Conservatoire et Jardin botaniques de la Ville de Genève
Spéciation, diversification et structuration intraspécifiques
L'étude de la diversité génétique à l’intérieur des espèces ainsi que l’analyse des relations génétiques entre espèces proches est le fil rouge de ce programme. Les domaines d’applications sont divers et concernent l’étude de la structuration génétique intraspécifique (application à des programmes de conservation), la phylogéographie (inférence de l’histoire démographique et spatiale des espèces), ainsi que l’étude des patterns de différentiation et de spéciation (application à la stratégie des codes barres génétiques).
Analysis of the genetic structure and phylogeography of Eryngium alpinum
Eryngium alpinum is a long-lived hemicryptophyte perennial Apiaceae growing at the subalpine stage of the European Alps and the Balkans. This species is protected, not only in Switzerland, but in all Europe, because it is attractive (intensive picking, culture...) and because the number of sites where it can still be found continues to diminish. In Switzerland, Eryngium alpinum is mostly found in the Prealpine region (Fribourg and Vaud) but also in the cantons of Wallis, Grisons and Uri. The plant is cultivated for commercial purposes or for gardening and it is thought to have escaped in the wild many times. It has also been introduced on purpose by amateurs in several sites of the Prealpes or the Jura mountains.
The aim of the project is to study the genetic structure of Eryngium alpinum using chloroplast and nuclear microsatellite markers (Gaudeul et al., 2002). The results using microsatellites show that the species decline in Switzerland is not due to inbreeding but rather the a major change in land-use that occurred since the fifties (Naciri & Lambelet, 2011; Naciri et al., in prep).
The phylogeographic recontruction using the two chloroplast markers psbA-trnH et trnS-trnG revealed the existence of six lineages, that probably originated from two major glacial refugia during the Quaternary glaciations, one located in Southwestern Alps and the second one in the Balkans (Naciri & Gaudeul, 2007).
The results obtained with the two types of markers now allow the identification of the populations or regions that deserve specific protection.
Differentiation of Silene patula and colonization scenario in North Africa.
The genus Silene (Caryophyllaceae) is mostly known from the bladder campion which is commonly found in our regions. Silenes are however more frequent in the Mediterranean region, one of their centers of diversification and speciation. Within the genus, the perennial species of the group 'italica' have been the focus of a detailed monography from 1981 to 1985 (Jeanmonod et Mascherpa, 1982; Jeanmonod, 1984a; 1984b; 1985). This study, that used classical methods of biosystematics, suggested that the group expanded and diversified from a European species (S. Italica) that crossed the Mediterranean Sea when it dried during the Messianian crisis, five million years ago. The morphological comparative analyses suggested the following scenario for North Africa: a migration from East to West, starting from Tunisia to Algeria and Morocco along the two mostly parallel mountain massifs : the Saharan Atlas in the South and the costal chain in the North. Each of these two routes were suspected to have allowed for the differentiation of two subspecies : S. patula ssp. amurensis in the South and S. patula ssp. patula in the North. In Morocco, these two subspecies met again leading to phenomenon of introgression. From Morocco, the migration was suspected to have continued toward the Iberian Peninsula through the Gibraltar Channel, leading to the diversification of additional species (S. mellifera, S. longicila, S. coutinhoi et S. fernandezii). In this scenario, the Pyrenees would have constituted a efficient physical barrier, unlike the Gibraltar Channel. Using three chloroplast markers (trnH-psbA, trnS-trnG and rps12-rpl20), we tested the former hypotheses, and used several hundred herbarium specimens collected in the eighties by D. Jeanmonod in Algeria and Morrocco. The results showed that the differentiation between the two subspecies patula and amurensis is not supported by the genetic data (Naciri et al., 2010). Moreover we have shown that the region including the Kabylies in Algeria gathers a large proportion of the species genetic diversity. This region is therefore suspected to be a probable refugium or place of origin from which spatial expansions have subsequently occurred in two directions: towards West (Morocco) and East (the Aurès Mountains).
