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. 2018 Jun 18;84(13):e00219-18.
doi: 10.1128/AEM.00219-18. Print 2018 Jul 1.

Effects of PslG on the Surface Movement of Pseudomonas Aeruginosa

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Free PMC article

Effects of PslG on the Surface Movement of Pseudomonas Aeruginosa

Jingchao Zhang et al. Appl Environ Microbiol. .
Free PMC article

Abstract

PslG attracted a lot of attention recently due to its great potential abilities in inhibiting biofilms of Pseudomonas aeruginosa However, how PslG affects biofilm development still remains largely unexplored. Here, we focused on the surface motility of bacterial cells, which is critical for biofilm development. We studied the effects of PslG on bacterial surface movement in early biofilm development at a single-cell resolution by using a high-throughput bacterial tracking technique. The results showed that compared with no exogenous PslG addition, when PslG was added to the medium, bacterial surface movement was significantly (4 to 5 times) faster and proceeded in a more random way with no clear preferred direction. A further study revealed that the fraction of walking mode increased when PslG was added, which then resulted in an elevated average speed. The differences of motility due to PslG addition led to a clear distinction in patterns of bacterial surface movement and retarded microcolony formation greatly. Our results provide insight into developing new PslG-based biofilm control techniques.IMPORTANCE Biofilms of Pseudomonas aeruginosa are a major cause for hospital-acquired infections. They are notoriously difficult to eradicate and pose serious health hazards to human society. So, finding new ways to control biofilms is urgently needed. Recent work on PslG showed that PslG might be a good candidate for inhibiting/disassembling biofilms of Pseudomonas aeruginosa through Psl-based regulation. However, to fully explore PslG functions in biofilm control, a better understanding of PslG-Psl interactions is needed. Toward this end, we examined the effects of PslG on the surface movement of Pseudomonas aeruginosa in this work. The significance of our work is in greatly enhancing our understanding of the inhibiting mechanism of PslG on biofilms by providing a detailed picture of bacterial surface movement at a single-cell level, which will allow a full understanding of PslG abilities in biofilm control and thus present potential applications in biomedical fields.

Keywords: Pseudomonas aeruginosa; Psl; PslG; bacterial movement; biofilm.

Figures

FIG 1
FIG 1
Effects of PslG addition on the surface motility of bacteria. (A) Cell speeds with and without exogenous PslG addition for both WT and Psl++ bacteria. (B) Fractions of cells that crawl only, walk only, or both crawl and walk. (C) Residence times of cells with and without PslG addition for both WT and Psl++ bacteria. The data shown in this figure were collected for time periods of 12 h, 17 h, 13 h, and 20 h after inoculation for WT, WT + PslG, Psl++, and Psl++ + PslG bacteria, respectively. For all results with PslG addition, 50 nM PslG was used. The Psl++ label refers to △Ppsl/PBAD-psl cells with 1% arabinose. Statistical significance was measured using one-way analysis of variance (ANOVA). *, P < 0.05; **, P < 0.01; ***, P < 0.001. All data are the averaged results for three repeats, and more than 1,000 tracked cells were examined for each condition.
FIG 2
FIG 2
Effects of PslG addition on the direction of bacterial surface motion. (A) Representative probability distributions of moving angle, p(θ), for a WT cell (red) and a Psl++ cell (green) without exogenous PslG addition. θ is defined as the angle of a cell's displacement vector between two consecutive frames relative to a horizontal reference axis. (B) Representative p(θ) for a WT cell (red) and a Psl++ cell (green) with PslG addition. (C and D) Representative trajectories of △Ppsl/PBAD-psl cells under 1% arabinose without (C) and with (D) PslG addition.
FIG 3
FIG 3
Efficiency of surface coverage and bacterial visit distribution. (A) Surface coverage maps at a total of 7,500 (corresponding to incubation times of 1.7 h, 2.2 h, 3.3 h, and 3.4 h for WT, WT + PslG, Psl++, and Psl++ + PslG cells, respectively) and 50,000 (corresponding to incubation times of 3.8 h, 10.4 h, 10.7 h, and 15.8 h for WT, WT + PslG, Psl++, and Psl++ + PslG cells, respectively) bacterial visits for both WT and Psl++ cells with and without PslG addition. Red indicates the surface area that has been visited or contaminated, while black indicates a “fresh” surface area. Bacteria in the current frame are shown in blue. The surface coverages are 2.7%, 9.2%, 1.8%, and 9.2% at 7,500 visits and 37.5%, 85.7%, 13.5%, and 55.4% at 50,000 visits for WT, WT with PslG, Psl++, and Psl++ with PslG cells, respectively. (B) Bright-field images and visit frequency maps at a total of ∼200,000 visits (corresponding to incubation times of 5.6 h, 16.3 h, 13.9 h, and 19.4 h for WT, WT + PslG, Psl++, and Psl++ + PslG cells, respectively), when microcolonies began to form. (C) The graph on the left displays the visit frequency distributions for WT cells without (red) and with (magenta) PslG addition, and the one on the right shows the visit frequency distributions for Psl++ cells without (olive) and with (green) PslG addition. Bars, 10 μm. Lines in panel C are power law fittings.
FIG 4
FIG 4
Measurement of the time required for microcolony formation. (A) An example of a microcolony formed by WT cells cultured in a flow cell for about 9 h in the case of no PslG addition (marked in red) and for about 18 h in the case of PslG addition (marked in magenta). (B) Time required to form the first microcolony (defined as clusters of 30 cells in this work) that appeared in the chosen field of view for both WT and Psl++ cells with and without PslG addition. Statistical significance was measured using one-way ANOVA. *, P < 0.05; **, P < 0.01; ***, P < 0.001. Bars, 10 μm.

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