Physiological Factors Influencing Bacillus thuringiensis Susceptibility in Laboratory and Field-Collected Bollworm, Helicoverpa zea (Lepidoptera: Noctuidae)

Authors: Du, Yuzhe; Scheibener, Shane; Little, Nathan; Elkins, Blake; Zhu, Yu Cheng

Submitted to: Agrochemicals
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/30/2026
Publication Date: 2/3/2026
Citation: Du, Y., Scheibener, S.A., Little, N., Elkins, B.H., Zhu, Y. 2026. Physiological Factors Influencing Bacillus thuringiensis Susceptibility in Laboratory and Field-Collected Bollworm, Helicoverpa zea (Lepidoptera: Noctuidae). Agrochemicals. 5(9):1-12. https://doi.org/10.3390/agrochemicals5010009.
DOI: https://doi.org/10.3390/agrochemicals5010009

Interpretive Summary: The corn earworm (Helicoverpa zea) is a major agricultural pest that affects crops such as corn and cotton. To control these pests, researchers have developed genetically modified (Bt) crops that produce insecticidal proteins like Cry toxin. However, some corn earworm populations have devolved resistance to Cry toxin over time, leading to the adoption of a new Bt protein, Vip. IIn this study, we analyzed three strains of corn earworm—two susceptible to Bt toxins and one that exhibited field resistance to Cry. We investigated the activity of two serine proteases and two binding receptor proteins in the insect's gut. Results revealed that resistant insects displayed altered protease activity and gene expression, likely diminishing the effectiveness of Cry toxin. Exposure to Vip led to an increase in one protease activity, indicating its role in activating Vip. Additionally, we assessed the expression levels of Cry binding receptor genes and detoxification enzymes. Their findings highlighted changes in protease and receptor gene expression—rather than detoxification enzymes—as key factors contributing to Cry toxin resistance in field-derived corn earworm populations. This study suggests that resistance mechanisms in corn earworms stem from modifications in proteolytic processing and gene expression.

Technical Abstract: Field-evolved resistance of Helicoverpa zea to the Bacillus thuringiensis (Bt) crystalline (Cry) toxins was first documented over a decade ago, prompting the adoption of genetically engineered crops expressing the Bt vegetative insecticidal protein (Vip). To better understand the molecular and biochemical mechanisms underlying Cry resistance, and to compare the distinct actions to Vip in H. zea, we analyzed two laboratory strains (Benzon and SIMRU) and one field population collected from commercial corn near Pickens, AR. In midgut brush-border membrane vesicles (BBMVs) isolated from larval midguts, the activities of chymotrypsin-like proteases, aminopeptidase N (APN) and alkaline phosphatase (ALP) were significantly altered in the Pickens strain compared to the SIMRU or Benzon populations in both 5th and 4th instar larvae, respectively. In contrast, no significant differences in trypsin-like activities were detected among three colonies. In Benzon larvae, exposure of neonates to a sublethal dose of Vip3A significantly increased chymotrypsin-like activity in 3rd instars but had no effect in later instars. Gene expression analysis of two serine protease genes (trypsin CFT-1 like serine and chymotrypsin-1-like protease), and five receptor binding genes (cadherin, ATP-binding cassette family C2, APN1, APN2, ALP) revealed significant downregulation of chymotrypsin-1-like protease and APN2 gene in the tolerant Pickens population. In contrast, transcript levels of gut detoxification enzymes genes (esterase, GST, CYP6B8 and CYP6B28) showed no significant differences among the three colonies. These findings suggest that chymotrypsin protease may play an important role in the activation of both Cry1Ac and Vip3A toxins in H. zea. The altered chymotrypsin-like and APN activities and downregulation of chymotrypsin-1-like protease and APN2 in the Pickens population may contribute to Cry tolerance. These insights into the biochemical and molecular basis of Bt tolerance may enhance our understanding of resistance mechanisms and support the development of more effective insect resistance management strategies.