Journal: bioRxiv

Article Title: Coping with multiple enemies: pairwise interactions do not predict evolutionary change in complex multitrophic communities

doi: 10.1101/492132

Figure Lengend Snippet: Abundance (log-transformed, mean ± std. dev., n = 7) of the focal species B. subtilis (orange circles), competitor bacterium S. marcescens (red circles), predator P. caudatum (green circles) and B. subtilis-specific parasite phage SPP1 (purple circles) in each treatment over the duration of the experiment (Days 1-10).

Article Snippet: Our experiment consisted of seven sets of species combinations of Bacillus subtilis (NCIB3610) with (1) SPP1 phage parasite in isolation; (2) Paramecium caudatum , a generalist [ ; ; ] bacterial predator in isolation; (3) Serratia marcescens (ATCC 29632), a competitor of B. subtilis , in isolation; (4) P. caudatum and SPP1 phage; (5) S. marcescens and SPP1 phage; (6) S. marcescens and P. caudatum; and (7) S. marcescens, P. caudatum and SPP1 phage, resulting in a total of 7 experimental treatments, each replicated seven times.

Techniques: Transformation Assay

Journal: bioRxiv

Article Title: Coping with multiple enemies: pairwise interactions do not predict evolutionary change in complex multitrophic communities

doi: 10.1101/492132

Figure Lengend Snippet: The effect of community composition on B. subtilis evolution relative to the ancestral strain (solid line). a . Competitive ability of B. subtilis isolates, measured as the ratio of B. subtilis to S. marcescens (dashed line indicates a 1:1 ratio, solid line represents ancestral resistance). b . Relative resistance to parasite in B. subtilis across treatment groups. Resistance to phage SPP1 was measured as the difference in growth (optical density) of B. subtilis in the presence or absence of the ancestral phage parasite. The solid line represents the resistance of ancestral B. subtilis and the resistance of our experimental treatment isolates relative to ancestral parasite resistance. c . Growth of B. subtilis isolates in the presence of the predator P. caudatum (measured as log 10 optical density of biofilm production). Solid line indicates resistance of the ancestral B. subtilis grown in the presence of P. caudatum ). In all cases, values to the right of the solid lines indicate higher relative resistance than the ancestral strain, values to the left of the solid line indicate lower resistance relative to ancestral strain of B. subtilis .

Article Snippet: Our experiment consisted of seven sets of species combinations of Bacillus subtilis (NCIB3610) with (1) SPP1 phage parasite in isolation; (2) Paramecium caudatum , a generalist [ ; ; ] bacterial predator in isolation; (3) Serratia marcescens (ATCC 29632), a competitor of B. subtilis , in isolation; (4) P. caudatum and SPP1 phage; (5) S. marcescens and SPP1 phage; (6) S. marcescens and P. caudatum; and (7) S. marcescens, P. caudatum and SPP1 phage, resulting in a total of 7 experimental treatments, each replicated seven times.

Techniques:

Journal: PloS one

Article Title: Differential expression of secreted phosphoprotein 1 in the motor cortex among primate species and during postnatal development and functional recovery.

doi: 10.1371/journal.pone.0065701

Figure Lengend Snippet: Figure 1. SPP1 mRNA expression in the sensorimotor cortex of the capuchin monkey. A: A photograph showing SPP1 mRNA expression around the central sulcus (CS) in the capuchin monkey. B, C: Nissl-stained (B) and adjacent sections in the primary motor cortex (M1) of the capuchin monkey showing the distribution of SPP1 mRNA- positive neurons (C). D: A control section hybridized with the SPP1 mRNA sense probe. E, F: Double-labeling study with SMI32, an antibody

Article Snippet: IHC was performed with a mouse anti-SPP1 monoclonal antibody (sc21742, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) by using the Vectastain Elite ABC Mouse IgG Kit (PK-6102, Vector Laboratories, Inc., Burlingame, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Staining, Control, Labeling

Journal: PloS one

Article Title: Differential expression of secreted phosphoprotein 1 in the motor cortex among primate species and during postnatal development and functional recovery.

