Characterization of Resistance to the Cry3Bb11 Toxin from Bacillus thuringiensis in Western Corn Rootworm, Diabrotica virgifera virgifera (Leconte) (Coleoptera: Chrysomelidae)

The western corn rootworm (WCR) Diabrotica virgifera virgifera LeConte is a major pest of corn. It has developed resistance against multiple chemical compounds and cultural practices used for management. One recent event is the resistance to transgenic corn expressing toxins from Bacillus thuringiensis. This study investigates molecular mechanisms involved in the resistance to Cry3Bb1 in WCR. For this purpose, neonates susceptible and resistant to Cry3Bb1 were studied. Larvae were exposed either to Cry3Bb1-expressing corn or near-isoline corn, for 4 or 8 hrs. RNA from whole larvae was extracted and sequenced for differential expression analysis of RNA sequencing. The results show different levels of expression between strains and between treatments for transcripts corresponding to proteins such as ABC transporters, amylases, proteases and protease inhibitors, detoxifying enzymes (cytochrome P450) and immunity related proteins. The proteins corresponding to the identified transcripts were consistent with results on the mode of action and resistance mechanisms of Cry toxins in other Coleoptera, Lepidoptera and Diptera. To confirm the results obtained by RNA sequencing and gain more information on the transcripts of interest, a quantitative Reverse Transcriptase PCR was performed, and their expression patterns and location were examined. Tissue location of transcripts is crucial in the interaction with Cry3Bb1, which is active in the midgut. The results were complemented by a detailed characterization of transcripts. Domains were identified for the membership to a family of protein. The combination of the two characterization methods assessed the likelihood of a transcript (protein) being involved in the interaction with Cry3Bb1 and resistance. Only two transcripts were located in the midgut, an α-amylase and an aspartic protease. RNA sequencing results were also used in the detection of polymorphisms that can be used as markers of the resistance to Cry3Bb1 in field populations. Polymorphisms were used to detect the effect of laboratory reselection on resistant populations of WCR collected in the field. However, the tested markers did not detect reselection, but a subset of loci was characteristic of the distinction between the susceptible non-diapausing strain and the resistant Hopkinton strain