In the mid-southern region of the United States, sugarcane borer, Diatraea saccharalis (F.), is a major target pest of transgenic maize expressing Bacillus thuringiensis (Bt) proteins. Novel transgenic maize technologies containing two or more pyramided Bt genes for controlling lepidopteran pests have recently become commercially available. Insect resistance management (IRM) is an important issue in the sustainable use of Bt crop technologies. The objective of this study was to determine the frequency of resistance alleles in field populations of D. saccharalis to the new pyramided Bt maize technologies. A total of 382 F(2) family lines derived from 735 feral larvae/pupae of D. saccharalis collected from six locations in Louisiana and Mississippi during 2008 and 2009 were screened for resistance to three new Bt maize technologies: MON 89034, Genuity™ VT Triple Pro™ and SmartStax™. None of the 382 F(2) isoline families survived on the Bt maize leaf tissue for ≥ 12 days in the F(2) screen. The joint frequency for two- or three-gene resistance models with 95% probability in these populations was estimated to be < 0.0063 to MON 89034 and < 0.003 to VT Triple Pro™ and SmartStax™. These results suggest that the resistance allele frequency in D. saccharalis to the three pyramided Bt maize technologies is low in the mid-southern region of the United States, which should meet the rare resistance assumption of the currently used IRM strategy for Bt maize.