Abstract
Background Bacillus anthracis is known to have low genetic variability. In spite of this lack of diversity, multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA) and single nucleotide polymorphisms (SNPs) including the canonical SNPs assay (canSNP) have proved to be highly effective to differentiate strains. Five different MLVA schemes based on a collection of 31 VNTR loci (MLVA8, MLVA15, MLVA20, MLVA25 and MLVA31) with increased resolving power have been described.ResultsMLVA31 was applied to characterize the French National Reference Laboratory collection. The total collection of 130 strains is resolved in 35 genotypes. The 119 veterinary and environmental strains collection in France were resolved into 26 genotypes belonging to three canSNP lineages and four MLVA clonal complexes (CCs) with particular geographical clustering. A subset of seven loci (MLVA7) is proposed to constitute a first line assay. The loci are compatible with moderate resolution equipment such as agarose gel electrophoresis and show a good congruence value with MLVA31. The associated MLVA and SNP data was imported together with published genotyping data by taking advantage of major enhancements to the MLVAbank software and web site.ConclusionsThe present report provides a wide coverage of the genetic diversity of naturally occurring B. anthracis strains in France as can be revealed by MLVA. The data obtained suggests that once such coverage is achieved, it becomes possible to devise optimized first-line MLVA assays comprising a sufficiently low number of loci to be typed either in one multiplex PCR or on agarose gels. Such a selection of seven loci is proposed here, and future similar investigations in additional countries will indicate to which extend the same selection can be used worldwide as a common minimum set. It is hoped that this approach will contribute to an efficient and low-cost routine surveillance of important pathogens for biosecurity such as B. anthracis.
Highlights
Bacillus anthracis is a spore forming Gram positive bacterium that causes anthrax, a zoonosis with a worldwide distribution
Taking advantage of the knowledge and data accumulated in the past thirteen years, the main aims of the present study are (1) to provide an update of the genetic diversity of B. anthracis strains naturally present in France, using the most recently developed MLVA31 assay, (2) to propose a selection of variable-number tandem repeat (VNTR) loci which could represent a reasonable first-line assay for B. anthracis MLVA genotyping (3) to present the new B. anthracis genotyping database made by taking advantage of major developments to the underlying MLVAbank software
Because databases are a key issue in terms of genotyping, we have significantly improved the software behind the http://mlva.u-psud.fr prototype by introducing three major functionalities: firstly, the new version is able to merge a number of independently curated databases so that they can be queried and browsed as a single entity, secondly, the database can host any kind of numeric genotyping data, such as canonical SNPs assay (canSNP), and thirdly, a tool has been included to automatically deduce a MLVA or single nucleotide polymorphisms (SNPs) genotype from genome sequence data
Summary
Bacillus anthracis is a spore forming Gram positive bacterium that causes anthrax, a zoonosis with a worldwide distribution. Especially ruminants, are infected by ingestion of soilborne spores while grazing [1]. In France, animal cases are regularly reported [2]. Anthrax is a professional disease and humans are infected through exposure to animals or contaminated animal products when such material is incidentally ingested, inhaled or comes into contact with an open wound [3,4]. Bacillus anthracis is known to have low genetic variability. In spite of this lack of diversity, multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA) and single nucleotide polymorphisms (SNPs) including the canonical SNPs assay (canSNP) have proved to be highly effective to differentiate strains. Five different MLVA schemes based on a collection of 31 VNTR loci (MLVA8, MLVA15, MLVA20, MLVA25 and MLVA31) with increased resolving power have been described
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