Sensitive and Specific Enzyme-Linked Immunosorbent Assay for Detecting Serum Antibodies against Mycobacterium avium subsp. paratuberculosis in Fallow Deer

Abstract

bThe enzyme-linked immunosorbent assay (ELISA) is the diagnostic test most commonly used in efforts to control paratuberculosis in domestic ruminants. However, commercial ELISAs have not been validated for detecting antibodies against Mycobacterium avium subsp. paratuberculosis in wild animals. In this study, we compared the sensitivities and specificities of five ELISAs using individual serum samples collected from 41 fallow deer with or without histopathological lesions consistent with paratuberculosis. Two target antigenic preparations were selected, an ethanol-treated protoplasmic preparation obtained from a fallow deer M. avium subsp. paratuberculosis isolate (ELISAs A and B) and a paratuberculosis protoplasmic antigen (PPA3) (ELISAs C and D). Fallow deer antibodies bound to the immobilized antigens were detected by using a horseradish peroxidase (HRP)-conjugated anti-fallow deer IgG antibody (ELISAs A and C) or HRP-conjugated protein G (ELISAs B and D). A commercially available assay, ELISA-E, which was designed to detect M. avium subsp. paratuberculosis antibodies in cattle, sheep, and goats, was also tested. Although ELISAs A, C, and E had the same sensitivity (72%), ELISAs A and C were more specific (100%) for detecting fallow deer with lesions consistent with paratuberculosis at necropsy than was the ELISA-E (87.5%). In addition, the ELISA-A was particularly sensitive for detecting fallow deer in the latent stages of infection (62.5%). The antibody responses detected with the ELISA-A correlated with both the severity of enteric lesions and the presence of acid-fast bacteria in gut tissue samples. In summary, our study shows that the ELISA-A can be a cost-effective diagnostic tool for preventing the spread of paratuberculosis among fallow deer populations.

First description of sarcoptic mange in the endangered Iberian lynx (Lynx pardinus): clinical and epidemiological features

“…The ELISA was adapted for the diagnosis of the disease in lynx by determining the best conjugate and its optimal concentration performing a preliminary checkerboard titration using reference sera samples. The best signal-to-noise ratio of the optical densities (ODs) of the positive and negative reference sera (Prieto et al 2014) was obtained using goat anti-cat horseradish peroxidase-conjugated (SIGMA, St. Louis, USA) diluted 1500 times in blocking solution. The peroxidase substrat e employed was 3, 3′ , 5,5′tetramethylbenzidine liquid substrate (TMB) (SIGMA, St. Louis, USA).…”

Molecular Characterization and Serodiagnostic Potential of Two Serpin Proteins in Psoroptes Ovis

“…Compared to rPsoSP2-iELISA, the rPsoSP1-iELISA displayed better diagnostic e ciency with higher values of sensitivity, speci city and AUC (rPsoSP1 -rPsoSP2: 96.0% − 90.0%; 90.91% − 78.18%; 0.988 − 0.964, respectively). Although rPsoSP1 showed cross-reaction with sera from 3/10 S. scabiei-infestation, the cross-reaction between these two ectoparasites have been proved commonly in other studies [3,32,33].…”

Validation of laboratory tests for infectious diseases in wild mammals: review and recommendations

“…Serum ELISAs for paratuberculosis detection in fallow deer ( Dama dama ) were evaluated, and their predictive values were interpreted together with the presence or absence of histologic lesions. 83 In the same study, ELISA-based results were compared pairwise to results from other testing techniques, including Ziehl–Neelsen staining, quantitative PCR, and bacterial culture. These multi-test comparisons for paratuberculosis indicated that pathogen exposure likely triggered antibody production without progressive infection.…”