bluuebunny/crossref_metadata_2025_split huggingface dataset:
Enhanced Protective Antibody Responses to PspA after Intranasal or Subcutaneous Injections of PspA Genetically Fused to Granulocyte-Macrophage Colony-Stimulating Factor or Interleukin-2
Authors: Wortham Charles , Grinberg Luba , Kaslow David C. , Briles David E. , McDaniel Larry S. , Lees Andrew , Flora Michael , Snapper Clifford M. , Mond James J. 
Published: December 2019

ABSTRACTAntibody to pneumococcal surface protein A (PspA) has been shown to be protective forStreptococcus pneumoniaeinfections in mice. In an attempt to define a model for inducing protective antibody to PspA in the absence of adjuvant, we designed two genetic fusions, PspA–interleukin-2 [IL-2]) and PspA–granulocyte-macrophage colony-stimulating factor (GM-CSF). These constructs maintained high cytokine function in vitro, as tested by their activity on IL-2 or GM-CSF-dependent cell lines. While intranasal immunization with PspA induced no detectable anti-PspA response, both PspA–IL-2 and PspA–GM-CSF stimulated high immunoglobulin G1 (IgG1) antibody responses. Interestingly, only the PspA–IL-2, not the PspA–GM-CSF, construct stimulated IgG2a antibody responses, suggesting that this construct directed the response along a TH1-dependent pathway. Comparable enhancement of the anti-PspA response with similar isotype profiles was observed after subcutaneous immunization as well. The enhancement observed with PspA–IL-2 was dependent on IL-2 activity in that it was not seen in IL-2 receptor knockout mice, while PspA in alum induced high-titer antibody in these mice. The antibody was tested for its protective activity in a mouse lethality model usingS. pneumoniaeWU-R2. Passive transfer of 1:90 dilutions of sera from mice immunized with PspA–IL-2 and PspA–GM-CSF elicited protection of CBA/N mice against intravenous challenge with over 170 50% lethal doses of capsular type 3 strain WU2. Only 0.17 μg or less of IgG antibody to PspA was able to provide passive protection against otherwise fatal challenge withS. pneumoniae. The data demonstrate that designing protein-cytokine fusions may be a useful approach for mucosal immunization and can induce high-titer systemic protective antibody responses.
Vaccination with Plasmid DNA Encoding Mycobacterial Antigen 85A Stimulates a CD4+and CD8+T-Cell Epitopic Repertoire Broader than That Stimulated byMycobacterium tuberculosisH37Rv Infection
Authors: Denis Olivier , Tanghe Audrey , Palfliet Kamiel , Jurion Fabienne , van den Berg Thierry-P. , Vanonckelen Albert , Ooms Josette , Saman Eric , Ulmer Jeffrey B. , Content Jean , Huygen Kris 
Published: December 2019

ABSTRACTVaccination of mice with plasmid DNA carrying the gene for the major secreted mycobacterial antigen 85A (Ag85A) fromMycobacterium tuberculosisis a powerful technique for generating robust specific Th1 helper T-cell responses, CD8+-mediated cytotoxicity, and protection againstM. tuberculosischallenge (K. Huygen et al., Nat. Med. 2:893–898, 1996). We have now analyzed in more detail the antigen-specific immune CD4+- and CD8+-T-cell responses induced in BALB/c mice vaccinated with Ag85A DNA and have compared these responses to those generated by intravenous infection withM. tuberculosis. T-cell-epitope mapping, as measured by interleukin-2 and gamma interferon secretion from splenic T cells restimulated in vitro with synthetic 20-mer peptides spanning the complete mature sequence of Ag85A, demonstrated that DNA vaccination stimulated a stronger and broader T-cell response than didM. tuberculosisinfection. Moreover, elevated cytotoxic T lymphocyte (CTL) activity against Ag85A-transfected and peptide-pulsed P815 target cells could be generated exclusively by vaccination with plasmid DNA, not followingM. tuberculosisinfection. By using DNA vaccination, three Ag85A CTL epitopes with predicted major histocompatibility complex class I binding motifs were defined. One of them was previously reported as a dominant, promiscuously recognized T-cell epitope in healthy humans with primary infections. These data strengthen the potential of DNA vaccination with respect to inducing antituberculous immunity in humans.
Helper T-Cell Epitopes Encoded by theBabesia bigemina rap-1Gene Family in the Constant and Variant Domains Are Conserved among Parasite Strains
Authors: Brown Wendy C. , McElwain Terry F. , Hötzel Isidro , Suarez Carlos E. , Palmer Guy H. 
Published: December 2019

