Spray products based on Bacillus thuringiensis (Bt) (Berliner) (Bacillales: Bacillaceae) are widely used as control agents of the fall armyworm, Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae), in some major crops such as corn, cotton, and soy. However, there is little information about the toxicity of Bt formulations to control corn and rice-infesting S. frugiperda strains. These S. frugiperda strains are morphologically indistinguishable, so they are genetically identified as corn and rice S. frugiperda strains. The main objective of this study was to evaluate the susceptibility of third instar larvae of both S. frugiperda biotypes to either the native Bt RT or the reference Bt HD1 subsp. kurstaki (Btk) formulations. In addition, the S. frugiperda populations were characterized by their esterase profile in polyacrylamide gels. This study also allowed differentiation of the S. frugiperda strains. Concerning the susceptibility assays, a significant difference in mortality values was found within some corn or rice S. frugiperda colonies when Bt RT treatment was used, whereas there were no significant differences in Btk HD1 toxicity within colonies that share the same biotype. When toxicity of either the native Bt RT or the reference Btk HD1 to each S. frugiperda biotype was compared, significant differences in mortality values also were found in the corn and the rice S. frugiperda strains. Finally, the statistical analysis of the mortality data revealed that there was a significant difference between the 2 S. frugiperda strains, regardless of the Bt preparation. These results show that the S. frugiperda control could depend not only on the bioinsecticide used, but also on the S. frugiperda biotype being treated.