showed significant genetic differentiation (0.288) among the 31 locations. However, dendrograms of results from cluster analysis did not provide support of a high level of genetic structuring among regions. The isolation by distance analysis for all sample locations showed the absence of significant correlation between genetic dissimilarity and geographic distance except for fall armyworm samples collected within Argentina. Moreover, the estimate of modest gene flow (Nm >1) may prevent gene fixation within regions. These results indicate that fall armyworm shows little genetic differentiation and high genetic diversity over its range, important information for the development of management strategies and monitoring the development of resistance to management techniques.