bluuebunny/crossref_metadata_2025 huggingface dataset:
Aggregation ofCryptococcus neoformansby Surfactant Protein D Is Inhibited by Its Capsular Component Glucuronoxylomannan
Authors: van de Wetering J. K. , Coenjaerts F. E. J. , Vaandrager A. B. , van Golde L. M. G. , Batenburg J. J.
Published: December 2003

Cryptococcus neoformansis an opportunistic pathogen invading the immunocompromised host. Infection starts with the inhalation of acapsular or sparsely encapsulated cells, after which capsule synthesis is initiated. The capsule is the main virulence factor of this yeast-like fungus. Pulmonary surfactant protein D (SP-D) is an important component of the local innate defense system. In the present study, interactions of SP-D with intactC. neoformanscells and their isolated capsular components were investigated. Although encapsulated cryptococci were bound, SP-D showed the highest affinity for acapsularC. neoformans. Only acapsular cryptococci were aggregated by SP-D. Furthermore, the cryptococcal capsular components glucuronoxylomannan (GXM) and mannoprotein 1 (MP1) were bound with relatively high affinity, in contrast to GalXM and MP2. Binding as well as aggregation of acapsularC. neoformansby SP-D could be inhibited by GXM in concentrations that are likely to be present in the lung after infection, suggesting that not only the capsule hampers SP-D function within the innate defense system of the lung but also the secreted capsular component GXM.
Apical Membrane Antigen 1, a Major Malaria Vaccine Candidate, Mediates the Close Attachment of Invasive Merozoites to Host Red Blood Cells
Authors: Mitchell G. H. , Thomas A. W. , Margos G. , Dluzewski A. R. , Bannister L. H.
Published: December 2003

Apical membrane antigen 1 (AMA-1) of
Plasmodium
merozoites is established as a candidate molecule for inclusion in a human malaria vaccine and is strongly conserved in the genus. We have investigated its function in merozoite invasion by incubating
Plasmodium knowlesi
merozoites with red cells in the presence of a previously described rat monoclonal antibody (MAb R31C2) raised against an invasion-inhibitory epitope of
P. knowlesi
AMA-1 and then fixing the material for ultrastructural analysis. We have found that the random, initial, long-range (12 nm) contact between merozoites and red cells occurs normally in the presence of the antibody, showing that AMA-1 plays no part in this stage of attachment. Instead, inhibited merozoites fail to reorientate, so they do not bring their apices to bear on the red cell surface and do not make close junctional apical contact. We conclude that AMA-1 may be directly responsible for reorientation or that the molecule may initiate the junctional contact, which is then presumably dependent on Duffy binding proteins for its completion.
Delayed Clearance ofEhrlichia chaffeensisInfection in CD4+T-Cell Knockout Mice†
Authors: Ganta Roman R. , Cheng Chuanmin , Wilkerson Melinda J. , Chapes Stephen K.
Published: December 2003

Human monocytic ehrlichiosis is an emerging tick-borne disease caused by the rickettsiaEhrlichia chaffeensis. To examine the role of helper T cells in host resistance to this macrophage-tropic bacterium, we assessedE. chaffeensisinfections in three mouse strains with differing functional levels of helper T cells. Wild-type, C57BL/6J mice resolved infections in approximately 2 weeks. Major histocompatibility complex class II (MHCII) knockout, B6.129-Abbtm1mice lacking helper T cells developed persistent infections that were not resolved even after several months. CD4+T-cell-deficient, B6.129S6-Cd4tm1Knwmice cleared the infection, but the clearance took 2 weeks longer than it did for wild-type mice. C57BL/6J mice resolved infection more rapidly following a second experimental challenge, but B6.129S6-Cd4tm1Knwmice did not. The B6.129S6-Cd4tm1Knwmice also developed activeE. chaffeensis-specific immunoglobulin G responses that were slightly lower in concentration and slower to develop than that observed in C57BL/6J mice.E. chaffeensis-specific cytotoxic T cells were not detected following a single bacterial challenge in any mouse strain, including wild-type C57BL/6J mice. However, the cytotoxic T-cell activity developed in all three mouse strains, including the MHCII and CD4+T-cell knockouts, when challenged with a secondE. chaffeensisinfection. The data reported here suggest that the cell-mediated immunity, orchestrated by CD4+T cells is critical for conferring rapid clearance ofE. chaffeensis.
Plant Cell-Based Intimin Vaccine Given Orally to Mice Primed with Intimin Reduces Time of
Escherichia coli
O157:H7 Shedding in Feces
Authors: Judge Nicole A. , Mason Hugh S. , O'Brien Alison D.
Published: December 2003

Intimin is the primary adhesin of
Escherichia coli
O157:H7, the most common infectious cause of bloody diarrhea in the United States and the leading cause of acute kidney failure in children who develop hemolytic uremic syndrome. Cattle are the primary reservoir of
E. coli
O157:H7. Indeed, most cases of
E. coli
O157:H7 infection in the United States occur after ingestion of contaminated undercooked hamburger or produce that had contact with bovine manure. Because intimin is required for persistent colonization of neonatal calves and adult cattle, we hypothesized that an intimin-based vaccination strategy in calves would reduce colonization of cattle with
E. coli
O157:H7. To test this concept in a small-animal model, we developed transgenic tobacco plant cells that express the carboxy-terminal host cell-binding domain of
E. coli
O157:H7 intimin. Mice were either immunized intraperitoneally with intimin expressed from the plant cells, fed transgenic plant cells, or both. Here we show that these mice generated an intimin-specific mucosal immune response when primed parenterally and then boosted orally and also exhibited a reduced duration of
E. coli
O157:H7 fecal shedding after challenge.
Role of Toll-Like Receptor 4 in Induction of Cell-Mediated Immunity and Resistance toBrucella abortusInfection in Mice
Authors: Campos Marco A. , Rosinha Gracia M. S. , Almeida Igor C. , Salgueiro Xirlene S. , Jarvis Bruce W. , Splitter Gary A. , Qureshi Nilofer , Bruna-Romero Oscar , Gazzinelli Ricardo T. , Oliveira Sergio C.
Published: December 2003

