bluuebunny/crossref_metadata_2025 huggingface dataset:
Trypanosoma cruziInfection Activates Extracellular Signal-Regulated Kinase in Cultured Endothelial and Smooth Muscle Cells
Authors: Mukherjee Shankar , Huang Huan , Petkova Stefka B. , Albanese Chris , Pestell Richard G. , Braunstein Vicki L. , Christ George J. , Wittner Murray , Lisanti Michael P. , Berman Joan W. , Weiss Louis M. , Tanowitz Herbert B.
Published: August 2004

Trypanosoma cruziinfection causes cardiomyopathy and vasculopathy. We examined the consequence of this infection for the mitogen-activated protein kinase (MAPK) pathways, which regulate cell proliferation in cultured human umbilical vein endothelial and vascular smooth muscle cells. Infection of these cells resulted in activation of extracellular signal-regulated kinases 1and 2 (ERK1/2) but not c-Jun N-terminal kinase or p38 MAPK. Treatment of these cells with the MAPK kinase inhibitor PD98059 prior to infection blocked the increase in phosphorylated ERK1/2 seen with infection. Heat-killed parasites did not activate ERK1/2, indicating that activation of ERK1/2 was dependent on infection of these cells by live parasites. Furthermore, transfection with dominant-negative Raf(301) or Ras(N17) constructs reduced the infection-associated levels of phospho-ERK1/2, indicating that the activation of ERK1/2 involved the Ras-Raf-ERK pathway. Infection also resulted in an increase in activator protein 1 (AP-1) activity, which was inhibited by transfection with a dominant-negative Raf(301) construct.T. cruzi-infected endothelial cells secreted endothelin-1 and interleukin-1β, which activated ERK1/2 and induced cyclin D1 expression in uninfected smooth muscle cells. These data suggest a possible molecular paradigm for the pathogenesis of the vasculopathy and the cardiovascular remodeling associated withT. cruziinfection.
Monocytes from Patients with Indeterminate and Cardiac Forms of Chagas' Disease Display Distinct Phenotypic and Functional Characteristics Associated with Morbidity
Authors: Souza Paulo E. A. , Rocha Manoel O. C. , Rocha-Vieira Etel , Menezes Cristiane A. S. , Chaves Andréa C. L. , Gollob Kenneth J. , Dutra Walderez O.
Published: August 2004

Many studies have demonstrated that monocyte-derived macrophages display critical activities in immunity to parasites. The ability of these cells to process and present antigens, produce cytokines, and provide costimulatory signals demonstrates their pivotal role in initiating immune responses. Although potential modulatory function has been attributed to monocytes from patients with Chagas' disease, a systematic phenotypic and functional analysis of these cells has not been performed. In this work, we analyzed the ex vivo expression of important surface molecules (CD11b and HLA-DR) and immunoregulatory cytokines (interleukin-10 [IL-10], IL-12 and tumor necrosis factor alpha [TNF-α]) in CD14+and CD14−monocytes from Chagas' disease patients with polar clinical forms of the disease: indeterminate or severe cardiac. We also evaluated the influence of in vitro infection withT. cruziin the expression of such molecules. We observed that monocytes from indeterminate-disease patients display lower levels of HLA-DR than those from noninfected individuals both ex vivo and after in vitro infection withT. cruzi.Although ex vivo expression of CD11b was similar among the groups, in vitro infection led to decreased expression of this molecule by monocytes from Chagas' disease patients but not from noninfected individuals. Analysis of the expression of immunoregulatory cytokines showed that while monocytes from indeterminate-disease patients are committed to IL-10 expression, a higher percentage of monocytes from cardiac-disease patients express TNF-α after exposure to live parasites. These results suggest that monocytes from indeterminate-disease patients display modulatory characteristics related to low HLA-DR and high IL-10 expression whereas monocytes from cardiac-disease, patients may be committed to induction of inflammatory responses related to high TNF-α expression.
Induction of Protection against Porcine Cysticercosis by Vaccination with Recombinant Oncosphere Antigens
Authors: Flisser Ana , Gauci Charles G. , Zoli André , Martinez-Ocaña Joel , Garza-Rodriguez Adriana , Dominguez-Alpizar Jose Luis , Maravilla Pablo , Rodriguez-Canul Rossana , Avila Guillermina , Aguilar-Vega Laura , Kyngdon Craig , Geerts Stanny , Lightowlers Marshall W.
Published: August 2004

Two recombinant
Taenia solium
oncosphere antigens, designated TSOL18 and TSOL45-1A, were investigated as vaccines to prevent transmission of the zoonotic disease cysticercosis through pigs. Both antigens were effective in inducing very high levels of protection (up to 100%) in three independent vaccine trials in pigs against experimental challenge infection with
T. solium
eggs, which were undertaken in Mexico and Cameroon. This is the highest level of protection that has been achieved against
T. solium
infection in pigs by vaccination with a defined antigen. TSOL18 and TSOL45-1A provide the basis for development of a highly effective practical vaccine that could assist in the control and, potentially, the eradication of human neurocysticercosis.
A
Salmonella fim
Homologue in
Citrobacter freundii
Mediates Invasion In Vitro and Crossing of the Blood-Brain Barrier in the Rat Pup Model
Authors: Hess Petra , Altenhöfer Artur , Khan A. Salam , Daryab Neda , Kim Kwang Sik , Hacker Jörg , Oelschlaeger Tobias A.
Published: August 2004

