Subtyping ofNeisseria meningitidisstrains isolated in Quebec, Canada: correlation between deduced amino acid sequences and serosubtyping techniques

1997 ◽  
Vol 43 (3) ◽  
pp. 234-238 ◽  
Author(s):  
Francis F. Arhin ◽  
France Moreau ◽  
Elaine L. Mills ◽  
James W. Coulton
Keyword(s):  
Amino Acid ◽  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Outer Membrane Proteins ◽  
Outer Membrane Vesicles ◽  
Amino Acid Sequences ◽  
Complete Agreement ◽  
Enzyme Linked Immunosorbent Assay ◽  
Variable Regions ◽  
Whole Cells

Routine serosubtyping of Neisseria meningitidis relies upon reactivity of whole cells to monoclonal antibodies (mAbs). This procedure is limited in providing maximum serosubtype information because some epitopes in whole cells are masked and because mAbs are currently unavailable for some epitopes. To address masking of epitopes in whole cells, we isolated outer membrane vesicles (OMVs) from nine representative meningococcal strains that were isolated (1991–1993) in the province of Quebec, Canada; the OMVs were used in enzyme-linked immunosorbent assay for reactivity to mAbs, and improved serosubtyping information was obtained. A recent proposal assigns subtypes based on deduced amino acid sequences in the variable regions of the class 1 outer membrane protein. This scheme maintains the subtyping nomenclature that is based on reactivity to mAbs by defining the sequences in the epitopes recognized by the mAbs. We used this technique to assign subtypes to the meningococcal strains isolated in Quebec. For the strains tested, serosubtyping using mAbs and subtyping based on deduced amino acid sequences were in complete agreement. Subtyping using deduced amino acid sequences is superior because it does not depend on the availability of mAbs.Key words: Neisseria meningitidis, outer membrane proteins, serosubtyping, PorA.

scholarly journals Sequence Polymorphism, Predicted Secondary Structures, and Surface-Exposed Conformational Epitopes of Campylobacter Major Outer Membrane Protein

2000 ◽  
Vol 68 (10) ◽  
pp. 5679-5689 ◽  
Author(s):  
Qijing Zhang ◽  
Jerrel C. Meitzler ◽  
Shouxiong Huang ◽  
Teresa Morishita
Keyword(s):  
Amino Acid ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Outer Membrane Proteins ◽  
Major Outer Membrane Protein ◽  
Amino Acid Sequences ◽  
Variable Regions ◽  
Conserved Sequences ◽  
Conformational Epitopes

ABSTRACT The major outer membrane protein (MOMP), a putative porin and a multifunction surface protein of Campylobacter jejuni, may play an important role in the adaptation of the organism to various host environments. To begin to dissect the biological functions and antigenic features of this protein, the gene (designatedcmp) encoding MOMP was identified and characterized from 22 strains of C. jejuni and one strain of C. coli. It was shown that the single-copy cmp locus encoded a protein with characteristics of bacterial outer membrane proteins. Prediction from deduced amino acid sequences suggested that each MOMP subunit consisted of 18 β-strands connected by short periplasmic turns and long irregular external loops. Alignment of the amino acid sequences of MOMP from different strains indicated that there were seven localized variable regions dispersed among highly conserved sequences. The variable regions were located in the putative external loop structures, while the predicted β-strands were formed by conserved sequences. The sequence homology of cmp appeared to reflect the phylogenetic proximity of C. jejuni strains, since strains with identical cmp sequences had indistinguishable or closely related macrorestriction fragment patterns. Using recombinant MOMP and antibodies recognizing linear or conformational epitopes of the protein, it was demonstrated that the surface-exposed epitopes of MOMP were predominantly conformational in nature. These findings are instrumental in the design of MOMP-based diagnostic tools and vaccines.

scholarly journals Cross-reactivity with Brazilian strains of Neisseria meningitidis B after immunization with outer membrane vesicles

2019 ◽  
Vol 7 ◽  
pp. 251513551989482
Author(s):  
Gabriela Trzewikoswki de Lima ◽  
Amanda Izeli Portilho ◽  
Elizabeth De Gaspari
Keyword(s):  
Public Health ◽  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Immunosorbent Assay ◽  
Humoral Response ◽  
Membrane Vesicles ◽  
Cross Reactivity ◽  
Outer Membrane Vesicles ◽  
Antigen Recognition ◽  
Enzyme Linked Immunosorbent Assay

Background: Immunization against Neisseria meningitidis is important for public health. Vaccines composed of cross-reactivity antigens avoid strain-specific responses, ensuring more comprehensive protection. Methods: The cross-reactivity between three strains from the last outbreak of N. meningitidis in Brazil was assessed in our studies, using enzyme-linked immunosorbent assay (ELISA) and immunoblotting assays. Results: Both assays verifed a similar humoral response between the strains evaluated. Patterns of antigen recognition differed with each dose evaluated. Conclusions: We observed that immunization with N. meningitidis B outer membrane vesicles (OMVs) led to the production of antibodies that recognized antigens of heterologous strains, indicating possible protection against these evaluated strains.

