Abstract: 【Objective】The objective of this study is to analyze the heterogeneity of Ralstonia solanacearum mutants by Tn5 transposon and screen out an avirulent mutant with high control efficiency. 【Method】 The pathogenicity of sixty R. solanacearum Tn5 transposon mutants was identified by using the attenuation index (AI, a quantitative index for calculating the pathogenicity of R. solanacearum) and tomato plant inoculation bioassay. The chromatographic behavior heterogeneity of different pathogenic mutants of R. solanacearum was analyzed by using high performance ion-exchange chromatography. A chromatography titer index (CTI_i) was expressed as CTI_i=S_i/ (S₁+S₂+S₃)×100% (i=1, 2 or 3; S₁, S₂ and S₃ represent peak areas of P₁, P₂ and P₃, respectively), and the relationship of CTI_i with the pathogenicity and control efficiency of tested mutants was also analyzed based on Pearson correlation coefficient (PCC). 【Result】 Based on the value of AI and the results of bioassay, sixty R. solanacearum mutants were divided into three pathogenic types: virulent, avirulent and interim. Thirty-three mutants belonged to virulent mutant with the value of AI ranged from 0.49 to 0.63 and disease incidence ranged from 66.33% to 100% (15 days after inoculation). Twenty mutants were avirulent with the value of AI ranged from 0.78 to 0.89 and the disease incidence was 0. There were only seven interim mutants with the value of AI ranged from 0.68 to 0.73 and disease incidence ranged from 24.17% to 45.92%. The control efficiency of twenty avirulent mutants against tomato bacterial wilt disease ranged from 16.68% to 92.45%. The best control efficiency was obtained by T831 of 91.74%, followed by T780 (87.51%). The mutant of T497 had the worst control efficiency of 16.68%. Using high performance ion-exchange chromatography, R. solanacearum mutants were successfully separated into three chromatographic peaks: P₁ (retention time of 0.6 min), P₂ (retention time of 4.4 or 4.5 min) and P₃ (retention time of 5.9 or 6.0 min). Both virulent and avirulent mutants had three different peaks: single peak, double and three peaks. However, the interim mutants only had double  peaks and three peaks types. For virulent mutants, the intense peak is P₃, while for avirulent mutants, P₁ is the majority. The virulent mutants with CTI₃ of 100% had high pathogenicity, which could induce tomato plant bacterial wilt with disease incidence up to 85%. The virulent mutants with CTI₃ less than 100% had the disease incidence ranged from 66.33% to 84.14%. The avirulent mutants with CTI₁ of 100% had high control efficiency which reached up to 80%, and the avirulent mutants with CTI₁ less than 90% had low control efficiency which ranged from 16.68% to 63.62%. The correlations of CTI₃ and the pathogenicity of virulent mutants were significantly positive (P＜0.01), with PCC of 0.62. The relationships between CTI₁ and control efficiency against bacterial wilt disease of the avirulent mutants were also significantly positive (P＜0.01), with PCC of 0.80. 【Conclusion】 R. solanacearum mutants by Tn5 transposon have three pathogenic types, which resulted in different chromatographic behaviors. CTI₁ constructed in this study can be used for rapidly selecting an avirulent mutant with high control efficiency and CTI₃ can be used for calculating the pathogenicity of R. solanacearum.

Key words: high performance ion-exchange chromatography; Ralstonia solanacearum')">