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      1. Author :
        Kadurugamuwa, J. L.; Sin, L.; Albert, E.; Yu, J.; Francis, K.; DeBoer, M.; Rubin, M.; Bellinger-Kawahara, C.; Jr, T. R. Parr; Contag, P. R.
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2003
      5. Publication :
        Infection and Immunity
      6. Products :
      7. Volume :
        71
      8. Issue :
        N/A
      9. Page Numbers :
        N/A
      10. Research Area :
        N/A
      11. Keywords :
        Animals, Bioware, Xen29, Xen5, Biofilms/ growth & development, Catheterization, Central Venous/adverse effects, Chemiluminescent Measurements, Colony Count, Microbial, Disease Models, Animal, Female, Humans, Luciferases/genetics/metabolism, Mice, Mice, Inbred BALB C, Pseudomonas Infections/ microbiology, Pseudomonas aeruginosa/genetics/ growth & development, Staphylococcal Infections/ microbiology, Staphylococcus aureus/genetics/ growth & development IVIS, Xenogen
      12. Abstract :
        We have developed a rapid, continuous method for real-time monitoring of biofilms, both in vitro and in a mouse infection model, through noninvasive imaging of bioluminescent bacteria colonized on Teflon catheters. Two important biofilm-forming bacterial pathogens, Staphylococcus aureus and Pseudomonas aeruginosa, were made bioluminescent by insertion of a complete lux operon. These bacteria produced significant bioluminescent signals for both in vitro studies and the development of an in vivo model, allowing effective real-time assessment of the physiological state of the biofilms. In vitro viable counts and light output were parallel and highly correlated (S. aureus r = 0.98; P. aeruginosa r = 0.99) and could be maintained for 10 days or longer, provided that growth medium was replenished every 12 h. In the murine model, subcutaneous implantation of the catheters (precolonized or postimplant infected) was well tolerated. An infecting dose of 10 (3) to 10 (5) CFU/catheter for S. aureus and P. aeruginosa resulted in a reproducible, localized infection surrounding the catheter that persisted until the termination of the experiment on day 20. Recovery of the bacteria from the catheters of infected animals showed that the bioluminescent signal corresponded to the CFU and that the lux constructs were highly stable even after many days in vivo. Since the metabolic activity of viable cells could be detected directly on the support matrix, nondestructively, and noninvasively, this method is especially appealing for the study of chronic biofilm infections and drug efficacy studies in vivo.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/12540570
      14. Call Number :
        139339
      15. Serial :
        5926
      1. Author :
        N/A
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2009
      5. Publication :
        Infection and immunity
      6. Products :
      7. Volume :
        77
      8. Issue :
        7
      9. Page Numbers :
        N/A
      10. Research Area :
        N/A
      11. Keywords :
        Animal Structures; Animals; Antibodies, Bacterial; Antigens, Bacterial; Bacterial Proteins; Bioware; Cell Wall; Colony Count, Microbial; Female; Humans; Mice; Mice, Inbred BALB C; Neutrophils; Opsonin Proteins; Proteome; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Staphylococcal Infections; Staphylococcal Vaccines; Staphylococcus aureus; Vaccines, Subunit; Vaccines, Synthetic; Whole Body Imaging; Xen29
      12. Abstract :
        Staphylococcus aureus is an important human pathogen with increasing clinical impact due to the extensive spread of antibiotic-resistant strains. Therefore, development of a protective polyvalent vaccine is of great clinical interest. We employed an intravenous immunoglobulin (IVIG) preparation as a source of antibodies directed against anchorless S. aureus surface proteins for identification of novel vaccine candidates. In order to identify such proteins, subtractive proteome analysis (SUPRA) of S. aureus anchorless cell wall proteins was performed. Proteins reacting with IVIG but not with IVIG depleted of S. aureus-specific opsonizing antibodies were considered vaccine candidates. Nearly 40 proteins were identified by this preselection method using matrix-assisted laser desorption ionization--time of flight analysis. Three of these candidate proteins, enolase (Eno), oxoacyl reductase (Oxo), and hypothetical protein hp2160, were expressed as glutathione S-transferase fusion proteins, purified, and used for enrichment of corresponding immunoglobulin Gs from IVIG by affinity chromatography. Use of affinity-purified