Construction and characterization of extracellular polysaccharide deletion mutant of Ralstonia solanacearum

Abstract:

[Objective] Bacterial wilt is a soil-borne disease caused by Ralstonia solanacearum. Extracellular polysaccharides are one of the key pathogenic factors of R. solanacearum. The physiological functions of extracellular polysaccharides on the pathogenesis of bacterial wilt were studied by constructing a mutant with synthesis of extracellular polysaccharides deficiency. [Methods] First, we cloned the homologous arm of epsD from the genome of R. solanacearum FJAT-91, then inserted into suicide plasmid pK18mobsacB. Second, the Gm gene was inserted into homologous arm to obtain recombinant plasmid pK18-epsD. Third, the recombinant plasmid was transformed into R. solanacearum FJAT-91 competent cells. The epsD gene deletion mutant was constructed by homologous recombination. Finally, we detected the differences of the biological characteristics of mutant strains and wild-type strains. [Results] Compared with the wild-type strains, the mutant strains showed different features. The mutant showed: grew slowly and decreased the yield of extracellular polysaccharides; significantly reduced the abilities of swimming motility and swarming motility; significantly decreased the colonization ability in roots and stems of tomato; had the Attenuation Index (AI) with 0.905 and the avirulence to tomato which indicated a nonpathogenic strain. [Conclusion] These results suggested that extracellular polysaccharides played the key role in pathogenicity of R. solanacearum. This research provides excellent materials and research foundation for the development of plant vaccines.