Association of Hemolytic Activity ofPseudomonas entomophila, a Versatile Soil Bacterium, with Cyclic Lipopeptide Production

Abstract

Pseudomonas entomophila is an entomopathogenic bacterium that is able to infect and kill Drosophila melanogaster upon ingestion. Its genome sequence suggests that it is a versatile soil bacterium closely related to Pseudomonas putida. The GacS/GacA two-component system plays a key role in P. entomophila pathogenicity, controlling many putative virulence factors and AprA, a secreted protease important to escape the fly immune response. P. entomophila secretes a strong diffusible hemolytic activity. Here, we showed that this activity is linked to the production of a new cyclic lipopeptide containing 14 amino acids and a 3-C 10 OH fatty acid that we called entolysin. Three nonribosomal peptide synthetases (EtlA, EtlB, EtlC) were identified as responsible for entolysin biosynthesis. Two additional components (EtlR, MacAB) are necessary for its production and secretion. The P. entomophila GacS/GacA two-component system regulates entolysin production, and we demonstrated that its functioning requires two small RNAs and two RsmA-like proteins. Finally, entolysin is required for swarming motility, as described for other lipopeptides, but it does not participate in the virulence of P. entomophila for Drosophila. While investigating the physiological role of entolysin, we also uncovered new phenotypes associated with P. entomophila, including strong biocontrol abilities.

Genetic and Functional Characterization of Cyclic Lipopeptide White-Line-Inducing Principle (WLIP) Production by Rice Rhizosphere Isolate Pseudomonas putida RW10S2

“…2 is quite similar to those of some three-membered NRPS systems identified in Pseudomonas strains: a first NRPS gene is linked with a cognate luxRlike gene but not linked with the two other NRPS genes that occur as a tandem in a separate operon. This analogy in genomic organization with the genes involved in biosynthesis of a viscosin-like LP (9), massetolide A (8), and entolysin (50) suggests that the three RW10S2 NRPSs equally operate consecutively to synthesize a nonribosomal peptide.…”

“…This putative regulatory gene was located upstream of the first NRPS gene, positioned divergently to it. The RW10S2 luxR-like gene product showed about 70% identity with the NRPS-linked LuxR-type regulators PsoR of P. putida PCL1445 (13) and EtlR of P. entomophila (50).…”

“…According to Balibar et al (4), the D configuration of amino acids in the Pseudomonas LP arthrofactin is generated by a specific condensation domain, with additional epimerization activity, acting on the amino acid loaded on the previous module. Such C/E domains constitute a distinct cluster separated from the clusters with regular condensation domains and the lipo-initiation domain cluster in phylogenetic analysis of their sequences (8,41,50). Application of this sequence comparison to the RW10S2 C domains (see Fig.…”

Screening concepts, characterization and structural analysis of microbial-derived bioactive lipopeptides: a review

“…[12,[117][118][119] For example, purification of LP by HPLC was carried out by reversed phase (RP)-HPLC using a semi-preparative C18 column and 0.1% trifluoroacetic acid/methanol/H 2 O as a mobile phase. The elution was monitored using UV detection at 214 and 280 nm.…”

Novel Antiphytopathogenic Compound 2-Heptyl-5-Hexylfuran-3-Carboxylic Acid, Produced by Newly Isolated Pseudomonas sp. Strain SJT25

“…P. putida was found to be an effective biocontrol agent (15,25,27). P. entomophila, being close to P. putida at the genome sequence level (31), is an entomopathogenic bacterium that has been previously reported to produce substances toxic for insects (4). P. entomophila also has ACC deaminase activity and can use ACC as a sole nitrogen source (13).…”