The species of the 'mollisima' group: common ancestor or convergent adaptation ?We are also interested in the 'mollissima' group, which comprises seven species (S. mollissima, S. andryalifolia, S. tomentosa, S. velutina, S. hifacensis, S. hicesiae and S. auriculifolia). All these species grow on cliffs, display similar morphological feartues and have restricted distributions in remote places in the Western mediterranean area, except S. andryalifolia which is found in North African (janmood, 1984c). We wonder, using the same chloroplast markers as before, whereas the seven species are issued form a single ancestral species which got fragmented during the Messinian Crisis or whether they result from parallel speciation events, and all derive from different species located not far geographically. The later hypothesis would explain the similarity among the species of the 'mollissima' group by a morphological convergent evolution (Naciri et al., 2008; 2010).
Taxonomic and phylogeographic study of the 'italica' group of Silenes in the Eastern Mediterranean Basin
Studying the Western species of the 'italica' group showed several features that led us extending the study to the Western Mediterranean Basin: S. italica which was thought to be basal in the group is in fact a recent species. On the contrary, S. nemoralis, the only biennial species of the group seems to be closer to the ancestral species. Since these two species are found in the whole mediterranean basin, the Greek and Turkish species have been added to our sampling (S. cythnia, S. goulimyi, S. sieberi, S. splendens, S. pseudonutans, S. gigantea, S. damboldtiana, S. spinescens, S. ispartensis and S. phrygia). The morphological and genetical analyses of the Western silenes of the 'italica' group is presently the subject of a PhD thesis conducted by Pierre-Emmanuel Du Pasquier.
Post glacial recolonisation patterns in South America: Analysis of the genetic structure and phylogeography of Astronium urundeuva et Geoffroea spinosa.
The last glaciations (-50'000 to -10'000) led to important changes in vegetation on almost all emerged lands. In Europe, the extension of the glaciers forced the majority of the species to move southward. It is however more difficult to describe precisely the history of vegetation successions in South America. This is almost due to the complexity of the climatic phenomenon that occurred during this period in the continent and to the scarcity of fossil remains in the Amazon Basin and elsewhere. According to recent data, the Amazon Basin was almost covered by a dry forest during the glacial periods. This forest has been afterwards restrained to the margins of the wet Amazonian forest during the last 10'000 years. We tested this hypothesis using two trees (Astronium urundeuva et Geoffroea spinosa) that are characteristic of the Seasonally Dry Tropical Forest (SDTF) using chloroplast and nuclear microsatellites (Caetano et al., 2005; Naciri-Graven et al., 2005) and extensive samplings in South America: Brazil, Paraguay, Argentina, Bolivia, Ecuador and the Galapagos, Colombia (Naciri et al., 2006; Caetano & Naciri, 2011). The results revealed a high spatial structure for Astronium urundeuva with signs of vicariance of two genetic lineages, progressive divergence due to isolation and a consecutive secondary contact South of the Caatinga in Brazil. All together, the study provides arguments that favor the existence of a previously more continuous formation of SDTF in eastern South America (Caetano et al., 2008a). This conclusion is all the more true for Geoffroea spinosa, for which little differentiation was found for the species all around the Amazon Basin (Caetano & Naciri 2011; Caetano et al., in prep). This project gave us the additional opportunity to study species boundaries in the Astronium genus (Caetano et al., 2008b) and to question the status of Geoffroea spinosa in the Galapagos using Bayesian simulations. The species has been shown to have colonized the Galapagos very recently, which might indicate that it has been introduced by Humans (Caetano et al., 2012).
