Fig. 4.

Fig. 4. Neutrophils influence granuloma mycobacterial burden via devR regulon modulation.

(A) PCA of M. marinum transcriptome from WT and Rac2D57N granulomas and log-phase broth culture. (B) Volcano plot showing the differential expression of M. marinum genes in Rac2D57N versus WT granulomas. Horizontal line: P_adj of 0.1; vertical lines: twofold changes. (C to E) Heatmaps for genes associated with antibiotic resistance, NAD⁺ biosynthesis, and devR regulon in M. marinum from WT and Rac2D57N granulomas. (F) Box plots showing median log-normalized counts of representative transcripts from (C) to (E). Each data point represents log-normalized count for the transcript from each experiment. Benjamini-Hochberg (BH) correction for adjusted P values. (G) Bacterial burden in WT M. marinum and devR mutant–infected granulomas from WT fish, as mean arbitrary fluorescence units (AU) (WT M. marinum 36 granulomas; devR mutant 33 granulomas). Unpaired t test with Welch’s correction. (H) Images showing WT M. marinum and devR mutant burden in granulomas from infected WT fish. (I) Granuloma bacterial burden in WT and Rac2D57N fish infected with WT M. marinum (red data points) or M. marinum overexpressing devR/devS (cyan data points). Mean AU. WT M. marinum infection: 32 granulomas from WT fish; 27 granulomas from Rac2D57N fish. M. marinum devRS overexpressor infection: 35 granulomas from WT fish; 36 granulomas from Rac2D57N fish. Brown-Forsythe and Welch analysis of variance (ANOVA) tests, followed by Dunnett’s T3 multiple comparisons test. (J) Images showing the burden of WT M. marinum and M. marinum devRS overexpressor in granulomas from WT and Rac2D57N fish. [(G) and (I)] Granulomas from 14 dpi animals. Each point represents mycobacterial fluorescence from a single granuloma; granulomas from three animals per group. Error bars indicate SD. [(H) and (J)] Scale bars, 50 μm.