Initial host defense to bacterial infection is executed by innate immunity, and therefore the main goal of this study was to examine the contribution of Toll-like receptors (TLRs) duringBrucella abortusinfection. CHO reporter cell lines transfected with CD14 and TLRs showed thatB. abortustriggers both TLR2 and TLR4. In contrast, lipopolysaccharide (LPS) and lipid A derived fromBrucellarough (R) and smooth (S) strains activate CHO cells only through TLR4. Consistently, macrophages from C3H/HePas mice exposed to R and S strains and their LPS produced higher levels of tumor necrosis factor alpha (TNF-α) and interleukin-12 compared to C3H/HeJ, a TLR4 mutant mouse. The essential role of TLR4 for induction of proinflammatory cytokines was confirmed with diphosphoryl lipid A fromRhodobacter sphaeroides. Furthermore, to determine the contribution of TLR2 and TLR4 in bacterial clearance, numbers ofBrucellawere monitored in the spleen of C3H/HeJ, C3H/HePas, TLR2 knockout, and wild-type mice at 1, 3, and 6 weeks followingB. abortusinfection. Interestingly, murine brucellosis was markedly exacerbated at weeks 3 and 6 after infection in animals that lacked functional TLR4 (C3H/HeJ) compared to C3H/HePas that paralleled the reduced gamma interferon production by this mouse strain. Finally, by mass spectrometry analysis we found dramatic differences on the lipid A profiles of R and S strains. In fact, S lipid A was shown to be more active to trigger TLR4 than R lipid A in CHO cells and more effective in inducing dendritic cell maturation. In conclusion, these results indicate that TLR4 plays a role in resistance toB. abortusinfection and that S lipid A has potent adjuvant activity.
Immunization of Female Mice with Glycoconjugates Protects Their Offspring against Encapsulated Bacteria
Authors: Richter Margret Y. , Jakobsen Håvard , Birgisdottir Alda , Haeuw Jean-François , Power Ultan F. , Del Giudice Giuseppe , Bartoloni Antonella , Jonsdottir Ingileif
Published: December 2003

The immune system of the newborn is immature, and therefore it is difficult to induce protective immunity by vaccination in the neonatal period. Immunization of mothers during pregnancy against infections caused by encapsulated bacteria could thus be particularly attractive, as infants do not respond to polysaccharide (PS) antigens. Transmission of maternal vaccine-specific antibodies and protection of offspring against pneumococcal bacteremia and/or lung infection were studied in a neonatal murine model of pneumococcal immunization and infections. Adult female mice were immunized with native pneumococcal PS (PPS) of serotypes 1, 6B, and 19F or PPS conjugated to tetanus protein (Pnc-TT), and PPS-specific antibodies were measured in sera of mothers and their offspring. Effective transmission of maternal antibodies was observed, as PPS-specific immunoglobulin G levels in 3-week-old offspring of immunized mothers were 37 to 322% of maternal titers, and a significant correlation between maternal and offspring antibody levels was observed. The PPS-specific antibodies persisted for several weeks but slowly decreased over time. Offspring of Pnc-TT-immunized mothers were protected against pneumococcal infections with homologous serotypes, whereas PPS immunization of mothers did not protect their offspring, in agreement with the low titer of maternal PPS specific antibodies. When adult female mice were immunized with a meningococcal serogroup C conjugate vaccine (MenC-CRM), antibody response and transmission were similar to those observed for pneumococcal antibodies. Importantly, bactericidal activity was demonstrated in offspring of MenC-CRM-immunized mothers. These results demonstrate that this murine model of pneumococcal immunization and infections is suitable to study maternal immunization strategies for protection of offspring against encapsulated bacteria.
Immunogenicity and Efficacy ofCryptococcus neoformansCapsular Polysaccharide Glucuronoxylomannan Peptide Mimotope-Protein Conjugates in Human Immunoglobulin Transgenic Mice
Authors: Maitta Robert W. , Datta Kausik , Lees Andrew , Belouski Shelley Sims , Pirofski Liise-anne
Published: December 2003

