bluuebunny/crossref_metadata_2025_split huggingface dataset:
Induction of Cell-Mediated Immunity againstMycobacterium tuberculosisUsing DNA Vaccines Encoding Cytotoxic and Helper T-Cell Epitopes of the 38-Kilodalton Protein
Authors: Fonseca Dora P. A. J. , Benaissa-Trouw Barry , van Engelen Marloes , Kraaijeveld Cees A. , Snippe Harm , Verheul André F. M. 
Published: July 2002

ABSTRACTCell-mediated immune responses are crucial in the protection against tuberculosis. In this study, we constructed DNA vaccines encoding cytotoxic T lymphocytes (CTL) and T helper cell (Th) epitopes of the 38-kDa lipoglycoprotein ofMycobacterium tuberculosisand analyzed and compared their immunogenicities with that of pXJ38, a DNA vaccine encoding the entire 38-kDa protein (X. Zhu, N. Venkataprasad, H. S. Thangaraj, M. Hill, M. Singh, J. Ivanyi, and H. M. Vordermeier, J. Immunol. 158:5921–5926, 1997). Plasmid DNAs encoding a CTL epitope, P3 (pP3), a Th epitope (vTh), or both the Th and the P3 epitopes (pThP3) were prepared and tested in C57BL6/J (H-2b) mice. Our results confirmed that DNA immunization with pXJ38 induces strong CD8+CTL and Th1 responses (high gamma interferon [IFN-γ], low interleukin-4 [IL-4]). Coadministration of plasmid DNAs encoding a Th epitope with those encoding a CTL epitope (vTh+pP3) elicited both antigen-specific CD8+CTL and Th1 responses. High levels of IFN-γ were secreted by spleen cells from all plasmid DNA-vaccinated mice after in vitro stimulation with the recombinant 38-kDa protein. Small or undetectable amounts of IL-4 were observed, which indicates the induction of a Th1-like response. Multiple-epitope vaccination by vTh+pP3 or pThP3 resulted in a broader Th1 response to peptide or epitopes than the single-epitope plasmid DNAs. Antigen-specific immunoglobulin G2a was only detected in sera from mice immunized with the plasmid pXJ38, and not in mice immunized with the epitope-based DNA vaccines. Thus, the absence of an antibody response after immunization with epitope plasmid DNAs and their ability to trigger only a specific cellular immune response may prove to be important advantages for a vaccine against tuberculosis.
Reciprocal, Temporal Expression of SpeA and SpeB by Invasive M1T1 Group A Streptococcal Isolates In Vivo
Authors: Kazmi Shahana U. , Kansal Rita , Aziz Ramy K. , Hooshdaran Massoumeh , Norrby-Teglund Anna , Low D. E. , Halim Abdel-Baset , Kotb Malak 
Published: July 2002

ABSTRACT

The streptococcal pyrogenic exotoxins (Spes) play a central role in the pathogenesis of invasive group A streptococcal (GAS) infections. The majority of recent invasive GAS infections have been caused by an M1T1 strain that harbors the genes for several streptococcal superantigens, including
speA
,
speB
,
speF
,
speG
, and
smeZ
. However, considerable variation in the expression of Spe proteins among clonal M1 isolates has been found, and many of the
speA
-positive M1 strains do not produce detectable amounts of SpeA in vitro. This study was designed to test the hypothesis that
speA
gene expression can be induced in vivo. A mouse infection chamber model that allows sequential sampling of GAS isolates at various time points postinfection was developed and used to monitor the kinetics of Spe production in vivo. Micropore Teflon diffusion chambers were implanted subcutaneously in BALB/c mice, and after 3 weeks the pores became sealed with connective tissue and sterile fluid containing a white blood cell infiltrate accumulated inside the infection chambers. Representative clonal M1T1 isolates expressing no detectable SpeA were inoculated into the implanted chambers, and the expression of SpeA in the aspirated aliquots of the chamber fluid was analyzed on successive days postinfection. Expression of SpeA was detected in the chamber fluid as early as days 3 to 5 postinfection in most animals, with a significant increase in expression by day 7 in all infected mice. Isolates recovered from the chamber and grown in vitro continued to produce SpeA even after 21 passages in vitro, suggesting stable switch on of the
speA
gene. A temporal relation between the upregulation of SpeA expression and the downregulation of SpeB expression was observed in vivo. These data suggest that in vivo host and/or environmental signals induced
speA
gene expression and suppressed
speB
gene expression. This underscores the role of the host-pathogen interaction in regulating the expression of streptococcal virulence factors in vivo. The model described here should facilitate such studies.
Predominance of Serotype-Specific Mucosal Antibody Response in
Shigella flexneri
-Infected Humans Living in an Area of Endemicity
Authors: Rasolofo-Razanamparany Voahangy , Cassel-Beraud Anne-Marie , Roux Jean , Sansonetti Philippe J. , Phalipon Armelle 
Published: July 2002

