Moraxella catarrhalis Outer Membrane Protein CD Elicits Antibodies That Inhibit CD Binding to Human Mucin and Enhance Pulmonary Clearance of M. catarrhalis in a Mouse Model

ABSTRACT The outer membrane protein CD of Moraxella catarrhalis is considered to be a potential vaccine antigen against Moraxella infection. We purified the native CD from isolate O35E, administered it to mice, and detected considerable titers of anti-CD antibodies. Anti-CD sera were cross-reactive towards six different M. catarrhalis isolates and promoted bacterial clearance of O35E in a pulmonary challenge model. To circumvent the difficulty of generating large quantities of CD from M. catarrhalis for vaccine use, the CD gene from O35E was cloned into Escherichia coli, and the recombinant CD, expressed without a signal sequence or fusion tags, represented ∼70% of the total E. coli proteins. The recombinant CD formed inclusion bodies that were solubilized with 6 M urea and then purified by ion-exchange chromatography, a procedure that produced soluble CD of high purity and yield. Mice immunized with the purified recombinant CD had significant titers of anti-CD antibodies that were cross-reactive towards 24 different M. catarrhalis isolates. Upon challenge, these mice showed enhanced bacterial clearance of both O35E and a heterologous M. catarrhalis isolate, TTA24. In an in vitro assay, antisera to either the native or the recombinant CD inhibited the binding activity of CD to human tracheobronchial mucin in a serum concentration-dependent manner, and the extent of inhibition appeared to correlate with the corresponding anti-CD antibody titer and whole-cell enzyme-linked immunosorbent assay titer. Our results demonstrate that the recombinant CD is a promising vaccine candidate for preventing Moraxella infection.

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Human Immune Response to Outer Membrane Protein CD of Moraxella catarrhalis in Adults with Chronic Obstructive Pulmonary Disease

Infection and Immunity ◽

10.1128/iai.71.3.1288-1294.2003 ◽

2003 ◽

Vol 71 (3) ◽

pp. 1288-1294 ◽

Cited By ~ 29

Author(s):

Timothy F. Murphy ◽

Charmaine Kirkham ◽

Dai-Fang Liu ◽

Sanjay Sethi

Keyword(s):

Chronic Obstructive Pulmonary Disease ◽

Immune Response ◽

Membrane Protein ◽

Outer Membrane ◽

Pulmonary Disease ◽

Outer Membrane Protein ◽

Moraxella Catarrhalis ◽

Enzyme Linked Immunosorbent Assay ◽

Chronic Obstructive ◽

Obstructive Pulmonary Disease

ABSTRACT Moraxella catarrhalis is a common cause of lower respiratory tract infection in adults with chronic obstructive pulmonary disease (COPD). The antibody response to outer membrane protein (OMP) CD, a highly conserved surface protein of M. catarrhalis under consideration as a vaccine antigen, was studied in adults with COPD following 40 episodes of infection or colonization. Following infection or colonization, 9 of 40 patients developed new serum immunoglobulin G (IgG) to OMP CD, as measured by enzyme-linked immunosorbent assay. Adsorption assays revealed that a proportion of the serum IgG was directed toward surface-exposed epitopes on OMP CD in six of the nine patients who developed new IgG to OMP CD. Immunoblot assays with fusion peptide constructs indicated that the new antibodies that developed after infection or colonization recognized conformational epitopes, particularly in the carboxy region of the protein. Three of 28 patients developed new mucosal IgA to OMP CD in sputum supernatants. This study establishes that OMP CD is a target of a systemic and mucosal immune response following infection and colonization in some patients with COPD.

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Identification and Characterization of a Novel Outer Membrane Protein (OMP J) of Moraxella catarrhalis That Exists in Two Major Forms

Journal of Bacteriology ◽

10.1128/jb.187.23.7977-7984.2005 ◽

2005 ◽

Vol 187 (23) ◽

pp. 7977-7984 ◽

Cited By ~ 12

Author(s):

John P. Hays ◽

Saskia van Selm ◽

Theo Hoogenboezem ◽

Silvia Estevão ◽

Kimberly Eadie ◽

...

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Moraxella Catarrhalis ◽

Outer Membrane Proteins ◽

Acute Otitis Media ◽

Resistant Isolate ◽

Genetic Lineage ◽

Bacterial Clearance ◽

Significant Difference

ABSTRACT Moraxella catarrhalis is a common commensal of the human respiratory tract that has been associated with a number of disease states, including acute otitis media in children and exacerbations of chronic obstructive pulmonary disease in adults. During studies to investigate the outer membrane proteins of this bacterium, two novel major proteins, of approximately 19 kDa and 16 kDa (named OMP J1 and OMP J2, respectively), were identified. Further analysis indicated that these two proteins possessed almost identical gene sequences, apart from two insertion/deletion events in predicted external loops present within the putative barrel-like structure of the proteins. The development of a PCR screening strategy found a 100% (96/96) incidence for the genes encoding the OMP J1 and OMP J2 proteins within a set of geographically diverse M. catarrhalis isolates, as well as a significant association of OMP J1/OMP J2 with both the genetic lineage and the complement resistance phenotype (Fisher's exact test; P < 0.01). Experiments using two ΔompJ2 mutants (one complement resistant and the other complement sensitive) indicated that both were less easily cleared from the lungs of mice than were their isogenic wild-type counterparts, with a significant difference in bacterial clearance being observed for the complement-resistant isolate but not for its isogenic ΔompJ2 mutant (unpaired Student's t test; P < 0.001 and P = 0.32). In this publication, we characterize a novel outer membrane protein of Moraxella catarrhalis which exists in two variant forms associated with particular genetic lineages, and both forms are suggested to contribute to bacterial clearance from the lungs.