doi: 10.1371/journal.pone.0065701

Figure Lengend Snippet: Figure 2. SPP1 expression in the motor cortex of the marmoset and squirrel monkey, and the brainstem and spinal cord of New World monkeys. A, B: Nissl-stained (A) and adjacent sections of the marmoset primary motor cortex (M1) hybridized with the SPP1 antisense probe (B). C, D: Nissl-stained (C) and adjacent sections of the squirrel monkey M1 hybridized with the SPP1 antisense probe (D). E, F: Low- (E) and high- (F) magnification photomicrographs of SPP1 mRNA- positive neurons in the red nucleus of the capuchin monkey. G, H: Low-

Article Snippet: IHC was performed with a mouse anti-SPP1 monoclonal antibody (sc21742, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) by using the Vectastain Elite ABC Mouse IgG Kit (PK-6102, Vector Laboratories, Inc., Burlingame, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Staining

Journal: PloS one

Article Title: Differential expression of secreted phosphoprotein 1 in the motor cortex among primate species and during postnatal development and functional recovery.

doi: 10.1371/journal.pone.0065701

Figure Lengend Snippet: Figure 3. Summary of the results from the species differences analysis. A: The density of SPP1 mRNA-positive neurons in each layer of the primary motor cortex (M1) in five animals: rat, marmoset, squirrel monkey, capuchin monkey, and rhesus macaque (the number of sections in each species was 6, 6, 10, 4, and 6, respectively). The mean density of SPP1 mRNA-positive neurons (6 SE) is shown. *P,0.05, **P,0.01, according to a Friedman’s one-way analysis of variance (ANOVA) with Dunn’s post-hoc tests. {P,0.05, {{P,0.01, according to a Kruskal–Wallis one-way ANOVA with Dunn’s post-hoc tests. B: The density of SPP1 protein-positive neurons in each layer of M1 in human (n = 6 sections). The mean density of SPP1 protein-positive neurons (6 SE) is shown. *P,0.05, **P,0.01, according to a Friedman’s one-way analysis of variance (ANOVA) with Dunn’s post-hoc tests. C–E: Schematic diagram of the interspecies differences in SPP1 expression in the projection neurons of motor-related structures including M1, red nucleus (RN), and lower cervical spinal segments. The motor systems are organized in a functional hierarchy from bottom to top: the spinal cord, brainstem, and motor cortex, each associated with increasing levels of complexity. SPP1 was commonly expressed in motor-related areas among all species examined; however, the level of motor hierarchy in which this gene was expressed varied among species. The regions with intense or moderate SPP1 signals are shown as red-filled circles, and those with weak or no signals are shown as gray-filled circles. See Table 4 for expression data details from the RN and spinal cord of each species. doi:10.1371/journal.pone.0065701.g003

Article Snippet: IHC was performed with a mouse anti-SPP1 monoclonal antibody (sc21742, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) by using the Vectastain Elite ABC Mouse IgG Kit (PK-6102, Vector Laboratories, Inc., Burlingame, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Functional Assay

Journal: PloS one

Article Title: Differential expression of secreted phosphoprotein 1 in the motor cortex among primate species and during postnatal development and functional recovery.

doi: 10.1371/journal.pone.0065701

Figure Lengend Snippet: Figure 4. SPP1 expression in the human motor cortex. A, B: Nissl- stained (A) and adjacent sections of the human M1 showing the distribution of SPP1protein-positive neurons (B). C, D: Control section incubated without primary antibody for the SPP1 protein (C) and incubated with primary antibody preabsorbed with SPP1 protein (D). E– H: High-magnification photomicrographs of SPP1 protein-positive neurons and scattergram showing the relationship between SPP1 signal intensity and size of the neuronal cell bodies in layers V (E, F) and III (G, H). Arrows and double arrowheads in (E) and (G) indicate neurons showing intense and weak signals, respectively. In (F) and (H), the number of neurons examined was 33, 56, and 42 in layer V, and 41, 41, and 31 in layer III; **P,0.01, ***P,0.0001, according to linear regression analysis. Squares, triangles, and small rectangles in (F) and (H) are data points from each human tissue sample. The solid, dashed, and dotted lines are linear approximations of the data represented by the squares, triangles, and small rectangles, respectively. Scale bars = 200 mm in A– D; 50 mm in E, G doi:10.1371/journal.pone.0065701.g004

