scholarly journals Composition of the Surface Proteome of Anaplasma marginale and Its Role in Protective Immunity Induced by Outer Membrane Immunization

2008 ◽  
Vol 76 (5) ◽  
pp. 2219-2226 ◽  
Author(s):  
Susan M. Noh ◽  
Kelly A. Brayton ◽  
Wendy C. Brown ◽  
Junzo Norimine ◽  
Gerhard R. Munske ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Protective Immunity ◽  
Vaccine Development ◽  
Bacterial Pathogen ◽  
Protein Composition ◽  
Anaplasma Marginale ◽  
Surface Proteins ◽  
Mammalian Host ◽  
Surface Complexes ◽  
Surface Proteome

ABSTRACT Surface proteins of tick-borne, intracellular bacterial pathogens mediate functions essential for invasion and colonization. Consequently, the surface proteome of these organisms is specifically relevant from two biological perspectives, induction of protective immunity in the mammalian host and understanding the transition from the mammalian host to the tick vector. In this study, the surface proteome of Anaplasma marginale, a tick-transmitted bacterial pathogen, was targeted by using surface-specific cross-linking to form intermolecular bonds between adjacent proteins. Liquid chromatography and tandem mass spectroscopy were then employed to characterize the specific protein composition of the resulting complexes. The surface complexes of A. marginale isolated from erythrocytes of the mammalian host were composed of multiple membrane proteins, most of which belong to a protein family, pfam01617, which is conserved among bacteria in the genus Anaplasma and the closely related genus Ehrlichia. In contrast, the surface proteome of A. marginale isolated from tick cells was much less complex and contained a novel protein, AM778, not identified within the surface proteome of organisms from the mammalian host. Immunization using the cross-linked surface complex induced protection against high-level bacteremia and anemia upon A. marginale challenge of cattle and effectively recapitulated the protection induced by immunization with whole outer membranes. These results indicate that a surface protein subset of the outer membrane is capable of inducing protective immunity and serves to direct vaccine development. Furthermore, the data support that remodeling of the surface proteome accompanies the transition between mammalian and arthropod hosts and identify novel targets for blocking transmission.

scholarly journals Simultaneous Variation of the Immunodominant Outer Membrane Proteins, MSP2 and MSP3, during Anaplasma marginale Persistence In Vivo

2003 ◽  
Vol 71 (11) ◽  
pp. 6627-6632 ◽  
Author(s):  
Kelly A. Brayton ◽  
Patrick F. M. Meeus ◽  
Anthony F. Barbet ◽  
Guy H. Palmer
Keyword(s):  
Membrane Proteins ◽  
Outer Membrane ◽  
Outer Membrane Proteins ◽  
Bacterial Pathogen ◽  
Anaplasma Marginale ◽  
Surface Proteins ◽  
Mammalian Host ◽  
Surface Coat ◽  
Vector Borne

ABSTRACT Vector-borne bacterial pathogens persist in the mammalian host by varying surface antigens to evade the existing immune response. To test whether the model of surface coat switching and immune evasion can be extended to a vector-borne bacterial pathogen with multiple immunodominant surface proteins, we examined Anaplasma marginale, a rickettsia with two highly immunogenic outer membrane proteins, major surface protein 2 (MSP2) and MSP3. The simultaneous clearance of variants of the two most immunodominant surface proteins of A. marginale followed by emergence of unique variants indicates that the switch rates and immune selection for MSP2 and MSP3 are sufficiently similar to explain the cyclic bacteremia observed during infection in the immunocompetent host.

scholarly journals Differential Expression and Sequence Conservation of the Anaplasma marginale msp2 Gene Superfamily Outer Membrane Proteins

2006 ◽  
Vol 74 (6) ◽  
pp. 3471-3479 ◽  
Author(s):  
Susan M. Noh ◽  
Kelly A. Brayton ◽  
Donald P. Knowles ◽  
Joseph T. Agnes ◽  
Michael J. Dark ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Outer Membrane ◽  
Differential Expression ◽  
Outer Membrane Proteins ◽  
Anaplasma Marginale ◽  
Surface Proteins ◽  
Transcriptional Analysis ◽  
Mammalian Host ◽  
Anaplasma Ovis ◽  
Sequence Comparisons

