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Simultaneous identification and comparison of pathogenic Brucella species in human serum and blood samples by means of multiplex PCR and confirmation of the results by PCR-RFLP
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Elnaz Akbarzadeh1   , Jamileh Noroosi1 , Vahab Piranfar2 , Dariush Ghasemi2 , Shiva Mirkalantari3 , Reza Mirnejad4 |
1- Islamic Azad University, Tehran North Branch
2- Baqiyatallah University of Medical Sciences
3- Semnan University of Medical Sciences
4- Baqiyatallah University of Medical Sciences , rmirnejad@yahoo.com |
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Abstract: (8106 Views) |
| Background and Aim: There are few studies on the isolation of Brucella abortus and Brucella melitensis from patients´ sera. The aim of this study was simultaneous isolation of pathogenic brucella species from human serum samples by multiplex PCR method.
Material andMethods: 50 blood and serum samples isolated from patients with clinically suspected brucellosis were inoculated into brucella agar medium and we used multiplex-PCR, with three primers to detect brucella species. We incubated 0.5 ml of serum in Brucella broth for 72 hours at 37 °C in 5% carbon dioxide. To confirm the results of PCR, the PCR products were restricted by restriction enzymes, TaqI and RsaI.
Result: From 50 blood samples 4 (8%) cultures were positive. Using biochemical tests and after determination of the characteristics of the positive cases, they were identified as B. melitensis. After multiplex PCR, 9 cases (18%) were positive and 41 cases (82%) were negative. Among the positive sera, B. melitensis was identified in 7 cases (78%) and B. abortus in 2 cases (22%). Of 50 blood samples, 5 (10%) were positive and 45 (90%) were negative for B. melitensis. All the results were confirmed by PCR-RFLP.
Conclusion: Our study showed that multiplex PCR method was simple, rapid and is more sensitive for isolation of brucella from serum (especially B. abortus and B. melitensis) in comparison to complete blood.
Keywords: Brucella spp., Multiplex-PCR, Serum blood, PCR-RFLP
Received: Jul 7, 2014 Accepted: Dec 10, 2014 |
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| Keywords: Brucella spp., Multiplex-PCR, Serum blood, PCR-RFLP |
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Full-Text [PDF 1257 kb]
(2113 Downloads)
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Type of Study: Original Research |
Subject:
General
Received: 2015/03/14 | Accepted: 2015/03/14 | Published: 2015/03/14
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