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Journal of Clinical Microbiology, August 2005, p. 3811-3817, Vol. 43, No. 8
0095-1137/05/$08.00+0     doi:10.1128/JCM.43.8.3811-3817.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Computational Approach Involving Use of the Internal Transcribed Spacer 1 Region for Identification of Mycobacterium Species

Amr M. Mohamed,^1, Dan J. Kuyper,² Peter C. Iwen,^1* Hesham H. Ali,² Dhundy R. Bastola,¹ and Steven H. Hinrichs¹

Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, Nebraska 68198-6495,¹ Department of Computer Science, University of Nebraska at Omaha, Omaha, Nebraska 68182-0116²

Received 9 November 2004/ Returned for modification 20 December 2004/ Accepted 17 April 2005

The rapid and reliable identification of clinically significant Mycobacterium species is a challenge for diagnostic laboratories. This study evaluates a unique sequence-dependent identification algorithm called MycoAlign for the differential identification of Mycobacterium species. The MycoAlign system uses pan-Mycobacterium-specific primer amplification in combination with a customized database and algorithm. The results of testing were compared with conventional phenotypic assays and GenBank sequence comparisons using the 16S rRNA target. Discrepant results were retested and evaluated using a third independent database. The custom database was generated using the hypervariable sequences of the internal transcribed spacer 1 (ITS-1) region of the rRNA gene complex from characterized Mycobacterium species. An automated sequence-validation process was used to control quality and specificity of evaluated sequence. A total of 181 Mycobacterium strains (22 reference strains and 159 phenotypically identified clinical isolates) and seven nonmycobacterial clinical isolates were evaluated in a comparative study to validate the accuracy of the MycoAlign algorithm. MycoAlign correctly identified all referenced strains and matched species in 94% of the phenotypically identified Mycobacterium clinical isolates. The ITS-1 sequence target showed a higher degree of specificity in terms of Mycobacterium identification than the 16S rRNA sequence by use of GenBank BLAST. This study showed the MycoAlign algorithm to be a reliable and rapid approach for the identification of Mycobacterium species and confirmed the superiority of the ITS-1 region sequence over the 16S rRNA gene sequence as a target for sequence-based species identification.

Present address: Department of Animal Medicine, Faculty of Veterinary Medicine, Assiut University, Assiut, Egypt.

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