ABSTRACTAmong important candidates for babesial vaccines are apical complex proteins, including rhoptry-associated protein 1 (RAP-1) fromBabesia bovisandB. bigemina, which have been shown to induce partial immunity. Four variantB. bigemina rap-1transcripts identified in a clone of the Mexico strain have highly conserved sequence in the central region but vary in sequence at the amino and carboxy termini (NT and CT) of the predicted proteins, resulting in different combinations of NT and CT domains in the individual gene products. Cattle were immunized with native protein consisting of the RAP-α1 variant, which contains NT-1 and CT-1 domains, and T-cell responses were characterized. We previously reported the identification of two T helper (Th) cell epitopes inB. bigeminaRAP-1α1 protein (I. Hötzel, W. C. Brown, T. F. McElwain, S. D. Rodriguez, and G. H. Palmer, Mol. Biochem. Parasitol. 81:89–99, 1996). One epitope mapped to the constant domain of RAP-1 (amino acids [aa] 144 to 187), and one mapped to the CT-1 variable domain (aa 386 to 480). Th1-like clones responding to these epitopes proliferated differentially to different strains ofB. bigemina, raising the possibilities that the T-cell epitopes may vary antigenically and that CT-1 may be differentially expressed with respect to the other RAP-1 CT domains in the different strains. In this report, we definitively map the T-cell epitope identified in the constant domain of RAP-1 to aa 159 to 187 (FVVSLLKKNVVRDPESNDVENFASQYFYM) and show that the predicted amino acid sequence is completely conserved among seven strains. The T-cell epitope in the CT-1 domain was mapped to aa 436 to 465 (VNSEKVDADDAGNAETQQLPDAENEVRADD), which is also completely conserved among eight strains ofB. bigemina. We further show that the RAP-1α1-immunized cattle were protected against homologousB. bigeminachallenge, thus suggesting an association between protective immunity and the helper T-cell response against the two epitopes. The immunogenic and highly conserved nature of these T-cell epitopes and their ability to stimulate functionally relevant Th cells that express gamma interferon support their inclusion in a vaccine.
Copy Number of Pilus Gene Clusters in
Haemophilus influenzae
and Variation in the
hifE
Pilin Gene
Authors: Read Timothy D. , Satola Sarah W. , Opdyke Jason A. , Farley Monica M. 
Published: December 2019

ABSTRACT

Brazilian purpuric fever (BPF)-associated
Haemophilus influenzae
biogroup aegyptius strain F3031 contains two identical copies of a five gene cluster (
hifA
to
hifE
) encoding pili similar to well-characterized Hif fimbriae of
H. influenzae
type b. HifE, the putative pilus tip adhesin of F3031, shares only 40% amino acid sequence similarity with the same molecule from type b strains, whereas the other four proteins have 75 to 95% identity. To determine whether pilus cluster duplication and the
hifE
F3031
allele were special features of BPF-associated bacteria, we analyzed a collection of
H. influenzae
strains by PCR with
hifA
- and
hifE
-specific oligonucleotides, by Southern hybridization with a
hifC
gene probe, and by nucleotide sequencing. The presence of two pilus clusters was limited to some
H. influenzae
biogroup aegyptius strains. The
hifE
F3031
allele was limited to
H. influenzae
biogroup aegyptius. Two strains contained one copy of
hifE
F3031
and one copy of a variant
hifE
allele. We determined the nucleotide sequences of four
hifE
genes from
H. influenzae
biogroup aegyptius and
H. influenzae
capsule serotypes a and c. The predicted proteins produced by these genes demonstrated only 35 to 70% identity to the three published HifE proteins from nontypeable
H. influenzae
, serotype b, and BPF strains. The C-terminal third of the molecules implicated in chaperone binding was the most highly conserved region. Three conserved domains in the otherwise highly variable N-terminal putative receptor-binding region of HifE were similar to conserved portions in the N terminus of
Neisseria
pilus adhesin PilC. We concluded that two pilus clusters and
hifE
F3031
were not specific for BPF-causing
H. influenzae
, and we also identified portions of HifE possibly involved in binding mammalian cell receptors.
Decreased Antitoxic Activities among Children with Clinical Episodes of Malaria
Authors: Jakobsen Palle Høy , McKay Veronica , N’Jie Ramou , Olaleye Ben O. , D’Alessandro Umberto , Zhang Gui-Hang , Eggelte Teunis A. , Koch Claus , Greenwood Brian M. 
Published: December 2019

