The guanidinobutyrase GbuA is essential for the alkylquinolone‐regulated pyocyanin production during parasitic growth of Pseudomonas aeruginosa in co‐culture with Aeromonas hydrophila

Abstract

The opportunistic pathogen Pseudomonas aeruginosa controls the production of virulence factors by quorum sensing (QS). Besides cell density, QS in P. aeruginosa is co-regulated by metabolic influences, especially nutrient limitation. Previously, a co-culture model system was established consisting of P. aeruginosa and the chitinolytic bacterium Aeromonas hydrophila, in which parasitic growth of P. aeruginosa is strictly dependent on the QS-controlled production of pyocyanin in response to nutrient limitation (Jagmann et al., ). In this study, the co-culture was employed to identify novel genes involved in the regulation of pyocyanin production. Via transposon mutagenesis, the gene gbuA encoding a guanidinobutyrase was identified, deletion of which led to a loss of pyocyanin production in co-cultures and to a reduced pyocyanin production in single cultures. Addition of the natural substrate of GbuA to the mutant strain enhanced the negative effect on pyocyanin production in single cultures. The gbuA mutant showed a reduced transcription of the pqsABCDE operon and could be complemented by PqsE overexpression and addition of alkylquinolone signal molecules. The strong effect of gbuA deletion on the QS-controlled pyocyanin production in co-cultures showed the value of this approach for the discovery of novel gene functions linking metabolism and QS in P. aeruginosa.

The aliphatic amidase AmiE is involved in regulation of Pseudomonas aeruginosa virulence

“…Remarkably, AmiE over-production induced also a decrease in PQS synthesis at least through a lower accumulation of i) its biosynthetic enzymes PqsB, PqsC and PqsD, and ii) the kynunerinase KinU (Supplementary Table S1) that is involved in the degradation of kynurenine leading to the production of the PQS precursor anthranilate44. Taken together, these data suggest strong relationships between AmiE and QS signals production, thus linking metabolism and QS in P. aeruginosa , as previously suggested43.…”

“…This could explain the strong production level decrease of 3-oxo-C 12 -HSL. In parallel it is interesting to note that retrieving AmiE’s in silico functional interactions from STRING database42, led to predict interactions of AmiE with PA14_63110, as well as with the guanidinobutyrase (GbuA), a mutant of which was shown to be strongly affected in PQS production43. Remarkably, AmiE over-production induced also a decrease in PQS synthesis at least through a lower accumulation of i) its biosynthetic enzymes PqsB, PqsC and PqsD, and ii) the kynunerinase KinU (Supplementary Table S1) that is involved in the degradation of kynurenine leading to the production of the PQS precursor anthranilate44.…”

LaoABCR, a Novel System for Oxidation of Long-Chain Alcohols Derived from SDS and Alkane Degradation in Pseudomonas aeruginosa

“…Purified genomic DNA of strain PAO1 (Puregene tissue core kit B; Qiagen, Hilden, Germany) was used as the template. Deletion mutants were generated via splicing by overlap extension (SOE) PCR (60). The required plasmids for the gene deletions were constructed with genomic DNA flanking the respective sequence of interest by ligating them into the vector pEX18Ap (Table 1) within the restriction sites XbaI/BamHI and HindIII (primers 1 to 20; Table 2).…”

2,3‐Butanediol catabolism in Pseudomonas aeruginosa PAO1

“…S12). While pyocyanin and pyoverdine can drive the production of 2,3-BD by the cohabiting fermenters (Filkins et al, 2015), the loss of the pyocyanin production in P. aeruginosa PAO1 ΔgbuA mutant can lower the transcription of PA4148-PA4153 in 2,3-BD catabolism pathway (Jagmann et al, 2016). Thus, catabolism of 2,3-BD by P. aeruginosa through the pathway identified in this work might enhance the anabolism of 2,3-BD by cohabiting fermenters and have a role in its competition with other microorganisms in the airways of CF patients.…”

“…P. aeruginosa is a widespread and metabolically versatile gram-negative bacterium that typically exists in the soil and in aqueous environments (Stover et al, 2000;Jagmann et al, 2016). It is also an important opportunistic pathogen that can cause serious respiratory infections, especially in cystic fibrosis (CF) patients (Feinbaum et al, 2012;Korgaonkar et al, 2013).…”