bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of aMoraxella catarrhalisOuter Membrane Protein Exhibiting Both Adhesin and Lipolytic Activities
Authors: Timpe Jennifer M. , Holm Melissa M. , Vanlerberg Serena L. , Basrur Venkatesha , Lafontaine Eric R. 
Published: July 2003

ABSTRACTThe UspA1 and Hag proteins have previously been shown to be involved in the ability of theMoraxella catarrhaliswild-type strain O35E to bind to human Chang and A549 cells, respectively. In an effort to identify novel adhesins, we generated a plasmid library ofM. catarrhalisDNA fragments, which was introduced into a nonadherentEscherichia colistrain. RecombinantE. colibacteria were subsequently enriched for clones that gained the ability to bind to Chang and A549 cells, yielding the plasmid pELFOS190. Transposon mutagenesis of this plasmid identified the potential adhesin genemcaP(M. catarrhalisadherence protein). Sequence analysis revealed that McaP is related to autotransporter proteins and has substantial similarity with the GDSL family of lipolytic enzymes, particularly theMoraxella bovisphospholipase B. Expression of themcaPgene product byE. coliincreased adherence to Chang, A549, and 16HBE14o−polarized human bronchial cells 50- to 100-fold. Spectrophotometric assays withp-nitrophenol derivatives also demonstrated that McaP is an esterase. Furthermore, thin-layer chromatography revealed that McaP cleaves both phosphatidylcholine and lysophosphatidylcholine. McaP releases fatty acids and glycerophosphorylcholine upon cleavage of phosphatidylcholine, thus exhibiting phospholipase B activity. The construction and characterization of isogenicM. catarrhalisO35E mutants demonstrated that the lack of McaP expression abolishes esterase activity and considerably decreases adherence to several human cell lines.
Effect of an Orphan Response Regulator on
Streptococcus mutans
Sucrose-Dependent Adherence and Cariogenesis
Authors: Idone Vincent , Brendtro Stacy , Gillespie Robert , Kocaj Steve , Peterson Erica , Rendi Mara , Warren Wayne , Michalek Suzanne , Krastel Kirsten , Cvitkovitch Dennis , Spatafora Grace 
Published: July 2003

ABSTRACT

Streptococcus mutans
is the principal acidogenic component of dental plaque that demineralizes tooth enamel, leading to dental decay. Cell-associated glucosyltransferases catalyze the sucrose-dependent synthesis of sticky glucan polymers that, together with glucan binding proteins, promote
S. mutans
adherence to teeth and cell aggregation. We generated an
S. mutans
Tn
916
transposon mutant, GMS315, which is defective in sucrose-dependent adherence and significantly less cariogenic than the UA130 wild-type progenitor in germfree rats. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western blotting, and N-terminal sequence analysis confirmed the absence of a 155-kDa glucosyltransferase S (Gtf-S) from GMS315 protein profiles. Mapping of the unique transposon insertion in GMS315 revealed disruption of a putative regulatory region located upstream of
gcrR
, a gene previously described by Sato et al. that shares significant amino acid identity with other bacterial response regulators (Y. Sato, Y. Yamamoto, and H. Kizaki, FEMS Microbiol. Lett. 186: 187-191, 2000). The
gcrR
regulator, which we call “
tarC
,” does not align with any of the 13 proposed two-component signal transduction systems derived from in silico analysis of the
S. mutans
genome, but rather represents one of several orphan response regulators in the genome. The results of Northern hybridization and/or real-time reverse transcription-PCR experiments reveal increased expression of both Gtf-S and glucan binding protein C (GbpC) in a
tarC
knockout mutant (GMS900), thereby supporting the notion that TarC acts as a negative transcriptional regulator. In addition, we noted that GMS900 has altered biofilm architecture relative to the wild type and is hypocariogenic in germfree rats. Taken collectively, these findings support a role for signal transduction in
S. mutans
sucrose-dependent adherence and aggregation and implicate TarC as a potential target for controlling
S. mutans
-induced cariogenesis.
Kinetics of Cellular Responses to IntraperitonealBrugia pahangiInfections in Normal and Immunodeficient Mice
Authors: Ramalingam Thirumalai , Rajan Bhargavi , Lee James , Rajan T. V. 
Published: July 2003

