BACKGROUNDPractical resistance of Helicoverpa zea to Cry proteins has become widespread in the US, making Vip3Aa the only effective Bacillus thuringiensis (Bt) protein for controlling this pest. Understanding the genetic basis of Vip3Aa resistance in H. zea is essential in sustaining the long‐term efficacy of Vip3Aa. The objectives of this study were to characterize the inheritance of Vip3Aa resistance in four distinct field‐derived H. zea strains (M1‐RR, AC4‐RR, R2‐RR and R15‐RR), and to test for shared genetic basis among these strains and a previously characterized Texas resistant strain (LT#70‐RR).RESULTSMaternal effects and sex linkage were absent, and the effective dominance level (DML) was 0.0 across Vip3Aa39 concentrations ranging from 1.0 to 31.6 μg cm−2, in all H. zea resistant strains. Mendelian monogenic model tests indicated that Vip3Aa resistance in each of the four strains was controlled by a single gene. However, interstrain complementation tests indicated that three distinct genetic loci are involved in Vip3Aa resistance in the five resistant H. zea strains: one shared by M1‐RR and LT#70‐RR; another shared by R2‐RR and R15‐RR; and a distinct one for AC4‐RR.CONCLUSIONResults of this study indicate that Vip3Aa resistance in all H. zea strains was controlled by a single, recessive and autosomal gene. However, there were three distinct genetic loci associated with Vip3Aa resistance in the five resistant H. zea strains. The information generated from this study is valuable for exploring mechanisms of Vip3Aa resistance, monitoring the evolution of Vip3Aa resistance, and devising effective strategies for managing Vip3Aa resistance in H. zea. © 2024 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.