Metapopulations in temporary streams - the role of drought-flood cycles in promoting high genetic diversity in a critically endangered freshwater fish and its consequences for the future.

Genetic factors have direct and indirect impacts in the viability of endangered species. Assessing their genetic diversity levels and population structure is thus fundamental for conservation and management. In this paper we use mitochondrial and nuclear markers to address phylogeographic and demographic data on the critically endangered Anaecypris hispanica, using a broad sampling set which covered its known distribution area in the Iberian Peninsula. Our results showed that the populations of A. hispanica are strongly differentiated (high and significant ФST and FST values, corroborated by the results from AMOVA and SAMOVA) and genetically diversified. We suggest that the restricted gene flow between populations may have been potentiated by ecological, hydrological and anthropogenic causes. Bayesian skyline plots revealed a signal for expansion for all populations (tMRCA between 68kya and 1.33Mya) and a genetic diversity latitudinal gradient was detected between the populations from the Upper (more diversified) and the Lower (less diversified) Guadiana river basin. We postulate a Pleistocenic westwards colonization route for A. hispanica in the Guadiana river basin, which is in agreement with the tempo and mode of paleoevolution of this drainage. The colonization of River Guadalquivir around 60kya with migrants from the Upper Guadiana, most likely by stream capture, is also suggested. This study highlights the view that critically endangered species facing range retreats (about 47% of its known populations have disappeared in the last 15years) are not necessarily small and genetically depleted. However, the extinction risk is not negligible since A. hispanica faces the combined effect of several deterministic and stochastic negative factors and, moreover, recolonization events after localized extinctions are very unlikely to occur due to the strong isolation of populations and to the patchily ecologically-conditioned distribution of fish. The inferred species distribution models highlight the significant contribution of temperature seasonality and isothermality to A. hispanica occurrence in Guadiana environments and emphasize the importance of stable climatic conditions for the preservation of this species. Given the strong population structure, high percentage of private haplotypes and virtual absence of inter-basin gene flow we suggest that each A. hispanica population should be considered as an independent Operational Conservation Unit and that ex-situ and in-situ actions should be conducted in parallel to allow for the long-term survival of the species and the preservation of the genetic integrity of its populations.