From the invasive
Citrobacter freundii
strain 3009, an invasion determinant was cloned, sequenced, and expressed. Sequence analysis of the determinant showed high homology with the
fim
determinant from
Salmonella enterica
serovar Typhimurium. The genes of the invasion determinant directed invasion of recombinant
Escherichia coli
K-12 strains into human epithelial cell lines of the bladder and gut as well as mannose-sensitive yeast agglutination and were termed
fim
Cf
genes. Expression of the Fim
Cf
proteins was shown by
35
S labeling and/or Western blotting. In the infant rat model of experimental hematogenous meningitis,
C. freundii
strain 3009 and the in vitro invasive recombinant
E. coli
K-12 strain harboring the
fim
Cf
determinant reached the cerebrospinal fluid, in contrast to the case for the control strain. The
fim
determinant was also necessary for efficient in vitro invasion by
C. freundii
, because a deletion mutant was strongly reduced in its invasion efficiency. The mutation could be complemented in
trans
by the corresponding genes. Invasion by
C. freundii
could be blocked only by
d
-mannose, GlcNAc, and chitin hydrolysate and not by other carbohydrates tested. In contrast, yeast agglutination was not affected by GlcNAc or chitin hydrolysate. This finding indicated mannose residues to be essential for both yeast agglutination and invasion, whereas GlcNAc (oligomer) residues of host cells are involved exclusively in invasion. These results showed the
fim
determinant of
C. freundii
to be responsible for
d
-mannose- and GlcNAc-dependent in vitro invasion without being assembled into pili and for crossing of the blood-brain barrier in the infant rat model.
Live
Lactobacillus reuteri
Is Essential for the Inhibitory Effect on Tumor Necrosis Factor Alpha-Induced Interleukin-8 Expression
Authors: Ma Donglai , Forsythe Paul , Bienenstock John
Published: August 2004

The mechanism of the apparent anti-inflammatory action of probiotic organisms is unclear.
Lactobacillus reuteri
is effective in inhibiting colitis in interleukin-10 (IL-10)-deficient mice. Nerve growth factor (NGF), in addition to its activity on neuronal cell growth, has significant anti-inflammatory effects in several experimental systems in vitro and in vivo, including a model of colitis. Our experiments were designed to explore the mechanism of effect of
L. reuteri
in the human epithelial cell lines T84 and HT29 on cytokine and NGF synthesis and IL-8 response to tumor necrosis factor alpha (TNF-α). Epithelial cells were cultured for various times with live and killed
L. reuteri
and examined by reverse transcription-PCR for NGF, IL-10, and TNF-α-induced IL-8 expression. An enzyme-linked immunosorbent assay was used to quantitate intracellular IL-8 and secreted product. Western blotting and confocal microscopy were used to determine the effects on IκB and NF-κB, respectively. Live but not heat-killed or gamma-irradiated
L. reuteri
upregulated NGF and dose dependently inhibited constitutive synthesis by T84 and HT29 cells of IL-8 and that induced by TNF-α in terms of mRNA and intracellular and secreted protein. Similarly,
L. reuteri
inhibited IL-8 synthesis induced by
Salmonella enterica
serovar Typhimurium.
L. reuteri
required preincubation and adherence for effect, inhibited translocation of NF-κB to the nuclei of HeLa cells, and prevented degradation of IκB. Neither cellular lysates nor media supernatants had any effect on TNF-α-induced IL-8. The conclusion is that
L. reuteri
has potent direct anti-inflammatory activity on human epithelial cells, which is likely to be related to the activity of ingested probiotics.
L. reuteri
also upregulates an unusual anti-inflammatory molecule, NGF, and inhibits NF-κB translocation to the nucleus.
Identification ofMycobacterium tuberculosisCounterimmune (cim) Mutants in Immunodeficient Mice by Differential Screening
Authors: Hisert Katherine B. , Kirksey Meghan A. , Gomez James E. , Sousa Alexandra O. , Cox Jeffery S. , Jacobs William R. , Nathan Carl F. , McKinney John D.
Published: August 2004

Tuberculosis (TB) is characterized by lifetime persistence ofMycobacterium tuberculosis. Despite the induction of a vigorous host immune response that curtails disease progression in the majority of cases, the organism is not eliminated. Subsequent immunosuppression can lead to reactivation after a prolonged period of clinical latency. Thus, while it is clear that protective immune mechanisms are engaged duringM. tuberculosisinfection, it also appears that the pathogen has evolved effective countermechanisms. Genetic studies with animal infection models and with patients have revealed a key role for the cytokine gamma interferon (IFN-γ) in resistance to TB. IFN-γ activates a large number of antimicrobial pathways. Three of these IFN-γ-dependent mechanisms have been implicated in defense againstM. tuberculosis: inducible nitric oxide synthase (iNOS), phagosome oxidase (phox), and the phagosome-associated GTPase LRG-47. In order to identify bacterial genes that provide protection against specific host immune pathways, we have developed the strategy of differential signature-tagged transposon mutagenesis. Using this approach we have identified threeM. tuberculosisgenes that are essential for progressiveM. tuberculosisgrowth and rapid lethality in iNOS-deficient mice but not in IFN-γ-deficient mice. We propose that these genes are involved in pathways that allowM. tuberculosisto counter IFN-γ-dependent immune mechanisms other than iNOS.
EncapsulatedStreptococcus suisInhibits Activation of Signaling Pathways Involved in Phagocytosis
Authors: Segura Mariela , Gottschalk Marcelo , Olivier Martin
Published: August 2004

