This MLST study similarly evidenced (Figure 3) three clusters of L. interrogans (corresponding to isolates grouped in L. interrogans clusters 1, 4 and 5). The clustering of isolates was in agreement with the lfb1-derived phylogeny. This result suggests that in the New Pifithrin-�� price Caledonian context, these lfb1-derived L. interrogans clusters are monophyletic

and probably each correspond to a single serovar. Again, L. interrogans cluster 5 did not contain any sequence of a known reference isolate, suggesting that it might correspond to a serovar not yet described, or at least not included in public sequence databases. Though the MLST phylogeny suggests that strains from this latter cluster could be related to the serovar Australis, seroconversions observed in New Caledonian patients infected with this strain merely point to Pyrogenes, a serogroup regarded as serologically related to Australis (data not shown). Whether ATPase inhibitor this cluster corresponds to a serovar not yet described or to a serovar described but which corresponding gene sequences have not been published GDC-0449 nmr remains to be studied. To further identify L. interrogans clusters 2 and 3 and to evaluate the feasibility of direct MLST from clinical specimen DNA extracts, we then tried to evaluate

the sequence polymorphism of the MLST targets using these clinical samples. Unfortunately, though both glmU and pntA could successfully be amplified and sequenced from extracts of patients containing ca. 200 leptospires per serum ml or more, none of the patients identified in these 2 clusters had leptospiraemia higher than 50 leptospires per ml. Interestingly, none of the isolate of our collection had lfb1 sequences identical to any of these two clusters. Because our isolate collection contains only strains collected until the year 2000, it cannot be known whether strains from these clusters were present in New Caledonia before 2001. They most probably already represented a limited part of the human cases

during this earlier period, as suggested by their low incidence over more than 2 years from 2008-february 2010 (see Table 4). It can also be hypothesized that strains from these clusters are of limited virulence to humans, therefore only associated with low leptospiraemia and would therefore seldom Liothyronine Sodium be evidenced, either by cultures (before 2001) or PCR (after 2001). Within L. borgpetersenii isolates, only two of the seven genes used in the MLST study of L. interrogans could be amplified. Actually, the set of primers used here was described by Thaipadungpanit et al [20] for use in L. interrogans isolates and was not supposed to amplify these genes in isolates from other species. Other MLST schemes have been used over a wider range of Leptospira species [18, 19]. These could have allowed a better typing of New Caledonian L. borgpetersenii isolates or clinical specimens. An ongoing program aimed at sequencing the complete genomes of a very large number of pathogenic Leptospira isolates (Vinetz J., com. pers.