Analysis of mono, duo and multispecies biofilms of Enterococcus faecalis, Pseudomonas aeruginosa, and Staphylococcus
Nycole da Silva Guerra1, Belisa Avila Rodrigues1, Isadora Pinto Loureiro1
1Laboratório de Biofilmes e Modelos Alternativos (BioModal), Department of Basic Health Sciences, Federal University of Health Sciences of Porto Alegre (UFCSPA), Sarmento Leite, 245, Porto Alegre, Rio Grande do Sul, 90050-170, Brazil; Postgraduate Program in Biosciences, Federal University of Health Sciences of Porto Alegre (UFCSPA), Sarmento Leite, 245, Porto Alegre, Rio Grande do Sul, 90050-170, Brazil.
Abstract:
Pseudomonas aeruginosa, Staphylococcus aureus, and Enterococcus faecalis are nosocomial pathogens often associated with hard-to-treat infections due to their ability to form biofilms, a key virulence factor. Within biofilms, microbial interactions modulate virulence and antimicrobial resistance, resulting in therapeutic failures and poor patient outcomes. Biofilms account for nearly 80% of chronic infections, highlighting their persistence and difficulty of eradication. This study evaluated the interactions of P. aeruginosa, E. faecalis, and S. aureus in mono-, duo-, and multi-species cultures, focusing on growth, biofilm formation, and virulence in an in vivo model. In vitro assays assessed bacterial growth (OD620 nm) and mono, duo, and multispecies biofilms through biomass quantification (crystal violet), viable cell counts (CFU/mL), and cell viability (XTT). Additionally, infection assays using Caenorhabditis elegans were used to evaluate the virulence of these pathogens by monitoring host survival. Duo and multispecies biofilms showed greater growth than monospecies, except for S. aureus + E. faecalis. Biofilm biomass was also higher in most combinations, except for S. aureus + P. aeruginosa. E. faecalis predominated in CFU counts of duo- and multispecies biofilms, showing levels similar to S. aureus in biofilm-associated cells and to P. aeruginosa in planktonic cells. XTT results indicated a higher viability of S. aureus cells in the biofilm and P. aeruginosa in the planktonic form. Finally, in vivo infections showed higher mortality in monospecies groups, due to the complexity of interspecies interactions that prioritize survival and persistence within the biofilm over pathogenicity. This study provides, for the first time, the analysis of S. aureus, P. aeruginosa, and E. faecalis in multispecies biofilm and in vivo model infection, advancing the understanding of interspecies dynamics and biofilm-associated infections.
Insights
This study reveals how Pseudomonas aeruginosa, Staphylococcus aureus, and Enterococcus faecalis interact in biofilms, impacting infection outcomes. Interspecies dynamics within biofilms prioritize survival over pathogenicity, affecting treatment strategies for chronic infections.
Area of Science:
- Microbiology
- Infectious Diseases
- Biofilm Research
Background:
- Nosocomial pathogens like Pseudomonas aeruginosa, Staphylococcus aureus, and Enterococcus faecalis form biofilms, a key factor in persistent, hard-to-treat infections.
- Microbial interactions within biofilms significantly influence virulence and antimicrobial resistance, leading to treatment failures and adverse patient outcomes.
- Biofilms are implicated in approximately 80% of chronic infections, underscoring their resilience and the challenges in their eradication.
Purpose of the Study:
- To investigate the growth, biofilm formation, and virulence dynamics of P. aeruginosa, E. faecalis, and S. aureus in mono-, duo-, and multi-species cultures.
- To analyze interspecies interactions within biofilms using both in vitro and in vivo models.
- To advance the understanding of multispecies biofilm formation and its impact on pathogenicity and host survival.
Main Methods:
- In vitro assays included bacterial growth measurements (OD620 nm), biofilm biomass quantification (crystal violet), viable cell counts (CFU/mL), and cell viability (XTT).
- Interspecies interactions were assessed in mono-, duo-, and multi-species cultures.
- In vivo virulence was evaluated using Caenorhabditis elegans infection models to monitor host survival rates.
Main Results:
- Duo- and multispecies biofilms generally exhibited enhanced growth and biomass compared to monospecies biofilms, with exceptions noted for specific combinations.
- Enterococcus faecalis demonstrated predominant CFU counts in mixed biofilms, with varying levels compared to Staphylococcus aureus and Pseudomonas aeruginosa in different cellular states.
- In vivo studies indicated higher mortality in monospecies infections, suggesting that interspecies interactions in biofilms shift pathogen behavior towards persistence rather than acute pathogenicity.
Conclusions:
- Interspecies interactions within multispecies biofilms significantly modulate the virulence and survival strategies of P. aeruginosa, E. faecalis, and S. aureus.
- The complexity of biofilm communities influences pathogenicity, with survival and persistence being prioritized over aggressive infection in mixed-species settings.
- This research provides novel insights into the dynamics of S. aureus, P. aeruginosa, and E. faecalis in multispecies biofilms and their implications for treating chronic biofilm-associated infections.
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