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Journal of Clinical Microbiology, December 2004, p. 5582-5587, Vol. 42, No. 12
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.12.5582-5587.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Pulsed-Field Gel Electrophoresis Study of Mycobacterium abscessus Isolates Previously Affected by DNA Degradation

Yansheng Zhang,^1* Mitchell A. Yakrus,² Edward A. Graviss,³ Natalie Williams-Bouyer,³ Christine Turenne,⁴ Amin Kabani,⁴ and Richard J. Wallace Jr.¹

Department of Microbiology, University of Texas Health Center, Tyler, Texas,¹ National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia,² Department of Pathology, Baylor College of Medicine, Houston, Texas,³ National Reference Center for Mycobacteriology, National Microbiology Laboratory, Health Canada, Winnipeg, Manitoba, Canada⁴

Received 6 February 2004/ Returned for modification 17 March 2004/ Accepted 25 August 2004

DNA degradation (which results in a smear pattern) occurs with almost 50% of Mycobacterium abscessus strains during pulsed-field gel electrophoresis (PFGE). We assessed the potential benefit of using thiourea-containing buffer with M. abscessus by studying 69 isolates not previously typeable by PFGE (i.e., those with a smear pattern). Random (epidemiologically unrelated) isolates that were typeable (no smear pattern) were included as controls. Genomic DNA was digested with DraI, XbaI, and AseI. PFGE gels were run in regular gel buffer with and without 100 µM thiourea. All 69 isolates that generated smear patterns had clear band profiles when the thiourea buffer was used. These isolates were divided into only 30 patterns with DraI, 20 patterns with XbaI, and 20 patterns with AseI. The molecular profiles were all closely or possibly related, and the differences between the isolates ranged from zero to six bands. By multilocus enzyme electrophoresis (MEE), 45 of 53 smear isolates (85%) belonged to two closely related electrophoretic types. These isolates contained at least one enzyme allele seen almost exclusively in this group. Isolates without smear patterns were unaffected by thiourea and produced unrelated PFGE profiles, as well as multiple MEE types. The hsp65 and 16S rRNA gene sequences of the isolates with smear patterns were identical to those of M. abscessus type strain ATCC 19977, which had a nonsmear pattern, suggesting that this clone is a subgroup within M. abscessus. This demonstrates that the inability to type M. abscessus by PFGE is associated with a single clone of organisms.

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* Corresponding author. Mailing address: Department of Microbiology, University of Texas Health Center, 11937 U.S. Hwy. 271, Tyler, TX 75708. Phone: (903) 877-7683. Fax: (903) 877-7652. E-mail: yansheng.zhang{at}uthct.edu.

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Journal of Clinical Microbiology, December 2004, p. 5582-5587, Vol. 42, No. 12
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.12.5582-5587.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

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