anti-Eno, anti-Oxo, and anti-hp2160 antibodies resulted in opsonization, phagocytosis, and killing of S. aureus by human neutrophils. High specific antibody titers were detected in mice immunized with recombinant antigens. In mice challenged with bioluminescent S. aureus, reduced staphylococcal spread was measured by in vivo imaging. The recovery of S. aureus CFU from organs of immunized mice was diminished 10- to 100-fold. Finally, mice immunized with hp2160 displayed statistically significant higher survival rates after lethal challenge with clinically relevant S. aureus strains. Taken together, our data suggest that anchorless cell wall proteins might be promising vaccine candidates and that SUPRA is a valuable tool for their identification.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/19364833
      14. Call Number :
        PKI @ catherine.lautenschlager @
      15. Serial :
        9039
      1. Author :
        Georgel, Philippe; Crozat, Karine; Lauth, Xavier; Makrantonaki, Evgenia; Seltmann, Holger; Sovath, Sosathya; Hoebe, Kasper; Du, Xin; Rutschmann, Sophie; Jiang, Zhengfan; Bigby, Timothy; Nizet, Victor; Zouboulis, Christos C; Beutler, Bruce
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2005
      5. Publication :
        Infection and immunity
      6. Products :
      7. Volume :
        73
      8. Issue :
        8
      9. Page Numbers :
        N/A
      10. Research Area :
        N/A
      11. Keywords :
        Animals; Anti-Bacterial Agents; Bioware; Chromosome Mapping; Eye Diseases; Fatty Acids, Monounsaturated; Likelihood Functions; Lod Score; Mice; Mice, Inbred C57BL; Oleic Acid; Receptors, Immunologic; Sequence Analysis, DNA; Skin; Staphylococcal Skin Infections; Stearoyl-CoA Desaturase; Streptococcus pyogenes; Time Factors; Toll-Like Receptor 2; Xen8.1, Xen20, Xen14
      12. Abstract :
        flake (flk), an N-ethyl-N-nitrosourea-induced recessive germ line mutation of C57BL/6 mice, impairs the clearance of skin infections by Streptococcus pyogenes and Staphylococcus aureus, gram-positive pathogens that elicit innate immune responses by activating Toll-like receptor 2 (TLR2). Positional cloning and sequencing revealed that flk is a novel allele of the stearoyl coenzyme A desaturase 1 gene (Scd1). flake homozygotes show reduced sebum production and are unable to synthesize the monounsaturated fatty acids (MUFA) palmitoleate (C(16:1)) and oleate (C(18:1)), both of which are bactericidal against gram-positive (but not gram-negative) organisms in vitro. However, intradermal MUFA administration to S. aureus-infected mice partially rescues the flake phenotype, which indicates that an additional component of the sebum may be required to improve bacterial clearance. In normal mice, transcription of Scd1-a gene with numerous NF-kappaB elements in its promoter--is strongly and specifically induced by TLR2 signaling. Similarly, the SCD1 gene is induced by TLR2 signaling in a human sebocyte cell line. These observations reveal the existence of a regulated, lipid-based antimicrobial effector pathway in mammals and suggest new approaches to the treatment or prevention of infections with gram-positive bacteria.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/16040962
      14. Call Number :
        PKI @ catherine.lautenschlager @
      15. Serial :
        9990
      1. Author :
        Kuklin, Nelly A; Clark, Desmond J; Secore, Susan; Cook, James; Cope, Leslie D; McNeely, Tessie; Noble, Liliane; Brown, Martha J; Zorman, Julie K; Wang, Xin Min; Pancari, Gregory; Fan, Hongxia; Isett, Kevin; Burgess, Bruce; Bryan, Janine; Brownlow, Michelle; George, Hugh; Meinz, Maria; Liddell, Mary E; Kelly, Rosemarie; Schultz, Loren; Montgomery, Donna; Onishi, Janet; Losada, Maria; Martin, Melissa; Ebert, Timothy; Tan, Charles Y; Schofield, Timothy L; Nagy, Eszter; Meineke, Andreas; Joyce, Joseph G; Kurtz, Myra B; Caulfield, Michael J; Jansen, Kathrin U; McClements, William; Anderson, Annaliesa S
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2006
      5. Publication :
        Infection and immunity
      6. Products :
      7. Volume :
        74
      8. Issue :
        4
      9. Page Numbers :
        N/A
      10. Research Area :
        N/A
      11. Keywords :
        Animals; Antibodies, Bacterial; Antigens, Bacterial; Bioware; Cation Transport Proteins; Disease Models, Animal; Female; Humans; Macaca mulatta; Mice; Mice, Inbred BALB C; Mice, Inbred ICR; Sepsis; Sequence Homology, Amino Acid; Staphylococcal Infections; Staphylococcal Vaccines; Staphylococcus aureus; Survival Rate; Xen8.1
      12. Abstract :