ABSTRACTFilarial infections evoke exuberant inflammatory responses in the peritoneal cavities of immunocompetent mice. Clearance of infection appears to be dependent on complex interactions between B1 and B2 B lymphocytes, T cells, eosinophils, macrophages, and the products of these cells. In an earlier communication, we described the course of infection in normal immunocompetent mice. In this study, we utilize mice with well-characterized mutations that disable one or more effector components of adaptive immunity in order to determine their roles in host protection. We characterize peritoneal exudate cells by flow cytometry and determine the kinetics of accumulation of each of the different cell types following infection withBrugia pahangi. We find that (i) four-color flow-cytometric analysis of peritoneal exudate cells using anti-CD3, -CD11b, -CD19, and -Gr1 can distinguish up to six different populations of cells; (ii) an initial influx of neutrophils occurs within 24 h of infection, independent of the adaptive immune status of mice, and these cells disappear by day 3; (iii) an early influx of eosinophils is seen at the site of infection in all strains studied, but a larger, second wave occurs only in mice with T cells; (iv) the presence of T cells and eosinophils is important in causing an increase in macrophage size during the course of infection; and (v) most unexpectedly, T-cell recruitment appears to be optimal only if B cells are present, since JHD mice recruit significantly fewer T cells to the site of infection.
The
Arcanobacterium pyogenes
Collagen-Binding Protein, CbpA, Promotes Adhesion to Host Cells
Authors: Esmay Paula A. , Billington Stephen J. , Link Malen A. , Songer J. Glenn , Jost B. Helen 
Published: July 2003

ABSTRACT

Arcanobacterium pyogenes
is an opportunistic pathogen associated with suppurative diseases in economically important food animals such as cattle, pigs, and turkeys.
A. pyogenes
adheres to host epithelial cells, and adhesion is promoted by the action of neuraminidase, which is expressed by this organism. However, a neuraminidase-deficient mutant of
A. pyogenes
only had a reduced ability to adhere to host epithelial cells, indicating that other factors are involved in adhesion. Far Western blotting revealed the presence of an approximately 120-kDa
A. pyogenes
cell wall protein that binds collagen type I. The 3.5-kb gene that encodes the 124.7-kDa CbpA protein was cloned, and sequence analysis indicated that CbpA contains a typical MSCRAMM protein domain structure. Recombinant, six-His-tagged CbpA (HIS-CbpA) was capable of binding collagen types I, II, and IV but not fibronectin. In addition, CbpA was involved in the ability of
A. pyogenes
to adhere to HeLa and 3T6 cells, as a
cbpA
knockout strain had 38.2 and 57.0% of wild-type adhesion, respectively. This defect could be complemented by providing
cbpA
on a multicopy plasmid. Furthermore, HIS-CbpA blocked
A. pyogenes
adhesion to HeLa or 3T6 cells in a dose-dependent manner.
cbpA
was only present in 48% of the
A. pyogenes
strains tested (
n
= 75), and introduction of plasmid-encoded
cbpA
into a naturally
cbpA
-deficient strain increased the ability of this strain to bind to HeLa and 3T6 cells 2.9- and 5.7-fold, respectively. These data indicate that CbpA, a collagen-binding protein of
A. pyogenes
, plays a role in the adhesion of this organism to host cells.
TransfectedPlasmodium knowlesiProduces Bioactive Host Gamma Interferon: a New Perspective for Modulating Immune Responses to Malaria Parasites
Authors: Ozwara Hastings , Langermans Jan A. M. , Kocken Clemens H. M. , van der Wel Annemarie , van der Meide Peter H. , Vervenne Richard A. W. , Mwenda Jason M. , Thomas Alan W. 
Published: July 2003

ABSTRACTTransgenic pathogenic microorganisms expressing host cytokines such as gamma interferon (IFN-γ) have been shown to manipulate host-pathogen interaction, leading to immunomodulation and enhanced protection. Expression of host cytokines in malaria parasites offers the opportunity to investigate the potential of an immunomodulatory approach by generating immunopotentiated parasites. Using the primate malaria parasitePlasmodium knowlesi, we explored the conditions for expressing host cytokines in malaria parasites.P. knowlesiparasites transfected with DNA constructs for expressing rhesus monkey (Macaca mulatta) IFN-γ under the control of the heterologousP. bergheiapical membrane antigen 1 promoter, produced bioactive IFN-γ in a developmentally regulated manner. IFN-γ expression had no marked effect on in vitro parasite development. Bioactivity of the parasite-produced IFN-γ was shown through inhibition of virus cytopathic effect and confirmed by usingM. mulattaperipheral blood cells in vitro. These data indicate for the first time that it is feasible to generate malaria parasites expressing bioactive host immunomodulatory cytokines. Furthermore, cytokine-expressing malaria parasites offer the opportunity to analyze cytokine-mediated modulation of malaria during the blood and liver stages of the infection.
Human Alveolar Macrophages Infected by Virulent Bacteria Expressing SipB Are a Major Source of Active Interleukin-18
Authors: Obregon Carolina , Dreher Donatus , Kok Menno , Cochand Laurence , Kiama Gitahi S. , Nicod Laurent P. 
Published: July 2003