ABSTRACT

Healthy Gambian children, children with clinical
Plasmodium falciparum
malaria, and children with asymptomatic
P. falciparum
infections were studied to investigate whether antitoxic activities may contribute to protection against malarial symptoms. Markers of inflammatory reactions, soluble tumor necrosis factor receptor I, and C-reactive protein were found in high concentrations in children with symptomatic
P. falciparum
malaria compared with levels in children with asymptomatic
P. falciparum
infections or in healthy children, indicating that inflammatory reactions are induced only in children with clinical symptoms. Concentrations of soluble tumor necrosis factor receptor I and C-reactive protein were associated with levels of parasitemia. We detected antitoxic activities in sera as measured by their capacity to block toxin-induced
Limulus
amoebocyte lysate (LAL) activation. Symptomatic children had decreased capacity to block induction of LAL activation by
P. falciparum
exoantigen. The decreased blocking activity was restored in the following dry season, when the children had no clinical malaria. Symptomatic children also had the highest immunoglobulin G (IgG) reactivities to conserved
P. falciparum
erythrocyte membrane protein 1 and “Pfalhesin” (band #3) peptides, indicating that such IgG antibodies are stimulated by acute disease but are lost rapidly after the disease episode. Half of the children with symptomatic infections had low levels of haptoglobin, suggesting that these children had chronic
P. falciparum
infections which may have caused symptoms previously. Only a few of the children with asymptomatic
P. falciparum
infections had high parasite counts, and antitoxic immunity in the absence of antiparasite immunity appears to be rare among children in this community.
Role of Adenylate Cyclase-Hemolysin in Alveolar Macrophage Apoptosis duringBordetella pertussisInfection In Vivo
Authors: Gueirard Pascale , Druilhe Anne , Pretolani Marina , Guiso Nicole 
Published: December 2019

ABSTRACTBordetella pertussisinduces in vitro apoptosis of murine alveolar macrophages by a mechanism that is dependent on expression of bacterial adenylate cyclase-hemolysin. Using a murine respiratory model, we found in this study that intranasal infection with a parentalB. pertussisstrain, but not with an isogenic variant deficient in the expression of all toxins and adhesins, induced a marked neutrophil accumulation in the bronchoalveolar lavage fluid and an early decrease in macrophage numbers. These phenomena paralleled a time-dependent rise in the proportion of apoptotic nuclei, as detected by flow cytometry, and of macrophages which had engulfed apoptotic bodies. Apoptotic death of bronchopulmonary cells was observed exclusively following intranasal infection with bacteria reisolated from lungs of infected animals and not withB. pertussiscollected after in vitro subculture. Using the terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling technique coupled to fluorescence microscopy and morphological analysis, we established that the apoptotic cells in bronchoalveolar lavage fluids were neutrophils and macrophages. Histological analysis of the lung tissues fromB. pertussis-infected mice showed increased numbers of apoptotic cells in the alveolar compartments. Cellular accumulation in bronchoalveolar lavage fluids and apoptosis of alveolar macrophages were significantly attenuated in mice infected with a mutant deficient in the expression of adenylate cyclase-hemolysin, indicating a role of this enzyme in these processes.
Synthesis and Characterization of Lipooligosaccharide-Based Conjugates as Vaccine Candidates for
Moraxella
(
Branhamella
)
catarrhalis
Authors: Gu Xin-Xing , Chen Jing , Barenkamp Stephen J. , Robbins John B. , Tsai Chao-Ming , Lim David J. , Battey James 
Published: December 2019

