bluuebunny/crossref_metadata_2025 huggingface dataset:
Iha from an
Escherichia coli
Urinary Tract Infection Outbreak Clonal Group A Strain Is Expressed In Vivo in the Mouse Urinary Tract and Functions as a Catecholate Siderophore Receptor
Authors: Léveillé Simon , Caza Mélissa , Johnson James R. , Clabots Connie , Sabri Mourad , Dozois Charles M.
Published: May 2006

Virulence factors of pathogenic
Escherichia coli
belonging to a recently emerged and disseminated clonal group associated with urinary tract infection (UTI), provisionally designated clonal group A (CGA), have not been experimentally investigated. We used a mouse model of ascending UTI with CGA member strain UCB34 in order to identify genes of CGA that contribute to UTI.
iha
was identified to be expressed by strain UCB34 in the mouse kidney using selective capture of transcribed sequences.
iha
from strain UCB34 demonstrated a siderophore receptor phenotype when cloned in a catecholate siderophore receptor-negative
E. coli
K-12 strain, as shown by growth promotion experiments and uptake of
55
Fe complexed to enterobactin or its linear 2, 3-dihydroxybenzoylserine (DHBS) siderophore derivatives. Siderophore-mediated growth promotion by Iha was TonB dependent. Growth and iron uptake were more marked with linear DHBS derivatives than with purified enterobactin. The reported phenotype of adherence to epithelial cells conferred by expressing
iha
from a multicopy cloning vector in a poorly adherent
E. coli
K-12 host strain was confirmed to be specific to
iha
, in comparison with other siderophore receptor genes.
iha
expression was regulated by the ferric uptake regulator Fur and by iron availability, as shown by real-time reverse transcriptase PCR. In a competitive infection experiment using the mouse UTI model, wild-type strain UCB34 significantly outcompeted an isogenic
iha
null mutant. Iha thus represents a Fur-regulated catecholate siderophore receptor that, uniquely, exhibits an adherence-enhancing phenotype and is the first described urovirulence factor identified in a CGA strain.
Constitutive Differences in Gene Expression Profiles Parallel Genetic Patterns of Susceptibility to Tuberculosis in Mice
Authors: Orlova Marianna O. , Majorov Konstantin B. , Lyadova Irina V. , Eruslanov Eugenii B. , M'lan Cyr E. , Greenwood Celia M. T. , Schurr Erwin , Apt Alexander S.
Published: May 2006

Interstitial lung macrophages from tuberculosis-susceptible I/St and tuberculosis-resistant A/Sn mice demonstrated significant constitutive differences in gene expression levels, whereas in vitro infection of these cells withMycobacterium tuberculosishad only a modulatory impact on gene expression. We conclude that intrinsic gene expression profiles are an important determinant of tuberculosis pathogenesis in mice.
Environmental Signals Generate a Differential and Coordinated Expression of the Heme Receptor Gene Family of
Bartonella quintana
Authors: Battisti James M. , Sappington Kate N. , Smitherman Laura S. , Parrow Nermi L. , Minnick Michael F.
Published: May 2006

Of all bacteria,
Bartonella quintana
has the highest reported in vitro hemin requirement, yet an explanation for this remains elusive. To produce diseases such as trench fever, endocarditis, and bacillary angiomatosis,
B. quintana
must survive and replicate in the disparate environments of the
Pediculus humanus corporis
(body louse) gut and the human vasculature. We previously identified a five-member family of hemin binding proteins (Hbps) synthesized by
B. quintana
that bind hemin on the outer surface but share no similarity to known bacterial heme receptors. In the present study, we examine the transcription, regulation, and synthesis of this virulence factor family by cultivation of the bacterium in environments that simulate natural heme, oxygen, and temperature conditions encountered in the host and insect vector. First, quantitative real-time PCR data show that
hbpC
expression is regulated by temperature, where a >100-fold increase in transcript quantity was seen at 30°C relative to 37°C, suggesting that HbpC synthesis would be greatest in the cooler temperature of the louse. Second, cultivation at human bloodstream oxygen concentration (5% relative to 21% atmospheric) significantly decreases the transcript quantity of all
hbp
genes, indicating that expression is influenced by O
2
and/or reactive oxygen species. Third, a differential expression pattern within the
hbp
family is revealed when
B. quintana
is grown in a range of hemin concentrations: subgroup I (
hbpC
and
hbpB
) predominates in a simulated louse environment (high heme), and subgroup II (
hbpA
,
hbpD
, and
hbpE
) is preferentially expressed in a simulated human background (low heme). By using two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblotting, and matrix-assisted laser desorption ionization—time of flight mass spectrometry fingerprinting, we demonstrate that synthesis of HbpA correlates with
hbpA
transcript increases observed at low hemin concentrations. Finally, an
hbpA
promoter-
lacZ
reporter construct in
B. quintana
demonstrates that a transcriptional regulator(s) is controlling the expression of
hbpA
through a
cis
-acting regulatory element located in the
hbpA
promoter region.
Characterization of a Novel High-Affinity Monoclonal Immunoglobulin G Antibody against the Ricin B Subunit
Authors: McGuinness Carolyn R. , Mantis Nicholas J.
Published: May 2006