Streptococcus suiscapsular type 2 is an important zoonotic agent of meningitis. Previous studies reported that, in contrast to nonencapsulated mutants, encapsulatedS. suisis able to resist phagocytosis. However, the mechanisms by whichS. suisavoids phagocytosis are unknown. To elucidate the signaling pathway(s) involved inS. suisantiphagocytosis, we compared the ability of an encapsulated strain and its nonencapsulated mutant to induce the activation of Akt and protein kinase C (PKC), which are downstream kinases of the phosphatidylinositol 3-kinase (PI-3K) pathway, known to be involved in the phagocytosis processes. The results demonstrated high levels of Akt and PKCα phosphorylation after infection of J774 macrophages with the nonencapsulated mutant, whereas the encapsulated strain showed reduced activation of PI-3K/Akt/PKCα signaling pathway, as well as several protein tyrosine events. These results correlated with the number of intracellular bacteria. Macrophages pretreated with specific PI-3K or PKC inhibitors showed reduced levels of Akt and PKCα phosphorylation, resulting in 50% reduction of phagocytosis. The role of phosphatases in the antiphagocytic mechanisms was evaluated by using phosphatase inhibitors, as well as SHP-1-deficient macrophages. Only in the absence of SHP-1 did the phagocytosis of encapsulatedS. suissignificantly increase, leading to Akt phosphorylation levels similar to those observed with the nonencapsulated strain, indicating activation of this important SH2 domain-containing tyrosine phosphatase by encapsulatedS. suis. Finally, when purifiedS. suiscapsular polysaccharide (CPS) was added to macrophages, no phosphorylation events were observed. In addition, CPS and encapsulatedS. suiswere able to inhibit the uptake of the nonencapsulated mutant. These results suggest the importance of CPS in the mechanisms, wherebyS. suisdownmodulates phagocytosis.
Dendritic Cells Induce Immunity and Long-Lasting Protection against Blood-Stage Malaria despite an In Vitro Parasite-Induced Maturation Defect
Authors: Pouniotis Dodie S. , Proudfoot Owen , Bogdanoska Violeta , Apostolopoulos Vasso , Fifis Theodora , Plebanski Magdalena
Published: August 2004

Dendritic cells (DC) suffer a maturation defect following interaction with erythrocytes infected with malaria parasites and become unable to induce protective malaria liver-stage immunity. Here we show that, by contrast, maturation-arrested DC in vitro are capable of the successful induction of antigen-specific gamma interferon (IFN-γ) and interleukin 4 (IL-4) T-cell responses, antibody responses, and potent protection against lethal blood-stage malaria challenge in vivo. Similar results were found with DC pulsed with intact parasitizedPlasmodium yoeliiorPlasmodium chabaudierythrocytes. Cross-strain protection was also induced. High levels of protection (80 to 100%) against lethal challenge were evident from 10 days after a single immunization and maintained up to 120 days. Interestingly, correlation studies versus blood-stage protection at different time points suggest that the immune effector mechanisms associated with protection could change over time. Antibody-independent, T-cell- and IL-12-associated protection was observed early after immunization, followed by antibody and IL-4-associated, IFN-γ-independent protection in long-term studies. These results indicate that DC, even when clearly susceptible to parasite-induced maturation defect effects in vitro, can be central to the induction of protection against blood-stage malaria in vivo.
Novel Modification of Lipid A of
Francisella tularensis
Authors: Phillips Nancy J. , Schilling Birgit , McLendon Molly K. , Apicella Michael A. , Gibson Bradford W.
Published: August 2004

We have investigated the lipid A of
Francisella tularensis
subsp.
holarctica
strain 1547-57, a type B strain, by using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry, nanoelectrospray quadrupole ion-trap mass spectrometry, and chemical methods. In accordance with the previously published structures of the lipid A from
F. tularensis
live vaccine strain (LVS) (ATCC 29684) (E. Vinogradov et al., Eur. J. Biochem.
269:
6112-6118, 2002), all of the major lipid A forms from strain 1547-57 were tetraacylated. As in the LVS strain, the major fatty acids detected in the
F. tularensis
1547-57 lipid A sample included 3-hydroxyoctadecanoic acid, 3-hydroxyhexadecanoic acid, hexadecanoic acid, and tetradecanoic acid. However, several of the lipid A components present in strain 1547-57 were of higher molecular weight than the previously published structures. A major component with an
M
r
of 1,666 was found to contain three C
18:0
(3-OH) fatty acids, one C
16:0
fatty acid, one phosphate group, and one 161-Da moiety. This 161-Da moiety could be removed from the lipid A by treatment with aqueous hydrofluoric acid and was identified as galactosamine following peracetylation and analysis by gas chromatography-mass spectrometry. Detailed investigations of the
M
r
-1,666 species by ion-trap mass spectrometry with multiple stages of fragmentation suggested that the galactosamine-1-phosphate was linked to the reducing terminus of the lipid A. Similar to the modification of lipid A with arabinosamine, lipopolysaccharide species from
F. tularensis
containing a phosphate-linked galactosamine could potentially influence its intracellular survival by conferring resistance to antimicrobial peptides.
Involvement of Raft-Like Plasma Membrane Domains of
Entamoeba histolytica
in Pinocytosis and Adhesion
Authors: Laughlin Richard C. , McGugan Glen C. , Powell Rhonda R. , Welter Brenda H. , Temesvari Lesly A.
Published: August 2004

Lipid rafts are highly ordered, cholesterol-rich, and detergent-resistant microdomains found in the plasma membrane of many eukaryotic cells. These domains play important roles in endocytosis, secretion, and adhesion in a variety of cell types. The parasitic protozoan
Entamoeba histolytica
, the causative agent of amoebic dysentery, was determined to have raft-like plasma membrane domains by use of fluorescent lipid analogs that specifically partition into raft and nonraft regions of the membrane. Disruption of raft-like membrane domains in
Entamoeba
with the cholesterol-binding agents filipin and methyl-β-cyclodextrin resulted in the inhibition of several important virulence functions, fluid-phase pinocytosis, and adhesion to host cell monolayers. However, disruption of raft-like domains did not inhibit constitutive secretion of cysteine proteases, another important virulence function of
Entamoeba
. Flotation of the cold Triton X-100-insoluble portion of membranes on sucrose gradients revealed that the heavy, intermediate, and light subunits of the galactose-
N
-acetylgalactosamine-inhibitible lectin, an important cell surface adhesion molecule of
Entamoeba
, were enriched in cholesterol-rich (raft-like) fractions, whereas EhCP5, another cell surface molecule, was not enriched in these fractions. The subunits of the lectin were also observed in high-density, actin-rich fractions of the sucrose gradient. Together, these data suggest that pinocytosis and adhesion are raft-dependent functions in this pathogen. This is the first report describing the existence and physiological relevance of raft-like membrane domains in
E. histolytica
.
Development of a Noninvasive Method for Detecting and Monitoring the Time Course of
Helicobacter pylori
Infection
Authors: Nyan Dougbeh C. , Welch Anthony R. , Dubois Andre , Coleman William G.
Published: August 2004

