bluuebunny/crossref_metadata_2025 huggingface dataset:
In Vivo Interleukin-6 Protects Neutrophils from Apoptosis in Osteomyelitis
Authors: Asensi Víctor , Valle Eulalia , Meana Alvaro , Fierer Joshua , Celada Antonio , Alvarez Victoria , Paz José , Coto Eliecer , Carton José Antonio , Maradona José Antonio , Dieguez Angeles , Sarasúa Julián , Ocaña Marcos G. , Arribas José Manuel
Published: June 2004

Polymorphonuclear neutrophils are critical for resolution of bacterial infections. In tissues, most of the neutrophils quickly die through apoptosis. Using propidium iodide DNA staining and DNA gel electrophoresis, we found that spontaneous apoptosis of neutrophils from patients suffering osteomyelitis (n= 52) was significantly decreased in relation to control neutrophils (n= 20) (40.2% ± 25.2% versus 54.5% ± 23.5%;P< 0.03). Incubation of neutrophils from normal volunteers with sera from patients with osteomyelitis reduced apoptosis from 79.1% ± 14.8% in control sera to 62.2% ± 18.7% in osteomyelitis sera. A significant increase of serum interleukin-6 (IL-6) and IL-1α was found in osteomyelitis (IL-6, 8.8 ± 11.9 pg/ml versus 1.8 ± 1.2 pg/ml in controls [P< 0.004]; IL-1α, 3.8 ± 6.4 pg/ml versus 1.0 ± 2.2 pg/ml in controls [P< 0.02]). No differences in the levels of other cytokines, such as tumor necrosis factor alpha, were found. There was an inverse correlation between IL-6 levels and neutrophil apoptosis (r= −0.855;P< 0.007), but this was not the case for other cytokines. The antiapoptotic effect of the osteomyelitis sera was reversed with anti-IL-6 antibodies (P< 0.03) and was reproduced with recombinant human IL-6 (P< 0.001). The longer life span of neutrophils in osteomyelitis induced by IL-6 could contribute to the tissue damage that occurs in these chronic bone infections.
Intestinal P Glycoprotein Acts as a Natural Defense Mechanism againstListeria monocytogenes
Authors: Neudeck Brien L. , Loeb Jennifer M. , Faith Nancy G. , Czuprynski Charles J.
Published: June 2004

Mechanisms by which the intestinal epithelium resists invasion by food-borne pathogens such asListeria monocytogenesare an evolving area of research. Intestinal P glycoprotein is well known to limit the absorption of xenobiotics and is believed to act as a cytotoxic defense mechanism. The aim of this study was to determine if intestinal P glycoprotein is involved in host defense againstL. monocytogenes. Caco-2 cells and a P-glycoprotein-overexpressing subclone (Caco-2/MDR) were employed in addition tomdr1a−/−mice and wild-type controls. In vitro invasion assays and in vivo experiments were employed to measure bacterial invasion and dissemination. In addition,L. monocytogenesproteins were labeled with [35S]methionine, and the transepithelial transport across Caco-2 monolayers was characterized in both directions. Overexpression of P glycoprotein in Caco-2/MDR cells led to increased resistance toL. monocytogenesinvasion, whereas P-glycoprotein inhibition led to increased invasion. Flux of [35S]methionine-labeledL. monocytogenesproteins was significantly greater in the basolateral-to-apical direction than in the apical-to-basolateral direction, indicating dependence on an apically located efflux transporter. Moreover, inhibiting P glycoprotein reduced the basolateral-to-apical flux of the proteins. Early dissemination ofL. monocytogenesfrom the gastrointestinal tract was significantly greater in themdr1a−/−mice than in wild-type controls. Expression and function of intestinal P glycoprotein is an important determinant in resistance to early invasion ofL. monocytogenes.
Anaplasma phagocytophilum
Has a Functional
msp2
Gene That Is Distinct from
p44
Authors: Lin Quan , Rikihisa Yasuko , Felek Suleyman , Wang Xueqi , Massung Robert F. , Woldehiwet Zerai
Published: June 2004

The
msp2
and
p44
genes encode polymorphic major outer membrane proteins that are considered unique to the intraerythrocytic agent of
Anaplasma marginale
and the intragranulocytic agent of
Anaplasma phagocytophilum
, respectively. In the present study, however, we found an
msp2
gene in
A. phagocytophilum
that was remarkably conserved among
A. phagocytophilum
strains from human granulocytic anaplasmosis (HGA) patients, ticks, and a horse from various regions in the United States, but the gene was different in a sheep isolate from the United Kingdom. The
msp2
gene in the
A. phagocytophilum
strain HZ genome was a single-copy gene and was located downstream of two
Ehrlichia chaffeensis omp-1
homologs and a decarboxylase gene (
ubiD
). The
msp2
gene was expressed by
A. phagocytophilum
in the blood from HGA patients NY36 and NY37 and by
A. phagocytophilum
isolates from these patients cultured in HL-60 cells at 37°C. The
msp2
gene was also expressed in a DBA/2 mouse infected by attaching ticks infected with strain NTN-1 and in a horse experimentally infected by attaching strain HZ-infected ticks. However, the transcript of the
msp2
gene was undetectable in
A. phagocytophilum
strain HZ in SCID mice and
Ixodes scapularis
ticks infected with strain NTN-1. These results indicate that
msp2
is functional in various strains of
A. phagocytophilum
, and relative expression ratios of
msp2
to
p44
vary in different infected hosts. These findings may be important in understanding roles that Msp2 proteins play in granulocytic ehrlichia infection and evolution of the polymorphic major outer membrane protein gene families in
Anaplasma
species.
Identification of an I-Ed-Restricted T-Cell Epitope ofEscherichia coliOuter Membrane Protein F
Authors: Williams Kristina M. , Bigley Elmer C.
Published: June 2004

