Les intégrons sont des éléments génétiques d'ADN mobiles tout comme les transposons et les plasmides R. Ils sont grandement impliqués dans la dissémination par transfert horizontal de gènes de résistance qui se retrouvent sous forme de cassette. Une cassette est constituée d'un gène associé à un site de recombinaison spécifique nommé attC reconnu par une intégrase. L'intégrase est l'enzyme capable d'exciser des gènes sous forme de cassettes et de les intégrer dans son intégron au niveau du site attl. Il existe deux types d'intégrons : les intégrons de résistance et les intégrons chromosomiques. L'intégron chromosomique de Treponema denticola est le premier à avoir été identifié chez un spirochète et est le premier exemple où le gène codant pour l'intégrase est dans le même sens que les cassettes. De plus, cet intégron semble plus près des intégrons de résistance que les autres intégrons chromosomiques. Des expériences in vivo chez E. coli nous permettent de constater que des interactions entre l'intégron chromosomique de T. denticola et les intégrons de résistance sont possibles et viennent appuyer notre hypothèse que les intégrons chromosomiques sont à l'origine des intégrons de résistance.

The conditions by which the production, in liquid media, of hemolytic activity by growing cells of T. hyodysenteriae and T. innocens are described. / From a ribonucleic acid added culture broth of T. hyodysenteriae, two hemolysins are shown in the supernatant. For each of them, a purification scheme allowing the obtention of an homogeneous hemolytic preparation by gel filtration and electrophoresis, is proposed. One hemolysin is associated with ribonucleic acid (HN) while the later is associated with bovine serum albumin (HP). They share some properties: their synthesis is blocked by chloramphenicol, they do not have proteolytic or phospholipasic activities, they are insensitive to oxygen and their activity does not require bivalent cations or is sensitive to EDTA's action; they are lytic to all red blood cell's type tested. They differ by the size of their apparent molecular weight, their mode of action, the stability of their activity to temperature and different pH and, their pattern of sensitivity to proteolytic enzymes. / The isolation of a non-hemolytic mutant strain of T. hyodysenteriae and its use in a ligated ileal loop model in rabbit has shown that there is a relation between the loss of the hemolytic activity and the loss of the enterotoxic activity.

Two-dimensional electrophoretic protein profiles of in vivo and in vitro propagated T.pallidum subsps. pallidum Nichols strain were analyzed and compared. This comparative analysis revealed two in vitro synthesized, cytoplasmic cylinder-associated polypeptides with molecular masses 29.5 and 34.7 kDa, pI 5.62, and one in vitro "lost" polypeptide with molecular mass 34.7 kDa, pI 5.34. integral membrane proteins of in vitro and in vivo propagated T. pallidum was identified by phase partitioning with the nonionic Triton X-114, and twelve outer membrane-associated, antigenic proteins were identified in western blots probed with pooled human secondary syphilitic sera. The solubilization of the outer membrane of T. pallidum with Triton X-114 were monitored by electron microscopy. Treatment of freshly harvested 35S labeled T. pallidum with 1% Triton X-114 resulted in solubilization of the outer membrane and reduction of the diameter of the treponemes from .14 +/- .02 micrometers to .095 +/- .003 micrometers. Examination of thin sections of untreated organisms showed integrity of outer and cytoplasmic membranes. In contrast, thin sections of Triton X-114-treated trponemes showed integrity of the cytoplasmic membrane but the loss of the outer membrane. The cytoplasmic cylinders generated by detergent treatment retained their periplasmic flagella, as judged by electron microscopy and immunoblotting. Integral membrane proteins of Treponema phagedenis were also identified by phase partitioning with Triton X-114, and sizteen cross-reactive, outer membrane-associated, outer membrane-associated, antigenic polypeptides were identified in western blots probed with pooled human secondary syphilitic sera. The results of this study indicate that tow-dimensional protein profiles of in vivo and in vitro propagated T.pallidum are almost identical except for the differences mentioned. This results also indicate that 1% Triton X-114 selectively solubilizes the outer membrane, and the antigenic hydrophobic proteins present in the detergent phrase are located exclusively in the outer membrane.

No description available.