Peptide mimotopes of capsular polysaccharides have been proposed as antigens for vaccines against encapsulated pathogens. In this study, we determined the antibody response to and efficacy of P13, a peptide mimetic of theCryptococcus neoformanscapsular polysaccharide glucuronoxylomannan (GXM), in mice that produce human antibodies. P13 was conjugated to tetanus toxoid (TT) or diphtheria toxoid (DT) and administered subcutaneously in Alhydrogel with or without CpG to mice transgenic for human immunoglobulin loci (XenoMouse mice) and expressing either immunoglobulin G2 (IgG2) (G2 mice) or IgG4 (G4 mice). Mice were vaccinated and revaccinated two or three times. The serum antibody responses of the mice to GXM and P13 and antibody idiotype expression were analyzed by an enzyme-linked immunosorbent assay. The results showed that both P13-TT and P13-DT were antigenic, inducing a mimetic response to P13 in both G2 and G4 mice, and immunogenic, inducing a mimotope response including VH3 (idiotype)-positive antibodies to GXM in G2 but not G4 mice. CpG led to higher titers of IgG to P13 and GXM in P13-TT-vaccinated G2 mice.C. neoformanschallenge of P13-protein conjugate-vaccinated and control G2 mice induced anamnestic IgG- and VH3-positive responses to GXM and was associated with a significantly decreased risk of death and a prolongation of survival in P13-DT-vaccinated mice compared to phosphate-buffered saline-treated or protein carrier-vaccinated mice. These findings reveal that P13 elicited a human antibody response with VH3 expression in human immunoglobulin transgenic mice that has been observed for human antibodies to GXM and support the concept that peptide mimotope-based vaccines may hold promise for the treatment ofC. neoformansinfections.
Biological Activities of Naturally Occurring Antibodies Reactive withCandida albicansMannan
Authors: Kozel Thomas R. , MacGill Randall S. , Percival Ann , Zhou Qing
Published: December 2003

Sera from normal adult humans may contain high levels of antibody reactive withCandida albicansmannan. This study examined selected biological activities of such antibodies, focusing on sera that were collected from 34 donors and analyzed individually. The results showed that antimannan titers were normally distributed. Reactivity as determined by enzyme-linked immunosorbent assay with serotype A mannan generally paralleled reactivity with serotype B. Analysis of the kinetics for activation of the complement system and deposition of complement component 3 (C3) onto serotype A and serotype B cells showed a decrease in the lag time that occurred before the onset of rapid accumulation of C3 that correlated with increasing antimannan titers. In contrast, there was a decrease in the overall rate of accumulation of C3 on serotype A cells that was strongly correlated with increasing antibody titers; serotype B cells showed no such decrease. An evaluation of the contribution of mannan antibody to opsonophagocytic killing showed that mannan antibody in individual sera and antimannan immunoglobulin G (IgG) affinity purified from human plasma contributed to killing by neutrophils in a dose-dependent fashion in the absence of a functional complement system. However, affinity-purified antibody in very high concentrations was inhibitory to both complement-dependent and complement-independent opsonophagocytosis, and this finding suggests a prozone-like effect. In contrast, if the complement system was functional, antimannan IgG was not needed for opsonophagocytic killing. These results suggest that naturally occurring mannan antibodies and the complement system are functionally redundant for opsonophagocytic killing by neutrophils.
Camelysin Is a Novel Surface Metalloproteinase from
Bacillus cereus
Authors: Grass Gregor , Schierhorn Angelika , Sorkau Eduard , Müller Helmut , Rücknagel Peter , Nies Dietrich H. , Fricke Beate
Published: December 2003

Bacillus cereus
frequently causes food poisoning or nosocomial diseases. Vegetative cells express the novel surface metalloproteinase camelysin (casein-cleaving metalloproteinase) during exponential growth on complex, peptide-rich media. Camelysin is strongly bound to the cell surface and can be solubilized only by detergents or butanol. Camelysin spontaneously migrates from the surface of intact bacterial cells to preformed liposomes. The complete sequence of the camelysin-encoding gene,
calY
, was determined by reverse PCR on the basis of the N-terminal sequence and some internal tryptic cleavage peptides. The
calY
gene codes for a polypeptide of 21.569 kDa with a putative signal peptide of 27 amino acids (2.513 kDa) preceding the mature protein (19.056 kDa). Although the predicted amino acid sequence of CalY does not exhibit a typical metalloprotease consensus sequence, high-pressure liquid chromatography-purified camelysin contains one zinc ion per protein molecule. Matrix-assisted laser desorption ionization-time-of-flight mass spectrometry and tryptic peptide mass fingerprinting confirmed the identity of this zinc-binding protein as CalY. Disruption of the
calY
gene results in a strong decrease in the cell-bound proteolytic activity on various substrates.
Salmonella enterica
Serovar Paratyphi C Carries an Inactive Shufflon
Authors: Tam Connie K. P. , Hackett Jim , Morris Christina
Published: December 2003

Salmonella enterica
serovar Typhi uses type IVB pili to facilitate bacterial self-association, but only when the PilV proteins (potential minor pilus proteins) are not synthesized. This pilus-mediated event may be important in typhoid fever pathogenesis. We initially show that
S. enterica
serovar Paratyphi C strains harbor a
pil
operon very similar to that of serovar Typhi. An important difference, however, is located in the shufflon which concludes the
pil
operon. In serovar Typhi, the Rci recombinase acts upon two 19-bp inverted repeats to invert the terminal region of the
pilV
gene, thereby disrupting PilV synthesis and permitting bacterial self-association. In serovar Paratyphi C, however, the shufflon is essentially inactive because each of the Rci 19-bp substrates has acquired a single base pair insertion. A PilV protein is thus synthesized whenever the
pil
operon is active, and bacterial self-association therefore does not occur in serovar Paratyphi C. The data thus suggest that serovar Typhi bacterial self-association using type IVB pili may be important in the pathogenesis of epidemic enteric fever.
Melanin fromFonsecaea pedrosoiInduces Production of Human Antifungal Antibodies and Enhances the Antimicrobial Efficacy of Phagocytes
Authors: Alviano Daniela S. , Franzen Anderson J. , Travassos Luiz R. , Holandino Carla , Rozental Sonia , Ejzemberg Regina , Alviano Celuta S. , Rodrigues Marcio L.
Published: December 2003

