bluuebunny/crossref_metadata_2025 huggingface dataset:
An Immunoglobulin Superfamily-Like Domain Unique to the
Yersinia pseudotuberculosis
Invasin Protein Is Required for Stimulation of Bacterial Uptake via Integrin Receptors
Authors: Dersch Petra , Isberg Ralph R.
Published: July 2002

The binding of the
Yersinia pseudotuberculosis
and
Yersinia enterocolitica
invasin proteins to β
1
integrin receptors allows internalization of these organisms by cultured cells. The C-terminal 192-residue superdomain of the
Y. pseudotuberculosis
invasin is necessary and sufficient for integrin recognition, while a region located outside, and N-terminal to, this superdomain strongly enhances the efficiency of bacterial uptake. Within the enhancer region is a domain called D2 that allows invasin-invasin interaction. To investigate the role of the enhancer region, bacterial cell binding and entry mediated by the
Y. pseudotuberculosis
invasin protein (invasin

pstb

) was compared to that of
Y. enterocolitica
invasin (invasin

ent

), which lacks the D2 self-association domain. Invasin

ent

was shown to be unable to promote self-interaction, using the DNA binding domain of λ repressor as a reporter. Furthermore, two genetically engineered in-frame deletion mutations that removed D2 from invasin

pstb

were significantly less proficient than wild-type invasin

pstb

at promoting uptake, although the amount of surface-exposed invasin as well as the cell binding capacity of the recombinant
Escherichia coli
strains remained similar. Competitive uptake assays showed that
E. coli
cells expressing invasin

pstb

had a significant advantage in the internalization process versus either
E. coli
cells expressing invasin

ent

or the invasin

pstb

derivatives deleted for D2, further demonstrating the importance of invasin self-interaction for the efficiency of invasin-mediated uptake.
Protection against VirulentMycobacterium aviumInfection following DNA Vaccination with the 35-Kilodalton Antigen Is Accompanied by Induction of Gamma Interferon-Secreting CD4+T Cells
Authors: Martin Ela , Kamath Arun T. , Triccas James A. , Britton Warwick J.
Published: July 2002

Mycobacterium aviumis an opportunistic pathogen that primarily infects immunocompromised individuals, although the frequency ofM. aviuminfection is also increasing in the immunocompetent population. The antigen repertoire ofM. aviumvaries from that ofMycobacterium tuberculosis, with the immunodominant 35-kDa protein being present inM. aviumandMycobacterium lepraebut not in members of theM. tuberculosiscomplex. Here we show that a DNA vector encoding thisM. avium35-kDa antigen (DNA-35) induces protective immunity against virulentM. aviuminfection, and this protective effect persists over 14 weeks of infection. In C57BL/6 mice, DNA vaccines expressing the 35-kDa protein as a cytoplasmic or secreted protein, both induced strong T-cell gamma interferon (IFN-γ) and humoral immune responses. Furthermore, the antibody response was to conformational determinants, confirming that the vector-encoded protein had adopted the native conformation. DNA-35 immunization resulted in an increased activated/memory CD4+T-cell response, with an accumulation of CD4+CD44hiCD45RBloT cells and an increase in antigen-specific IFN-γ production. The protective effect of the DNA-35 vectors againstM. aviuminfection was comparable to that of vaccination withMycobacterium bovisBCG and significantly greater than that for previous treated infection withM. avium. These results illustrate the importance of the 35-kDa protein in the protective response toM. aviuminfection and indicate that DNA vaccination successfully promotes a sustained level of protection during chronicM. aviuminfection.
Adhesion Molecule Deficiencies IncreasePorphyromonas gingivalis-Induced Alveolar Bone Loss in Mice
Authors: Baker Pamela J. , DuFour Lisa , Dixon Mark , Roopenian Derry C.
Published: July 2002

Alveolar bone resorption can be induced in specific-pathogen-free mice by oral infection withPorphyromonas gingivalis(P. J. Baker, R. T. Evans, and D. C. Roopenian, Arch. Oral Biol. 39:1035–1040, 1994). Here we used a mouse strain, C57BL/6J, which is relatively resistant toP. gingivalis-induced bone loss to examine whether partial or complete deletion of various adhesion molecules would increase susceptibility. Complete deletion of P-selectin or nearly complete lack of expression of intercellular adhesion molecule 1 (ICAM-1) led to increased susceptibility to bone resorption after oral infection, while a hypomorphic defect in β2-integrins did not. Both the total amount of bone lost and the number of sites at which there was significant loss were increased in mice deficient in either ICAM-1 or P-selectin. Each of the three adhesion molecule deficiencies was sufficient to decreaseP. gingivalis-specific serum immunoglobulin G responses, but lower antibody titers did not lead to increased bone loss in partially β2-integrin-deficient mice. In conclusion, P-selectin and ICAM-1 deficiencies increase susceptibility to and severity of alveolar bone loss afterP. gingivalisinfection. This finding underscores the importance of innate immunity in protection againstP. gingivalis-induced alveolar bone resorption.
Carcinoembryonic Antigen Family Receptor Specificity of
Neisseria meningitidis
Opa Variants Influences Adherence to and Invasion of Proinflammatory Cytokine-Activated Endothelial Cells
Authors: Muenzner Petra , Dehio Christoph , Fujiwara Taku , Achtman Mark , Meyer Thomas F. , Gray-Owen Scott D.
Published: July 2002

