bluuebunny/crossref_metadata_2025 huggingface dataset:
Significance of Heat-Stable and Heat-Labile Enterotoxins in Porcine Colibacillosis in an Additive Model for Pathogenicity Studies
Authors: Zhang Weiping , Berberov Emil M. , Freeling Jessica , He D. , Moxley Rodney A. , Francis David H.
Published: May 2006

Although heat-stable (ST) and heat-labile (LT) enterotoxins produced by enterotoxigenicEscherichia coli(ETEC) have been documented as important factors associated with diarrheal diseases, investigations assessing the contributions of individual enterotoxins to the pathogenesis ofE. coliinfection have been limited. To address the individual roles of enterotoxins in the diarrheal disease caused by K88-positive ETEC in young pigs, enterotoxin-positive and -negative isogenicE. colistrains were constructed by using pBR322 to clone and express LT and STb. Four strains, K88+astA, K88+astA/pBR322, K88+astASTb+, and K88+astALT+, were constructed and subsequently included in gnotobiotic piglet challenge studies, and their pathogenesis was assessed. The results indicated that all K88+isogenic strains were able to colonize the small intestines of piglets exhibiting the K88 receptor. However, only LT- and STb-positive strains caused appreciable diarrhea. Piglets inoculated with the K88+astALT+strain became dehydrated within 18 h, while those inoculated with the K88+astASTb+strain did not, although diarrhea developed in several piglets. The changes in the blood packed-cell volume and plasma total protein of gnotobiotic piglets inoculated with the LT-positive strains were significantly greater than those of pigs inoculated with the K88astA/pBR322 strain (P= 0.012,P= 0.002). Immunochemistry image analysis also suggested that LT enhanced bacterial colonization in a gnotobiotic piglet model. This investigation suggested that LT is a major contributor to the virulence of K88+ETEC and that isogenic constructs are a useful tool for studying the pathogenesis of ETEC infection.
Salmonella enterica
Serovar Typhimurium Requires the Lpf, Pef, and Tafi Fimbriae for Biofilm Formation on HEp-2 Tissue Culture Cells and Chicken Intestinal Epithelium
Authors: Ledeboer Nathan A. , Frye Jonathan G. , McClelland Michael , Jones Bradley D.
Published: May 2006

Recent work has demonstrated that
Salmonella enterica
serovar Typhimurium forms biofilms on HEp-2 tissue culture cells in a type 1 fimbria-dependent manner. To investigate how biofilm growth of HEp-2 tissue culture cells affects gene expression in
Salmonella
, we compared global gene expression during planktonic growth and biofilm growth. Microarray results indicated that the transcription of ∼100 genes was substantially altered by growth in a biofilm. These genes encode proteins with a wide range of functions, including antibiotic resistance, central metabolism, conjugation, intracellular survival, membrane transport, regulation, and fimbrial biosynthesis. The identification of five fimbrial gene clusters was of particular interest, as we have demonstrated that type 1 fimbriae are required for biofilm formation on HEp-2 cells and murine intestinal epithelium. Mutations in each of these fimbriae were constructed in
S. enterica
serovar Typhimurium strain BJ2710, and the mutants were found to have various biofilm phenotypes on plastic, HEp-2 cells, and chicken intestinal tissue. The
pef
and
csg
mutants were defective for biofilm formation on each of the three surfaces tested, while the
lpf
mutant exhibited a complete loss of the ability to form a biofilm on chicken intestinal tissue but only an intermediate loss of the ability to form a biofilm on tissue culture cells and plastic surfaces. The
bcf
mutant displayed increased biofilm formation on both HEp-2 cells and chicken intestinal epithelium, while the
sth
mutant had no detectable biofilm defects. In all instances, the mutants could be restored to a wild-type phenotype by a plasmid carrying the functional genes. This is the first work to identify the genomic responses of
Salmonella
to biofilm formation on host cells, and this work highlights the importance of fimbriae in adhering to and adapting to a eukaryotic cell surface. An understanding of these interactions is likely to provide new insights for intervention strategies in
Salmonella
colonization and infection.
Variation in the Presence of Neuraminidase Genes among
Streptococcus pneumoniae
Isolates with Identical Sequence Types
Authors: Pettigrew Melinda M. , Fennie Kristopher P. , York Matthew P. , Daniels Janeen , Ghaffar Faryal
Published: May 2006

Streptococcus pneumoniae
frequently colonizes the upper respiratory tract of young children and is an important cause of otitis media and invasive disease. Carriage is more common than disease, yet the genetic factors that predispose a given clone for disease are not known. The relationship between capsule type, genetic background, and virulence is complex, and important questions remain regarding how pneumococcal clones differ in their ability to cause disease. Pneumococcal neuraminidase cleaves sialic acid-containing substrates and is thought to be important for pneumococcal virulence. We describe the distribution of multilocus sequence types (ST), capsule type, and neuraminidase genes among 342 carriage, middle ear, blood, and cerebrospinal fluid (CSF) pneumococcal strains from young children. We found 149 STs among our
S. pneumoniae
isolates.
nanA
was present in all strains, while
nanB
and
nanC
were present in 96% and 51% of isolates, respectively. The distribution of
nanC
varied among the strain collections from different tissue sources (
P
= 0.03). The prevalence of
nanC
was 1.41 (95% confidence interval, 1.11, 1.79) times higher among CSF isolates than among carriage isolates. We identified isolates of the same ST that differed in the presence of
nanB
and
nanC
. These studies demonstrate that virulence determinants, other than capsule loci, vary among strains of identical ST. Our studies suggest that the presence of
nanC
may be important for tissue-specific virulence. Studies that both incorporate MLST and take into account additional virulence determinants will provide a greater understanding of the pneumococcal virulence potential.
Th1 Cytokines Facilitate CD8-T-Cell-Mediated Early Resistance to Infection withMycobacterium tuberculosisin Old Mice
Authors: Vesosky Bridget , Flaherty David K. , Turner Joanne
Published: May 2006

