© 2005 Blackwell Publishing Ltd www.blackwellpublishing.com/ddi 257 Diversity and Distributions, (Diversity Distrib.) (2005) 11, 257–263 BIODIVERSITY RESEARCH ABSTRACT Several studies have reported glacial refugia and migration pathways for different pine species in the Iberian Peninsula, all of them based on a single-species approach. In this paper, chloroplast microsatellites (cpSSRs) are used to compare population genetic structure and diversity estimates for interspecific pairs of populations located along a cline from southwestern (latitude 36°32′ N, longitude 5°17′ W) to northeastern Spain (latitude 42°14′ N, longitude 2°47′ E) in two widely distributed Mediterranean pines, Pinus halepensis Mill. and Pinus pinaster Ait. Some cpSSRs were shared between species, facilitating comparison of levels of gene diversity at the species level and inferences about within and among species differentiation. P. pinaster showed a much higher number of variants (29) and haplotypes (69) than P. halepensis (20 and 21, respectively). Moreover, genetic diversity estimates for interspecific pairs of populations along the cline were negatively correlated. Three main causes may explain the differences between species in the present-day dis- tribution of genetic diversity: (1) the distribution of genetic variability before the Quaternary glaciations, with an earlier presence of P. pinaster in the Iberian Peninsula and a late spread of P. halepensis from eastern and central Europe, (2) the location of the Holocene glacial refugia and the migration pathways from these refugia to the present-day range (from northeast to southwest in P. halepensis and from southwest to northeast and northwest in P. pinaster) and (3) the interactions between species during the postglacial spread. Keywords Chloroplast markers, conifers, Mediterranean region, migration, palaeogeography. INTRODUCTION The analysis of the distribution of genetic diversity in a species provides useful information for conservation programs and management at the species level (Petit et al., 1998; Savolainen, 2000). Field experiments (common-garden tests) have been traditionally used to measure genetic diversity in plant species. However, these experiments are time consuming and relatively expensive, especially in forest trees, where life cycles are very long. As a consequence, the use of molecular markers in conser- vation genetics — in some cases analysed in combination with adaptive traits — significantly increased, in particular of those markers displaying high polymorphism such as microsatellites. The relatively high rate of sequence evolution at chloroplast microsatellites (cpSSRs; Provan et al., 1999), the small genome size of cpDNA and the absence of sexual recombination make cpSSRs ideal markers for population and ecological genetics studies (Morgante et al ., 1996). Pine species are widely distributed across the Mediterranean region, where they are the main components of the landscape and have a high economic value. Maritime pine (Pinus pinaster Ait.) occurs naturally from southwestern Europe (France, Portugal, Spain, and Italy) to northwestern Africa (Algeria, Tunisia and Morocco) showing several adaptations to local conditions (Alía et al., 1997). Aleppo pine (Pinus halepensis Mill.) is distributed all around the Mediterranean basin, mainly along the coast and exceptionally in inland Spain, Tunisia and Italy. Both species have a scattered distribution due to ecological disturbances (e.g. forest fires) and habitat fragmentation caused by long-term human impact in the Mediterranean basin. The situation of 1 Unidad de Genética Forestal, CIFOR-INIA, Carretera de La Coruña km 7.5, 28040 Madrid, Spain, 2 Istituto di Genetica Vegetale, Sezione di Firenze, CNR, Via Madonna del Piano, 50019 Sesto Fiorentino, Firenze, Italy *Correspondence: G. G. Vendramin, Institute of Plant Genetics, CNR, Via Madonna del Piano, 50019 Sesto Fiorentino (Firenze), Italy. Tel.: +39 055 5225725; Fax: +39 055 5225729; E-mail: giovanni.vendramin@igv.cnr.it Blackwell Publishing, Ltd. Genetic diversity and differentiation of two Mediterranean pines (Pinus halepensis Mill. and Pinus pinaster Ait.) along a latitudinal cline using chloroplast microsatellite markers A. Gómez 1 , G. G. Vendramin 2 *, S. C. González-Martínez 1 and R. Alía 1