Article Snippet: IHC was performed with a mouse anti-SPP1 monoclonal antibody (sc21742, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) by using the Vectastain Elite ABC Mouse IgG Kit (PK-6102, Vector Laboratories, Inc., Burlingame, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Staining, Control, Incubation

Journal: PloS one

Article Title: Differential expression of secreted phosphoprotein 1 in the motor cortex among primate species and during postnatal development and functional recovery.

doi: 10.1371/journal.pone.0065701

Figure Lengend Snippet: Figure 5. SPP1 expression in the developing macaque monkey. A–D: Nissl-stained and adjacent sections showing the distribution of SPP1 mRNA-positive neurons in the primary motor cortex (M1) of macaque monkey at postnatal day 182 (P182; A, B) and P730 (C, D). E, F:

Article Snippet: IHC was performed with a mouse anti-SPP1 monoclonal antibody (sc21742, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) by using the Vectastain Elite ABC Mouse IgG Kit (PK-6102, Vector Laboratories, Inc., Burlingame, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Staining

Journal: PloS one

Article Title: Differential expression of secreted phosphoprotein 1 in the motor cortex among primate species and during postnatal development and functional recovery.

doi: 10.1371/journal.pone.0065701

Figure Lengend Snippet: Figure 6. Changes of SPP1 expression after lesion of the lateral corticospinal tract. A–H: Nissl-stained sections and adjacent sections showing the distribution of SPP1 mRNA-positive neurons in the primary motor cortex (M1; A, B) and ventral premotor cortex (PMv; C, D) of the intact monkey, and in the contralesional M1 (co-M1; E, F) and PMv (co-PMv; G, H) of the lesioned monkey (early stage). II–VI, layers II–VI of the cerebral cortex. I, J: Double-labeling study with SMI 32, an antibody against non-phosphorylated neurofilament H. I: Localization of SPP1 mRNA–positive neurons in layer V of the lesioned monkey (late stage). J: SMI 32 immunoreactivity in the same section as (I). Arrows indicate SPP1 mRNA–positive neurons showing SMI 32 immunoreactivity. Double arrowheads indicate SMI 32-immunoreactive neurons with no SPP1 expression. K: A bar chart showing the average density of SPP1 mRNA–positive neurons in M1, PMv, and dorsal premotor cortex (PMd) of an intact monkey, and in the co-M1, co-PMv, and contralesional PMd (co-PMd) of the lesioned monkey, with standard error. *P,0.05, **P,0.01, according to the Mann-Whitney U test. L: Scattergram showing the relationship between food retrieval success rate with precision grip and the density of SPP1 mRNA–positive neurons in the co-PMv. **P,0.01, according to linear regression analysis. The dashed and solid lines are linear approximations of the data represented by triangles and circles, respectively. The triangles and circles are data points from each lesioned monkey at early and late recovery stages, respectively. Scale bars = 200 mm in A–H; 50 mm in I, J. doi:10.1371/journal.pone.0065701.g006

Article Snippet: IHC was performed with a mouse anti-SPP1 monoclonal antibody (sc21742, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) by using the Vectastain Elite ABC Mouse IgG Kit (PK-6102, Vector Laboratories, Inc., Burlingame, CA, USA) according to the manufacturer’s instructions.

Techniques: Expressing, Staining, Labeling, MANN-WHITNEY

Journal: The Journal of infectious diseases

Article Title: Osteopontin impairs host defense during pneumococcal pneumonia.

doi: 10.1093/infdis/jir185

Figure Lengend Snippet: Figure 1. Pulmonary and plasma osteopontin concentrations are elevated during pneumococcal pneumonia. Osteopontin concentrations in (A) lung and (B) plasma before and 6, 24, and 48 h after infection with 104 colony-forming units of Streptococcus pneumoniae. Data are expressed as mean 6 standard error of the mean (SEM); n 5 8 mice per group. Asterisk, P , .05; double asterisk, P , .01; triple asterisk, P , .001, compared with t 5 0. Osteopontin concentrations in culture supernatants after incubation of (C) MH-S cells and (D) primary alveolar macrophages with medium or growth- arrested S. pneumoniae (multiplicity of infection, 1:6 and 1:60 for MH-S cells; 1:20 and 1:200 for primary alveolar macrophages) for 4 h (MH-S cells) or 20 h (primary alveolar macrophages). Data are expressed as mean 6 SEM; n 5 3 per group. Asterisk, P ,.05, compared with medium. OPN, osteopontin.