Numerous immunological defects begin to emerge as an individual ages, the consequence of which is heightened susceptibility to infectious diseases. Despite this decline in immune function, old mice display an early transient resistance toMycobacterium tuberculosisinfection in the lung, which is dependent on CD8 T cells and gamma interferon (IFN-γ) production. In this study, we investigated the mechanism of resistance by examining the CD8-T-cell phenotype and function in old naïve andM. tuberculosis-infected mice. Pulmonary CD8 T cells from naïve old mice expressed cell surface markers of memory in addition to receptors for several Th1 cytokines. Stimulation of lung cells from naïve old mice with a combination of Th1 cytokines (interleukin-2 [IL-2], IL-12, and IL-18) resulted in nonspecific production of IFN-γ by memory CD8 T cells. Following aerosol infection withM. tuberculosis, the lungs of old mice contained significantly more IL-12, IL-18, and IFN-γ than the lungs of young mice contained. Together, these data demonstrate that the increased and early production of Th1 cytokines in the lungs ofM. tuberculosis-infected old mice, in combination with CD8 T cells that can nonspecifically produce IFN-γ, leads to transient control ofM. tuberculosisgrowth in the lungs of old mice. Further characterization of this mechanism should provide essential information regarding the aging immune system and should contribute to the development of novel strategies to decrease the morbidity and mortality of the aging population associated with infectious diseases.
Heterogeneity in Tandem Octanucleotides within
Haemophilus influenzae
Lipopolysaccharide Biosynthetic Gene
losA
Affects Serum Resistance
Authors: Erwin Alice L. , Bonthuis Paul J. , Geelhood Jennifer L. , Nelson Kevin L. , McCrea Kirk W. , Gilsdorf Janet R. , Smith Arnold L.
Published: May 2006

Haemophilus influenzae
is subject to phase variation mediated by changes in the length of simple sequence repeat regions within several genes, most of which encode either surface proteins or enzymes involved in the synthesis of lipopolysaccharides (LPS). The translational repeat regions that have been described thus far all consist of tandemly repeated tetranucleotides. We describe an octanucleotide repeat region within a putative LPS biosynthetic gene,
losA
. Approximately 20 percent of nontypeable
H. influenzae
strains contain copies of
losA
and
losB
in a genetic locus flanked by
infA
and
ksgA
. Of 30 strains containing
losA
at this site, 24 contained 2 tandem copies of the octanucleotide CGAGCATA, allowing full-length translation of
losA
(on), and 6 strains contained 3, 4, 6, or 10 tandem copies (
losA
off). For a serum-sensitive strain, R3063, with
losA
off (10 repeat units), selection for serum-resistant variants yielded a heterogeneous population in which colonies with increased serum resistance had
losA
on (2, 8, or 11 repeat units), and colonies with unchanged sensitivity to serum had 10 repeats. Inactivation of
losA
in strains R3063 and R2846 (strain 12) by insertion of the
cat
gene decreased the serum resistance of these strains compared to
losA
-on variants and altered the electrophoretic mobility of LPS. We conclude that expression of
losA
, a gene that contributes to LPS structure and affects serum resistance, is determined by octanucleotide repeat variation.
Mycobacterium tuberculosisBinding to Human Surfactant Proteins A and D, Fibronectin, and Small Airway Epithelial Cells under Shear Conditions
Authors: Hall-Stoodley Luanne , Watts Gayle , Crowther Joy E. , Balagopal Ashwin , Torrelles Jordi B. , Robison-Cox James , Bargatze Robert F. , Harmsen Allen G. , Crouch Erika C. , Schlesinger Larry S.
Published: May 2006

A crucial step in infection is the initial attachment of a pathogen to host cells or tissue.Mycobacterium tuberculosishas evolved multiple strategies for establishing an infection within the host. The pulmonary microenvironment contains a complex milieu of pattern recognition molecules of the innate immune system that play a role in the primary response to inhaled pathogens. Encounters ofM. tuberculosiswith these recognition molecules likely influence the outcome of the bacillus-host interaction. Here we use a novel fluid shear assay to investigate the binding ofM. tuberculosisto innate immune molecules that are produced by pulmonary epithelial cells and are thought to play a role in the lung innate immune response. Virulent and attenuatedM. tuberculosisstrains bound best to immobilized human fibronectin (FN) and surfactant protein A (SP-A) under this condition. Binding under fluid shear conditions was more consistent and significant compared to binding under static conditions. Soluble FN significantly increased the adherence of both virulent and attenuatedM. tuberculosisstrains to human primary small airway epithelial cells (SAEC) under fluid shear conditions. In contrast, SP-A and SP-D effects on bacterial adherence to SAEC differed between the two strains. The use of a fluid shear model to simulate physiological conditions within the lung and select for high-affinity binding interactions should prove useful for studies that investigate interactions betweenM. tuberculosisand host innate immune determinants.
Macrophage Migration Inhibitory Factor Contributes to Host Defense against AcuteTrypanosoma cruziInfection
Authors: Reyes José L. , Terrazas Luis I. , Espinoza Bertha , Cruz-Robles David , Soto Virgilia , Rivera-Montoya Irma , Gómez-García Lorena , Snider Heidi , Satoskar Abhay R. , Rodríguez-Sosa Miriam
Published: May 2006