ABSTRACT

The mucosal humoral immune response elicited following
Shigella flexneri
infection in patients living in Antananarivo districts (Madagascar Island) was evaluated by measuring the gut-derived, circulating immunoglobulin A (IgA) antibody-secreting cells (ASC) specific for the major bacterial antigen lipopolysaccharide (LPS). Fifty, 34, 11, and 5% of the
S. flexneri
-positive patients were infected with serotypes 2a, 1a, 4a, and 3a, respectively. The total number of IgA ASC in infected patients increased significantly, compared to the number in healthy controls, early after the onset of disease. The number of anti-homologous LPS IgA ASC varied among individuals and peaked between days 5 and 10 after the onset of the disease. In the
S. flexneri
1a- and 2a-infected patients, the level of IgA ASC cross-reactivity to heterologous
S. flexneri
serotypes was weak. These data indicate that
S. flexneri
2a and 1a are the predominant strains responsible for shigellosis in this area of endemicity and that the anti-LPS antibody response following natural infection is mainly directed against serotype-specific determinants.
Roles of Fructosyltransferase and Levanase-Sucrase of
Actinomyces naeslundii
in Fructan and Sucrose Metabolism
Authors: Bergeron Lori J. , Burne Robert A. 
Published: July 2002

ABSTRACT

The ability of
Actinomyces naeslundii
to convert sucrose to extracellular homopolymers of fructose and to catabolize these types of polymers is suspected to be a virulence trait that contributes to the initiation and progression of dental caries and periodontal diseases. Previously, we reported on the isolation and characterization of the gene,
ftf
, encoding the fructosyltransferase (FTF) of
A. naeslundii
WVU45. Allelic exchange mutagenesis was used to inactivate
ftf
, revealing that FTF-deficient stains were completely devoid of the capacity to produce levan-type (β2,6-linked) polysaccharides. A polyclonal antibody was raised to a histidine-tagged, purified
A. naeslundii
FTF, and the antibody was used to localize the enzyme in the supernatant fluid. A sensitive technique was developed to detect levan formation by proteins that had been separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the method was used to confirm that the levan-synthesizing activity of
A. naeslundii
existed predominantly in a cell-free form, that a small amount of the activity was cell associated, and that the
ftf
mutant was unable to produce levans. By using the nucleotide sequence of the levanase gene of a genospecies 2
A. naeslundii
, formerly
Actinomyces viscosus
, a portion of a homologue of this gene (
levJ
) was amplified by PCR and inserted into a suicide vector, and the resulting construct was used to inactivate the
levJ
gene in the genospecies 1 strain WVU45. A variety of physiologic and biochemical studies were performed on the wild-type and LevJ-deficient strains to demonstrate that (i) this enzyme was the dominant levanase and sucrase of
A. naeslundii
; (ii) that LevJ was inducible by growth in sucrose; (iii) that the LevJ activity was found predominantly (>90%) in a cell-associated form; and (iv) that there was a second, fructose-inducible fructan hydrolase activity produced by these strains. The data provide the first detailed molecular analysis of fructan production and catabolism in this abundant and important oral bacterium.
Interleukin-2 and Loss of Immunity in ExperimentalMycobacterium aviumInfection
Authors: Mannering Stuart I. , Cheers Christina 
Published: July 2002

