Evaluation of a rapid automated assay for the isolation of Mycobacterium tuberculosis DNA directly from clinical samples, using magnetic particles

Abstract number: 1733_1485

Lysén J., Høidal Berthelsen H.K., Espelund M., Jeansson S., Mengshoel A.T., Refseth U.H.

Objectives: The objective of this study was to evaluate the performance of a fully automated DNA preparation system, the BUGS'n BEADS™ MYCOBACTERIUM (Genpoint, Norway), to isolate Mycobacterium tuberculosis complex DNA from clinical respiratory samples.

Method: The BUGS'n BEADS™ principle was used to develop an automated customised application. Briefly, the automatic isolation begins with addition of specially coated magnetic particles to Nalc-NaOH-treated clinical samples. Following bacterial capture, the sample matrix is removed. A rapid lysis at RT releases DNA, which is then adsorbed on to the same magnetic particles. Beads are washed and the purified DNA is robotically transferred for NAAT analysis. Samples were analysed both by using an in-house developed real-time PCR targeting the M. tuberculosis complex, and the Strand Displacement Amplification (SDA) part of the ProbeTec™ ET Mycobacterium tuberculosis Complex (DTB) Direct Detection (Becton Dickinson).

Results: Up to 48 samples can be simultaneously analysed using the automated BUGS'n BEADS™ system, with as little as 20 minutes of hands-on for each run, and requiring a total time to results of four hours. For smear positive samples, the results for the automated BUGS'n BEADS™ system in combination with real-time PCR were comparable to culture. The combination of SDA with the automated BUGS'n BEADS™ was comparable to the in-house Real-Time PCR. For the smear negative samples, we have observed so far that lowering the predefined BD cut-off from 3400 to 1000, enabled to improve sensitivity without worsening specificity. No inhibition was recorded for the SDA.

Conclusions: We here present a fully automated system for preparation of M. tuberculosis complex DNA from clinical respiratory samples, compatible with in-house developed and commercial downstream detection systems. Used with respiratory samples, the system will give clinically relevant results within one working day, requiring minimal hands-on.

The combination with SDA demonstrated successful removal of inhibition, while opening for further improvement of sensitivity. To further challenge the system, and evaluate lowering cut off, non-respiratory samples will be included in a more extensive study.

Session Details

Date: 31/03/2007
Time: 00:00-00:00
Session name: European Society of Clinical Microbiology and Infectious Diseases
Location: ICC, Munich, Germany
Presentation type:
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