bluuebunny/crossref_metadata_2025_split huggingface dataset:
Positive and Negativecis-Acting Regulatory Sequences Control Expression of Leukotoxin inActinobacillus actinomycetemcomitans652
Authors: Mitchell Christine , Gao Ling , Demuth Donald R. 
Published: September 2003

ABSTRACTIntegration of IS1301into an AT-rich inverted repeat located upstream of theltxoperon was previously shown to confer a hyperleukotoxic phenotype inActinobacillus actinomycetemcomitansIS1 (T. He, T. Nishihara, D. R. Demuth, and I. Ishikawa, J. Periodontol.70:1261-1268, 1999), but the mechanism leading to increased leukotoxin production was not determined. We show that an IS1ltxpromoter::lacZreporter construct expresses 12-fold higher levels of β-galactosidase activity than a reporter containing theltxpromoter fromA. actinomycetemcomitans652, suggesting that IS1301increases transcription of theltxoperon. Examination of the IS1301sequence identified a potential outwardly directed promoter. However, site-specific mutagenesis of the −35 element of the putative promoter had no effect on the transcriptional activity of the IS1 reporter construct. Furthermore, reverse transcriptase PCR and real-time PCR experiments did not detect a transcript that was initiated within IS1301. These results suggest that increased expression of leukotoxin in strain IS1 does not arise from an outwardly directed IS1301promoter. To determine how IS1301alters transcriptional regulation of theltxoperon,cis-acting sequences that regulate leukotoxin expression were identified. The AT-rich sequence that resides downstream from the site of IS1301insertion was shown to function as a positivecis-acting regulator of leukotoxin expression. This sequence resembles an UP element in its location, AT-rich content, and activity and is homologous to the consensus UP element sequence. In addition, a negativecis-acting sequence was identified upstream from the site of IS1301insertion, and deletion of this region increased promoter activity by fourfold. Mobility shift experiments showed that this region bound to a protein(s) in extracts fromA. actinomycetemcomitans652. The specific sequences required for this interaction were localized to a 26-nucleotide segment of theltxpromoter that resides 17 bp upstream from the site of IS1301insertion. Together, these results suggest that positive and negativecis-acting sequences regulate leukotoxin expression and that IS1301may increase transcription of theltxoperon inA. actinomycetemcomitansIS1 by displacing a negativecis-acting regulator approximately 900 bp upstream from the basal elements of theltxpromoter.
Mechanisms of Variable
p44
Expression by
Anaplasma phagocytophilum
Authors: Lin Quan , Rikihisa Yasuko , Ohashi Norio , Zhi Ning 
Published: September 2003

ABSTRACT

The human intragranulocytic bacterium
Anaplasma phagocytophilum
promotes variation of P44s, which are surface-exposed proteins encoded by a
p44
multigene family. In the present study, the specific
p44
gene expression loci in four strains of
A. phagocytophilum
were identified and it was determined that each consisted of four tandem genes,
tr1
,
omp-1X
,
omp-1N
, and
p44
. A putative σ
70
-type promoter was found upstream of
tr1
. The
p44
genes include a central hypervariable region flanked by conserved regions. The hypervariable region sequence in the
p44
expression locus was duplicated and, regardless of the expression status, conserved at another locus in both low- and high-passage cell cultures of strain NY-37. No significant differences in the hypervariable region were found when we compared
p44
sequences, at the level of cDNA, within the expression locus and within other loci in the genomes of strains NY-37 and HZ. Similarly, in cDNA isolated from patients and from assorted cultures of strains NY-31, NY-36, and NY-37, hypervariable regions of 450 deduced amino acid sequences of various
p44
s within each strain were found to be identical, as were those of
p44
sequences in the genome of strain HZ. These data suggest that variations in
p44
sequences at the level of the
p44
expression locus occur through unidirectional conversion of the entire (nonsegmental)
p44
hypervariable region including flanking regions with a corresponding sequence copied from one of the conserved donor
p44
genomic loci. The data suggest that the P44 antigenic repertoire within the hypervariable region is restricted.
Oral Immunization of BALB/c Mice with
Giardia duodenalis
Recombinant Cyst Wall Protein Inhibits Shedding of Cysts
Authors: Larocque R. , Nakagaki K. , Lee P. , Abdul-Wahid A. , Faubert G. M. 
Published: September 2003

