Molecular evidence for allopolyploid speciation and a single origin of the western Mediterranean orchid Dactylorhiza insularis (Orchidaceae)

The hybrid origin of the western Mediterranean orchid Dactylorhiza insularis was demonstrated by genetic markers. Allozyme data showed that throughout its range I). insularis has an allotriploid constitution and reproduces apomictically. The parental species of D. insularis were identified as D. romana and D. sambucina; they contributed 2 alleles and 1 allele, respectively, at the allozyme loci studied. The maternal species of D. insularis was D. romana, as inferred from cpDNA (trnL(UAA) intron). High genetic similarities were found when comparing present populations of D. romana and D. sambucina with their respective genomes 'frozen' in D. insularis. Dactylorhiza insularis showed fixed (or nearly fixed) heterozygosity at 11 out of the 19 loci studied, and poor genetic variation: eight multilocus genotypes were detected at allozyme level. No multilocus genotype differs from the most similar one by more than one allele substitution. All D. insularis individuals showed the same cpDNA haplotype (I), regardless of their geographic origin and multilocus genotype. The I haplotype is similar, but not identical to that found in D. romana (R). No recurrent formation of D. insularis was observed in hybrid zones between D. romana and D. sambucina, where diploid sexual hybrids (F-1, F-n, backcrosses) were detected. Available data agree with a single origin for D. insularis, which possibly occurred in the present postglacial, when L). romana and D. sambucina, expanding from their glacial refugia, came into contact. The genetic homogeneity found between D. romana and D. markusii, both from their locus classicus, indicates that the latter is a junior synonym of D. romana; on the other hand, D. romana and D. sambucina are well differentiated species(D-Nei=0.59)