Article Snippet: Effect of Osteopontin on S. pneumoniae Viability, Phagocytosis, and Phagolysosomal Fusion S. pneumoniae or Staphylococcus (S.) aureus (Newman strain) (1 3 106 bacteria/mL) was incubated in sterile normal saline in the presence of 0.8–800 ng/mL recombinant mouse osteopontin (rOPN;,1.0 endotoxin units per 1 lg as determined by the LAL assay; R&D Systems), 800 ng/mL boiled rOPN (30 min at 100 C), 800 ng/mL bovine serum albumin (BSA), or normal saline only at 37 C for 6 h. At indicated time points the number of bacteria was determined.

Techniques: Clinical Proteomics, Infection, Incubation

Journal: The Journal of infectious diseases

Article Title: Osteopontin impairs host defense during pneumococcal pneumonia.

doi: 10.1093/infdis/jir185

Figure Lengend Snippet: Figure 2. Prolonged survival and reduced bacterial growth in osteopontin knockout (KO) mice. A, Percentage survival of wild-type (WT) mice (filled symbols) and osteopontin KO mice (open symbols) after intranasal infection with 104 colony-forming units (CFU) of Streptococcus pneumoniae (n 5 14 mice per group). P value indicates the difference between groups. WT (gray) and osteopontin KO (white) mice were infected with 104 CFU of S. pneumoniae, and bacterial loads were determined 6, 24, and 48 h after infection in (B) lung, (C) blood, and (D) spleen. Data are expressed as box-and- whisker diagrams depicting the smallest observation, lower quartile, median, upper quartile, and largest observation; n 5 8 mice per group; asterisk, P ,.05; double asterisk, P ,.01; triple asterisk, P ,.001, compared with WT mice. Note to panels C and D: none of the mice in either group displayed positive blood or spleen culture results 6 h after infection; at 24 h, S. pneumoniae could be cultured from samples of the blood of only 3 of 8 osteopontin KO mice, compared with 7 of 8 WT mice and from the spleen tissue of only 1 of 7 osteopontin KO mice, compared with 7 of 8 WT mice (P , .05 and P , .01, respectively). OPN, osteopontin.

Article Snippet: Effect of Osteopontin on S. pneumoniae Viability, Phagocytosis, and Phagolysosomal Fusion S. pneumoniae or Staphylococcus (S.) aureus (Newman strain) (1 3 106 bacteria/mL) was incubated in sterile normal saline in the presence of 0.8–800 ng/mL recombinant mouse osteopontin (rOPN;,1.0 endotoxin units per 1 lg as determined by the LAL assay; R&D Systems), 800 ng/mL boiled rOPN (30 min at 100 C), 800 ng/mL bovine serum albumin (BSA), or normal saline only at 37 C for 6 h. At indicated time points the number of bacteria was determined.

Techniques: Knock-Out, Infection, Whisker Assay, Cell Culture

Journal: The Journal of infectious diseases

Article Title: Osteopontin impairs host defense during pneumococcal pneumonia.

doi: 10.1093/infdis/jir185

Figure Lengend Snippet: Figure 3. Decreased lung histopathology in osteopontin knockout (KO) mice. Representative lung histology of wild-type (WT) (A, D, G) and osteopontin KO (B, E, H ) mice at 6 h (A–C ), 24 h (D–F ), and 48 h (G–I) after intranasal infection with 104 CFU of Streptococcus pneumoniae. The lung sections are representative for 8 mice per group per time point. Hematoxilin and eosin staining, original magnification, 310. Inflammation scores are expressed as mean 6 standard error of the mean (WT mice, black bars; osteopontin KO mice, white bars; n 5 8 mice per group). Double asterisk, P ,.01, compared with WT mice. OPN, osteopontin.