The major outer sheath protein (Msp) is the most prominent surface antigen of the periodontal pathogen Treponema denticola. It mediates adhesion to extracellular matrix and dysregulation of cytoskeletal homeostasis of host cells. Disassembly of actin filaments and the coincident subcortical de novo synthesis of actin filaments in fibroblasts upon exposure to Msp were investigated with a barbed-end fluorescent labeling method. The functional impact of actin cytoskeleton disorganization was determined with a scratch wound migration assay in fibroblast monolayers and a videomicroscopy migration assay in neutrophils. Msp pretreatment had a significant inhibitory effect on the migration of the fibroblasts across a collagen substratum and inhibited the neutrophil chemotactic migration towards a chemoattractant. In a study originally aimed to find the biologically active domains of Msp that may perturb actin, short peptides were selected from the deduced and predicted surface exposed regions of Msp and investigated for their role in actin dynamics and cell motility. A novel BSA-conjugated peptide (P34BSA) was found serendipitously to induce stress fiber formation and stability in fibroblasts. This activity was found to be mediated by Rho activation and cofilin phosphorylation, which are important tandem signaling pathways in the regulation of a variety of actin-dependent cellular functions. P34BSA was internalized by the cells. Yet, a mechanistic study using low-temperature treatments and depletion of cholesterol with methyl-β-cyclodextrin (MβCD) revealed that P34BSA most likely induces actin stress fiber formation extracellularly through a Rho-dependent signaling pathway. P34BSA induced Rho activation via binding of guanosine nucleotide exchange factor p114RhoGEF to RhoA, one of many exchange factors that have been shown to play a role in activating Rho signaling. Pretreatment with P34BSA partially protected the fibroblasts against the actin-disrupting effects of cytochalasin D and latrunculin B, and the cells maintained most of their actin filaments. P34BSA treatment caused retardation of fibroblast migration on a collagen substratum. It also inhibited the chemotactic movement of neutrophils towards the chemoattractant fMLP. P34 may represent a novel amino acid sequence of a bacterial virulence protein that, when conjugated to BSA, can be used as a chemical reagent to investigate RhoA signaling pathways in host cells.

The major outer sheath protein (Msp) is the most prominent surface antigen of the periodontal pathogen Treponema denticola. It mediates adhesion to extracellular matrix and dysregulation of cytoskeletal homeostasis of host cells. Disassembly of actin filaments and the coincident subcortical de novo synthesis of actin filaments in fibroblasts upon exposure to Msp were investigated with a barbed-end fluorescent labeling method. The functional impact of actin cytoskeleton disorganization was determined with a scratch wound migration assay in fibroblast monolayers and a videomicroscopy migration assay in neutrophils. Msp pretreatment had a significant inhibitory effect on the migration of the fibroblasts across a collagen substratum and inhibited the neutrophil chemotactic migration towards a chemoattractant. In a study originally aimed to find the biologically active domains of Msp that may perturb actin, short peptides were selected from the deduced and predicted surface exposed regions of Msp and investigated for their role in actin dynamics and cell motility. A novel BSA-conjugated peptide (P34BSA) was found serendipitously to induce stress fiber formation and stability in fibroblasts. This activity was found to be mediated by Rho activation and cofilin phosphorylation, which are important tandem signaling pathways in the regulation of a variety of actin-dependent cellular functions. P34BSA was internalized by the cells. Yet, a mechanistic study using low-temperature treatments and depletion of cholesterol with methyl-β-cyclodextrin (MβCD) revealed that P34BSA most likely induces actin stress fiber formation extracellularly through a Rho-dependent signaling pathway. P34BSA induced Rho activation via binding of guanosine nucleotide exchange factor p114RhoGEF to RhoA, one of many exchange factors that have been shown to play a role in activating Rho signaling. Pretreatment with P34BSA partially protected the fibroblasts against the actin-disrupting effects of cytochalasin D and latrunculin B, and the cells maintained most of their actin filaments. P34BSA treatment caused retardation of fibroblast migration on a collagen substratum. It also inhibited the chemotactic movement of neutrophils towards the chemoattractant fMLP. P34 may represent a novel amino acid sequence of a bacterial virulence protein that, when conjugated to BSA, can be used as a chemical reagent to investigate RhoA signaling pathways in host cells.

