title: HspX is present within Mycobacterium paratuberculosis-infected macrophages and is recognized by sera from some infected cattle.
abstract: A portion of the gene encoding HspX has been previously identified as a sequence specific to Mycobacterium avium subspecies paratuberculosis (hereafter referred to as M. paratuberculosis) based on DNA hybridization experiments. In this study, rabbit antisera were raised against a recombinant protein of HspX fused to the Escherichia coli maltose binding protein (MBP/HspX). Immunoblots of lysates of M. paratuberculosis-infected macrophages probed with the rabbit antisera showed that HspX was present within infected macrophages of bovine and murine origin. This observation was confirmed by immunofluorescence microscopy of infected macrophages. Lysates of E. coli expressing HspX without the MBP fusion partner were loaded onto preparative SDS-PAGE gels and used to determine whether infected cattle generated a humoral immune response to the antigen. Sera from four of 24 paratuberculous cows (17%) detected HspX. No reactivity was present in sera from control cows. While HspX may be immunogenic during infection in some cows, the protein is not secreted and it does not stimulate cell-mediated immunity. Collectively, these data give a preliminary characterization of the first described M. paratuberculosis protein identified within infected macrophages.
authors.affiliation: National Animal Disease Center, ARS-USDA, Ames, IA 50010, USA. jbannant@nadc.ars.usda.gov
authors.forename: J P
authors.initials: JP
authors.lastname: Bannantine
authors.forename: J R
authors.initials: JR
authors.lastname: Stabel
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000032:analysis / Q000096:biosynthesis; D000942:Antigens, Bacterial; D001426:Bacterial Proteins / Q000032:analysis*; D002417:Cattle; D002418:Cattle Diseases / Q000276:immunology / Q000382:microbiology*; D002478:Cells, Cultured; D007163:Immunosorbent Techniques / Q000662:veterinary; D008264:Macrophages / Q000737:chemistry / Q000276:immunology / Q000382:microbiology*; D008856:Microscopy, Fluorescence / Q000662:veterinary; D016927:Mycobacterium avium subsp. paratuberculosis / Q000302:isolation & purification*; D010283:Paratuberculosis / Q000276:immunology / Q000382:microbiology*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000032:analysis
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C118127:HspX protein, Mycobacterium tuberculosis; D011993:Recombinant Fusion Proteins