Fonsecaea pedrosoiis a fungal pathogen that produces melanin. The functions of melanin and its possible influence in the protective immunological response during infection byF. pedrosoiare not known. In this work, treatment ofF. pedrosoimycelia with proteases and glycosidases followed by a denaturing agent and hot concentrated acid left a black residue. Scanning electron microscopy demonstrated that this processed melanized residue resembled very closely the intact mycelium in shape and size. Melanin particles were also isolated from culture fluids of conidia or sclerotic forms ofF. pedrosoi. Secreted melanins were reactive with sera from infected human patients, suggesting thatF. pedrosoisynthesizes melanin in vivo. The antibodies against melanin were purified from patients’ sera and analyzed by indirect immunofluorescence. They reacted with sclerotic cells from patients’ lesions as well as with sclerotic bodies cultivated in vitro, conidia, mycelia, and digested residues. Treatment ofF. pedrosoiwith purified antibodies against melanin inhibited fungal growth in vitro. The interaction ofF. pedrosoiwith phagocytes in the presence of melanin resulted in higher levels of fungal internalization and destruction by host cells, which was accompanied by greater degrees of oxidative burst. Taken together, these results indicate that melanin fromF. pedrosoiis an immunologically active fungal structure that activates humoral and cellular responses that could help the control of chromoblastomycosis by host defenses.
Single Intranasal MucosalMycobacterium bovisBCG Vaccination Confers Improved Protection Compared to Subcutaneous Vaccination against Pulmonary Tuberculosis
Authors: Chen Lihao , Wang Jun , Zganiacz Anna , Xing Zhou
Published: December 2003

Whether the intranasal (i.n.) route ofMycobacterium bovisBCG vaccination provides better protection against pulmonary tuberculosis than subcutaneous (s.c.) vaccination remains an incompletely solved issue. In the present study, we compared both immune responses and protection elicited by single BCG vaccinations via the i.n. or s.c. route in BALB/c mice. While both i.n. and s.c. vaccination triggered comparable levels of primary immune activation in the spleen and draining lymph nodes, i.n. vaccination led to a greater antigen-specific gamma interferon recall response in splenocytes than s.c. vaccination upon secondary respiratory mycobacterial challenge, accompanied by an increased frequency of antigen-specific lymphocytes. There was also a quicker cellular response in the lungs of i.n. vaccinated mice upon mycobacterial challenge. Mice vaccinated i.n. were found to be much better protected, particularly in the lung, than s.c. vaccinated counterparts against pulmonary tuberculosis at both 3 and 6 months postvaccination. These results suggest that the i.n. route of vaccination improves the protective effect of the current BCG vaccine.
Association between Protection against Clinical Malaria and Antibodies to Merozoite Surface Antigens in an Area of Hyperendemicity in Myanmar: Complementarity between Responses to Merozoite Surface Protein 3 and the 220-Kilodalton Glutamate-Rich Protein
Authors: Soe Soe , Theisen Michael , Roussilhon Christian , Aye Khin-Saw- , Druilhe Pierre
Published: December 2003

We performed a longitudinal clinical and parasitological follow-up study in OoDo, a village in southeast Asia in which malaria is hyperendemic, in order to assess the association between protection against malaria attacks and antibodies to three currently evaluated vaccine candidates, merozoite surface protein 1 (MSP1), MSP3, and the 220-kDa glutamate-rich protein (GLURP) from
Plasmodium falciparum.
Our results showed that the levels of cytophilic immunoglobulin G3 (IgG3) antibodies against conserved regions of MSP3 and GLURP were significantly correlated with protection against clinical
P. falciparum
malaria. In contrast, the levels of noncytophilic IgG4 antibodies against GLURP increased with the number of malaria attacks. Furthermore, we observed a complementary effect of the MSP3- and GLURP-specific IgG3 antibodies in relation to malaria protection. In the individuals that did not respond to one of the antigens, a strong response to the other antigen was consistently detected and was associated with protection, suggesting that induction of antibodies against both MSP3 and GLURP could be important for the development of protective immunity. The complementarity of the responses to the two main targets of antibody-dependent cellular inhibition identified to date provides the first rational basis for combining these two antigens in a hybrid vaccine formulation.
Induction of
Plasmodium falciparum
Transmission-Blocking Antibodies in Nonhuman Primates by a Combination of DNA and Protein Immunizations
Authors: Coban Cevayir , Philipp Mario T. , Purcell Jeanette E. , Keister David B. , Okulate Mobolaji , Martin Dale S. , Kumar Nirbhay
Published: December 2003

Malaria transmission-blocking vaccination can effectively reduce and/or eliminate transmission of parasites from the human host to the mosquito vector. The immunity achieved by inducing an antibody response to surface antigens of male and female gametes and parasite stages in the mosquito. Our laboratory has developed DNA vaccine constructs, based on Pfs25 (a
Plasmodium falciparum
surface protein of 25 kDa), that induce a transmission-blocking immune response in mice (C. A. Lobo, R. Dhar, and N. Kumar, Infect. Immun. 67:1688-1693, 1999). To evaluate the safety, immunogenicity, and efficacy of the Pfs25 DNA vaccine in nonhuman primates, we immunized rhesus macaques (
Macaca mulatta
) with a DNA vaccine plasmid encoding Pfs25 or a Pfg27-Pfs25 hybrid or with the plasmid (empty plasmid) alone. Immunization with four doses of these DNA vaccine constructs elicited antibody titers that were high but nonetheless unable to reduce the parasite's infectivity in membrane feeding assays. Further boosting of the antibody response with recombinant Pfs25 formulated in Montanide ISA-720 increased antibody titers (30-fold) and significantly blocked transmission of
P. falciparum
gametocytes to
Anopheles
mosquitoes (∼90% reduction in oocyst numbers in the midgut). Our data show that a DNA prime-protein boost regimen holds promise for achieving transmission-blocking immunity in areas where malaria is endemic and could be effective in eradicating malaria in isolated areas where the level of malaria endemicity is low.
l
-Serine Catabolism via an Oxygen-Labile
l
-Serine Dehydratase Is Essential for Colonization of the Avian Gut by
Campylobacter jejuni
Authors: Velayudhan Jyoti , Jones Michael A. , Barrow Paul A. , Kelly David J.
Published: December 2003