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

Infection and Immunity ◽

10.1128/iai.66.2.540-548.1998 ◽

1998 ◽

Vol 66 (2) ◽

pp. 540-548 ◽

Cited By ~ 39

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.

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Conservation of Outer Membrane Protein E among Strains of Moraxella catarrhalis

Infection and Immunity ◽

10.1128/iai.69.6.3576-3580.2001 ◽

2001 ◽

Vol 69 (6) ◽

pp. 3576-3580 ◽

Cited By ~ 18

Author(s):

Timothy F. Murphy ◽

Aimee L. Brauer ◽

Norine Yuskiw ◽

Erin R. McNamara ◽

Charmaine Kirkham

Keyword(s):

Monoclonal Antibodies ◽

Membrane Protein ◽

Respiratory Tract ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Moraxella Catarrhalis ◽

Vaccine Antigen ◽

Effective Vaccine ◽

Human Respiratory Tract ◽

The Stability

ABSTRACT Outer membrane protein E (OMP E) is a 50-kDa protein ofMoraxella catarrhalis which has several features that suggest that the protein may be an effective vaccine antigen. To assess the conservation of OMP E among strains of M. catarrhalis,22 isolates were studied with eight monoclonal antibodies which recognize epitopes on different regions of the protein. Eighteen of 22 strains were reactive with all eight antibodies. The sequences ofompE from 16 strains of M. catarrhalis were determined, including the 4 strains which were nonreactive with selected monoclonal antibodies. Analysis of sequences indicate a high degree of conservation among strains, with sequence differences clustered in limited regions of the gene. To assess the stability ofompE during colonization of the human respiratory tract, the sequences of ompE of isolates collected from patients colonized with the same strain for 3 to 9 months were determined. The sequences remained unchanged. These results indicate that OMP E is highly conserved among strains of M. catarrhalis, and preliminary studies indicate that the gene which encodes OMP E remains stable during colonization of the human respiratory tract.

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The Moraxella catarrhalis Porin-Like Outer Membrane Protein CD Is an Adhesin for Human Lung Cells

Infection and Immunity ◽

10.1128/iai.72.4.1906-1913.2004 ◽

2004 ◽

Vol 72 (4) ◽

pp. 1906-1913 ◽

Cited By ~ 63

Author(s):

Melissa M. Holm ◽

Serena L. Vanlerberg ◽

Ian M. Foley ◽

Darren D. Sledjeski ◽

Eric R. Lafontaine

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Gene Product ◽

Outer Membrane Protein ◽

Moraxella Catarrhalis ◽

Human Lung ◽

A549 Cells ◽

Vaccine Antigen ◽

Lung Cells ◽

Human Lung Cells

ABSTRACT The outer membrane protein CD (OMPCD) of Moraxella catarrhalis is an outer membrane protein with several attributes of a potential vaccine antigen. We isolated four transposon mutants of strain O35E on the basis of their reduced binding to A549 human lung cells in microcolony formation assays, and we determined that they contain a transposon in ompCD. We also found that these transposon insertions had pleiotropic effects: mutants grew slower, became serum sensitive, bound ∼10-fold less to A549 cells, and appeared transparent when grown on solid medium. We confirmed that these various phenotypes could be attributed solely to disruption of ompCD by constructing the isogenic strain O35E.CD1. O35E-ompCD was cloned, and recombinant Escherichia coli bacteria expressing the gene product exhibited a 10-fold increase in adherence to A549 cells. This is the first report of M. catarrhalis ompCD mutants, and our findings demonstrate that this gene product is an adhesin for human lung cells.

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Intranasal Vaccination with Recombinant Outer Membrane Protein CD and Adamantylamide Dipeptide as the Mucosal Adjuvant Enhances Pulmonary Clearance of Moraxella catarrhalis in an Experimental Murine Model

Infection and Immunity ◽

10.1128/iai.01081-06 ◽

2006 ◽

Vol 75 (4) ◽

pp. 1778-1784 ◽

Cited By ~ 28

Author(s):

Pablo D. Becker ◽

Gustavo M. Bertot ◽

David Souss ◽

Thomas Ebensen ◽

Carlos A. Guzmán ◽

...

