Detection of Brucella melitensis in goats by conventional and molecular assays
Title: Detection of Brucella melitensis in goats by conventional and molecular assays
Authors: NK Sandhu, NS Sharma, Paviter Kaur and Y Sandhu
Source: Ruminant Science (2019)-8(1):13-18
Cite this reference as: Sandhu NK, Sharma NS, Kaur Paviter and Sandhu Y (2019). Detection of Brucella melitensis in goats by conventional and molecular assays. Ruminant Science 8(1):13-18.
Brucellosis in goats is an economically important disease characterised by abortion, retained placenta, weak offspring and infertility. Though Brucella spp. show a host specificity, but some strains can be transmitted among a variety of animals, including humans. In the present study, a total of six isolates of B. melitensis isolated from 30 clinical samples of fetal stomach contents, vaginal swabs and discharges, placenta, uterine discharges and aborted material of goats were characterized using biochemical and molecular assays. Bruce ladder PCR was employed for detection and species identification of various Brucella isolates to study the prevalence patterns and to check whether the organisms were isolated from their natural hosts or not. By PCR using B4/B5 primer pair, an amplicon size of 223 bp was obtained thus confirming the isolates to be that of Brucella spp. By Bruce Ladder Multiplex PCR, amplified product size of 1682bp, 1071bp, 794 bp, 587bp, 450bp and 152bp was obtained in all the isolates thus confirming all the isolates as B. melitensis. DNA extraction directly from clinical samples of foetal stomach content, vaginal mucus, aborted material (spleen, liver), and cotyledons were carried out with the help of commercially available DNA extraction kit. Of the 30 DNA extracted directly from clinical samples, nine samples were positive for Brucella spp. by PCR using B4/B5 primer pair and amplicon size of 223 bp was obtained. Thus, species identification of Brucella spp. using multiplex assays will further help in understanding the epidemiology of brucellosis for effective control programs.
Alton GG, Jones LM, Angus ER and Verger JM (1988). Techniques for the Brucellosis Laboratory. 1st Edn, Paris. 22-63.
Baily GG, Krahn JB, Drasar BS and Stocker NG (1992). Detection of Brucella melitensis and B. abortus by DNA amplification. Journal of Tropical Medicine and Hygiene 95:271-275.
Bricker BJ and Halling SM (1994). Differentiation of Brucella abortus bv.1, 2, and 4, Brucella melitensis, Brucella ovis, and Brucella suis bv. 1 by PCR. Journal of Clinical Microbiology 32:2660-2666.
Bricker BJ (2002). PCR as a diagnostic tool for brucellosis. Veterinary Microbiology 90:435-446.
Dadawala AI, Chandel BS, Chauhan HC, Kumar Pankaj, Bhagat AG and Shah NM (2012). Detection of brucella antibody by using different serological tests in cattle from north Gujarat. Ruminant Science 1(2):145-148.
Ewalt DR and Bricker BJ (2000). Validation of the abbreviated Brucella AMOS PCR as a rapid screening method for differentiation of Brucella abortus field strain isolates and the vaccine strains, 19 and RB51. Journal of Clinical Microbiology 38(8):3085-3086.
Fekete A, Bantle JA and Halling SM (1990). Detection of Brucella by polymerase chain reaction in bovine fetal and maternal tissues. Journal of Veterinary Diagnostic Investigation 4:79-83.
Garcia-Yoldi D, Marin CM, De Miguel PM, Munoz PM, Vizmanos JL and Lopez-Goni I (2006). Multiplex PCR assay for the identification and differentiation of all Brucella species and the vaccine strains Brucella abortus S19 and RB51 and Brucella melitensis Rev1. Clinical Chemistry 52:779-781.
Godfroid J, Nielsen K and Saegerman C (2010) Diagnosis of brucellosis in livestock and wildlife. Croation Medicine Journal 51:296-305.
Gupta VK, Verma DK, Rout PK, Singh SV and Vihan VS (2006). Polymerase chain reaction (PCR) for detection of Brucella melitensis in goat milk. Small Ruminant Research 65(1-2):79-84.
