bluuebunny/crossref_metadata_2025_split huggingface dataset:
Contribution of the
Shigella flexneri
Sit, Iuc, and Feo Iron Acquisition Systems to Iron Acquisition In Vitro and in Cultured Cells
Authors: Runyen-Janecky L. J. , Reeves S. A. , Gonzales E. G. , Payne S. M. 
Published: March 2003

ABSTRACT

Shigella flexneri
possesses multiple iron acquisition systems, including proteins involved in the synthesis and uptake of siderophores and the Feo system for ferrous iron utilization. We identified an additional
S
.
flexneri
putative iron transport gene,
sitA
, in a screen for
S
.
flexneri
genes that are induced in the eukaryotic intracellular environment.
sitA
was present in all
Shigella
species and in most enteroinvasive
Escherichia coli
strains but not in any other
E
.
coli
isolates tested. The
sit
locus consists of four genes encoding a potential ABC transport system. The deduced amino acid sequence of the
S
.
flexneri sit
locus was homologous to the
Salmonella enterica
serovar Typhimurium Sit and
Yersinia pestis
Yfe systems, which mediate both manganese and iron transport. The
S
.
flexneri sit
promoter was repressed by either iron or manganese, and the iron repression was partially dependent upon Fur. A
sitA
::
cam
mutation was constructed in
S
.
flexneri
. The
sitA
mutant showed reduced growth, relative to the wild type, in Luria broth containing an iron chelator but formed wild-type plaques on Henle cell monolayers, indicating that the
sitA
mutant was able to acquire iron and/or manganese in the host cell. However, mutants defective in two of these iron acquisition systems (
sitA iucD
,
sitA feoB
, and
feoB iucD
) formed slightly smaller plaques on Henle cell monolayers. A strain carrying mutations in
sitA
,
feoB
, and
iucD
did not form plaques on Henle cell monolayers.
The Apa Protein ofMycobacterium tuberculosisStimulates Gamma Interferon-Secreting CD4+and CD8+T Cells from Purified Protein Derivative-Positive Individuals and Affords Protection in a Guinea Pig Model
Authors: Kumar Priti , Amara Rama Rao , Challu Vijay Kumar , Chadda Vineet Kumar , Satchidanandam Vijaya 
Published: March 2003

ABSTRACTThe search to identifyMycobacterium tuberculosisantigens capable of conferring protective immunity against tuberculosis has received a boost owing to the resurgence of tuberculosis over the past two decades. It has long been recognized that lymphoid cells are required for protection againstM. tuberculosis. While traditionally the CD4+populations of T cells were believed to predominantly serve this protective function, a pivotal role for CD8+T cells in this task has been increasingly appreciated. We show that the 50- to 55-kDa Apa protein, specified by the Rv1860 gene ofM. tuberculosis, can elicit both lymphoproliferative response and gamma interferon (IFN-γ) production from peripheral blood mononuclear cells (PBMC) of purified protein derivative (PPD)-positive individuals, with significant differences recorded in the levels of responsiveness between PPD-positive healthy controls and pulmonary tuberculosis patients. Flow cytometric analysis of whole blood stimulated with the recombinant Apa protein revealed a sizeable proportion of CD8+T cells in addition to CD4+T cells contributing to IFN-γ secretion. PBMC responding to the Apa protein produced no interleukin-4, revealing a Th1 phenotype. A DNA vaccine and a poxvirus recombinant expressing the Apa protein were constructed and tested for their ability to protect immunized guinea pigs against a challenge dose of virulentM. tuberculosis. Although the DNA vaccine afforded little protection, the poxvirus recombinant boost after DNA vaccine priming conferred a significant level of protective immunity, bringing about a considerable reduction in mycobacterial counts from the challenge bacilli in spleens of immunized guinea pigs, a result comparable to that achieved by BCG vaccination.
The Stress-Responsive
dgk
Gene from
Streptococcus mutans
Encodes a Putative Undecaprenol Kinase Activity
Authors: Lis Maciej , Kuramitsu Howard K. 
Published: March 2003