        Staphylococcus aureus is a major cause of nosocomial infections worldwide, and the rate of resistance to clinically relevant antibiotics, such as methicillin, is increasing; furthermore, there has been an increase in the number of methicillin-resistant S. aureus community-acquired infections. Effective treatment and prevention strategies are urgently needed. We investigated the potential of the S. aureus surface protein iron surface determinant B (IsdB) as a prophylactic vaccine against S. aureus infection. IsdB is an iron-sequestering protein that is conserved in diverse S. aureus clinical isolates, both methicillin resistant and methicillin sensitive, and it is expressed on the surface of all isolates tested. The vaccine was highly immunogenic in mice when it was formulated with amorphous aluminum hydroxyphosphate sulfate adjuvant, and the resulting antibody responses were associated with reproducible and significant protection in animal models of infection. The specificity of the protective immune responses in mice was demonstrated by using an S. aureus strain deficient for IsdB and HarA, a protein with a high level of identity to IsdB. We also demonstrated that IsdB is highly immunogenic in rhesus macaques, inducing a more-than-fivefold increase in antibody titers after a single immunization. Based on the data presented here, IsdB has excellent prospects for use as a vaccine against S. aureus disease in humans.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/16552052
      14. Call Number :
        PKI @ catherine.lautenschlager @
      15. Serial :
        9989
      1. Author :
        Kadurugamuwa, J. L.; Modi, K.; Yu, J.; Francis, K. P.; Purchio, T.; Contag, P. R.
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2005
      5. Publication :
        Infection and Immunity
      6. Products :
      7. Volume :
        73
      8. Issue :
        N/A
      9. Page Numbers :
        N/A
      10. Research Area :
        N/A
      11. Keywords :
        Animals, Diagnostic Imaging/ methods, Female, Mice, Microscopy, Electron, Scanning, Photons, Proteus Infections/ diagnosis, Proteus mirabilis/drug effects/isolation & purification, Pseudomonas Infections/ diagnosis, Pseudomonas aeruginosa/drug effects/isolation & purification, Urinary Catheterization/ adverse effects, Urinary Tract Infections/ diagnosis IVIS, Xenogen, Xen5, Xen44
      12. Abstract :
        Urinary tract infections (UTIs) are among the most common bacterial infections acquired by humans, particularly in catheterized patients. A major problem with catheterization is the formation of bacterial biofilms on catheter material and the risk of developing persistent UTIs that are difficult to monitor and eradicate. To better understand the course of UTIs and allow more accurate studies of in vivo antibiotic efficacy, we developed a catheter-based biofilm infection model with mice, using bioluminescently engineered bacteria. Two important urinary tract pathogens, Pseudomonas aeruginosa and Proteus mirabilis, were made bioluminescent by stable insertion of a complete lux operon. Segments of catheter material (precolonized or postimplant infected) with either pathogen were placed transurethrally in the lumen of the bladder by using a metal stylet without surgical manipulation. The bioluminescent strains were sufficiently bright to be readily monitored from the outside of infected animals, using a low-light optical imaging system, including the ability to trace the ascending pattern of light-emitting bacteria through ureters to the kidneys. Placement of the catheter in the bladder not only resulted in the development of strong cystitis that persisted significantly longer than in mice challenged with bacterial suspensions alone but also required prolonged antibiotic treatment to reduce the level of infection. Treatment of infected mice for 4 days with ciprofloxacin at 30 mg/kg of body weight twice a day cured cystitis and renal infection in noncatheterized mice. Similarly, ciprofloxacin reduced the bacterial burden to undetectable levels in catheterized mice but did not inhibit rebound of the infection upon cessation of antibiotic therapy. This methodology easily allows spatial information to be monitored sequentially throughout the entire disease process, including ascending UTI, treatment efficacy, and relapse, all without exogenous sampling, which is not possible with conventional methods.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/15972473
      14. Call Number :
        139333
      15. Serial :
        7110
      1. Author :
        Kadurugamuwa, J. L.; Modi, K.; Coquoz, O.; Rice, B.; Smith, S.; Contag, P. R.; Purchio, T.
