Our objectives were to refine the phylogeographic assessment within South America puma alife and to investigate the demographic history of pumas using a coalescent approach. Our results extend previous phylogeographic findings, reassessing the delimitation of historical population units in South America and demonstrating that this species experienced a considerable demographic expansion in the Holocene, ca. 8,000 years ago. Our analyses indicate that this expansion occurred in South America, prior to the hypothesized re-colonization of North America, which was therefore inferred to be even more recent.

In addition, the genetic diversity within South America was found to be larger than in Central and North America, suggesting that pumas from the latter subcontinent actually derive from a recent re-colonization event, following extinction in North America in the Late Pleistocene (ca. 10,000 12,000 puma athletic shoes years ago).Within the mtDNA, the NADH dehydrogenase subunit 5 ( ND5 ) gene has been successfully used in phylogenetic and phylogeographic studies of felids and other carnivores ( e.g. Culver et al. , 2000 ; Trinca et al. , 2012 ). In a puma basket black previous study focusing on pumas ( Culver et al. , 2000 ), three mtDNA segments were employed ( ND5 , 16S and ATP8 ). Of these, ND5 showed the highest polymorphic content in this species, based on a segment spanning 318 bp.

Given that the geographic sampling of South American pumas was limited in that first study, we aimed here to expand the representation of the various regions of this sub-continent, so as to allow refined inferences of population structure, maternal gene flow and demographic puma basket bow history. In addition to expanding the geographic coverage of South American regions to refine inferences on patterns of matrilineal subdivision, we have performed novel analyses on puma demographic history, which revealed consistent evidence of a recent population expansion in South America, prior to re-colonization of North America.

We applied a uniform prior using these ages as conservative minimum and maximum boundaries, respectively. The final run was based on 10 8 MCMC generations, with samples taken every 10 4 steps, and the first 10 4 steps removed as burn-in.The second set of Beast analyses aimed to infer the time of the most recent common ancestor ( t MRCA ) of different groups of samples, as well as to reconstruct the demographic history of pumas by estimating past fluctuations in population size via the Bayesian Skyline Plot (BSP) approach ( Drummond et al. , 2005 ). We assumed a Bayesian skyline tree prior and a strict molecular clock, puma basket heart black whose evolutionary rate was based on the estimate derived from the first round of analyses.

We assessed the t MRCA for four different sample sets: (i) complete sample; (ii) South America only; (iii) North and Central Americas (NA NCA); and (iv) a subset of 24 NA NCA samples that formed a regionally endemic subclade in the haplotype network, suggestive of autochthonous diversification (see Results). The BSP analyses were performed only for sets (i) and (ii), as their larger sample size allowed for more robust demographic inferences. The MCMC parameters were the same as in the first set of Beast analyses.