Helicobacter pylori
infection status following experimental inoculation of mice presently requires euthanasia. The purpose of this study was to develop a method for following the time course of
H. pylori
infection in live experimental animals. Twenty-six C57BL/6,
Helicobacter
-free female mice were inoculated with
H. pylori
Sydney strain 1, and 16 mice were sham inoculated. The mice were repeatedly tested during a period of about 1 year with an
H. pylori
species-specific primer-based PCR analysis of DNA extracted from fecal pellets of mice. The mice were euthanized at 6 months (
n
= 15) and 10 months (
n
= 15) to determine their infection status by histology, culture, and PCR of gastric specimens.
H. pylori
-inoculated mice were tested via the PCR method at 6 and 10 months prior to necropsy. Nine of 13 (69%) and 10 of 13 (77%) mice tested at 6 and 10 months, respectively, were positive. All sham-inoculated mice were negative. These two PCR results suggested a specificity of 100% with a sensitivity range between 69 and 77%. In contrast, sensitivity and specificity rose to 90 and 100% if groups of mice were tested once daily for 4 days. Seventy-seven to 85% of the experimental mice were also positive for
H. pylori
by culture. The histopathology demonstrated mild to severe gastritis. These findings demonstrate that the persistence or transience of
H. pylori
infection in live mice can be repeatedly evaluated over time. This method could allow the determination of the time course of infection and the efficacy of medications and/or vaccine without necropsy.
Analysis of Subassemblies of Pertussis Toxin Subunits In Vivo and Their Interaction with the Ptl Transport Apparatus
Authors: Burns Drusilla L. , Fiddner Stefanie , Cheung Anissa M. , Verma Anita
Published: August 2004

Pertussis toxin (PT) has an AB
5
structure that is typical of many bacterial protein toxins; however, this toxin is more complex than many toxins since it is composed of five different subunit types, subunits S1 to S5. Little is known about how PT assembles in vivo and how and when it interacts with its secretion apparatus, known as the Ptl transporter. In order to better understand these events, we expressed subsets of the genes encoding the S1, S2, and/or S4 subunits of PT in strains of
Bordetella pertussis
that either did or did not produce the Ptl proteins. We found evidence to suggest that certain subassemblies of the toxin, including subassemblies consisting of the S1 subunit and incomplete forms of the B oligomer, can form in vivo, at least transiently. These results suggest that the B oligomer of the toxin does not need to completely form before interactions between the S1 subunit and B-oligomer subunits can occur in vivo. All subassemblies localized primarily to the membrane fraction of the cell. Moreover, we found that Ptl-mediated secretion occurs in a strain that produces S1 and an incomplete complement of B-oligomer subunits. These results indicate that subassemblies of the toxin consisting of the S1 subunit and a partial B oligomer can interact with the Ptl system.
Involvement of CD14, Toll-Like Receptors 2 and 4, and MyD88 in the Host Response to the Fungal PathogenCryptococcus neoformansIn Vivo
Authors: Yauch Lauren E. , Mansour Michael K. , Shoham Shmuel , Rottman James B. , Levitz Stuart M.
Published: August 2004

The major capsular polysaccharide ofCryptococcus neoformans, glucuronoxylomannan (GXM), is recognized by Toll-like receptor 2 (TLR2), TLR4, and CD14. In these studies, mice deficient in CD14, TLR2, TLR4, and the TLR-associated adaptor protein, MyD88, were utilized to investigate the contribution of TLRs and CD14 to in vivo host defenses againstC. neoformans. MyD88−/−mice had significantly reduced survival compared with wild-type C57BL/6 mice after intranasal (i.n.) and intravenous (i.v.) infection with liveC. neoformans. CD14−/−mice had reduced survival when infected i.v., while TLR2−/−mice died significantly earlier after i.n. infection. Mortality was similar comparing TLR4 mutant C3H/HeJ mice and control C3H/HeOuJ mice following i.v. or i.n. challenge withC. neoformans. The course of pulmonary cryptococcosis was studied in more detail in the CD14−/−, TLR2−/−, and MyD88−/−mice. MyD88−/−mice infected i.n. had higher numbers of CFU in the lungs as well as higher GXM levels in the sera and lungs 7 days after infection than wild-type mice did. Surprisingly, there were no major differences in the levels of tumor necrosis factor alpha, interleukin-4 (IL-4), IL-10, IL-12p70, or gamma interferon in the lungs ofC. neoformans-infected knockout mice compared with wild-type mice. Histopathologic analysis of the lungs on day 7 postinfection revealed minimal inflammation in all mouse groups. These studies demonstrate a major role for MyD88 and relatively minor roles for CD14 and TLR2 in the response to cryptococcal infection, with the decreased survival of MyD88−/−mice correlating with increased numbers of lung CFU and serum and lung GXM levels.
Reduction of Antibody Response to an 11-Valent Pneumococcal Vaccine Coadministered with a Vaccine Containing Acellular Pertussis Components
Authors: Dagan Ron , Goldblatt David , Maleckar James R. , Yaïch Mansour , Eskola Juhani
Published: August 2004