ABSTRACT

The process of encystation is a key step in the
Giardia duodenalis
life cycle that allows this intestinal protozoan to survive between hosts during person-to-person, animal-to-person, waterborne, or food-borne transmission. The release of cysts from infected persons and animals is the main contributing factor to contamination of the environment. Genes coding for cyst wall proteins (CWPs), which could be used for developing a transmission-blocking vaccine, have been cloned. Since the immunogenicity of recombinant
Giardia
CWP is unknown, we have investigated the immunogenicity of recombinant CWP2 (rCWP2) and its efficacy in interfering with the phenomenon of encystation taking place in the small bowels of BALB/c mice vaccinated with the recombinant protein. Here we report that the immunization of BALB/c mice with rCWP2 stimulated the immune system in a manner comparable to that for a live infection with
Giardia muris
cysts. Fecal and serum anti-rCWP2 immunoglobulin A (IgA) antibodies were detected in the immunized mice. In addition, anti-rCWP2 IgG1 and IgG2a antibodies were detected in the serum. mRNAs coding for Th1 and Th2 types of cytokines were detected in spleen and Peyer's patch cells from immunized mice. When the vaccinated mice were challenged with live cysts, the animals shed fewer cysts. We conclude that rCWP2 is a possible candidate antigen for the development of a transmission-blocking vaccine.
Helicobacter pyloriOuter Membrane Vesicles Modulate Proliferation and Interleukin-8 Production by Gastric Epithelial Cells
Authors: Ismail Salim , Hampton Mark B. , Keenan Jacqueline I. 
Published: September 2003

ABSTRACTHelicobacter pyloriinfection, which is always associated with gastritis, can progress to ulceration or malignancy. The diversity in clinical outcomes is partly attributed to the expression of virulence factors and adhesins byH. pylori. However,H. pylorimay not have to adhere to the epithelium to cause gastritis. We hypothesize that outer membrane vesicles (OMV), which are constantly shed from the surface ofH. pylori, play a role as independent activators of host cell responses. In this study, we found that low doses of OMV fromcagPAI+toxigenic andcagPAI−nontoxigenic strains increased proliferation of AGS gastric epithelial cells. At higher doses, we detected growth arrest, increased toxicity, and interleukin-8 (IL-8) production. The only strain differences detected were vacuolation with the toxigenic strain and higher levels of IL-8 production with OMV from thecagPAI−nontoxigenic strain. In summary, we suggest that constitutively shed OMV play a role in promoting the low-grade gastritis associated withH. pyloriinfection.
Specific Antibody Responses to Three Schistosome-Related Carbohydrate Structures in Recently Exposed Immigrants and Established Residents in an Area ofSchistosoma mansoniEndemicity
Authors: Naus Cynthia W. A. , van Remoortere Alexandra , Ouma John H. , Kimani Gachuhi , Dunne David W. , Kamerling Johannis P. , Deelder André M. , Hokke Cornelis H. 
Published: September 2003

ABSTRACTBy the use of surface plasmon resonance spectroscopy, immunoglobulin G (IgG) subclass and IgM antibodies against three schistosome-derived carbohydrate structures, FLDN (Fucα1-3GalNAcβ1-4GlcNAcβ1-3Galα1), LDN-DF [GalNAcβ1-4(Fucα1-2Fucα1-3)GlcNAcβ1], and LDNF [GalNAcβ1-4(Fucα1-3)GlcNAcβ1-3Galα1], were measured in 184 previously unexposed Kenyan immigrants who moved into the Masongaleni area, whereSchistosoma mansoniis endemic. They were sampled within their first year of exposure and again 2 years later. A cohort selected out of the original residents of the area, who had been exposed for many years, served as controls. Associations with responses toS. mansoniworm, egg (SEA), and cercarial (CERC) antigens were examined. In addition, we measured responses to keyhole limpet hemocyanin, a glycoprotein which carries glycan epitopes that are also expressed by schistosomes. Specific IgG1 responses were most pronounced against FLDN and LDN-DF and strongly associated with those previously measured to SEA and CERC. Similarly to previously published age profiles of IgG1 and IgG2 responses to SEA, levels of IgG1 against LDN-DF decreased with age. In contrast, specific IgM responses against the three schistosome-derived carbohydrate structures were most marked against LDNF. Our results indicate that, of the three glycan structures tested, the acute response against schistosome glycoconjugate antigens in young children is mainly directed against the LDN-DF epitope. The response to LDN-DF in older individuals and the responses to the two other epitopes were similar in the two cohorts, suggesting that these antigens are recognized in the early stages of infection and that the immune response persists. The biological significance of these observations needs further elucidation.
The Aggregation Domain of Aggregation Substance, Not the RGD Motifs, Is Critical for Efficient Internalization by HT-29 Enterocytes
Authors: Waters Christopher M. , Wells Carol L. , Dunny Gary M. 
Published: September 2003