There is an urgent need for the development of a passive immunotherapy against the category B select agent ricin, a lethal ribosome-inactivating toxin composed of an enzymatic A subunit (RTA) and a single binding B subunit (RTB). To date, only one monoclonal antibody (MAb), a mouse immunoglobulin G (IgG1) against RTA called R70, has been deemed sufficiently potent in animal models to warrant further testing in humans. In this study, we have identified and characterized MAb 24B11, a murine IgG1 directed against RTB. In a Vero cell cytotoxicity assay, 24B11 was approximately two times more effective at neutralizing ricin than was R70. The equilibrium dissociation constants of 24B11 (KD= 4.2 × 10−9M) and R70 (KD= 3.2 × 10−9M) were virtually identical, suggesting that the difference in neutralization activity between the two MAbs was not due to differing affinities for the toxin. 24B11 blocked ricin attachment to galactoside receptors on primary mouse splenocytes and on the apical surfaces of human mucosal epithelial cell monolayers. Surprisingly, R70 also effectively interfered with ricin attachment to receptors on cell surfaces. Using a phage-displayed peptide library, we determined that 24B11 binds an epitope on RTB adjacent to, but not within, one of the two galactose binding domains. Finally, we demonstrate that R70 and 24B11, when combined, function synergistically to neutralize ricin in vitro, raising the possibility that these two MAbs could serve as a novel immunotherapeutic in vivo.
Human Recombinant Antibodies againstPlasmodium falciparumMerozoite Surface Protein 3 Cloned from Peripheral Blood Leukocytes of Individuals with Immunity to Malaria Demonstrate Antiparasitic Properties
Authors: Lundquist Rasmus , Nielsen Leif Kofoed , Jafarshad Ali , SoeSoe Daw , Christensen Lars Harder , Druilhe Pierre , Dziegiel Morten Hanefeld
Published: May 2006

Immunoglobulins from individuals with immunity to malaria have a strong antiparasitic effect when transferred toPlasmodium falciparummalaria infected patients. One prominent target of antiparasitic antibodies is the merozoite surface antigen 3 (MSP-3). We have investigated the antibody response against MSP-3 residues 194 to 257 (MSP-3194-257) on the molecular level. mRNA from peripheral blood leukocytes from clinically immune individuals was used as a source of Fab (fragment antibody) genes. A Fab-phage display library was made, and three distinct antibodies designated RAM1, RAM2, and RAM3 were isolated by panning. Immunoglobulin G1 (IgG1) and IgG3 full-length antibodies have been produced in CHO cells. Reactivity with the native parasite protein was demonstrated by immunofluorescence microscopy, flow cytometry, and immunoblotting. Furthermore, the antiparasitic effect of RAM1 has been tested in vitro in an antibody-dependent cellular inhibition (ADCI) assay. Both the IgG1 and the IgG3 versions of the antibody show an inhibitory effect on parasite growth.
Differential Localization of Complement Component 3 within the Capsular Matrix ofCryptococcus neoformans
Authors: Gates Marcellene A. , Kozel Thomas R.
Published: May 2006

The polysaccharide capsule ofCryptococcus neoformansis a powerful activator of the complement system. The goal of the present study was to assess serum and cellular variables that influence the sites for C3 binding within the capsular matrix. Confocal microscopy using fluorophore-labeled polyclonal anti-C3 and anticapsular monoclonal antibodies and rosetting of fluorescent microspheres coated with anti-C3 were used to identify sites of C3 binding relative to the capsular edge. The results showed that the source of serum was a major variable influencing localization of C3. C3 bound at or very near the capsular edge in the case of human serum. C3 deposition was further from the capsule edge with guinea pig and rat sera; in the case of mouse serum, there was no binding of C3 in the outer region of the capsule. Addition of human C3 to mouse serum led to deposition of the C3 at the capsular edge, indicating that distinct properties of mouse and human C3 account for the differential localization of C3. Finally, the density of the capsular matrix was an important variable in determining sites for C3 deposition. Yeast cells with a high concentration of polysaccharide near the capsule edge supported deposition of mouse C3 at or near the capsular edge, whereas cells with a low matrix density showed deposition well beneath the edge. Taken together, these results indicate that the spatial deposition of C3 within the capsular matrix is a complex process that is influenced by the serum source and the density of the capsular matrix.
Genetic Variation among Isolates of
Sarcocystis neurona
, the Agent of Protozoal Myeloencephalitis, as Revealed by Amplified Fragment Length Polymorphism Markers
Authors: Elsheikha H. M. , Schott H. C. , Mansfield L. S.
Published: May 2006

Sarcocystis neurona
causes serious neurological disease in horses and other vertebrates in the Americas. Based on epidemiological data, this parasite has recently emerged. Here, the genetic diversity of
Sarcocystis neurona
was evaluated using the amplified fragment length polymorphism (AFLP) method. Fifteen
S. neurona
taxa from different regions collected over the last 10 years were used; six isolates were from clinically diseased horses, eight isolates were from wild-caught opossums (
Didelphis virginiana
), and one isolate was from a cowbird (
Molothrus ater
). Additionally, four outgroup taxa were also fingerprinted. Nine primer pairs were used to generate AFLP patterns, with a total number of amplified fragments ranging from 30 to 60, depending on the isolate and primers tested. Based on the presence/absence of amplified AFLP fragments and pairwise similarity values, all the
S. neurona
isolates tested were clustered in one monophyletic group. No significant correlation could be found between genomic similarity and host origin of the
S. neurona
isolates. AFLP revealed significant intraspecific genetic variations, and
S. neurona
appeared as a highly variable species. Furthermore, linkage disequilibrium analysis suggested that
S. neurona
populations within Michigan have an intermediate type of population structure that includes characteristics of both clonal and panamictic population structures. AFLP is a reliable molecular technique that has provided one of the most informative approaches to ascertain phylogenetic relationships in
S. neurona
and its closest relatives, allowing them to be clustered by relative similarity using band matching and unweighted pair group method with arithmetic mean analysis, which may be applicable to other related protozoal species.
CD4+CD25+Regulatory T Cells Modulate the T-Cell and Antibody Responses inHelicobacter-Infected BALB/c Mice
Authors: Kaparakis Maria , Laurie Karen L. , Wijburg Odilia , Pedersen John , Pearse Martin , van Driel Ian R. , Gleeson Paul A. , Strugnell Richard A.
Published: May 2006