ABSTRACTExperimental infection of mice with a virulent strain ofMycobacterium aviumleads to a slowly progressive disease, which we have previously shown culminates in loss of gamma interferon (IFN-γ) production by T lymphocytes and death of the animals approximately 40 weeks after infection. Here we investigated the changes in T-cell activation, the production of interleukin-2 (IL-2), and the response to IL-2 throughoutM. aviuminfection as a possible explanation for this loss. We found that there is a steady increase in the percentage of T cells expressing activation markers right to the end of infection. However, in vivo T-cell proliferation, measured as a percentage of CD4+and CD8+cells incorporating 5-bromo-2′-deoxyuridine, initially increased but then remained constant. In the final stages of infection there was a decline in proliferation of activated (CD62L−) T cells. Since IL-2 is a major driver of T-cell proliferation, we asked whether this was due to loss of IL-2 responsiveness or production. However, CD25 (IL-2Rα) continued to be highly expressed in the terminal stages of infection, and although IL-2 production declined, addition of recombinant IL-2 to cultures could not rescue the final loss of IFN-γ production.
Genomic Approach for Analysis of Surface Proteins inChlamydia pneumoniae
Authors: Montigiani Silvia , Falugi Fabiana , Scarselli Maria , Finco Oretta , Petracca Roberto , Galli Giuliano , Mariani Massimo , Manetti Roberto , Agnusdei Mauro , Cevenini Roberto , Donati Manuela , Nogarotto Renzo , Norais Nathalie , Garaguso Ignazio , Nuti Sandra , Saletti Giulietta , Rosa Domenico , Ratti Giulio , Grandi Guido 
Published: July 2002

ABSTRACTChlamydia pneumoniae, a human pathogen causing respiratory infections and probably contributing to the development of atherosclerosis and heart disease, is an obligate intracellular parasite which for replication needs to productively interact with and enter human cells. Because of the intrinsic difficulty in working withC. pneumoniaeand in the absence of reliable tools for its genetic manipulation, the molecular definition of the chlamydial cell surface is still limited, thus leaving the mechanisms of chlamydial entry largely unknown. In an effort to define the surface protein organization ofC. pneumoniae, we have adopted a combined genomic-proteomic approach based on (i) in silico prediction from the available genome sequences of peripherally located proteins, (ii) heterologous expression and purification of selected proteins, (iii) production of mouse immune sera against the recombinant proteins to be used in Western blotting and fluorescence-activated cell sorter (FACS) analyses for the identification of surface antigens, and (iv) mass spectrometry analysis of two-dimensional electrophoresis (2DE) maps of chlamydial protein extracts to confirm the presence of the FACS-positive antigens in the chlamydial cell. Of the 53 FACS-positive sera, 41 recognized a protein species with the expected size on Western blots, and 28 of the 53 antigens shown to be surface-exposed by FACS were identified on 2DE maps of elementary-body extracts. This work represents the first systematic attempt to define surface protein organization inC. pneumoniae.
Structure ofCampylobacter jejuniLipopolysaccharides Determines Antiganglioside Specificity and Clinical Features of Guillain-Barré and Miller Fisher Patients
Authors: Ang C. W. , Laman J. D. , Willison H. J. , Wagner E. R. , Endtz H. P. , De Klerk M. A. , Tio-Gillen A. P. , Van den Braak N. , Jacobs B. C. , Doorn P. A. Van 
Published: July 2002

ABSTRACTGanglioside mimicry in the lipopolysaccharide (LPS) fraction ofCampylobacter jejuniisolated from Guillain-Barré syndrome (GBS) and Miller Fisher syndrome (MFS) patients was compared with isolates from patients with an uncomplicated enteritis. The antibody response toC. jejuniLPS and gangliosides in neuropathy patients and controls was compared as well. LPS from GBS and MFS-associated isolates more frequently contained ganglioside-like epitopes compared to control isolates. Almost all neuropathy patients showed a strong antibody response against LPS and multiple gangliosides in contrast to enteritis patients. Isolates from GBS patients more frequently had a GM1-like epitope than isolates from MFS patients. GQ1b-like epitopes were present in all MFS-associated isolates and was associated with anti-GQ1b antibody reactivity and the presence of oculomotor symptoms. These results demonstrate that the expression of ganglioside mimics is a risk factor for the development of post-Campylobacterneuropathy. This study provides additional evidence for the hypothesis that the LPS fraction determines the antiganglioside specificity and clinical features in post-Campylobacterneuropathy patients.
Osmolarity and pH Growth Conditions Regulate
fim
Gene Transcription and Type 1 Pilus Expression in Uropathogenic
Escherichia coli
Authors: Schwan William R. , Lee Jeffrey L. , Lenard Farrah A. , Matthews Brian T. , Beck Michael T. 
Published: July 2002

