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ABSTRACT: Meningococcal factor H-binding protein (fHbp) is a promising vaccine antigen. Previous studies described three fHbp antigenic variant groups and identified amino acid residues between 100 and 255 as important targets of variant-specific bactericidal antibodies. We investigated residues affecting expression of an epitope recognized by a murine IgG2a anti-fHbp mAb, designated JAR 4, which cross-reacted with fHbps in variant group 1 or 2 (95% of strains), and elicited human complement-mediated, cooperative bactericidal activity with other non-bactericidal anti-fHbp mAbs with epitopes involving residues between 121 and 216. From filamentous bacteriophage libraries containing random peptides that were recognized by JAR 4, we identified a consensus tripeptide, DHK that matched residues 25-27 in the N-terminal domain of fHbp. Since DHK was present in both JAR 4-reactive and non-reactive fHbps, the tripeptide was necessary but not sufficient for reactivity. Based on site-directed mutagenesis studies, the JAR 4 epitope could either be knocked out of a reactive variant 1 fHbp, or introduced into a non-reactive variant 3 protein. Collectively, the data indicated that the JAR 4 epitope was discontinuous and involved DHK residues beginning at position 25; YGN residues beginning at position 57; and a KDN tripeptide that was present in variant 3 proteins beginning at position 67 that negatively affected expression of the epitope. Thus, the region of fHbp encompassing residues 25-59 in the N-terminal domain is important for eliciting antibodies that can cooperate with other anti-fHbp antibodies for cross-reactive bactericidal activity against strains expressing fHbp from different antigenic variant groups.
Molecular Immunology 04/2009; 46(8-9):1647-53. · 2.90 Impact Factor
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ABSTRACT: Serum-complement-mediated bactericidal antibody (SBA) remains the serologic hallmark of protection against meningococcal disease, despite experimental and epidemiologic data that SBA may underestimate immunity. We measured bactericidal activity against three strains of Neisseria meningitidis group B in sera from 48 healthy adults and in whole blood from 15 subjects. Blood was anticoagulated with lepirudin, a specific thrombin inhibitor not known to activate complement. Depending on the test strain, protective SBA titers of >/=1:4 were present in only 8 to 15% of the subjects, whereas bactericidal activity was present in 40 to 87% of subjects according to the blood assay. Among SBA-negative subjects, blood from 23 to 42% gave a decrease of >/=2 log(10) CFU/ml after 1 h of incubation, and blood from 36 to 83% gave a decrease of >/=1 log(10) after 2 h. For most blood samples, bactericidal antibodies primarily were directed against noncapsular antigens, since activity was not inhibited by group B polysaccharide. For some SBA-negative subjects, white cells were not needed, since similar respective bactericidal activities were observed in blood and plasma. Bactericidal activity by whole blood of SBA-negative subjects can be rapid (<1 h) and effective (>/=2 log(10)) and, among all subjects, was four- to sixfold more prevalent than a positive SBA. Thus, while an SBA titer of >/=1:4 predicts protection against meningococcal disease, a titer of <1:4 is poorly predictive of susceptibility. More sensitive assays than SBA are needed to assess protective meningococcal immunity, or we risk underestimating the extent of immunity in the population and the effectiveness of new meningococcal vaccines.
Clinical and vaccine immunology: CVI 01/2008; 14(12):1596-602. · 2.37 Impact Factor
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Marzia M Giuliani,
Jeannette Adu-Bobie,
Maurizio Comanducci,
Beatrice Aricò,
Silvana Savino,
Laura Santini,
Brunella Brunelli,
Stefania Bambini,
Alessia Biolchi,
Barbara Capecchi, [......],
Maurizio Morandi,
Alessandro Bartalesi,
Vanda Cinotti,
Donatella Mannucci,
Francesca Titta,
Elisa Ovidi,
Jo Anne Welsch, Dan Granoff,
Rino Rappuoli,
Mariagrazia Pizza
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ABSTRACT: Meningitis and sepsis caused by serogroup B meningococcus are two severe diseases that still cause significant mortality. To date there is no universal vaccine that prevents these diseases. In this work, five antigens discovered by reverse vaccinology were expressed in a form suitable for large-scale manufacturing and formulated with adjuvants suitable for human use. The vaccine adjuvanted by aluminum hydroxide induced bactericidal antibodies in mice against 78% of a panel of 85 meningococcal strains representative of the global population diversity. The strain coverage could be increased to 90% and above by the addition of CpG oligonucleotides or by using MF59 as adjuvant. The vaccine has the potential to conquer one of the most devastating diseases of childhood.
Proceedings of the National Academy of Sciences 08/2006; 103(29):10834-9. · 9.68 Impact Factor
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ABSTRACT: The hallmark of immunity to meningococcal disease is a bactericidal titer in serum of > or =1:4 measured with human complement, but this threshold titer may underestimate the extent of protection. We used the infant rat model of meningococcal bacteremia to measure group C passive protective activity in serum samples from 91 unimmunized adults living in California. A total of 35 sera (38.5%) had passive protective activity. Sera with complement-mediated bactericidal titers of > or =1:4 were 3.4-fold more likely to confer protection (89%) than nonbactericidal sera (26%; P < 0.0001). Thus, bactericidal titers of > or =1:4 are a marker of protection, but this threshold lacks sensitivity for predicting protective activity. We investigated the 73 sera with bactericidal titers of <1:4 to determine the basis of protective activity. The 19 sera with protective activity had a higher geometric mean group C anticapsular antibody concentration (0.72 microg/ml) than the 54 sera that lacked protective activity (0.16 microg/ml; P < 0.001). Thus, protective activity in the absence of bactericidal activity was associated with higher concentrations of anticapsular antibodies, but not all sera with anticapsular antibodies conferred protection. Of 18 nonbactericidal sera with anticapsular antibody concentrations between 0.31 and 0.99 microg/ml, the 11 sera that conferred protection had a higher mean antibody avidity constant (21.9 nM(-1)) than the 7 nonprotective sera (14.6 nM(-1); P < 0.03). Thus, in sera with titers of <1:4, protective activity is associated with higher-avidity group C anticapsular antibodies, which are present in concentrations insufficient to elicit complement-mediated bacteriolysis in vitro but sufficient to confer protection in an in vivo bacteremia model.
Infection and Immunity 11/2004; 72(10):5903-9. · 4.16 Impact Factor
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ABSTRACT: GNA33 is a membrane-bound lipoprotein with murein hydrolase activity that is present in all Neisseria species and well conserved in different meningococcal isolates. The protein shows 33% identity to a lytic transglycolase (MltA) from Escherichia coli and has been shown to be involved in the degradation of both insoluble murein sacculi and unsubstituted glycan strands. To study the function of the gene and its role in pathogenesis and virulence, a knockout mutant of a Neisseria meningitidis serogroup B strain was generated. The mutant exhibited retarded growth in vitro. Transmission electron microscopy revealed that the mutant grows in clusters which are connected by a continuous outer membrane, suggesting a failure in the separation of daughter cells. Moreover, sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of culture supernatant revealed that the mutant releases several proteins in the medium. The five most abundant proteins, identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry analysis, belong to the outer membrane protein family. Finally, the mutant showed an attenuated phenotype, since it was not able to cause bacteremia in the infant rat model. We conclude that GNA33 is a highly conserved lipoprotein which plays an important role in peptidoglycan metabolism, cell separation, membrane architecture, and virulence.
Infection and Immunity 05/2004; 72(4):1914-9. · 4.16 Impact Factor
