Fig. 6: Biofilm differentially responds to distinct immune pressures.

A Integrated UMAP plot of biofilms co-cultured with MΦs, G-MDSCs, and PMNs. The biofilm control UMAP (Fig. 4B) was used as a template on which the co-culture samples were projected (using the ingest function of Scanpy). To the right, each individual co-culture condition is colored separately, with the other two co-culture conditions shown in yellow. B Bar plot showing the percentage of cells from each biofilm co-culture condition within each cluster. The pie chart shows combined cluster distributions for all co-culture samples. C (Left) Overlay of total mRNA counts on the integrated UMAP plot, with the highest number of captured transcripts present in cluster 1. (Middle) Trajectory analysis with the integrated biofilm-leukocyte co-culture samples (Palantir algorithm), which identified a differentiation pathway that converges to cluster 1 upon leukocyte exposure. (Right) Expression of atpA/B/C/D genes is concentrated in cluster 1, where the trajectory converges. D Transcriptional regulatory category expression within cluster 1 for biofilms co-cultured with immune cells compared to the biofilm control. Categories correspond to the schematic in Fig. 3A. Red dashed lines depict the average of the biofilm control for reference. E Top genes activated in biofilm cluster 1 in response to leukocyte exposure. Additional genes are listed in Supplementary Data 10–12. All genes have FDR-adjusted p-value ≤ 0.05 for differential expression (log₂ fold-change) compared to the other co-culture conditions. F–G Evaluation of respiration and ROS activity using CTC (F) and CM-H₂DCFDA (G) dyes (red) in biofilm (green)-leukocyte (blue) co-cultures with MΦs, G-MDSCs, or PMNs. Z-stack images were acquired (1 μm sections) and used to construct 3-D images. Color adjustments were applied uniformly across all images of the same experiment. Source data are provided as a Source Data file.