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Abstract Aim To test whether the genetic diversity of diadromous and landlocked populations of the small puyen Galaxias maculatus (known as jollytail in Australia and inanga in New Zealand) follow the same structuring patterns observed for migratory and non-migratory species of the genus Galaxias. This work also aimed to test whether the genetic structuring of a group of populations could be predicted from differences in the geomorphologic history of the region they inhabit. Location Eight landlocked populations were sampled from cold-temperate lakes in north-western Patagonia. The study area could be split latitudinally into two sectors that differed in their geomorphology, each of them hosting four populations. The southern sector shows evidence of a higher degree of glacial coverage, and the lakes are probably remnants of a big proglacial palaeolake. Lakes in the northern sector, on the other hand, suggest no common origin. Results Significant genetic structuring was found among the studied populations (Θ = 0.188), being the highest value reported to date for the species. Significant correlation was found between genetic diversity and lake area and perimeter. Diversity also showed a slight latitudinal variation suggesting the presence of genetically distinct groups of populations. The comparison of populations from the two geographical sectors showed that those from the north had a higher diversity, more private alleles and strong structuring, while those from the south were less diverse and much more homogeneous. Main conclusions Non-migratory populations of G. maculatus show much higher values of genetic structuring than those reported for diadromous populations. This follows the pattern seen when comparing migratory and non-migratory species of Galaxias. This agrees with population genetics theory which predicts that restricted gene flow would result in greater among-population divergence. Also, differences between northern and southern populations agreed with what was predicted by the geomorphologic history of the study area. During the Last Glacial Maximum ice cover in that region may have reduced the habitat of G. maculatus to a refuge with an impoverished gene pool. When the ice receded, leaving a great proglacial lake, that former population expanded and became fragmented after water levels descended. This resulted in present day lakes harbouring homogeneous populations with reduced diversity. The northern sector, in contrast, was less affected by glaciers, resulting in more geomorphologically stable lakes holding genetically diverse populations.