Genotyping of Bacillus anthracis strains based on automated capillary 25-loci Multiple Locus Variable-Number Tandem Repeats Analysis View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2006-04-06

AUTHORS

Florigio Lista, Giovanni Faggioni, Samina Valjevac, Andrea Ciammaruconi, Josée Vaissaire, Claudine le Doujet, Olivier Gorgé, Riccardo De Santis, Alessandra Carattoli, Alessandra Ciervo, Antonio Fasanella, Francesco Orsini, Raffaele D'Amelio, Christine Pourcel, Antonio Cassone, Gilles Vergnaud

ABSTRACT

BACKGROUND: The genome of Bacillus anthracis, the etiological agent of anthrax, is highly monomorphic which makes differentiation between strains difficult. A Multiple Locus Variable-number tandem repeats (VNTR) Analysis (MLVA) assay based on 20 markers was previously described. It has considerable discrimination power, reproducibility, and low cost, especially since the markers proposed can be typed by agarose-gel electrophoresis. However in an emergency situation, faster genotyping and access to representative databases is necessary. RESULTS: Genotyping of B. anthracis reference strains and isolates from France and Italy was done using a 25 loci MLVA assay combining 21 previously described loci and 4 new ones. DNA was amplified in 4 multiplex PCR reactions and the length of the resulting 25 amplicons was estimated by automated capillary electrophoresis. The results were reproducible and the data were consistent with other gel based methods once differences in mobility patterns were taken into account. Some alleles previously unresolved by agarose gel electrophoresis could be resolved by capillary electrophoresis, thus further increasing the assay resolution. One particular locus, Bams30, is the result of a recombination between a 27 bp tandem repeat and a 9 bp tandem repeat. The analysis of the array illustrates the evolution process of tandem repeats. CONCLUSION: In a crisis situation of suspected bioterrorism, standardization, speed and accuracy, together with the availability of reference typing data are important issues, as illustrated by the 2001 anthrax letters event. In this report we describe an upgrade of the previously published MLVA method for genotyping of B. anthracis and apply the method to the typing of French and Italian B. anthracis strain collections. The increased number of markers studied compared to reports using only 8 loci greatly improves the discrimination power of the technique. An Italian strain belonging to the B branch was described, and two new branches, D and E, are proposed. Owing to the upgrading achieved here, precise genotyping can now be produced either by automated capillary electrophoresis, or by the more accessible but slower and for some markers slightly less accurate agarose gel methodology. More... »

PAGES

33-33

Identifiers

URI

http://scigraph.springernature.com/pub.10.1186/1471-2180-6-33

DOI

http://dx.doi.org/10.1186/1471-2180-6-33

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1045604599

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/16600037


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