A predominant T-cell epitope ofEscherichia coliouter membrane protein F (OmpF) that encompasses amino acids 295 to 314 was identified in H-2dmice. BALB/c-derived T-cell hybridomas generated against this region were CD3+, CD4+, CD8−, and T-cell receptor αβ+and secreted TH-1-associated cytokines (interleukin-2 [IL-2] and gamma interferon), but not a TH-2-associated cytokine (IL-4), when restimulated with peptide 295-314. Class II+mouse lymphoma (A20) cells, but not class II(−) mouse mastocytoma (P815) cells, supported IL-2 secretion of hybridomas when substituted for syngeneic splenocytes as antigen-presenting cells (APCs). Antibodies specific for I-Edblocked IL-2 secretion by hybridomas, but I-Ad-specific antiserum did not. When transfected L cells expressing I-Ad(AαAβd), I-Ed(EαEβd), or the hybrid molecule I-EαAβdwere used as APCs, hybridomas recognized peptide only when presented by the I-Ed-transfected cells. When peptide 295-314 truncated at either the C or the N terminus of the sequence was used, the minimal epitope was determined. Critical residues were determined by using alanine-substituted peptide analogues. T-cell hybridomas were only stimulated by peptides that encompassed amino acids 295 to 303 (9-mer), and the core sequence required a minimum of three additional amino acids at either the amino or the carboxy terminus to induce IL-2 secretion. Critical residues were determined to be phenylalanine at position 295, threonine at position 300, and tyrosines at positions 301 and 302. This study is the first to identify a minimal T-cell epitope and major histocompatibility complex restriction element of the OmpF protein and confirms previous observations that there is considerable degeneracy in the length of peptides that can bind I-Edand variability in the amino acid composition of the C and N termini of these peptides.
Ultrastructure ofProteus mirabilisSwarmer Cell Rafts and Role of Swarming in Catheter-Associated Urinary Tract Infection
Authors: Jones Brian V. , Young Robert , Mahenthiralingam Eshwar , Stickler David J.
Published: June 2004

Proteus mirabilisis a common cause of catheter-associated urinary tract infection (C-UTI). It blocks indwelling urethral catheters through the formation of extensive crystalline biofilms. The obstruction of urine flow can induce episodes of pyelonephritis, septicemia, and shock.P. mirabilisexhibits a type of motility referred to as swarming, in which multicellular rafts of elongated, hyperflagellated swarmer cells form and move rapidly in concert over solid surfaces. It has been suggested that swarming is important in the pathogenesis of C-UTI. In this study we generated a set of stable transposon mutants deficient in swarming and used them to assess the role of swarming in the migration ofP. mirabilisover urinary catheters. Swarming was found to be essential for migration over all-silicone catheters. Swarming-deficient mutants were attenuated in migration over hydrogel-coated latex catheters, but those capable of swimming motility were able to move over and infect these surfaces. A novel vapor fixation technique for the preparation of specimens and scanning electron microscopy were used to resolve the ultrastructure ofP. mirabilismulticellular rafts. The flagellar filaments ofP. mirabiliswere found to be highly organized during raft migration and were interwoven in phase to form helical connections between adjacent swarmer cells. Mutants lacking these novel organized structures failed to swarm successfully. We suggest that these structures are important for migration and formation of multicellular rafts. In addition, the highly organized structure of multicellular rafts enablesP. mirabilisto initiate C-UTI by migration over catheter surfaces from the urethral meatus into the bladder.
Murine Oviduct Epithelial Cell Cytokine Responses toChlamydia muridarumInfection Include Interleukin-12-p70 Secretion
Authors: Johnson Raymond M.
Published: June 2004

Epithelial cells play an important role in host defense as sentinels for invading microbial pathogens.Chlamydia trachomatisis an intracellular bacterial pathogen that replicates in reproductive tract epithelium. Epithelial cells lining the reproductive tract likely play a key role in triggering inflammation and adaptive immunity duringChlamydiainfections. For this report a murine oviduct epithelial cell line was derived in order to determine how epithelial cells influence innate and adaptive immune responses duringChlamydiainfections. As expected, oviduct epithelial cells infected byChlamydia muridarumproduced a broad spectrum of chemokines, including CXCL16, and regulators of the acute-phase response, including interleukin-1α (IL-1α), IL-6, and tumor necrosis factor alpha. In addition, infected epithelial cells expressed cytokines that augment gamma interferon (IFN) production, including IFN-α/β and IL-12-p70. To my knowledge this is the first report of a non-myeloid/lymphoid cell type making IL-12-p70 in response to an infection. Equally interesting, infected epithelial cells significantly upregulated transforming growth factor alpha precursor expression, suggesting a mechanism by which they might play a direct role in the pathological scarring seen as a consequence ofChlamydiainfections. Data from these in vitro studies predict that infected oviduct epithelium contributes significantly to host innate and adaptive defenses but may also participate in the immunopathology seen withChlamydiainfections.
Identification of
Burkholderia cenocepacia
Genes Required for Bacterial Survival In Vivo
Authors: Hunt Tracey A. , Kooi Cora , Sokol Pamela A. , Valvano Miguel A.
Published: June 2004