In pneumococcal conjugate vaccines (PCVs), polysaccharide antigens are often conjugated to protein carriers related to other common vaccines. It is therefore important to test PCV interaction with other pediatric vaccines when administered simultaneously. We assessed the immune response to an 11-valent PCV conjugated to diphtheria and tetanus carriers (PncD/T11), administered concomitantly, but in separate sites, with a combined vaccine containing epitopes related antigenically to the carriers: polyribosylribitol phosphate-tetanus tox oid (PRP-T), diphtheria toxoid (DT), and tetanus toxoid (TT). In addition, these combinations contained inactivated poliovirus vaccine (IPV) and either whole-cell pertussis (wP) or acellular pertussis (aP) components. After coadministration of PncD/T11 with the combined vaccine containing wP (DTwP/IPV/PRP-T), the responses to all polysaccharides in the PncD/T11 were satisfactory. In contrast, when coadministered with an aP-containing combination (DTaP/IPV/PRP-T), the response to all seven pneumococcal conjugates to TT was significantly reduced after primary and booster immunization. The pneumococcal conjugates to DT were not significantly reduced after the primary series, but were somewhat reduced after booster. It is likely that some suppression of the tetanus-mediated response occurred even when the PncD/T11 was coadministered with wP, but this suppression was masked by the adjuvant effect of wP. By replacing wP with aP, this adjuvant effect was removed, unmasking the suppression of the tetanus-mediated response. With the increasing use of multiple aP-containing vaccines in infancy, novel approaches to adjuvants and carrier protein technology are likely to be required.
Neutrophil NADPH Oxidase Is Reduced at theAnaplasma phagocytophilumPhagosome
Authors: IJdo Jacob W. , Mueller Angel C.
Published: August 2004

The intracellular organismAnaplasma phagocytophilumcauses human granulocytic ehrlichiosis and specifically infects and multiplies in neutrophilic granulocytes. Previous reports have suggested that, for its survival, this bacterium suppresses the neutrophil respiratory burst. To investigate the mechanism of survival, we first assessed the kinetics ofA. phagocytophilumentry into neutrophils by using double-labeling confocal microscopy. At 30, 60, 120, and 240 min of incubation, 25, 50, 55, and 70% of neutrophils contained bacteria, respectively. The neutrophil respiratory burst in the presence ofA. phagocytophilumwas assessed by a kinetic cytochromecassay and by measurement of oxygen consumption. Neutrophils in the presence ofA. phagocytophilumdid not produce a significant respiratory burst, butA. phagocytophilumdid not inhibit the neutrophil respiratory burst when phorbol myristate acetate was added. Immunoelectron microscopy of neutrophils infected withA. phagocytophilumorEscherichia colirevealed that NADPH oxidase subunits gp91phoxand p22phoxwere significantly reduced at theA. phagocytophilumphagosome after 1 and 4 h of incubation. In neutrophils incubated simultaneously withA. phagocytophilumandE. colifor 30, 60, and 90 min, gp91phoxwas present on 20, 14, and 10% of theA. phagocytophilumphagosomes, whereas p22phoxwas present in 11, 5, and 4% of the phagosomes, respectively. Similarly, onE. coliphagosomes, gp91phoxwas present in 62, 64, and 65%, whereas p22phoxwas detected in 54, 48, and 48%. We conclude thatA. phagocytophilumdoes not suppress a global respiratory burst and that, under identical conditions in the same cells,A. phagocytophilum, but notE. coli, significantly reduces gp91phoxand p22phoxfrom its phagosome membrane.
Mutation of
toxB
and a Truncated Version of the
efa-1
Gene in
Escherichia coli
O157:H7 Influences the Expression and Secretion of Locus of Enterocyte Effacement-Encoded Proteins but not Intestinal Colonization in Calves or Sheep
Authors: Stevens Mark P. , Roe Andrew J. , Vlisidou Isabella , van Diemen Pauline M. , La Ragione Roberto M. , Best Angus , Woodward Martin J. , Gally David L. , Wallis Timothy S.
Published: August 2004

Enterohemorrhagic
Escherichia coli
(EHEC) strains comprise a broad group of bacteria, some of which cause attaching and effacing (AE) lesions and enteritis in humans and animals. Non-O157:H7 EHEC strains contain the gene
efa-1
(referred to in previous publications as
efa1
), which influences adherence to cultured epithelial cells. An almost identical gene in enteropathogenic
E. coli
(
lifA
) mediates the inhibition of lymphocyte proliferation and proinflammatory cytokine synthesis. We have shown previously that significantly lower numbers of EHEC O5 and O111
efa-1
mutants are shed in feces following experimental infection in calves and that these mutants exhibit reduced adherence to intestinal epithelia compared with isogenic wild-type strains.
E. coli
O157:H7 strains lack
efa-1
but encode a homolog on the pO157 plasmid (
toxB
/
l7095
) and contain a truncated version of the
efa-1
gene (
efa-1
′/
z4332
in O island 122 of the EDL933 chromosome). Here we report that
E. coli
O157:H7
toxB
and
efa-1
′ single and double mutants exhibit reduced adherence to cultured epithelial cells and show reduced expression and secretion of proteins encoded by the locus of enterocyte effacement (LEE), which plays a key role in the host-cell interactions of EHEC. The activity of LEE1, LEE4, and LEE5 promoters was not significantly altered in
E. coli
O157:H7 strains harboring
toxB
or
efa-1
′ mutations, indicating that the effect on the expression of LEE-encoded secreted proteins occurs at a posttranscriptional level. Despite affecting type III secretion, mutation of
toxB
and
efa-1
′ did not significantly affect the course of fecal shedding of
E. coli
O157:H7 following experimental inoculation of 10- to 14-day-old calves or 6-week-old sheep. Mutation of
tir
caused a significant reduction in fecal shedding of
E. coli
O157:H7 in calves, indicating that the formation of AE lesions is important for colonization of the bovine intestine.
Identification of a Polyclonal B-Cell Activator inPlasmodium falciparum
Authors: Donati Daria , Zhang Li Ping , Chen Qijun , Chêne Arnaud , Flick Kirsten , Nyström Maja , Wahlgren Mats , Bejarano Maria Teresa
Published: August 2004

