Detection and identification of Mycobacterium tuberculosis and Mycobacterium bovis from clinical species using DNA microarrays.

Abstract

The objectives of the current study were to evaluate the use of DNA microarray for the rapid and direct detection of Mycobacterium tuberculosis and Mycobacterium bovis in bovine milk, blood, and pharyngeal swab samples, and to compare the use of DNA microarrays with current molecular detection techniques. The present study describes a microarray assay based on mtp40 and pncA gene sequences, which can be used to detect M. tuberculosis and M. bovis species. Each probe was spotted onto a silylated glass slide with an arrayer and used for hybridization with fluorescently labeled DNA derived from amplified DNA samples. The detection limit for mycobacterial DNA using this DNA microarray method was 50 fg (5 tubercle bacilli). Mycobacterium tuberculosis and/or M. bovis was detected in 7.1% (24/336) of the cattle specimens using the DNA microarray compared to 6.0% (20/336) using culture methods. Mixed infections were detected in 3 animals using the DNA microarray method, whereas the mixed infections were detected in 2 animals using either culture or polymerase chain reaction methods. The use of ancillary in vitro tests alongside the DNA microarray enhanced the detection of cattle infected with M. tuberculosis and/or M. bovis and reduced the number of false-positive animals that would be culled. More species may be easily added to this system, and supplementary probes can be added to increase the simultaneous detection power.