Burkholderia cenocepacia
(formerly
Burkholderia cepacia
complex genomovar III) causes chronic lung infections in patients with cystic fibrosis. In this work, we used a modified signature-tagged mutagenesis (STM) strategy for the isolation of
B. cenocepacia
mutants that cannot survive in vivo. Thirty-seven specialized plasposons, each carrying a unique oligonucleotide tag signature, were constructed and used to examine the survival of 2,627
B. cenocepacia
transposon mutants, arranged in pools of 37 unique mutants, after a 10-day lung infection in rats by using the agar bead model. The recovered mutants were screened by real-time PCR, resulting in the identification of 260 mutants which presumably did not survive within the lungs. These mutants were repooled into smaller pools, and the infections were repeated. After a second screen, we isolated 102 mutants unable to survive in the rat model. The location of the transposon in each of these mutants was mapped within the
B. cenocepacia
chromosomes. We identified mutations in genes involved in cellular metabolism, global regulation, DNA replication and repair, and those encoding bacterial surface structures, including transmembrane proteins and cell surface polysaccharides. Also, we found 18 genes of unknown function, which are conserved in other bacteria. A subset of 12 representative mutants that were individually examined using the rat model in competition with the wild-type strain displayed reduced survival, confirming the predictive value of our STM screen. This study provides a blueprint to investigate at the molecular level the basis for survival and persistence of
B. cenocepacia
within the airways.
Mucosal Vaccination against Serogroup B Meningococci: Induction of Bactericidal Antibodies and Cellular Immunity following Intranasal Immunization with NadA of
Neisseria meningitidis
and Mutants of
Escherichia coli
Heat-Labile Enterotoxin
Authors: Bowe Frances , Lavelle Ed C. , McNeela Edel A. , Hale Christine , Clare Simon , Arico Beatrice , Giuliani Marzia M. , Rae Aaron , Huett Alan , Rappuoli Rino , Dougan Gordon , Mills Kingston H. G.
Published: June 2004

Conjugated polysaccharide vaccines protect against serogroup C meningococci. However, this approach cannot be applied to serogroup B, which is still a major cause of meningitis. We evaluated the immunogenicity of three surface-exposed proteins from serogroup B
Neisseria meningitidis
(App, NhhA, and NadA) identified during whole-genome sequencing. Mice were immunized intranasally with individual proteins in the presence of wild-type
Escherichia coli
heat-labile enterotoxin (LTwt), LTR72, a partially inactivated mutant, or LTK63, a completely nontoxic mutant, as the adjuvant. Each of the meningococcal proteins induced significant cellular responses; NhhA and NadA induced strong antibody responses, but only NadA induced bactericidal antibody when administered intranasally with mucosal adjuvants. In addition, immunoglobulin A and bactericidal antibodies were detected in the respiratory tract following intranasal delivery of NadA. Analysis of antigen-specific cytokine production by T cells from immunized mice revealed that intranasal immunization with NadA alone failed to generate detectable cellular immune responses. In contrast, LTK63, LTR72, and LTwt significantly augmented NadA-specific gamma interferon, interleukin-4 (IL-4), IL-5, and IL-10 production by spleen and lymph node cells, suggesting that both Th1 and Th2 cells were induced in vivo. The strongest cellular responses and highest bactericidal antibody titers were generated with LTR72 as the adjuvant. These findings demonstrate that the quality and magnitude of the immune responses generated by mucosal vaccines are influenced by the antigen as well as the adjuvant and suggest that nasal delivery of NadA with mucosal adjuvants has considerable potential in the development of a mucosal vaccine against serogroup B meningococci.
egc
-Encoded Superantigens from
Staphylococcus aureus
Are Neutralized by Human Sera Much Less Efficiently than Are Classical Staphylococcal Enterotoxins or Toxic Shock Syndrome Toxin
Authors: Holtfreter Silva , Bauer Kristin , Thomas Damien , Feig Christine , Lorenz Vera , Roschack Katharina , Friebe Erika , Selleng Kathleen , Lövenich Sandra , Greve Timm , Greinacher Andreas , Panzig Brigitte , Engelmann Susanne , Lina Gérard , Bröker Barbara M.
Published: June 2004

PCR was employed to determine the presence of all known superantigen genes (
sea
,
seq
, and
tst
) and of the exotoxin-like gene cluster (
set
) in 40
Staphylococcus aureus
isolates from blood cultures and throat swabs; 28 isolates harbored superantigen genes, five on average, and this strictly correlated with their ability to stimulate T-cell proliferation. In contrast, the
set
gene cluster was detected in every
S. aureus
strain, suggesting a nonredundant function for these genes which is different from T-cell activation. No more than 10% of normal human serum samples inhibited the T-cell stimulation elicited by
egc
-encoded enterotoxins (staphylococcal enterotoxins G, I, M, N, and O), whereas between 32 and 86% neutralized the classical superantigens. Similarly, intravenous human immunoglobulin G preparations inhibited
egc
-encoded superantigens with 10- to 100-fold-reduced potency compared with the classical enterotoxins. Thus, there are surprisingly large gaps in the capacity of human serum samples to neutralize
S. aureus
superantigens.
Nasal Delivery of Antigen with the B Subunit ofEscherichia coliHeat-Labile Enterotoxin Augments Antigen-Specific T-Cell Clonal Expansion and Differentiation
Authors: Apostolaki Maria , Williams Neil A.
Published: June 2004