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that is involved in the host defense against several pathogens. Here we used MIF−/−mice to determine the role of endogenous MIF in the regulation of the host immune response againstTrypanosoma cruziinfection. MIF−/−mice displayed high levels of blood and tissue parasitemia, developed severe heart and skeletal muscle immunopathology, and succumbed toT. cruziinfection faster than MIF+/+mice. The enhanced susceptibility of MIF−/−mice toT. cruziwas associated with reduced levels of proinflammatory cytokines, such as tumor necrosis factor alpha, interleukin-12 (IL-12), IL-18, gamma interferon (IFN-γ), and IL-1β, in their sera and reduced production of IL-12, IFN-γ, and IL-4 by spleen cells during the early phase of infection. At all time points, antigen-stimulated splenocytes from MIF+/+and MIF−/−mice produced comparable levels of IL-10. MIF−/−mice also produced significantly less Th1-associated antigen-specific immunoglobulin G2a (IgG2a) throughout the infection, but both groups produced comparable levels of Th2-associated IgG1. Lastly, inflamed hearts fromT. cruzi-infected MIF−/−mice expressed increased transcripts for IFN-γ, but fewer for IL-12 p35, IL-12 p40, IL-23, and inducible nitric oxide synthase, compared to MIF+/+mice. Taken together, our findings show that MIF plays a role in controlling acuteT. cruziinfection.
Infection of Human Dendritic Cells with aMycobacterium tuberculosis sigEMutant Stimulates Production of High Levels of Interleukin-10 but Low Levels of CXCL10: Impact on the T-Cell Response
Authors: Giacomini Elena , Sotolongo Ambar , Iona Elisabetta , Severa Martina , Remoli Maria Elena , Gafa Valerie , Lande Roberto , Fattorini Lanfranco , Smith Issar , Manganelli Riccardo , Coccia Eliana M.
Published: May 2006

TheMycobacterium tuberculosisgenome encodes 13 sigma factors. We have previously shown that mutations in some of these transcriptional activators renderM. tuberculosissensitive to various environmental stresses and can attenuate the virulence phenotype. In this work, we focused on extracytoplasmic factor σEand studied the effects induced by the deletion of its structural gene (sigE) in the infection of human monocyte-derived dendritic cells (MDDC). We found that the wild-typeM. tuberculosisstrain (H37Rv), thesigEmutant (ST28), and the complemented strain (ST29) were able to infect dendritic cells (DC) to similar extents, although at 4 days postinfection a reduced ability to grow inside MDDC was observed for thesigEmutant ST28. After mycobacterium capture, the majority of MDDC underwent full maturation and expressed both inflammatory cytokines, such as tumor necrosis factor alpha, and the regulatory cytokines interleukin-12 (IL-12), IL-18, and beta interferon (IFN-β). Conversely, a higher level of production of IL-10 was observed in ST28-infected MDDC compared to H37Rv- or ST29-infected cell results. However, in spite of the presence of IL-10, supernatants from ST28-infected DC induced IFN-γ production by T cells similarly to those from H37Rv-infected DC culture. On the other hand, IL-10 impaired CXCL10 production insigEmutant-infected DC and, indeed, its neutralization restored CXCL10 secretion. In line with these results, supernatants from ST28-infected cells showed a decreased capability to recruit CXCR3+CD4+T cells compared to those obtained from H37Rv-infected DC culture. Thus, our findings suggest that thesigEmutant-induced secretion of IL-10 inhibits CXCL10 expression and, in turn, the recruitment of activated-effector cells involved in the formation of granulomas.
Identification of a Novel Antigen ofSchistosoma mansoniShared withPlasmodium falciparumand Evaluation of Different Cross-Reactive Antibody Subclasses Induced by Human Schistosomiasis and Malaria
Authors: Pierrot Christine , Wilson Shona , Lallet Hélène , Lafitte Sophia , Jones Frances M. , Daher Wassim , Capron Monique , Dunne David W. , Khalife Jamal
Published: May 2006

Plasmodium falciparumandSchistosoma mansoniare often found in human coinfections, and cross-reactive antibodies to different components of the two parasites have been detected. In this work, we identified a cross-reactiveS. mansonigene product, referred to as SmLRR, that seems to belong to the leucine-rich repeat protein family. Comparative analysis of SmLRR revealed 57% similarity with a putative gene product encoded in theP. falciparumgenome. Antibodies to SmLRR were found in experimental infections and in bothS. mansoni- andP. falciparum-infected individuals. Correlative analysis of human anti-SmLRR responses in Kenya and Uganda suggested that malaria and schistosomiasis drive the immunoglobulin G3 (IgG3) and IgG4 isotypes, respectively, against SmLRR, suggesting that there is differential regulation of cross-reactive isotypes depending on the infection. In addition, the levels of anti-SmLRR IgG4, but not the levels of IgG3, correlated positively with the intensity ofS. mansoniinfection.
Differences in Gene Expression between the Classical and El Tor Biotypes of
Vibrio cholerae
O1
Authors: Beyhan Sinem , Tischler Anna D. , Camilli Andrew , Yildiz Fitnat H.
Published: May 2006

