Picture-winged flies (Diptera: Ulidiidae) are the most damaging insect pests of sweet corn (Zea mays L.) produced in Florida for the fresh market. Management of these pests, referred to as corn silk flies, relies on frequent pyrethroid applications targeting adults. In response to the need for an insecticide resistance management (IRM) program for corn silk flies in this highly intensive crop system, glass vial bioassays were conducted to determine the susceptibility of 12 corn silk fly populations to the pyrethroid beta-cyfluthrin. Two Euxesta eluta Loew and nine Euxesta stigmatias Loew populations were obtained by collecting infested ears in commercial and experimental fields in 2020 and 2021. One E. eluta laboratory colony was used as a susceptible reference population.The E. eluta reference colony was the most susceptible population, with an LC₅₀ value of 0.01 µg/vial.The E. stigmatias field populations were generally less susceptible to beta-cyfluthrin than the E. eluta field populations, with the highest LC₅₀ values attaining 3.51 µg/vial and 0.19 µg/vial, respectively. In addition, the five E. stigmatias populations from commercial sweet corn fields were as much as 17.6 times less susceptible than the four E. stigmatias populations from nontreated fields. Results suggest that E. stigmatias is less susceptible to pyrethroids than E. eluta. Results also suggest that corn silk flies in commercial sweet corn fields are selected for reduced pyrethroid susceptibility throughout the growing season.This study successfully used the glass vial bioassay method for corn silk flies, providing a new tool to initiate an IRM program.