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Identification of PblB mediating galactose-specific adhesion in a successful Streptococcus pneumoniae clone.

Hsieh YC, Lin TL, Lin CM, Wang JT - Sci Rep (2015)

Bottom Line: Preincubation of NTUH-P15 with D-galactose resulted in decreases of adherence to A549 cell in a dose-dependent manner.Challenge of mice with NTUH-P15, isogenic pblB mutant and pblB complementation strains determined that PblB was required for bacterial persistence in the nasopharynx and lung.PblB, as an adhesin mediating the galactose-specific adhesion activity of pneumococci, promote pneumococcal clonal success.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Chang Gung Children's Hospital, Chang Gung Memorial Hospital, Chang Gung University, College of Medicine, Taoyuan, Taiwan.

ABSTRACT
The pneumococcal genome is variable and there are minimal data on the influence of the accessory genome on phenotype. Pneumococcal serotype 14 sequence type (ST) 46 had been the most prevalent clone causing pneumonia in children in Taiwan. A microarray was constructed using the genomic DNA of a clinical strain (NTUH-P15) of serotype 14 ST46. Using DNA hybridization, genomic variations in NTUH-P15 were compared to those of 3 control strains. Microarray analysis identified 7 genomic regions that had significant increases in hybridization signals in the NTUH-P15 strain compared to control strains. One of these regions encoded PblB, a phage-encoded virulence factor implicated (in Streptococcus mitis) in infective endocarditis. The isogenic pblB mutant decreased adherence to A549 human lung epithelial cell compared to wild-type NTUH-P15 strain (P = 0.01). Complementation with pblB restored the adherence. PblB is predicted to contain a galactose-binding domain-like region. Preincubation of NTUH-P15 with D-galactose resulted in decreases of adherence to A549 cell in a dose-dependent manner. Challenge of mice with NTUH-P15, isogenic pblB mutant and pblB complementation strains determined that PblB was required for bacterial persistence in the nasopharynx and lung. PblB, as an adhesin mediating the galactose-specific adhesion activity of pneumococci, promote pneumococcal clonal success.

No MeSH data available.


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Adherence of wild-type NTUH-P15, pblB mutant, and pblB complementation strains to platelets/A549/HEp-2 cells in vitro.A. The pblB mutant decreased adherence to platelets compared to the wild-type strain. Complementation with pblB restored the adherence. B. The pblB mutant decreased adherence to A549 and HEp-2 cells compared to the wild-type strain. Complementation with pblB restored the adherence. C. Preincubation of wild-type NTUH-P15 with D-galactose decreased adherence to A549 cell. In contrast, preincubation of wild-type NTUH-P15 with D-glucoase and N-acetylneuraminic acid did not decreased adherence to A549 cell. D. Adherence of NTUH-P15 wild-type to A549 cells was inhibited in a dose-dependent manner by pretreatment of bacteria with D-galactose, but not in the isogenic pblB mutant. E. Adherence of NTUH-P15 wild-type to A549 cells pre-treated with ß-galactosidase decreased compared to that without ß-galactosidase, but not in the isogenic pblB mutant. Experiments were performed in triplicate. The standard error of the mean (SEM) are indicated as bar graph.
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f3: Adherence of wild-type NTUH-P15, pblB mutant, and pblB complementation strains to platelets/A549/HEp-2 cells in vitro.A. The pblB mutant decreased adherence to platelets compared to the wild-type strain. Complementation with pblB restored the adherence. B. The pblB mutant decreased adherence to A549 and HEp-2 cells compared to the wild-type strain. Complementation with pblB restored the adherence. C. Preincubation of wild-type NTUH-P15 with D-galactose decreased adherence to A549 cell. In contrast, preincubation of wild-type NTUH-P15 with D-glucoase and N-acetylneuraminic acid did not decreased adherence to A549 cell. D. Adherence of NTUH-P15 wild-type to A549 cells was inhibited in a dose-dependent manner by pretreatment of bacteria with D-galactose, but not in the isogenic pblB mutant. E. Adherence of NTUH-P15 wild-type to A549 cells pre-treated with ß-galactosidase decreased compared to that without ß-galactosidase, but not in the isogenic pblB mutant. Experiments were performed in triplicate. The standard error of the mean (SEM) are indicated as bar graph.

Mentions: Growth curves of the wild-type, mutant and complementation strains were comparable (data not shown). Since PblB is a platelet adhesin in S. mitis10, we evaluated whether PblB of NTUH-P15 mediates adhesion to platelets. We constructed an isogenic pblB insertion mutant in the NTUH-P15 strains. The pblB mutant decreased adherence to platelets (P = 0.01) compared to the wild-type strain (Fig. 3A). When the NTUH-P15 pblB mutant strain was complemented with the pblB gene, the adherence of the wild-type strain to platelets (P = 0.2) were not significantly different from the adherence of the complementation strain (Fig. 3A). Subsequently, we measured the impact of pblB on pneumococcal adherence to respiratory epithelial cell. The pblB mutant decreased adherence to lung epithelial cell (A549 cells) (P = 0.01) and laryngeal cell (HEp-2) (P = 0.04) compared to the wild-type strain (Fig. 3B). When the NTUH-P15 pblB mutant strain was complemented with the pblB gene, the adherence of the wild-type strain to A549 cells and HEp-2 were restored (Fig. 3B). Introduction of pblB into the pblB-negative NTUH-P3 strain did not increase adherence to A549 cells as compared to that of pblB-negative NTUH-P3 strain (P = 0.8) (data not shown)


Identification of PblB mediating galactose-specific adhesion in a successful Streptococcus pneumoniae clone.

