Jean-Christophe Pintaud

A set of 100 chloroplast DNA primer pairs to study population genetics and phylogeny in monocotyledons.

Chloroplast DNA sequences are of great interest for population genetics and phylogenetic studies. However, only a small set of markers are commonly used. Most of them have been designed for amplification in a large range of Angiosperms and are located in the Large Single Copy (LSC). Here we developed a new set of 100 primer pairs optimized for amplification in Monocotyledons. Primer pairs amplify coding (exon) and non-coding regions (intron and intergenic spacer). They span the different chloroplast regions: 72 are located in the LSC, 13 in the Small Single Copy (SSC) and 15 in the Inverted Repeat region (IR). Amplification and sequencing were tested in 13 species of Monocotyledons: Dioscorea abyssinica, D. praehensilis, D. rotundata, D. dumetorum, D. bulbifera, Trichopus sempervirens (Dioscoreaceae), Phoenix canariensis, P. dactylifera, Astrocaryum scopatum, A. murumuru, Ceroxylon echinulatum (Arecaceae), Digitaria excilis and Pennisetum glaucum (Poaceae). The diversity found in Dioscorea, Digitaria and Pennisetum mainly corresponded to Single Nucleotide Polymorphism (SNP) while the diversity found in Arecaceae also comprises Variable Number Tandem Repeat (VNTR). We observed that the most variable loci (rps15-ycf1, rpl32-ccsA, ndhF-rpl32, ndhG-ndhI and ccsA) are located in the SSC. Through the analysis of the genetic structure of a wild-cultivated species complex in Dioscorea, we demonstrated that this new set of primers is of great interest for population genetics and we anticipate that it will also be useful for phylogeny and bar-coding studies.

Phylogenetic relationships among arecoid palms (Arecaceae: Arecoideae)

BACKGROUND AND AIMS The Arecoideae is the largest and most diverse of the five subfamilies of palms (Arecaceae/Palmae), containing >50 % of the species in the family. Despite its importance, phylogenetic relationships among Arecoideae are poorly understood. Here the most densely sampled phylogenetic analysis of Arecoideae available to date is presented. The results are used to test the current classification of the subfamily and to identify priority areas for future research. METHODS DNA sequence data for the low-copy nuclear genes PRK and RPB2 were collected from 190 palm species, covering 103 (96 %) genera of Arecoideae. The data were analysed using the parsimony ratchet, maximum likelihood, and both likelihood and parsimony bootstrapping. KEY RESULTS AND CONCLUSIONS Despite the recovery of paralogues and pseudogenes in a small number of taxa, PRK and RPB2 were both highly informative, producing well-resolved phylogenetic trees with many nodes well supported by bootstrap analyses. Simultaneous analyses of the combined data sets provided additional resolution and support. Two areas of incongruence between PRK and RPB2 were strongly supported by the bootstrap relating to the placement of tribes Chamaedoreeae, Iriarteeae and Reinhardtieae; the causes of this incongruence remain uncertain. The current classification within Arecoideae was strongly supported by the present data. Of the 14 tribes and 14 sub-tribes in the classification, only five sub-tribes from tribe Areceae (Basseliniinae, Linospadicinae, Oncospermatinae, Rhopalostylidinae and Verschaffeltiinae) failed to receive support. Three major higher level clades were strongly supported: (1) the RRC clade (Roystoneeae, Reinhardtieae and Cocoseae), (2) the POS clade (Podococceae, Oranieae and Sclerospermeae) and (3) the core arecoid clade (Areceae, Euterpeae, Geonomateae, Leopoldinieae, Manicarieae and Pelagodoxeae). However, new data sources are required to elucidate ambiguities that remain in phylogenetic relationships among and within the major groups of Arecoideae, as well as within the Areceae, the largest tribe in the palm family.

Male‐specific DNA markers provide genetic evidence of an XY chromosome system, a recombination arrest and allow the tracing of paternal lineages in date palm

Whether sex chromosomes are differentiated is an important aspect of our knowledge of dioecious plants, such as date palm (Phoenix dactylifera). In this crop plant, the female individuals produce dates, and are thus the more valuable sex. However, there is no way to identify the sex of date palm plants before reproductive age, and the sex-determining mechanism is still unclear. To identify sex-linked microsatellite markers, we surveyed a set of 52 male and 55 female genotypes representing the geographical diversity of the species. We found three genetically linked loci that are heterozygous only in males. Male-specific alleles allowed us to identify the gender in 100% of individuals. These results confirm the existence of an XY chromosomal system with a nonrecombining XY-like region in the date palm genome. The distribution of Y haplotypes in western and eastern haplogroups allowed us to trace two male ancestral paternal lineages that account for all known Y diversity in date palm. The very low diversity associated with Y haplotypes is consistent with clonal paternal transmission of a nonrecombining male-determining region. Our results establish the date palm as a biological model with one of the most ancient sex chromosomes in flowering plants.

