bluuebunny/crossref_metadata_2025_split huggingface dataset:
Antimicrobial Activity of Murine Lung Cells against
Staphylococcus aureus
Is Increased In Vitro and In Vivo after Elafin Gene Transfer
Authors: McMichael J. W. , Maxwell A. I. , Hayashi K. , Taylor K. , Wallace W. A. , Govan J. R. , Dorin J. R. , Sallenave J.-M. 
Published: May 2005

ABSTRACT

Staphylococcus aureus
is a pathogen often found in pneumonia and sepsis. In the context of the resistance of this organism to conventional antibiotics, an understanding of the regulation of natural endogenous antimicrobial molecules is of paramount importance. Previous studies have shown that both human and mouse airways express a variety of these molecules, including defensins, cathelicidins, and the four-disulfide core protein secretory leukocyte protease inhibitor. We demonstrate here by culturing mouse tracheal epithelial cells at an air-liquid interface that, despite the production of
Defb1
,
Defb14
, and
Defr1
in this system, these cells are unable to clear
S. aureus
when exposed to this respiratory pathogen. Using an adenovirus (Ad)-mediated gene transfer strategy, we show that overexpression of elafin, an anti-elastase/antimicrobial molecule (also a member of the four-disulfide core protein family), dramatically improves the clearance of
S. aureus
. In addition, we also demonstrate that this overexpression is efficient in vivo and that intratracheal instillation of Ad-elafin significantly reduced the lung bacterial load and demonstrates concomitant anti-inflammatory activity by reducing neutrophil numbers and markers of lung inflammation, such as bronchoalveolar lavage levels of tumor necrosis factor and myeloperoxidase. These findings show that an increased antimicrobial activity phenotype is provided by the elafin molecule and have implications for its use in
S. aureus
-associated local and systemic infections.
Bordetella bronchisepticaAdherence to Cilia Is Mediated by Multiple Adhesin Factors and Blocked by Surfactant Protein A
Authors: Edwards Jessica A. , Groathouse Nathan A. , Boitano Scott 
Published: May 2005

ABSTRACTIn the virulent state (Bvg+),Bordetella bronchisepticaexpresses adhesins and toxins that mediate adherence to the upper airway epithelium, an essential early step in pathogenesis. In this study, we used a rabbit tracheal epithelial cell binding assay to test how specific host or pathogen factors contribute to ciliary binding. The host antimicrobial agent surfactant protein A (SP-A) effectively reduced ciliary binding by Bvg+B. bronchiseptica. To evaluate the relative contributions of bacterial adhesins and toxins to ciliary binding, we used mutant strains ofB. bronchisepticain the binding assay. When compared to Bvg+or Bvg−phase-lockedB. bronchisepticastrains, single-knockout strains lacking one of the known adhesins (filamentous hemagglutinin, pertactin, or fimbriae) displayed an intermediate ciliary binding capacity throughout the coincubation. AB. bronchisepticastrain deficient in adenylate cyclase-hemolysin toxin also displayed an intermediate level of adherence between Bvg+and Bvg−strains and had the lowest ciliary affinity of any of the Bvg+phase strains tested. AB. bronchisepticastrain that was missing dermonecrotic toxin also displayed intermediate binding; however, this strain displayed ciliary binding significantly higher than most of the adhesin knockouts tested. Taken together, these findings suggest that virulent-stateB. bronchisepticaexpresses multiple adhesins with overlapping contributions to ciliary adhesion and that host production of SP-A can provide innate immunity by blocking bacterial adherence to the ciliated epithelium.
Characterization of Fluorescent Chimeras of Cholera Toxin and
Escherichia coli
Heat-Labile Enterotoxins Produced by Use of the Twin Arginine Translocation System
Authors: Tinker Juliette K. , Erbe Jarrod L. , Holmes Randall K. 
Published: May 2005

