Cell aggregation promotes pyoverdine-dependent iron uptake and virulence in Pseudomonas aeruginosa

Abstract

In Pseudomonas aeruginosa the Gac signaling system and the second messenger cyclic diguanylate (c-di-GMP) participate in the control of the switch between planktonic and biofilm lifestyles, by regulating the production of the two exopolysaccharides Pel and Psl. The Gac and c-di-GMP regulatory networks also coordinately promote the production of the pyoverdine siderophore, and the extracellular polysaccharides Pel and Psl have recently been found to mediate c-di-GMP-dependent regulation of pyoverdine genes. Here we demonstrate that Pel and Psl are also essential for Gac–mediated activation of pyoverdine production. A pel psl double mutant produces very low levels of pyoverdine and shows a marked reduction in the expression of the pyoverdine-dependent virulence factors exotoxin A and PrpL protease. While the exopolysaccharide-proficient parent strain forms multicellular planktonic aggregates in liquid cultures, the Pel and Psl-deficient mutant mainly grows as dispersed cells. Notably, artificially induced cell aggregation is able to restore pyoverdine-dependent gene expression in the pel psl mutant, in a way that appears to be independent of iron diffusion or siderophore signaling, as well as of recently described contact-dependent mechanosensitive systems. This study demonstrates that cell aggregation represents an important cue triggering the expression of pyoverdine-related genes in P. aeruginosa, suggesting a novel link between virulence gene expression, cell–cell interaction and the multicellular community lifestyle.

Interdependence between iron acquisition and biofilm formation in Pseudomonas aeruginosa

“…Although the mechanism of biofilm-mediated regulation of pyoverdine remains unclear, there is a strong correlation between biofilm formation and cell aggregation (Visaggio et al ., 2015; Kang and Kirienko, 2017). This was initially identified and investigated by Imperi and colleagues, who demonstrated that the exopolysaccharides Pel and Psl were essential for planktonic cell aggregation and pyoverdine production (Visaggio et al ., 2015).…”

“…This was initially identified and investigated by Imperi and colleagues, who demonstrated that the exopolysaccharides Pel and Psl were essential for planktonic cell aggregation and pyoverdine production (Visaggio et al ., 2015). Artificially inducing cell aggregation, by adding agar to the media for example, was shown to restore pyoverdine in pel psl double mutants (Visaggio et al ., 2015).…”

“…This was initially identified and investigated by Imperi and colleagues, who demonstrated that the exopolysaccharides Pel and Psl were essential for planktonic cell aggregation and pyoverdine production (Visaggio et al ., 2015). Artificially inducing cell aggregation, by adding agar to the media for example, was shown to restore pyoverdine in pel psl double mutants (Visaggio et al ., 2015). Similarly, we observed that the supplementation of the quorum-sensing molecule Pseudomonas quinolone signal (PQS) to media rapidly stimulated aggregation of planktonic cells, causing high levels of pyoverdine (Kang et al ., 2017).…”

Role of Iron Uptake Systems in Pseudomonas aeruginosa Virulence and Airway Infection

“…For ToxA detection, cell-free culture supernatants were supplemented with 6ϫ SDS-PAGE loading dye (375 mM Tris-HCl [pH 6.8], 9% SDS, 50% glycerol, 0.03% bromophenol blue). The volumes of supernatants (or appropriate dilutions) loaded in SDS-PAGE gels were normalized to bacterial growth according to the following formula: loading volume (microliters) ϭ 10/OD 600 for the corresponding bacterial culture (49). Proteins resolved by SDS-PAGE were electrotransferred to a nitrocellulose filter (Hybond-C Extra; Amersham) and probed for ToxA by using a rabbit polyclonal anti-ToxA antibody (Sigma-Aldrich).…”

Bacillus cereus growth and biofilm formation: the impact of substratum, iron sources, and transcriptional regulator Sigma 54

“…Although these biofilms may contain low number of viable cells, from a contamination perspective, this could still be relevant as cells within the biofilm are protected against disinfectants by the biomass of dead cells. In P. aeruginosa cell aggregation, either via growth in a colony or by means of exopolysaccharide (EPS) production, has been shown to trigger the production of the siderophore pyoverdine and other virulence factors (Visaggio et al 2015). In Staphylococcus lugdunensis, the cell wall bound IsdC protein involved in iron transport promoted cell aggregation in iron depleted condition (Missineo et al 2014).…”