Hrp ^- Mutants of Pseudomonas solanacearum as Potential Biocontrol Agents of Tomato Bacterial Wilt

Abstract

There have been many attempts to control bacterial wilt with antagonistic bacteria or spontaneous nonpathogenic mutants of Pseudomonas solanacearum that lack the ability to colonize the host, but they have met with limited success. Since a large gene cluster ( hrp ) is involved in the pathogenicity of P. solanacearum , we developed a biological control strategy using genetically engineered Hrp - mutants of P. solanacearum. Three pathogenic strains collected in Guadeloupe (French West Indies) were rendered nonpathogenic by insertion of an ω-Km interposon within the hrp gene cluster of each strain. The resulting Hrp - mutants were tested for their ability to control bacterial wilt in challenge inoculation experiments conducted either under growth chamber conditions or under greenhouse conditions in Guadeloupe. Compared with the colonization by a pathogenic strain which spread throughout the tomato plant, colonization by the mutants was restricted to the roots and the lower part of the stems. The mutants did not reach the fruit. Moreover, the presence of the mutants did not affect fruit production. When the plants were challenge inoculated with a pathogenic strain, the presence of Hrp - mutants within the plants was correlated with a reduction in disease severity, although pathogenic bacteria colonized the stem tissue at a higher density than the nonpathogenic bacteria. Challenge inoculation experiments conducted under growth chamber conditions led, in some cases, to exclusion of the pathogenic strain from the aerial part of the plant, resulting in high protection rates. Furthermore, there was evidence that one of the pathogenic strains used for the challenge inoculations produced a bacteriocin that inhibited the in vitro growth of the nonpathogenic mutants.

Aggressiveness of French isolates of Ralstonia solanacearum and their potential use in biocontrol1

“…Biological control of bacterial wilt is based on microbial antagonism towards R. solanaceam (Trigalet et al, 1994) and we have long been developing techniques for biological control by using genetically engineered Hrp-mutants from pathogenic strains isolated from tomato in Guadeloupe (Trigalet & Trigalet-Demery, 1990;Frey et al, 1994) or from potato in Kenya (Smith et al, 1994). Competition for space in xylem vessels and induction of localized host resistance (tyloses and cell browning) are possible explanations for the protective ability of an Hrp-mutant strain against subsequent invasion by a pathogenic strain.…”

Secretion and regulation Hrp mutants of Erwinia amylovora trigger different responses in apple

“…When there was no protection, the virulent strain multiplied after a 3-day delay. In contrast, a study of the protective e¡ect of Hrp 3 mutants of Ralstonia solanacearum on bacterial wilt of tomato [12] has shown that the presence of Hrp 3 mutants within the plants was correlated with a reduction in the disease severity but was not associated with exclusion of the pathogenic strain, which remained at a much higher population density than the nonpathogenic bacteria.…”

An improved control efficacy against tobacco bacterial wilt by an engineered Pseudomonas mosselii expressing the ripAA gene from phytopathogenic Ralstonia solanacearum

“…Pseudomonas spp., Bacillus spp., Streptomyces spp., and other bacterial species are promising for controlling bacterial wilt (17). Interestingly, several avirulent R. solanacearum isolates are able to prevent bacterial wilt disease development (27). Pseudomonas spp.…”