Identification of proteins of potential diagnostic value for bovine paratuberculosis

Abstract

Previously we showed that Mycobacterium paratuberculosis culture filtrates (CFs) contain more antigens that react with sera from infected cattle than do cellular extracts of the organism. The goal of the present study was to identify proteins of potential diagnostic value among these CF proteins. Proteins of potential interest were first separated by 2-DE. Roughly 240 CF protein spots were detected on CBB-stained gels using Phoretix 2D software. Of these, 83% reacted with serum from M. paratuberculosis-infected cattle in immunoblots. When bovine serum was absorbed with M. phlei antigens, however, only 37 of these antigenic protein spots were reactive. Twenty-four of these spots were selected for identification based on their immunoblot staining intensity and differences in pI and mass. A total of 14 proteins were ultimately identified by MS and BLAST searches as ModD, PepA, ArgJ, CobT, Antigen 85C, and nine hypothetical proteins. N-terminal peptide analysis of PepA, Antigen 85C, ModD, MAP1693c, MAP2168c, and MAP1022c showed that each protein has 27-39 amino acids that may function as a signal sequence suggesting they are secreted through a Sec-dependent pathway. These 14 proteins from M. paratuberculosis CF are strong candidates for use as antigens for improved serodiagnostic tests for bovine paratuberculosis.

In Silico Identification of Epitopes in Mycobacterium avium subsp. paratuberculosis Proteins That Were Upregulated under Stress Conditions

“…Studies have identified several secreted M. avium subsp. paratuberculosis proteins in culture filtrates, but MAP2872c was not one of them (8), and its involvement in pathogenesis may be worth investigating.…”

Antigenicity of Recombinant Maltose Binding Protein-Mycobacterium avium subsp. paratuberculosis Fusion Proteins with and without Factor Xa Cleaving

“…paratuberculosis antigens have been investigated for their diagnostic potential in cell-and antibody-mediated assays (11,(13)(14)(15)(16)(17)(18)(19)(20)(21)(22). Some of these M. avium subsp.…”

Enzyme-Linked Immunospot: An Alternative Method for the Detection of Interferon Gamma in Johne's Disease

“…29 Numerous studies have been carried out to identify new antigens that may be used in the currently available and new MAP immunodiagnostic tests. 1,[7][8][9]17,23,24 To date, these recently identified antigens have examined only humoral and not cell-mediated immune responses.…”

“…The use of crude antigens has been identified as a problem for the immunological diagnosis of JD in many studies. 1,[7][8][9]23,24,43 There is a need for defined specific antigens that could enable extension of the assay to identify animals likely to be immune and those with a latent infection that will eventually succumb to disease. It may enable identification of sheep that go on to develop a specific lesion type.…”