The Gilbert’s potoroo (Potorous gilbertii) is a small marsupial endemic to the Two Peoples Bay Nature Reserve in the south-west of Western Australia. The Gilbert’s potoroo is classified as Australia’s most critically endangered mammal (IUCN 2006) with an estimated population of only 35 individuals. This thesis examines the health and disease status of the Gilbert’s potoroo, presenting a strong case for the relatively new concept of disease as a potential threatening factor and modifier of population decline. Specific diseases, including Cryptococcus, ectoparasitism, endoparasitism, haemoparasitism, Toxoplasma and a novel Treponema organism are extensively studied. An assessment of the clinical significance of these diseases is made, and management strategies are recommended to minimise the impact of these diseases on both the wild and captive population. The novel Treponema organism which clinically presents with tenacious, green discharge and an associated balanoposthitis in males is molecularly characterized. Epidemiological studies show the effects of this agent on reproductive function and a penicillin-based treatment regime is trialled in the analogous long-nosed potoroo (Potorous tridactylus) with a recommendation to then trial this treatment regime in the critically endangered Gilbert’s potoroo. Standard haematological and urinalysis findings are tabulated to form reference ranges for this species. A treatment regime for Cryptococcus in the analogous long-nosed potoroo is reported and parasitological findings, including the identification of a novel tick species are discussed. This thesis addresses key health issues, which have subsequently been incorporated into the Recovery Plan of the Gilbert’s potoroo. A document encompassing multiple disciplines and expertise to support the recovery of this critically endangered marsupial in its current environment. In addition, this thesis outlines a recommended health monitoring and treatment protocol for future translocation procedures and provides a working example of the emerging importance of health monitoring in threatened species recovery programs.

Zugl.: Berlin, Univ., Diss., 2008

Treponema denticola and Treponema vincentii were grown in a medium supplemented with 0.4% (wt/vol) alpha globulin in place of whole serum. Other serum fractions did not support growth. The growth factors in alpha globulin were destroyed by trypsin and by lipase. Lipid extraction of alpha globulin showed that both a protein and a lipid fraction were required for growth. Sodium salts of either oleic acid (cis-18:1) or elaidic acid (trans-18:1), added to 0.4% delipified alpha globulin supplemented media at a final concentration of 0.04 mg/ml, replaced the alpha globulin lipids required for optimal growth of these two oral treponemes. Tween 80 (polysorbitan monooleate) also supported growth in a medium containing protein. Short chain fatty acids plus 25 µg/ml thiamine pyrophosphate, added to either a basal medium or a medium containing 0.4% albumin, supported limited growth. The principal cellular fatty acids of T. denticola grown in an oleate medium were myristic, pentadecanoic, and palmitic acids. Treponema denticola appears capable of limited synthesis of cellular fatty acids from oleate. Fifty percent of the total protein content of commercial alpha globulin was found to be albumin. The protein required for T. denticola growth was separated from the other alpha globulin proteins by Affi-Gel Blue (Bio-Rad Laboratories) affinity chromatography which selectively adsorbed albumin. Serum albumin, added to a medium containing oleate, substituted for the alpha globulin protein required by these two treponemes. Trypsin destroyed the growth promoting activity of albumin. A weight ratio of albumin to sodium oleate of 50:1 (0.4% delipified albumin - 0.08 mg/ml oleate) supported optimal growth of T. denticola and T. vincentii. Starch, added to media containing oleate, could not replace albumin for optimal growth. Serum albumin solutions tightly bound added thiamine pyrophosphate (TPP). Optimal growth was achieved only when the TPP concentrations in albumin-oleate media were sufficient to provide excess TPP, unbound to albumin. Whole cells of T. denticola were shown to have proteolytic activity toward casein and alpha globulin proteins. Alpha globulin proteins were also found avidly attached to T. denticola cells that had been suspended in alpha globulin. / Ph. D.

Zugl.: Berlin, Freie Univ., Diss., 2004 / Dateiformat: zip, dateien im PDF-Format. _ Erscheinungsjahr an der Haupttitelstelle: 2004