ABSTRACT

A comparative study was performed to determine the effects of pH, osmolarity, and human urine on the transcription of several
fim
genes, as well as the overall expression of type 1 pili. Several
fim-lacZYA
fusions were constructed on single-copy plasmids to test a range of pHs and a range of osmolarities. Growth in acidic medium slightly reduced expression from all of the
fim
promoters (
fimA
,
fimB
, and
fimE
). Increased osmolarity in neutral-pH medium repressed
fimA
and
fimB
transcription by approximately 50% when 400 mM NaCl was used and nearly threefold when 800 mM NaCl was used, whereas
fimE
transcription rose slightly as the osmolarity increased. This effect was more pronounced in high-osmolarity acidic media;
fimB
and
fimA
expression decreased fivefold in growth media containing 800 mM NaCl compared to expression in growth media without added NaCl. Moreover,
fimE
expression doubled under the same high-osmolarity conditions compared to expression in a low-osmolarity acidic environment. When a
fimB-lacZ
or
fimE-lacZ
fusion was inserted into the chromosome of strain AAEC189,
fimE
expression changed slightly as the osmolarity increased, but
fimB
expression decreased by 50% in a low-pH high-osmolarity environment. When strain AAEC189 with either a plasmid-borne
fimB-lacZ
fusion or a plasmid-borne
fimE-lacZ
fusion was grown in human urine, similar changes in the levels of
fimB
and
fimE
expression were observed. Limiting-dilution reverse transcription-PCR confirmed that these changes in
fim
expression occurred in clinical isolates of uropathogenic
Escherichia coli
grown in media with different pHs and different osmolarities. Furthermore, the invertible switch region in uropathogenic strain NU149 shifted from favoring the phase-on position in a neutral-pH low-osmolarity environment to favoring the phase-off position in a low-pH high-osmolarity environment. Results obtained with an
ompR
mutant strain demonstrated that
fimB
expression was derepressed and that OmpR may neutralize repression by an acid response regulator of
fimE
expression in a low-pH environment. In addition, H-NS was verified to be important in regulation of
fimB
, but it had only a slight effect on
fimE
under the specific pH and osmotic growth conditions tested. Enzyme immunoassays with anti-type 1 pilus antibody and hemagglutination assays showed that fewer type 1 pili were detected with cells in a low-pH high-osmolarity environment. Together, these observations demonstrate that a combination of low pH and high osmolarity regulates the transcription of
fim
genes, which favors a shift in the invertible element to the phase-off orientation and a loss of type 1 pilus expression. Taken together, our data suggest that the environmental cues that we tested may regulate expression of type 1 pili in specific in vivo niches, such as murine kidneys and possibly human kidneys.
Lymphocyte Recruitment and Protective Efficacy against Pulmonary Mycobacterial Infection Are Independent of the Route of PriorMycobacterium bovisBCG Immunization
Authors: Palendira Umaimainthan , Bean Andrew G. D. , Feng Carl G. , Britton Warwick J. 
Published: July 2002

ABSTRACTMycobacterium tuberculosisinfects humans through the lung, and immunity to this chronic infection is mediated primarily by CD4+T lymphocytes. Recently we have demonstrated that the recruitment of lymphocytes to the lung during primary aerosolM. tuberculosisinfection in mice occurs predominantly through the interaction of α4β1integrin on CD4+T cells and vascular cell adhesion molecule-1 on the pulmonary endothelium. To investigate the effect of route of immunization withMycobacterium bovisBCG on the pattern of T-cell recruitment to the lung, we have analyzed the differences in expression of integrins on activated memory CD4+T cells infiltrating the lung following primary BCG immunization by aerosol, intravenous, and subcutaneous routes and after subsequent aerosol challenge withM. tuberculosis. There were marked differences in the patterns of recruitment of activated CD4+T cells to the lung following primary immunization by the three routes. Expansion of CD44hiCD62LlowCD4+T cells in the lung occurred following aerosol and intravenous BCG immunizations, and the lymphocyte recruitment was proportional to the pulmonary bacterial load. The majority of infiltrating CD4+T cells expressed α4β1integrin. On subsequent exposure to aerosol BCG rapid expansion of gamma interferon-secreting α4β1+CD4+T cells occurred to the same extent in all immunized mice, regardless of the route of immunization. Similar expansion of α4β1+CD4+memory T cells occurred followingM. tuberculosischallenge. The three routes of BCG immunization resulted in the same level of protection against aerosolM. tuberculosisor BCG challenge in both the lungs and spleen. Therefore, recruitment of effector T lymphocytes and protective efficacy against pulmonary mycobacterial infection are independent of the route of prior BCG immunization.
Role of Various Enterotoxins in
Aeromonas hydrophila-
Induced Gastroenteritis: Generation of Enterotoxin Gene-Deficient Mutants and Evaluation of Their Enterotoxic Activity
Authors: Sha Jian , Kozlova E. V. , Chopra A. K. 
Published: July 2002

