Actually, a diagnostic PCR using this target was later designed, validated according to international guidelines and confirmed to provide an epidemiologically relevant phylogeny . New Caledonia is an archipelago of the South-West Pacific (19-23°S; 164-167°E). Leptospirosis is known to FG-4592 chemical structure be endemic with epidemic bursts occurring during hot rainy periods [3, 10–12]. Presumptive serovars in New Caledonia based on MAT on human leptospirosis cases are Copenhageni, Icterohaemorragiae, Castellonis, Panama, Pomona, Australis and Pyrogenes
[10, 11, 13, 14]. The only native mammals are bats and flying foxes. Very few imported mammals are present: 4 rodent species (Rattus rattus, Rattus norvegicus, Rattus exulans and Vorinostat concentration Mus musculus) and domestic as well as feral dogs, cats, cattle, horses, goats, sheeps and the Rusa deer Cervus timorensis russa. The qPCR
technique used for leptospirosis diagnosis in New Caledonia amplifies a 331pb DNA fragment within the lfb1 gene, which sequence polymorphism allows the identification of the species of the infecting Leptospira strain using melting curve analysis . The Multi Locus Sequence Typing (MLST) technique uses sequence polymorphisms of multiple housekeeping genes for isolate Small molecule library concentration characterization and to investigate evolutionary relationships among closely-related bacteria. It is increasingly considered as the gold standard typing method, at least in species where sufficient sequence polymorphisms exists in housekeeping genes, because it relies on sequence data that are exchangeable and independent of the analytical platform [16, 17]. This technique, successfully applied to a number of bacterial pathogens,
was notably recently applied to the study of leptospires: various typing schemes based on the comparison of 2855-3165 bp concatenated sequences of housekeeping genes were proposed [18–20] and evaluated over Leptospira spp. reference strains and isolates. Because of the limited mammal diversity in New Caledonia, we hypothesized that a limited diversity of pathogenic Leptospira strains Janus kinase (JAK) would be present and aimed at evaluating if the sequence polymorphism of diagnostic PCR products would allow the identification of the infecting Leptospira. To better investigate this hypothesis and the epidemiology of leptospirosis in New Caledonia, we also performed a MLST study on a collection of isolates and evaluated its direct feasibility using leptospirosis patients’ serum DNA extracts. Additionally, extracts from Leptospira-infected deer kidneys contributed to a better description of the Leptospira strains currently involved in leptospirosis in New Caledonia. Methods Bacterial strains The strains studied were collected from 1989 to 2000 throughout mainland New-Caledonia. Eighteen were isolates from patients’ blood received at Institut Pasteur for diagnosis purpose, and 2 were isolated from deer in 1992, kindly provided by the New Caledonian Reference Veterinary Laboratory.