ABSTRACT

We analyzed a previously constructed stress-sensitive
Streptococcus mutans
mutant Tn-1 strain resulting from disruption by transposon Tn
916
of a gene encoding a protein exhibiting amino acid sequence similarity to the
Escherichia coli
diacylglycerol kinase. It was confirmed that the mutation led to significantly reduced lipid kinase activity, while expression of the intact gene on a plasmid restored both kinase activity and the wild-type phenotype. Further analysis revealed that the product of the
dgk
gene in
S. mutans
predominantly recognizes a lipid substrate other than diacylglycerol, most likely undecaprenol, as demonstrated by its efficient phosphorylation and the resistance of the product of the reaction to saponification. The physiological role of the product of the
dgk
gene as a putative undecaprenol kinase was further supported by a significantly higher sensitivity of the mutant to bacitracin compared with that of the parental strain.
Autodisplay: Development of an Efficacious System for Surface Display of Antigenic Determinants inSalmonellaVaccine Strains
Authors: Kramer Uwe , Rizos Konstantin , Apfel Heiko , Autenrieth Ingo B. , Lattemann Claus T. 
Published: March 2003

ABSTRACTTo optimize antigen delivery bySalmonellavaccine strains, a system for surface display of antigenic determinants was established by using the autotransporter secretion pathway of gram-negative bacteria. A modular system for surface display allowed effective targeting of heterologous antigens or fragments thereof to the bacterial surface by the autotransporter domain of AIDA-I, theEscherichia coliadhesin involved in diffuse adherence. A major histocompatibility complex class II-restricted epitope, comprising amino acids 74 to 86 of theYersinia enterocoliticaheat shock protein Hsp60 (Hsp6074-86), was fused to the AIDA-I autotransporter domain, and the resulting fusion protein was expressed at high levels on the cell surface ofE. coliandSalmonella entericaserovar Typhimurium. Colonization studies in mice vaccinated withSalmonellastrains expressing AIDA-I fusion proteins demonstrated high genetic stability of the generated vaccine strain in vivo. Furthermore, a pronounced T-cell response againstYersiniaHsp6074-86was induced in mice vaccinated with aSalmonellavaccine strain expressing the Hsp6074-86-AIDA-I fusion protein. This was shown by monitoringYersiniaHsp60-stimulated IFN-γ secretion and proliferation of splenic T cells isolated from vaccinated mice. These results demonstrate that the surface display of antigenic determinants by the autotransporter pathway deserves special attention regarding the application in live attenuatedSalmonellavaccine strains.
Synthetic Peptides Identify Promiscuous Human Th1 Cell Epitopes of the Secreted Mycobacterial Antigen MPB70
Authors: Al-Attiyah Raja , Shaban Fatema A. , Wiker Harald G. , Oftung Fredrik , Mustafa Abu S. 
Published: March 2003

ABSTRACTMPB70 is a secreted protein ofMycobacterium bovisandMycobacterium tuberculosiswhich stimulates both cellular and humoral immune responses during infection with bovine and human tubercle bacilli. In addition, vaccination with MPB70 has been shown to induce Th1 cell responses and protection in animal models of tuberculosis. The present study was carried out to map the dominant human Th1 cell epitopes of MPB70 in relation to major histocompatibility complex (MHC) class II restriction in healthy subjects showing strong T-cell responses to complex mycobacterial antigens. Peripheral blood mononuclear cells (PBMC) from HLA-DR-typed donors were tested with complex mycobacterial antigens (whole-cellM. tuberculosisandM. tuberculosisculture filtrates), with MPB70 purified from the culture filtrate ofM. bovisBCG Tokyo, and with 13 synthetic peptides (25-mers overlapping by 10 residues) covering the sequence of MPB70. The donors that responded to the complex antigens and MPB70 also responded to the cocktail of synthetic MPB70 peptides. Testing of PBMC with individual peptides showed that peptides p5 (amino acids [aa] 61 to 85), p6 (aa 76 to 100), p8 (aa 106 to 130), p12 (aa 166 to 190), and p13 (aa 181 to 193) were most frequently recognized in proliferation and gamma interferon (IFN-γ) assays. Testing of antigen-specific CD4+T-cell lines with the individual peptides of MPB70 confirmed that peptides p8, p12, and p13 contain immunodominant Th1 cell epitopes of MPB70. MHC restriction analysis with HLA-typed donors showed that MPB70 and its immunodominant peptides were presented to T cells promiscuously. The T-cell lines responding to MPB70 and peptides p8, p12, and p13 in IFN-γ assays mediated antigen-peptide-specific cytotoxic activity against monocytes/macrophages pulsed with the whole-protein antigen or the peptides. In conclusion, the promiscuous recognition of MPB70 and its immunodominant peptide defined epitopes (aa 106 to 130 and 166 to 193) by IFN-γ-producing Th1 cells supports possible application of this secreted antigen to subunit vaccine design.
Signaling through the T1/ST2 Molecule Is Not Necessary for Th2 Differentiation but Is Important for the Regulation of Type 1 Responses in NonhealingLeishmaniamajorInfection
Authors: Kropf P. , Herath S. , Klemenz R. , Müller I. 
Published: March 2003

