Fig. 3: Prefrailty associated microb-aging is linked with pathogenic traits of human biofilms.

Feces from individuals aged 30–70 years, 70+ years robust, and 70+ years prefrail, were cultured in vitro as a polymicrobial anaerobic biofilm. A, B The rate of dispersal of planktonic bacteria, biofilm-dispersed bacteria, during a 24-hour period from biofilms was measured. Raw values were normalized to the average dispersal rate of the 30–70 years group. D, E Normalized counts of biofilm-dispersed bacteria were cultured on the apical surface of the human Caco2/HT29MTX monolayer. After a 4-hour coculture, the number of bacteria adhered to the epithelium was quantified. A–D Pearson correlations between biofilm rate of dispersal (A) or adhesiveness (D) with the age of donors are presented. (B–E) Scatter plots depict for each individual’s its corresponding value of biofilm dispersal rate (B) or adhesiveness (E) and were grouped for each three cohort of individuals. B, E each dot is the average value of an individual donor biofilm (at least 12 replicates). The gender of each donor is indicated using open(male)/filled(female) circles. Statistical significance was determined by ANOVA followed by Tukey’s for multiple comparisons, where P < 0.05 was considered significant. C–F Heatmaps represent Kendall (F) or Pearson (C) correlation coefficients between biofilm pathogenic traits and clinical data (C) or biofilm taxa counts (F). Clinical parameters include discrete and continuous clinical features: household (1: solo 2: with a partner), gender (1: male, 2: female), MNA score, Charlson index, age, and Fried Frailty Index. The color gradient ranges from blue (negative) to red (positive) correlations, with statistically significant associations (P < 0.05) marked with *. For enhanced clarity, the heatmap in F was filtered to include only taxa rank with significantly positive or negative correlations (P < 0.05).