At change in the quantity of any of these components influences the 3D-structure of EPS. For example, biofilms of a fimbriae-deficient strain (flp-1-disrupted mutant) of the periodontal pathogen Aggregatibacter actinomycetemcomitans forms microcolonies in looser formation, and fibrils of fimbriae are not observed [27]. Furthermore, its adhesion to the surface was significantly blocked by sodium metaperiodate or DNase I treatment but not by proteases. This mutant secretes carbohydrates and DNA instead of fimbriae to coalescent on a surface [27]. Friedman and LED-209 Kolter screened for transposon insertion mutants of Pseudomonas aeruginosa PA14 that were unable to form pellicles that represent one type of biofilm formed at the air-liquid interface in static cultures [28]. They identified 7 flanking genesFigure 4. Strength of biofilms formed by P. gingivalis strains. Standardized cultures of P. gingivalis were inoculated into dGAM in saliva-coated 12-well polystyrene plate and incubated statically at 37uC for 60 h, and the resulting biofilms were sonicated for 1 s. Immediately after sonication, supernatants containing floating cells were removed by aspiration and the biofilm remnants were gently washed with PBS. P. gingivalis genomic DNA was isolated from the biofilms, and the numbers of P. gingivalis cells were determined using real-time PCR. Relative amounts of bacterial cell numbers were calculated based on the number of wild type cells without sonication and assigned a value of 1.0. The percentages shown indicate the amount of remaining biofilm after sonic disruption. Duplicate experiments were independently repeated three times with each strain. Standard error bars are shown. Statistical analysis was performed using Welch’s t test. *P,0.001 in comparison with the wild type strain. doi:10.1371/journal.pone.0056017.gThe Role of sinR in P. gingivalis Biofilms(pel) that contribute to the formation of the pellicle, and revealed that the products of these genes are involved in the construction of the EPS [28]. In B. subtilis, the structures of the biofilms formed by the eps (required for production of exopolysaccharide) mutant and tasA (forms amyloid fibers) mutant were flat. In contrast, the biofilms produced by the sinR (inhibitor of eps and tasA) mutant were extremely wrinkly [19,21]. The CLSM (Figure 2B) and SEM (Figure 3) images acquired in the present study show that the mutation of sinR induces morphological changes of the EPS from a laminar to a mesh-like structure. Thus, the SinR produced by P. gingivalis ATCC 33277 might be indirectly involved in the 3Dconformation of the EPS in biofilms by controlling the expression of genes associated with the EPS components. Xylella fastidiosa, a bacterium responsible for Pierce’s disease in grapevines, possesses both type I and type IV pili at the same cell pole. De La Fuente et al. [29] evaluated the attachment of the bacteria to a glass substratum using a microfluidic flow chamber in conjunction with pilus-defective mutants. The adhesion force required to disperse X. fastidiosa mutant ML240 site possessing only type I pili was significantly higher, whereas that of mutant cells possessing only type IV pili was significantly lower than those of wild type cells [29]. In contrast, Kuboniwa et al. [19] revealed that the exopolysaccharide per cell ratio of biofilms formed by a fimA mutant was significantly higher than that of wild type and 1407003 that the mutant formed a tough and cohesive biofilm. Furthermore, the exopol.At change in the quantity of any of these components influences the 3D-structure of EPS. For example, biofilms of a fimbriae-deficient strain (flp-1-disrupted mutant) of the periodontal pathogen Aggregatibacter actinomycetemcomitans forms microcolonies in looser formation, and fibrils of fimbriae are not observed [27]. Furthermore, its adhesion to the surface was significantly blocked by sodium metaperiodate or DNase I treatment but not by proteases. This mutant secretes carbohydrates and DNA instead of fimbriae to coalescent on a surface [27]. Friedman and Kolter screened for transposon insertion mutants of Pseudomonas aeruginosa PA14 that were unable to form pellicles that represent one type of biofilm formed at the air-liquid interface in static cultures [28]. They identified 7 flanking genesFigure 4. Strength of biofilms formed by P. gingivalis strains. Standardized cultures of P. gingivalis were inoculated into dGAM in saliva-coated 12-well polystyrene plate and incubated statically at 37uC for 60 h, and the resulting biofilms were sonicated for 1 s. Immediately after sonication, supernatants containing floating cells were removed by aspiration and the biofilm remnants were gently washed with PBS. P. gingivalis genomic DNA was isolated from the biofilms, and the numbers of P. gingivalis cells were determined using real-time PCR. Relative amounts of bacterial cell numbers were calculated based on the number of wild type cells without sonication and assigned a value of 1.0. The percentages shown indicate the amount of remaining biofilm after sonic disruption. Duplicate experiments were independently repeated three times with each strain. Standard error bars are shown. Statistical analysis was performed using Welch’s t test. *P,0.001 in comparison with the wild type strain. doi:10.1371/journal.pone.0056017.gThe Role of sinR in P. gingivalis Biofilms(pel) that contribute to the formation of the pellicle, and revealed that the products of these genes are involved in the construction of the EPS [28]. In B. subtilis, the structures of the biofilms formed by the eps (required for production of exopolysaccharide) mutant and tasA (forms amyloid fibers) mutant were flat. In contrast, the biofilms produced by the sinR (inhibitor of eps and tasA) mutant were extremely wrinkly [19,21]. The CLSM (Figure 2B) and SEM (Figure 3) images acquired in the present study show that the mutation of sinR induces morphological changes of the EPS from a laminar to a mesh-like structure. Thus, the SinR produced by P. gingivalis ATCC 33277 might be indirectly involved in the 3Dconformation of the EPS in biofilms by controlling the expression of genes associated with the EPS components. Xylella fastidiosa, a bacterium responsible for Pierce’s disease in grapevines, possesses both type I and type IV pili at the same cell pole. De La Fuente et al. [29] evaluated the attachment of the bacteria to a glass substratum using a microfluidic flow chamber in conjunction with pilus-defective mutants. The adhesion force required to disperse X. fastidiosa mutant possessing only type I pili was significantly higher, whereas that of mutant cells possessing only type IV pili was significantly lower than those of wild type cells [29]. In contrast, Kuboniwa et al. [19] revealed that the exopolysaccharide per cell ratio of biofilms formed by a fimA mutant was significantly higher than that of wild type and 1407003 that the mutant formed a tough and cohesive biofilm. Furthermore, the exopol.
