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MICROBIAL IMMUNOLOGY

Assessment of Cross-Reactivity between Mycobacterium bovis and M. kansasii ESAT-6 and CFP-10 at the T-Cell Epitope Level

H. Martin Vordermeier, Jemma Brown, Paul J. Cockle, Willeke P. J. Franken, Sandra M. Arend, Tom H. M. Ottenhoff, Keith Jahans, R. Glyn Hewinson
H. Martin Vordermeier
1TB Research Group
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  • For correspondence: m.vordermeier@vla.defra.gsi.gov.uk
Jemma Brown
2TB Diagnostic Section, Veterinary Laboratories Agency—Weybridge, New Haw, Addlestone KT15 3NB, United Kingdom
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Paul J. Cockle
1TB Research Group
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Willeke P. J. Franken
3Department of Infectious Diseases
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Sandra M. Arend
3Department of Infectious Diseases
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Tom H. M. Ottenhoff
4Department of Immunohematology and Blood Transfusion, Leiden University Medical Center, Leiden, The Netherlands
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Keith Jahans
2TB Diagnostic Section, Veterinary Laboratories Agency—Weybridge, New Haw, Addlestone KT15 3NB, United Kingdom
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R. Glyn Hewinson
1TB Research Group
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DOI: 10.1128/CVI.00116-07
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  • FIG. 1.
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    FIG. 1.

    Alignment of the amino acid sequences and positions of dominant T-cell epitopes of ESAT-6 (A) and CFP-10 (B) of M. bovis/M. tuberculosis and M. kansasii genotypes I to V (MKA-1 to MKA-V). Amino acids are represented by the one-letter code. Promiscuous recognition of M. bovis-derived ESAT-6 and CFP-10 peptides (defined as giving rise to T-cell responses in ≥50 of the cattle tested that recognized M. bovis ESAT-6 or CFP-10) is represented by open bars above the sequences (27, 31). The proportions of animals (responder frequencies) that recognized these peptides are shown above the respective bars (27, 31). (Adapted from reference 4 with permission of the publisher.)

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    FIG. 2.

    Recognition of M. bovis and M. kansasii ESAT-6-derived peptides by T cells from cattle with BTB. PBMCs from naturally infected cattle were stimulated with synthetic peptides (see Table 1 for sequence details). The readout system indicates the IFN-γ production measured by the ex vivo ELISPOT assay. The results are expressed as the mean SFCs/well, and the standard errors of the means for all values are <20% of the means. Horizontal lines indicate the cutoff for positivity (10 ΔSFCs/well).

  • FIG. 3.
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    FIG. 3.

    Recognition of M. bovis and M. kansasii CFP-10-derived peptides by T cells from cattle with BTB. PBMCs from naturally infected cattle were stimulated with synthetic peptides (see Table 1 for sequence details). The readout system indicates the IFN-γ production measured by the ex vivo ELISPOT assay. The results are expressed as the mean SFCs/well, and the standard errors of the means for all values are <20% of the means. Horizontal lines indicate the cutoff for positivity (10 ΔSFCs/well).

  • FIG. 4.
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    FIG. 4.

    BoLA class II restriction of the recognition of M. bovis ESAT-6 peptide 3. Cross-reactive recognition of peptides 3, 3A, and 3B in animal 2 (A) was BoLA-DR restricted (B). M. bovis-specific recognition of peptide 3 in animal 1 (C) was BoLA-DQ restricted (D). Anti-BoLA-DR or anti-BoLA-DQ monoclonal antibody was added to peptide 3-stimulated PBMC cultures (B and D). The results are expressed as the mean numbers of IFN-γ-producing SFCs/well ± standard errors of the means. *, P < 0.05 compared to the results for the other groups (Student's t test).

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  • TABLE 1.

    Peptides used in this study

    Protein and peptidePeptide sequenceaSpecies (Mka genotype)b
    ESAT-6
        1 EAYQGVQQKWDATATE M. bovis
        1A --------------Q- M. kansasii (I-V)
        2 KWDATATELNNALQNL M. bovis
        2A ------Q--------- M. kansasii (I-IV)
        2B ------Q---S----- M. kansasii (V)
        3 LNNALQNLARTISEAG M. bovis
        3A --------S------- M. kansasii (I-IV)
        3B --S-----S------- M. kansasii (V)
    CFP-10
        4 QEAGNFERISGDLKTQ M. bovis
        4A ------------M--- M. kansasii (IV)
        5 IDQVESTAGSLQGQWR M. bovis
        5A --------A---A--- M. kansasii (I-V)
        6 GSLQGQWRGAAGTAAQAA M. bovis
        6A A---A------------- M. kansasii (I-V)
        6B A---A------A------ M. kansasii (IV)
        7 AGTAAQAAVVRFQEAANK M. bovis
        7A --A--------------- M. kansasii (IV)
        8 VVRFQEAANKQKQELDEI M. bovis
        8A ------------A--E-- M. kansasii (I-V)
        9 QKQELDEISTNIRQAGVQYS M. bovis
        9A --A----E--------------- M. kansasii (I-V)
        10 NIRQAGVQYSRADEEQQQ M. bovis
        10A ----------K------- M. kansasii (I-V)
    • ↵ a -, amino acid residue in M. kansasii-derived peptide identical to the residue in the M. bovis sequence. Amino acids are represented by the one-letter code.

    • ↵ b M. kansasii genotype according to Arend et al. (4).

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Assessment of Cross-Reactivity between Mycobacterium bovis and M. kansasii ESAT-6 and CFP-10 at the T-Cell Epitope Level
H. Martin Vordermeier, Jemma Brown, Paul J. Cockle, Willeke P. J. Franken, Sandra M. Arend, Tom H. M. Ottenhoff, Keith Jahans, R. Glyn Hewinson
Clinical and Vaccine Immunology Sep 2007, 14 (9) 1203-1209; DOI: 10.1128/CVI.00116-07

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Assessment of Cross-Reactivity between Mycobacterium bovis and M. kansasii ESAT-6 and CFP-10 at the T-Cell Epitope Level
H. Martin Vordermeier, Jemma Brown, Paul J. Cockle, Willeke P. J. Franken, Sandra M. Arend, Tom H. M. Ottenhoff, Keith Jahans, R. Glyn Hewinson
Clinical and Vaccine Immunology Sep 2007, 14 (9) 1203-1209; DOI: 10.1128/CVI.00116-07
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KEYWORDS

Antigens, Bacterial
Bacterial Proteins
Epitopes, T-Lymphocyte
Mycobacterium Infections, Nontuberculous
Mycobacterium bovis
Mycobacterium kansasii

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