Differences in whole-genome expression patterns between the classical and El Tor biotypes of
Vibrio cholerae
O1 were determined under conditions that induce virulence gene expression in the classical biotype. A total of 524 genes (13.5% of the genome) were found to be differentially expressed in the two biotypes. The expression of genes encoding proteins required for biofilm formation, chemotaxis, and transport of amino acids, peptides, and iron was higher in the El Tor biotype. These gene expression differences may contribute to the enhanced survival capacity of the El Tor biotype in environmental reservoirs. The expression of genes encoding virulence factors was higher in the classical than in the El Tor biotype. In addition, the
vieSAB
genes, which were originally identified as regulators of
ctxA
transcription, were expressed at a fivefold higher level in the classical biotype. We determined the VieA regulon in both biotypes by transcriptome comparison of wild-type and
vieA
deletion mutant strains. VieA predominantly regulates gene expression in the classical biotype; 401 genes (10.3% of the genome), including those encoding proteins required for virulence, exopolysaccharide biosynthesis, and flagellum production as well as those regulated by σ
E
, are differentially expressed in the classical
vieA
deletion mutant. In contrast, only five genes were regulated by VieA in the El Tor biotype. A large fraction (20.8%) of the genes that are differentially expressed in the classical versus the El Tor biotype are controlled by VieA in the classical biotype. Thus, VieA is a major regulator of genes in the classical biotype under virulence gene-inducing conditions.
Rhamnolipids Are Virulence Factors That Promote Early Infiltration of Primary Human Airway Epithelia byPseudomonas aeruginosa
Authors: Zulianello Laurence , Canard Coralie , Köhler Thilo , Caille Dorothée , Lacroix Jean-Silvain , Meda Paolo
Published: May 2006

The opportunistic bacteriumPseudomonas aeruginosacauses chronic respiratory infections in cystic fibrosis and immunocompromised individuals. Bacterial adherence to the basolateral domain of the host cells and internalization are thought to participate inP. aeruginosapathogenicity. However, the mechanism by which the pathogen initially modulates the paracellular permeability of polarized respiratory epithelia remains to be understood. To investigate this mechanism, we have searched for virulence factors secreted byP. aeruginosathat affect the structure of human airway epithelium in the early stages of infection. We have found that only bacterial strains secreting rhamnolipids were efficient in modulating the barrier function of an in vitro-reconstituted human respiratory epithelium, irrespective of their release of elastase and lipopolysaccharide. In contrast to previous reports, we document thatP. aeruginosawas not internalized by epithelial cells. We further report that purified rhamnolipids, applied on the surfaces of the epithelia, were sufficient to functionally disrupt the epithelia and to promote the paracellular invasion of rhamnolipid-deficientP. aeruginosa. The mechanism involves the incorporation of rhamnolipids within the host cell membrane, leading to tight-junction alterations. The study provides direct evidence for a hitherto unknown mechanism whereby the junction-dependent barrier of the respiratory epithelium is selectively altered by rhamnolipids.
Role for Dendritic Cells in Immunoregulation during Experimental Vaginal Candidiasis
Authors: LeBlanc Dana M. , Barousse Melissa M. , Fidel Paul L.
Published: May 2006

Vulvovaginal candidiasis (VVC) caused by the commensal organismCandida albicansremains a significant problem among women of childbearing age, with protection against and susceptibility to infection still poorly understood. While cell-mediated immunity by CD4+Th1-type cells is protective against most forms of mucosal candidiasis, no protective role for adaptive immunity has been identified against VVC. This is postulated to be due to immunoregulation that prohibits a more profoundCandida-specific CD4+T-cell response against infection. The purpose of this study was to examine the role of dendritic cells (DCs) in the induction phase of the immune response as a means to understand the initiation of the immunoregulatory events. Immunostaining of DCs in sectioned murine lymph nodes draining the vagina revealed a profound cellular reorganization with DCs becoming concentrated in the T-cell zone throughout the course of experimental vaginalCandidainfection consistent with cell-mediated immune responsiveness. However, analysis of draining lymph node DC subsets revealed a predominance of immunoregulation-associated CD11c+B220+plasmacytoid DCs (pDCs) under both uninfected and infected conditions. Staining of vaginal DCs showed the presence of both DEC-205+and pDCs, with extension of dendrites into the vaginal lumen of infected mice in close contact withCandida. Flow cytometric analysis of draining lymph node DC costimulatory molecules and activation markers from infected mice indicated a lack of upregulation of major histocompatibility complex class II, CD80, CD86, and CD40 during infection, consistent with a tolerizing condition. Together, the results suggest that DCs are involved in the immunoregulatory events manifested during a vaginalCandidainfection and potentially through the action of pDCs.
Topical Application of
Escherichia coli
-Vectored Vaccine as a Simple Method for Eliciting Protective Immunity
Authors: Zhang Jianfeng , Shi Zhongkai , Kong Fan-kun , Jex Edward , Huang Zhigang , Watt James M. , Van Kampen Kent R. , Tang De-chu C.
Published: May 2006

We report here that animals can be protected against lethal infection by
Clostridium tetani
cells and
Bacillus anthracis
spores following topical application of intact particles of live or γ-irradiated
Escherichia coli
vectors overproducing tetanus and anthrax antigens, respectively. Cutaneous γδT cells were rapidly recruited to the administration site. Live
E. coli
cells were not found in nonskin tissues after topical application, although fragments of
E. coli
DNA were disseminated transiently. Evidence suggested that intact
E. coli
particles in the outer layer of skin may be disrupted by a γδT-cell-mediated innate defense mechanism, followed by the presentation of
E. coli
ligand-adjuvanted intravector antigens to the immune system and rapid degradation of
E. coli
components. The nonreplicating
E. coli
vector overproducing an exogenous immunogen may foster the development of a new generation of vaccines that can be manufactured rapidly and administered noninvasively in a wide variety of disease settings.
The ABC Transporter Protein OppA Provides Protection against Experimental
Yersinia pestis
Infection
Authors: Tanabe Mikio , Atkins Helen S. , Harland David N. , Elvin Stephen J. , Stagg Anthony J. , Mirza Osman , Titball Richard W. , Byrne Bernadette , Brown Katherine A.
Published: May 2006

