Evaluation of the Recombinant 10-Kilodalton Immunodominant Region of the BP26 Protein of Brucella abortus for Specific Diagnosis of Bovine Brucellosis

Abstract

Brucellosis is a disease with worldwide distribution affecting animals and human beings. Brucella abortus is the causative agent of bovine brucellosis. The cross-reactions of currently available diagnostic procedures for B. abortus infection result in false-positive reactions, which make the procedures unreliable. These tests are also unable to differentiate Brucella-infected and -vaccinated animals. The present work is focused on the use of a nonlipopolysaccharide (LPS) diagnostic antigen, a recombinant 10-kDa (r10-kDa) protein of B. abortus, for specific diagnosis of brucellosis. The purified recombinant protein was used as a diagnostic antigen in plate enzyme-linked immunosorbent assay (p-ELISA) format to screen 408 bovine serum samples (70 presumptively negative, 308 random, and 30 vaccinated), and the results were compared with those of the Rose Bengal plate agglutination test (RBPT) and the standard tube agglutination test (STAT). Statistical analysis in presumptive negative samples revealed 100 and 98.41% specificity of p-ELISA with RBPT and STAT, and an agreement of 91.43% with the tests using Cohen's kappa statistics. In random samples, the agreement of p-ELISA was 77.92% and 80.52% with RBPT and STAT, respectively. p-ELISA investigation of vaccinated samples reported no false-positive results, whereas RBPT and STAT reported 30% and 96.6% false-positive results, respectively. The data suggest that p-ELISA with r10-kDa protein may be a useful method for diagnosis of bovine brucellosis. Furthermore, p-ELISA may also be used as a tool for differentiating Brucella-vaccinated and naturally infected animals.Brucellosis, a contagious disease primarily affecting animals, is caused by members of the genus Brucella, namely, Brucella abortus (cattle, bison, and buffalo), B. melitensis (goats and sheep), B. suis (swine), B. canis (dogs), B. ovis (sheep), and B. neotomae (rodents). Bovine brucellosis is caused by biovars of B. abortus. The disease is asymptomatic in animals that have not conceived. Following infection with B. abortus, pregnant adult females develop placentitis, resulting in abortion between the fifth and ninth months of pregnancy and infertility. Adult male cattle may develop orchitis, leading to infertility. Brucellosis has zoonotic potential in terms of its transmissibility to human beings attending infected livestock, showing symptoms in the form of low-grade undulant fever, night sweats, early fatigue, joint pain leading to spondylitis, and orchitis, and it is commonly known as "undulant fever," "Mediterranean fever," or "Malta fever" (7). The disease remains a public health problem and results in severe economic losses in terms of loss of fetuses and loss of animal products. In India, bovine brucellosis is widespread and appears to be on the increase in recent times, perhaps due to increased trade, free grazing, and the preponderance of natural bull service and rapid movement of livestock (17).Due to its heterogeneous and poorly specific symptoms, diagnosis of brucellosis always requires labora...

Identification of a protective protein from stationary-phase exoproteome ofBrucella abortus

“…Omp25, Omp2b and a hypothetical protein were predicted as outer membrane proteins. Another protein identified in the present work, Omp28 (also called BP26), has been described to have the diagnostic potential in bovine brucellosis by our group earlier (Tiwari et al, 2011). One of the predicted unknown proteins of this study, 31-kDa surface protein, has been shown to be protective in nature (Cassataro et al, 2005).…”

“…One of the predicted unknown proteins of this study, 31-kDa surface protein, has been shown to be protective in nature (Cassataro et al, 2005). Another protein identified in the present work, Omp28 (also called BP26), has been described to have the diagnostic potential in bovine brucellosis by our group earlier (Tiwari et al, 2011). Three proteins in this study were predicted as cytoplasmic proteins.…”

Limitations of the BP26 Protein-Based Indirect Enzyme-Linked Immunosorbent Assay for Diagnosis of Brucellosis

“…Chaudhuri et al found that the Omp28-based ELISA showed 88.7% sensitivity and 93.8% specificity when used in the detection of bovine brucellosis (16). In 2011, Tiwari et al (28) found that the a For the detection of infection in sheep, the relative specificity and sensitivity of BP26-based i-ELISA to LPS-based i/c-ELISAs were 90% and 82.2%, respectively; (agreement) ϭ 0.5677 for the BP26-based i-ELISA and LPS-based i/c-ELISAs. For the detection in goats, the relative specificity and sensitivity of BP26-based i-ELISA to LPS-based i/c-ELISAs were 100% and 80%, respectively; ϭ 0.5246 for the BP26-based i-ELISA and LPS-based i/c-ELISAs.…”

Biological Warfare Agents and their Detection and Monitoring Techniques (Review Paper)

“…The unbound reagents are removed by washing and the enzyme is allowed to interact with its substrate to yield a coloured product that is quantitatively measured by optical density or can be visualised directly. ELISA is a simple, 20,21 , Brucella abortus 22 , Ebola and Marburg viruses 23 or to detect bio-threat toxins 24 . Besides, some other variants of immunological assays like fluorescent microscopy 25,26 are also used for detection of BWAs.…”