Rapid variable-number tandem-repeat genotyping for Mycobacterium leprae clinical specimens

Miyako Kimura, Rama Murthy Sakamuri, Nathan A. Groathouse, Becky L. Rivoire, David Gingrich, Susan Krueger-Koplin, Sangnae Cho, Patrick J. Brennan, Varalakshmi Vissa

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

Mycobacterium leprae is the noncultivable pathogen of leprosy. Since the genome sequence of an isolate of M. leprae has become available, multiple-locus variable-number tandem-repeat (VNTR) analysis (MLVA) has been explored as a tool for strain typing and identification of chains of transmission of leprosy. In order to discover VNTRs and develop methods transferable to clinical samples, MLVA was applied to a global collection of M. leprae isolates derived from leprosy patients and propagated in armadillo hosts. PCR amplification, agarose gel electrophoresis, and sequencing methods were applied to DNA extracts from these infected armadillo tissues (n = 21). We identified polymorphisms in 15 out of 25 short-tandem-repeat (STR) loci previously selected by in silico analyses of the M. leprae genome. We then developed multiplex PCR for amplification of these 15 loci in four separate PCRs suitable for fluorescent fragment length analysis and demonstrated STR profiles highly concordant with those from the sequencing methods. Subsequently, we extended this method to DNA extracts from human clinical specimens, such as skin biopsy specimens (n = 30). With these techniques, mapping of multiple loci and differentiation of genotypes have been possible using total DNA extracts from limited amounts of clinical samples at a reduced cost and with less time. These practical methods are therefore available and applicable to answer focused epidemiological questions and to allow monitoring of the transmission of M. leprae in different countries where leprosy is endemic.

Original languageEnglish
Pages (from-to)1757-1766
Number of pages10
JournalJournal of Clinical Microbiology
Volume47
Issue number6
DOIs
Publication statusPublished - 2009 Jun 1

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Mycobacterium leprae
Minisatellite Repeats
Leprosy
Armadillos
Microsatellite Repeats
DNA
Genome
Polymerase Chain Reaction
Agar Gel Electrophoresis
Multiplex Polymerase Chain Reaction
Computer Simulation
Genotype
Biopsy
Costs and Cost Analysis
Skin

All Science Journal Classification (ASJC) codes

  • Microbiology (medical)

Cite this

Kimura, M., Sakamuri, R. M., Groathouse, N. A., Rivoire, B. L., Gingrich, D., Krueger-Koplin, S., ... Vissa, V. (2009). Rapid variable-number tandem-repeat genotyping for Mycobacterium leprae clinical specimens. Journal of Clinical Microbiology, 47(6), 1757-1766. https://doi.org/10.1128/JCM.02019-08
Kimura, Miyako ; Sakamuri, Rama Murthy ; Groathouse, Nathan A. ; Rivoire, Becky L. ; Gingrich, David ; Krueger-Koplin, Susan ; Cho, Sangnae ; Brennan, Patrick J. ; Vissa, Varalakshmi. / Rapid variable-number tandem-repeat genotyping for Mycobacterium leprae clinical specimens. In: Journal of Clinical Microbiology. 2009 ; Vol. 47, No. 6. pp. 1757-1766.
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Kimura, M, Sakamuri, RM, Groathouse, NA, Rivoire, BL, Gingrich, D, Krueger-Koplin, S, Cho, S, Brennan, PJ & Vissa, V 2009, 'Rapid variable-number tandem-repeat genotyping for Mycobacterium leprae clinical specimens', Journal of Clinical Microbiology, vol. 47, no. 6, pp. 1757-1766. https://doi.org/10.1128/JCM.02019-08

Rapid variable-number tandem-repeat genotyping for Mycobacterium leprae clinical specimens. / Kimura, Miyako; Sakamuri, Rama Murthy; Groathouse, Nathan A.; Rivoire, Becky L.; Gingrich, David; Krueger-Koplin, Susan; Cho, Sangnae; Brennan, Patrick J.; Vissa, Varalakshmi.

In: Journal of Clinical Microbiology, Vol. 47, No. 6, 01.06.2009, p. 1757-1766.

Research output: Contribution to journalArticle

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Kimura M, Sakamuri RM, Groathouse NA, Rivoire BL, Gingrich D, Krueger-Koplin S et al. Rapid variable-number tandem-repeat genotyping for Mycobacterium leprae clinical specimens. Journal of Clinical Microbiology. 2009 Jun 1;47(6):1757-1766. https://doi.org/10.1128/JCM.02019-08