Escherichia coliheat-labile enterotoxin has unique immunogenic and adjuvant properties when administered mucosally to mice. These properties have revealed the potential for its use in the development of mucosal vaccines, an area of increasing interest. However, the inherent toxicity mediated by the A subunit precludes its widespread use. This problem has led to attempts to dissociate toxicity from adjuvant function by use of the B subunit. The ability of the B subunit ofE. coliheat-labile enterotoxin (EtxB) to enhance responses against antigens coadministered intranasally is demonstrated here with the use of the DO11.10 adoptive-transfer model, in which ovalbumin (OVA)-specific adoptively transferred T cells can be monitored directly by flow cytometry. Intranasal delivery of OVA with EtxB resulted in increased T-cell proliferative and systemic antibody responses against antigens. The increased Th2 cytokine production detected following in vitro restimulation of splenocyte and cervical lymph node (CLN) cells from the immunized mice correlated with increased OVA-specific immunoglobulin G1 antibody production. Flow cytometric analysis of T cells from mice early after immunization directly revealed the ability of EtxB to support antigen-specific clonal expansion and differentiation. Furthermore, while responses were first detected in the CLNs, they rapidly progressed to the spleen, where they were further sustained. Examination of CD69 expression on dividing cells supported the notion that activation induced by the presence of antigens is not sufficient to drive T-cell differentiation. Furthermore, a lack of CD25 expression on dividing cells suggested that EtxB-mediated T-cell clonal expansion may occur without a sustained requirement for interleukin 2.
Description of the Transcriptomes of Immune Response-Activated Hemocytes from the Mosquito Vectors
Aedes aegypti
and
Armigeres subalbatus
Authors: Bartholomay Lyric C. , Cho Wen-Long , Rocheleau Thomas A. , Boyle Jon P. , Beck Eric T. , Fuchs Jeremy F. , Liss Paul , Rusch Michael , Butler Katherine M. , Wu Roy Chen-Chih , Lin Shih-Pei , Kuo Hang-Yen , Tsao I-Yu , Huang Chiung-Yin , Liu Tze-Tze , Hsiao Kwang-Jen , Tsai Shih-Feng , Yang Ueng-Cheng , Nappi Anthony J. , Perna Nicole T. , Chen Chen-Cheng , Christensen Bruce M.
Published: June 2004

Mosquito-borne diseases, including dengue, malaria, and lymphatic filariasis, exact a devastating toll on global health and economics, killing or debilitating millions every year (54). Mosquito innate immune responses are at the forefront of concerted research efforts aimed at defining potential target genes that could be manipulated to engineer pathogen resistance in vector populations. We aimed to describe the pivotal role that circulating blood cells (called hemocytes) play in immunity by generating a total of 11,952
Aedes aegypti
and 12,790
Armigeres subalbatus
expressed sequence tag (EST) sequences from immune response-activated hemocyte libraries. These ESTs collapsed into 2,686 and 2,107 EST clusters, respectively. The clusters were used to adapt the web-based interface for annotating bacterial genomes called A Systematic Annotation Package for Community Analysis of Genomes (ASAP) for analysis of ESTs. Each cluster was categorically characterized and annotated in ASAP based on sequence similarity to five sequence databases. The sequence data and annotations can be viewed in ASAP at
https://asap.ahabs.wisc.edu/annotation/php/ASAP1.htm
. The data presented here represent the results of the first high-throughput in vivo analysis of the transcriptome of immunocytes from an invertebrate. Among the sequences are those for numerous immunity-related genes, many of which parallel those employed in vertebrate innate immunity, that have never been described for these mosquitoes.
Flagella and Chemotaxis Are Required for Efficient Induction ofSalmonella entericaSerovar Typhimurium Colitis in Streptomycin-Pretreated Mice
Authors: Stecher Bärbel , Hapfelmeier Siegfried , Müller Catherine , Kremer Marcus , Stallmach Thomas , Hardt Wolf-Dietrich
Published: June 2004

Salmonella entericasubspecies 1 serovar Typhimurium is a common cause of gastrointestinal infections. The host's innate immune system and a complex set ofSalmonellavirulence factors are thought to contribute to enteric disease. The serovar Typhimurium virulence factors have been studied extensively by using tissue culture assays, and bovine infection models have been used to verify the role of these factors in enterocolitis. Streptomycin-pretreated mice provide an alternative animal model to study enteric salmonellosis. In this model, theSalmonellapathogenicity island 1 type III secretion system has a key virulence function. Nothing is known about the role of other virulence factors. We investigated the role of flagella in murine serovar Typhimurium colitis. A nonflagellated serovar Typhimurium mutant (fliGHI) efficiently colonized the intestine but caused little colitis during the early phase of infection (10 and 24 h postinfection). In competition assays with differentially labeled strains, thefliGHImutant had a reduced capacity to get near the intestinal epithelium, as determined by fluorescence microscopy. A flagellated but nonchemotacticcheYmutant had the same virulence defects as thefliGHImutant for causing colitis. In competitive infections, both mutants colonized the intestine of streptomycin-pretreated mice by day 1 postinfection but were outcompeted by the wild-type strain by day 3 postinfection. Together, these data demonstrate that flagella are required for efficient colonization and induction of colitis in streptomycin-pretreated mice. This effect is mostly attributable to chemotaxis. Recognition of flagellar subunits (i.e., flagellin) by innate immune receptors (i.e., Toll-like receptor 5) may be less important.
Tritrichomonas foetusInduces Apoptotic Cell Death in Bovine Vaginal Epithelial Cells
Authors: Singh B. N. , Lucas J. J. , Hayes G. R. , Kumar Ish , Beach D. H. , Frajblat Marcel , Gilbert R. O. , Sommer U. , Costello C. E.
Published: June 2004