Hsieh YC, Lin TL, Lin CM, Wang JT - Sci Rep (2015)

Adherence of wild-type NTUH-P15, pblB mutant, and pblB complementation strains to platelets/A549/HEp-2 cells in vitro.A. The pblB mutant decreased adherence to platelets compared to the wild-type strain. Complementation with pblB restored the adherence. B. The pblB mutant decreased adherence to A549 and HEp-2 cells compared to the wild-type strain. Complementation with pblB restored the adherence. C. Preincubation of wild-type NTUH-P15 with D-galactose decreased adherence to A549 cell. In contrast, preincubation of wild-type NTUH-P15 with D-glucoase and N-acetylneuraminic acid did not decreased adherence to A549 cell. D. Adherence of NTUH-P15 wild-type to A549 cells was inhibited in a dose-dependent manner by pretreatment of bacteria with D-galactose, but not in the isogenic pblB mutant. E. Adherence of NTUH-P15 wild-type to A549 cells pre-treated with ß-galactosidase decreased compared to that without ß-galactosidase, but not in the isogenic pblB mutant. Experiments were performed in triplicate. The standard error of the mean (SEM) are indicated as bar graph.
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4508584&req=5

f3: Adherence of wild-type NTUH-P15, pblB mutant, and pblB complementation strains to platelets/A549/HEp-2 cells in vitro.A. The pblB mutant decreased adherence to platelets compared to the wild-type strain. Complementation with pblB restored the adherence. B. The pblB mutant decreased adherence to A549 and HEp-2 cells compared to the wild-type strain. Complementation with pblB restored the adherence. C. Preincubation of wild-type NTUH-P15 with D-galactose decreased adherence to A549 cell. In contrast, preincubation of wild-type NTUH-P15 with D-glucoase and N-acetylneuraminic acid did not decreased adherence to A549 cell. D. Adherence of NTUH-P15 wild-type to A549 cells was inhibited in a dose-dependent manner by pretreatment of bacteria with D-galactose, but not in the isogenic pblB mutant. E. Adherence of NTUH-P15 wild-type to A549 cells pre-treated with ß-galactosidase decreased compared to that without ß-galactosidase, but not in the isogenic pblB mutant. Experiments were performed in triplicate. The standard error of the mean (SEM) are indicated as bar graph.
Mentions: Growth curves of the wild-type, mutant and complementation strains were comparable (data not shown). Since PblB is a platelet adhesin in S. mitis10, we evaluated whether PblB of NTUH-P15 mediates adhesion to platelets. We constructed an isogenic pblB insertion mutant in the NTUH-P15 strains. The pblB mutant decreased adherence to platelets (P = 0.01) compared to the wild-type strain (Fig. 3A). When the NTUH-P15 pblB mutant strain was complemented with the pblB gene, the adherence of the wild-type strain to platelets (P = 0.2) were not significantly different from the adherence of the complementation strain (Fig. 3A). Subsequently, we measured the impact of pblB on pneumococcal adherence to respiratory epithelial cell. The pblB mutant decreased adherence to lung epithelial cell (A549 cells) (P = 0.01) and laryngeal cell (HEp-2) (P = 0.04) compared to the wild-type strain (Fig. 3B). When the NTUH-P15 pblB mutant strain was complemented with the pblB gene, the adherence of the wild-type strain to A549 cells and HEp-2 were restored (Fig. 3B). Introduction of pblB into the pblB-negative NTUH-P3 strain did not increase adherence to A549 cells as compared to that of pblB-negative NTUH-P3 strain (P = 0.8) (data not shown)

Bottom Line: Preincubation of NTUH-P15 with D-galactose resulted in decreases of adherence to A549 cell in a dose-dependent manner.Challenge of mice with NTUH-P15, isogenic pblB mutant and pblB complementation strains determined that PblB was required for bacterial persistence in the nasopharynx and lung.PblB, as an adhesin mediating the galactose-specific adhesion activity of pneumococci, promote pneumococcal clonal success.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Chang Gung Children's Hospital, Chang Gung Memorial Hospital, Chang Gung University, College of Medicine, Taoyuan, Taiwan.

ABSTRACT
The pneumococcal genome is variable and there are minimal data on the influence of the accessory genome on phenotype. Pneumococcal serotype 14 sequence type (ST) 46 had been the most prevalent clone causing pneumonia in children in Taiwan. A microarray was constructed using the genomic DNA of a clinical strain (NTUH-P15) of serotype 14 ST46. Using DNA hybridization, genomic variations in NTUH-P15 were compared to those of 3 control strains. Microarray analysis identified 7 genomic regions that had significant increases in hybridization signals in the NTUH-P15 strain compared to control strains. One of these regions encoded PblB, a phage-encoded virulence factor implicated (in Streptococcus mitis) in infective endocarditis. The isogenic pblB mutant decreased adherence to A549 human lung epithelial cell compared to wild-type NTUH-P15 strain (P = 0.01). Complementation with pblB restored the adherence. PblB is predicted to contain a galactose-binding domain-like region. Preincubation of NTUH-P15 with D-galactose resulted in decreases of adherence to A549 cell in a dose-dependent manner. Challenge of mice with NTUH-P15, isogenic pblB mutant and pblB complementation strains determined that PblB was required for bacterial persistence in the nasopharynx and lung. PblB, as an adhesin mediating the galactose-specific adhesion activity of pneumococci, promote pneumococcal clonal success.

No MeSH data available.


Related in: MedlinePlus