Campylobacter jejuni
is a microaerophilic, asaccharolytic bacterium. The identity of the carbon and energy sources used by
C. jejuni
in vivo is unknown, but the genome sequence of strain NCTC11168 indicates the presence of genes for catabolism of a limited range of amino acids, including serine. Specific omission of
l
-serine from a defined medium containing a mixture of amino acids led to a dramatic decrease in cell yields. As
C. jejuni
does not have a biosynthetic serine requirement, this supports earlier suggestions that
l
-serine is a preferentially catabolized amino acid. Serine transport was found to be mediated by at least two systems in strain 11168; a high-capacity, low-affinity
l
-serine-specific system encoded by
Cj1625c
(
sdaC
) and a higher-affinity
l
-serine/
l
-threonine system responsible for residual
l
-serine transport in an
sdaC
mutant. Catabolism of
l
-serine to pyruvate and ammonia is carried out by SdaA (encoded by
Cj1624c
), which was overexpressed, purified, and shown to be an oxygen-labile iron-sulfur enzyme.
l
-Serine dehydratase activity in an
sdaA
mutant was reduced 10-fold compared to that in the wild type, but the residual activity (due to the anabolic
l
-threonine dehydratase) could not support either growth on or utilization of
l
-serine in defined media. However, although
sdaA
mutants showed no obvious growth defect in complex media, they completely failed to colonize 3-week-old chickens as assayed both by cloacal swabs taken over a 6-week period and by cecal colony counts postmortem. In contrast, the isogenic parent strain colonized chickens to high levels within 1 week of inoculation. The results show that an active SdaA is essential for colonization of the avian gut by
C. jejuni
and imply that catabolism of
l
-serine is crucially important for the growth of this bacterium in vivo.
Tumor Necrosis Factor Alpha Enhances
Actinobacillus actinomycetemcomitans
Leukotoxin-Induced HL-60 Cell Apoptosis by Stimulating Lymphocyte Function-Associated Antigen 1 Expression†
Authors: Yamaguchi Noboru , Kubo Chie , Masuhiro Yoshikazu , Lally Edward T. , Koga Toshihiko , Hanazawa Shigemasa
Published: December 2003

We demonstrated previously that
Actinobacillus actinomycetemcomitans
leukotoxin (Ltx) is greatly able to induce apoptotic signaling in cells that are positive for lymphocyte function-associated antigen 1 (LFA-1), a cell receptor of Ltx. We investigated in this study whether inflammatory cytokines can regulate apoptosis of human leukemic HL-60 cells induced by Ltx. Of the cytokines tested, tumor necrosis factor alpha (TNF-α) significantly enhanced the Ltx-induced cell apoptosis. Northern and Western blotting analyses showed that TNF-α enhanced the expression of CD11a in the cells at both the mRNA and protein levels but did not do so for CD18 expression. TNF-α also enhanced the binding of Ltx to the cells. We also observed by measuring the mitochondrial transmembrane potential and the generation of superoxide anion that the cytokine enhanced Ltx-induced apoptosis in HL-60 cells. In addition, interleukin-1β significantly enhanced Ltx-induced cell apoptosis, although the enhancing activity was lower than that of TNF-α. These stimulatory effects of both cytokines were also observed for human polymorphonuclear leukocytes. The ability of TNF-α to increase cell susceptibility to Ltx could be inhibited by preincubation of the cells with a monoclonal antibody against TNF receptor 1 but not by preincubation of the cells with a monoclonal antibody against anti-TNF receptor 2. Furthermore, the results of an assay of caspase 3 intracellular activity (PhiPhiLuxG
1
D
2
) showed that Ltx-induced caspase 3 activation was completely neutralized by CD18 antibody treatment, although significant neutralization was also observed with anti-CD11a antibody. Taken together, the results of the present study indicate that TNF-α acts as a potent stimulator of Ltx-induced HL-60 cell apoptosis via TNF receptor 1-mediated upregulation of LFA-1 expression.
Immunogenicity of Gonococcal Transferrin Binding Proteins during Natural Infections
Authors: Price Gregory A. , Hobbs Marcia M. , Cornelissen Cynthia Nau
Published: December 2003

In this study, we examined the immune response during gonococcal infection to the individual transferrin binding proteins by using a quantitative enzyme-linked immunosorbent assay (ELISA). Recombinant transferrin binding protein A (rTbpA) and rTbpB were purified under nondenaturing conditions for use as ELISA antigens. Sera and secretions from culture-positive individuals were analyzed for antibodies to rTbpA and rTbpB and compared to samples from individuals with no history of gonococcal infection. Although antibodies to both rTbpA and rTbpB were detected in serum, in most cases the antibody levels were not significantly different from those measured in the control population. Also, previous history of gonococcal infection did not increase antibody levels in serum, suggesting the lack of an anamnestic response. Analysis of secretion samples revealed antibody levels that were generally below the limits of detection in our assay. Overall, this study demonstrated a paucity of systemic and local antibody responses to rTbps as a result of natural infection and represents a baseline over which a protective antibody response will have to be generated in order to develop an efficacious gonococcal vaccine.
Immunoglobulin G Isotype Responses to Variant Surface Antigens ofPlasmodium falciparumin Healthy Gabonese Adults and Children during and after Successive Malaria Attacks
Authors: Cabrera Gerardo , Yone Clarisse , Tebo Anne E. , van Aaken Jan , Lell Bertrand , Kremsner Peter G. , Luty Adrian J. F.
Published: December 2003

