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Molecular characterization of endangered Iranian terrestrial orchids using ISSR markers and association with floral and tuber-related phenotypic traits.

ABSTRACT: We aimed to study the genetic diversity, population structure, and phylogeny of Iranian orchids using inter-simple sequence repeat (ISSR) markers to find markers associated with phenotypic traits. Based on the phenotypic analysis, the inflorescence length and the flower number of studied accessions ranged from 3.92 to 27.13 cm and 5 to 50, respectively. On the other hand, the tuber length ranged from 1.80 to 9.35 cm. A total of 310 reproducible ISSR fragments with a size range of 150 to 3000 bp were amplified. ISSR primers provided an average polymorphism information content of 0.391, varied from 0.488 for UBC-876 to 0.351 for UBC-842. Os.J population showed the lowest genetic diversity (H = 0.057 and I = 0.075), while Oyst.JA population showed the highest genetic diversity (H = 0.114 and I = 0.158). At species level, the average coefficient of genetic differentiation (G _ST) ranged from 0.265 for Orchis simia to 0.587 for Himantoglossum affine. Gene flow (Nm) varied from 1.38 (O. simia) to 0.756 (Anacamptis collina). The UPGMA genetic similarity dendrogram using Jaccard coefficients (r = 0.973) revealed six main clusters. Based on the Bayesian clustering method, the highest probability of the data was achieved when accessions were divided into eight groups. Floral and tuber-related phenotypic traits represented high correlations together, and they were associated with some ISSR bands based on the multiple association analysis. Altogether, ISSR markers proved to be useful for discrimination and clarification of the relationships among species and populations collected from geographically different locations. Furthermore, it could identify the polymorphism among accessions within each population and species.

Supplementary information

The online version of this article (10.1007/s12298-020-00920-0) contains supplementary material, which is available to authorized users.

SUBMITTER: Gholami S

PROVIDER: S-EPMC7873147 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Background and aimsThe deception strategies of orchids remain poorly understood, especially in regard to the chemical compounds emitted from their flowers and their interaction with various taxonomic groups of pollinators. We investigated the phylogenetic relationships and compared the variation of floral chemical compounds between food-deceptive Dactylorhiza taxa (D. incarnata var. incarnata and D. incarnata var. ochroleuca, D. fuchsii and D. majalis) from populations in north-eastern Poland. We propose a model of the evolution of deception based on floral chemical signals in this genus.MethodsA Bayesian approach based on polymorphic plastid DNA (trnL, trnF and psbC-trnK), internal transcribed spacer (ITS) sequences and flow cytometry data was applied to confirm the taxonomic status of the studied orchids. We also identified and classified the pollinators and flower visitors in each Dactylorhiza population to the taxonomic level and compared our results with literature data. The chemical composition of pentane and diethyl ether extracts from the flowers was analysed by gas chromatography-mass spectrometry. Variation of the floral chemical components was visualized by non-metric multidimensional scaling based on Bray-Curtis dissimilarity.Key resultsThe genetic distinctiveness of D. incarnata, D. fuchsii and D. majalis was confirmed. No hybrids between them were found, but the chloroplast DNA (cpDNA), ITS haplotypes and flow cytometry showed genetic similarity between D. incarnata var. incarnata and D. incarnata var. ochroleuca. We determined that Apis mellifera (Hymenoptera) was the only shared pollinator of these taxa. Strangalia attenuata and Alosterna tabacicolor (Coleoptera) and Volucella pellucens and V. bombylans (Hymenoptera) were observed pollinating D. fuchsii. Visualization of the emission rates of the 61 floral chemical compounds detected from pentane extracts (mainly hydrocarbons and aldehydes) and the 51 from diethyl extracts (with abundant groups of benzenoids and non-aromatic acids) strongly differentiated D. incarnata, D. fuchsii and D. majalis, while those of the two varieties of D. incarnata (var. incarnata and var. ochroleuca) were almost identical.ConclusionsWhile the genetic data clearly supported the distinct lineages of D. incarnata, D. fuchsii and D. majalis, the patterns of emission of their flower chemical compounds were more complex within the series of shared compounds (alkanes and aldehydes) and taxon-specific compounds (benzenoids and esters). Their floral bouquet can influence the sexual, social and feeding behaviour of pollinators in different ways. We observed that the floral chemical compounds attracted both shared and species-specific pollinators to Dactylorhiza, confirming the multidirectional character of floral chemical signals in these food-deceptive taxa. Reduction of species-specific pollination levels in Dactylorhiza orchid taxa may promote hybridization between them.

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Molecular characterization of endangered Iranian terrestrial orchids using ISSR markers and association with floral and tuber-related phenotypic traits.

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