DNA barcodes in vascular plants: a population genetics approach
The DNA Barcode ideally allows the identification of new specimens to known species using a single DNA fragment. In animals, the mitochondrial cox1 sequence has been identified as a potential candidate in many lineages. In land plants, the situation is somehow more complex due to the mitochondria being less variable in plants than in animals. Moreover, the use of a single gene rapidly appeared to be inefficient in unambiguously assigning plant specimens to species. This therefore led to a more mitigated proposal for plant barcode, where the use of several sequences has been suggested (Savolainen et al., 2005; Chase et al., 2005). The Barcode effort in plants has consequently focused on identifying optimal sequences to be used from the nuclear and the chloroplast genomes and recent publications suggest to use a minimum of two chloroplast sequences (Chase et al., 2007; Kress and Erickson, 2007 ; CBOL Plant Working Group 2009). The DNA Barcode has promoted a large debate about its usefulness and the way it should be applied. One pitfall of the method that has regularly been pointed out is the estimation of intra- versus interspecific variability, and related parameters such as the minimal sampling sizes to figure in the reference database.
In thisproject, we specifically address this question, using groups of closely related species from seven genera. Four chloroplast sequences were selected (trnH-psbA, matK, rpoB and rpoC1) that have been suggested as the more suitable barcode tools. We consequently plan to use these loci, together with a sampling scheme of at least 50 individuals per species in order to be able to capture the maximum intraspecific variability. The goal of this proposal is to supply the scientific community with sampling rules that might serve as indications to be used in the Barcode databasing effort for plant. Our hypothesis is that the sampling scheme to be applied to fully uncover the species variability might be different between plants having different life cycles, because of their respective evolutionary history traits, among which their different generation times and dispersal ability. This hypothesis also relies on the observation that species delimitations might be more blurred within woody and perennial plants than in annuals. Within trees and shrubs, three genera were selected:
- Acer (A. campestre, A. monspessulanum, A. opalus, A. platanoides and A. pseudoplatanus)
- Lonicera (L. alpigena, L. caerulea, L. nigra and L. xylosteum)
- Salix (S. herbaceae, S. randiculata, S. randusa and S. serpillifolia)
For perennials, two genera were chosen :
- Adenostyles (A. alliariae, A. glabra and A. leucophylla)
- Gentiana (G. acaulis, G. alpina, G. angustifolia and G. clusii)
For annuals, two genera were also selected:
- Geranium (G. columbinum, G. dissectum and G. pusillum),
- Veronica (V. agrestis, V. arvensis, V. hederifolia, V persica and V. polita).
Phylogeography of three relict tree species of Zelkova (Ulmaceae)
The genus Zelkova has been very successful during the Tertiary (from -65 to -7 miliion years). Only six species remained, three of which are located in Soutwest Eurasia. This project aimed at analysing the genetic diversity of Zelkova sicula (a critically endangered endemic of Sicily), Z. abelicea (endemic of Crete, endangered) and Z. carpinifolia (endemic of Caucasus), using chloroplast and nuclear markers. Because a high genetic diversity and structuration was found within species, the results were successfully used to infer the geographic origin of trees cultivated in botanical gardens for which records were scarce or absent. For Z. abiliceaand Z. carpinifolia, it was possible to infer the origin with a high precision (the massif scale). The results also show that not all the within species diversity is correctly sampled in the botanical gerdens collections. This means that a concerted action should be taken to reach a higher quality of the genetic ressources gathered by the ex-situ collections of Zelkova. This project is the result of a tight collaboration with the University of Fribourg (Dr. G. Kozlowski and C. Christe) with the financial support of BGCI (Botanic Gardens Conservation International).
Phylogeographic studies of the Paracaryum (Boraginaceae) species of Turkey
Boraginaceae are one of the richest family of Turkey and the genus Paracaryum is the fourth one in this country in terms of species richness. Following the taxonomic revision she conducted for her thesis on the genus Paracaryum at the Hacettepe University in Ankara, the Dr Aslo Koca came to our laboratory to test the hypotheses that came out from her study. The main aim is to study the relashionships of 22 Paracaryum species using molecular markers, to understand their respective evolutionary and diversification patterns in the Anatolian domain, which is located at the junction of three major diversity hotspots : the Mediterranean, the Irano-Anatolian and the caucasion hotspots.