The carcinoembryonic antigen (CEA) family member CEACAM1 (previously called biliary glycoprotein or CD66a) was previously shown to function as a receptor that can mediate the binding of Opa protein-expressing
Neisseria meningitidis
to both neutrophils and epithelial cells. Since neutrophils and polarized epithelia have both been shown to coexpress multiple CEACAM receptors, we have now extended this work to characterize the binding specificity of meningococcal Opa proteins with other CEA family members. To do so, we used recombinant
Escherichia coli
expressing nine different Opa variants from three meningococcal strains and stably transfected cell lines expressing single members of the CEACAM family. These infection studies demonstrated that seven of the nine Opa variants bound to at least one CEACAM receptor and that binding to each of these receptors is sufficient to trigger the Opa-dependent bacterial uptake by these cell lines. The other two Opa variants do not appear to bind to either CEACAM receptors or heparan sulfate proteoglycan receptors, which are bound by some gonococcal Opa variants, thus implying a novel class of Opa proteins. We have also extended previous studies by demonstrating induction of CEACAM1 expression after stimulation of human umbilical vein endothelial cells with the proinflammatory cytokine tumor necrosis factor alpha, which is present in high concentrations during meningococcal disease. This induced expression of CEACAM1 leads to an increased Opa-dependent bacterial binding and invasion into the primary endothelia, implying that these interactions may play an important role in the pathogenesis of invasive meningococcal disease.
Correlations between Antibody Immune Responses at Different Mucosal Effector Sites Are Controlled by Antigen Type and Dosage
Authors: Externest Dörthe , Meckelein Barbara , Schmidt M. Alexander , Frey Andreas
Published: July 2002

Monitoring specific secretory immunoglobulin A (IgA) responses in the intestines after mucosal immunization or infection is impeded by the fact that sampling of small intestinal secretions requires invasive methods not feasible for routine diagnostics. Since IgA plasma cells generated after intragastric immunization are known to populate remote mucosal sites as well, secretory IgA responses at other mucosal surfaces may correlate to those in the intestines and could serve as proxy measures for IgA secretion in the gut. To evaluate the practicability of this approach, mice were immunized intragastrically with 0.2, 2, and 20 mg of ovalbumin plus 10 μg of cholera toxin, and the antigen-specific local secretory IgA responses in duodenal, ileal, jejunal, rectal, and vaginal secretions, saliva, urine, and feces, as well as serum IgG and IgA responses were analyzed by enzyme-linked immunosorbent assay. Correlation analysis revealed significant relationships between serum IgG and IgA, urinary IgA, salivary IgA, and secretory IgA in duodenal, jejunal, ileal, and rectal secretions for the 0.2-mg but not for the 20-mg ovalbumin dose. Fecal samples were poor predictors for intestinal antiovalbumin IgA responses, and no correlations could be established for cholera toxin, neither between local anti-cholera toxin levels nor to the antiovalbumin responses. Thus, specific IgA in serum, saliva, or urine can serve as a predictor of the release of specific IgA at intestinal surfaces after intragastric immunization, but the lack of correlations for high ovalbumin doses and for cholera toxin indicates a strong dependency on antigen type and dosage for these relationships.
Selection of Recombinant Antibodies Specific for Pathogenic
Streptococcus suis
by Subtractive Phage Display
Authors: de Greeff Astrid , van Alphen Loek , Smith Hilde E.
Published: July 2002

A semisynthetic antibody phage display library was used to select recombinant antibodies directed against surface components of a pathogenic strain of
Streptococcus suis
serotype 2 and against extracellular factor (EF), a protein known to be exclusively associated with pathogenic
S. suis
serotype 2 strains. Three distinct monoclonal phage antibodies directed against conformational epitopes of surface protein components of
S. suis
were selected. In addition, three different monoclonal phage antibodies were isolated that recognized EF. To isolate antibody fragments that recognize epitopes specific for a pathogenic
S. suis
serotype 2 strain, compared to a nonpathogenic serotype 2 strain, we applied a subtractive selection procedure. With this procedure, only one distinct phage antibody was found, and it was shown to be directed against EF. This demonstrates the selectivity of the applied procedure and confirms that EF is indeed differentially expressed by pathogenic and nonpathogenic strains. It also shows that EF is a very dominant antigen in phage antibody selections.
Genetic Locus for Streptolysin S Production by Group A Streptococcus
Authors: Nizet Victor , Beall Bernard , Bast Darrin J. , Datta Vivekananda , Kilburn Laurie , Low Donald E. , De Azavedo Joyce C. S.
Published: July 2002

Group A streptococcus (GAS) is an important human pathogen that causes pharyngitis and invasive infections, including necrotizing fasciitis. Streptolysin S (SLS) is the cytolytic factor that creates the zone of beta-hemolysis surrounding GAS colonies grown on blood agar. We recently reported the discovery of a potential genetic determinant involved in SLS production,
sagA
, encoding a small peptide of 53 amino acids (S. D. Betschel, S. M. Borgia, N. L. Barg, D. E. Low, and J. C. De Azavedo, Infect. Immun. 66:1671–1679, 1998). Using transposon mutagenesis, chromosomal walking steps, and data from the GAS genome sequencing project (
www.genome.ou.edu/strep.html
), we have now identified a contiguous nine-gene locus (
sagA
to
sagI
) involved in SLS production. The
sag
locus is conserved among GAS strains regardless of M protein type. Targeted plasmid integrational mutagenesis of each gene in the
sag
operon resulted in an SLS-negative phenotype. Targeted integrations (i) upstream of the
sagA
promoter and (ii) downstream of a terminator sequence after
sagI
did not affect SLS production, establishing the functional boundaries of the operon. A rho-independent terminator sequence between
sagA
and
sagB
appears to regulate the amount of
sagA
transcript produced versus transcript for the entire operon. Reintroduction of the nine-gene
sag
locus on a plasmid vector restored SLS activity to the nonhemolytic
sagA
knockout mutant. Finally, heterologous expression of the intact
sag
operon conferred the SLS beta-hemolytic phenotype to the nonhemolytic
Lactococcus lactis
. We conclude that gene products of the GAS
sag
operon are both necessary and sufficient for SLS production. Sequence homologies of
sag
operon gene products suggest that SLS is related to the bacteriocin family of microbial toxins.
Nonopsonic Phagocytosis of Zymosan andMycobacterium kansasiiby CR3 (CD11b/CD18) Involves Distinct Molecular Determinants and Is or Is Not Coupled with NADPH Oxidase Activation
Authors: Le Cabec Véronique , Cols Carine , Maridonneau-Parini Isabelle
Published: July 2002

