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 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 Amikacin and bacteriophage treatment modulates outer membrane proteins composition in Proteus mirabilis biofilm

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Agnieszka Maszewska ◽  
Magdalena Moryl ◽  
Junli Wu ◽  
Bin Liu ◽  
Lu Feng ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Outer Membrane ◽  
Proteus Mirabilis ◽  
Protein Biosynthesis ◽  
Outer Membrane Proteins ◽  
Wild Strain ◽  
Elongation Factor ◽  
Resistance Mechanisms ◽  
Trna Synthetases ◽  
Cytoplasmic Proteins

AbstractModification of outer membrane proteins (OMPs) is the first line of Gram-negative bacteria defence against antimicrobials. Here we point to Proteus mirabilis OMPs and their role in antibiotic and phage resistance. Protein profiles of amikacin (AMKrsv), phage (Brsv) and amikacin/phage (AMK/Brsv) resistant variants of P. mirabilis were compared to that obtained for a wild strain. In resistant variants there were identified 14, 1, 5 overexpressed and 13, 5, 1 downregulated proteins for AMKrsv, Brsv and AMK/Brsv, respectively. Application of phages with amikacin led to reducing the number of up- and downregulated proteins compared to single antibiotic treatment. Proteins isolated in AMKrsv are involved in protein biosynthesis, transcription and signal transduction, which correspond to well-known mechanisms of bacteria resistance to aminoglycosides. In isolated OMPs several cytoplasmic proteins, important in antibiotic resistance, were identified, probably as a result of environmental stress, e.g. elongation factor Tu, asparaginyl-tRNA and aspartyl-tRNA synthetases. In Brsv there were identified: NusA and dynamin superfamily protein which could play a role in bacteriophage resistance. In the resistant variants proteins associated with resistance mechanisms occurring in biofilm, e.g. polyphosphate kinase, flagella basal body rod protein were detected. These results indicate proteins important in the development of P. mirabilis antibiofilm therapies.

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 Genetic Similarity of Gonococcal Homologs to Meningococcal Outer Membrane Proteins of Serogroup B Vaccine

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
Author(s):  
Henju Marjuki ◽  
Nadav Topaz ◽  
Sandeep J. Joseph ◽  
Kim M. Gernert ◽  
Ellen N. Kersh ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Outer Membrane ◽  
Vaccine Development ◽  
Membrane Vesicle ◽  
Outer Membrane Proteins ◽  
The United States ◽  
Cross Protection ◽  
Outer Membrane Vesicle ◽  
Content Type ◽  
Sequence Identity

ABSTRACT The human pathogens Neisseria gonorrhoeae and Neisseria meningitidis share high genome identity. Retrospective analysis of surveillance data from New Zealand indicates the potential cross-protective effect of outer membrane vesicle (OMV) meningococcal serogroup B vaccine (MeNZB) against N. gonorrhoeae. A licensed OMV-based MenB vaccine, MenB-4C, consists of a recombinant FHbp, NhbA, NadA, and the MeNZB OMV. Previous work has identified several abundantly expressed outer membrane proteins (OMPs) as major components of the MenB-4C OMV with high sequence similarity between N. gonorrhoeae and N. meningitidis, suggesting a mechanism for cross-protection. To build off these findings, we performed comparative genomic analysis on 970 recent N. gonorrhoeae isolates collected through a U.S surveillance system against N. meningitidis serogroup B (NmB) reference sequences. We identified 1,525 proteins that were common to both Neisseria species, of which 57 proteins were predicted to be OMPs using in silico methods. Among the MenB-4C antigens, NhbA showed moderate sequence identity (73%) to the respective gonococcal homolog, was highly conserved within N. gonorrhoeae, and was predicted to be surface expressed. In contrast, the gonococcal FHbp was predicted not to be surface expressed, while NadA was absent in all N. gonorrhoeae isolates. Our work confirmed recent observations (E. A. Semchenko, A. Tan, R. Borrow, and K. L. Seib, Clin Infect Dis, 2018, https://doi.org/10.1093/cid/ciy1061) and describes homologous OMPs from a large panel of epidemiologically relevant N. gonorrhoeae strains in the United States against NmB reference strains. Based on our results, we report a set of OMPs that may contribute to the previously observed cross-protection and provide potential antigen targets to guide the next steps in gonorrhea vaccine development. IMPORTANCE Gonorrhea, a sexually transmitted disease, causes substantial global morbidity and economic burden. New prevention and control measures for this disease are urgently needed, as strains resistant to almost all classes of antibiotics available for treatment have emerged. Previous reports demonstrate that cross-protection from gonococcal infections may be conferred by meningococcal serogroup B (MenB) outer membrane vesicle (OMV)-based vaccines. Among 1,525 common proteins shared across the genomes of both N. gonorrhoeae and N. meningitidis, 57 proteins were predicted to be surface expressed (outer membrane proteins [OMPs]) and thus preferred targets for vaccine development. The majority of these OMPs showed high sequence identity between the 2 bacterial species. Our results provide valuable insight into the meningococcal antigens present in the current OMV-containing MenB-4C vaccine that may contribute to cross-protection against gonorrhea and may inform next steps in gonorrhea vaccine development.

