Abstract/Summary

Ophrys is amongst the best known orchid genera and is an established system for the study of pollinator mediated floral evolution. Two species, Ophrys fusca s.l. and Ophrys dyris (= O. omegaifera subsp. dyris), belonging to Ophrys section Pseudophrys are the focus of this study. An integrative study of morphological and genetic diversity of O. fusca and O. dyris was made, based on data collected from six populations in Central Portugal in five years. The populations were characterized a priori as “mixed” or pure”. The data included morpho-anatomical traits (272 plants) and phenological records and pollination success assessments (887 flowers from 260 plants). Additionally, sampling for DNA (165 plants) and leaf and pollinarium collection for cytological study (67 plants) was carried out. Multivariate analysis of the whole morphological dataset recovered two groups. When including only the plants common both to morphological and genetic datasets and a reduced number of diagnostic characters, three groups were found. Traits that better discriminate morphological groups are qualitative ones, namely the pilosity of the different lobes of the labellum. Univariate analysis of morpho-anatomical traits showed that vigour characters and depth of labellum indentation were the most variable, including between populations. Labellum indentation also showed greatest differences between the three types of populations (O. fusca/mixed/O. dyris). The study of flowering behaviour revealed incomplete floral isolation between the two taxa. Discrepancy between morphotypes in the average time for the flowering period in different years might threaten the synchrony with the supposed specialist pollinator, required for an effective pollination by sexual deception. This type of disruption being confirmed, it could represent an advantage for the fusca plants regarding the availability of pollinators. Percentages of pollination success obtained range from 2% to 20%. No significant differences in pollination success were found between plants confirmed as parental species or hybrids based on genetic data. Flow cytometry methods revealed constancy of nuclear DNA content (1C = 11.19 pg) in all the specimens analysed, including species and putative hybrids. Constancy of cytotypes was also confirmed, based on chromosome counts from the roots of two specimens, one of each species (2n = 4x = 72, 74). 72 for fusca, 74 for dyris. Following microsatellite data analysis, all the populations considered included individuals with mixed genotypes, representing a total of 44.8% of the plants. Genetic hybrids were found in all but one population, refuting the initial assumption of “pure” (one taxon) versus “mixed” populations. When comparing genetic and morphological datasets, despite a positive correlation, strong discrepancies have been found between the composition of the morphological groups and the genetic clusters. Implications are considered, in terms of predicting the persistence of putative hybrids and the conservation of evolutionary processes.