Tritrichomonas foetusis a serious veterinary pathogen, causing bovine trichomoniasis, a sexually transmitted disease leading to infertility and abortion.T. foetusinfects the mucosal surfaces of the reproductive tract. Infection withT. foetusleads to apoptotic cell death of bovine vaginal epithelial cells (BVECs) in culture. An affinity-purified cysteine protease (CP) fraction yielding on sodium dodecyl sulfate-polyacrylamide gel electrophoresis a single band with an apparent molecular mass of 30 kDa (CP30) also induces BVEC apoptosis. Treatment of CP30 with the protease inhibitors TLCK (Nα-p-tosyl-l-lysine chloromethyl ketone) and E-64 [l-trans-epoxysuccinyl-leucylamide-(4-guanido)-butane] greatly reduces induction of BVEC apoptosis. Matrix-assisted laser desorption ionization-time-of-flight MALDI-TOF mass spectrometry analysis of CP30 reveals a single peak with a molecular mass of 23.7 kDa. Mass spectral peptide sequence analysis of proteolytically digested CP30 reveals homologies to a previously reported cDNA clone, CP8 (D. J. Mallinson, J. Livingstone, K. M. Appleton, S. J. Lees, G. H. Coombs, and M. J. North, Microbiology141:3077-3085, 1995). Induction of apoptosis is highly species specific, since the related human parasiteTrichomonas vaginalisand associated purified CPs did not induce BVEC death. Fluorescence microscopy along with the Cell Death Detection ELISAPLUSassay and flow cytometry analyses were used to detect apoptotic nuclear condensation, DNA fragmentation, and changes in plasma membrane asymmetry in host cells undergoing apoptosis in response toT. foetusinfection or incubation with CP30. Additionally, the activation of caspase-3 and inhibition of cell death by caspase inhibitors indicates that caspases are involved in BVEC apoptosis. These results imply that apoptosis is involved in the pathogenesis ofT. foetusinfection in vivo, which may have important implications for therapeutic interference with host cell death that could alter the course of the pathology in vivo.
Responses of Well-Differentiated Airway Epithelial Cell Cultures from Healthy Donors and Patients with Cystic Fibrosis to
Burkholderia cenocepacia
Infection
Authors: Sajjan Umadevi , Keshavjee Shaf , Forstner Janet
Published: June 2004

Well-differentiated cultures established from airway epithelia of patients with cystic fibrosis (CF cultures) exhibited goblet cell hyperplasia, increased secretion of mucus, and higher basal levels of interleukin-8 than similarly cultured cells from healthy donors. Upon apical infection with low doses (10
4
to 10
5
CFU) of
Burkholderia cenocepacia
isolate BC7, the two cultures gave different responses. While normal cultures trapped the added bacteria in the mucus layer, killed and/or inhibited bacterial replication, and prevented bacterial invasion of the cells, CF cultures failed to kill and/or supported the growth of bacteria, leading to invasion of underlying epithelial cells, compromised transepithelial permeability, and cell damage. Depletion of the surface mucus layer prior to bacterial infection rendered the normal cultures susceptible to bacterial invasion, but the invading bacteria were mainly confined to vacuoles within the cells and appeared to be nonviable. In contrast, bacteria that invaded cells in CF cultures were found free in the cytoplasm surrounded by intermediate filaments and also between cells. Cultured CF airway epithelium was therefore more susceptible to infection than normal epithelium. This mimics CF tissue in vivo and illustrates differences in the way epithelia in CF patients and normal subjects handle bacterial infection. In addition, we found that the CF and normal cell cultures responded differently not only to isolate BC7 but also to isolates of other
B. cepacia
complex species. We therefore conclude that this cell culture model is suitable for investigation of
B. cepacia
complex pathogenesis in CF patients.
New Murine Model for the Study of
Chlamydia trachomatis
Genitourinary Tract Infections in Males
Authors: Pal Sukumar , Peterson Ellena M. , de la Maza Luis M.
Published: June 2004

The lack of an experimental model has significantly limited the understanding of the pathogenesis of
Chlamydia trachomatis
infections in males. In an attempt to establish a model using the natural route of infection, we inoculated male mice in the meatus urethra. To establish the 50% infectious dose (ID
50
), C3H/HeN (

H-2
k

) male mice were inoculated in the meatus urethra with doses ranging from 10
1
to 10
7
inclusion-forming units (IFU) of
C. trachomatis
mouse pneumonitis biovar (MoPn) and were euthanized at 10 days postinfection (p.i.). Approximately 50% of the animals inoculated with 5 × 10
4
IFU had positive cultures of the urethra, urinary bladder, epididymides, and/or testes. Subsequently, to characterize the course of the infection, a group of animals was inoculated with 10
6
IFU/mouse (20 times the ID
50
). Positive cultures from the urethra, urinary bladder, epididymides, and testes were obtained from the animals. The infection peaked in the first 2 weeks p.i. and subsequently declined over the 7 weeks of observation.
C. trachomatis
-specific antibodies were first detected in serum by 2 weeks p.i. and rose over the period of observation. The titers of immunoglobulin G2a (IgG2a) were 16-fold higher than those of IgG1. A lymphoproliferative assay using splenocytes and local lymph nodes showed a strong cell-mediated immune response. Levels of gamma interferon were significantly higher than those of interleukin-4 in the supernatants from stimulated lymphocytes. An acute inflammatory infiltrate consisting of polymorphonuclear leukocytes was detected in the urethra at 1 week p.i. At 3 weeks p.i., a mixed acute and chronic inflammatory infiltrate was observed in the urethra that by 5 to 6 weeks was mainly composed of mononuclear cells. Similar findings were also observed in the urinary bladder, although the inflammatory infiltrate was delayed by approximately a week relative to that in the urethra. Sections of the epididymides showed a focal acute inflammatory infiltrate at 2 weeks p.i. Immunohistochemical staining demonstrated multiple chlamydial inclusions in the epithelium of the urethra and urinary bladder. No chlamydial inclusions were observed in the epididymides or testes. In conclusion, inoculation of male mice in the meatus urethra with
C. trachomati
s MoPn results in an infection of the genitourinary tract that closely parallels that described in humans. This model should help to characterize the pathogenesis of chlamydial infections in males and to test therapeutic and preventive measures.
The
galU
Gene of
Pseudomonas aeruginosa
Is Required for Corneal Infection and Efficient Systemic Spread following Pneumonia but Not for Infection Confined to the Lung
Authors: Priebe Gregory P. , Dean Charles R. , Zaidi Tanweer , Meluleni Gloria J. , Coleman Fadie T. , Coutinho Yamara S. , Noto Michael J. , Urban Teresa A. , Pier Gerald B. , Goldberg Joanna B.
Published: June 2004

