Susceptibility of the sugarcane borer, Diatraea saccharalis (F.), to tebufenozide was measured using a feeding bioassay, and values obtained were compared with baselines generated before the use of this insecticide in Louisiana sugarcane, Saccharum spp. Results from our study suggest that susceptibility to tebufenozide is decreasing in field-collected sugarcane borers. Inflections in the log dosage-probit lines were detected for many of the field collections, indicating increased heterogeneity within these populations in response to tebufenozide. Where appropriate, probit transformation was used to estimate susceptibility, and significant differences (1.6–2.7-fold) were measured in LC₅₀ values between some field-collected cohorts and the previously measured baseline. In addition, a discriminating concentration (0.5 ppm) was used to estimate resistance frequencies in cohorts for which probit transformation was not appropriate. Results from these tests suggest that frequencies of resistance were high (49% in one cohort) in populations from some locations. Lighter weight pupae of the survivors from one of the more resistant cohorts suggests that tebufenozide resistance mechanisms may have a biological cost in terms of ecological fitness at early stages of resistance development. As a result of continued resistance monitoring, alternation of management chemistry is expected to help preserve this valuable sugarcane integrated pest management tactic.