Complement receptor type 3 (CR3) was initially described as an opsonic receptor. Subsequently, CR3-mediated lectin-sugar recognition mechanisms have been shown to play a major role in the nonopsonic phagocytosis of several pathogens, among themMycobacterium tuberculosis. Little is known about the binding and signal transduction mechanisms operating during nonopsonic ingestion through CR3 of different microorganisms. In the present study, we used CHO cells stably transfected with CR3 to show that CR3 was able to mediate internalization of zymosan and pathogenic mycobacteria (Mycobacterium kansasiiandMycobacterium avium) but not that of nonpathogenic species (Mycobacterium smegmatisandMycobacterium phlei). A combination of mannan and β-glucan inhibited the phagocytosis of zymosan but had no effect onM. kansasiiingestion. Among six monoclonal antibodies (MAbs) directed against the CD11b subunit of CR3 that decreased zymosan ingestion, only three inhibitedM. kansasiiphagocytosis. In particular, MAbs known to block the CR3 lectin site affected only internalization of zymosan. Using U937 macrophages, we observed that zymosan ingestion through CR3 induced superoxide production measured by cytochromecreduction and by translocation of the NADPH oxidase cytosolic component p47phox to the phagosomal membrane, whereas phagocytosis of viable or heat-killedM. kansasiidid not. Furthermore, lack of superoxide anion production during phagocytosis ofM. kansasiiwas not due to inhibition of NADPH oxidase per se or superoxide anion scavenging. Together, our results indicate that (i) nonopsonic phagocytosis of zymosan andM. kansasiiby CR3 implicates different molecular mechanisms involving multiple and distinct epitopes of CD11b and (ii) CR3 may transduce different cellular responses depending on the sites mediating nonopsonic phagocytosis.
Sunlight-Induced Propagation of the Lysogenic Phage Encoding Cholera Toxin
Authors: Faruque Shah M. , Asadulghani  , Rahman M. Mostafizur , Waldor Matthew K. , Sack David A.
Published: July 2002

In toxigenic
Vibrio cholerae
, the cholera enterotoxin (CT) is encoded by CTXΦ, a lysogenic bacteriophage. The propagation of this filamentous phage can result in the origination of new toxigenic strains. To understand the nature of possible environmental factors associated with the propagation of CTXΦ, we examined the effects of temperature, pH, salinity, and exposure to direct sunlight on the induction of the CTX prophage and studied the transmission of the phage to potential recipient strains. Exposure of cultures of CTXΦ lysogens to direct sunlight resulted in ∼10,000-fold increases in phage titers. Variation in temperature, pH, or salinity of the culture did not have a substantial effect on the induction of the prophage, but these factors influenced the stability of CTXΦ particles. Exposure of mixed cultures of CTXΦ lysogens and potential recipient strains to sunlight significantly increased both the in vitro and in vivo (in rabbit ileal loops) transduction of the recipient strains by CTXΦ. Included in these transduction experiments were two environmental nontoxigenic (CTXΦ
−
) strains of
V. cholerae
O139. These two O139 strains were transduced at high efficiency by CTXΦ, and the phage genome integrated into the O139 host chromosome. The resulting CTXΦ lysogens produced biologically active CT both in vitro and in rabbit ileal loops. This finding suggests a possible mechanism explaining the origination of toxigenic
V. cholerae
O139 strains from nontoxigenic progenitors. This study indicates that sunlight is a significant inducer of the CTX prophage and suggests that sunlight-induced transmission of CTXΦ may constitute part of a natural mechanism for the origination of new toxigenic strains of
V. cholerae
.
Interleukin 18 Restores Defective Th1 Immunity toCandida albicansin Caspase 1-Deficient Mice
Authors: Mencacci Antonella , Bacci Angela , Cenci Elio , Montagnoli Claudia , Fiorucci Sabrina , Casagrande Andrea , Flavell Richard A. , Bistoni Francesco , Romani Luigina
Published: July 2002

Caspase 1, formerly designated interleukin 1β (IL-1β)-converting enzyme, processes pro-IL-1β and pro-IL-18 to yield active cytokines that play a pivotal role in inflammation and cell activation. We show here the effect of caspase 1 deficiency on the inflammatory and adaptive immune responses to the fungusCandida albicans. Caspase 1 deficiency did not affect susceptibility to primary systemic infection with the fungus, as revealed by survival and fungal growth. However, Th1-mediated resistance to reinfection was greatly impaired in caspase 1-deficient mice, and this correlated with low-level production of IL-12 and gamma interferon. Early in infection, production of these cytokines and that of tumor necrosis factor alpha, IL-6, and, interestingly, IL-1β occurred normally in caspase 1-deficient mice, while that of IL-18 was severely impaired. Exogenous administration of IL-18, more than IL-12, restored the Th1-mediated resistance to the infection. We conclude that, while caspase 1 is not indispensable for release of mature IL-1β in candidiasis, the caspase 1-dependent production of IL-18 may represent an important and novel pathway for the expression of sustained Th1 reactivity to the fungus.
Group A Streptococcal
rofA
Gene Is Involved in the Control of Several Virulence Genes and Eukaryotic Cell Attachment and Internalization
Authors: Beckert Susanne , Kreikemeyer Bernd , Podbielski Andreas
Published: July 2002