GastricHelicobacterspp. induce chronic gastritis that may lead to ulceration and dysplasia. The host elicits a T helper 1 (Th1) response that is fundamental to the pathogenesis of these bacteria. We analyzed immune responses inHelicobacter-infected, normal mice depleted of CD4+CD25+T cells to investigate the in vivo role of regulatory T cells (Tregs) in the modulation ofHelicobacterimmunopathology. BALB/c and transgenic mice were depleted of CD4+CD25+T cells by administration of an anti-CD25 antibody either at the time of infection withHelicobacteror during chronic infection and gastritis. Depletion of CD25+Tregs prior to and during infection of mice withHelicobacterspp. did not affect either bacterial colonization or severity of gastritis. Depletion of CD25+Tregs was associated with increasedHelicobacter-specific antibody levels and an altered isotype distribution. Paragastric lymph node cells from CD25+Treg-depleted and control infected mice showed similar proliferation toHelicobacterantigens, but only cells from anti-CD25-treated animals secreted Th2 cytokines. CD25+Tregs do not control the level of gastritis induced by gastricHelicobacterspp. in normal, thymus-intact BALB/c mice. However, CD25+Tregs influence the cytokine and antibody responses induced by infection. Autoimmune gastritis is not induced inHelicobacter-infected mice depleted of CD25+Tregs but is induced in CD25+Treg-depleted mice, which have a higher frequency of autoreactive T cells.
Membrane Vesicles Shed byLegionella pneumophilaInhibit Fusion of Phagosomes with Lysosomes
Authors: Fernandez-Moreira Esteban , Helbig Juergen H. , Swanson Michele S.
Published: May 2006

When cultured in broth to the transmissive phase,Legionella pneumophilainfects macrophages by inhibiting phagosome maturation, whereas replicative-phase cells are transported to the lysosomes. Here we report that the ability ofL. pneumophilato inhibit phagosome-lysosome fusion correlated with developmentally regulated modifications of the pathogen's surface, as judged by its lipopolysaccharide profile and by its binding to a sialic acid-specific lectin and to the hydrocarbon hexadecane. Likewise, the composition of membrane vesicles shed byL. pneumophilawas developmentally regulated, based on binding to the lectin and to the lipopolysaccharide-specific monoclonal antibody 3/1. Membrane vesicles were sufficient to inhibit phagosome-lysosome fusion by a mechanism independent of type IV secretion, since only ∼25% of beads suspended with or coated by vesicles from transmissive phase wild type ordotAsecretion mutants colocalized with lysosomal probes, whereas ∼75% of beads were lysosomal when untreated or presented with vesicles from theL. pneumophila letAregulatory mutant orE. coli. As observed previously forL. pneumophilainfection of mouse macrophages, vesicles inhibited phagosome-lysosome fusion only temporarily; by 10 h after treatment with vesicles, macrophages delivered ∼72% of ingested beads to lysosomes. Accordingly, in the context of the epidemiology of the pneumonia Legionnaires' disease and virulence mechanisms ofLeishmaniaandMycobacteria, we discuss a model here in whichL. pneumophiladevelopmentally regulates its surface composition and releases vesicles into phagosomes that inhibit their fusion with lysosomes.
Attenuation and Persistence of and Ability To Induce Protective Immunity to a
Staphylococcus aureus aroA
Mutant in Mice
Authors: Buzzola Fernanda R. , Barbagelata María Sol , Caccuri Roberto L. , Sordelli Daniel O.
Published: May 2006

Staphylococcus aureus
is the most important etiological agent of bovine mastitis, a disease that causes significant economic losses to the dairy industry. Several vaccines to prevent the disease have been tested, with limited success. The aim of this study was to obtain a suitable attenuated
aro
mutant of
S. aureus
by transposon mutagenesis and to demonstrate its efficacy as a live vaccine to induce protective immunity in a murine model of intramammary infection. To do this, we transformed
S. aureus
RN6390 with plasmid pTV1
ts
carrying Tn
917
. After screening of 3,493 erythromycin-resistant colonies, one mutant incapable of growing on plates lacking phenylalanine, tryptophan, and tyrosine was isolated and characterized. Molecular characterization of the mutant showed that the affected gene was
aroA
and that the insertion occurred 756 bp downstream of the
aroA
start codon. Complementation of the
aroA
mutant with a plasmid carrying
aroA
recovered the wild-type phenotype. The mutant exhibited a 50% lethal dose (1 × 10
6
CFU/mouse) higher than that of the parental strain (4.3 × 10
4
CFU/mouse). The
aroA
mutant showed decreased ability to persist in the lungs, spleens, and mammary glands of mice. Intramammary immunization with the
aroA
mutant stimulated both Th1 and Th2 responses in the mammary gland, as ascertained by reverse transcription-PCR, and induced significant protection from challenge with either the parental wild-type or a heterologous strain isolated from a cow with mastitis.
Development of Luminescent
Mycobacterium avium
subsp.
paratuberculosis
for Rapid Screening of Vaccine Candidates in Mice
Authors: Rosseels Valérie , Roupie Virginie , Zinniel Denise , Barletta Raúl G. , Huygen Kris
Published: May 2006

Mycobacterium avium
subsp.
paratuberculosis
is a slowly growing mycobacterial species, requiring 6 to 8 weeks of culture before colonies can be counted visually. Here, we describe the development of luminescent
M. avium
subsp.
paratuberculosis
expressing
luxAB
genes of
Vibrio harveyi
and its use for vaccine testing in an experimental mouse model, replacing fastidious CFU counting by rapid luminometry.
Babesia microti
Primarily Invades Mature Erythrocytes in Mice
Authors: Borggraefe Ingo , Yuan Jie , Telford Sam R. , Menon Sanjay , Hunter Rouette , Shah Sohela , Spielman Andrew , Gelfand Jeffrey A. , Wortis Henry H. , Vannier Edouard
Published: May 2006