ABSTRACT

Cholera toxin (CT) is an AB
5
toxin responsible for the profuse secretory diarrhea resulting from
Vibrio cholerae
infection. CT consists of a pentameric, receptor-binding B subunit (CTB) and a monomeric A subunit (CTA) that has latent enzymatic activity. In addition to its enterotoxicity, CT has potent mucosal adjuvant activity and can also function as a carrier molecule with many potential applications in cell biology. In earlier studies, the toxic CTA
1
domain was replaced by several other antigenic protein domains to produce holotoxin-like chimeras for use as potential mucosal vaccines. In the present study we utilized the twin arginine translocation (
tat
) system to produce fluorescent CT chimeras, as well as fluorescent chimeras of
Escherichia coli
heat-labile toxins LTI and LTIIb. Fusion proteins containing either green fluorescent protein (GFP) or monomeric red fluorescent protein (mRFP) and the A
2
domain of CT, LTI, or LTIIb were transported to the periplasm of
E. coli
by the
tat
system, and the corresponding B polypeptides of CT, LTI, and LTIIb were transported to the periplasm by the
sec
system. The fluorescent fusion proteins were shown to assemble spontaneously and efficiently with the corresponding B polypeptides in the periplasm to form chimeric holotoxin-like molecules, and these chimeras bound to and entered cultured cells in a manner similar to native CT, LTI, or LTIIb. The GFP and mRFP derivatives of CT, LT, and LTIIb developed here are useful tools for studies on the cell biology of trafficking of the CT/LT family of bacterial enterotoxins. In addition, these constructs provide proof in principle for the development of novel chimeric CT-like or LT-like vaccine candidates containing CTA
2
fusion proteins that cannot be delivered to the periplasm of
E. coli
by use of the
sec
secretion pathway.
Infection of Newborn Piglets with
Bordetella pertussis
: a New Model for Pertussis
Authors: Elahi S. , Brownlie R. , Korzeniowski J. , Buchanan R. , O'Connor B. , Peppler M. S. , Halperin S. A. , Lee S. F. , Babiuk L. A. , Gerdts V. 
Published: May 2005

ABSTRACT

Bordetella pertussis
is the causative agent of pertussis or whooping cough. This bacterium is a human pathogen that under experimental conditions also infects selected rodents and primates. Here, we show for the first time that newborn piglets can be infected with
B. pertussis
when it is delivered intrapulmonarily. Infected piglets displayed fever and respiratory symptoms, such as nasal discharge, nonparoxysmal coughing, and breathing difficulties. Eventually, all infected animals developed severe bronchopneumonia, which in some cases was combined with a fibrinous pleuritits. Immunohistochemical staining revealed the presence of large numbers of
B. pertussis
cells within airways, adhering to the epithelial lining or phagocytosed by macrophages and neutrophils. Viable bacteria were reisolated from bronchoalveolar lavages and lung lesions for more than 10 days postinfection. The systemic presence of pertussis toxin was shown by hypoglycemia, lymphocytosis, and induction of a clustered pattern of CHO cells by serum and bronchoalveolar lavage samples. Thus, a large-animal model for pertussis was developed, which should complement existing rodent models for identifying the immune responses relevant to the design of new vaccines. In particular, this model should help researchers analyze the roles of both maternal and mucosal immunity in disease protection against pertussis and should ultimately assist in the design of new vaccines for early life protection.
Comparative Secretome Analyses of Three
Bacillus anthracis
Strains with Variant Plasmid Contents
Authors: Lamonica Janine M. , Wagner MaryAnn , Eschenbrenner Michel , Williams Leanne E. , Miller Tabbi L. , Patra Guy , DelVecchio Vito G. 
Published: May 2005

ABSTRACT

Bacillus anthracis
, the causative agent of anthrax, secretes numerous proteins into the extracellular environment during infection. A comparative proteomic approach was employed to elucidate the differences among the extracellular proteomes (secretomes) of three isogenic strains of
B. anthracis
that differed solely in their plasmid contents. The strains utilized were the wild-type virulent
B. anthracis
RA3 (pXO1
+
pXO2
+
) and its two nonpathogenic derivative strains: the toxigenic, nonencapsulated RA3R (pXO1
+
pXO2
−
) and the totally cured, nontoxigenic, nonencapsulated RA3:00 (pXO1
−
pXO2
−
). Comparative proteomics using two-dimensional gel electrophoresis followed by computer-assisted gel image analysis was performed to reveal unique, up-regulated, or down-regulated secretome proteins among the strains. In total, 57 protein spots, representing 26 different proteins encoded on the chromosome or pXO1, were identified by peptide mass fingerprinting. S-layer-derived proteins, such as Sap and EA1, were most frequently observed. Many sporulation-associated enzymes were found to be overexpressed in strains containing pXO1
+
. This study also provides evidence that pXO2 is necessary for the maximal expression of the pXO1-encoded toxins lethal factor (LF), edema factor (EF), and protective antigen (PA). Several newly identified putative virulence factors were observed; these include enolase, a high-affinity zinc uptake transporter, the peroxide stress-related alkyl hydroperoxide reductase, isocitrate lyase, and the cell surface protein A.
Characterization of CD8+T Cells and Microenvironment in Oral Lesions of Human Immunodeficiency Virus-Infected Persons with Oropharyngeal Candidiasis
Authors: McNulty Kelly M. , Plianrungsi Jananya , Leigh Janet E. , Mercante Donald , Fidel Paul L. 
Published: May 2005