The serotype M6 group A streptococcal RofA regulator was previously shown to exert a direct positive control of protein F1 expression and, concomitantly, fibronectin binding. Using a serotype M6
rofA
mutant, we demonstrate here that this regulator has a potentially indirect negative influence on the expression of the
mga
,
emm
6,
pel-sagA
, and
speA
virulence genes. Additionally, the
rofA
mutant exhibited reduced eukaryotic cell internalization rates in combination with decreased host cell viability.
Critical Role for Tumor Necrosis Factor Alpha in Controlling the Number of Lumenal Pathogenic Bacteria and Immunopathology in Infectious Colitis
Authors: Gonçalves Nathalie S. , Ghaem-Maghami Marjan , Monteleone Giovanni , Frankel Gad , Dougan Gordon , Lewis David J. M. , Simmons Cameron P. , MacDonald Thomas T.
Published: July 2002

Infection of mice with the intestinal bacterial pathogenCitrobacter rodentiumresults in colonic mucosal hyperplasia and a local Th1 inflammatory response similar to that seen in mouse models of inflammatory bowel disease. In these latter models, and in patients with Crohn's disease, neutralization of tumor necrosis factor alpha (TNF-α) is of therapeutic benefit. Since there is no information on the role of TNF-α in either immunity to noninvasive bacterial pathogens or on the role of TNF-α in the immunopathology of infectious colitis, we investigatedC. rodentiuminfection in TNFRp55−/−mice. In TNFRp55−/−mice, there were higher colonic bacterial burdens, but the organisms were cleared at the same rate as C57BL/6 mice, showing that TNF-α is not needed for protective antibacterial immunity. The most striking feature of infection in TNFRp55−/−mice, however, was the markedly enhanced pathology, with increased mucosal weight and thickness, increased T-cell infiltrate, and a markedly greater mucosal Th1 response. Interleukin-12 p40 transcripts were markedly elevated inC. rodentium-infected TNFRp55−/−mice, and this was associated with enhanced mucosal STAT4 phosphorylation. TNF-α is not obligatory for protective immunity toC. rodentiumin mice; however, it appears to play some role in downregulating mucosal pathology and Th1 immune responses.
Nasal and Vaginal Vaccinations Have Differential Effects on Antibody Responses in Vaginal and Cervical Secretions in Humans
Authors: Johansson Eva-Liz , Wassén Lotta , Holmgren Jan , Jertborn Marianne , Rudin Anna
Published: July 2002

Sexually transmitted diseases are a major health problem worldwide, but there is still a lack of knowledge about how to induce an optimal immune response in the genital tract of humans. In this study we vaccinated 21 volunteers nasally or vaginally with the model mucosal antigen cholera toxin B subunit and determined the level of specific immunoglobulin A (IgA) and IgG antibodies in vaginal and cervical secretions as well as in serum. To assess the hormonal influence on the induction of antibody responses after vaginal vaccination, we administered the vaccine either independently of the stage in the menstrual cycle or on days 10 and 24 in the cycle in different groups of subjects. Vaginal and nasal vaccinations both resulted in significant IgA and IgG anti-cholera toxin B subunit responses in serum in the majority of the volunteers in the various vaccination groups. Only vaginal vaccination given on days 10 and 24 in the cycle induced strong specific antibody responses in the cervix with 58-fold IgA and 16-fold IgG increases. In contrast, modest responses were seen after nasal vaccination and in the other vaginally vaccinated group. Nasal vaccination was superior in inducing a specific IgA response in vaginal secretions, giving a 35-fold increase, while vaginal vaccination induced only a 5-fold IgA increase. We conclude that a combination of nasal and vaginal vaccination might be the best vaccination strategy for inducing protective antibody responses in both cervical and vaginal secretions, provided that the vaginal vaccination is given on optimal time points in the cycle.
Gravidity-Dependent Production of Antibodies That Inhibit Binding ofPlasmodium falciparum-Infected Erythrocytes to Placental Chondroitin Sulfate Proteoglycan during Pregnancy
Authors: O'Neil-Dunne Iona , Achur Rajeshwara N. , Agbor-Enoh Sean T. , Valiyaveettil Manojkumar , Naik Ramachandra S. , Ockenhouse Christian F. , Zhou Ainong , Megnekou Rosette , Leke Rose , Taylor Diane W. , Gowda D. Channe
Published: July 2002

During pregnancy,Plasmodium falciparum-infected erythrocytes sequester in the placenta by adhering to chondroitin 4-sulfate, creating a risk factor for both the mother and the fetus. The primigravidae are at higher risk for placental malaria than the multigravidae. This difference in susceptibility has been attributed to the lack of antibodies that block the adhesion of infected erythrocytes to placental chondroitin 4-sulfate in primigravid women. However, recent results show that many primigravidae at term have antibody levels similar to those of multigravidae, and thus the significance of antiadhesion antibodies in providing protection against malaria during pregnancy remains unclear. In this study, we analyzed plasma samples from women of various gravidities at different gestational stages for antiadhesion antibodies. The majority of women, regardless of gravidity, had similar levels of antibodies at term. Most primigravidae had low levels of or no antiadhesion antibodies prior to ∼20 weeks of pregnancy and then produced antibodies. Multigravidae also lacked antibodies until ∼12 weeks of pregnancy, but thereafter they efficiently produced antibodies. In pregnant women who had placental infection at term, higher levels of antiadhesion antibodies correlated with lower levels of placental parasitemia. The difference in kinetics of antibody production between primigravidae and multigravidae correlated with the prevalence of malaria in these groups, suggesting that antibodies are produced during pregnancy in response to placental infection. The early onset of efficient antibody response in multigravidae and the delayed production to antibodies in primigravidae appear to account for the gravidity-dependent differential susceptibilities of pregnant women to placental malaria.
Protective KilledLeptospira borgpeterseniiVaccine Induces Potent Th1 Immunity Comprising Responses by CD4 and γδ T Lymphocytes
Authors: Naiman Brian M. , Alt David , Bolin Carole A. , Zuerner Richard , Baldwin Cynthia L.
Published: July 2002