Babesia microti
is a tick-borne red blood cell parasite that causes babesiosis in people. Its most common vertebrate reservoir is the white-footed mouse. To determine whether
B. microti
invades reticulocytes, as does the canine pathogen
B. gibsoni
, we infected the susceptible inbred mouse strains C.B-17.scid and DBA/2 with a clinical isolate of
B. microti
. Staining of fixed permeabilized red blood cells with 4′,6′-diamidino-2-phenylindole or YOYO-1, a sensitive nucleic acid stain, revealed parasite nuclei as large bright dots. Flow cytometric analysis indicated that parasite DNA is primarily found in mature erythrocytes that expressed
Babesia
antigens but not the transferrin receptor CD71. In contrast, CD71-positive reticulocytes rarely contained
Babesia
nuclei and failed to express
Babesia
antigens. Accordingly, the frequency of YOYO-1-positive, CD71-negative cells strongly correlated with parasitemia, defined as the frequency of infected red blood cells assessed on Giemsa-stained blood smears. Importantly, the absolute numbers generated by the two techniques were similar. Parasitemia was modest and transient in DBA/2 mice but intense and sustained in C.B-17.scid mice. In both strains, parasitemia preceded reticulocytosis, but reticulocytes remained refractory to
B. microti
. In immunocompetent C.B-17 mice, reticulocytosis developed early, despite a marginal and short-lived parasitemia. Likewise, an early reticulocytosis developed in resistant BALB/cBy and B10.D2 mice. These studies establish that
B. microti
has a tropism for mature erythrocytes. Although reticulocytes are rarely infected, the delayed reticulocytosis in susceptible strains may result from parasite or host activities to limit renewal of the mature erythrocyte pool, thereby preventing an overwhelming parasitemia.
Plasmodium falciparum-Infected Erythrocytes Increase Intercellular Adhesion Molecule 1 Expression on Brain Endothelium through NF-κB
Authors: Tripathi Abhai K. , Sullivan David J. , Stins Monique F.
Published: May 2006

Sequestration ofPlasmodium falciparum-infected erythrocytes (Pf-IRBC) in postcapillary brain endothelium is a hallmark of cerebral malaria (CM) pathogenesis. There is a correlation between adherent Pf-IRBC and increased expression of intercellular cell adhesion molecule 1 (ICAM-1), which is also a receptor for Pf-IRBC on human brain microvascular endothelial cells (HBMEC). The underlying mechanism for the increased ICAM-1 expression has not been clearly defined. Therefore, we investigated the mechanisms of ICAM-1 expression on isolated HBMEC after exposure to Pf-IRBC. Ultrastructural characterization of the model confirmed that there was attachment through both Pf-IRBC knobs and HBMEC microvillus protrusions. Pf-IRBC induced a dose- and time-dependent increase in ICAM-1 expression on HBMEC that was specific for human brain endothelium and was not observed with human umbilical vein endothelium. Involvement of both membrane-associated Pf-IRBC proteins and parasite-derived soluble factors with the increase in ICAM-1 expression was demonstrated by surface trypsinization and fractionation. Pf-IRBC exposure induced nuclear translocation of NF-κB in HBMEC, which was linked to ICAM-1 expression, as shown by use of specific inhibitors of the transcription factor NF-κB and immunocytochemistry. In addition, inhibition of reactive oxygen species decreased Pf-IRBC-induced ICAM-1 expression on HBMEC. Parasite-induced ICAM-1 expression explains the localization of this molecule on brain endothelium in postmortem CM brain samples. By increasing ICAM-1 expression, Pf-IRBC may increase their sequestration, thereby perpetuating CM.
Organ-Specific Role of MyD88 for Gene Regulation during Polymicrobial Peritonitis
Authors: Weighardt Heike , Mages Jörg , Jusek Gabriela , Kaiser-Moore Simone , Lang Roland , Holzmann Bernhard
Published: May 2006

Sepsis leads to the rapid induction of proinflammatory signaling cascades by activation of the innate immune system through Toll-like receptors (TLR). To characterize the role of TLR signaling through MyD88 for sepsis-induced transcriptional activation, we investigated gene expression during polymicrobial septic peritonitis by microarray analysis. Comparison of gene expression profiles for spleens and livers from septic wild-type and MyD88-deficient mice revealed striking organ-specific differences. Whereas MyD88 deficiency strongly reduced sepsis-induced gene expression in the liver, gene expression in the spleen was largely independent of MyD88, indicating organ-specific transcriptional regulation during polymicrobial sepsis. In addition to genes regulated by MyD88 in an organ-dependent manner, we also identified genes that exhibited an organ-independent influence of MyD88 and mostly encoded cytokines and chemokines. Notably, the expression of interferon (IFN)-regulated genes was markedly increased in septic MyD88-deficient mice compared to that in septic wild-type controls. Expression of IFN-regulated genes was dependent on the adapter protein TRIF. These results suggest that the influence of MyD88 on gene expression during sepsis strongly depends on the organ compartment affected by inflammation and that the lack of MyD88 may lead to disbalance of the expression of IFN-regulated genes.
Immunological and Molecular Characterization of Susceptibility in Relationship to Bacterial Strain Differences inMycobacterium aviumsubsp.paratuberculosisInfection in the Red Deer (Cervus elaphus)
Authors: O'Brien R. , Mackintosh C. G. , Bakker D. , Kopecna M. , Pavlik I. , Griffin J. F. T.
Published: May 2006