ABSTRACT

Aggregation substance (AS), a surface protein encoded on the pheromone-inducible plasmids of
Enterococcus faecalis
, has been shown to increase adherence and internalization into a number of different cell types, presumably through integrin binding mediated by the N-terminal RGD motif of AS. Here, defined mutations constructed in Asc10, the AS encoded by the plasmid pCF10, are analyzed for their ability to promote increased internalization levels into HT-29 enterocytes. The results clearly show that the previously identified Asc10 functional domain, not the RGD motifs, is critical for Asc10-directed internalization of
E. faecalis
into HT-29 enterocytes. Also, expression of Asc10 in the nonaggregating
E. faecalis
strain INY3000 is unable to mediate HT-29 internalization. However, Asc10-expressing
E. faecalis
cells are not internalized as bacterial aggregates, suggesting bacterial aggregation is not a prerequisite for HT-29 internalization. These data show that Asc10 directs internalization of
E. faecalis
into HT-29 enterocytes through a non-RGD-dependent mechanism.
Deregulated Production of Protective Cytokines in Response toCandida albicansInfection in Patients with Chronic Mucocutaneous Candidiasis
Authors: Lilic Desa , Gravenor Ian , Robson Neil , Lammas David A. , Drysdale Pam , Calvert Jane E. , Cant Andrew J. , Abinun Mario 
Published: September 2003

ABSTRACTPatients with chronic mucocutaneous candidiasis (CMC) are selectively unable to clear the yeastCandida, which results in persistent debilitating infections affecting the skin, nails, and mucous membranes. The underlying defect is unknown. Recent animal studies highlighted the importance of type 1 cytokines in protection againstCandida, and previous work suggested that CMC patients may exhibit altered cytokine production in response toCandida.Based on these findings, in this study we investigated cytokine production in CMC patients by assessing a range of inflammatory, anti-inflammatory, type 1, and type 2 cytokines (interleukin-2 [IL-2], IL-4, IL-5, IL-6, IL-10, IL-12, gamma interferon [IFN-γ], tumor necrosis factor alpha [TNF-α]) in whole-blood cultures in response to five different fractions ofCandida albicans(carbohydrate, purified mannan, and protein-rich fractions, etc.), as well as non-Candidaantigens. Our results demonstrate that cytokine production is deregulated in aCandida-specific way for some cytokines (IL-2, IL-10), is deregulated more generally for other cytokines (IL-12, IL-6, IFN-γ), and is not markedly altered for still other cytokines (TNF-α, IL-4, IL-5). The most notable finding in CMC patients was the markedly impaired production of IL-12 in parallel with dramatically increased levels of IL-6 and IL-10 that occurred selectively in response toCandida.These results suggest that patients with CMC have impaired production of type 1-inducing cytokines (possibly a macrophage or dendritic cell defect?), which could result in an inability to mount protective cell-mediated responses and a failure to clearCandida.Continued tissue damage and inflammation may trigger production of high levels of inhibitory cytokines, such as the IL-10 production seen in our study, which would further reduce production of type 1-inducing cytokines in a positive feedback loop leading to persistent infection.
Analysis of Immunological Nonresponsiveness to the 19-Kilodalton Fragment of Merozoite Surface Protein 1 ofPlasmodium yoelii: Rescue by Chemical Conjugation to Diphtheria Toxoid (DT) and Enhancement of Immunogenicity by Prior DT Vaccination
Authors: Stanisic Danielle I. , Martin Laura B. , Liu Xue Q. , Jackson David , Cooper Juan , Good Michael F. 
Published: September 2003

ABSTRACTThePlasmodiummerozoite surface protein 1 (MSP1) is a leading vaccine candidate for protecting against the blood stage of malaria. Previous studies have shown that the 19-kDa carboxyl terminus of this protein is able to induce protective immunity in some monkey and mouse strains. We show that immunization with the recombinantPlasmodium yoelii19-kDa fragment of MSP1 (MSP119) expressed inSaccharomyces cerevisiae(yMSP119) can induce protective antibodies in several inbred mouse strains and one outbred mouse strain. However, mice expressing theH-2smajor histocompatibility complex haplotype are unable to generate yMSP119-specific antibodies. While synthetic peptides derived from MSP119are immunogenic in B10.S mice, they cannot function as helper epitopes, and immunization with yMSP119does not induce T cells that recognize the recombinant protein or synthetic peptides corresponding to its sequence. Nonresponsiveness could be overcome by using chemical linkers to conjugate yMSP119to diphtheria toxoid (DT), resulting in immunogens capable of inducing protective yMSP119-specific antibodies in both MSP119-responsive and otherwise nonresponsive mouse strains. The ability of sera from mice immunized with the conjugate to inhibit binding of a protective monoclonal antibody (MAb 302) to yMSP119correlated strongly with a delay in the prepatent period. Chemical conjugation of yMSP119to DT may be a preferred method to enhance immunogenicity, as carrier priming experiments demonstrated that an existing immune response to DT enhanced a subsequent antibody response to yMSP119after vaccination with yMSP119-DT. These results have important implications for the development of a malaria vaccine to protect a population with diverse HLAs.
Construction and Functional Activities of Chimeric Mouse-Human Immunoglobulin G and Immunoglobulin M Antibodies against theNeisseria meningitidisPorA P1.7 and P1.16 Epitopes
Authors: Michaelsen Terje E. , Ihle Øistein , Beckstrøm Karen Johanne , Herstad Tove K. , Kolberg Jan , Høiby E. Arne , Aase Audun 
Published: September 2003

