Cry proteins of Bacillus thuringiensis (Bt) have been successfully used as biopesticides and in transgenic crops throughout the world. However, resources against the most serious agricultural pathogens, plant root-knot nematodes, are limited. The genomes of several highly nematicidal virulent Bt strains from our laboratory have been sequenced, facilitating the identification of novel Cry proteins and other virulence factors. We identified two novel Cry proteins, Cry5Ca1 and Cry5Da1, that exhibit high toxicity against Meloidogyne incognita. Using the Caenorhabditis elegans model, the two Cry5 toxins were shown to negatively affect nematode life span, fertility, and survival. The 50% lethal concentrations (LC50s) of Cry5Ca1 and Cry5Da1 were 57.22 μg/ml and 36.69 μg/ml, respectively. Moreover, a synergistic effect (synergism factor, 1.61 to 2.04) was observed for nematicidal toxicity of Cry5Ca1 and Cry5Da1, which is accordant with the phylogenetic results suggesting that domain II of the two novel Cry5 toxins evolved into two independent clades. Through comparison of the depressed degree of toxicity in the β-methylgalactoside detoxification test, we found that the novel toxin Cry5D possesses a different galactosebinding epitope; meanwhile, the finding that Cry5D does not share a motif (GXXXE) in the corresponding loop of domain II with Cry5B could explain the different galactose binding performance. Additionally, low-level cross-resistance of C. elegans bre mutant strains was evident between Cry5B and Cry5D. These results suggest that Cry5D can be used as an alternative to delay the potential resistance of nematodes to Cry5B.
Geng, C., Liu, Y., Li, M., Tang, Z., Muhammad, S., Zheng, J., … Sun, M. (2017). Dissimilar crystal proteins Cry5Ca1 and Cry5Da1 synergistically act against Meloidogyne incognita and delay Cry5Ba-based nematode resistance. Applied and Environmental Microbiology, 83(18). https://doi.org/10.1128/AEM.03505-16