Distribution and diversity of type III secretion system-like genes in saprophytic and phytopathogenic fluorescent pseudomonads

Abstract

Type three secretion systems (TTSSs) are protein translocation mechanisms associated with bacterial pathogenicity in host plants, and hypersensitive reactions in non-host plants. Distribution and diversity of TTSS-like genes within a collection of saprophytic and phytopathogenic fluorescent pseudomonads were characterized. This collection included 16 strains belonging to 13 pathogenic species, and 87 strains belonging to five saprophytic species isolated from plant rhizosphere and soil. Presence of conserved hypersensitive reaction/pathogenicity (hrp) genes (hrc RST) was assessed both by PCR using primers designed to amplify the corresponding sequence and by dot-blot hybridization using a PCR-amplified hrc RST fragment as a probe. PCR allowed the detection of TTSS-like genes in 75% and 32% of the phytopathogenic and saprophytic strains, respectively, and dot-blot hybridization in 100% and 49% of the phytopathogenic and saprophytic strains, respectively. The restriction fragment length polymorphism (RFLP) of 26 amplified hrc RST fragments revealed a considerable diversity. Twenty-one distinct RFLP types were identified and one hrc RST fragment was sequenced per RFLP type. The obtained hrc RST sequences clustered into three groups. Two of these groups included both phytopathogenic and saprophytic strains. The diversity of 16S rRNA genes, commonly used as an evolution marker, was characterized using PCR-RFLP. Polymorphism of the 16S rRNA genes corresponded to that of hrc RST genes, suggesting that these genes have followed a similar evolution. However, the occurrence of few mismatches suggests that sometimes TTSS-like genes might have undergone horizontal genetic transfer.

Type III Secretion System and Virulence Markers Highlight Similarities and Differences between Human- and Plant-Associated Pseudomonads Related to Pseudomonas fluorescens and P. putida

“…None of these isolates produced the pyocyanin pigment in King A medium or the quorum-sensing Pseudomonas quinolone signal (data not shown), traits shared only by P. aeruginosa strains (46,47). A second set of 12 strains associated with plants consisted of (i) 3 reference strains harboring T3SSs, i.e., C7R12, SBW25, and F113, well described for their beneficial effects on plant growth and health and for the presence of T3SSs (22,23,48), and (ii) 9 additional strains isolated from the rhizosphere or phyllosphere of four plant species and chosen to cover the diversity of hrcRST (rscRST) genes described so far in P. fluorescens (23,49). A third set was composed of three reference strains and included (i) the type strain of P. fluorescens species (ATCC 13525 T type strain), which does not harbor any T3SS gene, and (ii) two pathogenic strains of Pseudomonas extensively studied for their T3SSs, one strain pathogenic to humans, P. aeruginosa PAO1, and one strain pathogenic to plants, P. syringae pv.…”

“…hrcRST (rscRST) and pscRST, genes encoding structural components of T3SSs belonging to the Hrp1 and to the Ysc family, respectively, were amplified. hrcRST (rscRST) DNA fragments were amplified using HRCR8092 5=-CCITT(C/T)ATCGT(C/T)AT(C/T)GA(C/T)(C/T)T-3= and HRCT8986 5=-CTGTCCCAGATIAICTGIGT-3= (where I indicates d-inosine), as previously described by Mazurier et al (23). pscRST DNA fragments were amplified with HRCRPa, 5=-CCNTT(C/T)ATCGAT (C/T)GA(C/T)C-3=, and HRCTPa, 5=-AC(A/C)GGCCAACTTGS(A/G) TAGC-3=.…”

“…In the human pathogen P. aeruginosa, T3SSs belong to the Ysc-T3SS family; they form a short needle embedded within the two Gram-negative bacteria membranes, which serves as a secretion channel for the injection of exoenzymes, playing key roles in eukaryotic cell intoxication, into the target cell cytosol (17-19). In the plant pathogen P. syringae, the T3SS involved in plant pathogenicity belongs to the Hrp1-T3SS family; it produces a long and flexible pilus, similar to a flagellum, which is capable of transpiercing the thick wall of plant cells and allows the injection of multiple effector proteins into the cells that suppress the plant's innate immune defenses, manipulate hormone signaling, and elicit cell death (20,21).Although the P. fluorescens species is considered nonphytopathogenic, T3SS genes related to the Hrp1-T3SS family (15, 16) have been reported in rhizosphere populations (22)(23)(24). More generally, the presence of this secretion system involved in cell-tocell interactions with eukaryotic organisms appears to be quite a Citation Mazurier S, Merieau A, Bergeau D, Decoin V, Sperandio D, Crépin A, Barbey C, Jeannot K, Vicré-Gibouin M, Plésiat P, Lemanceau P, Latour X.…”

“…Although the P. fluorescens species is considered nonphytopathogenic, T3SS genes related to the Hrp1-T3SS family (15,16) have been reported in rhizosphere populations (22)(23)(24). More generally, the presence of this secretion system involved in cell-tocell interactions with eukaryotic organisms appears to be quite a common feature in nonpathogenic strains of plant-associated P. fluorescens (16,23,25).…”

“…More generally, the presence of this secretion system involved in cell-tocell interactions with eukaryotic organisms appears to be quite a common feature in nonpathogenic strains of plant-associated P. fluorescens (16,23,25).…”