Morphological Innovations and Vast Extensions of Mountain Habitats Triggered Rapid Diversification Within the Species-Rich Irano-Turanian Genus Acantholimon (Plumbaginaceae)

The Irano-Turanian floristic region spans a topographically complex and climatically continental territory, which has served as a source of xerophytic taxa for neighboring regions and is represented by a high percent of endemics. Yet, a comprehensive picture of the abiotic and biotic factors that have driven diversification within this biota remains to be established due to the scarcity of phylogenetic studies. Acantholimon is an important component of the subalpine steppe flora of the Irano-Turanian region, containing c. 200 cushion-forming sub-shrubby pungent-leaved species. Our recent molecular phylogenetic study has led to enlarging the circumscription of this genus to include eight mono- or oligospecific genera lacking the characteristic life-form and leaves. Using the same molecular phylogeny, here we investigate the tempo and mode of diversification as well as the biogeographic patterns in this genus, to test the hypothesis that a combination of key morphological innovations and abiotic factors is behind Acantholimon high species diversity. Molecular dating analysis indicates that Acantholimon s.l. started to diversify between the Late Miocene and the Pliocene and the biogeographic analysis points to an Eastern Iran–Afghanistan origin. Macroevolutionary models support the hypothesis that the high diversity of the genus is explained by accelerated diversification rates in two clades associated with the appearance of morphological key innovations such as a cushion life-form and pungent leaves; this would have favored the colonization of water-stressed, substrate-poor mountainous habitats along the newly uplifted IT mountains during the Mio-Pliocene. Given the apparent similarity of mountain habitats for most species of Acantholimon, we hypothesize that its current high species diversity responds to a scenario of non-adaptive radiation fueled by allopatric speciation rather than evolutionary radiation driven by ecological opportunity. Similar scenarios might underlie the high diversity of other speciose genera in the topographically complex Irano-Turanian landscape, though this remains to be tested with fine-grained distribution and climatic data

Cladistic analysis of the genus Ebenus (Fabaceae-Hedysareae) based on morphological data

In this study, cladistic analysis of characteristics morphological dataset using 21 characters for 26 taxa, including 19 taxa of Ebenus, 2 taxa of Taverniera, 2 taxa of Onobrychis and 2 taxa of Hedysarum as ingroups plus Alhagi persarum as an outgroup were chosen in phylogenetic analyses. Maximum parsimony approach as implemented in PAUP* with a heuristic search was employed. Fifty shortest trees with L=17.88 steps and a CI=0.709 and an RI=0.901 were generated. The analysis revealed that Ebenus was monophyletic. None of its multi-specific sections, Ebenidium and Euebenus, were monophyletic. E. cretica and E. stellata were basalmost branches that followed by a clade consisting of the remaining species. Onobrychis and Hedysarum were strongly allied with Ebenus

Extensive survey of the ycf4 plastid gene throughout the IRLC legumes: Robust evidence of its locus and lineage specific accelerated rate of evolution, pseudogenization and gene loss in the tribe Fabeae.

The genome organization and gene content of plastome (plastid genome) are highly conserved among most flowering plant species. Plastome variation (in size and gene order) is rare in photosynthetic species but size variation, rearrangements and gene/intron losses is attributed to groups of seed plants. Fabaceae (legume family), in particular the subfamily Papilionoideae and the inverted repeat lacking clade (IRLC), a largest legume lineage, display the most dramatic and structural change which providing an excellent model for understanding of mechanisms of genomic evolution. The IRLC comprises 52 genera and ca 4000 species divided into seven tribes. In present study, we have sampled several representatives from each tribe across the IRLC from various herbaria and field. The ycf4 gene, which plays a role in regulating and assembly of photosystem I, is more variable in the tribe Fabeae than in other tribes. In certain species of Lathyrus, Pisum and Vavilovia, all belonging to Fabeae, the gene is either absent or a pseudogene. Our study suggests that ycf4 gene has undergone positive selection. Furthermore, the rapid evolution of the gene is locus and lineage specific and is not a shared character of the IRLC in legumes

Characterization and comparative analysis of the complete plastid genomes of four Astragalus species.

Astragalus is the largest flowering plant genus. We assembled the plastid genomes of four Astragalus species (Astragalus iranicus, A. macropelmatus, A. mesoleios, A. odoratus) using next-generation sequencing and analyzed their plastomes including genome organization, codon usage, nucleotide diversity, prediction of RNA editing and etc. The total length of the newly sequenced Astragalus plastomes ranged from 121,050 bp to 123,622 bp, with 110 genes comprising 76 protein-coding genes, 30 transfer RNA (tRNA) genes and four ribosome RNA (rRNA) genes. Comparative analysis of the chloroplast genomes of Astragalus revealed several hypervariable regions comprising three non-coding sites (trnQ(UUG)-accD, rps7 -trnV(GAC) and trnR(ACG)-trnN(GUU)) and four protein-coding genes (ycf1, ycf2, accD and clpP), which have potential as molecular markers. Positive selection signatures were found in five genes in Astragalus species including rps11, rps15, accD, clpP and ycf1. The newly sequenced species, A. macropelmatus, has an approximately 13-kb inversion in IR region. Phylogenetic analysis based on 75 protein-coding gene sequences confirmed that Astragalus form a monophyletic clade within the tribe Galegeae and Oxytropis is sister group to the Coluteoid clade. The results of this study may helpful in elucidating the chloroplast genome structure, understanding the evolutionary dynamics at genus Astragalus and IRLC levels and investigating the phylogenetic relationships. Moreover, the newly plastid genomes sequenced have been increased the plastome data resources on Astragalus that can be useful in further phylogenomic studies