Acute pneumonias and corneal infections due to
Pseudomonas aeruginosa
are typically caused by lipopolysaccharide (LPS)-smooth strains. In cystic fibrosis patients, however, LPS-rough strains of
P. aeruginosa
, which lack O antigen, can survive in the lung and cause chronic infection. It is not clear whether an LPS-rough phenotype affects cytotoxicity related to the type III secretion system (TTSS). We previously reported that interruption of the
galU
gene in
P. aeruginosa
results in production of a rough LPS and truncated LPS core. Here we evaluated the role of the
galU
gene in the pathogenesis of murine lung and eye infections and in cytotoxicity due to the TTSS effector ExoU. We studied
galU
mutants of strain PAO1, of its cytotoxic variant expressing ExoU from a plasmid, and of the inherently cytotoxic strain PA103. The
galU
mutants were more serum sensitive than the parental strains but remained cytotoxic in vitro. In a corneal infection model, the
galU
mutants were significantly attenuated. In an acute pneumonia model, the 50% lethal doses of the
galU
mutants were higher than those of the corresponding wild-type strains, yet these mutants could cause mortality and severe pneumonia, as judged by histology, even with minimal systemic spread. These findings suggest that the
galU
gene is required for corneal infection and for efficient systemic spread following lung infection but is not required for infection confined to the lung. Host defenses in the lung appear to be insufficient to control infection with LPS-rough
P. aeruginosa
when local bacterial levels are high.
Pseudomonas aeruginosa
Pyocyanin Is Critical for Lung Infection in Mice
Authors: Lau Gee W. , Ran Huimin , Kong Fansheng , Hassett Daniel J. , Mavrodi Dimitri
Published: June 2004

Pseudomonas aeruginosa
secretes copious amounts of the redox-active phenazine, pyocyanin (PCN), during cystic fibrosis lung infection. PCN has been shown to interfere with a variety of cellular processes in cultured lung epithelial cells. Here, by using two respiratory tract models of infection, we demonstrate that PCN mediates tissue damage and necrosis during lung infection.
Increased Susceptibility of Secretor Factor Gene
Fut2
-Null Mice to Experimental Vaginal Candidiasis
Authors: Hurd Elizabeth A. , Domino Steven E.
Published: June 2004

Fut2-LacZ-null mice, which are a model of the human ABO and Lewis nonsecretor group, display increased susceptibility to experimental yeast vaginitis, indicating a role for α(1,2)fucosylated cervical glycans in mucosal defense. However, the lack of significant effect of competitive inhibition by exogenous neoglycoproteins in this study emphasizes the complexity of
Candida
-epithelial cell adhesion events.
Multiserotype Protection of Mice against Pneumococcal Colonization of the Nasopharynx and Middle Ear by Killed Nonencapsulated Cells Given Intranasally with a Nontoxic Adjuvant
Authors: Malley Richard , Morse Sarah C. , Leite Luciana C. C. , Areas Ana Paula Mattos , Ho Paulo Lee , Kubrusly Flavia S. , Almeida Igor C. , Anderson Porter
Published: June 2004

Intranasal challenge of C57BL/6 mice with
Streptococcus pneumoniae
serotypes 6B, 14, and 23F produced colonization of the middle ear and NP. Intranasal vaccination with ethanol-killed nonencapsulated cells with adjuvant protected both sites. Of four nontoxic adjuvants tested, the cholera toxin B subunit was most effective and least nonspecifically protective.
Expression of a C Terminally Truncated Form of Pertussis Toxin S1 Subunit Effectively Induces Protection against Pertussis Toxin following DNA-Based Immunization
Authors: Kamachi Kazunari , Arakawa Yoshichika
Published: June 2004

Four plasmids encoding different C terminally and N terminally truncated pertussis toxin S1 subunits of
Bordetella pertussis
were constructed and tested for inducibility of protection against pertussis toxin in mice after DNA-based immunization. The region encoding an N-terminal 180-amino-acid fragment of the S1 subunit had the most potent ability to induce protective immunity.
Oral Immunization with an
rfaH
Mutant Elicits Protection against Salmonellosis in Mice
Authors: Nagy Gábor , Dobrindt Ulrich , Hacker Jörg , Emödy Levente
Published: June 2004

Loss of the transcriptional antiterminator RfaH results in virulence attenuation (>10
4
-fold increase in 50% lethal dose) of the archetypal
Salmonella enterica
serovar Typhimurium strain SL1344 by both orogastric and intraperitoneal routes of infection in BALB/c mice. Oral immunization with the mutant efficiently protects mice against a subsequent oral infection with the wild-type strain. Interestingly, in vitro immunoreactivity is not confined to strain SL1344; rather, it is directed also towards other serovars of
S. enterica
and even
Salmonella bongori
strains.
Partial safety factors for designing and assessing flexible pavement performance
Authors: Wang S S , Hong H P
Published: June 2004