ABSTRACTOropharyngeal candidiasis (OPC), the most common oral infection in human immunodeficiency virus-positive persons, correlates with reduced blood CD4+T cells. In those with OPC, CD8+T cells accumulate at the lamina propria-epithelium interface at a distance from the organism at the outer epithelium. The present study aimed to characterize the tissue-associated CD8+T cells and tissue microenvironment in both OPC+and OPC−persons. The results show that the majority of CD8+T cells possess the αβ T-cell receptor, the thymus-derived αβ CD8 antigen heterodimer, and similar levels of the α4β7, α4β1, and αeβ7homing receptors. Studies to evaluate the tissue microenvironment showed that in OPC+persons, the adhesion molecule for T cells to enter mucosa, mucosal addressin cell adhesion molecule, is significantly increased, whereas E-cadherin, which allows T cells to migrate through mucosa, is significantly decreased compared to OPC−persons. These results continue to support a role for CD8+T cells against OPC under conditions of reduced numbers of CD4+T cells, with susceptibility to infection potentially associated with a dysfunction in mucosal CD8+T-cell migration by reduced tissue-associated E-cadherin.
Differential Effects of Total and Partial Neutralization of Tumor Necrosis Factor on Cell-Mediated Immunity toMycobacterium bovisBCG Infection
Authors: Guler Reto , Olleros Maria L. , Vesin Dominique , Parapanov Roumen , Garcia Irene 
Published: May 2005

ABSTRACTThe effects of total and partial inhibition of tumor necrosis factor (TNF) on sensitivityto Mycobacterium bovisBCG infection were investigated by using transgenic mice in which hepatocytes produced different amounts of human soluble TNF receptor 1 (sTNFR1) fused to the Fc fragment of human immunoglobulin G3 that could be detected in the serum. Transgenic mice expressing high serum levels of sTNFR1, neutralizing all circulating TNF, failed to develop differentiated granulomas and bactericidal mechanisms, and they succumbed to BCG infection. sTNFR1 transgenic mice did not activate BCG-induced Th1-type cytokines early in infection, but uncontrolled cytokine release was found late in infection. In this work we also evaluated the effect of partial inhibition of TNF on resistance to BCG infection. Transgenic mice expressing low levels of sTNFR1 were protected against BCG infection, and they developed increased bactericidal mechanisms, such as enhanced inducible nitric oxide synthase activity, increased macrophage activation, and showed higher numbers of liver granulomas early in infection compared to their negative littermates. Our data suggest that while total inhibition of TNF prevented BCG-induced cell-mediated immune responses, partial inhibition of TNF could contribute to macrophage activation, induction of bactericidal mechanisms, and granuloma formation in the early phase of BCG infection.
Phase 1 Clinical Trial of Apical Membrane Antigen 1: an Asexual Blood-Stage Vaccine for
Plasmodium falciparum
Malaria
Authors: Malkin Elissa M. , Diemert David J. , McArthur Julie H. , Perreault John R. , Miles Aaron P. , Giersing Birgitte K. , Mullen Gregory E. , Orcutt Andrew , Muratova Olga , Awkal May , Zhou Hong , Wang Jin , Stowers Anthony , Long Carole A. , Mahanty Siddhartha , Miller Louis H. , Saul Allan , Durbin Anna P. 
Published: May 2005