Molecular Phylogeny and Divergence Time Estimation of Onobrychis sect. Onobrychis (Fabaceae) based on nrDNA ITS

Phylogenetic analysis and estimating divergence time of O. sect. Onobrychis were conducted using nrDNA ITS sequence data. A total of 60 species from the section Onobrychis, seven species of the five sister sections belonging to O. subgen. Sisyrosema and eight species of other genera of the tribe Hedysareae (Eversmannia subspinosa ، Greuteria argyreum، Corethrodendron multijugum ، Taverniera cuneifolia، Hedysarum polybotrys، Sulla spinosissima و Alhagi maurorum) as outgroups were chosen. Phylogenetic analyses were performed by maximum likelihood and Bayesian methods. The results showed that O. sect. Onobrychis is monophyletic and composed of two main well-supported clades. The first clade includes annual and perennial species with long wings, often with the basic chromosome number x= 8. The second clade comprises perennial species with short wings and two annuals with long wings, having the basic chromosome number x= 7. Sect. The divergence time estimation of this section was analyzed with BEAST. The section was originated at 11.96 Mya in middle Miocene and subsequently diverged into two linages at 9.29-9.58 Mya.IntroductionThe genus Onobrychis Mill. comprises ca. 170 annual or perennial herbs divided into two subgenera and nine sections. At the current status, O. sect. Onobrychis with merging sects. Lophobrychis and Dendrobrychis is the largest section of the genus. Materials and MethodsIn this study, phylogenetic analysis and estimating divergence time of O. sect. Onobrychis were conducted using nrDNA ITS. A total of 60 species from the section Onobrychis, seven species of the five sister sections belonging to O. subgen. Sisyrosema and eight species of other genera of the tribe Hedysareae (Eversmannia subspinosa, Greuteria argyreum, Corethrodendron multijugum, Taverniera cuneifolia, Hedysarum polybotrys, Sulla spinosissima, and Alhagi maurorum) as out-groups were chosen. Phylogenetic analyses were performed by maximum likelihood and Bayesian methods. The divergence times estimation of this section was analyzed with BEAST. FindingsThe results showed that O. sect. Onobrychis is monophyletic and composed of two main well-supported clades including clade “A” and clade “B”. Each clad was divided into several subclades. Results and ConclusionsThe first clade includes annual and perennial species with long wings, often with the basic chromosome number x= 8, and the second one composes species with the basic chromosome number x= 7. It was also found that the annual species that were previously classified in the sect. Lophobrychis were not monophyletic, and three long-winged annual species were placed in clade “A”, and two annual long-winged species were related to short-winged perennial species, are placed in clade “B”. But the two long-winged species O. cornuta and O. elymaitica, were previously classified in the sect. Dendrobrychis, are placed in the clade “A”. Sect. Onobrychis originated at 11.96 Mya in the middle Miocene and was subsequently diverged into two lineages at 9.29-9.58 Mya

The GC content of three ITS regions among clades.

This is the first study to systematically evaluate rRNA secondary structures of Hedysareae with an emphasis on Hedysarum. ITS2 and 5.8S regions of the genus shared a common secondary structure with a four-fingered central loop, whereas ITS1 possessed five distinct structures. The secondary structural features of the two regions provided advantageous data for clades, species groups, and closely related species. Hemi-CBCs were mostly observed in the reconstruction of species groups, and Nsts, mostly between closely related species. The investigations showed that ITS1 varied more than ITS2 in length, GC content, and most of the diversity indices within the tribe. Maximum likelihood analyses of the synchronized sequence-structure tree of ITS1 were performed. The accuracy and phylogenetic signals of ITS1 were higher than ITS2. The similar GC content, and no CBC, in both spacers, fortified the close relationship of CEGO and H. sections Stracheya and Hedysarum clades in the synchronized sequence-structure tree topology of ITS1. In both regions, no inter-generic CBCs were detected inside the CEGO clade and the inter-sectional level of Hedysarum. But, in the ITS2 region, a CBC was detected between H. section Multicaulia, and Taverniera versus H. sections Hedysarum, and Stracheya. The lowest inter-sectional genetic distance and structural features were found between H. sect. Hedysarum and H. sect. Stracheya clades in the ITS2 region.</div