A novel Lawsonia intracellularis autotransporter protein is a prominent antigen

Eleanor Watson, Ewan M. Clark, M. Pilar Alberdi, Neil F. Inglis, Megan E. Porter, Lisa Imrie, Kevin McLean, Erin D. T. Manson, Alex F. Lainson, David George Emslie Smith

    Research output: Contribution to journalArticle

    Abstract

    Investigation of antigenic determinants of the microaerophilic obligate intracellular bacterium Lawsonia intracellularis using a mass spectrometry approach identified a novel bacterial protein present in an extract of cell culture medium aspirated from heavily infected in vitro cell cultures. Western immunoblotting analysis of SDS-PAGE-resolved proteins using immune sera pooled from L. intracellularis-infected pigs revealed the presence of a strongly immunoreactive band of ∼ 72 kDa. Liquid chromatography-electrospray ionization-tandem mass spectrometry analysis of this component and database mining using a fully annotated L. intracellularis genome sequence and the comprehensive GenBank prokaryotic genomic database highlighted the presence of a protein that shares little sequence similarity with other prokaryotic proteins and appears to be highly species specific. Detailed bioinformatic analyses identified the protein as member of the autotransporter protein family of surface-exposed proteins, and the designation LatA (Lawsonia autotransporter protein A) is suggested. Recognition of recombinant LatA on Western blots by a panel of sera from infected and control pigs corresponded 100% with a commercial serodiagnostic that relies on in vitro culture of this fastidious organism. LatA therefore represents a potential candidate for the development of a rapid and species-specific serodiagnostic reagent.

    Original languageEnglish
    Pages (from-to)1282-1287
    Number of pages6
    JournalClinical and Vaccine Immunology
    Volume18
    Issue number8
    DOIs
    Publication statusPublished - Aug 2011

    Fingerprint

    Lawsonia Bacteria
    Staphylococcal Protein A
    Antigens
    Proteins
    Swine
    Cell Culture Techniques
    Western Blotting
    Databases
    Bacterial Proteins
    Electrospray Ionization Mass Spectrometry
    Nucleic Acid Databases
    Tandem Mass Spectrometry
    Computational Biology
    Liquid Chromatography
    Culture Media
    Immune Sera
    Epitopes
    Polyacrylamide Gel Electrophoresis
    Mass Spectrometry
    Membrane Proteins

    Keywords

    • Animals
    • Antibodies, Bacterial
    • Antigens, Bacterial
    • Blotting, Western
    • Computational Biology
    • Desulfovibrionaceae Infections
    • Immunoassay
    • Lawsonia Bacteria
    • Mass Spectrometry
    • Membrane Transport Proteins
    • Swine

    Cite this

    Watson, E., Clark, E. M., Alberdi, M. P., Inglis, N. F., Porter, M. E., Imrie, L., ... Smith, D. G. E. (2011). A novel Lawsonia intracellularis autotransporter protein is a prominent antigen. Clinical and Vaccine Immunology, 18(8), 1282-1287. https://doi.org/10.1128/CVI.05073-11
    Watson, Eleanor ; Clark, Ewan M. ; Alberdi, M. Pilar ; Inglis, Neil F. ; Porter, Megan E. ; Imrie, Lisa ; McLean, Kevin ; Manson, Erin D. T. ; Lainson, Alex F. ; Smith, David George Emslie. / A novel Lawsonia intracellularis autotransporter protein is a prominent antigen. In: Clinical and Vaccine Immunology. 2011 ; Vol. 18, No. 8. pp. 1282-1287.
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    abstract = "Investigation of antigenic determinants of the microaerophilic obligate intracellular bacterium Lawsonia intracellularis using a mass spectrometry approach identified a novel bacterial protein present in an extract of cell culture medium aspirated from heavily infected in vitro cell cultures. Western immunoblotting analysis of SDS-PAGE-resolved proteins using immune sera pooled from L. intracellularis-infected pigs revealed the presence of a strongly immunoreactive band of ∼ 72 kDa. Liquid chromatography-electrospray ionization-tandem mass spectrometry analysis of this component and database mining using a fully annotated L. intracellularis genome sequence and the comprehensive GenBank prokaryotic genomic database highlighted the presence of a protein that shares little sequence similarity with other prokaryotic proteins and appears to be highly species specific. Detailed bioinformatic analyses identified the protein as member of the autotransporter protein family of surface-exposed proteins, and the designation LatA (Lawsonia autotransporter protein A) is suggested. Recognition of recombinant LatA on Western blots by a panel of sera from infected and control pigs corresponded 100{\%} with a commercial serodiagnostic that relies on in vitro culture of this fastidious organism. LatA therefore represents a potential candidate for the development of a rapid and species-specific serodiagnostic reagent.",
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    author = "Eleanor Watson and Clark, {Ewan M.} and Alberdi, {M. Pilar} and Inglis, {Neil F.} and Porter, {Megan E.} and Lisa Imrie and Kevin McLean and Manson, {Erin D. T.} and Lainson, {Alex F.} and Smith, {David George Emslie}",
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    Watson, E, Clark, EM, Alberdi, MP, Inglis, NF, Porter, ME, Imrie, L, McLean, K, Manson, EDT, Lainson, AF & Smith, DGE 2011, 'A novel Lawsonia intracellularis autotransporter protein is a prominent antigen', Clinical and Vaccine Immunology, vol. 18, no. 8, pp. 1282-1287. https://doi.org/10.1128/CVI.05073-11

