Molecular genetics of Pseudomonas syringae pathovar pisi: plasmid involvement in cultivar-specific incompatibility

Abstract

A mutant (PF24) of the race 1 strain, 2WA, of Pseudomonas syringrre pv.pisi has been characterized in terms of its interactions with pea (Pisum satiuum) cultivars. The mutant showed a changed reaction (avirulence to virulence) with a gruup of pea cultivars, including cvs. Belinda and Puget, previously thought to contain resistance genes R1 and R3. Avirulence towards cv. Puget was restored by transfer of any one of five cosmid clones from a race 3 (strain 870A) gene library to a rifampicin-resistant derivative of PF24. These observations were in agreement with a revised race-specific resistance genotype for Belinda and similar cultivars comprising a single resistance gene, R3. An incompatible interaction was observed between strain PF24 and cvs. Vinco (postulated to harbour race-specific resistance genes R1, R2, R 3 and R5) and Hurst's Greenshaft (R4 and possibly Rl), indicating that the mutant retains at least one avirulence gene (A1 or A1 and A4). Mutant PF24 showed loss of a cryptic plasmid (pAV212) compared with its progenitor, strain 299A. A subclone (pAV233) of one of the race 3 restoration clones showed strong hybridization with similar-sized digestion fragments in race 3 plasmid DNA, confirming the A3 gene to be plasmid-borne. Strong cross-hybridization was also observed with a single 3.27 kb EcoRI fragment of plasmid DNA present in strain 2WA but absent from strain PF24. This is consistent with the corresponding A3 determinant being located on pAV212 in the race 1 strain 2WA. The novel avirulence gene corresponding to A3 in strain 870A is provisionally designated aurPpi3. A spontaneous race-change variant (strain 1759, which expressed no avirulence phenotype toward the pea differential cultivars) was derived from the race 3 strain 870A. This race 6 strain and a wild isolate of a race 6 strain both lacked plasmid DNA sequences corresponding to the insert in pAV233.

Changes in cultivar-specificity toward pea can result from transfer of plasmid RP4 and other incompatibility group P1 replicons to Pseudomonas syringae pv. pisi.

“…Strains of Escherichia coli and P. syringae pv. pisi were cultured as described by Bavage et al (1991). Fluorescent pigment production was observed after growth (25 "C, 2 d) of strains of P. syringae pv.…”

“…Plant reactions were recorded 7 d after inoculation. A susceptible response was characterized by an area of water-soaking around the site of inoculation while a hypersensitive response was characterized by a necrotic area around the site of inoculation (Bavage et al, 1991).…”

“…These were based on the methods of Sambrook et al (1989), modified by Bavage et al (1991). Plasmid DNA \ for digestion with restriction endonucleases was isolated by the method of Bavage et al (1991).…”

“…pisi and pea (Pisum sativum) a gene-forgene model has been proposed Bavage et al, 1991). Of the five avr genes postulated to occur, two (A2 and A3 in the scheme of Taylor et al, 1989) have been cloned (Vivian et al, 1989 ;Bavage et al, 1991). The A2 gene has been shown to interact with a single R2 gene in P. sativum (Vivian et al, 1989) and also with the R P M l gene in Arabidopsis thaliana (Dangl et al, 1992).…”

“…pisi and pea (Pisum sativum) a gene-forgene model has been proposed Bavage et al, 1991). Of the five avr genes postulated to occur, two (A2 and A3 in the scheme of Taylor et al, 1989) have been cloned (Vivian et al, 1989 ;Bavage et al, 1991).…”