Regulation of AHL Production and Its Contribution to Epiphytic Fitness in Pseudomonas syringae

Abstract

Pseudomonas syringae forms large cell aggregates that are more stress tolerant than solitary cells during epiphytic growth on plants. The differential survival of aggregates on leaves suggests that epiphytic fitness traits may be controlled in a density-dependent manner via cell-cell signaling. We investigated this hypothesis in P. syringae B728a. Synthesis of N-acyl-homoserine lactone (AHL), 3-oxo-hexanoyl homoserine lactone, and the expression of the gene encoding AHL synthase ahlI were maximal at high cell concentrations. The expression of the AHL regulator ahlR, in contrast, was similar at all cell concentrations. A screen of Tn5 mutants revealed that P. syringae B728a requires a novel transcriptional activator for AHL production. This regulator, which belongs to the TetR family, was also required for epiphytic fitness and has been designated AefR (for AHL and epiphytic fitness regulator). The expression of ahlI was greatly reduced in both aefR- and gacA- mutants and was completely restored in either mutant after addition of exogenous AHL. In contrast, the expression of aefR was not reduced in either gacS- or gacA- mutants. Thus, AefR appears to positively regulate AHL production independently of the regulators GacS/GacA and also controls traits in P. syringae B728a that are required for epiphytic colonization.

Acyl-homoserine lactone-mediated cross talk among epiphytic bacteria modulates behavior of Pseudomonas syringae on leaves

“…Individual leaves were placed in sterile plastic bags with 10 ml of 10 mM potassium phosphate buffer (KPO 4 buffer; pH 7.0) and sonicated for 7 min to remove bacteria from leaves as in other studies (Quinones et al, 2004. Appropriate dilutions of leaf washings were plated on KB and the plates were incubated at 28 1C for 3 days.…”

“…Bacterial colonies differing in appearance were spotted on KB and LA plates for assay of AHL production. The plates were then immediately sprayed with a dense suspension (410 9 cells per ml) of BHSL (pBQ9) (Quinones et al, 2004) with an artists' airbrush and incubated at 28 1C overnight. Induction of QS was visualized by illuminating plates with a handheld UV lamp and observing the green fluorescent halos in the lawn of BHSL (pBQ9).…”

“…Disease incidence was assessed on at least 14 individual leaves from each of five or more replicate pots. Epiphytic population sizes were determined by plating appropriate dilutions of washings of individual trifoliate leaves 24 h after inoculation as previously described (Quinones et al, 2004.…”

“…Such density-dependent behavior is apparently controlled by QS. AhlI confers production of 3OC6HSL under the control of ahlR and other regulatory genes in Pss (Quinones et al, 2004). A variety of traits including extracellular polysaccharide production, oxidative stress tolerance, swarming motility, promotion of water-soaked lesions and extent of internal tissue maceration in bean pods are regulated by QS in Pss Quinones et al, 2005).…”

The ppuI-rsaL-ppuR quorum-sensing system regulates cellular motility, pectate lyase activity, and virulence in potato opportunistic pathogen Pseudomonas sp. StFLB209

“…11) When AHL concentrations increase and reach a threshold, AHL receptor proteins belonging to the LuxR protein family bind AHL and regulate the expression of numerous genes. AHL-mediated quorum sensing regulates various phenotypes in plants, including the production of exoenzymes 12) and antimicrobial substances, 13) motility, 14−15) biofilm formation, 16) epiphytic fitness, 17) and pathogenicity. 12,18) Although many AHL-producing bacteria have been isolated from the phyllosphere, 4−7) the role of quorum sensing in adaptation to survival on the plant surface and its contribution to shaping certain community structures has not yet been elucidated.…”