ABSTRACT Bacterial pathogens in the genera Anaplasma and Ehrlichia encode a protein superfamily, pfam01617, which includes the predominant outer membrane proteins (OMPs) of each species, major surface protein 2 (MSP2) and MSP3 of Anaplasma marginale and Anaplasma ovis, Anaplasma phagocytophilum MSP2 (p44), Ehrlichia chaffeensis p28-OMP, Ehrlichia canis p30, and Ehrlichia ruminantium MAP1, and has been shown to be involved in both antigenic variation within the mammalian host and differential expression between the mammalian and arthropod hosts. Recently, complete sequencing of the A. marginale genome has identified an expanded set of genes, designated omp1-14, encoding new members of this superfamily. Transcriptional analysis indicated that, with the exception of the three smallest open reading frames, omp2, omp3, and omp6, these superfamily genes are transcribed in A. marginale-infected erythrocytes, tick midgut and salivary glands, and the IDE8 tick cell line. OMPs 1, 4, 7 to 9, and 11 were confirmed to be expressed as proteins by A. marginale within infected erythrocytes, with expression being either markedly lower (OMPs 1, 4, and 7 to 9) or absent (OMP11) in infected tick cells, which reflected regulation at the transcript level. Although the pfam01617 superfamily includes the antigenically variable MSP2 and MSP3 surface proteins, analysis of the omp1-14 sequences throughout a cycle of acute and persistent infection in the mammalian host and tick transmission reveals a high degree of conservation, an observation supported by sequence comparisons between the St. Maries strain and Florida strain genomes.

scholarly journals Linkage between Anaplasma marginale Outer Membrane Proteins Enhances Immunogenicity but Is Not Required for Protection from Challenge

2013 ◽  
Vol 20 (5) ◽  
pp. 651-656 ◽  
Author(s):  
Susan M. Noh ◽  
Joshua E. Turse ◽  
Wendy C. Brown ◽  
Junzo Norimine ◽  
Guy H. Palmer
Keyword(s):  
Membrane Proteins ◽  
Outer Membrane ◽  
Protective Immunity ◽  
Bacterial Infections ◽  
Vaccine Development ◽  
Outer Membrane Proteins ◽  
Anaplasma Marginale ◽  
Anamnestic Response ◽  
Covalent Bonds ◽  
Content Type

ABSTRACTThe prevention of bacterial infections via immunization presents particular challenges. While outer membrane extracts are often protective, they are difficult and expensive to isolate and standardize and thus are often impractical for development and implementation in vaccination programs. In contrast, individual proteins, which are easily adapted for use in subunit vaccines, tend to be poorly protective. Consequently, identification of the specific characteristics of outer membrane-based immunogens, in terms of the antigen contents and contexts that are required for protective immunity, represents a major gap in the knowledge needed for bacterial vaccine development. Using as a modelAnaplasma marginale, a persistent tick-borne bacterial pathogen of cattle, we tested two sets of immunogens to determine whether membrane context affected immunogenicity and the capacity to induce protection. The first immunogen was composed of a complex of outer membrane proteins linked by covalent bonds and known to be protective. The second immunogen was derived directly from the first one, but the proteins were individualized rather than linked. The antibody response induced by the linked immunogen was much greater than that induced by the unlinked immunogen. However, both immunogens induced protective immunity and an anamnestic response. These findings suggest that individual proteins or combinations of proteins can be successfully tested for the ability to induce protective immunity with less regard for overall membrane context. Once protective antigens are identified, immunogenicity could be enhanced by cross-linking to allow a reduced immunogen dose or fewer booster vaccinations.

scholarly journals Subdominant Antigens in Bacterial Vaccines: AM779 Is Subdominant in the Anaplasma marginale Outer Membrane Vaccine but Does Not Associate with Protective Immunity

PLoS ONE ◽  
2012 ◽  
Vol 7 (9) ◽  
pp. e46372 ◽  
Author(s):  
Saleh M. Albarrak ◽  
Wendy C. Brown ◽  
Susan M. Noh ◽  
Kathryn E. Reif ◽  
Glen A. Scoles ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Protective Immunity ◽  
Anaplasma Marginale

scholarly journals Identification of Novel Antigenic Proteins in a Complex Anaplasma marginale Outer Membrane Immunogen by Mass Spectrometry and Genomic Mapping

