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Enterococcus spp. produces slime and survives in rat peritoneal macrophages
L Baldassarri1, R Cecchini, L Bertuccini
1Laboratorio di Ultrastrutture, Istituto Superiore di Sanità, Rome, Italy. baldassa@iss.it
Abstract:
Enterococcal clinical isolates were investigated for the ability to form biofilm on inert surfaces, as a measure of slime production, in an attempt to find new possible virulence factors for these microorganisms. This property was commonly found among Enterococcus faecalis. Also E. faecium isolates were able to form biofilm, although to a lesser extent; for this species, however, biofilm formation seemed more frequently associated with isolates from infection rather than with environmental strains or isolates from healthy individuals. Biofilm formation was strongly affected by the presence of an additional carbohydrate source in the medium, or by iron deprivation, indicating a role of slime for survival in stressful conditions. Slime-producing E. faecalis were able to survive inside peritoneal macrophages for extended periods compared to slime-negative strains or to slime-positive bacteria grown in conditions depressing slime production. In particular, slime-producing and slime-negative cells showed a decrease of 1 and 2 log units, respectively, at 1 h after infection; slime-negative cells were then rapidly killed, with clearance of bacterial cells at 24 h. Slime-producing bacteria persisted up to 48 h, which was the last time point examined, as after that time viability of both infected and non-infected macrophages started to decline. Scanning electron microscopy observations showed the presence of abundant amorphous extracellular material, of possible polysaccharide nature, embedding bacterial cells to form a multilayered biofilm. Even in conditions not supporting biofilm formation, bacterial cells appeared capsulated, suggesting that capsule and slime might represent different structures. Genes belonging to the epa locus or to a putative icaA homolog did not seem to be involved in synthesis and export of slime.
Insights
Slime production, a key virulence factor in Enterococcus, enhances bacterial survival within macrophages. This biofilm formation ability is common in Enterococcus faecalis and linked to infections in Enterococcus faecium.
Area of Science:
- Microbiology
- Bacterial Pathogenesis
- Biofilm Formation
Background:
- Enterococcal infections pose significant public health challenges.
- Understanding bacterial virulence factors is crucial for developing effective treatments.
- Biofilm formation is a potential virulence factor in Enterococcus species.
Purpose of the Study:
- To investigate biofilm formation as a slime production indicator in Enterococcal clinical isolates.
- To explore the role of slime production in Enterococcal virulence, particularly in host-pathogen interactions.
- To identify factors influencing biofilm formation and its association with infection.
Main Methods:
- Culturing of Enterococcal isolates (Enterococcus faecalis, Enterococcus faecium) on inert surfaces.
- Assessing biofilm formation quantitatively.
- Evaluating the impact of nutrient availability (carbohydrate source) and iron levels on biofilm production.
- Investigating bacterial survival within peritoneal macrophages using in vitro infection models.
- Microscopic analysis (Scanning Electron Microscopy) of biofilm structure.
Main Results:
- Biofilm formation was prevalent in Enterococcus faecalis and observed to a lesser extent in Enterococcus faecium, especially in clinical isolates.
- Slime production was enhanced by additional carbohydrate sources and iron deprivation, suggesting a role in stress survival.
- Slime-producing Enterococcus faecalis demonstrated significantly enhanced survival within macrophages compared to slime-negative strains.
- Slime-negative bacteria were rapidly cleared, while slime-producing bacteria persisted for extended periods (up to 48 hours).
- Scanning Electron Microscopy revealed multilayered biofilms with extracellular matrix, distinct from capsular structures.
Conclusions:
- Slime production, indicated by biofilm formation, is a significant virulence factor for Enterococcus species, particularly Enterococcus faecalis.
- Biofilm formation contributes to bacterial resistance against host immune defenses, specifically macrophage clearance.
- Environmental conditions like nutrient availability and iron levels modulate slime production and bacterial survival.
- The study suggests slime and capsule are distinct structures, and genes like epa or icaA homologs are not directly involved in slime synthesis.