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Evaluation of Four Serological Tests for the Detection of Brucellosis in Goats and Cattle Under the Field Condition of Bangladesh

Muktaderul Ahmed
Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh.

Md. Ariful Islam
Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh.

Mst. Minara Khatun
Department of Microbiology and Hygiene, Faculty of Veterinary Science, Bangladesh Agricultural University, Mymensingh-2202, Bangladesh.

Byeong-Kirl Baek
Korean Zoonoses Research Institute, Chonbuk National University, Jeonju-756, Republic of Korea.

Abstract/ Executive Summary:

As the prompt and accurate diagnosis is important for undertaking an effective control measure, the present study was undertaken to evaluate four serological tests for diagnosis of Brucella infection in goats and cattle in some selected areas of Bangladesh. Sera were collected from goats (n = 108) and cattle (n = 60). All sera were screened for Brucella specific antibody response by the rose bengal plate test (RBT), slow agglutination test (SAT), indirect enzyme-linked immunosorbent assay (i-ELISA) and competitive ELISA (c-ELISA). Isolation of Brucella organisms was performed by culturing of blood samples onto bacteriological media and identification of the organism was conducted by Gram’s staining and biochemical tests. The sensitivity and specificity of the serological tests were estimated as follows: in goats, RBT: 100 and 93.40%, SAT: 100 and 96.22%, i-ELISA: 66.67 and 92.38%, c-ELISA: 100%, respectively and in cattle, RBT: 100 and 94.92%, SAT: 100 and 96.61%, i-ELISA: 50 and 93.10%, c-ELISA: 100%, respectively. RBT was found as a suitable screening test and c-ELISA could be used as a confirmatory test for the diagnosis of Brucella infection.

Keywords: Brucellosis, Goats, Cattle, Serological tests

Location: Mymensingh district in Bangladesh.

Start Date:

End Date:

Program Area: Animal Health and Management

Commodity: Goat, Cattle

Objectives:

To compare the sensitivity and specificity of four serological tests such as: RBT, SAT, indirect ELISA (i-ELISA) and competitive ELISA (c-ELISA) for diagnosis of brucellosis in goats and cattle in Bangladesh.

Methodology:

Goat (n = 108) and cattle (n = 60) sera were randomly collected from some selected areas of Mymensingh district in Bangladesh. All sera were taken from the sexually matured animals. The RBT was performed as per standard procedure with little modifications. Briefly, 75 μL of serum was taken on a rose bengal plate or glass plate by micropipette. The rose bengal coloured antigen bottle was shaken well to ensure homogenous suspension and then 25 μL of the Rose Bengal coloured antigen was added to the serum. The antigen and serum were mixed thoroughly and waited for 5 min. The result was observed immediately after 5 min. Definite clumping/ agglutination was considered as positive reaction, whereas no clumping/agglutination was regarded as negative. Slow Agglutination Test (SAT) was carried out with EDTA as described with slight modifications. Briefly 168 μL of Serum Agglutination de Wright (SAW) buffer was pipetted in the first well and 100 μL in the second and third wells of a 96 wells microplate. Then 32 μL of serum was added in the first well (dilution 1/6.25). After mixing 100 μL was transferred to the second well (1/12.5). Then 100 μL of diluted serum was transferred to the third well and finally, 100 μL of liquid was discarded from the third well. Thus, all wells contained 100 μL of diluted serum samples. 100 μL of standardized SAW antigen was added to all wells. The plate was then incubated at 37°C for 24 h (+/- 4 h). After 24 h agglutination reaction was recorded by using a magnifying mirror against illumination source. The i-ELISA and c-ELISA were performed using commercial kits according the manufacturer’s protocol (Svanova Biotech AB, art. No. 10-2700-10, SE-751 83 and No. 10-2701-02, SE-751 83, Uppsala, Sweden). Blood samples of goat and cattle tested positive in the RBT were cultured onto nutrient agar, blood agar and brucella agar media (Becton, Dickinson and company, Sparks, MD, USA). The media were inoculated with 100 μL of processed blood sample. The inoculated media were incubated at 37°C for 3-5 days in presence of 5% CO₂. The media were examined daily. Colony morphology characteristic of Brucella were subcultured onto brucella agar to obtain pure culture. Brucella suspected pure culture was stained with Gram’s staining method. Biochemical characterization of the Brucella suspected isolates was performed by catalase, oxidase and urease tests.

Source of Information: Asian Journal of Biological Sciences, Volume: 4, Issue: 5, Page No.: 477-482, Year: 2011, ISSN 1996-3351

Article Link (Online Journal): DOI: 10.3923/ajbs.2011.477.482

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

In this study, RBT detected the highest positive reactors for Brucella specific antibody response compared to other serological tests. Thus, RBT could be used as a suitable screening test for diagnosis of brucellosis in goat and cattle. All culture positive samples were tested positive by the c-ELISA indicating that this test could be used for confirmatory diagnosis of brucellosis.

Knowledge Management: Journal