ABSTRACT

Moraxella
(
Branhamella
)
catarrhalis
is an important cause of otitis media and sinusitis in children and of lower respiratory tract infections in adults. Lipooligosaccharide (LOS) is a major surface antigen of the bacterium and elicits bactericidal antibodies. Treatment of the LOS from strain ATCC 25238 with anhydrous hydrazine reduced its toxicity 20,000-fold, as assayed in the
Limulus
amebocyte lysate (LAL) test. The detoxified LOS (dLOS) was coupled to tetanus toxoid (TT) or high-molecular-weight proteins (HMP) from nontypeable
Haemophilus influenzae
through a linker of adipic acid dihydrazide to form dLOS-TT or dLOS-HMP. The molar ratios of dLOS to TT and HMP conjugates were 19:1 and 31:1, respectively. The antigenicity of the two conjugates was similar to that of the LOS, as determined by double immunodiffusion. Subcutaneous or intramuscular injection of both conjugates elicited a 50- to 100-fold rise in the geometric mean of immunoglobulin G (IgG) to the homologous LOS in mice after three injections and a 350- to 700-fold rise of anti-LOS IgG in rabbits after two injections. The immunogenicity of the conjugate was enhanced by formulation with monophosphoryl lipid A plus trehalose dimycolate. In rabbits, conjugate-induced antisera had complement-mediated bactericidal activity against the homologous strain and heterologous strains of
M. catarrhalis
. These results indicate that a detoxified LOS-protein conjugate is a candidate for immunization against
M. catarrhalis
diseases.
Genetic Control of Immune Response to Recombinant Antigens Carried by an AttenuatedSalmonella typhimuriumVaccine Strain:Nramp1Influences T-Helper Subset Responses and Protection against Leishmanial Challenge
Authors: Soo Shiu-Shing , Villarreal-Ramos Bernardo , Khan C. M. Anjam , Hormaeche Carlos E. , Blackwell Jenefer M. 
Published: December 2019

ABSTRACTAttenuated strains ofSalmonella typhimuriumhave been widely used as vehicles for delivery and expression of vaccine antigens in murine models of infectious disease. In mice, early bacterial replication following infection withS. typhimuriumis controlled by the gene (Nramp1, formerlyIty/Lsh/Bcg) encoding the natural-resistance-associated macrophage protein (Nramp1). Nramp1 regulates macrophage activation and has multiple pleiotropic effects, including regulation of tumor necrosis factor alpha, interleukin 1β (IL-1β), and major histocompatibility complex class II molecules, all of which influence antigen processing and presentation. Nramp1 also has a direct effect on antigen processing, possibly by regulating the activity of proteases in the late endosomal compartment. Hence, there are multiple ways (regulation of bacterial load or recombinant antigen dose, class II molecule expression, costimulatory or adjuvant activity, and antigen processing) that Nramp1 might influence responses to recombinant salmonella vaccines. To test the hypothesis that Nramp1 influences responses to vaccination, congenic mouse strains have been used to analyze immune responses to recombinant antigens (tetanus toxoid antigen and leishmanial gp63) carried by live attenuatedS. typhimurium aroA aroDmutants. Results show that congenic mice carrying the wild-type (S. typhimuriumresistance)Nramp1allele mount a predominantly T-helper-1 (IL-2 and gamma interferon) response to vaccination and show enhanced resolution of lesions following challenge infection withLeishmania major. In contrast, mice carrying mutant (S. typhimuriumsusceptibility)Nramp1mount a T-helper-2 (immunoglobulin E and IL-4) response and show exacerbated lesion growth upon challenge.
Campylobacter jejuni
Cytolethal Distending Toxin Causes a G
2
-Phase Cell Cycle Block
Authors: Whitehouse Chris A. , Balbo Paul B. , Pesci Everett C. , Cottle Daniel L. , Mirabito Peter M. , Pickett Carol L. 
Published: December 2019

ABSTRACT

Cytolethal distending toxin (CDT) from the diarrheagenic bacterium
Campylobacter jejuni
was shown to cause a rapid and specific cell cycle arrest in HeLa and Caco-2 cells. Within 24 h of treatment, CDT caused HeLa cells to arrest with a 4
N
DNA content, indicative of cells in G
2
or early M phase. Immunofluorescence studies indicated that the arrested cells had not entered M phase, since no evidence of tubulin reorganization or chromatin condensation was visible. CDT treatment was also shown to cause HeLa cells to accumulate the inactive, tyrosine-phosphorylated form of CDC2. These results indicated that CDT treatment results in a failure to activate CDC2, which leads to cell cycle arrest in G
2
. This mechanism of action is novel for a bacterial toxin and provides a model for the generation of diarrheal disease by
C. jejuni
and other diarrheagenic bacteria that produce CDT.
Identification of Epitope and Surface-Exposed Domains of
Shigella flexneri
Invasion Plasmid Antigen D (IpaD)
Authors: Turbyfill K. Ross , Mertz Jennifer A. , Mallett Corey P. , Oaks Edwin V. 
Published: December 2019