scholarly journals Immunization withSalmonella entericaSerovar Typhimurium-Derived Outer Membrane Vesicles Delivering the Pneumococcal Protein PspA Confers Protection against Challenge withStreptococcus pneumoniae

2010 ◽  
Vol 79 (2) ◽  
pp. 887-894 ◽  
Author(s):  
Maneesha Muralinath ◽  
Meta J. Kuehn ◽  
Kenneth L. Roland ◽  
Roy Curtiss
Keyword(s):  
Membrane Proteins ◽  
Outer Membrane ◽  
Outer Membrane Proteins ◽  
Lethal Dose ◽  
Serum Antibody ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
Gram Negative Bacteria ◽  
Vesicle Preparation ◽  
Serovar Typhimurium

ABSTRACTGram-negative bacteria produce outer membrane vesicles (OMVs) that serve a variety of functions related to survival and pathogenicity. Periplasmic and outer membrane proteins are naturally captured during vesicle formation. This property has been exploited as a method to derive immunogenic vesicle preparations for use as vaccines. In this work, we constructed aSalmonella entericaserovar Typhimurium strain that synthesized a derivative of the pneumococcal protein PspA engineered to be secreted into the periplasmic space. Vesicles isolated from this strain contained PspA in the lumen. Mice intranasally immunized with the vesicle preparation developed serum antibody responses against vesicle components that included PspA andSalmonella-derived lipopolysaccharide and outer membrane proteins, while no detectable responses developed in mice immunized with an equivalent dose of purified PspA. Mucosal IgA responses developed against theSalmonellacomponents, while the response to PspA was less apparent in most mice. Mice immunized with the vesicle preparation were completely protected against a 10× 50% lethal dose (LD50) challenge ofStreptococcus pneumoniaeand significantly protected against a 200× LD50challenge, while control mice immunized with purified PspA or empty vesicles were not protected. These results establish that vesicles can be used to mucosally deliver an antigen from a Gram-positive organism and induce a protective immune response.

scholarly journals Simultaneous detection of LipL32 and LipL21 genes of pathogenic leptospira from serum samples of bovines by multiplex PCR

2011 ◽  
Vol 1 (1) ◽  
pp. 15
Author(s):  
Timiri V. Meenambigai ◽  
Gopalakrishnan Ravikumar ◽  
Andy Srithar ◽  
Govindan Balakrishnan ◽  
Chidambaram Saranya ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Multiplex Pcr ◽  
Outer Membrane ◽  
Vaccine Development ◽  
Outer Membrane Proteins ◽  
Simultaneous Detection ◽  
Serum Samples ◽  
Pathogenic Leptospira ◽  
Annealing Conditions ◽  
Reference Strains

<p>Leptospirosis is a worldwide zoonotic disease of cattle associated with pathogenic leptospiral infection. This study focuses in the use of a molecular tool to detect pathogenic leptospiral infection in bovines by targeting the outer membrane proteins LipL32 and LipL21 simultaneously in a multiplex PCR. Sixteen pathogenic reference strains and 10 bovine serum samples were analyzed for simultaneous detection of both genes at appropriate annealing conditions. These findings are suggestive of the fact that multiplex PCR can be used to detect major outer membrane proteins of pathogenic leptospira from serum samples. Further it aided in the differentiation of pathogenic and non-pathogenic species of leptospires too. This study will definitely serve as a valuable tool, as it suggests the importance of <em>LipL32</em> genes as potential candidates for vaccine development to control animal Leptospirosis.</p>

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