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2005
      5. Publication :
        Infection and Immunity
      6. Products :
      7. Volume :
        73
      8. Issue :
        N/A
      9. Page Numbers :
        N/A
      10. Research Area :
        N/A
      11. Keywords :
        IVIS, Xenogen, Xen10
      12. Abstract :
        We developed a method for simultaneous in vivo biophotonic monitoring of pneumococcal meningitis and the accompanying neuronal injury in live transgenic mice. Streptococcus pneumoniae engineered for bioluminescence (lux) was used for direct visualization of disease progression and antibiotic treatment in a mouse model of meningitis. The host response was monitored in transgenic mice containing an inducible firefly luciferase (luc) reporter gene under transcriptional control of the mouse glial fibrillary acidic protein (GFAP) promoter. Based on the different spectra of light emission and substrate requirements for lux and luc, we were able to separately monitor the two reporters using a highly sensitive in vivo imaging system. The level of neuronal damage and recovery following antibiotic treatment was dependent on the time of treatment. This model has potential for simultaneous multiparameter monitoring and testing of therapies that target the pathogen or host response to prevent neuronal injury and recovery.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/16299273
      14. Call Number :
        139327
      15. Serial :
        7497
      1. Author :
        Srivastava, Amit; Henneke, Philipp; Visintin, Alberto; Morse, Sarah C; Martin, Victoria; Watkins, Claire; Paton, James C; Wessels, Michael R; Golenbock, Douglas T; Malley, Richard
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2005
      5. Publication :
        Infection and immunity
      6. Products :
      7. Volume :
        73
      8. Issue :
        10
      9. Page Numbers :
        6479-6487
      10. Research Area :
        N/A
      11. Keywords :
        Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Bacterial Proteins; Caspases; Lipopolysaccharides; Macrophages; Mice; Mice, Inbred Strains; Nasopharynx; Pneumococcal Infections; Streptococcus pneumoniae; Streptolysins; Xen10
      12. Abstract :
        Pneumolysin, the cholesterol-dependent cytolysin of Streptococcus pneumoniae, induces inflammatory and apoptotic events in mammalian cells. Toll-like receptor 4 (TLR4) confers resistance to pneumococcal infection via its interaction with pneumolysin, but the underlying mechanisms remain to be identified. In the present study, we found that pneumolysin-induced apoptosis is also mediated by TLR4 and confers protection against invasive disease. The interaction between TLR4 and pneumolysin is direct and specific; ligand-binding studies demonstrated that pneumolysin binds to TLR4 but not to TLR2. Involvement of TLR4 in pneumolysin-induced apoptosis was demonstrated in several complementary experiments. First, macrophages from wild-type mice were significantly more prone to pneumolysin-induced apoptosis than cells from TLR4-defective mice. In gain-of-function experiments, we found that epithelial cells expressing TLR4 and stimulated with pneumolysin were more likely to undergo apoptosis than cells expressing TLR2. A specific TLR4 antagonist, B1287, reduced pneumolysin-mediated apoptosis in wild-type cells. This apoptotic response was also partially caspase dependent as preincubation of cells with the pan-caspase inhibitor zVAD-fmk reduced pneumolysin-induced apoptosis. Finally, in a mouse model of pneumococcal infection, pneumolysin-producing pneumococci elicited significantly more upper respiratory tract cell apoptosis in wild-type mice than in TLR4-defective mice, and blocking apoptosis by administration of zVAD-fmk to wild-type mice resulted in a significant increase in mortality following nasopharyngeal pneumococcal exposure. Overall, our results strongly suggest that protection against pneumococcal disease is dependent on the TLR4-mediated enhancement of pneumolysin-induced apoptosis.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/16177320
      14. Call Number :
        PKI @ catherine.lautenschlager @
      15. Serial :
        10001
      1. Author :
        Mann, B.; Orihuela, C.; Antikainen, J.; Gao, G.; Sublett, J.; Korhonen, T. K.; Tuomanen, E.