The identification of
Yersinia pestis
as a potential bioterrorism agent and the emergence of antibiotic-resistant strains have highlighted the need for improved vaccines and treatments for plague. The aim of this study was to evaluate the potential for ATP-binding cassette (ABC) transporter proteins to be exploited as novel vaccines against plague. Western blotting of ABC transporter proteins using sera from rabbits immunized with killed whole
Y. pestis
cells or human convalescent-phase sera identified four immunologically reactive proteins: OppA, PstS, YrbD, and PiuA. Mice immunized with these proteins developed antibody to the immunogen. When the immunized mice were challenged with
Y. pestis
, the OppA-immunized mice showed an increased time to death compared to other groups, and protection appeared to correlate with the level of immunoglobulin G antibody to OppA.
Differential Genome Contents of Nontypeable
Haemophilus influenzae
Strains from Adults with Chronic Obstructive Pulmonary Disease
Authors: Fernaays Matthew M. , Lesse Alan J. , Sethi Sanjay , Cai Xueya , Murphy Timothy F.
Published: May 2006

Haemophilus influenzae
is an important cause of otitis media in children and lower respiratory infection in adults with chronic obstructive pulmonary disease (COPD). Patients with COPD experience periodic exacerbations that are associated with acquisition of new bacterial strains. However, not every strain acquisition is associated with exacerbation. To test the hypothesis that genetic differences among strains account for differences in pathogenic potential, a microarray consisting of 4,992 random 1.5- to 3-kb genomic fragments of an exacerbation strain was constructed. Competitive hybridization was performed using six strains associated with exacerbation as well as five strains associated with asymptomatic colonization. Seven sequences that were absent in all five colonization strains and present in at least two exacerbation strains were identified. One such sequence was a previously unreported gene with high homology to the meningococcal immunoglobulin A (IgA) protease gene, which is distinct from the previously described
H. influenzae
IgA protease. To assess the distribution of the seven sequences among well-characterized strains of
H. influenzae
, 59 exacerbation strains and 73 asymptomatic colonization strains were screened by PCR for the presence of these sequences. The presence or absence of any single sequence was not significantly associated with exacerbations of COPD. However, logistic regression and subgroup analysis identified combinations of the presence and absence of genes that are associated with exacerbations. These results indicate that patterns of genes are associated with the ability of strains of
H. influenzae
to cause exacerbations of COPD, supporting the concept that differences in pathogenic potential are based in part on genomic differences among infecting strains, not merely host factors.
Borrelia burgdorferi
OspC Protein Required Exclusively in a Crucial Early Stage of Mammalian Infection
Authors: Tilly Kit , Krum Jonathan G. , Bestor Aaron , Jewett Mollie W. , Grimm Dorothee , Bueschel Dawn , Byram Rebecca , Dorward David , VanRaden Mark J. , Stewart Philip , Rosa Patricia
Published: May 2006

This study demonstrates a strict temporal requirement for a virulence determinant of the Lyme disease spirochete
Borrelia burgdorferi
during a unique point in its natural infection cycle, which alternates between ticks and small mammals. OspC is a major surface protein produced by
B. burgdorferi
when infected ticks feed but whose synthesis decreases after transmission to a mammalian host. We have previously shown that spirochetes lacking OspC are competent to replicate in and migrate to the salivary glands of the tick vector but do not infect mice. Here we assessed the timing of the requirement for OspC by using an
ospC
mutant complemented with an unstable copy of the
ospC
gene and show that
B. burgdorferi
's requirement for OspC is specific to the mammal and limited to a critical early stage of mammalian infection. By using this unique system, we found that most bacterial reisolates from mice persistently infected with the initially complemented
ospC
mutant strain no longer carried the wild-type copy of
ospC
. Such spirochetes were acquired by feeding ticks and migrated to the tick salivary glands during subsequent feeding. Despite normal behavior in ticks, these
ospC
mutant spirochetes did not infect naive mice.
ospC
mutant spirochetes from persistently infected mice also failed to infect naive mice by tissue transplantation. We conclude that OspC is indispensable for establishing infection by
B. burgdorferi
in mammals but is not required at any other point of the mouse-tick infection cycle.
Global Gene Expression Profiling of the Asymptomatic BacteriuriaEscherichia coliStrain 83972 in the Human Urinary Tract
Authors: Roos Viktoria , Klemm Per
Published: May 2006

Urinary tract infections (UTIs) are an important health problem worldwide, with many million cases each year.Escherichia coliis the most common organism causing UTIs in humans. The asymptomatic bacteriuriaE. colistrain 83972 is an excellent colonizer of the human urinary tract, where it causes long-term bladder colonization. The strain has been used for prophylactic purposes in patients prone to more severe and recurrent UTIs. For this study, we used DNA microarrays to monitor the expression profile of strain 83972 in the human urinary tract. Significant differences in expression levels were seen between the in vivo expression profiles of strain 83972 in three patients and the corresponding in vitro expression profiles in lab medium and human urine. The data revealed an in vivo lifestyle of microaerobic growth with respiration of nitrate coupled to degradation of sugar acids and amino acids, with no signs of attachment to host tissues. Interestingly, genes involved in NO protection and metabolism showed significant up-regulation in the patients. This is one of the first studies to address bacterial whole-genome expression in humans and the first study to investigate global gene expression of anE. colistrain in the human urinary tract.
High Levels of Interleukin-10 Impair Resistance to Pulmonary Coccidioidomycosis in Mice in Part through Control of Nitric Oxide Synthase 2 Expression
Authors: Jimenez Maria del Pilar , Walls Lorraine , Fierer Joshua
Published: May 2006