ABSTRACTRecent publications have demonstrated that the protease caspase-1 is responsible for the processing of pro-interleukin 18 (IL-18) into the active form. Studies on cell lines and murine macrophages have shown that the bacterial invasion factor SipB activates caspase-1, triggering cell death. Thus, we investigated the role of SipB in the activation and release of IL-18 in human alveolar macrophages (AM), which are the first line of defense against inhaled pathogens. Under steady-state conditions, AM are a more important source of IL-18 than are dendritic cells (DC) and monocytes. Cytokine production by AM and DC was compared after both types of cells had been infected with a virulent strain ofSalmonella entericaserovar Typhimurium and an isogenicsipBmutant, which were used as an infection model. Infection with virulentSalmonellaled to marked cell death with features of apoptosis while both intracellular activation and release of IL-18 were demonstrated. In contrast, thesipBmutant did not induce such cell death or the release of active IL-18. The specific caspase-1 inhibitor Ac-YVAD-CMK blocked the early IL-18 release in AM infected with the virulent strain. However, the type ofSalmonellainfection did not differentially regulate IL-18 gene expression. We concluded that the bacterial virulence factor SipB plays an essential posttranslational role in the intracellular activation of IL-18 and the release of the cytokine in human AM.
Biochemical and Functional Analyses of the Mip Protein: Influence of the N-Terminal Half and of Peptidylprolyl Isomerase Activity on the Virulence of
Legionella pneumophila
Authors: Köhler Rolf , Fanghänel Jörg , König Bettina , Lüneberg Edeltraud , Frosch Matthias , Rahfeld Jens-Ulrich , Hilgenfeld Rolf , Fischer Gunter , Hacker Jörg , Steinert Michael 
Published: July 2003

ABSTRACT

The virulence factor Mip (macrophage infectivity potentiator) contributes to the intracellular survival of
Legionella pneumophila
, the causative agent of Legionnaires' disease. The protein consists of two domains that are connected via a very long α-helix (A. Riboldi-Tunnicliffe et al., Nat. Struct. Biol.
8:
779-783, 2001). The fold of the C-terminal domain (residues 100 to 213) is closely related to human FK506-binding protein (FKBP12), and like FKBP12, Mip exhibits peptidylprolyl
cis
/
trans
isomerase (PPIase) activity. The α-helical N-terminal domain is responsible for the formation of very stable Mip homodimers. In order to determine the importance of the homodimeric state of Mip for its biochemical activities and for infectivity of
Legionella
, a truncated, monomeric Mip variant [Mip
(77-213)
] was overexpressed in
Escherichia coli
and characterized biochemically. In vitro isomerase activity assays revealed that the altered protein exhibits full isomerase activity towards peptide substrates. However, the deletion resulted in a dramatic loss in the efficiency of refolding of reduced and carboxy-methylated RNase T
1
. By
cis
complementation of the Mip-negative mutant strain
L. pneumophila
JR32-2, we constructed the strain
L. pneumophila
JR32-2.4, which expresses an N-terminally truncated variant of Mip. Infection studies with these strains revealed that the N-terminal part and the dimerization of Mip but not its PPIase activity are necessary for full virulence in
Acanthamoeba castellanii
. Infection of guinea pigs showed that strains with dimerization-deficient Mip (JR32-2.4) or a very low PPIase activity (JR32-2.2) were significantly attenuated in the animal model. These results suggest a different role of the PPIase activity and the N-terminally mediated dimeric state of Mip in monocellular systems and during the infection of guinea pigs.
Oral Pretreatment of Mice with CpG DNA Reduces Susceptibility to Oral or Intraperitoneal Challenge with VirulentListeria monocytogenes
Authors: Ray Nancy B. , Krieg Arthur M. 
Published: July 2003