Leptospira borgpeterseniiserovar hardjo is the most common cause of bovine leptospirosis and also causes zoonotic infections of humans. A protective killed vaccine against serovar hardjo was shown to induce strong antigen-specific proliferative responses by peripheral blood mononuclear cells (PBMC) from vaccinated cattle by 2 months after the first dose of vaccine. This response was absent from nonvaccinated control cattle. The mean response peaked by 2 months after completion of the two-dose vaccination regimen, and substantial proliferation was measured in in vitro cultures throughout the 7 months of the study period. Variations in magnitude of the response occurred among the vaccinated animals, but by 7 months postvaccination there was a substantial antigen-specific response with PBMC from all vaccinated animals. Up to one-third of the PBMC from vaccinated animals produced gamma interferon (IFN-γ) after 7 days in culture with antigen, as ascertained by flow cytometric analysis, and significant levels of IFN-γ were measured in culture supernatants by enzyme-linked immunosorbent assay. Two-color immunofluorescence revealed that one-third of the IFN-γ-producing cells were γδ T cells, with the remaining cells being CD4+T cells. The significance of this study is the very potent Th1-type immune response induced and sustained following vaccination with a killed bacterial vaccine adjuvanted with aluminum hydroxide and the involvement of γδ T cells in the response. Moreover, induction of this Th1-type cellular immune response is associated with the protection afforded by the bovine leptospiral vaccine againstL. borgpeterseniiserovar hardjo.
Protective Effect ofLactobacillus caseiStrain Shirota on Shiga Toxin-ProducingEscherichia coliO157:H7 Infection in Infant Rabbits
Authors: Ogawa Michinaga , Shimizu Kensuke , Nomoto Koji , Takahashi Masatoshi , Watanuki Masaaki , Tanaka Ryuichiro , Tanaka Tetsuya , Hamabata Takashi , Yamasaki Shinji , Takeda Yoshifumi
Published: July 2002

We examined colonization patterns of Shiga toxin-producingEscherichia coli(STEC), concentrations of Shiga toxins (Stxs) and specific immunoglobulin A (lgA) against Stxs and STEC bacterial cell surface antigen in various portions of the gastrointestinal tract in an infant rabbit infection model. After inoculation of 3-day-old infant rabbits with STEC strain 89020087 at low doses (∼103CFU/body), numbers of colonizing STEC bacteria and concentrations of Stxs in the intestine increased dramatically and the animals developed diarrhea within a couple of days after infection. Daily administration ofLactobacillus caseifrom the day of birth dramatically decreased the severity of diarrhea and lowered STEC colonization levels in the gastrointestinal tract 100-fold day 7 after infection. Both Stx1 and Stx2 concentrations in the intestines and histological damage to the intestinal mucus induced by STEC infection were decreased by the administration ofL. casei. Examination of the concentrations of volatile fatty acids and pH of the intestinal contents revealed that the protective effect ofL. caseiadministration against STEC infection was not due to fermented products such as lactic acid in the gastrointestinal tract. Administration ofL. caseiincreased levels of lgAs against Stx1, Stx2, and formalin-killed STEC cells in the colon approximately two-, four-, and threefold, respectively, compared with those of the untreated controls by day 7 after infection. These results suggest that administration ofL. caseistrain Shirota enhances the local immune responses to STEC cells and Stxs and leads to elimination of STEC and thus decreases Stx concentrations in the intestines.
Synthetic 6B Di-, Tri-, and Tetrasaccharide-Protein Conjugates Contain Pneumococcal Type 6A and 6B Common and 6B- Specific Epitopes That Elicit Protective Antibodies in Mice
Authors: Jansen Wouter T. M. , Hogenboom Sietske , Thijssen Mark J. L. , Kamerling Johannis P. , Vliegenthart Johannes F. G. , Verhoef Jan , Snippe Harm , Verheul André F. M.
Published: July 2002

The immunogenicity and protective capacity of
Streptococcus pneumoniae
6B capsular polysaccharide (PS)-derived synthetic phosphate-containing disaccharide (Rha-ribitol-
P
-), trisaccharide (ribitol-
P
-Gal-Glc-), and tetrasaccharide (Rha-ribitol-
P
-Gal-Glc-)-protein conjugates in rabbits and mice were studied. In rabbits, all saccharides conjugated to keyhole limpet hemocyanin (KLH) evoked high levels of pneumococcal (Pn) type 6B antibodies that facilitated type-specific phagocytosis. Unlike the disaccharide rabbit antisera, tri- and tetrasaccharide rabbit antisera also reacted with 6A PS in an enzyme-linked immunosorbent assay (ELISA) and promoted phagocytosis of 6A pneumococci. All these rabbit antisera passively protected mice against a Pn 6B challenge. The disaccharide conjugate-induced antiserum, however, failed to protect mice against a 6A challenge. In mice, phagocytic and protective anti-Pn 6B antibodies were only induced by the tetrasaccharide conjugate and not by PS 6B or PS 6B-protein conjugates. These antibodies did not cross-react with 6A PS in ELISA and were unable to phagocytize 6A pneumococci. In conclusion, the disaccharide and tetrasaccharide conjugates already contain epitopes capable of inducing 6B-specific, fully protective antibodies in rabbits and mice, respectively.
Efficacy of Two Alternate Vaccines Based onPlasmodium falciparumMerozoite Surface Protein 1 in anAotusChallenge Trial
Authors: Stowers Anthony W. , Cioce Vittoria , Shimp Richard L. , Lawson Mark , Hui George , Muratova Olga , Kaslow David C. , Robinson Robin , Long Carole A. , Miller Louis H.
Published: July 2002