Genetic isolation of Cape Verde Island Phoenix atlantica (Arecaceae) revealed by microsatellite markers.

Increasing human pressure on the environment in the isolated Macaronesian island group of Cape Verde is threatening many endemic species with extinction. The status of Phoenix atlantica, the Cape Verde Island date palm, is one of the unresolved taxonomic issues not only of the archipelago’s flora but also in the genus Phoenix. We applied 15 nuclear microsatellite markers and one chloroplast minisatellite marker to individuals of Phoenix from the Cape Verde Islands, P. dactylifera, P. canariensis and P. sylvestris, in order to assess the taxonomic position of P. atlantica within the genus. Our analysis showed that P. atlantica is clearly distinct from its close relatives and that its closest relative is likely to be its nearest geographical neighbour, P. dactylifera. Comparable levels of genetic diversity were found in insular P. atlantica and continental P. dactylifera despite the large difference in geographic range size. Our findings highlight the importance of conserving the relatively fragmented and isolated populations of P. atlantica as one of only␣two endemic trees on the islands and emphasise the need for further studies into its evolution and relationship with P. dactylifera.

Phylogenetic utility of the nuclear genes AGAMOUS 1 and PHYTOCHROME B in palms (Arecaceae): an example within Bactridinae

BACKGROUND AND AIMS Molecular phylogenetic studies of palms (Arecaceae) have not yet provided a fully resolved phylogeny of the family. There is a need to increase the current set of markers to resolve difficult groups such as the Neotropical subtribe Bactridinae (Arecoideae: Cocoseae). We propose the use of two single-copy nuclear genes as valuable tools for palm phylogenetics. METHODS New primers were developed for the amplification of the AGAMOUS 1 (AG1) and PHYTOCHROME B (PHYB) genes. For the AGAMOUS gene, the paralogue 1 of Elaeis guineensis (EgAG1) was targeted. The region amplified contained coding sequences between the MIKC K and C MADS-box domains. For the PHYB gene, exon 1 (partial sequence) was first amplified in palm species using published degenerate primers for Poaceae, and then specific palm primers were designed. The two gene portions were sequenced in 22 species of palms representing all genera of Bactridinae, with emphasis on Astrocaryum and Hexopetion, the status of the latter genus still being debated. KEY RESULTS The new primers designed allow consistent amplification and high-quality sequencing within the palm family. The two loci studied produced more variability than chloroplast loci and equally or less variability than PRK, RPBII and ITS nuclear markers. The phylogenetic structure obtained with AG1 and PHYB genes provides new insights into intergeneric relationships within the Bactridinae and the intrageneric structure of Astrocaryum. The Hexopetion clade was recovered as monophyletic with both markers and was weakly supported as sister to Astrocaryum sensu stricto in the combined analysis. The rare Astrocaryum minus formed a species complex with Astrocaryum gynacanthum. Moreover, both AG1 and PHYB contain a microsatellite that could have further uses in species delimitation and population genetics. CONCLUSIONS AG1 and PHYB provide additional phylogenetic information within the palm family, and should prove useful in combination with other genes to improve the resolution of palm phylogenies.

Close Genetic Proximity Between Cultivated and Wild Bactris gasipaes Kunth Revealed by Microsatellite Markers in Western Ecuador

Bactris gasipaes Kunth (peach palm or Pejibaye) is the only domesticated palm of the Neotropics. The genetic relationships between the crop and its wild relatives are still unclear. We undertook field and laboratory work in order to describe differentiation and relationships between the wild and cultivated populations of the species in Western Ecuador, and their possible interactions. A volumetric study was undertaken on the fruits of both populations, as well as a population genetic analysis in order to clarify these relationships. Fruits from cultivated plants collected in the region of sympatry of wild and cultivated plants in North-West Ecuador showed intermediate volumes between those of reference samples for the wild and cultivated plants in allopatry. Using 8 microsatellite loci, we assessed 83 wild and cultivated individuals from Western Ecuador and cultivated plants from Amazonia and Central America as a reference for the cultivated form. We detected high polymorphism in the wild and cultivated samples and low, but significant level of genetic differentiation between wild and cultivated populations. The cultivated population in North-Western Ecuador showed close genetic proximity with the sympatric wild population, consistent with the volumetric study. These results have implications for hypotheses on evolution of this crop and for strategies of genetic conservation of the wild forms.

