Autoreactive antibodies are present in sheep with Johne's disease and cross-react with Mycobacterium avium subsp. paratuberculosis antigens

Abstract

Mycobacterial infection has been linked to the generation of autoantibodies, including anti-neutrophil cytoplasmic autoantibodies (ANCA), in clinical studies and small animal models. In an attempt to identify antibodies that react with both self and mycobacterial antigens in naturally infected ruminants, we generated a phage display library comprising single chain antibody fragments (scFv) from sheep with Johne's disease (JD). The library was screened simultaneously against ovine small intestinal tissue and Mycobacterium avium subsp. paratuberculosis (MAP). A clone (termed AutoH1) reacted strongly with host tissue and MAP, recognizing a proteinase-susceptible 32 kDa determinant in ovine gut tissues and lymphatics, and in blood granulocytes but not mononuclear cells. In granulocytes, binding was to cytoplasmic granules and cell membranes; in MAP, AutoH1 bound bacterial cell wall determinants. We further identified a synthetic peptide sequence recognized by AutoH1, using this to generate a carrier:peptide fusion protein (paH1). Sera of normal and JD sheep were screened for AutoH1-like autoantibody activity; 7/11 JD animals showed autoreactivity that could be blocked by paH1, while 0/21 normal animals showed no such serological reactivity. It is possible that the severe pathology observed in ruminant JD may have an autoimmune component, possibly due to ANCA-type binding; this remains to be further investigated.