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Journal of Clinical Microbiology, February 2004, p. 578-584, Vol. 42, No. 2
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.2.578-584.2004
Copyright © 2004, American Society of Microbiology. All Rights Reserved.

Evaluation of Partial 16S Ribosomal DNA Sequencing for Identification of Nocardia Species by Using the MicroSeq 500 System with an Expanded Database

Joann L. Cloud,1* Patricia S. Conville,2 Ann Croft,3 Dag Harmsen,4 Frank G. Witebsky,2 and Karen C. Carroll1,5,{dagger}

ARUP Institute for Clinical and Experimental Pathology,1 ARUP Laboratories,3 Department of Pathology, University of Utah, Salt Lake City, Utah,5 Microbiology Service, Department of Laboratory Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Department of Health and Human Services, Bethesda, Maryland,2 Institute of Hygiene, University of Münster, Münster, Germany4

Received 5 September 2003/ Returned for modification 23 October 2003/ Accepted 31 October 2003

Identification of clinically significant nocardiae to the species level is important in patient diagnosis and treatment. A study was performed to evaluate Nocardia species identification obtained by partial 16S ribosomal DNA (rDNA) sequencing by the MicroSeq 500 system with an expanded database. The expanded portion of the database was developed from partial 5' 16S rDNA sequences derived from 28 reference strains (from the American Type Culture Collection and the Japanese Collection of Microorganisms). The expanded MicroSeq 500 system was compared to (i) conventional identification obtained from a combination of growth characteristics with biochemical and drug susceptibility tests; (ii) molecular techniques involving restriction enzyme analysis (REA) of portions of the 16S rRNA and 65-kDa heat shock protein genes; and (iii) when necessary, sequencing of a 999-bp fragment of the 16S rRNA gene. An unknown isolate was identified as a particular species if the sequence obtained by partial 16S rDNA sequencing by the expanded MicroSeq 500 system was 99.0% similar to that of the reference strain. Ninety-four nocardiae representing 10 separate species were isolated from patient specimens and examined by using the three different methods. Sequencing of partial 16S rDNA by the expanded MicroSeq 500 system resulted in only 72% agreement with conventional methods for species identification and 90% agreement with the alternative molecular methods. Molecular methods for identification of Nocardia species provide more accurate and rapid results than the conventional methods using biochemical and susceptibility testing. With an expanded database, the MicroSeq 500 system for partial 16S rDNA was able to correctly identify the human pathogens N. brasiliensis, N. cyriacigeorgica, N. farcinica, N. nova, N. otitidiscaviarum, and N. veterana.

* Corresponding Author. Mailing address: ARUP Institute for Clinical and Experimental Pathology, 500 Chipeta Way, Salt Lake City, UT 84108. Phone: (801) 583-2787, ext. 2439. Fax: (801) 584-5207. E-mail: cloudjl{at}aruplab.com.

{dagger} Present address: Division of Microbiology, The Johns Hopkins Hospital, Baltimore, MD 21287.

Journal of Clinical Microbiology, February 2004, p. 578-584, Vol. 42, No. 2
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.2.578-584.2004
Copyright © 2004, American Society of Microbiology. All Rights Reserved.



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