Screening for Cry proteins with unique receptors in fall armyworm (Spodoptera frugiperda) and corn earworm (Helicoverpa zea)

Bacillus thuringiensis (Bt) proteins have been the primary method for controlling Lepidopteran pests of corn and cotton over the past 20 years, due to their effectiveness and high level of target specificity. The fall armyworm (Spodoptera frugiperda) developed resistance to Cry1f, a Bt protein produced by both corn and cotton, within three years of registration in Puerto Rico. The evolution of resistance in pest insects poses as a threat to food safety as well as potentially resulting in economic losses of over $1 billion. The majority of Cry proteins follow a three domain (I to III) structure that is important for their mode of action as well as this research. One of the key steps in the Cry mode of action, mostly controlled by domain II, is the recognition of receptors in target midgut cells. Domain III has also been found to be involved in some cases, though, and is an important topic in this project. The overarching objective of this project is to serve as a proof of concept for a high-throughput method of identifying Cry proteins that may be suitable for gene pyramiding. I have tested this by first producing individual Cry1F domains and testing their ability to prevent binding and toxicity of full length Cry1F toxin against larvae of the fall armyworm (Spodoptera frugiperda) and corn earworm (helicoverpa zea) through competition bioassays, as described in objectives one, two, and three. As confirmation that this method can be used to identify toxins whose insecticidal activity is unknown, in objective four, I have completed the actions in objective two with Cry1Ba, Cry1Da, and Cry9Aa and performed competition bioassays.