2005 ◽  
Vol 73 (12) ◽  
pp. 8109-8118 ◽  
Author(s):  
Job E. Lopez ◽  
William F. Siems ◽  
Guy H. Palmer ◽  
Kelly A. Brayton ◽  
Travis C. McGuire ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Membrane Proteins ◽  
Outer Membrane ◽  
Vaccine Development ◽  
Outer Membrane Proteins ◽  
Serum Antibody ◽  
Elongation Factor ◽  
Anaplasma Marginale ◽  
Antigenic Proteins ◽  
Associated Proteins

ABSTRACT Immunization with purified Anaplasma marginale outer membranes induces complete protection against infection that is associated with CD4+ T-lymphocyte-mediated gamma interferon secretion and immunoglobulin G2 (IgG2) antibody titers. However, knowledge of the composition of the outer membrane immunogen is limited. Recent sequencing and annotation of the A. marginale genome predicts at least 62 outer membrane proteins (OMP), enabling a proteomic and genomic approach for identification of novel OMP by use of IgG serum antibody from outer membrane vaccinates. Outer membrane proteins were separated by two-dimensional electrophoresis, and proteins recognized by total IgG and IgG2 in immune sera of outer membrane-vaccinated cattle were detected by immunoblotting. Immunoreactive protein spots were excised and subjected to liquid chromatography-tandem mass spectrometry. A database search of the A. marginale genome identified 24 antigenic proteins that were predicted to be outer membrane, inner membrane, or membrane-associated proteins. These included the previously characterized surface-exposed outer membrane proteins MSP2, operon associated gene 2 (OpAG2), MSP3, and MSP5 as well as recently identified appendage-associated proteins. Among the 21 newly described antigenic proteins, 14 are annotated in the A. marginale genome and include type IV secretion system proteins, elongation factor Tu, and members of the MSP2 superfamily. The identification of these novel antigenic proteins markedly expands current understanding of the composition of the protective immunogen and provides new candidates for vaccine development.

scholarly journals T Lymphocyte Immunity in Host Defence againstChlamydia trachomatisand Its Implication for Vaccine Development

1998 ◽  
Vol 9 (2) ◽  
pp. 99-109 ◽  
Author(s):  
X Yang ◽  
RC Brunham
Keyword(s):  
T Cell ◽  
Immune Responses ◽  
Protective Immunity ◽  
Vaccine Development ◽  
Research Effort ◽  
Dominant Role ◽  
Bacterial Pathogen ◽  
Cd4 T Cell ◽  
T Helper 1 ◽  
Humoral Immune

Chlamydia trachomatisis an obligate intracellular bacterial pathogen that causes several significant human infectious diseases, including trachoma, urethritis, cervicitis and salpingitis, and is an important cofactor for transmission of human immunodeficiency virus. Until very recently, over three decades of research effort aimed at developing aC trachomatisvaccine had failed, due mainly to the lack of a precise understanding of the mechanisms for protective immunity. Although most studies concerning protective immunity toC trachomatishave focused on humoral immune responses, recent studies have clearly shown that T helper-1 (Th1)-like CD4 T cell-mediated immune responses play the dominant role in protective immunity. These studies suggest a paradigm for chlamydial immunity and pathology based on the concept of heterogeneity (Th1/Th2) in CD4 T cell immune responses. This concept for chlamydial immunity offers a rational template on which to base renewed efforts for development of a chlamydial vaccine that targets the induction of cell-mediated Th1 immune responses.

scholarly journals Enterotoxigenic Escherichia coli Elicits Immune Responses to Multiple Surface Proteins

2010 ◽  
Vol 78 (7) ◽  
pp. 3027-3035 ◽  
Author(s):  
Koushik Roy ◽  
Scott Bartels ◽  
Firdausi Qadri ◽  
James M. Fleckenstein
Keyword(s):  
Outer Membrane ◽  
Dna Sequences ◽  
Vaccine Development ◽  
Membrane Vesicle ◽  
Outer Membrane Proteins ◽  
Surface Proteins ◽  
Enterotoxigenic Escherichia Coli ◽  
Virulence Determinants ◽  
Human Sera ◽  
Considerable Morbidity