Polyclonal B-cell activation and hypergammaglobulinemia are prominent features of human malaria. We report here thatPlasmodium falciparum-infected erythrocytes directly adhere to and activate peripheral blood B cells from nonimmune donors. The infected erythrocytes employ the cysteine-rich interdomain region 1α (CIDR1α) ofP. falciparumerythrocyte membrane protein 1 (PfEMP1) to interact with the B cells. Stimulation with recombinant CIDR1α induces proliferation, an increase in B-cell size, expression of activation molecules, and secretion of immunoglobulins (immunoglobulin M) and cytokines (tumor necrosis factor alpha and interleukin-6). Furthermore, CIDR1α binds to Fab and Fc fragments of human immunoglobulins and to immunoglobulins purified from the sera of different animal species. This binding pattern is similar to that of the polyclonal B-cell activatorStaphylococcus aureusprotein A. Our findings shed light on the understanding of the molecular basis of polyclonal B-cell activation during malaria infections. The results suggest that thevargene family encoding PfEMP1 has evolved not only to mediate the sequestration of infected erythrocytes but also to manipulate the immune system to enhance the survival of the parasite.
Combined Effects of Blood and Temperature Shift on
Borrelia burgdorferi
Gene Expression as Determined by Whole Genome DNA Array
Authors: Tokarz Rafal , Anderton Julie M. , Katona Laura I. , Benach Jorge L.
Published: August 2004

Borrelia burgdorferi
undergoes differential gene expression during transmission from its tick vector to a vertebrate host. The addition of blood to a spirochete culture at 35°C for 48 h had a dramatic effect on gene expression of this organism. Utilizing
B. burgdorferi
whole genome DNA arrays, we compared the transcriptomes of the spirochetes following a 2-day temperature shift with blood and without blood. Using combined data from three independent RNA isolations we demonstrated that the addition of blood led to a differential expression of 154 genes. Of these, 75 genes were upregulated, with 49 (65%) of them encoded on plasmids. Blood supplementation of cultures also resulted in the downregulation of 79 genes, where 56 (70%) were plasmid encoded. We verified our results by reverse transcriptase PCR of several genes in both flat and feeding ticks. In the 2-day experiment we observed the effect that exposure to increased temperature and blood combined had on
B. burgdorferi
gene expression at this crucial time when the spirochetes begin to move from the vector to a new vertebrate host. These changes, among others, coincide with the upregulation of the chemotaxis and sensing regulons, of the lp38-encoded ABC transporter, of proteases capable of remodeling the outer surface of the spirochetes, and of the recombination genes of cp32 as a transient or initial part of the stress response of the phage. These are all functions that could cause or facilitate the changes that spirochetes undergo following a blood meal in the tick.
Transcriptome Analysis of
Pseudomonas aeruginosa
after Interaction with Human Airway Epithelial Cells
Authors: Frisk Anders , Schurr Jill R. , Wang Guoshun , Bertucci Donna C. , Marrero Luis , Hwang Sung Hei , Hassett Daniel J. , Schurr Michael J.
Published: August 2004

The transcriptional profile of
Pseudomonas aeruginosa
after interactions with primary normal human airway epithelial cells was determined using Affymetrix GeneChip technology. Gene expression profiles indicated that various genes involved in phosphate acquisition and iron scavenging were differentially regulated.
Microarray Analysis ofAeromonas hydrophilaCytotoxic Enterotoxin-Treated Murine Primary Macrophages
Authors: Galindo C. L. , Fadl A. A. , Sha J. , Chopra A. K.
Published: August 2004

We performed microarray analyses of murine peritoneal macrophages to examine cellular transcriptional responses to a cytotoxic enterotoxin ofAeromonas hydrophila. While 66% of altered genes were common to both primary macrophages and the murine macrophage cell line RAW 264.7, Act caused expression changes of 28 genes specifically in murine peritoneal macrophages.
The Neuroendocrine Stress Hormone Norepinephrine Augments
Escherichia coli
O157:H7-Induced Enteritis and Adherence in a Bovine Ligated Ileal Loop Model of Infection
Authors: Vlisidou Isabella , Lyte Mark , van Diemen Pauline M. , Hawes Pippa , Monaghan Paul , Wallis Timothy S. , Stevens Mark P.
Published: August 2004

The role of the neuroendocrine environment in the pathogenesis of enteric bacterial infections is increasingly being recognized. Here we report that norepinephrine augments
Escherichia coli
O157:H7-induced intestinal inflammatory and secretory responses as well as bacterial adherence to intestinal mucosa in a bovine ligated ileal loop model of infection. Norepinephrine modulation of enteritis and adherence was dependent on the ability of
E. coli
O157:H7 to form attaching and effacing lesions.
Transcriptome of Enterohemorrhagic
Escherichia coli
O157 Adhering to Eukaryotic Plasma Membranes
Authors: Dahan Sivan , Knutton Stuart , Shaw Robert K. , Crepin Valerie F. , Dougan Gordon , Frankel Gad
Published: August 2004

Using a DNA microarray, we determined changes in enterohemorrhagic
Escherichia coli
O157:H7 gene expression during binding to plasma membranes. Analysis of the complete transcriptomes of the bound bacteria revealed increased levels of stress-associated mRNAs and decreased levels of mRNA encoding proteins involved in translation and type III secretion.
The Capsule of
Bacillus anthracis
Behaves as a Thymus-Independent Type 2 Antigen
Authors: Wang Taia T. , Lucas Alexander H.
Published: August 2004

Bacillus anthracis
elaborates a homopolymeric capsule composed of γ-
d
-glutamic acid residues. Mice were immunized with formalin-fixed encapsulated
B. anthracis
bacilli, and the serum antibody response to a γ-
d
-glutamyl capsular epitope was measured. Antiglutamyl antibodies were elicited in athymic BALB/c Nu/Nu, BALB/c Nu/+, and CBA/J mice but not in CBA/N
xid
mice. These response patterns define the capsule of
B. anthracis
as a thymus-independent type 2 antigen.
Helicobacter pylori
Infection of Human and Murine Primary Gastric Cells
Authors: Clyne Marguerite , Drumm Brendan
Published: August 2004