Johne's disease (JD) infection, caused byMycobacterium aviumsubsp.paratuberculosis, represents a major disease problem in farmed ruminants. Although JD has been well characterized in cattle and sheep, little is known of the infection dynamics or immunological response in deer. In this study, typing ofM. aviumsubsp.paratuberculosisisolates from intestinal lymphatic tissues from 74 JD-infected animals showed that clinical isolates ofM. aviumsubsp.paratuberculosisfrom New Zealand farmed red deer were exclusively of the bovine strain genotype. The susceptibility of deer toM. aviumsubsp.paratuberculosiswas further investigated by experimental oral-route infection studies using defined isolates of virulent bovine and ovineM. aviumsubsp.paratuberculosisstrains. Oral inoculation with high (109CFU/animal) or medium (107CFU/animal) doses of the bovine strain ofM. aviumsubsp.paratuberculosisestablished 100% infection rates, compared to 69% infection following inoculation with a medium dose of the ovine strain. The high susceptibility of deer to the bovine strain ofM. aviumsubsp.paratuberculosiswas confirmed by a 50% infection rate following experimental inoculation with a low dose of bacteria (103CFU/animal). This study is the first to report experimentalM. aviumsubsp.paratuberculosisinfection in red deer, and it outlines the strong infectivity of bovine-strainM. aviumsubsp.paratuberculosisisolates for cervines.
Staphylococcus aureus
Enterotoxin D Is Secreted in Milk and Stimulates Specific Antibody Responses in Cows in the Course of Experimental Intramammary Infection
Authors: Tollersrud Tore , Kampen Annette H. , Kenny Kevin
Published: May 2006

An enterotoxin D (SED)-producing strain of
Staphylococcus aureus
was used to infect one mammary gland of each of 17 lactating dairy cows. All glands became infected and shed bacteria over a sampling period of 3 weeks. Serum and milk antibodies specific for SED were monitored by an enzyme-linked immunosorbent assay for 12 weeks. Elevated anti-SED antibodies were detected in all cows after infection, and immunoglobulin of the G2 subclass comprised most of the specific serum response. SED was detected in mastitic milk samples from two cows at levels of 5 to 10 ng/ml. An in vitro lymphocyte proliferation assay showed that SED at levels below 10 pg/ml induced proliferation of bovine lymphocytes and that sheep antiserum specific for SED neutralized this proliferative response. Sera obtained from the cows pre- and postinfection inhibited lymphocyte proliferation at SED concentrations of 10 and 50 ng/ml, respectively. The addition of SED to whole blood or to isolated neutrophils had no significant effect on neutrophil function in vitro. The results show that SED is secreted during mammary gland infection, is mitogenic for bovine lymphocytes, and stimulates the production of specific antibodies.
Isolation of a Human Monoclonal Antibody with Strong Neutralizing Activity against Diphtheria Toxin
Authors: Kakita Mai , Takahashi Tsuyoshi , Komiya Takako , Iba Yoshitaka , Tsuji Takao , Kurosawa Yoshikazu , Takahashi Motohide
Published: May 2006

We isolated a human monoclonal antibody against diphtheria toxin (DT). It bound to fragment B with a binding activity (

K
d

) of 3.01 nM. The neutralizing activity assayed by the rabbit skin test was estimated to be 73,600 IU/g. This could be used as a therapeutic drug against DT in place of the traditional equine sera.
Flagellin-Independent Regulation of Chemokine Host Defense inCampylobacter jejuni-Infected Intestinal Epithelium
Authors: Johanesen Priscilla A. , Dwinell Michael B.
Published: May 2006

Campylobacter jejuniis a leading cause of bacterial food-borne diarrheal disease throughout the world and the most frequent antecedent of autoimmune neuropathy Guillain-Barré syndrome. While infection is associated with immune memory, little is known regarding the role of the epithelium in targeting dendritic cells (DC) for initiating the appropriate adaptive immune response toC. jejuni. The objective of this study was to define the role for the intestinal epithelium in the induction of the adaptive immune response inC. jejuniinfection by assessing the production of DC and T-cell chemoattractants. Human T84 epithelial cells were used as model intestinal epithelia. Infection of T84 cells withC. jejunidose- and time-dependently up-regulated DC and T-cell chemokine gene transcription and secretion. Induction required live bacteria and was in the physiologically relevant direction for attraction of mucosal immunocytes.C. jejuni-activated NF-κB signaling was shown to be essential for proinflammatory chemokine secretion. Notably,C. jejunisecretion occurred independently of flagellin identification by Toll-like receptor 5. Secretion of a DC chemoattractant by differing clinicalC. jejuniisolates suggested adherence/invasion were key virulence determinants of epithelial chemokine secretion. The regulated epithelial expression of DC and T-cell chemoattractants suggests a mechanism for the directed trafficking of immune cells required for the initiation of adaptive immunity in campylobacteriosis. Chemokine secretion occurs despiteCampylobacterevasion of the flagellin pattern recognition receptor, suggesting that alternate host defense strategies limit disease pathogenesis.
Differential Expression and Sequence Conservation of theAnaplasma marginale msp2Gene Superfamily Outer Membrane Proteins
Authors: Noh Susan M. , Brayton Kelly A. , Knowles Donald P. , Agnes Joseph T. , Dark Michael J. , Brown Wendy C. , Baszler Timothy V. , Palmer Guy H.
Published: May 2006

