Abstract
Background and Aim:The present serodiagnosis of brucellosis in livestock is based on the whole cell or smooth lipopolysaccharide of the Brucella organism in which specificity is hampered by the cross-reactivity, especially with the antibodies against Yersinia enterocolitica O:9 organism. The problem can be addressed by screening for better immunodominant antigens. Hence, the present study was undertaken to screen protein antigens of Brucella abortus for their diagnostic potential in cattle brucellosis.Materials and Methods:Protein antigens of B. abortus (n=10) non-reactive to antibodies against Y. enterocolitica O:9 were selected, expressed in Escherichia coli, assessed the reactivity of expressed recombinant proteins by Western blot, standardized indirect-enzyme-linked immunosorbent assay (ELISA) for detecting Brucella antibodies in cattle serum, and comparative evaluation was done.Results:All the selected protein antigens were expressed and in the Western blot with Brucella antibodies positive cattle serum, six recombinant (Brucella protein 26 [BP26], Cu-Zn Superoxide dismutase [SodC], B. abortus I-1885, Serine protease, Bacterioferritin, and Brucella Lumazine Synthase [BLS]) proteins showed reaction whereas none of the proteins showed reactivity with Brucella negative cattle serum. ELISA has been done using known Brucella positive and negative cattle sera samples (n=113 each) in which the performance of recombinant proteins in diagnosing brucellosis was in the order of BP26 > BLS > SodC followed by rest of the proteins. BP26 based ELISA was found to be better with area under the curve as 0.953, and diagnostic sensitivity, diagnostic specificity, and Youden’s index of 90.27%, 95.58%, and 0.8584, respectively, with the excellent agreement (k=0.85).Conclusion:BP26 could be a potential diagnostic antigen among the immunodominant proteins of B. abortus in ruling out Y. enterocolitica O:9 infection while diagnosing brucellosis in cattle herds.
Highlights
Brucellosis is a multifaceted zoonotic disease with significant animal and human health impact, caused by facultative, intracellular bacteria of genus Brucella, which comprises six main species, namely Brucella abortus, Brucella melitensis, Brucella suis, Brucella ovis, Brucella canis, and Brucella neotomae, and other recently identified species include Brucella pinnipedialis, Brucella ceti, Brucella microti, Brucella inopinata, Brucella papionis, and Brucella vulpis [1].Copyright: Nagalingam, et al Open Access
Hyperimmune serum raised against B. abortus S99 and Y. enterocolitica O:9 in rabbit, rabbit serum tested negative for Brucella antibodies, and serum samples collected from different cattle were used in this study
After confirmation of sequence identity, the plasmids of pETite N-His Kan vector containing individual inserts of twin-arginine, invB, thiB, bp26, sodC, bab-1885, serine protease, virB12, bfr, and bls, of the B. abortus S99 strain were successfully transformed into HI-Control BL21 (DE3) chemically competent cells, evident from the colony PCR with respective band size amplification for screened clones
Summary
Brucellosis is a multifaceted zoonotic disease with significant animal and human health impact, caused by facultative, intracellular bacteria of genus Brucella, which comprises six main species, namely Brucella abortus, Brucella melitensis, Brucella suis, Brucella ovis, Brucella canis, and Brucella neotomae, and other recently identified species include Brucella pinnipedialis, Brucella ceti, Brucella microti, Brucella inopinata, Brucella papionis, and Brucella vulpis [1].Copyright: Nagalingam, et al Open Access. The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. The common species involved in causing brucellosis in cattle is B. abortus. The present study was undertaken to screen protein antigens of Brucella abortus for their diagnostic potential in cattle brucellosis
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