Plant evolution in alkaline magnesium-rich soils: A phylogenetic study of the Mediterranean genus Hormathophylla (Cruciferae: Alysseae) based on nuclear and plastid sequences

Abstract

Habitats with alkaline edaphic substrates are often associated with plant speciation and diversification. The tribe Alysseae, in the family Brassicaceae, epitomizes this evolutionary trend. In this lineage, some genera, like Hormathophylla, can serve as a good case for testing the evolutionary framework. This genus is centered in the western Mediterranean. It grows on different substrates, but mostly on alkaline soils. It has been suggested that diversification in many lineages of the tribe Alysseae and in the genus Hormathophylla is linked to a tolerance for high levels of Mg+2 in xeric environments. In this study, we investigated the controversial phylogenetic placement of Hormathophylla in the tribe, the generic limits and the evolutionary relationships between the species using ribosomal and plastid DNA sequences. We also examined the putative association between the evolution of different ploidy levels, trichome morphology and the type of substrates. Our analyses demonstrated the monophyly of the genus Hormathophylla including all previously described species. Nuclear sequences revealed two lineages that differ in basic chromosome numbers (x = 7 and x = 8 or derived 11, 15) and in their trichome morphology. Contrasting results with plastid genes indicates more complex relationships between these two lineages involving recent hybridization processes. We also found an association between chloroplast haplotypes and substrate, especially in populations growing on dolomites. Finally, our dated phylogeny demonstrates that the origin of the genus took place in the mid-Miocene, during the establishment of temporal land bridges between the Tethys and Paratethys seas, with a later diversification during the upper Pliocene.

Fig 1. Distribution area of 11 taxa in Hormathophylla.

Sampled localities are represented by dots and numbers as detailed in Table 1. A) Hormathophylla species with a broad distribution in the Iberian Peninsula (H. spinosa, H. cochleata subsp. cochleata, H. lapeyrouseana, H. saxigena, H. ligustica and H. pyrenaica). B) Additional Hormathophylla species which are present exclusively in the southwestern Iberian Peninsula (H. cochleata subsp. Baetica, H. reverchonii, H. cadevalliana, H. longicaulis, and H. purpurea).

Fig 2. Phylogenetic relationships inferred by ML and BI analyses of the 114 samples belonging to Hormathophylla, based on the ribosomal sequence ITS1-5.8S-ITS2, shown as a majority rule consensus tree.

Posterior probability and bootstrap support values are shown above and below the branches. Trichome types, as described in S5 Table, across different species are coded as T1, T2, T3, and T4. Basic chromosome number (x) is indicated by colored bars. Substrate type of the populations where sequences come from are coded as follows: D: dolomite; G: gypsum; L: limestone; S: serpentine; Si: siliceous.

Fig 3. Statistical parsimony network based on plastid sequences (trnL-trnF, trnT-trnL and rpl32-trnL), for species of Hormathophylla, and Alyssoides, Brachypus and Fibigia, as outgroups.

Lineages obtained in BI trees are represented. Haplotypes are grouped into three groups I, II and III. The substrate type of the populations where the haplotypes occur is coded as follows: D: dolomite; G: gypsum; L: limestone; S: serpentine; Si: siliceous.

Fig 4. Phylogenetic relationships inferred through ML and BI analyses of the 96 samples belonging to the genus Hormathophylla, and the species Fibigia clypeata, Brachypus suffruticosus, and Alyssoides utriculata, of the chloroplast regions trnL-trnF, trnT-trnL and rpl32-trnL, shown as a majority rule consensus tree.

Posterior probability and bootstrap support values are shown over and below the branches, respectively.

Fig 5. Tree generated in estimation of divergence times with BEAST using nuclear ribosomal ITS sequences.

Time scale is shown in millions of years. Error bars show 95% HPD in each node. Values above each node show median with respect to probability in millions of years.

Fig 6. SEM photographs illustrating variation of leaf trichome morphological types in Hormathophylla.

. A) Type I: H. cadevalliana. B) Type II: H. lapeyrouseana. C) Type III: H. purpurea. D) Type IV: H. ligustica. Scale bar = 100 μm.