scholarly journals Serum Antibody Responses in Ethiopian Meningitis Patients Infected with Neisseria meningitidis Serogroup A Sequence Type 7

2007 ◽  
Vol 14 (4) ◽  
pp. 451-463 ◽  
Author(s):  
Gunnstein Norheim ◽  
Abraham Aseffa ◽  
Mohammed Ahmed Yassin ◽  
Getahun Mengistu ◽  
Afework Kassu ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Outer Membrane ◽  
Acute Phase ◽  
Outer Membrane Proteins ◽  
Serum Antibody ◽  
Outer Membrane Vesicles ◽  
Sequence Type ◽  
Enzyme Linked Immunosorbent Assay ◽  
Human Complement ◽  
Convalescent Phase

ABSTRACT To elucidate critical components of protective immune responses induced during the natural course of serogroup A meningococcal disease, we studied acute-, early-convalescent-, and late-convalescent-phase sera from Ethiopian patients during outbreaks in 2002 to 2003. Sera were obtained from laboratory-confirmed patients positive for serogroup A sequence type 7 (ST-7) meningococci (A:4/21:P1.20,9) (n = 71) and from Ethiopian controls (n = 113). The sera were analyzed using an enzyme-linked immunosorbent assay to measure levels of immunoglobulin G (IgG) against serogroup A polysaccharide (APS) and outer membrane vesicles (OMVs) and for serum bactericidal activity (SBA) using both rabbit and human complement sources. Despite relatively high SBA titers and high levels of IgG against APS and OMVs in acute-phase patient sera, significant increases were seen in the early convalescent phase. Antibody concentrations returned to acute-phase levels in the late convalescent phase. Considering all patients' sera, a significant but low correlation (r = 0.46) was observed between SBA with rabbit complement (rSBA) using an ST-5 reference strain and SBA with human complement (hSBA) using an ST-7 strain from Ethiopia. While rSBA demonstrated a significant linear relation with IgG against APS, hSBA demonstrated significant linear relationships with IgG against both APS and OMV. This study indicates that antibodies against both outer membrane proteins and APS may be important in providing the protection induced during disease, as measured by hSBA. Therefore, outer membrane proteins could also have a role as components of future meningococcal vaccines for the African meningitis belt.

scholarly journals Additive and Synergistic Bactericidal Activity of Antibodies Directed against Minor Outer Membrane Proteins of Neisseria meningitidis

2007 ◽  
Vol 75 (11) ◽  
pp. 5434-5442 ◽  
Author(s):  
Vincent E. Weynants ◽  
Christiane M. Feron ◽  
Karine K. Goraj ◽  
Martine P. Bos ◽  
Philippe A. Denoël ◽  
...  
Keyword(s):  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Bactericidal Activity ◽  
Outer Membrane Proteins ◽  
Critical Density ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Wild Type ◽  
Major Variable ◽  
Type Strains

ABSTRACT Neisseria meningitidis serogroup B is a major cause of bacterial meningitis in younger populations. The available vaccines are based on outer membrane vesicles obtained from wild-type strains. In children less than 2 years old they confer protection only against strains expressing homologous PorA, a major, variable outer membrane protein (OMP). We genetically modified a strain in order to eliminate PorA and to overproduce one or several minor and conserved OMPs. Using a mouse model mimicking children's PorA-specific bactericidal activity, it was demonstrated that overproduction of more than one minor OMP is required to elicit antibodies able to induce complement-mediated killing of strains expressing heterologous PorA. It is concluded that a critical density of bactericidal antibodies needs to be reached at the surface of meningococci to induce complement-mediated killing. With minor OMPs, this threshold is reached when more than one antigen is targeted, and this allows cross-protection.

scholarly journals Mapping of a Protective Epitope of the CopB Outer Membrane Protein of Moraxella catarrhalis

1998 ◽  
Vol 66 (2) ◽  
pp. 540-548 ◽  
Author(s):  
Christoph Aebi ◽  
Leslie D. Cope ◽  
Jo L. Latimer ◽  
Sharon E. Thomas ◽  
Clive A. Slaughter ◽  
...  
Keyword(s):  
Amino Acid ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Moraxella Catarrhalis ◽  
Amino Acid Sequences ◽  
Nucleotide Sequence Analysis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Protective Epitope

ABSTRACT A monoclonal antibody (MAb) (MAb 10F3) directed against the CopB outer membrane protein of Moraxella catarrhalis previously was found to enhance pulmonary clearance of M. catarrhalisin an animal model (M. Helminen, I. Maciver, J. L. Latimer, L. D. Cope, G. H. McCracken, Jr., and E. J. Hansen, Infect. Immun. 61:2003–2010, 1993). In the present study, this same MAb was shown to exert complement-dependent bactericidal activity against this pathogen in vitro. Nucleotide sequence analysis of thecopB gene from two MAb 10F3-reactive and two MAb 10F3-unreactive strains of M. catarrhalis revealed that the deduced amino acid sequences of these four CopB proteins were at least 90% identical. Comparison of the amino acid sequences of these proteins allowed localization of possible MAb 10F3 binding sites to five relatively small regions of the CopB protein from M. catarrhalis O35E. When five synthetic peptides representing these regions were tested for their ability to bind MAb 10F3 in a direct enzyme-linked immunosorbent assay system, an oligopeptide containing 26 amino acids was shown to bind this MAb. The actual binding region for MAb 10F3 was localized further through the use of overlapping decapeptides that spanned this 26-mer. A fusion protein containing the same 26-mer readily bound MAb 10F3 and was used to immunize mice. The resultant antiserum contained antibodies that reacted with the CopB protein of the homologous M. catarrhalis strain in Western blot analysis and bound to the surface of both homologous and heterologous strains of M. catarrhalis.

