8006. Cmm strains from the recent epidemics in Belgium in 2010–2012 showed identical MLVA haplotypes which suggests that a clonal population was responsible for these outbreaks. The presence of the same MLVA haplotypes of Cmm strains from 2011 and 2012 could mean that bacteria persisted in the used equipment, devices or soil and induced the outbreaks in the following years. Population of Belgian strains isolated from 2010–2011 is epidemiologically related to at least two French strains that exhibited the same

MLVA haplotype. Moreover, based on minimum spanning tree, Belgian strains were found to be evolutionary related to the French strain PD 5749. When MLVA data was analyzed taking into account differences in the number of repeats it appeared that two French and two Spanish strains were found to have a similar MLVA find more haplotype to the group Alvocidib research buy of Belgian strains from 2010–2012 suggesting that there might be a common origin of these strains (Additional file 1: Figure S1). It is worth mentioning that the strain

ES 2686.1 isolated in Spain in 2002 was linked to outbreaks of Cmm in 2002–2007 in Canary Islands [6]. Two French strains isolated in 2010 showed the same MLVA haplotype as strains from recent Belgian outbreaks which may imply that the contaminated material was spread also in France. Different MLVA patterns between strains from the recent Belgian outbreaks of 2010–2012 and Belgian strains isolated previously support our hypothesis about a novel introduction, presumably originating from a single lot of seeds or contaminated tomato seedlings. Remarkably, all

Belgian Cmm strains from 2010–2012 MK 2206 (Table 1), were purchased from the same nursery. In this study, VNTR loci were chosen to be longer than or equal to 20 bp to simplify the interpretation of the results from an agarose gel and to allow performing the analysis in standard laboratories not equipped in sophisticated tools (fragment analyzer or sequencer) required to analyze small (a few nucleotides) differences in an amplicon size. Shorter repeats are represented in a higher number of copies and are more likely to be polymorphic [49]. However, many studies showed successful application of longer repeats which gave satisfactory resolution and discriminatory power [16, 50]. Interleukin-2 receptor Moreover, in silico analysis of tandem repeats in the Cmm genome NCPPB 382 revealed only a few short repeats (6–8 bp) that had remarkably higher number of copies (around 10 copies).These microsatellite loci might be investigated in the future and combined with currently available MLVA scheme. MLVA can provide phylogenetic information even with a limited number of loci [51]. MLVA assays are relatively robust [17, 52] but as any other technique they have their limitations. In MLVA, a need to develop a new set of loci for every species or serovar under investigation might be necessary.