Screening of phenotypic conversion mutant strains of Ralstonia solanacearum for effective biological control of Verticillium wilt in eggplant

Abstract

Ralstonia solanacearum causes bacterial wilt and undergoes spontaneous phenotypic conversion (PC) from its wild-type form to non-pathogenic form. We previously reported that the pre-inoculation of PC mutants in Solanum plants suppressed bacterial wilt. In the present study, we clarified the inoculation method of PC mutants and PC mutant strains for effective control the eggplant Verticillium wilt caused by Verticillium dahliae, a soil-borne fungal plant pathogen. We first investigated three different methods for pre-inoculating PC mutants in the five-leaf-stage seedlings of eggplant: Method A involved soaking plant roots in the PC mutant suspension immediately before challenge-inoculation with V. dahliae; Method B involved irrigation with the PC mutant suspension in pots at 7 days before challenge-inoculation with the pathogen; and Method C involved soaking plant roots in the PC mutant suspension at 3 weeks before challenge-inoculation with the pathogen. The highest biological control was achieved with Method A. To quickly and easily select effective PC mutants for biological control of Verticillium wilt in a growth chamber, a simplified evaluation method based on Method A was applied using the young (two-leaf-stage) seedlings of eggplant. Furthermore, we investigated the biological control efficacies of 27 PC mutants against Verticillium wilt in eggplants using the simplified evaluation method in vivo, and growth inhibition of V. dahliae by PC mutants in vitro. However, no relationship existed between the biological control and growth inhibition of V. dahliae, suggesting that it is unlikely to be involved as the main factor affecting the biological control by PC mutants. Three PC mutants, 8345PC, 8109PC, and 8107PC, which showed high biological control efficacies in the simplified evaluation method (64.6%, 66.0%, and 68.9%, respectively), were evaluated for their biological control using five-leaf-stage eggplant seedlings in greenhouse test. Among these three mutants, the 8107PC strain showed the highest biological control of 43.3% and was selected as the most effective PC mutant for suppressing Verticillium wilt in eggplant. Thus, the use of PC mutants is expected to be effective in the control of Verticillium wilt in eggplant.