ABSTRACT

Three enterotoxins from the
Aeromonas hydrophila
diarrheal isolate SSU have been molecularly characterized in our laboratory. One of these enterotoxins is cytotoxic in nature, whereas the other two are cytotonic enterotoxins, one of them heat labile and the other heat stable. Earlier, by developing an isogenic mutant, we demonstrated the role of a cytotoxic enterotoxin in causing systemic infection in mice. In the present study, we evaluated the role of these three enterotoxins in evoking diarrhea in a murine model by developing various combinations of enterotoxin gene-deficient mutants by marker-exchange mutagenesis. A total of six isogenic mutants were prepared in a cytotoxic enterotoxin gene (
act
)-positive or -negative background strain of
A. hydrophila
. We developed two single knockouts with truncation in either the heat-labile (
alt
) or the heat-stable (
ast
) cytotonic enterotoxin gene; three double knockouts with truncations of genes encoding (i)
alt
and
ast
, (ii)
act
and
alt
, and (iii)
act
and
ast
genes; and a triple-knockout mutant with truncation in all three genes,
act
,
alt
, and
ast
. The identity of these isogenic mutants developed by double-crossover homologous recombination was confirmed by Southern blot analysis. Northern and Western blot analyses revealed that the expression of different enterotoxin genes in the mutants was correspondingly abrogated. We tested the biological activity of these mutants in a diet-restricted and antibiotic-treated mouse model with a ligated ileal loop assay. Our data indicated that all of these mutants had significantly reduced capacity to evoke fluid secretion compared to that of wild-type
A. hydrophila
; the triple-knockout mutant failed to induce any detectable level of fluid secretion. The biological activity of selected
A. hydrophila
mutants was restored after complementation. Taken together, we have established a role for three enterotoxins in
A. hydrophila
-induced gastroenteritis in a mouse model with the greatest contribution from the cytotoxic enterotoxin Act, followed by the Alt and Ast cytotonic enterotoxins.
Preexisting Inflammation Due toMycobacterium bovisBCG Infection Differentially Modulates T-Cell Priming against a Replicating or Nonreplicating Immunogen
Authors: Dudani Renu , Chapdelaine Yvan , van Faassen Henk , Smith Dean K. , Shen Hao , Krishnan Lakshmi , Sad Subash 
Published: July 2002

ABSTRACTInduction of T-cell memory by vaccination ensures long-term protection against pathogens. We determined whether on-going inflammatory responses during vaccination influenced T-cell priming. A preexposure of mice toMycobacterium bovisBCG impaired their subsequent ability to prime T cells againstListeria monocytogenes. This was characterized by a decrease inL. monocytogenes-specific gamma interferon (IFN-γ)-secreting CD4+and CD8+T cells. The intensity of T-cell priming towardsL. monocytogenesdepended on the extent ofL. monocytogenesexpansion, and a cessation of this expansion caused byM. bovisBCG-induced inflammation resulted in impairment in T-cell priming. A challenge ofM. bovisBCG-infected mice with a higherL. monocytogenesdose increasedL. monocytogenessurvival and restored T-cell priming towardsL. monocytogenes. Impairment in T-cell priming towardsL. monocytogenesdue toM. bovisBCG-induced inflammation resulted in a compromised protective efficacy in the long term after mice were rechallenged withL. monocytogenes. Preexisting inflammation selectively impaired T-cell priming for replicating immunogens as CD8+T-cell response to ovalbumin administered as an inert antigen (ovalbumin-archaeosomes) was enhanced byM. bovisBCG preimmunization, whereas priming towards ovalbumin administered as a live immunogen (L. monocytogenes-ovalbumin) was impaired. Thus, depending on the nature of the immunogen, the presence of prior inflammatory responses may either impede or boost vaccine efficacy.