A cutoff value of 50% similarity was applied to define MLVA clusters (named MLVA cluster 1 to MLVA cluster 9). The colors used are based on MLVA clusters. Figure 4 Minimum spanning tree (MST) representation of the MLVA clustering. The colors used in figure 4A are based on MLVA clusters. The colors used in figure 4B are based on MLST GSK872 clonal complexes. White circles correspond to genotypes not clustered by MLVA or MLST. The MLVA data for 189 strains, including 3 reference

strains, were analyzed in BioNumerics. Each circle represents an MLVA genotype and its size is proportional to the number of strains. A logarithmic scale was used when drawing branches. The thicker branches link the MLVA genotypes differing by only one allele, the thinner branches link MLVA GSK126 order genotypes

differing by more than one allele. Comparison of MLVA and MLST clustering MLVA clustering showed a clonal distribution CB-839 purchase of the population similar to that obtained by MLST (Figure 4). All human strains of MLST CC17 clustered together in MLVA cluster 9 and the bovine strains of MLST CC17 belonged to several MLVA clusters, suggesting greater heterogeneity of this population (Figure 4). With the exception of 3 strains, the MLST CC19 strains clustered into 2 linked MLVA clusters, MLVA cluster 6 and MLVA cluster 7. The MLST CC23 strains of serotype III and the MLST CC10 strains clustered into MLVA cluster 2. The strains from Tolmetin MLST CC23 serotype Ia also formed a separate group, the MLVA cluster 8. Discrimination of S. agalactiae strains by MLVA The diversity index obtained with MLVA was 0.960 (95% CI [0.943 - 0.978]), which is greater than that obtained with MLST

(0.881). For the population studied, MLVA distinguished 98 genotypes, whereas MLST distinguished 51 different STs. A much higher level of diversity was observed with MLVA, particularly within the major CCs. For example, the 73 CC17 strains were separated into 12 STs by MLST and 22 MLVA genotypes; the 63 CC19 strains were separated into 15 STs by MLST and 35 MLVA genotypes and the 15 CC23 strains were separated into 6 STs by MLST and 15 MLVA genotypes. Nevertheless, two genotypes (46 and 47) accounted for 76% (45/59) of CC17 strains of human origin. For this particular genogroup, the discriminatory power of the MLVA method was greater than that of MLST, although it remained low. Discussion In this study, we applied the multi locus VNTR analysis (MLVA) typing method to S. agalactiae. VNTR analysis, a method based on tandem repeat polymorphisms at multiple loci, has been successfully applied to many other bacterial species [30, 41]. We investigated the relevance of this tool for the genotyping of S. agalactiae, by testing this method on six VNTR loci in 189 strains previously characterized by MLST and serotyping. The MLVA-6 scheme is inexpensive and can be carried out with the equipment routinely used for PCR amplification and agarose gel electrophoresis.