Geographic isolation and climatic variability contribute to genetic differentiation in fragmented populations of the long-lived subalpine conifer Pinus cembra L. in the western Alps.

BACKGROUND: Genetic processes shape the modern-day distribution of genetic variation within and between populations and can provide important insights into the underlying mechanisms of evolution. The resulting genetic variation is often unequally partitioned within species' distribution range and especially large differences can manifest at the range limit, where population fragmentation and isolation play a crucial role in species survival. Despite several molecular studies investigating the genetic diversity and differentiation of European Alpine mountain forests, the climatic and demographic constrains which influence the genetic processes are often unknown. Here, we apply non-coding microsatellite markers to evaluate the sporadic peripheral and continuous populations of cembra pine (Pinus cembra L.), a long-lived conifer species that inhabits the subalpine treeline ecotone in the western Alps to investigate how the genetic processes contribute to the modern-day spatial distribution. Moreover, we corroborate our findings with paleoecological records, micro and macro-remains, to infer the species' possible glacial refugia and expansion scenarios. RESULTS: Four genetically distinct groups were identified, with Bayesian and FST based approaches, across the range of the species, situated in the northern, inner and south-western Alps. We found that genetic differentiation is substantially higher in marginal populations than at the center of the range, and marginal stands are characterized by geographic and genetic isolation due to spatial segregation and restricted gene flow. Moreover, multiple matrix regression approaches revealed effects of climatic heterogeneity in species' spatial genetic pattern. Also, population stability tests indicated that all populations had experienced a severe historical bottleneck, no heterozygosity excess was detected, suggesting that more recently population sizes have remained relatively stable. CONCLUSIONS: Our study demonstrated that cembra pine might have survived in multiple glacial refugia and subsequently recolonized the Alps by different routes. Modern-day marginal populations, at the edge of the species' range, could maintain stable sizes over long periods without inbreeding depression and preserve high amounts of genetic variation. Moreover, our analyses indicate that climatic variability has played a major role in shaping differentiation, in addition to past historical events such as migration and demographic changes.

Tree rings provide a new class of phenotypes for genetic associations that foster insights into adaptation of conifers to climate change.

Local adaptation in tree species has been documented through a long history of common garden experiments where functional traits (height, bud phenology) are used as proxies for fitness. However, the ability to identify genes or genomic regions related to adaptation to climate requires the evaluation of traits that precisely reflect how and when climate exerts selective constraints. We combine dendroecology with association genetics to establish a link between genotypes, phenotypes and interannual climatic fluctuations. We illustrate this approach by examining individual tree responses embedded in the annual rings of 233 Pinus strobus trees growing in a common garden experiment representing 38 populations from the majority of its range. We found that interannual variability in growth was affected by low temperatures during spring and autumn, and by summer heat and drought. Among-population variation in climatic sensitivity was significantly correlated with the mean annual temperature of the provenance, suggesting local adaptation. Genotype-phenotype associations using these new tree-ring phenotypes validated nine candidate genes identified in a previous genetic-environment association study. Combining dendroecology with association genetics allowed us to assess tree vulnerability to past climate at fine temporal scales and provides avenues for future genomic studies on functional adaptation in forest trees.