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Abstract. Polymorphism at nine microsatellite loci was examined to assess the level of genetic differentiation between four Anopheles arabiensis populations from Senegal, the high plateau of Madagascar, and Reunion and Mauritius islands. Eight of nine loci showed great polymorphism (2-16 alleles/locus) and significant genetic differentiation was revealed between all four populations by F-and R-statistics, with Fst estimates ranging from 0.080 to 0.215 and equivalent Rst values ranging between 0.022 and 0.300. These high amounts of genetic differentiation are discussed in relation to geographic distance including large bodies of water, and history of mosquito settlement, and insecticide use on the islands. The results suggest that historical events of drift rather than mutation are probably the forces generating genetic divergence between these populations, with homogenization of the gene pool by migration being drastically restricted across the ocean.

Background Trypanosoma cruzi has been classified into six Discrete Typing Units (DTUs), designated as TcI–TcVI. In order to effectively use this standardized nomenclature, a reproducible genotyping strategy is imperative. Several typing schemes have been developed with variable levels of complexity, selectivity and analytical sensitivity. Most of them can be only applied to cultured stocks. In this context, we aimed to develop a multiplex Real-Time PCR method to identify the six T. cruzi DTUs using TaqMan probes (MTq-PCR).Methods/Principal FindingsThe MTq-PCR has been evaluated in 39 cultured stocks and 307 biological samples from vectors, reservoirs and patients from different geographical regions and transmission cycles in comparison with a multi-locus conventional PCR algorithm. The MTq-PCR was inclusive for laboratory stocks and natural isolates and sensitive for direct typing of different biological samples from vectors, reservoirs and patients with acute, congenital infection or Chagas reactivation. The first round SL-IR MTq-PCR detected 1 fg DNA/reaction tube of TcI, TcII and TcIII and 1 pg DNA/reaction tube of TcIV, TcV and TcVI reference strains. The MTq-PCR was able to characterize DTUs in 83% of triatomine and 96% of reservoir samples that had been typed by conventional PCR methods. Regarding clinical samples, 100% of those derived from acute infected patients, 62.5% from congenitally infected children and 50% from patients with clinical reactivation could be genotyped. Sensitivity for direct typing of blood samples from chronic Chagas disease patients (32.8% from asymptomatic and 22.2% from symptomatic patients) and mixed infections was lower than that of the conventional PCR algorithm.Conclusions/SignificanceTyping is resolved after a single or a second round of Real-Time PCR, depending on the DTU. This format reduces carryover contamination and is amenable to quantification, automation and kit production.