We have shown previously that there is a direct correlation between IL-10 levels and susceptibility toCoccidioides immitisperitonitis in C57BL/6 (B6), DBA/2, and BXD recombinant inbred mice. We now show that B6 mice are also more susceptible toC. immitispneumonia and that interleukin-10 (IL-10)-deficient (IL-10−/−) B6 mice are more resistant toC. immitispneumonia. In addition, we established that high levels of IL-10 are sufficient to make genetically resistant mice susceptible to bothC. immitisperitonitis and pneumonia by infecting h.IL-10 transgenic mice. Infected h.IL-10 transgenic mice express lower levels of gamma interferon, IL-12 p40, and inducible nitric oxide synthetase 2 (NOS2) mRNA in their lungs, implicating inducible NOS as a defense mechanism in this disease. We treated DBA/2 mice with aminoguanidine, and they became more susceptible toC. immitisperitonitis and pneumonia. We conclude that high levels of IL-10 are both necessary and sufficient to make mice susceptible toC. immitis, regardless of the genetic background of the mice, and that IL-10 impairs resistance toC. immitisin part by suppressing NO synthesis.
Differential Expression of the Bhmp39 Major Outer Membrane Proteins of
Brachyspira hyodysenteriae
Authors: Witchell Timothy D. , Coutts Scott A. J. , Bulach Dieter M. , Adler Ben
Published: May 2006

The enteric, anaerobic spirochete
Brachyspira hyodysenteriae
is the causative agent of swine dysentery, a severe mucohemorrhagic diarrheal disease of pigs that has economic significance in every major pork-producing country. Recent investigation into potential vaccine candidates has focused on the outer membrane proteins of
B. hyodysenteriae
. Bhmp39 (formerly Vsp39) is the most abundant surface-exposed outer membrane protein of
B. hyodysenteriae
; its predicted gene sequence has previously been shown to share sequence similarity to eight genes divided evenly between two paralogous loci. The peptide sequence suggested that Bhmp39 is encoded by one of these genes,
bhmp39h
. The biological significance of maintaining eight homologous
bhmp39
genes is unclear, though it has been proposed that this may play a role in antigenic variation. In this study, real-time, reverse transcription-PCR was used to demonstrate that
bhmp39f
and
bhmp39h
were the transcripts most abundantly expressed by
B. hyodysenteriae
strain B204 cultured under in vitro growth conditions. Mass spectrometry data of the purified 39-kDa membrane protein showed that both Bhmp39f and Bhmp39h were present. Northern blot analysis across predicted Rho-independent terminators demonstrated that the genes of the
bhmp39efgh
locus result in monocistronic transcripts.
DifferentialRequirements for Soluble and Transmembrane Tumor Necrosis Factor in theImmunological Control of Primary and SecondaryListeria monocytogenesInfection
Authors: Musicki Korana , Briscoe Helen , Tran Stephen , Britton Warwick J. , Saunders Bernadette M.
Published: May 2006

The relative contributions of transmembrane tumor necrosis factor (memTNF) and soluble tumor necrosis factor (solTNF) in innate and adaptive immunity are poorly defined. We examined the capacities of wild-type (WT) mice, TNF−/−mice, and memTNF mice, which express only transmembrane TNF, to control primary and secondaryListeria monocytogenesinfections. Soluble TNF was not required for induction or maintenance of protective immunity against a low-dose (200-CFU)Listeriainfection. In contrast to TNF−/−mice, both WT and memTNF mice cleared the bacilli within 10 days and were fully protected against rechallenge with a lethal infective dose. Furthermore, T cells transferred from immune mice, but not from naïve, WT, and memTNF mice, protected TNF−/−recipients against an otherwise lethal infection. By contrast, infection with a higher dose ofListeria(2,000 CFU) clearly demonstrated that solTNF is required to coordinate an optimal protective inflammatory response. memTNF mice were more susceptible to a high-dose infection, and they exhibited delayed bacterial clearance, increased inflammation, and necrosis in the liver that resulted in 55% mortality. The dysregulated inflammation was accompanied by prolonged elevated expression of mRNAs for several chemokines as well as the macrophage effector molecules inducible nitric oxide synthase and LRG-47 in the livers of memTNF mice but not in the livers of WT mice. These data demonstrated that memTNF is sufficient for establishing protective immunity against a primary low-doseListeriainfection but that solTNF is required for optimal control of cellular inflammation and resistance to a primary high-dose infection. By contrast, memTNF alone is sufficient for resolution of a secondary, high-dose infection and for the transfer of protective immunity with memory T cells.
Basis for the Failure ofFrancisella tularensisLipopolysaccharide To Prime Human Polymorphonuclear Leukocytes
Authors: Barker Jason H. , Weiss Jerrold , Apicella Michael A. , Nauseef William M.
Published: May 2006

Francisella tularensisis the intracellular gram-negative coccobacillus that causes tularemia, and its virulence and infectiousness make it a potential agent of bioterrorism. Previous studies using mononuclear leukocytes have shown that the lipopolysaccharide (LPS) ofF. tularensisis neither a typical proinflammatory endotoxin nor an endotoxin antagonist. This inertness suggests thatF. tularensisLPS does not bind host LPS-sensing molecules such as LPS-binding protein (LBP). Using priming of the polymorphonuclear leukocyte (PMN) oxidase as a measure of endotoxicity, we found thatF. tularensislive vaccine strain LPS did not behave like either a classic endotoxin or an endotoxin antagonist in human PMNs, even when the concentration of LBP was limiting. Furthermore,F. tularensisLPS did not compete with a radiolabeled lipooligosaccharide fromNeisseria meningitidisfor binding to LBP or to the closely related PMN granule protein, bactericidal/permeability-increasing protein. Our results suggest that the inertness ofF. tularensisLPS and the resistance ofF. tularensisto oxygen-independent PMN killing may result from the inability ofF. tularensisLPS to be recognized by these important LPS-sensing molecules of the innate immune system.
Borrelia burgdorferi
Lacking BBK32, a Fibronectin-Binding Protein, Retains Full Pathogenicity
Authors: Li Xin , Liu Xianzhong , Beck Deborah S. , Kantor Fred S. , Fikrig Erol
Published: May 2006