We assessed immunoglobulin G (IgG) isotype responses with specificity for the variant surface antigens (VSA) of heterologousPlasmodium falciparumisolates by using flow cytometry and plasma from healthy Gabonese adults and from children during and after two consecutive malaria episodes. The individual isolate-specific antibody profiles differed markedly in terms of their isotype content but were similar for healthy adults and healthy uninfected children. In healthy adults, IgG3 and IgG2 responses were the highest, while in healthy children, IgG3 and IgG4 predominated. A transiently elevated IgG1 response was observed during the second of two successive malaria episodes in children, signalingP. falciparuminfection-induced cross-reactive anti-VSA responses. Our findings highlight the prominence of IgG3 in the overall profile of these responses but also indicate a marked age-related increase in the prevalence of anti-VSA antibodies of the classically noncytophilic IgG2 isotype, possibly reflecting the high frequency of the histidine-131 variant of FcγRIIA in the Gabonese population.
Role of Siderophore Biosynthesis in Virulence of
Staphylococcus aureus
: Identification and Characterization of Genes Involved in Production of a Siderophore
Authors: Dale Suzanne E. , Doherty-Kirby Amanda , Lajoie Gilles , Heinrichs David E.
Published: December 2003

Molecular determinants underlying the production of siderophores in the human and animal pathogen
Staphylococcus aureus
and the contribution of siderophore production to the virulence of this bacterium have, until now, remained undefined. Here, we show that
S. aureus
strains RN6390 and Newman produce siderophore when the cells are starved for iron. We further identified and characterized a nine-gene, iron-regulated operon, designated
sbn
and situated between
sirABC
and
galE
on the
S. aureus
chromosome, that is involved in the production of a siderophore. Mutation of the
sbnE
gene, in both RN6390 and Newman, eliminates the ability of these strains to produce a siderophore under iron-limited growth conditions, while introduction of multicopy
sbnE
into
sbnE
mutants complemented the inability of the mutants to produce the siderophore.
sbnE
mutants, in both the RN6390 and Newman backgrounds, displayed a drastic growth deficiency, compared to the wild type, in iron-restricted growth medium, whereas no such deficiency was observed during growth in iron-replete medium. Complemented mutants showed a restored ability to grow under iron restriction. We further showed that an
sbnE
mutant was compromised in a murine kidney abscess model of
S. aureus
infection, illustrating the importance of siderophore production to the pathogenicity of
S. aureus. sbn
genes were present in all
S. aureus
strains tested (and all
S. aureus
genome sequences) but were undetectable in any of the 13 coagulase-negative staphylococci tested, including
Staphylococcus epidermidis
.
Interleukin-18 Is an Essential Element in Host Resistance to Experimental Group B Streptococcal Disease in Neonates
Authors: Cusumano Vitaliano , Midiri Angelina , Valeria Cusumano Valentina , Bellantoni Antonella , De Sossi Giuseppe , Teti Giuseppe , Beninati Concetta , Mancuso Giuseppe
Published: December 2003

Previous studies demonstrated that interleukin-12 (IL-12)-dependent gamma interferon (IFN-γ) responses have a major role in restricting in vivo bacterial growth during infection of mice with group B streptococci (GBS), important human pathogens. Like IL-12, IL-18 is a potent IFN-γ inducer. The role of IL-18 in experimental GBS infection was investigated here. Significant elevations of IL-18 levels over baseline values were detected in plasma samples from neonatal mice rendered septic with GBS. Neutralization of IL-18 significantly increased mortality and bacterial burden (P< 0.05). In contrast, administration of recombinant IL-18 (rIL-18) before or after GBS challenge remarkably improved survival and decreased blood colony counts, in association with increased IFN-γ production by spleen cells. The beneficial effects of rIL-18 were counteracted by administration of neutralizing anti-IFN-γ monoclonal antibodies, indicating that the effects of IL-18 were mediated by IFN-γ. Finally, low rIL-18 doses that had no effect of their own on bacterial burden could act in synergy with rIL-12 to protect neonatal mice during GBS infection. Collectively, our data indicate that IL-18 responses have an important role in host defenses against GBS and that rIL-18 may be useful in alternative strategies to treat neonatal GBS disease.
Effects of Anti-Endothelial Cell Antibodies in Leprosy and Malaria
Authors: Dugué Christophe , Perraut Ronald , Youinou Pierre , Renaudineau Yves
Published: December 2003

