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Mucin promotes rapid surface motility in Pseudomonas aeruginosa.

Yeung AT, Parayno A, Hancock RE - MBio (2012)

Bottom Line: In this study, we added mucin to swimming media and found that it promoted the ability of P. aeruginosa to exhibit rapid surface motility.Interestingly, bacterial cells at the thick edge appeared piled up and lacked flagella, while cells at the motility center had flagella.Our data from various genetic and phenotypic studies suggest that mucin may be promoting a modified form of swarming or a novel form of surface motility in P. aeruginosa.

View Article: PubMed Central - PubMed

Affiliation: Centre for Microbial Diseases and Immunity Research, University of British Columbia, Vancouver, British Columbia, Canada.

ABSTRACT

Unlabelled: An important environmental factor that determines the mode of motility adopted by Pseudomonas aeruginosa is the viscosity of the medium, often provided by adjusting agar concentrations in vitro. However, the viscous gel-like property of the mucus layer that overlays epithelial surfaces is largely due to the glycoprotein mucin. P. aeruginosa is known to swim within 0.3% (wt/vol) agar and swarm on the surface at 0.5% (wt/vol) agar with amino acids as a weak nitrogen source. When physiological concentrations or as little as 0.05% (wt/vol) mucin was added to the swimming agar, in addition to swimming, P. aeruginosa was observed to undergo highly accelerated motility on the surface of the agar. The surface motility colonies in the presence of mucin appeared to be circular, with a bright green center surrounded by a thicker white edge. While intact flagella were required for the surface motility in the presence of mucin, type IV pili and rhamnolipid production were not. Replacement of mucin with other wetting agents indicated that the lubricant properties of mucin might contribute to the surface motility. Based on studies with mutants, the quorum-sensing systems (las and rhl) and the orphan autoinducer receptor QscR played important roles in this form of surface motility. Transcriptional analysis of cells taken from the motility zone revealed the upregulation of genes involved in virulence and resistance. Based on these results, we suggest that mucin may be promoting a new or highly modified form of surface motility, which we propose should be termed "surfing."

Importance: An important factor that dictates the mode of motility adopted by P. aeruginosa is the viscosity of the medium, often provided by adjusting agar concentrations in vitro. However, the gel-like properties of the mucous layers that overlay epithelial surfaces, such as those of the lung, a major site of Pseudomonas infection, are contributed mostly by the production of the glycoprotein mucin. In this study, we added mucin to swimming media and found that it promoted the ability of P. aeruginosa to exhibit rapid surface motility. These motility colonies appeared in a circular form, with a bright green center surrounded by a thicker white edge. Interestingly, bacterial cells at the thick edge appeared piled up and lacked flagella, while cells at the motility center had flagella. Our data from various genetic and phenotypic studies suggest that mucin may be promoting a modified form of swarming or a novel form of surface motility in P. aeruginosa.

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Related in: MedlinePlus

Swimming (A), swarming (B and C), and mucin-promoted (D) motilities of P. aeruginosa. Motilities were examined on plates containing 0.3% (wt/vol) agar (swim), 0.5% (wt/vol) agar (swarm), or 0.3% (wt/vol) agar with 0.4% (wt/vol) mucin.
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fig1: Swimming (A), swarming (B and C), and mucin-promoted (D) motilities of P. aeruginosa. Motilities were examined on plates containing 0.3% (wt/vol) agar (swim), 0.5% (wt/vol) agar (swarm), or 0.3% (wt/vol) agar with 0.4% (wt/vol) mucin.

