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Cell-associated hemolysis activity in the clinical strain of Pseudomonas fluorescens MFN1032
Daniel Sperandio1, Gaelle Rossignol, Josette Guerillon
1Laboratory of cold microbiology signals and the microenvironment, LMDF-SME, UPRES EA 4312, University of Rouen, 55 rue Saint Germain, 27000 Evreux, France.
Background:
MFN1032 is a clinical Pseudomonas fluorescens strain able to grow at 37 degrees C. MFN1032 cells induce necrosis and apoptosis in rat glial cells at this temperature. This strain displays secretion-mediated hemolytic activity involving phospholipase C and cyclolipopeptides. Under laboratory conditions, this activity is not expressed at 37 degrees C. This activity is tightly regulated and is subject to phase variation.
Results:
We found that MFN1032 displays a cell-associated hemolytic activity distinct from the secreted hemolytic activity. Cell-associated hemolysis was expressed at 37 degrees C and was only detected in vitro in mid log growth phase in the presence of erythrocytes. We studied the regulation of this activity in the wild-type strain and in a mutant defective in the Gac two-component pathway. GacS/GacA is a negative regulator of this activity. In contrast to the Pseudomonas fluorescens strains PfO-1 and Pf5, whose genomes have been sequenced, the MFN1032 strain has the type III secretion-like genes hrcRST belonging to the hrpU operon. We showed that disruption of this operon abolished cell-associated hemolytic activity. This activity was not detected in P.fluorescens strains carrying similar hrc genes, as for the P. fluorescens psychrotrophic strain MF37.
Conclusions:
To our knowledge this the first demonstration of cell-associated hemolytic activity of a clinical strain of Pseudomonas fluorescens. Moreover, this activity seems to be related to a functional hrpU operon and is independent of biosurfactant production. Precise link between a functional hrpU operon and cell-associated hemolytic activity remains to be elucidated.
Insights
This study reveals a novel cell-associated hemolytic activity in clinical Pseudomonas fluorescens MFN1032 at 37°C, linked to the hrpU operon and distinct from secreted toxins.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Molecular Biology
Background:
- Clinical Pseudomonas fluorescens strain MFN1032 grows at 37°C, inducing necrosis and apoptosis in rat glial cells.
- This strain exhibits secretion-mediated hemolytic activity (phospholipase C, cyclolipopeptides), but it's not expressed at 37°C.
- Secreted hemolytic activity is tightly regulated and subject to phase variation.
Purpose of the Study:
- To characterize a distinct hemolytic activity of Pseudomonas fluorescens MFN1032 at 37°C.
- To investigate the genetic regulation of this cell-associated hemolytic activity.
- To determine the relationship between this activity and known virulence factors or secretion systems.
Main Methods:
- Comparative analysis of hemolytic activity in wild-type and mutant MFN1032 strains.
- In vitro assays using erythrocytes to detect cell-associated hemolysis at 37°C.
- Genetic analysis involving disruption of the hrpU operon and comparison with other P. fluorescens strains.
Main Results:
- A novel cell-associated hemolytic activity was identified in MFN1032 at 37°C, expressed during mid-log phase with erythrocytes.
- The GacS/GacA two-component system acts as a negative regulator of this cell-associated hemolysis.
- Disruption of the type III secretion-like hrpU operon abolished cell-associated hemolytic activity; this activity was absent in other P. fluorescens strains lacking these genes.
Conclusions:
- This is the first report of cell-associated hemolytic activity in a clinical Pseudomonas fluorescens strain.
- The activity is linked to a functional hrpU operon and is independent of biosurfactant production.
- The precise molecular mechanism connecting the hrpU operon to cell-associated hemolysis requires further investigation.