Down Regulation of Virulence Factors of Pseudomonas aeruginosa by Salicylic Acid Attenuates Its Virulence on Arabidopsis thaliana and Caenorhabditis elegans

Department of Horticulture and Landscape Architecture, Colorado State University, 217 Shepardson Building, Fort Collins, CO 80523, USA.
Infection and Immunity (Impact Factor: 3.73). 10/2005; 73(9):5319-28. DOI: 10.1128/IAI.73.9.5319-5328.2005
Source: PubMed


Salicylic acid (SA) is a phenolic metabolite produced by plants and is known to play an important role in several physiological processes, such as the induction of plant defense responses against pathogen attack. Here, using the Arabidopsis thaliana-Pseudomonas aeruginosa pathosystem, we provide evidence that SA acts directly on the pathogen, down regulating fitness and virulence factor production of the bacteria. Pseudomonas aeruginosa PA14 showed reduced attachment and biofilm formation on the roots of the Arabidopsis mutants lox2 and cpr5-2, which produce elevated amounts of SA, as well as on wild-type Arabidopsis plants primed with exogenous SA, a treatment known to enhance endogenous SA concentration. Salicylic acid at a concentration that did not inhibit PA14 growth was sufficient to significantly affect the ability of the bacteria to attach and form biofilm communities on abiotic surfaces. Furthermore, SA down regulated three known virulence factors of PA14: pyocyanin, protease, and elastase. Interestingly, P. aeruginosa produced more pyocyanin when infiltrated into leaves of the Arabidopsis transgenic line NahG, which accumulates less SA than wild-type plants. This finding suggests that endogenous SA plays a role in down regulating the synthesis and secretion of pyocyanin in vivo. To further test if SA directly affects the virulence of P. aeruginosa, we used the Caenorhabditis elegans-P. aeruginosa infection model. The addition of SA to P. aeruginosa lawns significantly diminished the bacterium's ability to kill the worms, without affecting the accumulation of bacteria inside the nematodes' guts, suggesting that SA negatively affects factors that influence the virulence of P. aeruginosa. We employed microarray technology to identify SA target genes. These analyses showed that SA treatment affected expression of 331 genes. It selectively repressed transcription of exoproteins and other virulence factors, while it had no effect on expression of housekeeping genes. Our results indicate that in addition to its role as a signal molecule in plant defense responses, SA works as an anti-infective compound by affecting the physiology of P. aeruginosa and ultimately attenuating its virulence.

