A Bacillus thuringiensis ΔcdsR-based expression system for insecticidal proteins identified using AlphaFold3.

Abstract

Environmentally friendly biopesticides, such as those based on the bacterium Bacillus thuringiensis (Bt), have an important role to play in Integrated Pest Management systems. An increasingly diverse range of pesticidal proteins is being discovered, led by advances in genome sequencing and the bioinformatic identification of candidate genes. In this work, we utilized an AlphaFold3-based structure-prediction approach to identify candidate genes and, in parallel, developed a Bt chassis strain that can be universally used to express new pesticidal proteins. This chassis strain is based on a cdsR deletion strain with a controllable cell death pathway that decouples toxin production from sporulation, enabling cytoplasmic hyperexpression via strong promoters and encapsulation of recombinant proteins. These characteristics provide superior efficacy, UV stability, and environmental safety compared with conventional Bt formulations. Several of the identified proteins displayed insecticidal activity when expressed in this strain, providing a versatile framework for next-generation biocontrol and sustainable pest-management innovation.