    A novel Lawsonia intracellularis autotransporter protein is a prominent antigen. / Watson, Eleanor; Clark, Ewan M.; Alberdi, M. Pilar; Inglis, Neil F.; Porter, Megan E.; Imrie, Lisa; McLean, Kevin; Manson, Erin D. T.; Lainson, Alex F.; Smith, David George Emslie.

    In: Clinical and Vaccine Immunology, Vol. 18, No. 8, 08.2011, p. 1282-1287.

    Research output: Contribution to journalArticle

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    T1 - A novel Lawsonia intracellularis autotransporter protein is a prominent antigen

    AU - Watson, Eleanor

    AU - Clark, Ewan M.

    AU - Alberdi, M. Pilar

    AU - Inglis, Neil F.

    AU - Porter, Megan E.

    AU - Imrie, Lisa

    AU - McLean, Kevin

    AU - Manson, Erin D. T.

    AU - Lainson, Alex F.

    AU - Smith, David George Emslie

    PY - 2011/8

    Y1 - 2011/8

    N2 - Investigation of antigenic determinants of the microaerophilic obligate intracellular bacterium Lawsonia intracellularis using a mass spectrometry approach identified a novel bacterial protein present in an extract of cell culture medium aspirated from heavily infected in vitro cell cultures. Western immunoblotting analysis of SDS-PAGE-resolved proteins using immune sera pooled from L. intracellularis-infected pigs revealed the presence of a strongly immunoreactive band of ∼ 72 kDa. Liquid chromatography-electrospray ionization-tandem mass spectrometry analysis of this component and database mining using a fully annotated L. intracellularis genome sequence and the comprehensive GenBank prokaryotic genomic database highlighted the presence of a protein that shares little sequence similarity with other prokaryotic proteins and appears to be highly species specific. Detailed bioinformatic analyses identified the protein as member of the autotransporter protein family of surface-exposed proteins, and the designation LatA (Lawsonia autotransporter protein A) is suggested. Recognition of recombinant LatA on Western blots by a panel of sera from infected and control pigs corresponded 100% with a commercial serodiagnostic that relies on in vitro culture of this fastidious organism. LatA therefore represents a potential candidate for the development of a rapid and species-specific serodiagnostic reagent.

    AB - Investigation of antigenic determinants of the microaerophilic obligate intracellular bacterium Lawsonia intracellularis using a mass spectrometry approach identified a novel bacterial protein present in an extract of cell culture medium aspirated from heavily infected in vitro cell cultures. Western immunoblotting analysis of SDS-PAGE-resolved proteins using immune sera pooled from L. intracellularis-infected pigs revealed the presence of a strongly immunoreactive band of ∼ 72 kDa. Liquid chromatography-electrospray ionization-tandem mass spectrometry analysis of this component and database mining using a fully annotated L. intracellularis genome sequence and the comprehensive GenBank prokaryotic genomic database highlighted the presence of a protein that shares little sequence similarity with other prokaryotic proteins and appears to be highly species specific. Detailed bioinformatic analyses identified the protein as member of the autotransporter protein family of surface-exposed proteins, and the designation LatA (Lawsonia autotransporter protein A) is suggested. Recognition of recombinant LatA on Western blots by a panel of sera from infected and control pigs corresponded 100% with a commercial serodiagnostic that relies on in vitro culture of this fastidious organism. LatA therefore represents a potential candidate for the development of a rapid and species-specific serodiagnostic reagent.

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    KW - Antibodies, Bacterial

    KW - Antigens, Bacterial

    KW - Blotting, Western

    KW - Computational Biology

    KW - Desulfovibrionaceae Infections

    KW - Immunoassay

    KW - Lawsonia Bacteria

    KW - Mass Spectrometry

    KW - Membrane Transport Proteins

    KW - Swine

    U2 - 10.1128/CVI.05073-11

    DO - 10.1128/CVI.05073-11

    M3 - Article

    VL - 18

    SP - 1282

    EP - 1287

    JO - Clinical and Vaccine Immunology

    JF - Clinical and Vaccine Immunology

    SN - 1556-6811

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    ER -

    Watson E, Clark EM, Alberdi MP, Inglis NF, Porter ME, Imrie L et al. A novel Lawsonia intracellularis autotransporter protein is a prominent antigen. Clinical and Vaccine Immunology. 2011 Aug;18(8):1282-1287. https://doi.org/10.1128/CVI.05073-11