1. Volume 20, Issue 4, Pages 535-547 (October 2016)
Diverse Intestinal Bacteria Contain Putative Zwitterionic Capsular Polysaccharides with Anti-inflammatory Properties 
C. Preston Neff, Matthew E. Rhodes, Kathleen L. Arnolds, Colm B. Collins, Jody Donnelly, Nichole Nusbacher, Paul Jedlicka, Jennifer M. Schneider, Martin D. McCarter, Michael Shaffer, Sarkis K. Mazmanian, Brent E. Palmer, Catherine A. Lozupone 
Cell Host & Microbe 
Volume 20, Issue 4, Pages (October 2016)
DOI: /j.chom
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2. Cell Host & Microbe 2016 20, 535-547DOI: (10.1016/j.chom.2016.09.002)
Copyright © 2016 Elsevier Inc. Terms and Conditions

3. Figure 1
PSA Operon of B. fragilis NCTC 9343 and Chemical Structures of B. fragilis NCTC 9343 PSA, B. fragilis 638R PSA2, and S. pneumoniae SP1
(A) The PSA operon of B. fragilis and the conserved properties of B. fragilis AATGal-ZPS operons, as described by Coyne et al. (2001)
(B) Chemical structure of PSA of B. fragilis and related ZPSs. The shaded regions indicate the amino sugar AATGal that is synthesized by the wcfR gene and transferred by the wcfS gene to a polysaccharide backbone that varies in structure between these ZPSs. Genes shared across Bacteroidales AATGal-ZPSs, and examples of other AATGal-ZPS operons, are shown in Figures S2–S4.
Cell Host & Microbe  , DOI: ( /j.chom )
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4. Figure 2
A Neighbor-Joining Tree of BLAST Hits to the wcfR Gene of B. fragilis with an e-Value < 1 × 10−90
Highly related sequences from different strains of the same species (or undesignated strains that are likely the same species) are collapsed into a wedge with the number of strains (genomes) within that wedge noted. Colors indicate the taxonomic order of the bacteria from which the wcfR sequence came. The wcfR genes of bacterial strains tested in immune assays are indicated with a star, showing that tested bacteria include those with wcfR homologs throughout this phylogeny. Figure S7 shows electron micrograph evidence of a convergence in surface structures in bacteria that are very distantly related phylogenetically but share highly similar wcfR gene homologs in their genomes.
Cell Host & Microbe  , DOI: ( /j.chom )
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5. Figure 3
16S rRNA Trees of All Genomes That Were Screened in the Orders Bacteroidales and Erysipelotrichales Showing Presence/Absence of PSA-like Operons
(A) Strains surveyed in the Bacteroidales order.
(B) Strains surveyed in the Erysipelotrichales order.
Species shown in red text had genes encoding both wcfR and wcfS, and the genes were in adjacent or close positions within the operon. Those without a close homolog to wcfR are in black. Nodes in which all bacteria had the same color designation are collapsed into wedges with the number of strains noted in parentheses. The stars indicate predicted PSA-producing (in red) and non-PSA-producing (in black) bacterial strains selected for immunologic testing.
Cell Host & Microbe  , DOI: ( /j.chom )
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6. Figure 4
WT B. fragilis Induces Higher IL-10 Production and Proportion of Tregs Compared to B. fragilis ΔPSA in Human PBMCs and LPMCs
(A) Levels of IL-10 in the supernatant of PBMC cultured with WT B. fragilis and B. fragilis ΔPSA for 3 days as determined by ELISA.
(B and C) Percent IL-10+ CD4+ T cells (B) and CD4+IL-10+ T cells (C) that are CD25+/−FoxP3+/− after 1-day stimulation with WT B. fragilis.
(D) Proportion of CD4+ T cells that are CD25+FoxP3+ in PBMC after 3 days of culture with WT B. fragilis or B. fragilis ΔPSA.
(E) Proportion CD4+ T cells that are CD25+FoxP3+CD127-CTLA4+ in PBMC after 3 days of culture with WT B. fragilis and B. fragilis ΔPSA.
(F and G) Levels of IL-10 in the supernatant (F) and proportion of Tregs generated from purified naive T cells mixed with bacteria stimulated purified CD14+ monocytes (G).
(H and I) Levels of IL-10 in the supernatant (H) and proportion of CD4+ T cells that are CD25+FoxP3+CD127-CTLA4+ of LPMC (I) after 3 days of culture with bacterial lysates.
Statistical significance was calculated by paired t tests. Data for other cytokines (IL-6, TNF-α, IL-17, and IL-22) are in Figure S5A. Representative staining for CD25+FoxP3+CTLA4+CD127− Tregs in human PBMCs and LPMCs are in Figure S5B.
Cell Host & Microbe  , DOI: ( /j.chom )
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7. Figure 5
Putative PSA Producers Induce Increased Levels of IL-10 and Tregs
(A–D) Putative PSA producers in the Bacteroidales or Erysipelotrichales orders induce more IL-10 and Tregs compared to their non-PSA-producing relatives in PBMCs and LPMCs.
IL-10 levels (A) and proportion of CD25+FoxP3+CTLA4+CD127− cells of CD4+ T cells (B) in PBMCs cultured with bacteria from the Bacteroidales (circles) or Erysipelotrichales order (squares) for 3 days. IL-10 levels (C) and proportion CD25+FoxP3+CTLA4+CD127− cells of CD4+ T cells (D) in LPMC cultured with bacteria from the Bacteroidales (circles) or Erysipelotrichales order (squares) for 3 days. Predicted PSA producers are in red and those that are not are in black. Statistical significance between medians of predicted and non-predicted PSA producers was calculated by nonparametric Mann-Whitney tests.
(E–H) WT B. cellulosilyticus induces higher IL-10 production and proportion of Tregs compared to a wcfR knockout in whole PBMCs and purified naive T cells.
Levels of IL-10 (E) and proportion CD4+ T cells that are CD25+FoxP3+CD127-CTLA4+ in PBMCs (F) after 3 days of culture with B. cellulosilyticus and B. cellulosilyticus ΔZPS1. Levels of IL-10 in the supernatant (G) and proportion of Tregs generated from purified naive T cells mixed with bacteria stimulated purified CD14+ monocytes (H). Statistical significance was calculated by paired t tests. Comparisons of stimulations with WT B. cellulosilyticus and the closely related B. intestinalis are in Figure S6.
Cell Host & Microbe  , DOI: ( /j.chom )
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8. Figure 6
B. cellulosyliticus Administration Attenuates Murine Colitis In Vivo
(A) Representative micrographs of H&E stained colonic sections demonstrate that B. cellulosyliticus causes a decrease in intestinal inflammation relative to either vehicle controls or mice gavaged with B. intestinalis (black scale bar, 200 μm).
(B) Histological inflammatory index scores show protection from inflammation in mice gavaged with B. cellulosilyticus relative to vehicle control or B. intestinalis.
(C) Weight loss curve data correlate with attenuated inflammation seen in histological scores.
(D and E) Representative contour plots showing expression of FoxP3+ T cells from the colonic lamina propria of each group (D), then enumerated (E).
∗p < 0.05, ∗∗p < 0.01 with unpaired one-tailed t tests. Error bars in (B), (C), and (E) represent mean ± SEM.
Cell Host & Microbe  , DOI: ( /j.chom )
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