ABSTRACTWe studied the in vitro protective activities of human immunoglobulin G1 (IgG1), IgG3, and IgM antibodies against group B meningococci by constructing sets of chimeric mouse-human antibodies (chIgG1, chIgG3, and chIgM, respectively) with identical binding regions against the P1.7 and P1.16 epitopes on PorA. This was done by cloning the V genes of three mouse hybridoma antibodies and subsequently transfecting vectors containing the homologous heavy- and light-chain genes into NSO cells. Cell clones secreting intact human chIgG1, chIgG3, or chIgM antibodies originating from three parent mouse antibodies were isolated. The functional affinities appeared to be similar for all human isotypes and surprisingly also for the pentameric chIgM antibody. chIgG1 exhibited greater serum bactericidal activity (SBA) than chIgG3, while chIgG3 was more efficient in inducing a respiratory burst (RB) associated with opsonophagocytosis than chIgG1 was. On the other hand, chIgM exhibited SBA similar to that of chIgG1, but it exhibited much higher RB activity than chIgG3 and chIgG1 exhibited. The antibodies against the P1.16 epitope were more efficient in terms of SBA than the antibodies against the P1.7 epitope were; thus, 10- to 40-fold-lower concentrations of antibodies against P1.16 than of antibodies against P1.7 were needed to induce SBA. On the other hand, antibodies against these epitopes were equally effective in inducing RB. Our results revealed differences in the functional activities of human chIgG1, chIgG3, and chIgM antibodies against meningococci, which might influence their protective effects against meningococcal disease.
Serum Immunoglobulin G Response to Candidate Vaccine Antigens during Experimental Human Pneumococcal Colonization
Authors: McCool Tera L. , Cate Thomas R. , Tuomanen Elaine I. , Adrian Peter , Mitchell Tim J. , Weiser Jeffrey N. 
Published: September 2003

ABSTRACT
The immune response to pneumococcal surface structures during colonization was examined in a model of experimental human pneumococcal carriage. Healthy uncolonized adults were given a type 23F or 6B pneumococcus, and a portion of these subjects became colonized (6 of 14 with type 23F and 6 of 8 with type 6B). Sera from colonized and uncolonized subjects were used to determine the titer of antibody specific to pneumococcal surface components under consideration in development of noncapsular polysaccharide-based vaccines. These vaccine candidates included pneumococcal surface protein A (PspA), choline binding protein A (CbpA), lipoteichoic acid, immunoglobulin A1 (IgA1) protease, pneumolysin, proteinase maturation protein A, and pneumococcal surface adhesin A. Only the two related choline binding proteins, PspA and CbpA, were immunogenic in colonized subjects as determined by a statistically significant rise in the serum IgG titer. The serum IgG response to PspA was shown previously to correlate inversely with susceptibility to carriage and was localized to a region within the N-terminal portion of PspA. This region is highly variable in amino acid sequence between pneumococcal strains. Despite the sequence diversity in the immunodominant regions of both PspA and CbpA, a significant strain-to-strain cross-reactivity in the serum IgG response following experimental human carriage was observed. These findings support the need for further investigation of the human antibody response to PspA and CbpA and the potential use of one or both of these proteins as novel vaccine antigens for the prevention of pneumococcal colonization.
The Vsa Proteins Modulate Susceptibility of
Mycoplasma pulmonis
to Complement Killing, Hemadsorption, and Adherence to Polystyrene
Authors: Simmons Warren L. , Dybvig Kevin 
Published: September 2003