Disturbance and Resilience in Tropical American Palm Populations and Communities

We review resilience to natural and anthropogenic disturbance of palm populations and communities in tropical America. Response of palms to disturbance depends on their morphological traits, their reproductive strategies and the impacts of these traits and strategies on phenology and gene flow. Human impact induces changes in genetic structure, increasing endogamy and genetic drift in fragmented populations. Forest fragmentation and harvest of palm organs are well documented whereas effects of intermediate disturbance like selective logging, hunting or fire remain poorly known. We recommend emphasis on long-term experiments and on the use of mechanistic approaches in future research to facilitate integration of available data into a theoretical ecological framework.

Phylogenetic Relationships of the Cultivated Neotropical Palm Bactris gasipaes (Arecaceae) with its Wild Relatives Inferred from Chloroplast and Nuclear DNA Polymorphisms

Abstract Peach palm (Bactris gasipaes Kunth.) is the only Neotropical palm domesticated since pre-Columbian times. It plays an important role not only at the local level due to its very nutritious fruits, but also in the international market for its gourmet palm heart. Phylogenetic relationships of the peach palm with wild Bactris taxa are still in doubt, and have never been addressed using molecular sequence data. We generated a chloroplast DNA phylogeny using intergenic spacers from a sampling of cultivars of Bactris gasipaes as well as putative wild relatives and other members of the genus Bactris. We estimated phylogenetic relationships using maximum parsimony (MP), maximum likelihood (ML), and Bayesian analysis. Our results indicated a close affinity between three taxa: Bactris gasipaes var. gasipaes, B. gasipaes var. chichagui, and B. riparia. There was no clear differentiation between these three taxa at the level of chloroplast sequences, and they shared a unique inversion that we characterized in this paper. Bactris setulosa, a species potentially related to the Bactris gasipaes complex, appeared highly divergent, and seemed to be a composite taxon with affinities outside the complex. We also investigated nuclear microsatellite polymorphisms at 8 loci within Bactris gasipaes, B. riparia, and B. setulosa, finding a pattern of relationships in agreement with the cpDNA data. The results presented here are important for future studies on domestication and crop improvement of Bactris gasipaes.

Genetic structure of the date palm (Phoenix dactylifera) in the Old World reveals a strong differentiation between eastern and western populations

BACKGROUND AND AIMS Date palms (Phoenix dactylifera, Arecaceae) are of great economic and ecological value to the oasis agriculture of arid and semi-arid areas. However, despite the availability of a large date palm germplasm spreading from the Atlantic shores to Southern Asia, improvement of the species is being hampered by a lack of information on global genetic diversity and population structure. In order to contribute to the varietal improvement of date palms and to provide new insights on the influence of geographic origins and human activity on the genetic structure of the date palm, this study analysed the diversity of the species. METHODS Genetic diversity levels and population genetic structure were investigated through the genotyping of a collection of 295 date palm accessions ranging from Mauritania to Pakistan using a set of 18 simple sequence repeat (SSR) markers and a plastid minisatellite. KEY RESULTS Using a Bayesian clustering approach, the date palm genotypes can be structured into two different gene pools: the first, termed the Eastern pool, consists of accessions from Asia and Djibouti, whilst the second, termed the Western pool, consists of accessions from Africa. These results confirm the existence of two ancient gene pools that have contributed to the current date palm diversity. The presence of admixed genotypes is also noted, which points at gene flows between eastern and western origins, mostly from east to west, following a human-mediated diffusion of the species. CONCLUSIONS This study assesses the distribution and level of genetic diversity of accessible date palm resources, provides new insights on the geographic origins and genetic history of the cultivated component of this species, and confirms the existence of at least two domestication origins. Furthermore, the strong genetic structure clearly established here is a prerequisite for any breeding programme exploiting the effective polymorphism related to each gene pool.

A revision of the palm genera ( Arecaceae ) of New Caledonia

SummaryKnowledge of New Caledonian palms (Arecaceae) has increased dramatically during the past few years with a wealth of new morphological, anatomical, ecological and molecular data now available. This information makes necessary several changes in the definition of New Caledonian endemic genera. In this article, we present a synopsis of all palm genera of New Caledonia and the subtribes to which they belong, keys for their identification, and a checklist of the species with new combinations where necessary. We retain nine genera of Areceae instead of the 15 previously recognised. The monotypic genera Alloschmidia, Campecarpus, Moratia, Lavoixia and Veillonia, and the bispecific genus Brongniartikentia, are reduced into synonymy with other genera. We provide a description and discussion of the correspondingly enlarged genera Basselinia, Cyphophoenix, Cyphokentia and Clinosperma, as well as notes on the four subtribes now recognised on the island: Livistoninae, Archontophoenicinae, Clinospermatinae and Basseliniinae. In addition, we formally incorporate the monotypic genus Lepidorrhachis from Lord Howe Island within the Basseliniinae. As a result of the changes made here at the generic and specific levels, the family Arecaceae in New Caledonia now comprises 10 genera and 38 species. Five new combinations are made.