| Summary: | Biofilms are a major problem in hard-to-heal wounds. Moreover, they are composed of different species and are often tolerant to antimicrobial agents. At the same time, interspecific synergy and/or competition occurs when some bacterial species clash. For this reason, the tolerance of two dual-species wound biofilm models of <i>Pseudomonas aeruginosa</i> and <i>Staphylococcus aureus</i> or <i>Enterococcus faecium</i> against antimicrobials and antimicrobial dressings were analyzed quantitatively and by confocal laser scanning microscopy (CLSM). The results were compared to findings with planktonic bacteria. Octenidine-dihydrochloride/phenoxyethanol and polyhexamethylene biguanide (PHMB) irrigation solutions showed a significant, albeit delayed reduction in biofilm bacteria, while the PHMB dressing was not able to induce this effect. However, the cadexomer-iodine dressing caused a sustained reduction in and killed almost all bacteria down to 10<sup>2</sup> cfu/mL within 6 days compared to the control (10<sup>10</sup> cfu/mL). By means of CLSM in untreated human biofilm models, it became evident that <i>P. aeruginosa</i> dominates over <i>E. faecium</i> and <i>S. aureus</i>. Additionally, <i>P. aeruginosa</i> appeared as a vast layer at the bottom of the samples, while <i>S. aureus</i> formed grape-like clusters. In the second model, the distribution was even clearer. Only a few <i>E. faecium</i> were visible, in contrast to the vast layer of <i>P. aeruginosa</i>. It seems that the different species avoid each other and seek their respective niches. These mixed-species biofilm models showed that efficacy and tolerance to antimicrobial substances are nearly species-independent. Their frequent application appears to be important. The bacterial wound biofilm remains a challenge in treatment and requires new, combined therapy options.
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