Characterization of Genetic Diversity of Bacillus anthracis in France by Using High-Resolution Melting Assays and Multilocus Variable-Number Tandem-Repeat Analysis

Abstract Background: Molecular genotyping of Bacillus anthracis, the etiologic agent of anthrax, is important for differentiating and identifying strains from different geographic areas and for tracing strains deliberately released in a bioterrorism attack. We previously described a multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA) based on 25 marker loci. Although the method has great differentiating power and reproducibility, faster genotyping at low cost may be requested to accurately identify B. anthracis strains in the field. Methods: We used the High Resolution Melter-1 (Idaho Technology) and a saturating dye of double-stranded DNA (LCGreen I) to identify alleles via PCR and melting-curve analysis of the amplicons. We applied high-resolution melting analysis (HRMA) to a collection of 19 B. anthracis strains. Results: HRMA produced reproducible results for 6 of the 25 B. anthracis loci tested. These easily interpretable and distinguishable melting curve results were consistent with MLVA results obtained for the same alleles. The feasibility of this method was demonstrated in testing of different allelic variants for the 6 selected loci. Conclusions: The described HRMA application for screening B. anthracis VNTR loci is fast and widely accessible and may prove particularly useful under field conditions.

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Multiple-locus variable-number tandem repeat analysis and clinical characterization of Leptospira interrogans canine isolates

Journal of Medical Microbiology ◽

10.1099/jmm.0.000027 ◽

2015 ◽

Vol 64 (3) ◽

pp. 288-294 ◽

Cited By ~ 8

Author(s):

Nobuo Koizumi ◽

Maki Mizutani Muto ◽

Hidemasa Izumiya ◽

Motoi Suzuki ◽

Makoto Ohnishi

Keyword(s):

Tandem Repeat ◽

Variable Number Tandem Repeat ◽

Variable Number ◽

Leptospira Interrogans ◽

Multiple Locus ◽

Repeat Analysis

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A study of the variation in genetic diversity of Haemophilus Influenzae serotype b (Hib) strains in the UK between 1987 and 2010 using multilocus variable-number tandem repeat analysis (MLVA)

Journal of Infection ◽

10.1016/j.jinf.2011.04.169 ◽

2011 ◽

Vol 63 (6) ◽

pp. e100

Author(s):

Sapna Manglani ◽

David Litt ◽

Shona Neal ◽

Mary Slack

Keyword(s):

Genetic Diversity ◽

Haemophilus Influenzae ◽

Tandem Repeat ◽

Variable Number Tandem Repeat ◽

Variable Number ◽

Repeat Analysis ◽

Serotype B ◽

The Uk

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Characterization of MRSA isolates from Kazakhstan using a multiple locus variable-number tandem repeat analysis

Journal of Microbiology Immunology and Infection ◽

10.1016/j.jmii.2015.02.385 ◽

2015 ◽

Vol 48 (2) ◽

pp. S109

Author(s):

D.B. Babenko ◽

A.V. Lavrinenko ◽

I.S. Azizov

Keyword(s):

Tandem Repeat ◽

Variable Number Tandem Repeat ◽

Variable Number ◽

Multiple Locus ◽

Repeat Analysis

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Multiple-Locus Variable-Number Tandem Repeat Analysis Reveals Genetic Relationships within Bacillus anthracis

Journal of Bacteriology ◽

10.1128/jb.182.10.2928-2936.2000 ◽

2000 ◽

Vol 182 (10) ◽

pp. 2928-2936 ◽

Cited By ~ 454

Author(s):

P. Keim ◽

L. B. Price ◽

A. M. Klevytska ◽

K. L. Smith ◽

J. M. Schupp ◽

...

Keyword(s):

Bacillus Anthracis ◽

Tandem Repeat ◽

Genetic Relationships ◽

Variable Number Tandem Repeat ◽

Variable Number ◽

Group Method ◽

Multiple Alleles ◽

Multiple Locus ◽

Sequence Characterization

ABSTRACT Bacillus anthracis is one of the most genetically homogeneous pathogens described, making strain discrimination particularly difficult. In this paper, we present a novel molecular typing system based on rapidly evolving variable-number tandem repeat (VNTR) loci. Multiple-locus VNTR analysis (MLVA) uses the combined power of multiple alleles at several marker loci. In our system, fluorescently labeled PCR primers are used to produce PCR amplification products from eight VNTR regions in the B. anthracisgenome. These are detected and their sizes are determined using an ABI377 automated DNA sequencer. Five of these eight loci were discovered by sequence characterization of molecular markers (vrrC 1, vrrC 2,vrrB 1, vrrB 2, and CG3), two were discovered by searching complete plasmid nucleotide sequences (pXO1-aat and pXO2-at), and one was known previously (vrrA). MLVA characterization of 426 B. anthracis isolates identified 89 distinct genotypes. VNTR markers frequently identified multiple alleles (from two to nine), with Nei's diversity values between 0.3 and 0.8. Unweighted pair-group method arithmetic average cluster analysis identified six genetically distinct groups that appear to be derived from clones. Some of these clones show worldwide distribution, while others are restricted to particular geographic regions. Human commerce doubtlessly has contributed to the dispersal of particular clones in ancient and modern times.

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