BBK32, a fibronectin-binding protein of
Borrelia burgdorferi
, is one of many surface lipoproteins that are differentially expressed by the Lyme disease spirochete at various stages of its life cycle. The level of BBK32 expression in
B. burgdorferi
is highest during infection of the mammalian host and lowest in flat ticks. This temporal expression profile, along with its fibronectin-binding activity, strongly suggests that BBK32 may play an important role in Lyme pathogenesis in the host. To test this hypothesis, we constructed an isogenic BBK32 deletion mutant from wild-type
B. burgdorferi
B31 by replacing the BBK32 gene with a kanamycin resistance cassette through homologous recombination. We examined both the wild-type strain and the BBK32 deletion mutant extensively in the experimental mouse-tick model of the
Borrelia
life cycle. Our data indicated that
B. burgdorferi
lacking BBK32 retained full pathogenicity in mice, regardless of whether mice were infected artificially by syringe inoculation or naturally by tick bite. The loss of BBK32 expression in the mutant had no adverse effect on spirochete acquisition (mouse-to-tick) and transmission (tick-to-mouse) processes. These results suggest that additional
B. burgdorferi
proteins can complement the function of BBK32, fibronectin binding or otherwise, during the natural spirochete life cycle.
Members of a
Legionella pneumophila
Family of Proteins with ExoU (Phospholipase A) Active Sites Are Translocated to Target Cells
Authors: VanRheenen Susan M. , Luo Zhao-Qing , O'Connor Tamara , Isberg Ralph R.
Published: May 2006

Legionella
pneumophila
replicates within alveolar macrophages, causing a
severe pneumonia termed Legionnaires' disease. The bacterium resides
within a vacuole that escapes immediate transport to the host lysosome.
Instead, the vacuole interacts with the early secretory pathway to
establish an environment suitable for rapid multiplication. A type IV
secretion system is central to the pathogenicity of the bacterium, and
many protein substrates that are translocated by this system to the
host cell have been identified. One of these, VipD, was found to
interrupt the late secretory pathway when overproduced in
Saccharomyces cerevisiae
. We independently identified VipD in
a previous study and have further characterized this protein as well as
its three paralogs. The
vipD
gene belongs to a family of
L. pneumophila
open reading frames that are predicted to
contain a phospholipase A domain with sequence similarity to the type
III-secreted toxin ExoU from
Pseudomonas aeruginosa
. Similarly
to other known translocated proteins of
L. pneumophila
, VipD
is strongly induced in early stationary phase, a time when the
bacterium is most virulent. Detergent extraction studies of infected
macrophages confirm that VipD is translocated into host cells via the
type IV secretion system. A second assay for translocation revealed
that two paralogs of VipD, VpdA and VpdB, also have translocation
signals recognized by the type IV system. A strain lacking VipD and its
three paralogs grew at wild-type rates in murine macrophages, although
secondary mutations that cause growth defects in strains lacking VipD
accumulate. The quadruple mutant displayed a growth advantage in the
amoebal host
Dictyostelium discoideum
, indicating that the
protein family may modulate intracellular growth in a complex fashion.
VipD is mildly toxic when overproduced in eukaryotic cells, and the
toxicity is partially dependent on the putative phospholipase active
site. VipD and its paralogs therefore define a family of translocated
proteins that may assist in the establishment of a vacuole suitable for
bacterial replication through functioning as a
phospholipase.
Eosinophils Can Function as Antigen-Presenting Cells To Induce Primary and Secondary Immune Responses toStrongyloides stercoralis
Authors: Padigel Udaikumar M. , Lee James J. , Nolan Thomas J. , Schad Gerhard A. , Abraham David
Published: May 2006

Several studies have demonstrated roles for eosinophils during innate and adaptive immune responses to helminth infections. However, evidence that eosinophils are capable of initiating an immune response to parasite antigens is lacking. The goal of the present in vitro study was to investigate the potential of eosinophils to serve as antigen-presenting cells (APC) and initiate an immune response to parasite antigens. Purified eosinophils were exposed to solubleStrongyloides stercoralisantigens, and the expression of various surface markers involved in cell activation was examined. Antigen-exposed eosinophils showed a sixfold increase in expression levels of CD69 and major histocompatibility complex (MHC) class II, a fourfold increase in levels of T-cell costimulatory molecule CD86, and a twofold decrease in levels of CD62L compared to eosinophils cultured in medium containing granulocyte-macrophage colony-stimulating factor. The ability of eosinophils to present antigen to T cells was determined by culturing them with T cells in vitro. Eosinophils pulsed with antigen stimulated antigen-specific primed T cells and CD4+T cells to increase interleukin-5 (IL-5) production. The blocking of MHC class II expression on eosinophils inhibited their ability to induce IL-5 production by CD4+T cells in culture. Antigen-pulsed eosinophils were able to prime naïve T cells and CD4+T cells in culture and polarized them into Th2 cells producing IL-5 similar to that induced by antigen-loaded dendritic cells. These results demonstrate that eosinophils are capable of activating antigen-specific Th2 cells inducing the release of cytokines and assist in the priming of naïve T cells to initiate Th2 responses against infection. This study highlights the potential of eosinophils to actively induce immune responses against infection by amplifying antigen-specific Th2-cell responses.
Immunoglobulin A Antibodies against Ricin A and B Subunits Protect Epithelial Cells from Ricin Intoxication
Authors: Mantis Nicholas J. , McGuinness Carolyn R. , Sonuyi Oluwakemi , Edwards Gary , Farrant Stephanie A.
Published: May 2006