Mentions: In the presence of a low-percentage (0.2% to 0.35% [wt/vol]) agar, P. aeruginosa swims in the submerged water-filled spaces of the agar by the use of its single polar flagellum, resulting in the formation of a halo within the agar layer after overnight incubation at 37°C (Fig. 1A). Interestingly, when mucin was added to “swim” agar, in addition to swimming, P. aeruginosa was observed to move relatively rapidly across the surface of the agar. The surface motility zones could be observed when mucin was added at concentrations as low as 0.05% (wt/vol), and the diameter of the surface motility zone increased as the concentration of added mucin increased (up to 1% [wt/vol] mucin tested; Fig. 1D shows an example at 0.4% [wt/vol] mucin). Moreover, the addition of mucin to 0.5% (wt/vol) agar (0.5% agar normally promotes swarming motility of P. aeruginosa) changed the surface motility pattern from dendritic to circular, although the diameter of the motility colony remained similar (data not shown). The same surface motility patterns were observed when mucin was spread onto an agar slab. In the presence of mucin, the surface motility colonies of both P. aeruginosa strains PA14 and PAO1 appeared circular, with a green center surrounded by a thick white edge. This motility pattern somewhat resembles the solar flare-like colonial swarming pattern of strain PAO1 (Fig. 1C) but differs from the dendritic swarming colony of strain PA14 (Fig. 1B). To better mimic the nutritional composition of the CF sputum, we replaced the typical BM2 (62 mM potassium phosphate buffer [pH 7], 0.1% [wt/vol] Casamino Acids [CAA], 2 mM MgSO4, 10 µM FeSO4, 0.4% [wt/vol] glucose)-minimal medium in the swim plates with a modified version of the synthetic CF sputum medium (MSCFM) (SCFM [18] without NH4Cl) in which NH4Cl was excluded. When mucin was added to MSCFM, virtually identical surface motility colonies were observed (data not shown). Moreover, the same motility phenotype was observed when physiological amounts of DNA (1.4 mg/ml) were added to the mucin-MSCFM plates (data not shown). While we used the same DNA concentration (1.4 mg/ml) as Fung et al. used in their synthetic CF sputum growth medium (19), Sriramulu et al. had used a higher concentration of DNA (4 mg/ml) in their artificial CF sputum medium (20). However, in our hands the addition of DNA at ≥4 mg/ml inhibited the growth of P. aeruginosa in mucin-MSCFM (Fig. S1). Furthermore, when the surface motilities of P. aeruginosa were compared on mucin-containing MSCFM plates (without NH4Cl) and mucin-containing SCFM plates (with NH4Cl provided at 2.3 mM [18]), no differences were observed in the motility colony morphology, rate of motility zone expansion, or growth of the bacteria.


Mucin promotes rapid surface motility in Pseudomonas aeruginosa.

Yeung AT, Parayno A, Hancock RE - MBio (2012)

Swimming (A), swarming (B and C), and mucin-promoted (D) motilities of P. aeruginosa. Motilities were examined on plates containing 0.3% (wt/vol) agar (swim), 0.5% (wt/vol) agar (swarm), or 0.3% (wt/vol) agar with 0.4% (wt/vol) mucin.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3569861&req=5

fig1: Swimming (A), swarming (B and C), and mucin-promoted (D) motilities of P. aeruginosa. Motilities were examined on plates containing 0.3% (wt/vol) agar (swim), 0.5% (wt/vol) agar (swarm), or 0.3% (wt/vol) agar with 0.4% (wt/vol) mucin.
Mentions: In the presence of a low-percentage (0.2% to 0.35% [wt/vol]) agar, P. aeruginosa swims in the submerged water-filled spaces of the agar by the use of its single polar flagellum, resulting in the formation of a halo within the agar layer after overnight incubation at 37°C (Fig. 1A). Interestingly, when mucin was added to “swim” agar, in addition to swimming, P. aeruginosa was observed to move relatively rapidly across the surface of the agar. The surface motility zones could be observed when mucin was added at concentrations as low as 0.05% (wt/vol), and the diameter of the surface motility zone increased as the concentration of added mucin increased (up to 1% [wt/vol] mucin tested; Fig. 1D shows an example at 0.4% [wt/vol] mucin). Moreover, the addition of mucin to 0.5% (wt/vol) agar (0.5% agar normally promotes swarming motility of P. aeruginosa) changed the surface motility pattern from dendritic to circular, although the diameter of the motility colony remained similar (data not shown). The same surface motility patterns were observed when mucin was spread onto an agar slab. In the presence of mucin, the surface motility colonies of both P. aeruginosa strains PA14 and PAO1 appeared circular, with a green center surrounded by a thick white edge. This motility pattern somewhat resembles the solar flare-like colonial swarming pattern of strain PAO1 (Fig. 1C) but differs from the dendritic swarming colony of strain PA14 (Fig. 1B). To better mimic the nutritional composition of the CF sputum, we replaced the typical BM2 (62 mM potassium phosphate buffer [pH 7], 0.1% [wt/vol] Casamino Acids [CAA], 2 mM MgSO4, 10 µM FeSO4, 0.4% [wt/vol] glucose)-minimal medium in the swim plates with a modified version of the synthetic CF sputum medium (MSCFM) (SCFM [18] without NH4Cl) in which NH4Cl was excluded. When mucin was added to MSCFM, virtually identical surface motility colonies were observed (data not shown). Moreover, the same motility phenotype was observed when physiological amounts of DNA (1.4 mg/ml) were added to the mucin-MSCFM plates (data not shown). While we used the same DNA concentration (1.4 mg/ml) as Fung et al. used in their synthetic CF sputum growth medium (19), Sriramulu et al. had used a higher concentration of DNA (4 mg/ml) in their artificial CF sputum medium (20). However, in our hands the addition of DNA at ≥4 mg/ml inhibited the growth of P. aeruginosa in mucin-MSCFM (Fig. S1). Furthermore, when the surface motilities of P. aeruginosa were compared on mucin-containing MSCFM plates (without NH4Cl) and mucin-containing SCFM plates (with NH4Cl provided at 2.3 mM [18]), no differences were observed in the motility colony morphology, rate of motility zone expansion, or growth of the bacteria.