ABSTRACT

Apical membrane antigen 1 (AMA1), a polymorphic merozoite surface protein, is a leading blood-stage malaria vaccine candidate. A phase 1 trial was conducted with 30 malaria-naïve volunteers to assess the safety and immunogenicity of the AMA1-C1 malaria vaccine. AMA1-C1 contains an equal mixture of recombinant proteins based on sequences from the FVO and 3D7 clones of
Plasmodium falciparum
. The proteins were expressed in
Pichia pastoris
and adsorbed on Alhydrogel. Ten volunteers in each of three dose groups (5 μg, 20 μg, and 80 μg) were vaccinated in an open-label study at 0, 28, and 180 days. The vaccine was well tolerated, with pain at the injection site being the most commonly observed reaction. Anti-AMA1 immunoglobulin G (IgG) was detected by enzyme-linked immunosorbent assay (ELISA) in 15/28 (54%) volunteers after the second immunization and in 23/25 (92%) after the third immunization, with equal reactivity to both AMA1-FVO and AMA1-3D7 vaccine components. A significant dose-response relationship between antigen dose and antibody response by ELISA was observed, and the antibodies were predominantly of the IgG1 isotype. Confocal microscopic evaluation of sera from vaccinated volunteers demonstrated reactivity with
P. falciparum
schizonts in a pattern similar to native parasite AMA1. Antigen-specific in vitro inhibition of both FVO and 3D7 parasites was achieved with IgG purified from sera of vaccinees, demonstrating biological activity of the antibodies. To our knowledge, this is the first AMA1 vaccine candidate to elicit functional immune responses in malaria-naïve humans, and our results support the further development of this vaccine.
Strain-Specific Effects of Probiotics on Gut Barrier Integrity following Hemorrhagic Shock
Authors: Luyer Misha D. , Buurman Wim A. , Hadfoune M'hamed , Speelmans Gea , Knol Jan , Jacobs Jan A. , Dejong Cornelis H. C. , Vriesema Aldwin J. M. , Greve Jan Willem M. 
Published: May 2005

ABSTRACT

Probiotic therapy modulates the composition of the intestinal flora and inhibits the inflammatory response. These properties may be of benefit in the preservation of gut barrier integrity after injury or stress. In this study, we examined the effect of two
Lactobacillus
strains selected for their pathogen exclusion properties on intestinal barrier integrity following hemorrhagic shock. Additionally, the responsiveness of the macrophage cell line RAW 264.7 to combined exposure to
Lactobacillus
DNA or oligodeoxynucleotides containing CpG motifs (CpG-ODN) and endotoxin was assessed by measuring tumor necrosis factor alpha (TNF-α) release. Rats were administered lactobacilli (5 × 10
9
CFU) or vehicle for 7 days and were subjected subsequently to hemorrhagic shock by withdrawal of 2.1 ml blood/100 g tissue. Levels of plasma endotoxin, bacterial translocation to distant organs, and filamentous actin (F-actin) in the ileum were determined 24 h later. Rats treated with
Lactobacillus rhamnosus
showed reduced levels of plasma endotoxin (8 ± 2 pg/ml versus 24 ± 4 pg/ml;
P
= 0.01), bacterial translocation (2 CFU/gram versus 369 CFU/gram;
P
< 0.01), and disruption of F-actin distribution following hemorrhagic shock compared with nontreated control rats. In contrast, pretreatment with
Lactobacillus fermentum
had no substantial effect on gut barrier integrity. Interestingly, DNA preparations from both lactobacilli reduced endotoxin-induced TNF-α release dose dependently, whereas CpG-ODN increased TNF-α release. In conclusion, the pathogen exclusion properties of both
Lactobacillus
strains and the reduction of endotoxin-induced inflammation by their DNA in vitro are not prerequisites for a beneficial effect of probiotic therapy on gut barrier function following hemorrhagic shock. Although pretreatment with
Lactobacillus
spp. may be useful to preserve gut barrier integrity following severe hypotension, a thorough assessment of specific strains seems to be essential.
EnhancedPseudomonas aeruginosaBiofilm Development Mediated by Human Neutrophils
Authors: Walker Travis S. , Tomlin Kerry L. , Worthen G. Scott , Poch Katie R. , Lieber Jonathan G. , Saavedra Milene T. , Fessler Michael B. , Malcolm Kenneth C. , Vasil Michael L. , Nick Jerry A. 
Published: May 2005

ABSTRACTCystic fibrosis (CF) lung disease features persistent neutrophil accumulation to the airways from the time of infancy. CF children are frequently exposed toPseudomonas aeruginosa, and by adulthood, 80% of CF patients are chronically infected. The formation of biofilms is a particularly important phenotypic characteristic ofP. aeruginosathat allows for bacterial survival despite aggressive antibiotic therapy and an exuberant immune response. Here, we show that the presence of neutrophils enhances initialP. aeruginosabiofilm development over a period of 72 h through the formation of polymers comprised of actin and DNA. F-actin was found to be a site of attachment forP. aeruginosa. These actin and DNA polymers are present in CF sputum, and disruption of the polymers dispersed the associatedP. aeruginosacells and reduced biofilm development. These findings demonstrate a potential maladaptation of the primary innate response. When the host fails to eradicate the infection, cellular components from necrotic neutrophils can serve as a biological matrix to facilitateP. aeruginosabiofilm formation.
Opsonized Virulent
Brucella abortus
Replicates within Nonacidic, Endoplasmic Reticulum-Negative, LAMP-1-Positive Phagosomes in Human Monocytes
Authors: Bellaire Bryan H , Roop R. Martin , Cardelli James A. 
Published: May 2005

