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This study focused on using photodynamic inactivation (PDI) to eliminate mixed biofilms of C. albicans\u2013S. aureus formed on poly (urethane) (PU) discs functionalized with a nanocomposite layer containing phloxine B (PhB). Additionally, the effect of PDI on the ALS3 and HWP1 genes of C. albicans was examined in mixed biofilms. Spectral analysis showed a continuous release of PhB from the nanocomposite in Mueller\u2013Hinton broth within 48 h, with a released amount of PhB &lt; 5% of the total amount. The anti-biofilm effectiveness of the light-activated nanocomposite with PhB showed a reduction in the survival rate of biofilm cells to 0.35% and 31.79% for S. aureus and C. albicans, respectively, compared to the control biofilm on PU alone. Scanning electron microscopy images showed that the nanocomposite effectively reduced the colonization and growth of the mixed biofilm. While PDI reduced the regulation of the ALS3 gene, the HWP1 gene was upregulated. Nevertheless, the cell survival of the C. albicans\u2013S. aureus biofilm was significantly reduced, showing great potential for the elimination of mixed biofilms.<\/jats:p>","DOI":"10.3390\/jof11080582","type":"journal-article","created":{"date-parts":[[2025,8,5]],"date-time":"2025-08-05T10:50:21Z","timestamp":1754391021000},"page":"582","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Mixed Candida albicans\u2013Staphylococcus aureus Biofilm Is Reduced by Light-Activated Nanocomposite with Phloxine B"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5161-0157","authenticated-orcid":false,"given":"Jarmila","family":"Czucz Varga","sequence":"first","affiliation":[{"name":"Department of Microbiology and Virology, Faculty of Natural Sciences, Comenius University in Bratislava, Mlynsk\u00e1 dolina, Ilkovi\u010dova 6, 842 15 Bratislava, Slovakia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Juraj","family":"Bujd\u00e1k","sequence":"additional","affiliation":[{"name":"Department of Physical and Theoretical Chemistry, Faculty of Natural Sciences, Comenius University in Bratislava, Mlynsk\u00e1 dolina, Ilkovi\u010dova 6, 842 15 Bratislava, Slovakia"},{"name":"Institute of Inorganic Chemistry, Slovak Academy of Sciences, D\u00fabravsk\u00e1 cesta 9, 845 36 Bratislava, Slovakia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9219-0803","authenticated-orcid":false,"given":"Helena","family":"Bujd\u00e1kov\u00e1","sequence":"additional","affiliation":[{"name":"Department of Microbiology and Virology, Faculty of Natural Sciences, Comenius University in Bratislava, Mlynsk\u00e1 dolina, Ilkovi\u010dova 6, 842 15 Bratislava, Slovakia"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,8,5]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"e00043-22","DOI":"10.1128\/mmbr.00043-22","article-title":"White-Opaque Switching in Candida albicans: Cell Biology, Regulation, and Function","volume":"88","author":"Soll","year":"2024","journal-title":"Microbiol. 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