The effect of
Helicobacter pylori
infection on human and murine primary gastric cells was determined. CagA was phosphorylated following adherence of
H. pylori
to primary human gastric cells. However, it did not adhere to human primary duodenal cells or murine gastric cells, and CagA could not be detected in cell lysates. Identification of an easily available animal model of infection in which the organism adheres to gastric mucosal cells would enhance studies of the virulence of
H. pylori
.
Chlamydia trachomatis
Infection Inhibits Both Bax and Bak Activation Induced by Staurosporine
Authors: Xiao Yangming , Zhong Youmin , Greene Whitney , Dong Feng , Zhong Guangming
Published: August 2004

We have previously shown that
Chlamydia trachomatis
inhibits host cell apoptosis and blocks mitochondrial cytochrome
c
release. We now report that activation of both Bax and Bak, two proapoptotic members of the Bcl-2 family that regulate mitochondrial cytochrome
c
release, was inhibited in chlamydia-infected cells. This observation has provided new information on the mechanisms of chlamydial antiapoptotic activity.
Characterization of Binding of
Streptococcus oralis
Glyceraldehyde-3-Phosphate Dehydrogenase to
Porphyromonas gingivalis
Major Fimbriae
Authors: Maeda Kazuhiko , Nagata Hideki , Kuboniwa Masae , Kataoka Kosuke , Nishida Nobuko , Tanaka Muneo , Shizukuishi Satoshi
Published: August 2004

Binding of
Streptococcus oralis
glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to
Porphyromonas gingivalis
fimbriae was characterized via a biomolecular interaction analysis system. The interaction was specific, and the association constant value was 4.34 × 10
7
M
−1
, suggesting that
S. oralis
GAPDH functions as a dominant receptor for
P. gingivalis
and contributes to
P. gingivalis
colonization.
Competitive Growth Advantage of Nontoxigenic Mutants in the Stationary Phase in Archival Cultures of Pathogenic
Vibrio cholerae
Strains
Authors: Paul Kalidas , Ghosh Amalendu , Sengupta Nilanjan , Chowdhury Rukhsana
Published: August 2004

Spontaneous nontoxigenic mutants of highly pathogenic
Vibrio cholerae
O1 strains accumulate in large numbers during long-term storage of the cultures in agar stabs. In these mutants, production of the transcriptional regulator ToxR was reduced due to the presence of a mutation in the ribosome-binding site immediately upstream of the
toxR
open reading frame. Consequently, the ToxR-dependent virulence regulon was turned off, with concomitant reduction in the expression of cholera toxin and toxin-coregulated pilus. An intriguing feature of these mutants is that they have a competitive fitness advantage when grown in competition with the parent strains in stationary-phase cocultures which is independent of RpoS, the only locus known to be primarily associated with acquisition of a growth advantage phenotype in bacteria.
The AraC Family Transcriptional Regulator Rv1931c Plays a Role in the Virulence of
Mycobacterium tuberculosis
Authors: Frota Cristiane C. , Papavinasasundaram K. G. , Davis Elaine O. , Colston M. Joseph
Published: August 2004

A
Mycobacterium tuberculosis
strain disrupted in the AraC homologue
Rv1931c
was isolated. The mutant strain exhibited reduced survival both in macrophages and in a mouse infection model, with survival being restored on complementation with the
Rv1931c
gene. These results suggest that
Rv1931c
regulates genes important for virulence of
M. tuberculosis
.
Both Expansion of Regulatory GR1+CD11b+Myeloid Cells and Anergy of T Lymphocytes Participate in Hyporesponsiveness of the Lung-Associated Immune System during Acute Toxoplasmosis
Authors: Voisin Mathieu-Benoît , Buzoni-Gatel Dominique , Bout Daniel , Velge-Roussel Florence
Published: August 2004

Oral infection withToxoplasma gondiileads to transient systemic hyporesponsiveness. In this report, we characterized the presence in the lungs of GR1+CD11b+myeloid cells that have potent nitric oxide-dependent immunoregulatory properties. We also demonstrated the interleukin 2-reversible anergy of both pulmonary CD8+and CD4+activated T lymphocytes with infection.
Development of a Novel Chloramphenicol Resistance Expression Plasmid Used for Genetic Complementation of a
fliG
Deletion Mutant in
Treponema denticola
Authors: Slivienski-Gebhardt Linda L. , Izard Jacques , Samsonoff William A. , Limberger Ronald J.
Published: August 2004

A new expression plasmid containing the
fla
operon promoter and a staphylococcal chloramphenicol resistance gene, was constructed to help assess the role of
fliG
in
Treponema denticola
motility. Deletion of
fliG
resulted in a nonmotile mutant with a markedly decreased number of flagellar filaments. Wild-type
fliG
genes from
T. denticola
and from
Treponema pallidum
were cloned into this expression plasmid. In both cases, the gene restored the ability of the mutant to gyrate its cell ends and enabled colony spreading in agarose. This shuttle plasmid enables high-level expression of genes in
T. denticola
and possesses an efficient selectable marker that provides a new tool for treponemal genetics.
Host Response to a
dam
Mutant of
Salmonella enterica
Serovar Enteritidis with a Temperature-Sensitive Phenotype
Authors: Giacomodonato Mónica N. , Sarnacki Sebastián H. , Caccuri Roberto L. , Sordelli Daniel O. , Cerquetti M. Cristina
Published: August 2004

The temperature-sensitive
dam
mutant strain of
Salmonella enterica
serovar Enteritidis SD1 is highly attenuated and induces innate and protective immunity in mice. SD1 activates NF-κB and induces gamma interferon secretion. Early interaction of the SD1 mutant with intestinal epithelial cells was associated with ruffling of enterocytes. Invading bacteria were found inside Peyer's patches after inoculation.
Protection of Black-Tailed Prairie Dogs (Cynomys ludovicianus) against Plague after Voluntary Consumption of Baits Containing Recombinant Raccoon Poxvirus Vaccine
Authors: Mencher Jordan S. , Smith Susan R. , Powell Tim D. , Stinchcomb Dan T. , Osorio Jorge E. , Rocke Tonie E.
Published: August 2004

