Phylogeny, Speciation and Biogeography in South America

This proposal seeks support for continued studies on phylogeny, biogeography and speciation in Hypochaeris (Compositae) of Chile and Argentina. Morphological investigations reveal that, Hypochaeris contains 13 species native to Chile and 20-30 to Argentina. The genus is well suited for studies on speciation due to its herbaceous habit, low chromosome number (n = 4), and large chromosomes (2-8 micro-m) with a distinctive bimodal karyotype. Phylogenetic results from the first grant phase clearly show Hypochaeris in South America to be monophyletic, but more phylogenetic studies (using nuclear and chloroplast DNA sequences, plus AFLP data among very close relatives) of the remaining South American species are needed to determine the precise relationships among these recent and rapidly evolving taxa. Furthermore, and quite unexpected, based on our increased taxon sampling within the genus and outgroups, relationships among the European sections of Hypochaeris are now seen to interdigitate with the genus Leontodon, underscoring the need for reevaluation of generic limits among European Hypochaeris, Leontodon and Picris. I Proposed biogeographic studies will: (1) test isolation by distance and the role of dispersal in the complex populational system of H. apargioides; (2) assess levels of genetic variation in species with isolated and small populations (such as H. spathulata) in comparison with more widespread taxa. such as H. scorzonerae; (3) test the hypothesis of broader genetic variation in lowland older species in comparison with more recently evolved taxa with presumptively reduced genetic variation in open after Pleistocene glaciation; and (4) examine levels of genetic variation in the introduced and weedy species, H. radicata, in its home range in Europe and in comparison to populations from Japan, Chile, Argentina, and South Africa. We will continue to investigate modes of populational differentiation and speciation among taxa of Hypochaeris in several ways: (1) intensify our studies of the genetic bases species differences and the role of chromosomal differentiation during speciation by examining in more detail a case of . inferspecific hybridization (between H. tenuifolia and H. apargioides) in southern Chile in comparison with a known similar occurrence between H. glabra and H. radicata in Europe; (2) examine cytogenetic differentiation among all species in South America, emphasizing karyotypic analysis with fluorochrome staining and in-situ DNA hybridization (plus total DNA content as a correlate to observed karyotypic alterations), because initial studies have revealed valuable diagnostic features that are correlating with DNA sequence data in suggesting evolutionary groups; and (3) investigate the close relatives H. thrincinoides, H. apargioides and H. clarionoides (forming transitions from sea level to 3000 m) to determine direction of evolution and specific genetic, cytogenetic, and morphological changes during speciation.

This project on evolutionary and biogeographic relationships in the genus Hypochaeris of the sunflower family in South America has been very successful. While many in-depth studies on evolutionary patterns and process have been done in plants of North America and Europe, very few have been completed on plants of southern South America. We have used modern DNA techniques, involving sequences and population-level fingerprints, to resolve relationships among and within species with high levels of precision. Our ongoing research on Hypochaeris has revealed many interesting evolutionary aspects. First, we have determined that the species in South America have been derived from those in the Mediterranean region, and not the reverse, despite the large number of species in South America (c. 40 vs. 15 in Europe). Second, we have identified the closest relative of the South American complex, H. angustifolia, from Morocco. This suggests that the group dispersed to South America by long-distance dispersal (perhaps by birds). Application of a molecular clock allows estimation of age for this origin as between 1- 3 million years. Third, genetic divergence among the South American species is very low, in contrast to much higher levels among the Mediterranean species. DNA sequence divergence is extremely low, which suggests that the group rapidly radiated in parallel evolutionary lines throughout the temperate regions of the continent. By using a DNA fingerprint technique (AFLP: Amplified Fragment Length Polymorphism), however, we can see clearly different closely related subgroups, which now opens the possibility to examine specific mechanisms of speciation in consort with ecological and geographical information. Fourth, using populational-level AFLPs, we have examined genetic relationships within and among populations of H. acaulis and H. palustris, both being inbreeders. The former occurs on high volcanic peaks in Chile and adjacent Argentina. Data show a genetically divergent population structure due to geographic isolation and inbreeding. The latter occurs not only in the high Andes but also in the Coastal Range in Chile. We have demonstrated higher levels of genetic variation in the Coastal Range, and we hypothesize that this region may have been a refugium during Pleistocene glaciation in the Andes. Fifth, we have done detailed cytogenetic studies, which have allowed development of an evolutionary hypothesis for chromosomal change within the South American complex. Because all species have the same basic chromosome number (2n = 8), the differences are subtle, but consistent, and only by use of modern in-situ DNA techniques have we been able to formulate new and precise hypotheses involving translocations and inversions of chromosomal segments.