Epithelial cells of the respiratory and gastrointestinal tracts are extremely vulnerable to the cytotoxic effects of ricin, a Shiga-like toxin with ribosome-inactivating properties. While mucosal immunity to ricin correlates with secretory immunoglobulin A (IgA) antibody levels in vivo, the potential of IgA to protect epithelial cells from ricin in vitro has not been examined due to the unavailability of well-defined antitoxin IgA antibodies. Here we report the characterization of four monoclonal IgA antibodies (IgA MAbs) produced from the Peyer's patches and mesenteric lymph nodes of BALB/c mice immunized intragastrically with ricin toxoid. Two IgA MAbs (33G2 and 35H6) were active against ricin's lectin subunit (RTB), and two (23D7 and 25A4) reacted with the toxin's enzymatic subunit (RTA). All four IgA MAbs neutralized ricin in a Vero cell cytotoxicity assay, blocked toxin-induced interleukin-8 release by the human monocyte/macrophage cell line 28SC, and protected polarized epithelial cell monolayers from ricin-mediated protein synthesis inhibition. 33G2 and 35H6 reduced ricin binding to the luminal surfaces of human intestinal epithelial cells to undetectable levels in tissue section overlay assays, whereas 23D7 had no effect on toxin attachment. 23D7 and 25A4 did, however, reduce ricin transcytosis across MDCK II cell monolayers, possibly by interfering with intracellular toxin transport. We conclude that IgA antibodies against RTA and RTB can protect mucosal epithelial cells from ricin intoxication.
Inorganic Phosphate Induces Spore Morphogenesis and Enterotoxin Production in the Intestinal Pathogen
Clostridium perfringens
Authors: Philippe Valeria A. , Méndez Marcelo B. , Huang I-Hsiu , Orsaria Lelia M. , Sarker Mahfuzur R. , Grau Roberto R.
Published: May 2006

Clostridium perfringens
enterotoxin (CPE) is an important virulence factor for food poisoning and non-food borne gastrointestinal (GI) diseases. Although CPE production is strongly regulated by sporulation, the nature of the signal(s) triggering sporulation remains unknown. Here, we demonstrated that inorganic phosphate (P
i
), and not pH, constitutes an environmental signal inducing sporulation and CPE synthesis. In the absence of P
i
-supplementation,
C. perfringens
displayed a
spo0A
phenotype, i.e., absence of polar septation and DNA partitioning in cells that reached the stationary phase of growth. These results received support from our Northern blot analyses which demonstrated that P
i
was able to counteract the inhibitory effect of glucose at the onset of sporulation and induced
spo0A
expression, indicating that P
i
acts as a key signal triggering spore morphogenesis. In addition to being the first study reporting the nature of a physiological signal triggering sporulation in clostridia, these findings have relevance for the development of antisporulation drugs to prevent or treat CPE-mediated GI diseases in humans.
The triad of shift work, occupational noise, and physical workload and risk of coronary heart disease
Authors: Virkkunen H , Härmä M , Kauppinen T , Tenkanen L
Published: May 2006

Background:Shift work, noise, and physical workload are very common occupational exposures and they tend to cluster in the same groups of workers.

Objectives:To study the short and long term effects of these exposures on risk of coronary heart disease (CHD) and to estimate the joint effects of these factors.

Methods:The study population in this prospective 13 year follow up study of 1804 middle aged industrially employed men was collected at the first screening for the Helsinki Heart Study. The CHD end points (ICD-9 codes 410–414 and ICD-10 codes I20–I25) were obtained from official Finnish registers. The Finnish job-exposure matrix FINJEM provided information on occupational exposures. Relative risks (RR) of CHD for the exposures were estimated using Cox’s proportional hazard models adjusting for classical risk factors of CHD.

Results:The RR in the five year follow up for continuous noise combined with impulse noise was 1.28; for shift work it was 1.59, and for physical workload 1.18, while in the 13 year follow up the RRs were 1.58, 1.34, and 1.31, respectively. When adjusted for white-collar/blue-collar status the RRs decreased markedly. The RR in the 13 year follow up for those exposed to two risk factors was close to 1.7 and for those exposed to all three, 1.87.

Conclusion:Shift work and continuous noise entailed an excess risk for CHD in the shortest follow up with only a few retired workers but a decreasing risk during the longer follow up. For physical workload and impulse noise the trend was opposite: the CHD risk was increasing with increasing follow up time despite increasing numbers of retired workers.
NONLINEAR ASYMPTOTICS FOR HYPERBOLIC INTERNAL WAVES OF SMALL WIDTH
Authors: GUES OLIVIER , RAUCH JEFFREY
Published: May 2006

Semilinear hyperbolic problems with source terms piecewise smooth and discontinuous across characteristic surfaces yield similarly piecewise smooth solutions. If the discontinuous source is replaced with a smooth transition layer, the discontinuity of the solution is replaced by a smooth internal layer. In this paper we describe how the layer structure of the solution can be computed from the layer structure of the source. The key idea is to use a transmission problem strategy for the problem with the smooth internal layer. That leads to an anastz different from the obvious candidates. The obvious candidates lead to overdetermined equations for correctors. With the transmission problem strategy we compute infinitely accurate expansions.