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Moraxella Catarrhalis ◽

Interleukin 2 ◽

Immunoglobulin A ◽

Acute Otitis Media ◽

Vaccine Antigen ◽

Mucosal Adjuvant ◽

Tract Infections

ABSTRACT Moraxella catarrhalis causes acute otitis media in children and lower respiratory tract infections in adults and elderly. In children the presence of antibodies against the highly conserved outer membrane protein CD correlates with protection against infection, suggesting that this protein may be useful as a vaccine antigen. However, native CD is difficult to purify, and it is still unclear if recombinant CD (rCD) is a valid alternative. We performed a side-by-side comparison of the immunogenicities and efficacies of vaccine formulations containing native CD and rCD with adamantylamide dipeptide as the mucosal adjuvant. Intranasal vaccination of mice stimulated the production of high CD-specific antibody titers in sera and of secretory immunoglobulin A in mucosal lavages, which cross-recognized both antigens. While vaccination with native CD increased the number of interleukin-2 (IL-2)- and gamma interferon-producing cells, rCD mainly stimulated IL-4-secreting cells. Nevertheless, efficient bacterial clearance was observed in the lungs of challenged mice receiving native CD and in the lungs of challenged mice receiving rCD (96% and 99%, respectively). Thus, rCD is a promising candidate for incorporation in vaccine formulations for use against M. catarrhalis.

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Characterization of the Humoral Immune Response to Chlamydia Outer Membrane Protein 2 in Chlamydial Infection

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.10.1.103-107.2003 ◽

2003 ◽

Vol 10 (1) ◽

pp. 103-107 ◽

Cited By ~ 15

Author(s):

I. Portig ◽

J. C. Goodall ◽

R. L. Bailey ◽

J. S. H. Gaston

Keyword(s):

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Chlamydial Infection ◽

Chlamydia Pneumoniae ◽

Specific Protein ◽

Enzyme Linked Immunosorbent Assay ◽

Recent Infection ◽

Humoral Immune ◽

Immunodominant Antigen

ABSTRACT Detection of antibodies to an outer membrane protein 2 (OMP2) by enzyme-linked immunosorbent assay (ELISA) by using either the Chlamydia trachomatis- or the Chlamydia pneumoniae-specific protein was investigated. OMP2 is an immunodominant antigen giving rise to antibody responses in humans infected with different C. trachomatis serovars (A to C and D to K) or with C. pneumoniae, which could be detected by OMP2 ELISA. OMP2 ELISA is not species specific, but antibody titers were usually higher on the homologous protein. The sensitivity of this assay was high but varied according to the “gold standard” applied. Levels of antibody to C. pneumoniae OMP2 as detected by ELISA seem to return to background or near-background values within a shorter period of time compared to antibodies to C. pneumoniae detected by microimmunofluorescence (MIF), making it more likely that positive results in ELISA reflect recent infection. Thus, OMP2 ELISA has distinct advantages over MIF and commercially available ELISAs and might be a useful tool for the serodiagnosis of chlamydial infection.

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The Helicobacter pylori Autotransporter ImaA (HP0289) Modulates the Immune Response and Contributes to Host Colonization

Infection and Immunity ◽

10.1128/iai.00312-12 ◽

2012 ◽

Vol 80 (7) ◽

pp. 2286-2296 ◽

Cited By ~ 13

Author(s):

William E. Sause ◽

Andrea R. Castillo ◽

Karen M. Ottemann

Keyword(s):

Immune Response ◽

Helicobacter Pylori ◽

Membrane Protein ◽

Outer Membrane ◽

Outer Membrane Protein ◽

Dependent Manner ◽

Wild Type ◽

Content Type ◽

H Pylori ◽

Murine Infections

ABSTRACTThe human pathogenHelicobacter pyloriemploys a diverse collection of outer membrane proteins to colonize, persist, and drive disease within the acidic gastric environment. In this study, we sought to elucidate the function of the host-induced geneHP0289, which encodes an uncharacterized outer membrane protein. We first generated an isogenicH. pylorimutant that lacksHP0289and found that the mutant has a colonization defect in single-strain infections and is greatly outcompeted in mouse coinfection experiments with wild-typeH. pylori. Furthermore, we used protease assays and biochemical fractionation coupled with an HP0289-targeted peptide antibody to verify that the HP0289 protein resides in the outer membrane. Our previous findings showed that theHP0289promoter is upregulated in the mouse stomach, and here we demonstrate thatHP0289expression is induced under acidic conditions in an ArsRS-dependent manner. Finally, we have shown that theHP0289mutant induces greater expression of the chemokine interleukin-8 (IL-8) and the cytokine tumor necrosis factor alpha (TNF-α) in gastric carcinoma cells (AGS). Similarly, transcription of the IL-8 homolog keratinocyte-derived chemokine (KC) is elevated in murine infections with the HP0289 mutant than in murine infections with wild-typeH. pylori. On the basis of this phenotype, we renamed HP0289 ImaA forimmunomodulatoryautotransporter protein. Our work has revealed that genes inducedin vivoplay an important role inH. pyloripathogenesis. Specifically, the outer membrane protein ImaA modulates a component of the host inflammatory response, and thus may allowH. pylorito fine tune the host immune response based on ImaA expression.

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