Bottom Line: In this study, we added mucin to swimming media and found that it promoted the ability of P. aeruginosa to exhibit rapid surface motility.Interestingly, bacterial cells at the thick edge appeared piled up and lacked flagella, while cells at the motility center had flagella.Our data from various genetic and phenotypic studies suggest that mucin may be promoting a modified form of swarming or a novel form of surface motility in P. aeruginosa.

View Article: PubMed Central - PubMed

Affiliation: Centre for Microbial Diseases and Immunity Research, University of British Columbia, Vancouver, British Columbia, Canada.

ABSTRACT

Unlabelled: An important environmental factor that determines the mode of motility adopted by Pseudomonas aeruginosa is the viscosity of the medium, often provided by adjusting agar concentrations in vitro. However, the viscous gel-like property of the mucus layer that overlays epithelial surfaces is largely due to the glycoprotein mucin. P. aeruginosa is known to swim within 0.3% (wt/vol) agar and swarm on the surface at 0.5% (wt/vol) agar with amino acids as a weak nitrogen source. When physiological concentrations or as little as 0.05% (wt/vol) mucin was added to the swimming agar, in addition to swimming, P. aeruginosa was observed to undergo highly accelerated motility on the surface of the agar. The surface motility colonies in the presence of mucin appeared to be circular, with a bright green center surrounded by a thicker white edge. While intact flagella were required for the surface motility in the presence of mucin, type IV pili and rhamnolipid production were not. Replacement of mucin with other wetting agents indicated that the lubricant properties of mucin might contribute to the surface motility. Based on studies with mutants, the quorum-sensing systems (las and rhl) and the orphan autoinducer receptor QscR played important roles in this form of surface motility. Transcriptional analysis of cells taken from the motility zone revealed the upregulation of genes involved in virulence and resistance. Based on these results, we suggest that mucin may be promoting a new or highly modified form of surface motility, which we propose should be termed "surfing."

Importance: An important factor that dictates the mode of motility adopted by P. aeruginosa is the viscosity of the medium, often provided by adjusting agar concentrations in vitro. However, the gel-like properties of the mucous layers that overlay epithelial surfaces, such as those of the lung, a major site of Pseudomonas infection, are contributed mostly by the production of the glycoprotein mucin. In this study, we added mucin to swimming media and found that it promoted the ability of P. aeruginosa to exhibit rapid surface motility. These motility colonies appeared in a circular form, with a bright green center surrounded by a thicker white edge. Interestingly, bacterial cells at the thick edge appeared piled up and lacked flagella, while cells at the motility center had flagella. Our data from various genetic and phenotypic studies suggest that mucin may be promoting a modified form of swarming or a novel form of surface motility in P. aeruginosa.

Show MeSH
Related in: MedlinePlus