A Synthetic Polymicrobial Community Biofilm Model Demonstrates Spatial Partitioning, Tolerance to Antimicrobial Treatment, Reduced Metabolism, and Small Colony Variants Typical of Chronic Wound Biofilms

Abstract

Understanding chronic wound infection is key for successful treatment and requires accurate laboratory models. We describe a modified biofilm flow device that effectively mimics the chronic wound environment, including simulated wound fluid, a collagen-based 3D biofilm matrix, and a five-species mixture of clinically relevant bacteria (Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli, Enterococcus faecalis, and Citrobacter freundii). Mixed biofilms were cultured for between 3 and 14 days with consistent numbers of bacteria that exhibited reduced metabolic activity, which increased with a high dose of glucose. S. aureus was recovered from biofilms as a small colony variant, but as a normal colony variant if P. aeruginosa was excluded from the system. Bacteria within the biofilm did not co-aggregate but formed discrete, species-specific clusters. Biofilms demonstrated differential tolerance to the topical antimicrobials Neosporin and HOCl, consistent with protection due to the biofilm lifestyle. The characteristics exhibited within this model match those of real-world wound biofilms, reflecting the clinical scenario and yielding a powerful in vitro tool that is versatile, inexpensive, and pivotal for understanding chronic wound infection.

Solving polymicrobial puzzles: evolutionary dynamics and future directions

“…Previous research indicates that within wound environments, anaerobic bacteria tend to inhabit the middle region, sustained by oxygen consumers (Choi et al, 2019). Compelling in vivo evidence supports the notion that P. aeruginosa predominantly colonizes the deeper layers of wounds, while S. aureus tends to establish itself on the wound's surface (Fazli et al, 2009;Chen et al, 2021;Khalid et al, 2023). Pouget et al, showed aggregated biofilms of both S. aureus and P. aeruginosa at the specific locations still maintaining a mixed polymicrobial biofilm with enhanced characteristics (Pouget et al, 2022).…”

A growing battlefield in the war against biofilm-induced antimicrobial resistance: insights from reviews on antibiotic resistance

“…In chronic wound infections, biofilms consisting of S. aureus and P. aeruginosa are commonly found together leading to delayed healing and persistent infections (Borges et al, 2022). Laboratory models developed to monitor biofilm colonization in chronic wounds and biopsies reveal that bacteria form species specific aggregates in distinct regions of the tissue (Khalid et al, 2023). Real-time detection of volatile metabolites can be used to discriminate between different microbial species in biofilms (Slade et al, 2022).…”