scholarly journals Genetic Diversity of the 28-Kilodalton Outer Membrane Protein Gene in Human Isolates of Ehrlichia chaffeensis

1999 ◽  
Vol 37 (4) ◽  
pp. 1137-1143 ◽  
Author(s):  
Xue-Jie Yu ◽  
Jere W. McBride ◽  
David H. Walker
Keyword(s):  
Amino Acid ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Dna Sequence ◽  
Outer Membrane Protein ◽  
Signal Sequence ◽  
Outer Membrane Proteins ◽  
Protein A ◽  
Amino Acid Sequences ◽  
Ehrlichia Chaffeensis

The Ehrlichia chaffeensis 28-kDa outer membrane protein (p28) gene was sequenced completely by genomic walking with adapter PCR. The DNA sequence of the p28 gene was nearly identical to the previously reported sequence (N. Ohashi, N. Zhi, Y. Zhang, and Y. Rikihisa, Infect. Immun. 66:132–139, 1998), but analysis of a further 75 bp on the 5′ end of the gene revealed DNA that encoded a 25-amino-acid signal sequence. The leader sequence was removed from the N terminus of a 30-kDa precursor to generate the mature p28 protein. A monoclonal antibody (MAb), 1A9, recognizing four outer membrane proteins of E. chaffeensis (Arkansas strain) including the 25-, 26-, 27-, and 29-kDa proteins (X.-J. Yu, P. Brouqui, J. S. Dumler, and D. Raoult, J. Clin. Microbiol. 31:3284–3288, 1993) reacted with the recombinant p28 protein. This result indicated that the four proteins recognized by MAb 1A9 were encoded by the multiple genes of the 28-kDa protein family. DNA sequence alignment analysis revealed divergence of p28 among all five human isolates of E. chaffeensis. The E. chaffeensis strains could be divided into three genetic groups on the basis of the p28 gene. The first group consisted of the Sapulpa and St. Vincent strains. They had predicted amino acid sequences identical to each other. The second group contained strain 91HE17 and strain Jax, which only showed 0.4% divergence from each other. The third group contained the Arkansas strain only. The amino acid sequences of p28 differed by 11% between the first two groups, by 13.3% between the first and third groups, and by 13.1% between the second and third groups. The presence of antigenic variants of p28 among the strains of E. chaffeensis and the presence of multiple copies of heterogeneous genes suggest a possible mechanism by which E. chaffeensismight evade the host immune defenses. Whether or not immunization with the p28 of one strain of E. chaffeensis would confer cross-protection against other strains needs to be investigated.

Sequencing ofporAfrom clinical isolates ofNeisseria meningitidisdefines a subtyping scheme and its genetic regulation

1998 ◽  
Vol 44 (1) ◽  
pp. 56-63 ◽  
Author(s):  
Francis F Arhin ◽  
France Moreau ◽  
James W Coulton ◽  
Elaine L Mills
Keyword(s):  
Membrane Protein ◽  
Dna Sequencing ◽  
Neisseria Meningitidis ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Promoter Region ◽  
Outer Membrane Proteins ◽  
Coding Region ◽  
Variable Regions ◽  
Class 1

Subtyping Neisseria meningitidis by methods that rely on monoclonal antibody (mAb) reactivity results in an unusually high number of strains that are not subtypeable. To subtype 48 strains isolated (1993-1994) in the province of Quebec that were not subtypeable by mAb-based techniques, we used DNA sequencing of the variable regions of porA, a gene that encodes the class 1 outer membrane protein. We assigned subtypes to all the previously nonserosubtypeable isolates and identified some novel subtypes. Because our sequencing strategy included the promoter region of porA, different isolates were compared in their sequences of the porA promoter region. A poly(G) stretch lies between the -10 and -35 regions of the promoter; replacement of a G residue by an A residue in this region resulted in loss of expression of porA. No correlation was found between the number of G residues in the poly(G) stretch and the level of expression; a minimum of 10 G residues is required in this stretch for expression of porA. One isolate expressed no class 1 outer membrane protein because of the insertion sequence IS1301 in the coding region of porA. Another isolate did not express the protein owing to a frame-shift mutation within the coding region of porA. Sequencing of porA allowed assignments of subtypes to previously uncharacterized isolates and provided insights about the regulation of expression of this gene in N. meningitidis.Key words: Neisseria meningitidis, outer membrane proteins, subtyping, PorA, DNA sequencing.

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