Microsatellites have been a workhorse of evolutionary genetic studies for decades and are still commonly in use for estimating signatures of genetic diversity at the population and species level across a multitude of taxa. Yet, the very high mutation rate of these loci is a double-edged sword, conferring great sensitivity at shallow levels of analysis (e.g., paternity analysis) but yielding considerable uncertainty for deeper evolutionary comparisons. For the present study, we used reduced representation genome-wide data (RADseq) to test for patterns of interspecific hybridization previously characterized using microsatellite data in a contact zone between two closely related mouse lemur species in Madagascar (Microcebus murinus and M. griseorufus). We revisit this system by examining populations in, near, and far from the contact zone, including many of the same individuals that had previously been identified as hybrids with microsatellite data. Surprisingly, we find no evidence for admixed nuclear ancestry. Instead, re-analyses of microsatellite data and simulations suggest that previously inferred hybrids were false positives and that the program NewHybrids can be particularly sensitive to erroneously inferring hybrid ancestry. Combined with results from coalescent-based analyses and evidence for local syntopic co-occurrence, we conclude that the two mouse lemur species are in fact completely reproductively isolated, thus providing a new understanding of the evolutionary rate whereby reproductive isolation can be achieved in a primate.
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