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2006
      5. Publication :
        Infection and Immunity
      6. Products :
      7. Volume :
        74
      8. Issue :
        N/A
      9. Page Numbers :
        N/A
      10. Research Area :
        N/A
      11. Keywords :
        IVIS, Xenogen, Xen7
      12. Abstract :
        Members of the choline binding protein (Cbp) family are noncovalently bound to phosphorylcholine residues on the surface of Streptococcus pneumoniae. It has been suggested that CbpG plays a role in adherence and increase virulence both at the mucosal surface and in the bloodstream, but the function of this protein has been unclear. A new sequence analysis indicated that CbpG is a possible member of the S1 family of multifunctional surface-associated serine proteases. Clinical isolates contained two alleles of cbpG, and one-third of the strains expressed a truncated protein lacking the C-terminal, cell wall-anchoring choline binding domain. CbpG on the surface of pneumococci (full length) or released into the supernatant (truncated) showed proteolytic activity for fibronectin and casein, as did CbpG expressed on lactobacilli or as a purified full-length or truncated recombinant protein. Recombinant CbpG (rCbpG)-coated beads adhered to eukaryotic cells, and TIGR4 mutants lacking CbpG or having a truncated CbpG protein showed decreased adherence in vitro and attenuation of disease in mouse challenge models of colonization, pneumonia, and bacteremia. Immunization with rCbpG was protective in an animal model of colonization and sepsis. We propose that CbpG is a multifunctional surface protein that in the cell-attached or secreted form cleaves host extracellular matrix and in the cell-attached form participates in bacterial adherence. This is the first example of distinct functions in virulence that are dependent on natural variation in expression of a choline binding domain.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/16428724
      14. Call Number :
        140887
      15. Serial :
        6992
      1. Author :
        Orihuela, C. J.; Radin, J. N.; Sublett, J. E.; Gao, G.; Kaushal, D.; Tuomanen, E. I.
      2. Title :
      3. Type :
        Journal Article
      4. Year :
        2004
      5. Publication :
        Infection and Immunity
      6. Products :
      7. Volume :
        72
      8. Issue :
        N/A
      9. Page Numbers :
        N/A
      10. Research Area :
        N/A
      11. Keywords :
        IVIS, Xenogen, Xen7, Xen35
      12. Abstract :
        Streptococcus pneumoniae is a leading cause of invasive bacterial disease. This is the first study to examine the expression of S. pneumoniae genes in vivo by using whole-genome microarrays available from The Institute for Genomic Research. Total RNA was collected from pneumococci isolated from infected blood, infected cerebrospinal fluid, and bacteria attached to a pharyngeal epithelial cell line in vitro. Microarray analysis of pneumococcal genes expressed in these models identified body site-specific patterns of expression for virulence factors, transporters, transcription factors, translation-associated proteins, metabolism, and genes with unknown function. Contributions to virulence predicted for several unknown genes with enhanced expression in vivo were confirmed by insertion duplication mutagenesis and challenge of mice with the mutants. Finally, we cross-referenced our results with previous studies that used signature-tagged mutagenesis and differential fluorescence induction to identify genes that are potentially required by a broad range of pneumococcal strains for invasive disease.
      13. URL :
        http://www.ncbi.nlm.nih.gov/pubmed/15385455
      14. Call Number :
        141856
      15. Serial :
        6874
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