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25474103[PMID] - PMC - NCBI

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1.
Figure 2

Figure 2. Phylogeny of SRR glycoprotein BRs from a representative number of S. sanguinis and S. gordonii strains, and glycan binding to immobilized adhesin-BRs.. From: Oral Streptococci Utilize a Siglec-Like Domain of Serine-Rich Repeat Adhesins to Preferentially Target Platelet Sialoglycans in Human Blood.

(A) ClustalW2 was used for both the sequence alignment and the neighbor-joining tree reconstruction. Corresponding SRR adhesin-BR sequence accession numbers are provided in . (B) PAA-supported sTa binds to both the Hsa-BR and BRs of the four GspB-orthologues. (C) PAA-SLn only binds to Hsa-BR. (n = 6, SEM).

Lingquan Deng, et al. PLoS Pathog. 2014 Dec;10(12):e1004540.
2.
Figure 1

Figure 1. Sialoglycan microarray analysis of binding specificities of adhesin-BRs and corresponding whole bacteria.. From: Oral Streptococci Utilize a Siglec-Like Domain of Serine-Rich Repeat Adhesins to Preferentially Target Platelet Sialoglycans in Human Blood.

(A–C) Binding of the adhesin-BR proteins at 50 nM is presented. (n = 4, SD). (D) Adhesin-BR proteins and Cy3-labeled whole bacteria were assayed using the same sialoglycan microarrays. Heat map was generated using the method as previously described . R1 and R2 represent two different spacers.

Lingquan Deng, et al. PLoS Pathog. 2014 Dec;10(12):e1004540.
3.
Figure 6

Figure 6. Divalent cation effect on bacterial binding and preferential recognition of platelets over RBCs by Hsa-BR in whole blood.. From: Oral Streptococci Utilize a Siglec-Like Domain of Serine-Rich Repeat Adhesins to Preferentially Target Platelet Sialoglycans in Human Blood.

(A) DL1 and DL1Δhsa binding to either platelets or RBCs were comparable in EDTA- or heparin-anticoagulated whole blood. Bar values = 100×[number of bacterium-bound platelets (or RBCs)/total number of platelets (or RBCs)]. (n = 3, SEM). (B) Quantification and comparison of Hsa-BR binding to platelets and RBCs in whole blood. Bar values = 100×[number of HsaBR-bound platelets (or RBCs)/total number of platelets (or RBCs)]. (n = 6, SEM). (C) Q1 shows mainly RBCs, and Q4 platelets. Only 2° antibody, anti-GST-APC, alone was added in whole blood. (D–F) 6 to 96 pmol of Hsa-BR was added to whole blood, followed by anti-GST-APC. ****P<0.0001.

Lingquan Deng, et al. PLoS Pathog. 2014 Dec;10(12):e1004540.
4.
Figure 3

Figure 3. Bacterial binding to saliva and glycoconjugates, and bacterium-mediated hemagglutination.. From: Oral Streptococci Utilize a Siglec-Like Domain of Serine-Rich Repeat Adhesins to Preferentially Target Platelet Sialoglycans in Human Blood.

(A and B) Binding of DL1 vs. DL1Δhsa against immobilized whole human saliva, salivary ductal secretions, isolated and purified glycoproteins and related glycoconjugates, was tested by a dot blot assay. The binding was studied in the presence of EDTA (A) or divalent cations (B). (C–F) Bacterium-mediated hemagglutination using human RBCs and two-fold serial diluted bacterial suspensions. (C and D) DL1- and DL1Δhsa-mediated hemagglutination using non-sialidase treated (Ø) RBCs. (E and F) DL1- and DL1Δhsa-mediated hemagglutination using sialidase treated (S) RBCs.

Lingquan Deng, et al. PLoS Pathog. 2014 Dec;10(12):e1004540.
5.
Figure 7

Figure 7. Blocking of DL1-platelet binding by Hsa-BR in whole human blood.. From: Oral Streptococci Utilize a Siglec-Like Domain of Serine-Rich Repeat Adhesins to Preferentially Target Platelet Sialoglycans in Human Blood.

(A) Representative FSC/SSC plot showing whole blood with added DL1-Cy3. Platelets and bacteria are gated as R1. (B–E) are gated from R1: (B) Whole blood with DL1-Cy3 and CD41a-APC. Non-DL1-bound platelets appear in Q1 and DL1-bound in Q2, while free DL1-Cy3 in Q3. (C–E) Whole blood with different concentrations of Hsa-BR added first, followed by DL1-Cy3 and CD41a-APC. Increasing amount of Hsa-BR results in decreasing count of DL1-bound platelets (Q2 percentages) and increasing numbers of free bacteria (Q3 percentages). (F) Quantification of Hsa-BR blocking of DL1 binding to platelets in whole blood. The percentage values of DL1-bound platelets were normalized, with the highest single value being 100%. (n = 6, SEM). *P<0.05, ***P<0.001.

Lingquan Deng, et al. PLoS Pathog. 2014 Dec;10(12):e1004540.
6.
Figure 5

Figure 5. Representative flow cytometry profiles showing preferential platelet-bacterial adherence in intact whole human blood.. From: Oral Streptococci Utilize a Siglec-Like Domain of Serine-Rich Repeat Adhesins to Preferentially Target Platelet Sialoglycans in Human Blood.

(A–C) Whole blood (WB) in PBS. (D–F) Two antibodies, CD235a-FITC and CD41a-APC, were added to WB. (G–I) DL1-Cy3 incubated with CD235a-FITC and CD41a-APC: (G) DL1-Cy3 also appears in Q4 in the FSC/SSC plot, the same as platelets. (J–L) DL1-Cy3, CD235a-FITC and CD41a-APC were added to WB: (J) Platelets/bacteria are gated as R1 and RBCs/WBCs as R2. (K) Events from gate R1. (L) Events from gate R2. Bacteria preferentially recognize platelets compared to RBCs. (M–O) DL1Δhsa-Cy3, CD235a-FITC and CD41a-APC were added to WB: (N) Events from gate R3, and DL1Δhsa-Cy3 bound platelets are drastically reduced compared to DL1-Cy3 bound platelets. (O) Events from gate R4. Blood samples from multiple donors were tested and consistent results were obtained.

Lingquan Deng, et al. PLoS Pathog. 2014 Dec;10(12):e1004540.
7.
Figure 4

Figure 4. Bacterial interactions with separated erythrocytes, platelets, and whole human plasma.. From: Oral Streptococci Utilize a Siglec-Like Domain of Serine-Rich Repeat Adhesins to Preferentially Target Platelet Sialoglycans in Human Blood.

(A) Wild type bacteria only hemagglutinate non-sialidase treated RBCs. Mutant strains do not show any hemagglutination. Bars represent median values. (n = 3). (B) Bacterial adhesion to fixed, immobilized platelets. (n = 6, SEM). (C) Whole plasma samples were immobilized on dot blot in three-fold serial dilutions starting from the original concentration in whole blood. Each strain was tested to interact with either non-sialidase treated (Ø) or sialidase treated whole plasma (S). *P<0.05, ***P<0.001.

Lingquan Deng, et al. PLoS Pathog. 2014 Dec;10(12):e1004540.

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