ABSTRACTListeria monocytogenesis an enteroinvasive intracellular bacterial pathogen that infects humans and other animals, including mice, sometimes resulting in severe systemic infections. Previous studies showed that intraperitoneal (i.p.) pretreatment of susceptible BALB/c mice with immune-stimulatory CpG DNA 48 to 96 h prior to i.p. challenge with virulentL. monocytogenesreduces bacterial numbers in livers by greater than 100-fold, correlating with recovery from infection. Here we show that oral pretreatment of BALB/c mice with CpG DNA results in decreased susceptibility to either oral or i.p. challenge withL. monocytogenes. A single dose of 200 μg of CpG DNA administered to BALB/c mice orally by gavage 48 h or 7 days before oral challenge with virulentL. monocytogenesreduces bacterial numbers approximately 10- to 100-fold in livers and spleens. Lymphotoxin alpha knockout mice lacking Peyer's patches (PPs) and pretreated orally with CpG DNA 48 h prior to oral challenge withL. monocytogenesalso have reduced susceptibility to infection, suggesting that PPs are required neither for oral infection nor for CpG-induced resistance against oral infection withL. monocytogenes. Surprisingly, 48-h oral pretreatment of BALB/c mice with 100 to 200 μg of CpG DNA results in approximately 100-fold-decreased bacterial numbers in livers following i.p. challenge withL. monocytogenes, suggesting, along with other data in this report, that orally delivered CpG DNA induces systemic resistance to infection. These results indicate that oral administration of CpG DNA induces systemic innate immune defenses against either oral or systemic infection with virulentL. monocytogenes.
Contribution of a Response Regulator to the Virulence of
Streptococcus pneumoniae
Is Strain Dependent
Authors: Blue Clare E. , Mitchell Tim. J. 
Published: July 2003

ABSTRACT

Bacterial two-component signal transduction systems (TCS) enable bacteria to respond to environmental changes and regulate a range of genes accordingly. They have a crucial role in regulating many cellular responses and have excellent potential as antibacterial-drug targets. We have constructed mutations in a TCS response regulator gene for two different strains of the human pathogen
Streptococcus pneumoniae
. These mutants have been analyzed in our murine model of infection. Data suggest that in a D39 background the response regulator gene is essential for virulence; an isogenic mutant is avirulent via intraperitoneal, intranasal, and intravenous routes of infection. This mutant, which does not show impaired growth in vitro, is unable to grow in the lung tissue or in blood. Mutation of the response regulator in a 0100993 background results in a strain that is fully virulent intraperitoneally and intravenously but shows decreased levels of bacteremia and increased murine survival following intranasal infection. The ability to grow in the lung tissue is not impaired in this mutant, suggesting that it has an impaired ability to disseminate from the lungs to the systemic circulation. Our data highlight the importance of assessing the contribution of putative virulence factors to the infection process at different sites of infection and provide evidence that virulence determinants can behave very differently based on the genetic background of the bacterial strain. These important findings may be relevant to other bacterial pathogens.
Phosphoinositide 3 Kinase Mediates Toll-Like Receptor 4-Induced Activation of NF-κB in Endothelial Cells
Authors: Li Xianwu , Tupper Joan C. , Bannerman Douglas D. , Winn Robert K. , Rhodes Christopher J. , Harlan John M. 
Published: July 2003

ABSTRACT
Many of the proinflammatory effects of gram-negative bacteria are elicited by the interaction of bacterial lipopolysaccharide (LPS) with Toll-like receptor 4 (TLR4) expressed on host cells. TLR4 signaling leads to activation of NF-κB and transcription of many genes involved in the inflammatory response. In this study, we examined the signaling pathways involved in NF-κB activation by TLR4 signaling in human microvascular endothelial cells. Akt is a major downstream target of phosphoinositide 3 kinase (PI3-kinase), and PI3-kinase activation is necessary and sufficient for Akt phosphorylation. Consequently, Akt kinase activation was used as a measure of PI3-kinase activity. In a stable transfection system, dominant-negative mutants of myeloid differentiation factor 88 (MyD88) and interleukin-1 (IL-1) receptor-associated kinase 1 (IRAK-1) (MyD88-TIR and IRAK-DD, respectively) blocked Akt kinase activity in response to LPS and IL-1β. A dominant-negative mutant (Mal-P/H) of MyD88 adapter-like protein (Mal), a protein with homology to MyD88, failed to inhibit LPS- or IL-1β-induced Akt activity. Moreover, a dominant-negative mutant of p85 (p85-DN) inhibited the NF-κB luciferase activity, IL-6 production, and IκBα degradation elicited by LPS and IL-1β but not that stimulated by tumor necrosis factor alpha. The dominant-negative mutant of Akt partially inhibited the NF-κB luciferase activity evoked by LPS and IL-1β. However, expression of a constitutively activated Akt failed to induce NF-κB luciferase activity. These findings indicate that TLR4- and IL-1R-induced PI3-kinase activity is mediated by the adapter proteins MyD88 and IRAK-1 but not Mal. Further, these studies suggest that PI3-kinase is an important mediator of LPS and IL-1β signaling leading to NF-κB activation in endothelial cells and that Akt is necessary but not sufficient for NF-κB activation by TLR4.
Modification of In Vivo and In Vitro T- and B-Cell-Mediated Immune Responses by thePseudomonas aeruginosaQuorum-Sensing MoleculeN-(3-Oxododecanoyl)-l-Homoserine Lactone
Authors: Ritchie Adam J. , Yam Andrew O. W. , Tanabe Kara M. , Rice Scott A. , Cooley Margaret A. 
Published: July 2003