Bacterial pathogens in the generaAnaplasmaandEhrlichiaencode a protein superfamily, pfam01617, which includes the predominant outer membrane proteins (OMPs) of each species, major surface protein 2 (MSP2) and MSP3 ofAnaplasma marginaleandAnaplasma ovis,Anaplasma phagocytophilumMSP2 (p44),Ehrlichia chaffeensisp28-OMP,Ehrlichia canisp30, andEhrlichia ruminantiumMAP1, and has been shown to be involved in both antigenic variation within the mammalian host and differential expression between the mammalian and arthropod hosts. Recently, complete sequencing of theA. marginalegenome has identified an expanded set of genes, designatedomp1-14, encoding new members of this superfamily. Transcriptional analysis indicated that, with the exception of the three smallest open reading frames,omp2,omp3, andomp6, these superfamily genes are transcribed inA. marginale-infected erythrocytes, tick midgut and salivary glands, and the IDE8 tick cell line. OMPs 1, 4, 7 to 9, and 11 were confirmed to be expressed as proteins byA. marginalewithin infected erythrocytes, with expression being either markedly lower (OMPs 1, 4, and 7 to 9) or absent (OMP11) in infected tick cells, which reflected regulation at the transcript level. Although the pfam01617 superfamily includes the antigenically variable MSP2 and MSP3 surface proteins, analysis of theomp1-14sequences throughout a cycle of acute and persistent infection in the mammalian host and tick transmission reveals a high degree of conservation, an observation supported by sequence comparisons between the St. Maries strain and Florida strain genomes.
The Major Subunit, CfaB, of Colonization Factor Antigen I from Enterotoxigenic
Escherichia coli
Is a Glycosphingolipid Binding Protein
Authors: Jansson Lena , Tobias Joshua , Lebens Michael , Svennerholm Ann-Mari , Teneberg Susann
Published: May 2006

Bacterial adherence to mucosal surfaces is an important virulence trait of pathogenic bacteria. Adhesion of enterotoxigenic
Escherichia coli
(ETEC) to the intestine is mediated by a number of antigenically distinct colonization factors (CFs). One of the most common CFs is CFA/I. This has a fimbrial structure composed of a major repeating subunit, CfaB, and a single tip subunit, CfaE. The potential carbohydrate recognition by CFA/I was investigated by binding CFA/I-fimbriated bacteria and purified CFA/I fimbriae to a large number of variant glycosphingolipids separated on thin-layer chromatograms. For both fimbriated bacteria and purified fimbriae, specific interactions could be identified with a number of nonacid glycosphingolipids. These included glucosylceramide, lactosylceramide with phytosphingosine and/or hydroxy fatty acids, neolactotetraosylceramide, gangliotriaosylceramide, gangliotetraosylceramide, the H5 type 2 pentaglycosylceramide, the Le
a
-5 glycosphingolipid, the Le
x
-5 glycosphingolipid, and the Le
y
-6 glycosphingolipid. These glycosphingolipids were also recognized by recombinant
E. coli
expressing CFA/I in the absence of tip protein CfaE, as well as by purified fimbriae from the same strain. This demonstrates that the glycosphingolipid-binding capacity of CFA/I resides in the major CfaB subunit.
Localization of the C3-Like ADP-Ribosyltransferase from
Staphylococcus aureus
during Bacterial Invasion of Mammalian Cells
Authors: Molinari Gabriella , Rohde Manfred , Wilde Christian , Just Ingo , Aktories Klaus , Chhatwal Gursharan S.
Published: May 2006

The C3stau2 exoenzyme from
Staphylococcus aureus
is a C3-like ADP-ribosyltransferase which possesses no specific receptor-binding domain or translocation unit required for entry in target cells where its substrate is located. Here we show that C3stau2 can reach its target after invasion of staphylococci in eukaryotic cells without needing translocation.
Toll-Like Receptor 2 Is Required for Control of Pulmonary Infection withFrancisella tularensis
Authors: Malik Meenakshi , Bakshi Chandra Shekhar , Sahay Bikash , Shah Aaloki , Lotz Steven A. , Sellati Timothy J.
Published: May 2006

Toll-like receptor 2 (TLR2) deficiency enhances murine susceptibility to infection byFrancisella tularensisas indicated by accelerated mortality, higher bacterial burden, and greater histopathology. Analysis of pulmonary cytokine levels revealed that TLR2 deficiency results in significantly lower levels of tumor necrosis factor alpha and interleukin-6 but increased amounts of gamma interferon and monocyte chemoattractant protein 1. This pattern of cytokine production may contribute to the exaggerated pathogenesis seen in TLR2−/−mice. Collectively, these findings suggest that TLR2 plays an important role in tempering the host response to pneumonic tularemia.
MyD88-Dependent Responses Involving Toll-Like Receptor 2 Are Important for Protection and Clearance ofLegionella pneumophilain a Mouse Model of Legionnaires' Disease
Authors: Archer Kristina A. , Roy Craig R.
Published: May 2006

Legionella pneumophilais a gram-negative facultative intracellular parasite of macrophages. AlthoughL. pneumophilais the causative agent of a severe pneumonia known as Legionnaires' disease, it is likely that most infections caused by this organism are cleared by the host innate immune system. It is predicted that host pattern recognition proteins belonging to the Toll-like receptor (TLR) family are involved in the protective innate immune responses. We examined the role of TLR-mediated responses inL. pneumophiladetection and clearance using genetically altered mouse hosts in which the macrophages are permissive forL. pneumophilaintracellular replication. Our data demonstrate that cytokine production by bone marrow-derived macrophages (BMMs) in response toL. pneumophilainfection requires the TLR adapter protein MyD88 and is reduced in the absence of TLR2 but not in the absence of TLR4. Bacterial growth ex vivo in BMMs from MyD88-deficient mice was not enhanced compared to bacterial growth ex vivo in BMMs from heterozygous littermate controls. Wild-type mice were able to clearL. pneumophilafrom the lung, whereas respiratory infection of MyD88-deficient mice caused death that resulted from robust bacterial replication and dissemination. In contrast to an infection with virulentL. pneumophila, MyD88-deficient mice were able to clear infections withL. pneumophila dotAmutants, indicating that MyD88-independent responses in the lung are sufficient to clear bacteria that are unable to replicate intracellularly. In vivo growth ofL. pneumophilawas enhanced in the lungs of TLR2-deficient mice, which resulted in a delay in bacterial clearance. No significant differences were observed in the growth and clearance ofL. pneumophilain the lungs of TLR4-deficient mice and heterozygous littermate control mice. Our data indicate that MyD88 is crucial for eliciting a protective innate immune response against virulentL. pneumophilaand that TLR2 is one of the pattern recognition receptors involved in initiating this MyD88-dependent response.
Mycobacterium marinum
Erp Is a Virulence Determinant Required for Cell Wall Integrity and Intracellular Survival
Authors: Cosma Christine L. , Klein Kathryn , Kim Rosa , Beery Dana , Ramakrishnan Lalita
Published: May 2006