In designing and assessing pavement performance, the uncertainty in material properties and geometrical variables of pavement and in traffic and environmental actions should be considered. A single factor is employed to deal with these uncertainties in the current American Association of State Highway and Transportation Officials (AASHTO) guide for design of pavements. However, use of this single factor may not ensure reliability-consistent pavement design and assessment because different random variables that may have different degrees of uncertainty affect the safety and performance of pavement differently. Similar problems associated with structural design have been recognized by code writers and dealt with using partial safety factors or load resistance factors. The present study is focused on evaluating a set of partial safety factors to be used in conjunction with the flexible pavement deterioration model in the Ontario pavement analysis of cost and the model in the AASHTO guide for evaluating the flexible pavement performance or serviceability. Evaluation and probabilistic analyses are carried out using the first-order reliability method and simple simulation technique. The results of the analysis were used to suggest factors that could be used, in a partial safety factor format, for designing or assessing flexible pavement conditions to achieve a specified target safety level.Key words: deterioration, reliability, pavement, serviceability, stochastic process, performance, partial safety factor.
Analysis and design of laterally unsupported portal frames for out-of-plane stability
Authors: Zinoviev Ilian , Mohareb Magdi
Published: June 2004

A methodology for the analysis and design of laterally unsupported portal frames is proposed. A finite element model is developed to predict the elastic critical load and associated buckling mode. Regression analysis is then conducted to find lateral displacement and rotation field expressions that closely approximate the buckled configurations predicted by the finite element analysis. The obtained functions are then substituted into the total potential energy expression, and the stationarity conditions are evoked. The resulting eigen-value problem is solved for the out-of-plane buckling loads that are then compared with those based on the finite element model. The agreement between the two solutions provides an indication of the accuracy of the simplified energy solution. The member destabilizing effects induced by axial forces are separated from those induced by strong axis bending. The separation of these two effects is subsequently exploited in a two-step eigen-value procedure, aimed at determining the key member resistances defined in the interaction check of the standard CSA-S16-01, while accurately modeling the boundary conditions of the member. These are (i) compressive resistance of the member in the absence of bending effects and (ii) flexural resistance of the member in the absence of axial force effects.Key words: portal frames, lateral buckling, finite element analysis, wide flange sections, frame design, principle of stationary potential energy.
Economic gains around mergers and acquisitions in the construction industry of the United States of America
Authors: Choi Jongsoo , Russell Jeffrey S
Published: June 2004

As waves of mergers and acquisitions (M&A) have swept over American industrial business organizations, construction firms have been caught in the middle of the resulting turbulence. Nonetheless, no research has investigated these significant events in the construction industry. Built upon the financial theories and methodology, the overall success level of construction M&A transactions was assessed. The research findings, which were drawn from an analysis of 171 construction M&A transactions, indicate that the performance of construction M&A was positive at an insignificant level, as measured by equity market returns. Whereas the relationship between the type of diversification strategy and performance indicates that while the related diversification strategy has been slightly favored by both theories and empirical research findings over unrelated diversification, no significant performance difference was observed between two diversification strategies.Key words: mergers and acquisitions, diversification strategy, equity market returns.
Dynamic performance of the Confederation Bridge due to traffic and wind
Authors: Naumoski Nove , Cheung Moe S , Foo Simon
Published: June 2004

This paper describes results from a study of the dynamic performance of the Confederation Bridge during traffic load tests and a storm with high winds that occurred at the bridge site on 7 November 2001. The traffic load tests were conducted over a three-span segment of the bridge and consisted of single and double truck configurations at different speeds. The winds during the storm of 7 November 2001 were the strongest that have ever been measured at the bridge site since the opening of the bridge. The vibrations of the bridge during both the load tests and the storm were recorded by a network of 76 accelerometers. By processing the recorded vibrations, vertical deflections and dynamic amplification factors due to the traffic load tests and vertical and lateral deflections due to the wind during the storm were computed. The computed values were compared with those used in the design. It was found that both the deflections and the dynamic amplification factors were much smaller than the design values.Key words: bridge, acceleration, deflection, record, vibration, filtering, frequency, mode, Fourier spectrum.
Caractérisation d'une maçonnerie composée d'éléments en aggloméré de chanvre
Authors: Bütschi Pierre-Yves , Deschenaux Christian , Miao Buquan , Srivastava Narenda K
Published: June 2004

The tested masonry elements, made of an agglomeration of industrial hemp and inorganic binders, were prefabricated on a cement brick production with an objective to use them for light-weight load-bearing wall construction and for insulation purposes. The tests indicate that the load-bearing characteristics of such masonry construction is close to the present Swiss recommendations. Furthermore, the sound insulation characteristics are comparable to the numerous other traditional systems, but they are still not competitive compared to other existing systems from the point of view of thermal insulation.Key words: construction, brick, light weight, building materials, industrial hemp, mechanical properties, physical properties, sustainable development.
Endophytic bacterial communities of field-grown potato plants and their plant-growth-promoting and antagonistic abilities
Authors: Sessitsch Angela , Reiter Birgit , Berg Gabriele
Published: June 2004