ABSTRACTEnterotoxigenicEscherichiacoli(ETEC) causes considerable morbidity and mortality due to diarrheal illness in developing countries, particularly in young children. Despite the global importance of these heterogeneous pathogens, a broadly protective vaccine is not yet available. While much is known regarding the immunology of well-characterized virulence proteins, in particular the heat-labile toxin (LT) and colonization factors (CFs), to date, evaluation of the immune response to other antigens has been limited. However, the availability of genomic DNA sequences for ETEC strains coupled with proteomics technology affords opportunities to examine novel uncharacterized antigens that might also serve as targets for vaccine development. Analysis of whole or fractionated bacterial proteomes with convalescent-phase sera can potentially accelerate identification of secreted or surface-expressed targets that are recognized during the course of infection. Here we report results of an immunoproteomics approach to antigen discovery with ETEC strain H10407. Immunoblotting of proteins separated by two-dimensional electrophoresis (2DE) with sera from mice infected with strain H10407 or with convalescent human sera obtained following natural ETEC infections demonstrated multiple immunoreactive molecules in culture supernatant, outer membrane, and outer membrane vesicle preparations, suggesting that many antigens are recognized during the course of infection. Proteins identified by this approach included established virulence determinants, more recently identified putative virulence factors, as well as novel secreted and outer membrane proteins. Together, these studies suggest that existing and emerging proteomics technologies can provide a useful complement to ongoing approaches to ETEC vaccine development.

scholarly journals Anaplasma phagocytophilum Asp14 Is an Invasin That Interacts with Mammalian Host Cells via Its C Terminus To Facilitate Infection

2012 ◽  
Vol 81 (1) ◽  
pp. 65-79 ◽  
Author(s):  
Amandeep Kahlon ◽  
Nore Ojogun ◽  
Stephanie A. Ragland ◽  
David Seidman ◽  
Matthew J. Troese ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Anaplasma Phagocytophilum ◽  
Affinity Purification ◽  
Surface Protein ◽  
Surface Proteins ◽  
Host Cells ◽  
Mammalian Host ◽  
Content Type

Anaplasma phagocytophilum, a member of the familyAnaplasmataceae, is the tick-transmitted obligate intracellular bacterium that causes human granulocytic anaplasmosis. The life cycle ofA. phagocytophilumis biphasic, transitioning between the noninfectious reticulate cell (RC) and infectious dense-cored (DC) forms. We analyzed the bacterium's DC surface proteome by selective biotinylation of surface proteins, NeutrAvidin affinity purification, and mass spectrometry. Transcriptional profiling of selected outer membrane protein candidates over the course of infection revealed thataph_0248(designatedasp14[14-kDaA. phagocytophilumsurface protein]) expression was upregulated the most duringA. phagocytophilumcellular invasion.asp14transcription was induced during transmission feeding ofA. phagocytophilum-infected ticks on mice and was upregulated when the bacterium engaged its receptor, P-selectin glycoprotein ligand 1. Asp14 localized to theA. phagocytophilumsurface and was expressed duringin vivoinfection. Treating DC organisms with Asp14 antiserum or preincubating mammalian host cells with glutathioneS-transferase (GST)–Asp14 significantly inhibited infection of host cells. Moreover, preincubating host cells with GST-tagged forms of both Asp14 and outer membrane protein A, anotherA. phagocytophiluminvasin, pronouncedly reduced infection relative to treatment with either protein alone. The Asp14 domain that is sufficient for cellular adherence and invasion lies within the C-terminal 12 to 24 amino acids and is conserved among otherAnaplasmaandEhrlichiaspecies. These results identify Asp14 as anA. phagocytophilumsurface protein that is critical for infection, delineate its invasion domain, and demonstrate the potential of targeting Asp14 in concert with OmpA for protecting against infection byA. phagocytophilumand otherAnaplasmataceaepathogens.

scholarly journals The serogroup B meningococcal outer membrane vesicle-based vaccine 4CMenB induces cross-species protection against Neisseria gonorrhoeae

2020 ◽  
Vol 16 (12) ◽  
pp. e1008602
Author(s):  
Isabelle Leduc ◽  
Kristie L. Connolly ◽  
Afrin Begum ◽  
Knashka Underwood ◽  
Stephen Darnell ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Outer Membrane ◽  
Vaccine Development ◽  
Membrane Vesicle ◽  
Surface Proteins ◽  
Mouse Serum ◽  
Infection Model ◽  
Outer Membrane Vesicle ◽  
Species Protection ◽  
Significant Difference