Article Snippet: Effect of Osteopontin on S. pneumoniae Viability, Phagocytosis, and Phagolysosomal Fusion S. pneumoniae or Staphylococcus (S.) aureus (Newman strain) (1 3 106 bacteria/mL) was incubated in sterile normal saline in the presence of 0.8–800 ng/mL recombinant mouse osteopontin (rOPN;,1.0 endotoxin units per 1 lg as determined by the LAL assay; R&D Systems), 800 ng/mL boiled rOPN (30 min at 100 C), 800 ng/mL bovine serum albumin (BSA), or normal saline only at 37 C for 6 h. At indicated time points the number of bacteria was determined.

Techniques: Histopathology, Knock-Out, Infection, Staining

Journal: The Journal of infectious diseases

Article Title: Osteopontin impairs host defense during pneumococcal pneumonia.

doi: 10.1093/infdis/jir185

Figure Lengend Snippet: Figure 4. Osteopontin stabilizes Streptococcus pneumoniae viability in vitro. A, S. pneumoniae in saline (106 colony-forming units [CFU]/mL) was incubated with increasing doses (0.8–800 ng/mL) of recombinant osteopontin (black symbols) or saline (white symbols), and the viability of S. pneumoniae was determined over 6 h at 37C. B, S. pneumoniae in saline (106 CFU/mL) was incubated with 800 ng/mL recombinant osteopontin (filled squares), 800 ng/mL boiled recombinant osteopontin (open squares), 800 ng/mL bovine serum albumin (triangles), or saline (circles), and the viability of S. pneumoniae was determined over 6 h at 37C. Dashed lines depict detection limits. C, Osteopontin binds to S. pneumoniae. Enzyme-linked immunosorbent assay plates were coated or not coated with 1 3 108 CFU/mL S. pneumoniae type 3 (ATCC 6303) or serotype 2 (D39); coating with anti-osteopontin IgG was used as positive control. Binding was assessed using biotin-labeled recombinant mouse osteopontin. Data are means 6 standard error (n 5 4–6). Double asterisk, P , .01 vs buffer. OPN, osteopontin.

Article Snippet: Effect of Osteopontin on S. pneumoniae Viability, Phagocytosis, and Phagolysosomal Fusion S. pneumoniae or Staphylococcus (S.) aureus (Newman strain) (1 3 106 bacteria/mL) was incubated in sterile normal saline in the presence of 0.8–800 ng/mL recombinant mouse osteopontin (rOPN;,1.0 endotoxin units per 1 lg as determined by the LAL assay; R&D Systems), 800 ng/mL boiled rOPN (30 min at 100 C), 800 ng/mL bovine serum albumin (BSA), or normal saline only at 37 C for 6 h. At indicated time points the number of bacteria was determined.

Techniques: In Vitro, Saline, Incubation, Recombinant, Enzyme-linked Immunosorbent Assay, Positive Control, Binding Assay, Labeling

Journal: The Journal of infectious diseases

Article Title: Osteopontin impairs host defense during pneumococcal pneumonia.

doi: 10.1093/infdis/jir185

Figure Lengend Snippet: Figure 5. Similar bacterial growth during pneumococcal sepsis. Bacterial loads in (A) blood, (B) lung, (C) liver, and (D) spleen from wild-type (WT; gray) and osteopontin knockout (OPN KO; white) mice at 24 and 48 h after intravenous injection with 105 colony-forming units (CFU) of Streptococcus pneumoniae. Data are expressed as box-and-whisker diagrams depicting the smallest observation, lower quartile, median, upper quartile, and largest observation; n 5 8 mice per group. Dashed line depicts detection limit.

Article Snippet: Effect of Osteopontin on S. pneumoniae Viability, Phagocytosis, and Phagolysosomal Fusion S. pneumoniae or Staphylococcus (S.) aureus (Newman strain) (1 3 106 bacteria/mL) was incubated in sterile normal saline in the presence of 0.8–800 ng/mL recombinant mouse osteopontin (rOPN;,1.0 endotoxin units per 1 lg as determined by the LAL assay; R&D Systems), 800 ng/mL boiled rOPN (30 min at 100 C), 800 ng/mL bovine serum albumin (BSA), or normal saline only at 37 C for 6 h. At indicated time points the number of bacteria was determined.

Techniques: Knock-Out, Injection, Whisker Assay