In an attempt to produce a more defined, clinical-grade version of a vaccine based onPlasmodium falciparummerozoite surface protein 1 (MSP1), we evaluated the efficacy of two recombinant forms of MSP1 in anAotus nancymaichallenge model system. One recombinant vaccine, bvMSP142, based on the 42-kDa C-terminal portion of MSP1, was expressed as a secreted protein in baculovirus-infected insect cells. A highly pure baculovirus product could be reproducibly expressed and purified at yields in excess of 8 mg of pure protein per liter of culture. This protein, when tested for efficacy in theAotuschallenge model, gave significant protection, with only one of seven monkeys requiring treatment for uncontrolled parasitemia after challenge withP. falciparum. The second recombinant protein, P30P2MSP119, has been used in previous studies and is based on the smaller, C-terminal 19-kDa portion of MSP1 expressed inSaccharomyces cerevisiae. Substantial changes were made in its production process to optimize expression. The optimum form of this vaccine antigen (as judged by in vitro and in vivo indicators) was then evaluated, along with bvMSP142, for efficacy in theA. nancymaisystem. The new formulation of P30P3MSP119performed significantly worse than bvMSP142and appeared to be less efficacious than we have found in the past, with four of seven monkeys in the vaccinated group requiring treatment for uncontrolled parasitemia. With both antigens, protection was seen only when high antibody levels were obtained by formulation of the vaccines in Freund's adjuvant. Vaccine formulation in an alternate adjuvant, MF59, resulted in significantly lower antibody titers and no protection.
Analysis of Transcriptionally Active Gene Clusters of Major Outer Membrane Protein Multigene Family inEhrlichia canisandE. chaffeensis
Authors: Ohashi Norio , Rikihisa Yasuko , Unver Ahmet
Published: July 2002

Ehrlichia canisandE. chaffeensisare tick-borne obligatory intramonocytic ehrlichiae that cause febrile systemic illness in humans and dogs, respectively. The current study analyzed the pleomorphic multigene family encoding approximately 30-kDa major outer membrane proteins (OMPs) ofE. canisandE. chaffeensis. Upstream fromsecAand downstream of hypothetical transcriptional regulator, 22 paralogs of theompgene family were found to be tandemly arranged except for one or two genes with opposite orientations in a 28- and a 27-kb locus in theE. canisandE. chaffeensisgenomes, respectively. Each locus consisted of three highly repetitive regions with four nonrepetitive intervening regions.E. canis, in addition, had a 6.9-kb locus which contained a repeat of three tandem paralogs in the 28-kb locus. These total 47 paralogous and orthologous genes encoded OMPs of approximately 30 to 35 kDa consisting of several hypervariable regions alternating with conserved regions. In the 5′-end half of the 27-kb locus or the 28-kb locus of eachEhrlichiaspecies, 14 paralogs were linked by short intergenic spaces ranging from −8 bp (overlapped) to 27 bp, and 8 remaining paralogs in the 3′-end half were connected by longer intergenic spaces ranging from 213 to 632 bp. All 22 paralogs, five unknown genes, andsecAin theompcluster inE. caniswere transcriptionally active in the monocyte culture, and the paralogs with short intergenic spaces were cotranscribed with their adjacent genes, including the respective intergenic spaces at both the 5′ and the 3′ sides. Althoughompgenes are diverse, our results suggest that the gene organization of the clusters and the gene locus are conserved between two species ofEhrlichiato maintain a unique transcriptional mechanism for adaptation to environmental changes common to them.
Different Subsets of Enteric Bacteria Induce and Perpetuate Experimental Colitis in Rats and Mice
Authors: Rath Heiko C. , Schultz Michael , Freitag René , Dieleman Levinus A. , Li Fengling , Linde Hans-Jörg , Schölmerich Jürgen , Sartor R. Balfour
Published: July 2002

Resident bacteria are incriminated in the pathogenesis of experimental colitis and inflammatory bowel diseases. We investigated the relative roles of various enteric bacteria populations in the induction and perpetuation of experimental colitis. HLA-B27 transgenic rats received antibiotics (ciprofloxacin, metronidazole, or vancomycin-imipenem) in drinking water or water alone in either prevention or treatment protocols. Mice were treated similarly with metronidazole or vancomycin-imipenem before or after receiving 5% dextran sodium sulfate (DSS). Germfree transgenic rats were colonized with specific-pathogen-free enteric bacteria grown overnight either in anaerobic or aerobic atmospheres. Nontransgenic rats colonized with anaerobic bacteria served as negative controls. Although preventive metronidazole significantly attenuated colitis in transgenic rats and DSS-treated mice, it had no therapeutic benefit once colitis was established. Ciprofloxacin also partially prevented but did not treat colitis in B27 transgenic rats. In both animal models vancomycin-imipenem most effectively prevented and treated colitis. Germfree transgenic rats reconstituted with enteric bacteria grown under anaerobic conditions had more aggressive colitis than those associated with aerobic bacteria. These results suggest that a subset of resident luminal bacteria induces colitis, but that a complex interaction of commensal aerobic and anaerobic bacteria provides the constant antigenic drive for chronic immune-mediated colonic inflammation.
Profiles of Immunoglobulin M (IgM) and IgG Antibodies against Defined Carbohydrate Epitopes in Sera ofSchistosoma-Infected Individuals Determined by Surface Plasmon Resonance
Authors: van Remoortere Alexandra , van Dam Govert J. , Hokke Cornelis H. , van den Eijnden Dirk H. , van Die Irma , Deelder AndréM.
Published: July 2002