The
Mycobacterium tuberculosis
exported repetitive protein (Erp) is a virulence determinant required for growth in cultured macrophages and in vivo. To better understand the role of Erp in
Mycobacterium
pathogenesis, we generated a mutation in the
erp
homologue of
Mycobacterium marinum
, a close genetic relative of
M. tuberculosis. erp
-deficient
M. marinum
was growth attenuated in cultured macrophage monolayers and during chronic granulomatous infection of leopard frogs, suggesting that Erp function is similarly required for the virulence of both
M. tuberculosis
and
M. marinum
. To pinpoint the step in infection at which Erp is required, we utilized a zebrafish embryo infection model that allows
M. marinum
infections to be visualized in real-time, comparing the
erp
-deficient strain to a ΔRD1 mutant whose stage of attenuation was previously characterized in zebrafish embryos. A detailed microscopic examination of infected embryos revealed that bacteria lacking Erp were compromised very early in infection, failing to grow and/or survive upon phagocytosis by host macrophages. In contrast, ΔRD1 mutant bacteria grow normally in macrophages but fail to induce host macrophage aggregation and subsequent cell-to-cell spread. Consistent with these in vivo findings,
erp
-deficient but not RD1-deficient bacteria exhibited permeability defects in vitro, which may be responsible for their specific failure to survive in host macrophages.
High Frequency of CD4+T Cells Specific for the TB10.4 Protein Correlates with Protection againstMycobacterium tuberculosisInfection
Authors: Hervas-Stubbs Sandra , Majlessi Laleh , Simsova Marcela , Morova Jana , Rojas Marie-Jesus , Nouzé Clémence , Brodin Priscille , Sebo Peter , Leclerc Claude
Published: May 2006

TB10.4 is a newly identified antigen ofMycobacterium tuberculosisrecognized by human and murine T cells upon mycobacterial infection. Here, we show that immunization withMycobacterium bovisBCG induces a strong, genetically controlled, Th1 immune response against TB10.4 in mice. BALB/c and C57BL/6 strains behave as high and low responders to TB10.4 protein, respectively. The TB10.4:74-88 peptide was identified as an immunodominant CD4+T-cell epitope forH-2dmice. Since recent results, as well as the present study, have raised interest in TB10.4 as a subunit vaccine, we analyzed immune responses induced by this antigen delivered by a new vector, the adenylate cyclase (CyaA) ofBordetella pertussis. CyaA is able to target dendritic cells and to deliver CD4+or CD8+T-cell epitopes to the major histocompatibility complex class II/I molecule presentation pathways, triggering specific Th1 or cytotoxic T-lymphocyte (CTL) responses. Several CyaA harboring either the entire TB10.4 protein or various subfragments containing the TB10.4:20-28 CTL epitope were shown to induce TB10.4-specific Th1 CD4+and CD8+T-cell responses. However, none of the recombinant CyaA, injected in the absence of adjuvant, was able to induce protection againstM. tuberculosisinfection. In contrast, TB10.4 protein administered with a cocktail of strong adjuvants that triggered a strong Th1 CD4+T-cell response induced significant protection againstM. tuberculosischallenge. These results confirm the potential value of the TB10.4 protein as a candidate vaccine and show that the presence of high frequencies of CD4+T cells specific to this strong immunogen correlates with protection againstM. tuberculosisinfection.
Elevated Plasma Phenylalanine in Severe Malaria and Implications for Pathophysiology of Neurological Complications
Authors: Lopansri Bert K. , Anstey Nicholas M. , Stoddard Gregory J. , Mwaikambo Esther D. , Boutlis Craig S. , Tjitra Emiliana , Maniboey Helena , Hobbs Maurine R. , Levesque Marc C. , Weinberg J. Brice , Granger Donald L.
Published: May 2006

Cerebral malaria is associated with decreased production of nitric oxide and decreased levels of its precursor,l-arginine. Abnormal amino acid metabolism may thus be an important factor in malaria pathogenesis. We sought to determine if other amino acid abnormalities are associated with disease severity in falciparum malaria. Subjects were enrolled in Dar es Salaam, Tanzania (children) (n= 126), and Papua, Indonesia (adults) (n= 156), in two separate studies. Plasma samples were collected from subjects with WHO-defined cerebral malaria (children), all forms of severe malaria (adults), and uncomplicated malaria (children and adults). Healthy children and adults without fever or illness served as controls. Plasma amino acids were measured using reverse-phase high-performance liquid chromatography with fluorescence detection. Several plasma amino acids were significantly lower in the clinical malaria groups than in healthy controls. Despite the differences, phenylalanine was the only amino acid with mean levels outside the normal range (40 to 84 μM) and was markedly elevated in children with cerebral malaria (median [95% confidence interval], 163 [134 to 193] μM;P< 0.0001) and adults with all forms of severe malaria (median [95% confidence interval], 129 [111 to 155] μM;P< 0.0001). In adults who survived severe malaria, phenylalanine levels returned to normal, with clinical improvement (P= 0.0002). Maintenance of plasma phenylalanine homeostasis is disrupted in severe malaria, leading to significant hyperphenylalaninemia. This is likely a result of an acquired abnormality in the function of the liver enzyme phenylalanine hydroxylase. Determination of the mechanism of this abnormality may contribute to the understanding of neurological complications in malaria.
Atomic Iodine Generation via Fluorine Atoms for Chemical Oxygen-Iodine Laser
Authors: Čenský Miroslav , Špalek Otomar , Jirásek Vít , Kodymová Jarmila , Jakubec Ivo
Published: May 2006