As a result of damaging endothelial cells (ECs),Mycobacterium lepraetriggers the production of antibodies (Abs). These anti-EC Abs (AECAs) can be divided into two types. The first type nonspecifically reacts with components of the cytosol (CY) and can be detected by enzyme-linked immunosorbent assay (ELISA). The second specifically reacts with the EC membrane (MB) and requires fluorescence-activated cell sorter (FACS) analysis to be detected. The presence of both types of AECAs was determined in 68 leprosy patients. The ELISA was positive for 35 of them but also for 30 of 34 malaria patients and 17 of 50 healthy African controls. However, whereas FACS analysis showed MB reactivity in only three malaria patients and four controls, this reactivity was found in 27 leprosy patients, more of those having the lepromatous than the tuberculoid form. Specificity for MB, which we failed to absorb by incubation with CY lysates, predominated over that for CY in leprosy, unlike malaria, where the EC reactivity was restricted to the CY. Western blot analysis and two-dimensional electrophoresis revealed that calreticulin, vimentin, tubulin, and heat shock protein 70 were targeted by AECAs from leprosy patients, but other proteins remained unidentified. These auto-Abs, but not those from malaria patients, did activate ECs, as indicated by the E-selectin and intercellular adhesion molecule 1 upregulation, and/or induced them into apoptosis, as documented by four different methods. Our findings suggest that, in some but not all leprosy patients, AECAs may play a role in pathogenesis.
Legionella pneumophila
Type II Protein Secretion Promotes Virulence in the A/J Mouse Model of Legionnaires' Disease Pneumonia
Authors: Rossier Ombeline , Starkenburg Shawn R. , Cianciotto Nicholas P.
Published: December 2003

Legionella pneumophila
, the gram-negative agent of Legionnaires' disease, possesses type IV pili and a type II protein secretion (Lsp) system, both of which are dependent upon the PilD prepilin peptidase. By analyzing multiple
pilD
mutants and various types of Lsp mutants as well as performing
trans
-complementation of these mutants, we have confirmed that PilD and type II secretion genes are required for
L. pneumophila
infection of both amoebae and human macrophages. Based upon a complete analysis of
lspDE
,
lspF
, and
lspG
mutants, we found that the type II system controls the secretion of protease, RNase, lipase, phospholipase A, phospholipase C, lysophospholipase A, and tartrate-sensitive and tartrate-resistant acid phosphatase activities and influences the appearance of colonies. Examination of the developing
L. pneumophila
genome database indicated that the organism has two other loci (
lspC
and
lspLM
) that are predicted to promote secretion and thus a set of genes that is comparable to the type II secretion genes in other gram-negative bacteria. In contrast to
lsp
mutants,
L. pneumophila
pilus mutants lacking either the PilQ secretin, the PspA pseudopilin, or pilin were not defective for colonial growth, secreted activities, or intracellular replication.
L. pneumophila dot/icm
mutants were also not impaired for type II-dependent exoenzymes. Upon intratracheal inoculation into A/J mice,
lspDE
,
lspF
, and
pilD
mutants, but not pilus mutants, exhibited a reduced ability to grow in the lung, as measured by competition assays. The
lspF
mutant was also defective in an in vivo kinetic assay. Examination of infected mouse sera revealed that type II secreted proteins are expressed in vivo. Thus, the
L. pneumophila
Lsp system is a virulence factor and the only type II secretion system linked to intracellular infection.
Mice Deficient in Interleukin-4 (IL-4) or IL-4 Receptor α Have Higher Resistance to Sporozoite Infection withPlasmodium berghei(ANKA) than Do Naive Wild-Type Mice
Authors: Saeftel Michael , Krueger Andreas , Arriens Sandra , Heussler Volker , Racz Paul , Fleischer Bernhard , Brombacher Frank , Hoerauf Achim
Published: December 2003

BALB/c interleukin-4 (IL-4−/−) or IL-4 receptor-α (IL-4rα−/−) knockout (KO) mice were used to assess the roles of the IL-4 and IL-13 pathways during infections with the blood or liver stages of plasmodium in murine malaria. Intraperitoneal infection with the blood-stage erythrocytes ofPlasmodium berghei(ANKA) resulted in 100% mortality within 24 days in BALB/c mice, as well as in the mutant mouse strains. However, when infected intravenously with the sporozoite liver stage, 60 to 80% of IL-4−/−and IL-4rα−/−mice survived, whereas all BALB/c mice succumbed with high parasitemia. Compared to infected BALB/c controls, the surviving KO mice showed increased NK cell numbers and expression of inducible nitric oxide synthase (iNOS) in the liver and were able to eliminate parasites early during infection. In vivo blockade of NO resulted in 100% mortality of sporozoite-infected KO mice. In vivo depletion of NK cells also resulted in 80 to 100% mortality, with a significant reduction in gamma interferon (IFN-γ) production in the liver. These results suggest that IFN-γ-producing NK cells are critical in host resistance against the sporozoite liver stage by inducing NO production, an effective killing effector molecule againstPlasmodium. The absence of IL-4-mediated functions increases the protective innate immune mechanism identified above, which results in immunity againstP. bergheiinfection in these mice, with no major role for IL-13.
Immune Response to Meningococcal Serogroup C Conjugate Vaccine in Asplenic Individuals
Authors: Balmer Paul , Falconer Michelle , McDonald Paula , Andrews Nick , Fuller Emily , Riley Christine , Kaczmarski Edward , Borrow Raymond
Published: December 2003