Hamidi A, Scholl A, Dreshaj D, Robaj A, Sylejmani D, Ramadani N, Dahouk S and Nockler K (2015). Isolation and identification of Brucella melitensis biovar 3 from vaccinated small ruminants: A public health threat in Kosovo. Transboundary and Emerging Diseases 63(6):296-299.
Herman L and De-Ridder H (1992) Identification of Brucella spp. by using the polymerase chain reaction. Applied Environmental Microbiology 58:2099-3001.
Hinic V, Brodard I, Thomann A, Cevtnic Z, Makaya PV, Frey J and Abril C (2008). Novel identification and differentiation of Brucella melitensis, B. abortus, B. suis, B. ovis, B. canis, and B. neotomae suitable for both conventional and real-time PCR systems. Journal of Microbial Methods 75(2):375-8.
Lopez-Goni I, Garcia-Yoldi D, Marin M, De-Miguel J and Munoz M (2008). Evaluation of a Multiplex PCR assay (Bruce-ladder) for molecular typing of all Brucella species, including the vaccine strains. Journal of Clinical Microbiology 46:3484-3487.
Maymona AM, Mohamed TS, Abdulwahab YA and Musa TM (2014). Phenotypic characterization of Brucella melitensis isolated from livestock in Abu Dhabi Emirates. African Journal of Microbiology Research 8(39):3523-28.
Mirnejad R, Vahdati AR, Ahmadi A, Mortazavi SM and Piranfar V (2013). Comparison of culture and multiplex PCR technique for detection of Brucella abortus and Brucella melitensis from human blood samples. Zahedan Journal of Research in Medical Sciences 15(12):5-8.
OIE (2009). Bovine Brucellosis (Chapter 2.4.3). OIE Terrestrial Manual 1-35.
OIE (2016). Brucellosis (Brucella abortus, B.melitensis and B. suis) (Chapter 2.1.4.OIE Terrestrial Manual) 1- 44.
Panchasara HH, Patel JS and Patel PR (2015). Seroprevalence of brucella infection in organized farms and field conditions in Gujarat. Ruminant Science 4(1):37-41.
Patel KB, Patel SI, Patel BK, Chauhan HC, Bhagat AG and Chandel BS (2017). Comparative study on cultural and molecular methods for detection of Brucella melitensis in sheep and goat. Ruminant Science 6(2):309-314.
Preena P, Wasudeorao UW, Ganesan PI, Ronald BSM, Balakrishnan S and Vibin V (2018). Bilateral epididymo-orchitis caused by Brucella melitensis in a Madras red sheep. Ruminant Science 7(1):145-147.
Poester FP, Nielsen K, Samartino LE and Yu WL (2010). Diagnosis of brucellosis. Open Veterinary Science Journal 4:46-60.
Polding JB (1942). Brucellosis in India. Indian Journal of Veterinary Sciences 13:27-34.
Quinn PJ, Carter ME, Markey BR and Carter GR (1994). Clinical Veterinary Microbiology. Wolfe Publishing, an imprint of Mosky Year Book, Europe Limited, Spain: 268-272.
Sahin M (2008). Investigation of bovine brucellosis in Northeastern Turkey. Tropical Animal Health and Production 40(4):281-286.
Sung K, Moon H, Jong WK, Ji YK, Kyung YK, Yun MH and Suk CJ (2011). Advanced multiplex PCR assay for differentiation of Brucella species. Applied and Environmental Microbiology 77:6726-6728.
Teixeria-Gomes AP, Cloeckaert A and Zygmunt M (2000). Characterization of heat, oxidative and acid stress responses in Brucella melitensis. American Society for Microbiology 68(5):2954-2961.
Tekle M, Legesse M, Edao BM, Ameni G and Mamo G (2019). Isolation and identification of Brucella melitensis using bacteriological and molecular tools from aborted goats in the Afar region of North-eastern Ethiopia. BMC Microbiology 19:108.
Yu WL and Nielsen K (2010). Review of detection of Brucella spp. by polymerase chain reaction. Croatian Medical Journal 51:306-313.