Journal: mSphere

Article Title: Enterococcus faecalis induces H₂O₂-mediated epithelial cell death and enhances Candida albicans virulence in oropharyngeal candidiasis

doi: 10.1128/msphere.00822-25

Figure Lengend Snippet: C. albicans enhances enterococcal H₂O₂ release, and catalase limits fungal oxidative damage. ( A ) Kinetics of extracellular H₂O₂ release during aerobic culture of E. faecalis , C. albicans, and their combination in BHI broth media. H 2 O 2 in culture supernatants was measured with the Amplex Red hydrogen peroxide/peroxidase assay kit. E. faecalis alone (strain Ef13), C. albicans alone (strain SC5314), and paraformaldehyde-fixed C. albicans were used as controls. Results represent means ± SD from three independent experiments. Live C. albicans significantly increased H 2 O 2 from E. faecalis after 60 and 90 min of interaction. *** P < 0.001, **** P < 0.0001 (uncorrected two-way ANOVA with Fisher’s LSD test). ( B ) Effect of C. albicans catalase on oxidative damage from E. faecalis. C. albicans strain SN250 or an isogenic catalase homozygous deletion mutant ( cat1 Δ/Δ strain(X)) was allowed to interact with E. faecalis for 2 h at 1:10 fungal:bacterial ratio, and fungal metabolic activity was measured by the XTT assay. Catalase (40 U/well) was added to rescue the mutant phenotype. Fungal metabolic activity was expressed as %fungal viability = x / y *100, where x is the OD 450 of C. albicans with E. faecalis, and y is the OD 450 of C. albicans only. The metabolic activity of the catalase mutant is significantly compromised by E. faecalis , while the metabolic activity of the reference strain is not significantly affected. Adding catalase rescues the mutant from oxidative damage. Results represent means ± SD from three independent experiments with technical triplicates. P values shown are from Brown-Forsythe uncorrected one-way ANOVA (Welch’s t test correction).

Article Snippet: C. albicans strain SC5314 (ATCC MYA-2876) was sub-cultured in Yeast Extract (Sigma, USA)-Peptone (Gibco, USA)-Dextrose (J.T.Baker, US) (YPD) broth and incubated aerobically at 30°C in an orbital shaker, overnight.

Techniques: Mutagenesis, Activity Assay, XTT Assay

Journal: mSphere

Article Title: Enterococcus faecalis induces H₂O₂-mediated epithelial cell death and enhances Candida albicans virulence in oropharyngeal candidiasis

doi: 10.1128/msphere.00822-25

Figure Lengend Snippet: Co-infection with E. faecalis and C. albicans increases oral epithelial cell cytotoxicity. ( A ) Apoptosis: OKF6-TERT2 keratinocytes were exposed for 24 h to C. albicans SC5314, alone or together with Ef13 (1:10 fungal-bacteria cell ratio), with/without catalase (40 U/well). Apoptosis was quantified by luminescence using the RealTime-Glo Annexin V assay. Bars show mean luminescence A.U. ×1,000 ± SD in a representative of three independent experiments with technical triplicates. Dual-species infection did not increase apoptosis over the Ef13 mono-infection. Apoptosis in single- and dual-species infection was completely inhibited by catalase, confirming the role of H 2 O 2 in this process. ( B ) Necrosis. Necrosis was quantified in the same cultures after 48 h of infection by measuring fluorescence. Bars show fluorescence A.U. ×1,000 ± SD in a representative of three independent experiments, with technical triplicates. Ef13 alone significantly increased necrosis, and catalase attenuated this effect ( P < 0.0001). Dual infection induced greater necrosis than either organism alone ( P = 0.0001 vs Ef13; P < 0.0002 vs C. albicans ), and catalase partially rescued the epithelial cells from necrosis, showing that H 2 O 2 is partially responsible for necrosis. All P values are from Brown-Forsythe uncorrected one-way ANOVA (Welch’s t test correction).

Article Snippet: C. albicans strain SC5314 (ATCC MYA-2876) was sub-cultured in Yeast Extract (Sigma, USA)-Peptone (Gibco, USA)-Dextrose (J.T.Baker, US) (YPD) broth and incubated aerobically at 30°C in an orbital shaker, overnight.

Techniques: Infection, Bacteria, Annexin V Assay, Fluorescence