To study the effect of plant growth on potato-associated bacteria, the composition and properties of bacteria colonizing the endosphere of field-grown potato were analyzed by a multiphasic approach. The occurrence and diversity of potato-associated bacteria were monitored by a cultivation-independent approach, using terminal restriction fragment length polymorphism analysis of 16S rDNA. The patterns obtained revealed a high heterogeneity of community composition and suggested the existence of plant-specific communities. However, endophytic populations correlated to a certain extent with plant growth performance. Endophytes were also isolated from plants that grew well or grew poorly and were identified by partial sequencing of the 16S rRNA genes. A broad phylogenetic spectrum was found among isolates and differently growing plants hosted different bacterial populations. In an approach to investigate the plant-growth-promoting potential of potato-associated bacteria, a total of 35 bacteria were screened by dual testing for in vitro antagonism towards (i) the fungal pathogens Verticillium dahliae, Rhizoctonia solani, Sclerotinia sclerotiorum, and Phytophthora cactorum and (ii) the bacterial pathogens Erwinia carotovora, Streptomyces scabies, and Xanthomonas campestris. The proportion of isolates with antagonistic activity was highest against Streptomyces sp. (43%) followed by those against Xanthomonas sp. (29%). As all plants showed more or less severe disease symptoms of scab disease caused by Streptomyces scabies, we assume that the presence of the pathogen induced the colonization of antagonists. The antifungal activity of the isolates was generally low. The biotechnological potential of endophytic isolates assessed by their antagonistic activity and by in vitro production of enzymes, antibiotics, siderophores, and the plant growth hormone indole-1,3-acetic acid was generally high. Overall, seven endophytes were found to antagonize fungal as well as bacterial pathogens and showed a high production of active compounds and were therefore considered promising biological control agents.Key words: T-RFLP, 16S rRNA, siderophores, IAA, biocontrol.
Nitrogen transfer and assimilation between the arbuscular mycorrhizal fungus Glomus intraradices Schenck & Smith and Ri T-DNA roots of Daucus carota L. in an in vitro compartmented system
Authors: Toussaint Jean-Patrick , St-Arnaud Marc , Charest Christiane
Published: June 2004

Nitrogen metabolism was examined in monoxenic cultures of carrot roots (Daucus carota L.) colonized with the arbuscular mycorrhizal (AM) fungus Glomus intraradices Schenck & Smith. Glutamine synthetase and glutamate dehydrogenase activities were significantly increased in mycorrhizal roots for which only the extraradical mycelium had exclusive access to NH4NO3 in a distinct hyphal compartment inaccessible to the roots. This was in comparison with the water controls but was similar to the enzyme activities of non-arbuscular-mycorrhizal (non-AM) roots that had direct access to NH4NO3. In addition, glutamate dehydrogenase activity was significantly enhanced in AM roots compared with non-AM roots. Carrot roots took up 15NH4+ more efficiently than 15NO3–, and the extraradical hyphae transfered 15NH4+ to host roots from the hyphal compartment but did not transfer 15NO3–. The extraradical mycelium was shown, for the first time, to have a different glutamine synthetase monomer than roots. Our overall results highlight the active role of AM fungi in nitrogen uptake, transfer, and assimilation in their symbiotic root association.Key words: arbuscular mycorrhizal fungus, Ri T-DNA carrot roots, in vitro root-organ culture, nitrogen metabolism.
Influence des conditions de culture préalables et de la présence du gènerpoSpour la survie deSalmonella typhimuriumen eau de mer exposée à la lumière solaire
Authors: Maatouk K , Zaafrane S , Gauthier J M , Bakhrouf A
Published: June 2004

The effect of sunlight exposure on Salmonella typhimurium isogenic strains harboring an rpoS gene functional (rpoS+) or not functional (rpoS) was investigated in microcosms of sterile sea water at 20 °C. The two strains rapidly lost their ability to produce colonies on solid culture media. The detrimental action of sunlight was more important when the salinity of sea water increased. The survival of stationary phase cells was influenced by RpoS. Bacteria grown in media with high salinity or osmolarity and transferred to sea water in stationary phase were more resistant to irradiation than those grown in media with low salinity. Prior growth under oxidative (0.2 mmol/L of H2O2) or amino acid starved (minimal medium) conditions did not modify the survival of either strain when they were exposed to sunlight. Bacteria were more resistant when cells were incubated in sea water in the dark prior to being exposed to sunlight. The resistance to sunlight irradiation was also greater in clones of both strains isolated from microcosms exposed to sunlight for 90 min, then further inoculated into sea water and reexposed to sunlight.Key words: Salmonella typhimurium, sea water, sunlight, rpoS, σs.
Influences of σBandagron expression of staphylococcal enterotoxin B (seb) inStaphylococcus aureus
Authors: Schmidt Katherine A , Donegan Niles P , Kwan, Jr. William A , Cheung Ambrose
Published: June 2004

In Staphylococcus aureus, enterotoxin B (SEB) is a superantigen that activates host interleukins and induces adverse responses, ranging from food poisoning to toxic shock. The alternate sigma factor, σB(SigmaB), and agr are two known regulators of S. aureus. Northern blots of strain COL, a sigB-positive strain, showed an inverse correlation between σBexpression and seb message. seb expression was also measured as a function of a seb promoter linked to green fluorescent protein (GFP) expression in RN6390, COL, and Newman. In sigB mutants of RN6390, SH1000, COL, and Newman, seb promoter activities, as measured by GFP expression, increased relative to the respective parental types but at differing levels, suggesting alternate strain-specific regulation. In agr mutants of RN6390 and Newman, seb promoter activities were intermediate between the high level seen for the sigB mutant and the low level in the sigB active strains. A sigB agr double mutant of RN6390 displayed lower GFP expression than the agr mutant. These results suggest that while σBand agr regulate seb expression in a divergent manner, other activator(s) of seb that depend on sigB expression may be present in S. aureus.Key words: staphylococcal enterotoxin B, σBregulation, agr regulation.