ABSTRACT

The variable surface antigens (Vsa) of the murine respiratory pathogen
Mycoplasma pulmonis
are associated with the virulence of the microorganism in the lung. In strain UAB CT, the antigens consist of an N-terminal region that is combined with one of seven different C-terminal variable regions comprised of tandem repeats.
M. pulmonis
producing a VsaA protein with about 40 tandem repeats (R40) does not adhere to red blood cells or polystyrene. Strains that produce VsaH contain a short C-terminal region that lacks tandem repeats and adhere to red blood cells and plastic. We isolated and analyzed
M. pulmonis
strain CT variants (CT182 and derivatives) that produced a VsaA protein with only three tandem repeats (R3). These variants adhered to plastic and red blood cells similarly to the VsaH-producing strain. When the R3-producing CT182 strain or the VsaH-producing strains were incubated with normal guinea pig serum, they were efficiently killed. Killing was abolished when the serum was heat inactivated. In contrast, the
M. pulmonis
strains that produced VsaA R40 were highly resistant to complement killing. CT182R3 variants that survived the complement killing reactions all produced the R40 form of VsaA and were resistant to complement killing. VsaA R40 is the first mycoplasmal protein shown to be associated with resistance to complement. As both VsaH and VsaA can mediate adherence to plastic, cytadherence, and susceptibility to complement, we propose that Vsa modulates these phenotypes by nonspecific interactions.
gp63 Homologues inTrypanosoma cruzi: Surface Antigens with Metalloprotease Activity and a Possible Role in Host Cell Infection
Authors: Cuevas Ileana C. , Cazzulo Juan J. , Sánchez Daniel O. 
Published: September 2003

ABSTRACTgp63 is a highly abundant glycosylphosphatidylinositol (GPI)-anchored membrane protein expressed predominantly in the promastigote but also in the amastigote stage ofLeishmaniaspecies. InLeishmaniaspp., gp63 has been implicated in a number of steps in establishment of infection. Here we demonstrate thatTrypanosoma cruzi, the etiological agent of Chagas' disease, has a family of gp63 genes composed of multiple groups. Two of these groups,Tcgp63-Iand-II, are present as high-copy-number genes. The genomic organization and mRNA expression pattern were specific for each group.Tcgp63-Iwas widely expressed, while theTcgp63-IIgroup was scarcely detected in Northern blots, even though it is well represented in theT. cruzigenome. Western blots using sera directed against a synthetic peptide indicated that theTcgp63-Igroup produced proteins of ∼78 kDa, differentially expressed during the life cycle. Immunofluorescence staining and phosphatidylinositol-specific phospholipase C digestion confirmed that Tcgp63-I group members are surface proteins bound to the membrane by a GPI anchor. We also demonstrate the presence of metalloprotease activity which is attributable, at least in part, to Tcgp63-I group. Since antibodies against Tcgp63-I partially blocked infection of Vero cells by trypomastigotes, a possible role for this group in infection is suggested.
BrucellaLumazine Synthase Elicits a Mixed Th1-Th2 Immune Response and Reduces Infection in Mice Challenged withBrucella abortus544 Independently of the Adjuvant Formulation Used
Authors: Velikovsky Carlos A. , Goldbaum Fernando A. , Cassataro Juliana , Estein Silvia , Bowden Raúl A. , Bruno Laura , Fossati Carlos A. , Giambartolomei Guillermo H. 
Published: September 2003

ABSTRACTThe immunogenicity and protective efficacy of recombinant lumazine synthase fromBrucellaspp. (rBLS) administered with different adjuvants was evaluated in mice. Mice were immunized with rBLS in the absence or the presence of aluminum hydroxide gel (BLS-Al), monophosphoryl lipid A (BLS-MPA), or incomplete Freund's adjuvant (BLS-IFA). rBLS per se induced a vigorous immunoglobulin G (IgG) response, with high titers of IgG1 as well as IgG2. All the adjuvants increased this response; the BLS-IFA formulation was the most effective at inducing BLS-specific IgG antibodies. In addition, after in vitro stimulation with rBLS, spleen cells from BLS-IFA-, BLS-Al-, or BLS-MPA-immunized mice proliferated and produced interleukin-2 (IL-2), gamma interferon (IFN-γ), IL-10, and IL-4, suggesting the induction of a mixed Th1-Th2 response. Immunization with rBLS protected mice against challenge withB. abortus544. The levels of protection in the spleen were similar for all adjuvants, but only BLS-Al and BLS-IFA were effective in the liver. Our results indicate that BLS might be a useful candidate for the development of subunit vaccines against brucellosis, since it elicits antigen-specific cellular responses, with production of IFN-γ and protection, independently of the adjuvant formulation used.