Prairie dogs (Cynomysspp.) are highly susceptible toYersinia pestisand significant reservoirs of plague for humans in the western United States. A recombinant raccoon poxvirus, expressing the F1 antigen ofY. pestis, was incorporated into a palatable bait and offered to 18 black-tailed prairie dogs (Cynomys ludovicianus) for voluntary consumption; 18 negative control animals received placebo baits. Antibody titers againstY. pestisF1 antigen increased significantly (P< 0.01) in vaccinees, and their survival was significantly higher upon challenge withY. pestisthan that of negative controls (P< 0.01).
Growth Phase Regulation of
flaA
Expression in
Helicobacter pylori
Is
luxS
Dependent
Authors: Loh John T. , Forsyth Mark H. , Cover Timothy L.
Published: August 2004

LuxS plays a role in the synthesis of an extracellular signaling molecule, autoinducer 2 (AI-2). To analyze a possible role of AI-2 in regulating
Helicobacter pylori
gene expression, we constructed a panel of transcriptional reporter strains. We show that the expression of
H. pylori flaA
is growth phase dependent and that
flaA
transcription increases in association with increased culture density. Mutating the
luxS
gene eliminates growth-phase-dependent control of
flaA
, and this growth phase dependence is restored when the
luxS
mutant strain is complemented with the wild-type
luxS
gene.
Differential Monocyte Activation Underlies Strain-SpecificMycobacterium tuberculosisPathogenesis
Authors: Manca Claudia , Reed Michael B. , Freeman Sherry , Mathema Barun , Kreiswirth Barry , Barry Clifton E. , Kaplan Gilla
Published: August 2004

In vitro infection of monocytes withMycobacterium tuberculosisHN878 and related W/Beijing isolates preferentially induced interleukin-4 (IL-4) and IL-13, which characterize Th2 polarized immunity. In contrast, CDC1551 induced more IL-12 and other molecules associated with phagocyte activation and Th1 protective immunity. The differential cytokine-chemokine response was mediated by extracted lipids, suggesting that these molecules regulate host responses to infection.
The P Domain of the P0 Protein ofPlasmodium falciparumProtects against Challenge with Malaria Parasites
Authors: Rajeshwari K. , Patel Kalpesh , Nambeesan Savithri , Mehta Monika , Sehgal Alfica , Chakraborty Tirtha , Sharma Shobhona
Published: August 2004

Monoclonal antibodies (MAbs) specific for the P domain of thePlasmodium falciparumP0 phosphoriboprotein (PfP0) blocked the invasion of RBCs byP. falciparum. Vaccination with this P-domain peptide protected mice upon malaria parasite challenge. The absolute specificity of the MAbs and the PfP0 P peptide makes them potential protective malaria reagents.
TheSalmonella entericaSerovar Typhimurium Divalent Cation Transport Systems MntH and SitABCD Are Essential for Virulence in anNramp1G169Murine Typhoid Model
Authors: Zaharik Michelle L. , Cullen Vivian Li , Fung Angela M. , Libby Stephen J. , Kujat Choy Sonya L. , Coburn Bryan , Kehres David G. , Maguire Michael E. , Fang Ferric C. , Finlay B. Brett
Published: August 2004

Nramp1 is a transporter that pumps divalent cations from the vacuoles of phagocytic cells and is associated with the innate resistance of mice to diverse intracellular pathogens. We demonstrate thatsitAandmntH, genes encoding high-affinity metal ion uptake systems inSalmonella entericaserovar Typhimurium, are upregulated whenSalmonellais internalized by Nramp1-expressing macrophages and play an essential role in systemic infection of congenic Nramp1-expressing mice.
Parameters of the Attenuated Schistosome Vaccine Evaluated in the Olive Baboon
Authors: Kariuki Thomas M. , Farah Idle O. , Yole Dorcas S. , Mwenda Jason M. , Van Dam Govert J. , Deelder André M. , Wilson R. Alan , Coulson Patricia S.
Published: August 2004

Five exposures of baboons to the attenuated schistosome vaccine gave greater protection than three exposures, but this attenuation was not sustained when challenge was delayed. Within the scope of the data collected, fecal egg counts and circulating antigen levels did not accurately predict the observed worm burdens. Levels of immunoglobulin G at challenge correlated best with protection, but there was little evidence of a recall response.
Bordetella pertussisAdenylate Cyclase-Hemolysin Induces Interleukin-6 Secretion by Human Tracheal Epithelial Cells
Authors: Bassinet Laurence , Fitting Catherine , Housset Bruno , Cavaillon Jean-Marc , Guiso Nicole
Published: August 2004

After interaction with tracheal epithelial cells,Bordetella pertussisinduces the secretion of interleukin-6. This secretion is dependent on the expression of adenylate cyclase-hemolysin by the bacterium but not on the expression of other characterized bacterial toxins or adhesins. This finding confirms the important role of adenylate cyclase-hemolysin in the pathogenicity of the bacterium.
QUANTUM REFLECTION OF MASSLESS NEUTRINOS FROM A TORSION-INDUCED POTENTIAL
Authors: ALIMOHAMMADI M. , SHARIATI A.
Published: August 2004

In the context of the Einstein–Cartan–Dirac model, where the torsion of the space–time couples to the axial currents of the fermions, we study the effects of this quantum-gravitational interaction on a massless neutrino beam crossing through a medium with a high number density of fermions at rest. We calculate the reflection amplitude and show that a specific fraction of the incident neutrinos reflects from this potential if the polarization of the medium is different from zero. We also discuss the order of magnitude of the fermionic number density in which this phenomenon is observable, in other theoretical contexts, for example, the strong gravity regime and the effective field theory approach.