ABSTRACTT1/ST2 is a stable cell surface marker selectively expressed on type 2 T helper (Th2) effector cells. Since nonhealingLeishmaniamajorinfections in susceptible BALB/c mice have been ascribed to a polarized Th2 response, we used an anti-T1/ST2 monoclonal antibody (MAb) or a T1-Fc fusion protein to investigate the role of CD4+T1/ST2+Th2 cells in experimental leishmaniasis. We show that interfering with T1/ST2 signaling had no effect on lesion development or parasite replication; however, it induced a significantly higher type 1 response and an enhanced capacity of CD4+T cells to respond to interleukin 12 (IL-12). Surprisingly, even in the presence of an elevated Th1 response, the production of antigen-specific type 2 cytokines was not altered in the group of mice treated with the anti-T1/ST2 MAb or the T1-Fc fusion protein. To characterize further this Th2 response, we assessed the cytokine profile of CD4+T cells and found that interfering with T1/ST2 signaling did not alter the cytokine profile of CD4+T1/ST2+T cells. These results show that T1/ST2 signaling is not necessary for the differentiation of naive CD4+T cells into antigen-specific CD4+T1/ST2+Th2 cells. In addition to CD4+T1/ST2+T cells, we detected another subpopulation of CD4+Th2 cells, negative for the expression of T1/ST2, that could differentiate in vivo in response toL.majorinfection. Taken together, our results suggest that CD4+T1/ST2+Th2 cells but not CD4+T1/ST2−Th2 cells can downregulate the Th1 response during the course of a nonhealingL.majorinfection through a mechanism that is independent of IL-4 or IL-10.
Mutation ofluxSAffects Biofilm Formation inStreptococcus mutans
Authors: Merritt Justin , Qi Fengxia , Goodman Steven D. , Anderson Maxwell H. , Shi Wenyuan 
Published: March 2003

ABSTRACTQuorum sensing is a bacterial mechanism for regulating gene expression in response to changes in population density. Many bacteria are capable of acyl-homoserine lactone-based or peptide-based intraspecies quorum sensing andluxS-dependent interspecies quorum sensing. While there is good evidence about the involvement of intraspecies quorum sensing in bacterial biofilm, little is known about the role ofluxSin biofilm formation. In this study, we report for the first time thatluxS-dependent quorum sensing is involved in biofilm formation ofStreptococcus mutans. S. mutansis a major cariogenic bacterium in the multispecies bacterial biofilm commonly known as dental plaque. An ortholog ofluxSforS. mutanswas identified using the data available in theS. mutansgenome project (http://www.genome.ou.edu/smutans.html). Using an assay developed for the detection of the LuxS-associated quorum sensing signal autoinducer 2 (AI-2), it was demonstrated that this ortholog was able to complement theluxSnegative phenotype ofEscherichia coliDH5α. It was also shown that AI-2 is indeed produced byS. mutans. AI-2 production is maximal during mid- to late-log growth in batch culture. Mutant strains devoid of theluxSgene were constructed and found to be defective in producing the AI-2 signal. There are also marked phenotypic differences between the wild type and theluxSmutants. Microscopic analysis of in vitro-grown biofilm structure revealed that theluxSmutant biofilms adopted a much more granular appearance, rather than the relatively smooth, confluent layer normally seen in the wild type. These results suggest that LuxS-dependent signal may play an important role in biofilm formation ofS. mutans.
Recognition of Mycobacterial Epitopes by T Cells across Mammalian Species and Use of a Program That Predicts Human HLA-DR Binding Peptides To Predict Bovine Epitopes
Authors: Vordermeier Martin , Whelan Adam O. , Hewinson R. Glyn 
Published: March 2003

