Polyploidy in the olive complex (Olea europaea): evidence from flow cytometry and nuclear microsatellite analyses

Abstract

Background: Phylogenetic and phylogeographic investigations have been previously performed to study the evolution of the olive tree complex (Olea europaea). A particularly high genomic diversity has been found in north-west Africa. However, to date no exhaustive study has been addressed to infer putative polyploidization events and their evolutionary significance in the diversification of the olive tree and its relatives.

Methods: Representatives of the six olive subspecies were investigated using (a) flow cytometry to estimate genome content, and (b) six highly variable nuclear microsatellites to assess the presence of multiple alleles at co-dominant loci. In addition, nine individuals from a controlled cross between two individuals of O. europaea subsp. maroccana were characterized with microsatellites to check for chromosome inheritance.

Key results: Based on flow cytometry and genetic analyses, strong evidence for polyploidy was obtained in subspp. cerasiformis (tetraploid) and maroccana (hexaploid), whereas the other subspecies appeared to be diploids. Agreement between flow cytometry and genetic analyses gives an alternative approach to chromosome counting to determine ploidy level of trees. Lastly, abnormalities in chromosomes inheritance leading to aneuploid formation were revealed using microsatellite analyses in the offspring from the controlled cross in subsp. maroccana.

Conclusions: This study constitutes the first report for multiple polyploidy in olive tree relatives. Formation of tetraploids and hexaploids may have played a major role in the diversification of the olive complex in north-west Africa. The fact that polyploidy is found in narrow endemic subspecies from Madeira (subsp. cerasiformis) and the Agadir Mountains (subsp. maroccana) suggests that polyploidization has been favoured to overcome inbreeding depression. Lastly, based on previous phylogenetic analyses, we hypothesize that subsp. cerasiformis resulted from hybridization between ancestors of subspp. guanchica and europaea.

Fig. 1.

Flow cytometric analysis of the olive complex: comparative genome content (in pg per nucleus) between different taxa and populations of the olive complex. For each provenance, the mean genome size was estimated from the individuals analysed. Standard deviation based on two to five individuals (see Table 1) is given for each provenance, except for the sample of O. europaea subsp. laperrinei for which the only individual sampled was independently characterized three times.

Fig. 2.

Inheritance of alleles at locus ssrOe-DCA8 in two F₁ trees (m5 and m8) of the cross between individuals S1 × S2 of O. europaea subsp. maroccana. Electrophoresis of PCR products was carried out on a denaturing 5% acrylamide gel using an automated sequencer (ABI377; Applied Biosystems). For each individual, the size of each allele is indicated in the margin. The underlined sizes indicate alleles from the female (S1) and the sizes in bold-faced type indicate alleles from the male (S2). In individual m5, allele 161 is in italic because it was probably inherited from both parents. The F₁ trees m5 and m8 only received two of the six alleles revealed in parent S1 (m5: alleles 161 and 157; m8: alleles 135 and 161).