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Genetic History of the Remnant Population of the Rare Orchid Cypripedium calceolus Based on Plastid and Nuclear rDNA.

ABSTRACT: The lady's slipper orchid (Cypripedium calceolus), which inhabits shady deciduous and mixed forests and meadows, is now threatened with extinction in many European countries, and its natural populations have been dramatically declining in recent years. Knowledge of its evolutionary history, genetic variability, and processes in small populations are therefore crucial for the species' protection. Nowadays, in south-west Poland, it is only distributed in seven small remnant and isolated populations, which we examined. One nuclear (ITS rDNA) and two plastid (accD-psa1, trnL-F) markers were analyzed and compared globally in this study. Based on the nuclear marker, the most common ancestor of C. calceolus and Cypripedium shanxiense existed about 2 million years ago (95% HPD: 5.33-0.44) in Asia. The division of the C. calceolus population into the European and Asian lineages indicated by C/T polymorphism started about 0.5 million years ago (95% HPD: 1.8-0.01). The observed variation of plastid DNA, which arose during the Pleistocene glacial-interglacial cycles, is still diffuse in Poland. Its distribution is explained by the result of fragmentation or habitat loss due to human impact on the environment.

SUBMITTER: Gorniak M

PROVIDER: S-EPMC8235785 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND AND AIMS:Farfugium (Asteraceae) is a small genus that contains the two species F. japonicum and F. hiberniflorum and is distributed along a long archipelago in east Asia. The common taxon, F. japonicum, includes three varieties associated with a wide range of habitats, including forest understorey (sciophytes), coastal crag (heliophytes) and riverbed (rheophytes). Leaf shape is an important taxonomic character within this genus and is associated with the habitat. METHODS:Twenty populations that included all Farfugium taxa were collected throughout its range. Leaf morphology was measured to determine differences amongst the taxa. Phylogenetic analyses based on sequences of the internal transcribed spacer of nuclear rDNA and four plastid DNA regions (matK, trnL-trnF, trnH-psbA and rpl20-rps12) were conducted separately. KEY RESULTS:Leaf morphology was significantly different amongst taxa, but morphological variations were partly explained by adaptation to certain environmental conditions that each population inhabited. Molecular phylogenies for the nDNA internal transcribed spacer and cpDNA were consistent in classifying F. hiberniflorum and the Taiwanese var. formosanum, whilst suggesting polyphyletic origins for the rheophyte, sciophyte and heliophyte taxa. All samples from the southern Ryukyus (Japan) and Taiwan clustered into a monophyletic group, which corroborates the land configuration theory involving Quaternary land-bridge formation and subsequent fragmentation into islands. The incongruence between the two DNA datasets may imply traces of introgressive hybridization and/or incomplete lineage sorting. CONCLUSIONS:The occurrence of rheophyte, sciophyte and heliophyte plants within Farfugium may be attributable to their isolation on islands and subsequent adaptation to the riparian, coastal crag and forest understorey environments, following their migration over the Quaternary land-bridge formation along their distribution range. Nearly identical DNA sequences coupled with highly divergent morphologies amongst these taxa suggest that diversification was rapid.

Project description:Universal angiosperm enrichment probe sets designed to enrich hundreds of putatively orthologous nuclear single-copy loci are increasingly being applied to infer phylogenetic relationships of different lineages of angiosperms at a range of evolutionary depths. Studies applying such probe sets have focused on testing the universality and performance of the target nuclear loci, but they have not taken advantage of off-target data from other genome compartments generated alongside the nuclear loci. Here we do so to infer phylogenetic relationships in the orchid genus Epidendrum and closely related genera of subtribe Laeliinae. Our aims are to: 1) test the technical viability of applying the plant anchored hybrid enrichment (AHE) method (Angiosperm v.1 probe kit) to our focal group, 2) mine plastid protein coding genes from off-target reads; and 3) evaluate the performance of the target nuclear and off-target plastid loci in resolving and supporting phylogenetic relationships along a range of taxonomical depths. Phylogenetic relationships were inferred from the nuclear data set through coalescent summary and site-based methods, whereas plastid loci were analyzed in a concatenated partitioned matrix under maximum likelihood. The usefulness of target and flanking non-target nuclear regions and plastid loci was assessed through the estimation of their phylogenetic informativeness. Our study successfully applied the plant AHE probe kit to Epidendrum, supporting the universality of this kit in angiosperms. Moreover, it demonstrated the feasibility of mining plastome loci from off-target reads generated with the Angiosperm v.1 probe kit to obtain additional, uniparentally inherited sequence data at no extra sequencing cost. Our analyses detected some strongly supported incongruences between nuclear and plastid data sets at shallow divergences, an indication of potential lineage sorting, hybridization, or introgression events in the group. Lastly, we found that the per site phylogenetic informativeness of the ycf1 plastid gene surpasses that of all other plastid genes and several nuclear loci, making it an excellent candidate for assessing phylogenetic relationships at medium to low taxonomic levels in orchids.

Project description:Background and aimsAlthough abscisic acid (ABA) is commonly recognized as a primary cause of seed dormancy, there is a lack of information on the role of ABA during orchid seed development. In order to address this issue, the localization and quantification of ABA were determined in developing seeds of Cypripedium formosanum.MethodsThe endogenous ABA profile of seeds was measured by enzyme-linked immunosorbent assay (ELISA). Temporal and spatial distributions of ABA in developing seeds were visualized by immunohistochemical staining with monoclonal ABA antibodies. Fluoridone was applied to test the causal relationship between ABA content and seed germinability.Key resultsABA content was low at the proembryo stage, then increased rapidly from 120 to 150 days after pollination (DAP), accompanied by a progressive decrease in water content and seed germination. Immunofluorescence signals indicated an increase in fluorescence over time from the proembryo stage to seed maturation. From immunogold labelling, gold particles could be seen within the cytoplasm of embryo-proper cells during the early stages of seed development. As seeds approached maturity, increased localization of gold particles was observed in the periplasmic space, the plasmalemma between embryo-proper cells, the surface wall of the embryo proper, and the inner walls of inner seed-coat cells. At maturity, gold particles were found mainly in the apoplast, such as the surface wall of the embryo proper, and the shrivelled inner and outer seed coats. Injection of fluoridone into capsules resulted in enhanced germination of mature seeds.ConclusionsThe results indicate that ABA is the key inhibitor of germination in C. formosanum. The distinct accumulation pattern of ABA suggests that it is synthesized in the cytosol of embryo cells during the early stages of seed development, and then exported to the apoplastic region of the cells for subsequent regulatory processes as seeds approach maturity.

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Genetic History of the Remnant Population of the Rare Orchid Cypripedium calceolus Based on Plastid and Nuclear rDNA.

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