ABSTRACTBioinformatics tools have the potential to accelerate research into the design of vaccines and diagnostic tests by exploiting genome sequences. The aim of this study was to assess whether in silico analysis could be combined with in vitro screening methods to rapidly identify peptides that are immunogenic duringMycobacterium bovisinfection of cattle. In the first instance theM. bovis-derived protein ESAT-6 was used as a model antigen to describe peptides containing T-cell epitopes that were frequently recognized across mammalian species, including natural hosts for tuberculosis (humans and cattle) and small-animal models of tuberculosis (mice and guinea pigs). Having demonstrated that some peptides could be recognized by T cells from a number ofM. bovis-infected hosts, we tested whether a virtual-matrix-based human prediction program (ProPred) could identify peptides that were recognized by T cells fromM. bovis-infected cattle. In this study, 73% of the experimentally defined peptides from 10M. bovisantigens that were recognized by bovine T cells contained motifs predicted by ProPred. Finally, in validating this observation, we showed that three of five peptides from the mycobacterial antigen Rv3019c that were predicted to contain HLA-DR-restricted epitopes were recognized by T cells fromM. bovis-infected cattle. The results obtained in this study support the approach of using bioinformatics to increase the efficiency of epitope screening and selection.
Cryptococcus neoformans
with a Mutation in the Tetratricopeptide Repeat-Containing Gene,
CCN1
, Causes Subcutaneous Lesions but Fails To Cause Systemic Infection
Authors: Chung Seyung , Mondon Philippe , Chang Yun C. , Kwon-Chung Kyung J. 
Published: March 2003

ABSTRACT

We studied a
Cryptococcus neoformans
strain that caused feline chronic nasal granuloma without disseminated disease. This strain, B-4551, grows at temperatures up to 35°C and fails to cause systemic infection in mice. Many cells of B-4551 formed short hyphal elements in feline nasal tissue and occasionally at 35°C in vitro. A complementation and sequence analysis revealed that the temperature-sensitive (Ts) phenotype of B-4551 was due to deletion of a lysine residue in the cryptococcal
CCN1
gene. B-4551 complemented with the wild type
CCN1
gene grew at 37°C and caused fatal systemic infection in mice. The
CCN1
gene encodes a protein containing 16 copies of a tetratricopeptide repeat.
CCN1
is homologous to the
Saccharomyces cerevisiae CLF1
gene, which is required for pre-mRNA splicing, cell cycle progression, and DNA replication, and to the
Drosophila melanogaster crn
gene, which is involved in neurogenesis.
CLF1
complemented the Ts phenotype of B-4551.
CCN1
, however, failed to rescue the
clf1
mutant in
S. cerevisiae
. These results indicate that the Ccn1p may not be as functionally diverse as Clf1p in yeast.
Anaplasma phagocytophilumReduces Neutrophil Apoptosis In Vivo
Authors: Scaife Helena , Woldehiwet Zerai , Hart C. Anthony , Edwards Steven W. 
Published: March 2003

ABSTRACTOvine neutrophils spontaneously underwent apoptosis during culture in vitro, as assessed by morphological changes and exposure of annexin V binding sites on their cell surfaces. The addition of conditioned medium from concanavalin A-treated ovine peripheral blood mononuclear cells (PBMC) could partially protect against this progression into apoptosis, but dexamethasone and sodium butyrate could not. Actinomycin D accelerated the rate at which ovine neutrophils underwent apoptosis. Neutrophils isolated from sheep experimentally infected withAnaplasma phagocytophilumshowed significantly delayed apoptosis during culture ex vivo, and the addition of conditioned medium from PBMC to these cells could not delay apoptosis above the protective effects observed after in vivo infection. The ability of neutrophils fromA. phagocytophilum-infected sheep to activate a respiratory burst was increased compared to the activity measured in neutrophils from uninfected sheep, but chemotaxis was decreased in neutrophils from infected sheep. These data are the first demonstration that in vivo infection withA. phagocytophilumresults in changes in rates of apoptosis of infected immune cells. This may help explain how these bacteria replicate in these normally short-lived cells.
Resistance to Acute Babesiosis Is Associated with Interleukin-12- and Gamma Interferon-Mediated Responses and Requires Macrophages and Natural Killer Cells
Authors: Aguilar-Delfin Irma , Wettstein Peter J. , Persing David H. 
Published: March 2003