There is a pressing need for a gonorrhea vaccine due to the high disease burden associated with gonococcal infections globally and the rapid evolution of antibiotic resistance in Neisseria gonorrhoeae (Ng). Current gonorrhea vaccine research is in the stages of antigen discovery and the identification of protective immune responses, and no vaccine has been tested in clinical trials in over 30 years. Recently, however, it was reported in a retrospective case-control study that vaccination of humans with a serogroup B Neisseria meningitidis (Nm) outer membrane vesicle (OMV) vaccine (MeNZB) was associated with reduced rates of gonorrhea. Here we directly tested the hypothesis that Nm OMVs induce cross-protection against gonorrhea in a well-characterized female mouse model of Ng genital tract infection. We found that immunization with the licensed Nm OMV-based vaccine 4CMenB (Bexsero) significantly accelerated clearance and reduced the Ng bacterial burden compared to administration of alum or PBS. Serum IgG and vaginal IgA and IgG that cross-reacted with Ng OMVs were induced by 4CMenB vaccination by either the subcutaneous or intraperitoneal routes. Antibodies from vaccinated mice recognized several Ng surface proteins, including PilQ, BamA, MtrE, NHBA (known to be recognized by humans), PorB, and Opa. Immune sera from both mice and humans recognized Ng PilQ and several proteins of similar apparent molecular weight, but MtrE was only recognized by mouse serum. Pooled sera from 4CMenB-immunized mice showed a 4-fold increase in serum bactericidal50 titers against the challenge strain; in contrast, no significant difference in bactericidal activity was detected when sera from 4CMenB-immunized and unimmunized subjects were compared. Our findings directly support epidemiological evidence that Nm OMVs confer cross-species protection against gonorrhea, and implicate several Ng surface antigens as potentially protective targets. Additionally, this study further defines the usefulness of murine infection model as a relevant experimental system for gonorrhea vaccine development.

scholarly journals Immunogenicity of Anaplasma marginale Type IV Secretion System Proteins in a Protective Outer Membrane Vaccine

2007 ◽  
Vol 75 (5) ◽  
pp. 2333-2342 ◽  
Author(s):  
Job E. Lopez ◽  
Guy H. Palmer ◽  
Kelly A. Brayton ◽  
Michael J. Dark ◽  
Stephanie E. Leach ◽  
...  
Keyword(s):  
T Cell ◽  
Outer Membrane ◽  
Acute Infection ◽  
Anaplasma Marginale ◽  
Secretion System ◽  
Type Iv Secretion ◽  
Surface Proteins ◽  
Type Iv Secretion System ◽  
Type Iv ◽  
T Lymphocyte Proliferation

ABSTRACT Rickettsial pathogens in the genera Anaplasma and Ehrlichia cause acute infection in immunologically naive hosts and are major causes of tick-borne disease in animals and humans. Immunization with purified outer membranes induces protection against acute Anaplasma marginale infection and disease, and a proteomic and genomic approach recently identified 21 proteins within the outer membrane immunogen in addition to the well-characterized major surface proteins MSP1 to MSP5. Among the newly described proteins were the type IV secretion system (TFSS) proteins VirB9, VirB10, and conjugal transfer protein (CTP). In other gram-negative bacteria, TFSS proteins form channels, facilitate secretion of molecules, and are required for intracellular survival. However, TFSS proteins have not been explored as vaccine antigens. In this study we demonstrate that in Anaplasma marginale outer membrane-vaccinated cattle, VirB9, VirB10, and CTP are recognized by serum immunoglobulin G2 (IgG2) and stimulate memory T-lymphocyte proliferation and gamma interferon secretion. VirB9 induced the greatest proliferation in CD4+ T-cell lines, and VirB9-specific CD4+ T-cell clones responded to three A. marginale strains, confirming the VirB9-specific T-cell responses are directed against epitopes in the native protein. The three TFSS proteins are highly conserved with orthologous proteins in Anaplasma phagocytophilum, Ehrlichia chaffeensis, and Ehrlichia canis. Recognition of TFSS antigens by CD4+ T cells and by IgG2 from cattle immunized with the protective outer membrane fraction provides a rationale for including these proteins in development of vaccines against A. marginale and related pathogens.

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