We report here that sera of children and adults infected withSchistosoma mansoni, S. haematobium, orS. japonicumcontain antibodies against GalNAcβ1-4(Fucα1-2Fucα1-3)GlcNAc (LDN-DF) and to a lesser extent to Galβ1-4(Fucα1-3)GlcNAc (Lewisx) and GalNAcβ1-4GlcNAc (LDN). Surface plasmon resonance (SPR) spectroscopy was used to monitor the presence of serum antibodies to neoglycoconjugates containing these carbohydrate epitopes and to define the immunoglobulin M (IgM) and IgG subclass distribution of the antibodies. The serum levels of antibodies to LDN-DF are high related to LDN and Lewisxfor all examined groups ofSchistosoma-infected individuals. A higher antibody response to the LDN-DF epitope was found in sera of infected children than in sera of infected adults regardless of the schistosome species. With respect to the subclasses, we found surprisingly that individuals infected withS. japonicumhave predominantly IgG antibodies, while individuals infected withS. mansonimainly show an IgM response; high levels of both isotypes were measured in sera of individuals infected withS. haematobium. These data provide new insights in the human humoral immune response to schistosome-derived glycans.
Antibody Recognition of Rodent Malaria Parasite Antigens Exposed at the Infected Erythrocyte Surface: Specificity of Immunity Generated in Hyperimmune Mice
Authors: Mota Maria M. , Brown K. Neil , Do Rosário Virgilio E. , Holder Anthony A. , Jarra William
Published: July 2002

In regions where malaria is endemic, inhabitants remain susceptible to repeated reinfection as they develop and maintain clinical immunity. This immunity includes responses to surface-exposed antigens onPlasmodiumsp.-infected erythrocytes. Some of these parasite-encoded antigens may be diverse and phenotypically variable, and the ability to respond to this diversity and variability is an important component of acquired immunity. Characterizing the relative specificities of antibody responses during the acquisition of immunity and in hyperimmune individuals is thus an important adjunct to vaccine research. This is logistically difficult to do in the field but is relatively easily carried out in animal models. Infections in inbred mice with rodent malaria parasitePlasmodium chabaudi chabaudiAS represent a good model forPlasmodium falciparumin humans. This model has been used in the present study in a comparative analysis of cross-reactive and specific immune responses in rodent malaria. CBA/Ca mice were rendered hyperimmune toP. chabaudi chabaudi(AS or CB lines) orPlasmodium berghei(KSP-11 line) by repeated infection with homologous parasites. Serum fromP. chabaudi chabaudiAS hyperimmune mice reacted with antigens released from disruptedP. chabaudi chabaudiAS-infected erythrocytes, butP. chabaudi chabaudiCB andP. bergheiKSP-11 hyperimmune serum also contained cross-reactive antibodies to these antigens. However, antibody activity directed against antigens exposed at the surfaces of intactP. chabaudi chabaudi-infected erythrocytes was mainly parasite species specific and, to a lesser extent, parasite line specific. Importantly, this response included opsonizing antibodies, which bound to infected erythrocytes, leading to their phagocytosis and destruction by macrophages. The results are discussed in the context of the role that antibodies to both variable and invariant antigens may play in protective immunity in the face of continuous susceptibility to reinfection.
Protective Role of Lung Surfactant Protein D in a Murine Model of Invasive Pulmonary Aspergillosis
Authors: Madan Taruna , Kishore Uday , Singh Mamta , Strong Peter , Hussain Ejaj M. , Reid Kenneth B. M. , Sarma P. Usha
Published: July 2002

The protective effects of intranasal administration of amphotericin B (AmB), human SP-A, SP-D and a 60-kDa fragment of SP-D (rSP-D) were examined in a murine model of invasive pulmonary aspergillosis (IPA). The untreated group of IPA mice showed no survival at 7 days postinfection. Treatment with AmB, SP-D, and rSP-D increased the survival rate to 80, 60, and 80%, respectively, suggesting that SP-D (and rSP-D) can protect immunosuppressed mice from an otherwise fatal challenge with
Aspergillus fumigatus
conidia.
Interleukin-10 Regulates the Tissue Factor Activity of Monocytes in an In Vitro Model of Bacterial Endocarditis
Authors: Veltrop Marcel H. A. M. , Langermans Jan A. M. , Thompson Jan , Bancsi Maurice J. L. M. F.
Published: July 2002

Monocytes are important effector cells in the pathogenesis of bacterial endocarditis since they provide the tissue factor that activates the coagulation system and maintains established vegetations. Monocytes secrete cytokines that can modulate monocyte tissue factor activity (TFA), thereby affecting the formation and maintenance of vegetations. In this study, we show that monocytes cultured for 4 h on aStreptococcus sanguis-infected fibrin matrix mimicking the in vivo vegetational surface express high levels of TFA. This was accompanied by secretion of the proinflammatory cytokines tumor necrosis factor alpha (TNF-α), interleukin-1α (IL-1α), and IL-1β. After a 24-h incubation period the anti-inflammatory cytokine IL-10 could also be detected. Our data show that, whereas TNF-α and IL-1 have a minor role in the induction of TFA by monocytes cultured on a fibrin matrix, TNF-α but not IL-1 plays an important role in the induction of IL-10 by these cells. In turn, our data show that IL-10 is an important factor in the downregulation of monocyte TFA. In summary, we conclude that IL-10 is an important factor in the control of monocyte TFA in endocardial vegetations.
Vaccination against the Intracellular PathogensLeishmania majorandL. amazonensisby Directing CD40 Ligand to Macrophages
Authors: Chen Gang , Darrah Patricia A. , Mosser David M.
Published: July 2002

