Our objectives were all white puma sneakers to refine the phylogeographic assessment within South America 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.

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 history. In addition to expanding the geographic coverage of best puma shoes 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.

These samples were originated from a diverse array black and gold puma shoes of geographic regions throughout most of the puma range, with greater emphasis on South America (see Table S1 ). Two samples of Puma yagouaroundi were also included to be used as outgroups in some analyses.To select a marker that would provide suitable information levels, we initially examined the mtDNA fragments used in previous studies, especially those involving Neotropical felids ( e.g. Eizirik et al. , 1998 , 2001 ; Johnson et al. , 1998 , 1999 ; Culver et al. , 2000 ).

The resulting novel puma sequences were deposited black puma sandals in GenBank (accession numbers {"type":"entrez-nucleotide-range","attrs":{"text":"KF460496-KF460523","start_term":"KF460496","end_term":"KF460523","start_term_id":"572098819","end_term_id":"572098873"}} KF460496-KF460523 ).To investigate the evolutionary relationships among puma mtDNA lineages, haplotype networks were built using the median-joining approach ( Bandelt et al. , 1999 ), as implemented in the software Network 4.5.1.6. We explored two outgroup options for rooting the network, one with the P. yagouaroundi sequences generated here, and the other employing M.

Finally, we conducted a set of analyses to investigate the demographic history of pumas. We performed neutrality tests (Tajima's D, Fu & Li' D* and F*, Fu's Fs) with DnaSP, as well as a mismatch distribution analysis ( Rogers and Harpending, 1992 ; Schneider and Excoffier, 1999 ) with Arlequin. In addition, we used the program Beast 1.6.1 ( Drummond and Rambaut, 2007 ) to perform estimates of coalescence times and past demography. We defined the best model of nucleotide substitution for our data set, which was the HKY ( Hasegawa et al. , 1985 ) G model, using the Akaike Information Criterion (AIC, Akaike, 1974 ) implemented in jModelTest 0.1 burgundy puma shoes ( Posada, 2008 ).

Our first set of Beast runs was aimed at estimating the molecular clock rate for our segment in puma lineages. We assumed a Yule Process for the tree prior and incorporated an uncorrelated lognormal relaxed molecular clock. In order to better fit the expectations of the Yule process we only included five divergent puma haplotypes, along with the two jaguarundi sequences generated here. To calibrate the molecular clock we assumed that pumas and jaguarundis diverged between 3.16 and 6.01 MYA (95% HPD of the estimate reported by Johnson et al. (2006) using a nuclear supermatrix).