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Evaluation of PCR for identifying Brucella species isolated from blood from patients living in a non-endemic area

Abstract number: P1978

Aftab H., Dargis R., Christensen J.J., Kemp M.

Objectives: Brucellosis is rare in Denmark. Primary laboratories are not prepared for handling these BSL class 3 organisms, and there is a considerable risk of laboratory staff being exposed to the bacterium. Traditionally, isolates of Brucella species has been referred to our national reference laboratory and final identification has been made by a combination of culture and agglutination by antisera. As speed, safety and reliability are crucial factors in identification of Brucella species we evaluated these factors using PCR as an alternative.

Methods: Seventeen isolates from blood were analysed by PCR using primer sets for B. melitensis, B. abortus, and B. suis, respectively. The three PCR analyses were run in parallel and had PCR products of different size. Eight isolates previously identified by culture and agglutination by antiserum, were re-examined by PCR, and nine consecutively referred isolates were identified by PCR only. Two of the previously identified isolates had been reported as B. melitensis, four as B. abortus, and two as Brucella species.

The PCR test was completed in less than one working day, whereas cultivation and agglutination required at least overnight incubation.

Results: All isolates were postive for B. melitensis by PCR. One isolate was repeatedly positive in PCR for B. abortus in addition to B. melitensis, however the PCR product in the B. abortus assay did not show the expected size. The DNA sequence of the PCR product from the B. abortus PCR did not allow discrimination between the species. None of the isolates were identified as B. abortus or B. suis by PCR.

Conclusions:

1PCR is faster than culture and agglutination. A definite conclusion of whether a suspected isolate is a clinical relevant Brucella species can be obtained within a few hours.

2PCR is less hazardous than culture and agglutination. Isolates referred to the reference laboratory are immediately inactivated in DNA extraction buffer, allowing all subsequent procedures to be carried out in BSL class 2 facilities. The suspected isolate may even be inactivated, e.g. by heating, before shipment. In contrast to live Brucella bacteria, inactivated material may be send by ordinary means of transportation.

3PCR results in definite species identification and there are no difficulties in interpretation of agglutination obtained by more or less cross-reacting antisera.

PCR for Brucella species is a fast, safe, and reliable method for identification of clinical isolates.

Session Details

Date: 19/04/2008
Time: 00:00-00:00
Session name: 18th European Congress of Clinical Microbiology and Infectious Diseases
Subject:
Location: Barcelona, Spain
Presentation type:
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