Interspecies communication in Streptococcus gordonii-Veillonella atypica biofilms: signaling in flow conditions requires juxtaposition - PubMed (original) (raw)
Interspecies communication in Streptococcus gordonii-Veillonella atypica biofilms: signaling in flow conditions requires juxtaposition
Paul G Egland et al. Proc Natl Acad Sci U S A. 2004.
Abstract
During the development of human oral biofilm communities, the spatial arrangement of the bacteria is thought to be driven by metabolic interactions between them. Streptococcus gordonii and Veillonella atypica, two early colonizing members of the dental plaque biofilm, have been postulated to participate in metabolic communication; S. gordonii ferments carbohydrates to form lactic acid, which is a preferred fermentation substrate for V. atypica. We found that, during agar-plate coculture of these organisms, a signaling event occurs that results in increased expression of the S. gordonii alpha-amylase-encoding gene amyB. Confocal scanning laser microscopy of coculture flowcell-grown biofilms using human saliva as the sole nutrient showed that V. atypica caused S. gordonii to increase expression of a PamyB-'gfp transcriptional fusion in a spatially resolved fashion. In this open system, only those streptococci in mixed-species microcolonies containing V. atypica expressed GFP; nearby S. gordonii colonies that lacked V. atypica did not express GFP. In a closed system containing S. gordonii and V. atypica, flow cytometric analysis showed that S. gordonii containing the PamyB-'gfp reporter plasmid exhibited mean fluorescence levels 20-fold higher than did S. gordonii that had not been incubated with V. atypica. Thus, in a closed system where a diffusible signal can accumulate above a required threshold, interspecies signaling mediates a change in gene expression. We provide evidence that, in open systems like those that predominate in natural biofilms, diffusible signals between species are designed to function over short distances, on the order of 1 mum.
Figures
Fig. 1.
Detection of amylase activity in starch plates. (A) Starch hydrolysis occurs in a coculture of S. gordonii and V. atypica (Right) but not when the species are grown separately. (B) Starch hydrolysis by the amyB mutant alone and in coculture with V. atypica.
Fig. 2.
Amylase activity from culture supernatants. Activity is normalized for the fluorescence of labeled S. gordonii cells in each culture. Error bars represent standard deviations.
Fig. 3.
Confocal scanning laser microscopic analysis of dual-species biofilms. (A) Maximum projections (all confocal sections in a single field of view) of a single confocal stack showing fluorescence from Syto-59 (blue; all cells), Alexa Fluor 546-conjugated anti-V. atypica antibodies (red), and GFP (green; S. gordonii expressing amyB). The three fluorescence channels are shown separately and, in Lower Right, as overlay of GFP with V. atypica. (Inset) An enlargement of the boxed microcolony labeled with an asterisk. (B) Graphs of fluorescence intensity versus depth in a dual-species microcolony (Left) and a monospecies microcolony (Right) depicted in the upper right corner of each graph. The dual species microcolony is the same colony marked with an asterisk in A and is shown as an overlay of all three colors. The monospecies microcolony is labeled with a dagger in A. Microcolonies are shown as maximum projection images. Fluorescence of Syto-59 (blue triangles), Alexa Fluor 546-conjugated anti-V. atypica antibodies (open squares), and GFP (green triangles) are shown at each 0.5-μm-spaced optical slice of the confocal stack.
Fig. 4.
Confocal scanning laser microscopic analysis of S. gordonii (p_Pamy_-′gfp) monospecies (uninduced control) biofilms. Maximum projection images of fluorescence from channels for the detection of Syto-59 (Left) and GFP (Right) are shown.
Fig. 5.
Flow cytometric analysis of Pamy_-directed GFP expression. S. gordonii (p_Pamy_-′_gfp) and V. atypica as monospecies cultures (A and B, respectively) and coculture (C). No increase in fluorescence was seen when S. gordonii containing the gfp reporter plasmid with no promoter, pPE1010, was incubated with V. atypica (D). (E and F) Fluorescence from S. gordonii (p_Pamy_-′gfp) when separated by dialysis tubing from V. atypica (E) and sterile medium (F). Fluorescence (FL1-height) is graphed logarithmically on the x axis. Forward scatter (y axis) is indicative of particle size.
References
- Mikx, F. H. & Van der Hoeven, J. S. (1975) Arch. Oral Biol. 20**,** 407-410. -PubMed
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