[20]. The phylogeny was established independently for L. interrogans strains and isolates (7 genes providing a concatenate sequence of 3155 bp) and for L. borgpetersenii (2 genes for a total concatenate sequence of 968 bp). Both phylogenies are presented in Figure 3a and 3b respectively. These results evidenced three clusters among the L. interrogans New Caledonian isolates and two clusters among L. borgpetersenii isolates. Based on sequences of reference isolates available in databases, these clusters could putatively be assigned

to a few serogroups. Among L. interrogans isolates, one cluster could correspond to serovars SYN-117 ic50 Pomona, Selleck Acalabrutinib Canicola, Pyrogenes or Hebdomadis,

ATM Kinase Inhibitor purchase another one to the serovar Icterohaemorragiae or Copenhageni. Lastly, one L. interrogans cluster did not match to any known reference strain. Among L. borgpetersenii isolates, one clustered with L. borgpetersenii Hardjo-bovis JB197, whereas four other isolates clustered together, but no publicly available sequence allowed putatively identifying this cluster. Figure 3 MLST-deduced phylogeny of New Caledonian isolates and reference strains. Blue legends indicate reference strains, red legends indicate the putative unknown serovar.. GenBank accession numbers are provided as additional file 1 Tables S1 and S2. A: L. interrogans phylogeny based on a concatenate 3155 bp sequence. B. L. borgpetersenii phylogeny based on a pntA+glmU concatenate

968 bp sequence. Direct MLST from clinical Galactosylceramidase specimens To further confirm the existence of the 5 L. interrogans clusters identified with lfb1 polymorphism on clinical samples, we tried to amplify and sequence glmU and pntA from these clinical samples, using the MLST primers and PCR conditions. Actually, these 2 genes are correctly amplified from isolates belonging to both L. interrogans and L. borgpetersenii species and their polymorphism allows discriminating the same clusters within New Caledonian L. interrogans isolates as the 7 genes do (data not shown). When using L. interrogans-infected clinical specimens, these two genes were successfully amplified from samples infected with ≥ ca. 200 leptospires per ml. Discussion While studying the sequence polymorphism of our diagnostic lfb1 qPCR product [15] in clinical specimens and a collection of isolates, we identified 2 L. borgpetersenii clusters and 5 L. interrogans clusters (Figure 1). Interestingly, one L. interrogans cluster (cluster 5) contained only sequences from human clinical specimens and did not include any known sequence of a reference strain, even after extensive searches in public databases.