Small Ruminant Research 66 (2006) 169–174 Single-step PCR for detection of Brucella melitensis from tissue and blood of goats V.K. Gupta ∗ , Deepak K. Verma, Kalpana Singh, Ranjeeta Kumari, S.V. Singh, V.S. Vihan Central Institute for Research on Goats, Makhdoom, PO. Farah, Mathura, Uttar Pradesh 281122, India Received 8 June 2005; received in revised form 18 August 2005; accepted 1 September 2005 Available online 28 October 2005 Abstract To overcome some of the limitations of conventional microbiological techniques in the diagnosis of brucellosis in goats, a simple single-step tissue and blood PCR, based on amplification of the 720-bp sequence of a gene coding for synthesis of an immunogenic outer membrane protein specific for Brucella melitensis (omp31), was evaluated on samples collected from 76 goats at time of slaughter. PCR results were compared with serology performed on sera from the same animals. In controlled experiments, sensitivity and specificity of tissue PCR was found to be 86 and 100%, respectively. The sensitivity of blood PCR was slightly higher (93%), with a specificity of 100%. However, both serological tests exhibited a low level of sensitivity and specificity at 71 and 86%, respectively. Since this PCR assay can be performed in 1 day, is very reproducible, easily standardized and avoids the risk of infection in laboratory workers, it will complement serological testing and be a practical and reliable tool for diagnosis of brucellosis in goats. © 2005 Elsevier B.V. All rights reserved. Keywords: PCR; Brucellosis; Goats; Tissue; Blood 1. Introduction Brucellosis in goats, caused by Brucella melitensis, frequently result in abortions and diminished levels of milk production. Once the acute period of the disease is over, animals may present with little or no disease symp- toms and Brucella cells can chronically be located in the supramammary lymph nodes and mammary glands of 80% of infected animals; thus, animals continue to secrete the pathogen in their body fluids (Morgan ∗ Corresponding author. Tel.: +91 565 276 3260; fax: +91 565 276 3246. E-mail address: vivek@cirg.res.in (V.K. Gupta). and McDiarmid, 1960; Cordes and Carter, 1979; Ruiz- Castaneda, 1986; Breitmeyer et al., 1992). In rural areas of India, humans often consume milk in the form of raw milk and cheese; thus, the pathogen could easily be acquired by consumers. Therefore, there is no doubt that control of this disease in goats will have an imme- diate effect on the incidence of this disease in humans. Diagnosis of brucellosis in goats is currently based on serological and microbiological tests. It is well known that serological methods are not always sensitive or specific (Perry and Bundle, 1990; D´ ıaz-Aparicio et al., 1994). Moreover, they have repeatedly been reported to cross-react with antigens other than those from Bru- cella spp. (Perry and Bundle, 1990; D´ ıaz-Aparicio et al., 0921-4488/$ – see front matter © 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.smallrumres.2005.09.001