The Secret Life of the Anthrax Agent Bacillus anthracis: Bacteriophage-Mediated Ecological Adaptations
Figure 9
Phage-regulated loci in B. anthracis.
(A) Promoter-gfp fusions expressed on worm agar. Bacteria were plated for 16 hours at 30°C and resulting colonies are shown in 200X phase-contrast and fluorescence images (0.5 second exposures). Strains are ΔSterne or its Wip4 lysogen (“/Wip4”) transformed with pASD2 encoding gfp alone or indicated promoter fusions (P). (B) Expression of PBA3443-gfp on BHI agar in either the ΔSterne/Wip4 background or ΔSterne co-transformed with the compatible vector pWH1520 (either without and insert or that bearing bcp25,26 or wip48,49). Bacteria were plated for 16 hours at 30°C and colonies are shown in 200X phase-contrast or fluorescence images with 0.5 second exposures. 2000X fluorescence images of representative bacteria are shown with exposure times. RFUs are averages of three experiments for ∼1×108 mid-log phase cells grown in BHI and suspended in buffer. (C) BA3443 and earthworm colonization. One month after colonization, bacteria were recovered, visualized, and enumerated. Strains are ΔSterne/Wip4 transformed with indicated vectors or the ΔSterne/Wip4 mutant ΔBA3443. Hind-guts were examined by phase-contrast or fluorescence microscopy at 200X or 2000X magnification. Exposure times are indicated in 2000X images. Average CFUs per gram of gut is shown from three experiments, indicating total viability (vegetative cells and spores) and spores alone. (D) B. anthracis-encoded loci required in the soil. Bacteria were recovered and enumerated at the indicated times from inoculated soil microcosms. Strains include ΔSterne/Wip4 with pASD2 alone or the indicated mutations. Values are mean averages (n = 3) and error bars are standard deviations.