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Available from: Balakrishnan Prithiviraj, Dec 18, 2013
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    • "Phenolic acids are also known to regulate the infection efficiency during the exchange of signals between legumes and rhizobia (Mandal et al., 2010). However, despite the known effects of phenolic compounds on bacterial virulence, the mechanisms of action behind them are, as yet, unresolved (Chow et al., 2011; Mirzoeva et al., 1997; Prithiviraj et al., 2005). "
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    ABSTRACT: Several studies have reported effects of the plant phenolic acids cinnamic acid (CA) and salicylic acid (SA) on virulence of soft rot enterobacteria. However, the mechanisms involved in these processes are not yet fully understood. Here, we investigated whether CA and SA interfere with the quorum-sensing (QS) system of two Pectobacterium species, P. aroidearum and P. carotovorum subsp brasiliense, which are known to produce N-acyl-homoserine lactone (AHL) QS signals. Our results clearly indicate that both phenolic compounds affect the QS machinery of the two species, consequently altering the expression of bacterial virulence factors. While in control treatments, expression of QS-related genes increased over time, exposure of bacteria to nonlethal concentrations of CA or SA inhibited the expression of QS genes, including expI, expR, PC1_1442 (luxR transcriptional regulator) and luxS (a component of the AI-2 system). Other virulence genes known to be regulated by the QS system, such as pecS, pel, peh and yheO, were also down-regulated relative to the control. In agreement with the low levels of expression of expI and expR, CA and SA also reduced the level of AHL signal. The effects of CA and SA on AHL signaling were confirmed in compensation assays, in which exogenous application of N-(β-ketocaproyl)-L-homoserine lactone (eAHL) led to the recovery of the reduction in virulence caused by the two phenolic acids. Collectively, the results of gene expression studies, bioluminescence assays, virulence assays and compensation assays with eAHL clearly support a mechanism by which CA and SA interfere with Pectobacterium virulence via the QS machinery. This article is protected by copyright. All rights reserved.
    Molecular Plant Pathology 07/2015; DOI:10.1111/mpp.12295 · 4.72 Impact Factor
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    • "It is not well-understood which rhizodeposit compounds recruit or influence which groups of microbes and how. An effective approach is to examine microbial associations with plant mutants deficient in the biosynthesis and rhizodeposition of specific groups of compounds (Prithiviraj et al. 2005; Rudrappa et al. 2008). It is worth noting that composition of rhizodeposits varies substantially among different plant species (Czarnota et al. 2003; Warembourg et al. 2003). "
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    ABSTRACT: High bacterial density and diversity near plant roots has been attributed to rhizodeposit compounds that serve as both energy sources and signal molecules. However, it is unclear if and how specific rhizodeposit compounds influence bacterial diversity. We silenced the biosynthesis of isoflavonoids, a major component of soybean rhizodeposits, using RNA interference in hairy root composite plants, and examined changes in rhizosphere bacteriome diversity. We used successive sonication to isolate soil fractions from different rhizosphere zones at two different time points, and analyzed denaturing gradient gel electrophoresis profiles of 16S rRNA gene amplicons. Extensive diversity analysis of the resulting spatio temporal profiles of soybean bacterial communities indicated that indeed isoflavonoids significantly influenced soybean rhizosphere bacterial diversity. Our results also suggested a temporal gradient effect of rhizodeposit isoflavonoids on the rhizosphere. However, the hairy root transformation process itself significantly altered rhizosphere bacterial diversity, necessitating appropriate additional controls. Gene silencing in hairy root composite plants combined with successive sonication is a useful tool to determine the spatio temporal effect of specific rhizodeposit compounds on rhizosphere microbial communities.
    Molecular Plant-Microbe Interactions 10/2014; 28(1). DOI:10.1094/MPMI-08-14-0247-R · 3.94 Impact Factor
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    • "T-DNA transfer from A. tumefaciens to the plant cell requires activation of the vir regulon present on the Ti plasmid (Gelvin, 2006). Salicylic acid and ethylene, two plant signal molecules involved in the regulation of defence-regulated gene expression during plant–microbe interaction (Ecker and Davis, 1987; Prithiviraj et al., 2005) have been shown to inhibit expression of the vir regulon (Yuan et al., 2007; Nonaka et al., 2008). One possible hypothesis for the earlier appearance of tumours is that (hydroxylated) OLs can be detected by the plant and that the presence of (hydroxylated) OLs causes some kind of plant defence leading to the production of ethylene or salicylic acid. "
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    ABSTRACT: Ornithine lipids (OLs) are phosphorus-free membrane lipids that are widespread among Gram-negative bacteria. Their basic structure consists of a 3-hydroxy fatty acyl group attached in amide linkage to the α-amino group of ornithine and a second fatty acyl group ester-linked to the 3-hydroxy position of the first fatty acid. It has been shown that OLs can be hydroxylated within the amide-linked fatty acyl moiety, the secondary fatty acyl moiety or within the ornithine moiety. These modifications have been related to increased stress tolerance and symbiotic proficiency in different organisms such as Rhizobium tropici or Burkholderia cenocepacia. Analysing the membrane lipid composition of the plant pathogen Agrobacterium tumefaciens we noticed that it forms two different OLs. In the present work we studied if OLs play a role in stress tolerance and pathogenicity in A. tumefaciens. Mutants deficient in the OLs biosynthesis genes olsB or olsE were constructed and characterized. They either completely lack OLs (ΔolsB) or only form the unmodified OL (ΔolsE). Here we present a characterization of both OL mutants under stress conditions and in a plant transformation assay using potato tuber discs. Surprisingly, the lack of agrobacterial OLs promotes earlier tumour formation on the plant host.
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