ABSTRACTN-3-(oxododecanoyl)-l-homoserine lactone (OdDHL), a quorum-sensing molecule ofPseudomonas aeruginosa, plays an important role in the pathogenesis of the organism through its control of virulence factor expression. Several reports have suggested that OdDHL can also directly modulate host immune responses. However, the nature of the modulation is controversial, with different reports suggesting promotion of either humoral (Th2-mediated) or inflammatory (Th1-mediated) responses. This report describes a series of studies which demonstrate for the first time that in vivo administration of OdDHL can modulate the course of an antibody response, with an increase in ovalbumin (OVA)-specific immunogloblulin G1 (IgG1) but not IgG2a in OdDHL-treated OVA-immunized BALB/c mice compared to levels for controls. In vitro stimulation of lymphocytes from both Th1-biased C57Bl/6 and T-cell receptor transgenic mice and Th2-biased BALB/c mice in the presence of OdDHL demonstrated that OdDHL inhibits in vitro cytokine production in response to both mitogen and antigen, with gamma interferon (IFN-γ) tending to be more inhibited than interleukin-4 (IL-4). In vitro mitogen or antigen restimulation of cells from mice treated with OdDHL in vivo shows effects on cytokine production which depend on the underlying immune bias of the mouse strain used, with a relative increase of IFN-γ in Th1-biased C57Bl/6 mice and a relative increase of IL-4 in Th2-biased BALB/c mice. Thus, the mode of action of OdDHL on T-cell cytokine production is likely to be a relatively nonspecific one which accentuates an underlying immune response bias rather than one which specifically targets either Th1 or Th2 responses.
Effect of Complement Component C3 Deficiency on Experimental Lyme Borreliosis in Mice
Authors: Lawrenz Matthew B. , Wooten R. Mark , Zachary James F. , Drouin Scott M. , Weis Janis J. , Wetsel Rick A. , Norris Steven J. 
Published: July 2003

ABSTRACTMice deficient in complement component C3 (C3−/−) and syngeneic C57BL/6 control mice were challenged withBorrelia burgdorferito determine the role of complement in immune clearance and joint histopathology during experimental Lyme borreliosis. Tibiotarsal joint, ear, and heart tissues were monitored for spirochete numbers at 2, 4, 8, and 12 weeks postinoculation with 105B. burgdorferiB31 clone 5A4 by using quantitative real-time PCR. The spirochete load in joint and ear tissue remained higher in the C3−/−mice than in the wild-type counterparts throughout the 12-week study, whereas the numbers in heart tissue of both groups of mice decreased substantially at 8 to 12 weeks postinfection. Histopathology scores for joint tissue were generally higher in the C3−/−mice compared to C57BL/6 controls at 2 and 4 weeks postinfection, which may reflect the presence of higher numbers of bacteria in the joints at these early time points. Levels of anti-B. burgdorferiimmunoglobulin G tended to be reduced in the C3−/−mice compared to control mice. Furthermore, a 5.5-fold-lower number of the complement-sensitiveBorrelia gariniiwas needed to infect C3−/−mice compared to C57BL/6 mice, indicating that its sensitivity to complement is one barrier to infection of the mouse model byB. garinii. These results indicate that the complement system may be important in controlling the early dissemination and progression ofB. burgdorferiinfection.