Cell Wall Proteome of Candida albicans Reveals Proteins Associated with Tolerance to Antibiofilm Activity of a Lippia graveolens Kunth Stem Extract

<i>Candida albicans</i> is an opportunistic fungus capable of forming biofilms that are resistant to conventional antifungal treatments. This study evaluated the antibiofilm activity of an ethanolic extract from Mexican oregano (<i>Lippia graveolens</i> Kunth) stem and its impact on the protein composition of the <i>C. albicans</i> cell wall (CW). The proteomic analysis was restricted to the adherent cells that persisted after treatment, representing the more tolerant subpopulation. First, the biofilm-forming capacity of 18 clinical isolates was characterized. Subsequently, the effect of the <i>Lippia graveolens</i> Kunth stem extract on biofilm formation was assessed in clinical isolates of <i>C. albicans</i> with high and moderate biofilm-forming capacities. The results showed an MIC₉₀ of 16 mg/mL against <i>C. albicans</i> isolates in planktonic growth. Furthermore, the extract exhibited an antibiofilm effect, showing a 77% inhibition in the highly biofilm-forming <i>C. albicans</i> 2400H strain at 1.6 mg/mL. To assess how the extract influences CW composition, we analyzed the CW proteome of the adherent biofilm cells of <i>C. albicans</i> that persisted after exposure to the stem extract. Mass spectrometry identified 1006 total proteins, where 156 were up-regulated, and 94 were down-regulated. Of the differentially expressed proteins, we identify 40 CW proteins (CWP's) associated with dimorphic transition, adhesion, oxidative stress, and biofilm formation. These findings provide the potential of the <i>L. graveolens</i> Kunth stem as a natural antifungal agent against <i>C. albicans</i> biofilms. In addition, we identified CWP's associated with tolerance to the extract's antibiofilm activity, offering mechanistic insight into persistence and potential targets for improving antibiofilm interventions.