Chemical generation of atomic iodine for a chemical oxygen-iodine laser (COIL) was investigated experimentally. In the two-step reaction mechanism, molecular fluorine reacts with nitrogen oxide and formed fluorine atoms react then with hydrogen iodide to iodine atoms. The efficiency of this process was studied in dependence on mixing conditions, flow rate of reacting gases and pressure in reactor. A maximum concentration of atomic iodine was obtained at approximately equimolar ratio of reacting gases (F2, NO and HI), which agrees well with the stoichiometry of production reactions. A shortage of any reacting gases limits the rate of atomic iodine formation. An excess of F2 relative to NO at a simultaneous deficiency of HI had a most detrimental effect on atomic iodine production. High concentrations of atomic iodine (5-8 × 1015 cm-3) were achieved by this method at pressures 4-9 kPa, which are sufficient for a COIL operation. This makes it possible to use the above method as a source of iodine atoms and their injection into the primary flow of singlet oxygen in COIL.
Copolymer Micelles with Polyelectrolyte Shell in Aqueous Media. A Mean-Field Study
Authors: Jelínek Karel , Uhlík Filip , Limpouchová Zuzana , Matějíček Pavel , Procházka Karel
Published: May 2006

The multimolecular micelles formed by polystyrene-block-poly(methacrylic acid) (PS-PMA) copolymer and by hydrophobically modified PS-PMA copolymer with naphthalene and anthracene (PS-N-PMA-A) were studied by self-consistent field (SCF) calculations in aqueous media. The labeling with covalently bonded naphthalene between PS and PMA blocks and with anthracene at the free end of PMA blocks, which is suitable for experimental nonradiative excitation energy transfer (NRET) studies of PS-N-PMA-A micelles, modifies the structure of micellar shell. The study was aimed at understanding structure and behavior of micelles at different pH and ionic strength. The results show that the presence of hydrophobic tags has only a small influence on the overall structure of micelles but it strongly affects the distribution of PMA free ends. The hydrophobic labels (anthracenes) try to return into the shell and their certain fraction is localized close to the core/shell interface, which causes a fairly high NRET efficiency. The calculated and experimentally measured NRET efficiency were compared; their trends are reasonable at the semiquantitative level.
LOCAL SOLVABILITY OF AN INITIAL BOUNDARY VALUE PROBLEM FOR A QUASILINEAR HYPERBOLIC-PARABOLIC SYSTEM
Authors: KAGEI YOSHIYUKI , KAWASHIMA SHUICHI
Published: May 2006

This paper investigates the solvability of initial boundary value problem for a quasilinear hyperbolic-parabolic system which consists of a transport equation and strongly parabolic system. The characteristics of the transport equation are assumed to be outward on the boundary of the domain. The unique local (in time) existence of solutions is shown in the class of continuous functions with values in Hs, where s is an integer satisfying s ≥ [n/2]+1.
SELF-SIMILAR ISOTHERMAL IRROTATIONAL MOTION FOR THE EULER, EULER–POISSON SYSTEMS AND THE FORMATION OF THE PLASMA SHEATH
Authors: FELDMAN MIKHAIL , HA SEUNG-YEAL , SLEMROD MARSHALL
Published: May 2006

We consider the self-similar dynamics of a compressible irrotational isothermal fluid and show that a variant of Elling–Liu's ellipticity principle holds. Specially we prove that a closed pseudo-Mach surface is inconsistent with smooth flow. Furthermore, we give an application of our result to the formulation of a plasma sheath in a plasma consisting of cold ions and hot electrons.
SHORT-TIME EXISTENCE FOR SCALE-INVARIANT HAMILTONIAN WAVES
Authors: HUNTER JOHN K.
Published: May 2006

We prove short-time existence of smooth solutions for a class of nonlinear, and in general spatially nonlocal, Hamiltonian evolution equations that describe the self-interaction of weakly nonlinear scale-invariant waves. These equations include ones that describe weakly nonlinear hyperbolic surface waves, such as nonlinear Rayleigh waves in elasticity.
ASYMPTOTIC BEHAVIOR OF SOLUTIONS TO CONSERVATION LAWS WITH PERIODIC FORCING
Authors: AMADORI DEBORA , SERRE DENIS
Published: May 2006

We consider the asymptotic behavior of the solution of a forced scalar conservation law, where both the forcing and the initial data are periodic. We prove that there exists a steady state toward which the solution converges in the L1 norm, as t → ∞. We do not assume any smallness or smoothness of the initial data. The limit steady state can be discontinuous, as effect of resonance, and it can be identified when the potential of the forcing term has a unique global minimum, thanks to the conservation of mass. The flux is assumed to be strictly convex; the relevance of this assumption is justified by the construction of solutions with a lattice of periods for a flux with an inflection point.
Taste Receptors in the Gastrointestinal Tract. I. Bitter taste receptors and α-gustducin in the mammalian gut
Authors: Rozengurt Enrique
Published: May 2006

Molecular sensing by gastrointestinal (GI) cells plays a critical role in the control of multiple fundamental functions in digestion and also initiates hormonal and/or neural pathways leading to the regulation of caloric intake, pancreatic insulin secretion, and metabolism. Molecular sensing in the GI tract is also responsible for the detection of ingested harmful drugs and toxins, thereby initiating responses critical for survival. The initial recognition events and mechanism(s) involved remain incompletely understood. The notion to be discussed in this article is that there are important similarities between the chemosensensory machinery elucidated in specialized neuroepithelial taste receptor cells of the lingual epithelium and the molecular transducers localized recently in enteroendocrine open GI cells that sense the chemical composition of the luminal contents of the gut.