ABSTRACT

Cells in the
Brucella
spp. are intracellular pathogens that survive and replicate within host monocytes.
Brucella
maintains persistent infections in animals despite the production of high levels of anti-
Brucella
-specific antibodies. To determine the effect of antibody opsonization on the ability of
Brucella
to establish itself within monocytes, the intracellular trafficking of virulent
Brucella abortus
2308 and attenuated
hfq
and
bacA
mutants was followed in the human monocytic cell line THP-1. Early trafficking events of
B. abortus
2308-containing phagosomes (BCP) were indistinguishable from those seen for control particles (heat-killed
B. abortus
2308, live
Escherichia coli
HB101, or latex beads). All phagosomes transiently communicated the early-endosomal compartment and rapidly matured into LAMP-1
+
, cathepsin D
+
, and acidic phagosomes. By 2 h postinfection, however, the number of cathepsin D
+
BCP was significantly lower for live
B. abortus
2308-infected cells than for either
Brucella
mutant strains or control particles.
B. abortus
2308 persisted within these cathepsin D
−
, LAMP-1
+
, and acidic vesicles; however, at the onset of intracellular replication, the numbers of acidic
B. abortus
2308 BCP decreased while remaining cathepsin D
−
and LAMP-1
+
. In contrast to
B. abortus
2308, the isogenic
hfq
and
bacA
mutants remained in acidic, LAMP-1
+
phagosomes and failed to initiate intracellular replication. Notably, markers specific for the host endoplasmic reticulum were absent from the BCPs throughout the course of the infection. Thus, opsonized
B. abortus
in human monocytes survives within phagosomes that remain in the endosomal pathway and replication of virulent
B. abortus
2308 within these vesicles corresponds with an increase in intraphagosomal pH.
Expression of Genes Encoding Innate Host Defense Molecules in Normal Human Monocytes in Response toCandida albicans
Authors: Kim Hee Sup , Choi Eun Hwa , Khan Javed , Roilides Emmanuel , Francesconi Andrea , Kasai Miki , Sein Tin , Schaufele Robert L. , Sakurai Kenichi , Son Chang Gue , Greer Braden T. , Chanock Stephen , Lyman Caron A. , Walsh Thomas J. 
Published: May 2005

ABSTRACTLittle is known about the regulation and coordinated expression of genes involved in the innate host response toCandida albicans. We therefore examined the kinetic profile of gene expression of innate host defense molecules in normal human monocytes infected withC. albicansusing microarray technology. Freshly isolated peripheral blood monocytes from five healthy donors were incubated withC. albicansfor 0 to 18 h in parallel with time-matched uninfected control cells. RNA from monocytes was extracted and amplified for microarray analysis, using a 42,421-gene cDNA chip. Expression of genes encoding proinflammatory cytokines, including tumor necrosis factor alpha, interleukin 1 (IL-1), IL-6, and leukemia inhibitory factor, was markedly enhanced during the first 6 h and coincided with an increase in phagocytosis. Expression of these genes returned to near baseline by 18 h. Genes encoding chemokines, including IL-8; macrophage inflammatory proteins 1, 3, and 4; and monocyte chemoattractant protein 1, also were strongly up-regulated, with peak expression at 4 to 6 h, as were genes encoding chemokine receptors CCR1, CCR5, CCR7, and CXCR5. Expression of genes whose products may protect monocyte viability, such as BCL2-related protein, metallothioneins, CD71, and SOCS3, was up-regulated at 4 to 6 h and remained elevated throughout the 18-h time course. On the other hand, expression of genes encoding T-cell-regulatory molecules (e.g., IL-12, gamma interferon, and transforming growth factor β) was not significantly affected during the 18-h incubation. Moreover, genes encoding IL-15, the IL-13 receptor (IL-13Ra1), and CD14 were suppressed during the 18-h exposure toC. albicans. Thus,C. albicansis a potent inducer of a dynamic cascade of expression of genes whose products are related to the recruitment, activation, and protection of neutrophils and monocytes.