Skip to main content
. Author manuscript; available in PMC: 2020 Sep 2.
Published in final edited form as: Nat Microbiol. 2018 Sep 24;3(11):1243–1254. doi: 10.1038/s41564-018-0248-x

Figure 2. TtsA mediates the transport of typhoid toxin to the trans side of the peptidoglycan layer.

Figure 2.

(a-e) Wild-type and ΔttsA S. Typhi carrying chromosomally-encoded FLAG-tagged CdtB (as a surrogate for typhoid toxin), DsbD, or SlyA (as indicated) were grown in TTIM for 24 hs, fixed, mock treated or treated with lysozyme for 20 min (to permeabilize the PG layer), and stained as in Fig. 1 (a). To probe the TtsA-dependent translocation of typhoid toxin from the cis to the trans side of the PG layer, bacteria (wild-type and ΔttsA mutant) were first treated with proteinase K for 30 min, followed by lysozyme or mock treatment for 20 min (c and e), or first treated with lysozyme for 20 min, followed by proteinase K treatment for 30 min (d). In all cases, the number of bacterial cells positive for CdtB, DsbD, or SlyA signals (green), was quantified relative to the total number of bacteria analyzed (LPS, red signal). The average ratios of green positive cells to red total cells ± standard deviation from 3 independent experiments are shown. In all cases, a total of 30 images were collected from which 100 randomly selected bacteria (total number) per image were analyzed, resulting in 3,000 bacterial cells analyzed per bacterial strain or condition (**** p < 0.0001, (c) ns p = 0.131, (d) ns p = 0.2078, p = 0.28, (e) ns p = 0.2646, two-sided Student’s t Test) (Supplementary data set 2) (b-e).