ABSTRACTWe examined the role of the cytokines gamma interferon (IFN-γ) and interleukin-12 (IL-12) in the model of acute babesiosis with the WA1Babesia. Mice genetically deficient in IFN-γ-mediated responses (IFNGR2KO mice) and IL-12-mediated responses (Stat4KO mice) were infected with the WA1Babesia, and observations were made on the course of infection and cytokine responses. Levels of IFN-γ and IL-12 in serum increased 24 h after parasite inoculation. The augmented susceptibility observed in IFNGR2KO and Stat-4KO mice suggests that the early IL-12- and IFN-γ-mediated responses are involved in protection against acute babesiosis. Resistance appears to correlate with an increase in nitric oxide (NO) production. In order to assess the contribution of different cell subsets to resistance against the parasite, we also studied mice lacking B cells, CD4+T cells, NK cells, and macrophages. Mice genetically deficient in B lymphocytes or CD4+T lymphocytes were able to mount protective responses comparable to those of immunosufficient mice. In contrast, in vivo depletion of macrophages or NK cells resulted in elevated susceptibility to the infection. Our observations suggest that a crucial part of the response that protects from the pathogenicBabesiaWA1 is mediated by macrophages and NK cells, probably through early production of IL-12 and IFN-γ, and induction of macrophage-derived effector molecules like NO.
Effects of Zinc Deficiency and Pneumococcal Surface Protein A Immunization on Zinc Status and the Risk of Severe Infection in Mice
Authors: Strand Tor A. , Hollingshead Susan K. , Julshamn Kåre , Briles David E. , Blomberg Bjørn , Sommerfelt Halvor 
Published: March 2003

ABSTRACT

Streptococcus pneumoniae
is a major cause of illness and death in children in developing countries. In these children, zinc deficiency is associated with an increased risk of acute respiratory tract infections, which can be reduced by daily zinc administration. Severe infections decrease zinc levels in plasma and may thereby move individuals with preexisting low zinc stores into a vicious cycle of infection and unavailable zinc. Pneumococcal surface protein A (PspA) has emerged as a promising vaccine candidate, and immunization with this antigen protects animals from pneumococcal infection. In an animal experiment, we measured the effect of zinc depletion on the immune response to parenterally administrated PspA and assessed the effect of this PspA vaccination and zinc depletion on the severity of pneumococcal infection and on zinc status. Mice were kept on different diets for 5 weeks, immunized twice 14 days apart, and challenged intranasally with
S. pneumoniae
. Mice on the zinc-deficient diet showed substantially reduced immune responses to PspA, more extensive pneumococcal colonization in the nasal mucosa, more severe infections, and an increased risk of death. PspA immunization reduced the risk of severe disease, and the reduction in severity was reflected in substantially reduced zinc depletion from bones.
Pathogenic
Yersinia enterocolitica
Strains Increase the Outer Membrane Permeability in Response to Environmental Stimuli by Modulating Lipopolysaccharide Fluidity and Lipid A Structure
Authors: Bengoechea J. A. , Brandenburg K. , Arraiza M. D. , Seydel U. , Skurnik M. , Moriyón I. 
Published: March 2003

ABSTRACT

Pathogenic biotypes of
Yersinia enterocolitica
(serotypes O:3, O:8, O:9, and O:13), but not environmental biotypes (serotypes O:5, O:6, O:7,8, and O:7,8,13,19), increased their permeability to hydrophobic probes when they were grown at pH 5.5 or in EGTA-supplemented (Ca
2+
-restricted) media at 37°C. A similar observation was also made when representative strains of serotypes O:8 and O:5 were tested after brief contact with human monocytes. The increase in permeability was independent of the virulence plasmid. The role of lipopolysaccharide (LPS) in this phenomenon was examined by using
Y. enterocolitica
serotype O:8. LPS aggregates of bacteria grown in acidic or EGTA-supplemented broth took up more
N
-phenylnaphthylamine than LPS aggregates of bacteria grown in standard broth and also showed a marked increase in acyl chain fluidity which correlated with permeability, as determined by measurements obtained in the presence of hydrophobic dyes. No significant changes in O-antigen polymerization were observed, but lipid A acylation changed depending on the growth conditions. In standard medium at 37°C, there were hexa-, penta-, and tetraacyl lipid A forms, and the pentaacyl form was dominant. The amount of tetraacyl lipid A increased in EGTA-supplemented and acidic media, and hexaacyl lipid A almost disappeared under the latter conditions. Our results suggest that pathogenic
Y. enterocolitica
strains modulate lipid A acylation coordinately with expression of virulence proteins, thus reducing LPS packing and increasing outer membrane permeability. The changes in permeability, LPS acyl chain fluidity, and lipid A acylation in pathogenic
Y. enterocolitica
strains approximate the characteristics in
Yersinia pseudotuberculosis
and
Yersinia pestis
and suggest that there is a common outer membrane pattern associated with pathogenicity.