Asplenic individuals are known to be at increased risk of infection with encapsulated bacteria. Recent United Kingdom recommendations stated that this at-risk group should receive one dose of the meningococcal serogroup C conjugate (MCC) vaccine. However, the immune response of asplenic individuals to MCC vaccine is unknown. The immune response of asplenics (
n
= 130) to immunization with the MCC vaccine was investigated. Asplenic individuals had a significantly lower geometric mean titer (GMT) (157.8; 95% confidence interval [CI], 94.5 to 263.3) of bactericidal antibody in serum (SBA) than an age-matched control group (
n
= 48) (1448.2; 95% CI, 751.1 to 2792.0). However, 80% of asplenic individuals achieved the proposed protective SBA titer of ≥8. No differences were observed between the two groups in the serogroup C-specific immunoglobulin G geometric mean concentration. A significant reduction in SBA GMT or the number of responders achieving an SBA titer of ≥8 was observed if the reason for splenectomy was a medical cause or if MCC vaccination occurred <10 years after splenectomy. Individuals (
n
= 29) who did not achieve an SBA titer of ≥16 were offered a second dose of MCC vaccine. Analysis of the SBA response revealed that 61% (14 of 23) of the individuals who received a second dose achieved a protective titer. In total, 93% of asplenic individuals achieved a titer of ≥8 following MCC vaccination (one or two doses combined). We recommend that, following vaccination of asplenics, either the level of functional antibody should be determined, with a second dose of MCC vaccine offered to nonresponders, or two doses of MCC vaccine should be routinely offered.
Infection with an Avirulent
phoP
Mutant of
Neisseria meningitidis
Confers Broad Cross-Reactive Immunity
Authors: Newcombe J. , Eales-Reynolds L.-J. , Wootton L. , Gorringe A. R. , Funnell S. G. P. , Taylor S. C. , McFadden J. J.
Published: December 2003

Successful vaccines against serogroup A and C meningococcal strains have been developed, but current serogroup B vaccines provide protection against only a limited range of strains. The ideal meningococcal vaccine would provide cross-reactive immunity against the variety of strains that may be encountered in any community, but it is unclear whether the meningococcus possesses immune targets that have the necessary level of cross-reactivity. We have generated a
phoP
mutant of the meningococcus by allele exchange. PhoP is a component of a two-component regulatory system which in other bacteria is an important regulator of virulence gene expression. Inactivation of the PhoP-PhoQ system in
Salmonella
leads to avirulence, and
phoP
mutants have been shown to confer protection against virulent challenge. These mutants have been examined as potential live attenuated vaccines. We here show that a
phoP
mutant of the meningococcus is avirulent in a mouse model of infection. Moreover, infection of mice with the
phoP
mutant stimulated a bactericidal immune response that not only killed the infecting strain but also showed cross-reactive bactericidal activity against a range of strains with different serogroup, serotype, and serosubtyping antigens. Sera from the mutant-infected mice contained immunoglobulin G that bound to the surface of a range of meningococcal strains and mediated opsonophagocytosis of meningococci by human phagocytic cells. The meningococcal
phoP
mutant is thus a candidate live, attenuated vaccine strain and may also be used to identify cross-reactive protective antigens in the meningococcus.
Vaccination with Attenuated
Neisseria meningitidis
Strains Protects against Challenge with Live Meningococci
Authors: Li Yanwen , Sun Yao-hui , Ison Cathy , Levine Myron M. , Tang Christoph M.
Published: December 2003

Meningococcal disease is a life-threatening infection caused by
Neisseria meningitidis
. Currently, there are no vaccines to prevent infection with serogroup B
N. meningitidis
strains, the leading cause of meningococcal meningitis in Europe and North America. Here we describe the construction and characterization of two attenuated serogroup B
N. meningitidis
strains, YH102 (MC58Δ
sia
Δ
rfaF
) and YH103 (MC58Δ
sia
Δ
metH
). Both strains are markedly attenuated in their capacity to cause bacteremia in rodent models and have a reduced ability to survive in a human whole-blood assay. Immunization of adult mice with these strains leads to the development of bactericidal antibodies and confers sterilizing protection against challenge with homologous live bacteria. Furthermore, we show that the strains confer protection against infection by other serogroups. Use of the attenuated strains in animals with gene knockouts or after depletion of immunological effectors could be used to define the basis of protection, and human volunteer studies could be undertaken to examine the immune response following exposure to this important human pathogen.
Innate Immune Response of Oral and Foreskin Keratinocytes: Utilization of Different Signaling Pathways by Various Bacterial Species
Authors: Chung Whasun O. , Dale Beverly A.
Published: December 2003

The innate immune response is critical for the epithelial antimicrobial barrier. The human β-defensins are small, cationic antimicrobial peptides that are made by epithelial cells and that play a role in mucosal and skin defenses. Human β-defensin 1 (hBD-1) is expressed constitutively in epithelial tissues, whereas hBD-2 and hBD-3 are expressed in response to bacterial stimuli or inflammation. Previous studies showed that hBD-2 was induced byFusobacterium nucleatumcell wall extract without the involvement of the NF-κB transcription factors, which typically are associated with innate immunity and inflammation. The goal of this study was to characterize signaling pathways involved in hBD-2 induction in response to commensal and pathogenic bacteria. Cultured human oral and foreskin keratinocytes were treated separately with inhibitors of NF-κB, c-Jun N-terminal kinase (JNK), and p38 and then stimulated with oral commensalStreptococcus gordonii, oral pathogensPorphyromonas gingivalisandActinobacillus actinomycetemcomitans, skin commensalStaphylococcus epidermidis, or skin pathogenStreptococcus pyogenes. Different bacteria induced different levels of hBD-2 and in response to the various inhibitors tested, although certain common patterns were observed for commensal- and pathogen-stimulated cells. hBD-2 induction by all bacteria tested was partially or completely blocked by inhibitors of the JNK and p38 pathways. However, in addition, hBD-2 induction by pathogenic bacteria in both oral and foreskin keratinocytes was blocked by inhibitors of NF-κB. The results indicate that commensal and pathogenic bacteria utilize different pathways in hBD-2 induction and suggest that epithelial cells from different body sites have common signaling mechanisms to distinguish between commensal and pathogenic bacteria.