ABSTRACT

Transport and surface expression of the invasion plasmid antigens (Ipa proteins) is an essential trait in the pathogenicity of
Shigella
spp. In addition to the type III protein secretion system encoded by the
mxi/spa
loci on the large virulence plasmid, transport of IpaB and IpaC into the surrounding medium is modulated by IpaD. To characterize the structural topography of IpaD, the Geysen epitope-mapping system was used to identify epitopes recognized by surface-reactive monoclonal and polyclonal antibodies produced against purified recombinant IpaD or synthetic IpaD peptides. Surface-exposed epitopes of IpaD were confined to the first 180 amino acid residues, whereas epitopes in the carboxyl-terminal half were not exposed on the
Shigella
surface. By using convalescent-phase sera from 10
Shigella flexneri
-infected monkeys, numerous epitopes were mapped within a surface-exposed region of IpaD between amino acid residues 14 and 77. Epitopes were also identified in the carboxyl-terminal half of IpaD with a few convalescent-phase sera. Comparison of IpaD epitope sequences with
Salmonella
SipD sequences indicated that very similar epitopes may exist in the carboxyl-terminal region of each protein whereas the IpaD epitopes in the surface-exposed amino-terminal region were unique for the
Shigella
protein. Although the IpaD and SipD homologs may play similar roles in transport, the dominant serum antibody response to IpaD is against the unique region of this protein exposed on the surface of the pathogen.
Identification of a Homolog of CcpA Catabolite Repressor Protein in
Streptococcus mutans
Authors: Simpson Christine L. , Russell Roy R. B. 
Published: December 2019

ABSTRACT

A locus containing a gene with homology to
ccpA
of other bacteria has been cloned from
Streptococcus mutans
LT11, sequenced, and named
regM
. Upstream of the
regM
gene, on the opposite strand, is a gene encoding an X-Pro dipeptidase,
pepQ
. A 14-bp palindromic sequence with homology to the consensus catabolite-responsive element sequence lay in the promoter region between the two genes. To study the function of
regM
, the gene was inactivated by insertion of an antibiotic resistance marker. Diauxic growth of
S. mutans
on a number of sugars in the presence of glucose was not affected by disruption of
regM
. The loss of RegM increased glucose repression of α-galactosidase, mannitol-1-P dehydrogenase, and P-β-galactosidase activities. These results suggest that while RegM can affect catabolite repression in
S. mutans
, it does not conform to the model proposed for CcpA in
Bacillus subtilis.
Cloning of Syrian Hamster (Mesocricetus auratus) Cytokine cDNAs and Analysis of Cytokine mRNA Expression in Experimental Visceral Leishmaniasis
Authors: Melby Peter C. , Tryon Victor V. , Chandrasekar Bysani , Freeman Gregory L. 
Published: December 2019

ABSTRACTThe Syrian golden hamster (Mesocricetus auratus) is uniquely susceptible to a variety of intracellular pathogens and is an excellent model for a number of human infectious diseases. The molecular basis for this high level of susceptibility is unknown, and immunological studies related to this model have been limited by the lack of available reagents. In this report we describe the cloning and sequence analysis of portions of the Syrian hamster interleukin 2 (IL-2), IL-4, gamma interferon (IFN-γ), tumor necrosis factor alpha, IL-10, IL-12p40, and transforming growth factor β cDNAs. In addition, we examined the cytokine response to infection with the intracellular protozoanLeishmania donovaniin this animal model. Sequence analysis of the hamster cytokines revealed 69 to 93% homology with the corresponding mouse, rat, and human nucleotide sequences and 48 to 100% homology with the deduced amino acid sequences. The hamster IFN-γ, compared with the mouse and rat homologs, had an additional 17 amino acids at the C terminus that could decrease the biological activity of this molecule and thus contribute to the extreme susceptibility of this animal to intracellular pathogens. The splenic expression of these genes in response to infection withL. donovani, the cause of visceral leishmaniasis (VL), was determined by Northern blotting. VL in the hamster is a progressive, lethal disease which very closely mimics active human disease. In this model there was pronounced expression of the Th1 cytokine mRNAs, with transcripts being detected as early as 1 week postinfection. Basal expression of IL-4 in uninfected hamsters was prominent but did not increase in response to infection withL. donovani. IL-12 transcript expression was detected at low levels in infected animals and paralleled the expression of IFN-γ. Expression of IL-10, a potent macrophage deactivator, increased throughout the course of infection and could contribute to the progressive nature of this infection. These initial studies are the first to examine the molecular immunopathogenesis of a hamster model of VL infection and indicate that progressive disease in this model of VL is not associated with early polarization of the splenic cellular immune response toward a Th2 phenotype and away from a Th1 phenotype.