CD40 ligand (CD40L) is a potent inducer of interleukin-12 (IL-12) production from macrophages and dendritic cells. We show that combining CD40L with antigen derived fromLeishmaniais an effective way to preferentially induce type 1 immune responses to the antigen and to vaccinate mice against subsequent challenge with virulent organisms. Mice vaccinated in this way had smaller lesions, with more than 1,000-fold fewer parasites within them. To improve the efficiency of CD40L-induced immunopotentiation, we attempted to specifically direct CD40L to macrophages. We developed transfected cells expressing CD40L and a singleLeishmaniaantigen, gp63. These cells bound efficiently to macrophages and induced robust IL-12 production. Vaccination with these cotransfected cells provided a significant degree of protection against challenge with virulent organisms. CD40L was also adsorbed to the surface of virulentLeishmania. These organisms induced only modest lesions in genetically susceptible mice, and the lesions had an average of 105-fold fewer organisms within them relative to control mice. These studies suggest that CD40L could be exploited to improve vaccines against intracellular pathogens, especially those organisms that reside within cells expressing CD40 on their surface.
B Cells and Antibodies Are Required for Resistance to the Parasitic Gastrointestinal NematodeTrichuris muris
Authors: Blackwell Nathan M. , Else Kathryn J.
Published: July 2002

Previous studies using cell transfers and antibody receptor knockout mice have shown that B cells and antibodies are not essential components of the expulsion mechanism inTrichuris murisinfections. Serum transfer experiments have given mixed results regarding the importance of antibodies in this infection model, and the role of B cells in initiating or maintaining T-cell responses has not been addressed. We used B-cell-deficient μMT mice to determine if B cells play a role in anti-T. murisimmune responses. In contrast to wild-type C57BL/6 mice, μMT mice were susceptible to infection. Antigen-restimulated mesenteric lymph node cells from infected μMT mice produced only naive levels of Th2-associated cytokines but had increased levels of gamma interferon. However, these mice appeared capable of mounting a Th2-dependent mucosal mastocytosis, though this was significantly delayed compared to that seen in wild-type mice. Resistance toT. muriswas restored following reconstitution with naive C57BL/6 splenic B cells, as was in vitro Th2 cytokine production in response to parasite antigen. Treatment of μMT mice with anti-interleukin-12 monoclonal antibody during the first 2 weeks of infection also restored immunity, suggesting that μMT mice can be manipulated to expel worms at the time of T-cell priming. Additionally, treatment of μMT mice with parasite-specific immunoglobulin G1 purified from the serum of resistant NIH mice prevented worm establishment, suggesting an important role for antibodies. Our results as a whole describe the first detailed report of a critical role for B cells in resistance to an intestinal nematode.
Brucella abortus
Cyclic β-1,2-Glucan Mutants Have Reduced Virulence in Mice and Are Defective in Intracellular Replication in HeLa Cells
Authors: Briones Gabriel , Iñón de Iannino Nora , Roset Mara , Vigliocco Ana , Paulo Patricia Silva , Ugalde Rodolfo A.
Published: July 2002

Null cyclic β-1,2-glucan synthetase mutants (
cgs
mutants) were obtained from
Brucella abortus
virulent strain 2308 and from
B. abortus
attenuated vaccinal strain S19. Both mutants show greater sensitivity to surfactants like deoxycholic acid, sodium dodecyl sulfate, and Zwittergent than the parental strains, suggesting cell surface alterations. Although not to the same extent, both mutants display reduced virulence in mice and defective intracellular multiplication in HeLa cells. The
B. abortus
S19
cgs
mutant was completely cleared from the spleens of mice after 4 weeks, while the 2308 mutant showed a 1.5-log reduction of the number of brucellae isolated from the spleens after 12 weeks. These results suggest that cyclic β-1,2-glucan plays an important role in the residual virulence of the attenuated
B. abortus
S19 strain. Although the
cgs
mutant was cleared from the spleens earlier than the wild-type parental strain (
B. abortus
S19) and produced less inflammatory response, its ability to confer protection against the virulent strain
B. abortus
2308 was fully retained. Equivalent levels of induction of spleen gamma interferon mRNA and anti-lipopolysaccharide (LPS) of immunoglobulin G2a (IgG2a) subtype antibodies were observed in mice injected with
B. abortus
S19 or the
cgs
mutant. However, the titer of anti-LPS antibodies of the IgG1 subtype induced by the
cgs
mutant was lower than that observed with the parental S19 strain, thus suggesting that the
cgs
mutant induces a relatively exclusive Th1 response.
Immunoglobulin A-Mediated Protection againstBordetella pertussisInfection
Authors: Hellwig Sandra M. M. , van Spriel Annemiek B. , Schellekens Joop F. P. , Mooi Frits R. , van de Winkel Jan G. J.
Published: July 2002

Infection withBordetella pertussis, the causative agent of pertussis (whooping cough) in humans, is followed by the production of antibodies of several isotypes, including immunoglobulin A (IgA). Little is known, however, about the role of IgA in immunity against pertussis. Therefore, we studied targeting ofB. pertussisto the myeloid receptor for IgA, FcαRI (CD89), using either IgA purified from immune sera of pertussis patients or bispecific antibodies directed againstB. pertussisand FcαRI (CD89 BsAb). Both IgA and CD89 BsAb facilitated FcαRI-mediated binding, phagocytosis, and bacterial killing by human polymorphonuclear leukocytes (PMNL) and PMNL originating from human FcαRI-transgenic mice. Importantly, FcαRI targeting resulted in enhanced bacterial clearance in lungs of transgenic mice. These data support the capacity of IgA to induce anti-B. pertussiseffector functions via the myeloid IgA receptor, FcαRI. Increasing the amount of IgA antibodies induced by pertussis vaccines may result in higher vaccine efficacy.