Abstract
Cell envelope proteins from Mycobacterium avium subspecies paratuberculosis (MAP) that are antigenically distinct from closely related mycobacterial species are potentially useful for Johne's Disease (JD) diagnosis. We evaluated the potential of ELISAs, based on six antigenically distinct recombinant MAP cell envelope proteins (SdhA, FadE25_2, FadE3_2, Mkl, DesA2, and hypothetical protein MAP1233) as well as an extract of MAP total cell envelope proteins, to detect antibodies against MAP in the sera of infected cattle. The sensitivity (Se) and specificity (Sp) of an ELISA based on MAP total cell envelope proteins, when analyzing 153 bovine serum samples, was 75 and 96%, respectively. Analysis of the same samples, using a commercial serum ELISA resulted in a Se of 56% and Sp of 99%. Results of ELISA analysis using plates coated with recombinant cell envelope proteins ranged from a highest Se of 94% and a lowest Sp of 79% for Sdh A to a lowest Se of 67% and a highest Sp of 95% for hypothetical protein MAP1233. Using polyclonal antibodies to MAP total cell envelope proteins, immunohistochemical analysis of intestinal and lymph node tissues from JD-positive cattle detected MAP organisms whereas antibodies to recombinant proteins did not. Finally, polyclonal antibodies to MAP total cell envelope protein and to recombinant SdhA, FadE25_2, and DesA2 proteins immunomagnetically separated MAP microorganisms spiked in PBS. These results suggest that antigenically distinct MAP cell envelope proteins and antibodies to these proteins may have potential to detect MAP infection in dairy cattle.
Highlights
Johne’s disease (JD) is a non-treatable chronic granulomatous enteritis of cattle and small ruminants caused by Mycobacterium avium subspecies paratuberculosis (MAP) [1]
In the first part of this study, we investigated the use of MAP total cell envelope antigens and six recombinant MAP cell envelope proteins in ELISAs developed to identify serum antibodies to MAP bacteria in naturally infected cattle
We showed that ELISAs with MAP total cell envelope proteins using absorbed serum samples significantly increased specificity
Summary
Johne’s disease (JD) is a non-treatable chronic granulomatous enteritis of cattle and small ruminants caused by Mycobacterium avium subspecies paratuberculosis (MAP) [1]. In the silent stage I, infected animals are healthy without shedding of MAP in the feces [4]. In stage II, the disease is subclinical and infected animals appear healthy and can intermittently shed MAP in the feces, thereby contaminating the environment and acting as a source of infection to herd-mates [4]. Current laboratory tests have very limited sensitivity in the diagnosis of animals at stage I and II of infection and cattle may remain undiagnosed for several years [5]. In stages III (clinical disease) and IV (advanced clinical disease), infected animals exhibit typical clinical signs of JD such as intermittent to continuous diarrhea, weight loss, and emaciation and shed large numbers of MAP in the feces [4]
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