Fig. 4

PGL Promotes Intercellular Bacterial Transfer and Prevents Bacterial Clearance

(A) Mean (≥5 biological replicates) number of resident macrophages and monocytes occupying the HBV at each time point, quantified every 10 min from 1 to 11 hpi in Tg (mpeg1::yfp) fish with green fluorescent macrophages after infection with 1–3 wild-type or PDIM⁻ red fluorescent Mm.

(B) Percentage of fish where the infecting bacteria were phagocytosed by a resident macrophage (black) or a monocyte (gray) over the first 11 hr following infection of Tg (mpeg1:YFP) fish in the HBV with red fluorescent 1–3 wild-type or PDIM⁻ Mm. n = number of larvae per group. Significance testing done using Fisher’s exact test. Results representative of three independent experiments.

(C) Representative images from a time-lapse movie of a bacterial transfer event. Uninfected Hoechst positive (blue fluorescence) monocyte (yellow arrow) is seen phagocytosing an infected cell (yellow arrowhead). Scale bar, 50 μm. Time stamp, mpi.

(D) Representative images from a time-lapse movie showing an infected macrophage (green fluorescent) clearing red fluorescent PGL⁻ Mm (yellow arrowhead). Scale bar, 50μm. Time stamp, mpi. (See also Movies S1 and S2 and Tables S1 and S2.)

(E) Quantification of bacterial transfer events from experiments represented by (C) and (D). Percentage of animals demonstrating a transfer event during the designated imaging time block.

(F) Percentage of animals remaining infected over the first 5 days of infection with 1–3 wild-type, PGL⁻, or PDIM⁻ Mm into the HBV of wild-type fish. Numbers of fish infected with each Mm strain: 30 wild-type, 28 PGL⁻, and 28 PDIM⁻. Results representative of two separate experiments.

(G) Percentage of infected (black) or uninfected (gray) untreated, DMSO control, or QVD-OPH treated wild-type fish 5 dpi with 1-3 wild-type Mm into the HBV. n = number of larvae per group. Results representative of two separate experiments.