A protein A/G indirect enzyme-linked immunosorbent assay for the detection of anti-Brucellaantibodies in Arctic wildlife

Abstract

Abstract.A species-independent indirect enzyme-linked immunosorbent assay (iELISA) based on chimeric protein A/G was established for the detection of anti-Brucella antibodies in Arctic wildlife species and compared to previously established brucellosis serological tests for hooded seals (Cystophora cristata), minke whales (Balaenoptera acutorostrata), sei whales (Balaenoptera borealis), fin whales (Balaenoptera physalus), and polar bears (Ursus maritimus), as well as bacteriology results for reindeer and caribou (Rangifer tarandus sp.). The protein A/G iELISA results were consistent with the other serological tests with Cohen kappa values between 0.47 and 0.92, and the protein A/G iELISA can thus offer a technically simple method for these species yielding results consistent with established brucellosis serological tests. Receiver operator characteristics analysis proved that the reindeer and caribou protein A/G iELISA results were consistent with the bacteriological gold standard with an area under the curve of 0.99, and the protein A/G iELISA was thus validated as a sensitive and specific serological method for the detection of anti-Brucella antibodies in reindeer and caribou. The binding of the antibodies from the respective species to protein A and G were also evaluated in the iELISA. The antibodies from hooded seals and polar bears reacted stronger to protein A than to G. The sei whale, fin whale, reindeer, and caribou antibodies reacted stronger to protein G than to A. The minke whale antibodies reacted to both protein A and G. There was a strong correlation (r s = 0.88-0.98) between the optical density results obtained with the iELISA with protein A/G and protein A or G, showing that protein A/G is as well suited as protein A or G for the detection of anti-Brucella antibodies in these species with the iELISA.

Brucella antibody seroprevalence in Antarctic seals (Arctocephalus gazella, Leptonychotes weddellii and Mirounga leonina)

“…A previous study on hooded seals (Nymo et al 2013) utilizing the same iELISA as in the present study and the RBT showed a Cohen's kappa value (to assess pairwise coherence) between the 2 tests of 0.52. The PrA/G iELISA yielded a seroprevalence of 23.8% (41/172), while the seroprevalence found with the RBT was 9.9% (17/172; Nymo et al 2013). This further strengthens the evidence of sample quality affecting the outcome of the RBT, as some of the serum samples used in the present study were from animals found dead, and thus of poor quality, and the sera from lactating pups were highly lipemic, while the hooded seal sera in the previous study were of good quality (I. H. Nymo pers.…”

“…The mean optical density (OD) of duplicate wells was expressed as a percentage of the reactivity of a positive serum from a bacteriology-positive hooded seal ([OD sample/OD positive control] × 100) = percent positivity (%P). The cutoff for the iELISA was calculated by the mean value of 125 negative hooded seal serum samples (negative in the Complement Fixation Test, the Slow Agglutination of Wright [SAW EDTA] and the RBT) plus 2.58 standard deviations, pro viding a cutoff for the iELISA of 73.6%P (Nymo et al 2013). …”

“…with the RBT, with Brucella abortus Weybridge 99 strain as an antigen (IDEXX Laboratories, Pourquier) according to the manufacturer's instructions (OIE 2008). An iELISA previously shown to yield results in coherence with a panel of serological tests when utilized on hooded seal Cystophora cristata serum samples was also used (Nymo et al 2013). The mean optical density (OD) of duplicate wells was expressed as a percentage of the reactivity of a positive serum from a bacteriology-positive hooded seal ([OD sample/OD positive control] × 100) = percent positivity (%P).…”

Age-dependent prevalence of anti-Brucella antibodies in hooded seals Cystophora cristata

“…Serum samples were analysed for anti-Brucella antibodies with a Protein A/G indirect enzymelinked immunosorbent assay (ELISA) for the detection of anti-Brucella antibodies in hooded seals (Nymo et al 2013). …”

Exposure of harbour seals Phoca vitulina to Brucella in declining populations across Scotland

“…Similar to this study, Brucella antibodies have been detected in sera from a number of marine mammal species using Rose Bengal tests (Tryland et al 1999, Retamal et al 2000, Hernández-Mora et al 2008, Jensen et al 2013, and using both indirect ELISA (iELISA) and cELISAs primarily designed for ruminants (Tryland et al 1999, Nielsen et al 2001, Van Bressem et al 2001, Tachibana et al 2006, Roe et al 2010, Lynch et al 2011, Jensen et al 2013, Nymo et al 2013a). Here, differences in the prevalence estimates obtained from the RBT and the cELISA results highlight the need to consider test performance when conducting serological studies.…”