Investigation of Cry3Bb1 resistance and intoxication in Western Corn Rootworm by RNA sequencing

The western corn rootworm (WCR) Diabrotica virgifera virgifera LeConte is a major pest of corn that has evolved resistance to transgenic maize that produces insecticidal Cry toxins. The specific mode of action of Cry3Bb1 and mechanism of resistance in WCR are unknown. This study compared gene expression between Cry3Bb1-susceptible and Cry3Bb1-resistant WCR neonates, in the presence and absence of Cry3Bb1. RNA-Seq data were analysed to identify differentially expressed transcripts between strains of WCR, providing candidate transcripts for resistance to Cry3Bb1. Constitutive and Cry3Bb1-induced differences between strains caused the differential expression of 608 transcripts after 8hr. Differentially expressed transcripts between strains included ABC transporters, proteases and -amylases, which known to be receptors or activators of Cry toxins and involved in resistance to Cry toxins in other insects. The response to Cry3Bb1 treatment resulted in approximately 5,000 differentially expressed transcripts in the susceptible strain and included the same annotation categories found between strains but also included metalloproteases, cadherins and signalling proteins. None of these annotations were identified in the response of the resistant strain to Cry3Bb1, which was represented by only 12 transcripts. Tissue-specific expression analysis of selected transcripts revealed that an -amylase and a protease were expressed in the midgut, the target organ of Cry toxins. A protease inhibitor and two ABC transporters were expressed outside the midgut, suggesting a limited role in resistance. Numerous polymorphic sites were identified from the RNA-Seq data that showed allele frequency differences between the resistant and susceptible strains. Analysis of these polymorphisms in a larger set WCR strains suggested that the differences were due to genetic drift rather than being associated with resistance to Cry3Bb1. Polymorphisms identified in genes with known roles in resistance to Cry toxins did not appear to differ in frequency between resistant and susceptible strains.