Thierry Leroy

Genetics of coffee quality

Coffee quality, in the present context of overproduction worldwide, has to be considered as a main selection criterion for coffee improvement. After a definition of quality, and an overview of the non genetic factors affecting its variation, this review focuses on the genetic factors involved in the control of coffee quality variation. Regarding the complexity of this trait, the different types of quality are first presented. Then, the great variation within and between coffee species is underlined, mainly for biochemical compounds related to quality (caffeine, sugars, chlorogenic acids, lipids). The ways for breeding quality traits for cultivated species, Coffea arabica and Coffea canephora are discussed, with specific challenges for each species. For C. arabica, maintaining a good quality in F 1 intraspecific hybrids, introgressed lines from Timor hybrid, and grafted varieties are the main challenges. For C. canephora, the improvement is mainly based on intraspecific and interspecific hybrids, using the whole genetic variability available within this species. An improvement is obtained for bean size, with significant genetic gains in current breeding programmes. The content in biochemical compounds related to cup quality is another way to improve Robusta quality. Finally, ongoing programmes towards the understanding of the molecular determinism of coffee quality, particularly using coffee ESTs, are presented.

Differentially expressed genes and proteins upon drought acclimation in tolerant and sensitive genotypes of Coffea canephora

The aim of this study was to investigate the molecular mechanisms underlying drought acclimation in coffee plants by the identification of candidate genes (CGs) using different approaches. The first approach used the data generated during the Brazilian Coffee expressed sequence tag (EST) project to select 13 CGs by an in silico analysis (electronic northern). The second approach was based on screening macroarrays spotted with plasmid DNA (coffee ESTs) with separate hybridizations using leaf cDNA probes from drought-tolerant and susceptible clones of Coffea canephora var. Conilon, grown under different water regimes. This allowed the isolation of seven additional CGs. The third approach used two-dimensional gel electrophoresis to identify proteins displaying differential accumulation in leaves of drought-tolerant and susceptible clones of C. canephora. Six of them were characterized by MALDI-TOF-MS/MS (matrix-assisted laser desorption-time of flight-tandem mass spectrometry) and the corresponding proteins were identified. Finally, additional CGs were selected from the literature, and quantitative real-time polymerase chain reaction (qPCR) was performed to analyse the expression of all identified CGs. Altogether, >40 genes presenting differential gene expression during drought acclimation were identified, some of them showing different expression profiles between drought-tolerant and susceptible clones. Based on the obtained results, it can be concluded that factors involved a complex network of responses probably involving the abscisic signalling pathway and nitric oxide are major molecular determinants that might explain the better efficiency in controlling stomata closure and transpiration displayed by drought-tolerant clones of C. canephora.

Effects of shade on the development and sugar metabolism of coffee (Coffea arabica L.) fruits.

Coffee fruits grown in shade are characterized by larger bean size than those grown under full-sun conditions. The present study assessed the effects of shade on bean characteristics and sugar metabolism by analyzing tissue development, sugar contents, activities of sucrose metabolizing enzymes and expression of sucrose synthase-encoding genes in fruits of coffee (Coffea arabica L.) plants submitted to full-sun (FS) and shade (SH) conditions. Evolution of tissue fresh weights measured in fruits collected regularly from flowering to maturation indicated that this increase is due to greater development of the perisperm tissue in the shade. The effects of light regime on sucrose and reducing sugar (glucose and fructose) contents were studied in fresh and dry coffee beans. Shade led to a significant reduction in sucrose content and to an increase in reducing sugars. In pericarp and perisperm tissues, higher activities of sucrose synthase (EC 2.4.1.13) and sucrose-phosphate synthase (SPS: EC 2.4.1.14) were detected at maturation in the shade compared with full sun. These two enzymes also had higher peaks of activities in developing endosperm under shade than in full sun. It was also noted that shade modified the expression of SUS-encoding genes in coffee beans; CaSUS2 gene transcripts levels were higher in SH than in FS. As no sucrose increase accompanied these changes, this suggests that sucrose metabolism was redirected to other metabolic pathways that need to be identified.

Recurrent selection of tropical tree crops

The recurrent selection schemes recommended by CIRAD for 6 tropical tree crops are presented and compared. Breeding programmes are carried out under cooperation between CIRAD and its partners in regions where the crops are grown. The crops are cacao, coffee, rubber tree, oil palm, coconut and eucalyptus. After a short look at the background, the reasons behind the options chosen are given and the main characteristics of the schemes are described and illustrated. The state of progress and some major results are discussed. Over and above the differences linked to plant biology, the genetic diversity available and the type of varietal output, common characteristics are emphasized. The time taken for each generation means simplifying the intercrossing phases between successive breeding cycles, and thus, the parents tested are heterozygous: whenever possible, a clonal varietal output is an additional source of progress. The use of biotechnologies to study diversity is a valuable asset, and their application for early selection (QTL) is also promising. The preponderance of combining ability coexists with marked heterosis, which poses the problem of the latters origin. As a result, all of the schemes described, but one, are reciprocal recurrent selection schemes.

Molecular characterization of an ethylene receptor gene (CcETR1) in coffee trees, its relationship with fruit development and caffeine content.

To understand the importance of ethylene receptor genes in the quality of coffee berries three full-length cDNAs corresponding to a putative ethylene receptor gene (ETR1) were isolated from Coffea canephora cDNA libraries. They differed by their 3′UTR and contained a main ORF and a 5′UTR short ORF putatively encoding a small polypeptide. The CcETR1 gene, present as a single copy in the C. canephora genome, contained five introns in the coding region and one in its 5′UTR. Alternative splicing can occur in C. canephora and C. pseudozanguebariae, leading to a truncated polypeptide. C. pseudozanguebariaeETR1 transcripts showed various forms of splicing alterations. This gene was equally expressed at all stages of fruit development. A segregation study on an inter-specific progeny showed that ETR1 is related to the fructification time, the caffeine content of the green beans, and seed weight. Arabidopsis transformed etiolated seedlings, which over-expressed CcETR1, displayed highly reduced gravitropism, but the triple response was observed in an ethylene enriched environment. These plants behaved like a low-concentration ethylene-insensitive mutant thus confirming the receptor function of the encoded protein. This gene showed no induction during the climacteric crisis but some linkage with traits related to quality.

Stable resistance against the leaf miner #Leucoptera coffeella# expressed by genetically transformed #Coffea canephora# in a pluriannual field experiment in French Guiana

SummaryA pluriannual field trial of transgenic clones of Coffea canephora (the Robusta coffee tree) transformed for resistance to the lepidopteran coffee leaf miner Leucoptera coffeella was installed in French Guiana. Fifty-eight transformed clones produced by transformation of the C. canephora clone 126 were planted. They were harbouring the pEF1α constitutive promoter of Arabidopsis thaliana controlling either the Bacillus thuringiensis native gene for the cry1Ac insecticidal protein (eight clones) or a synthetic cry1Ac gene (53 clones). The vectors for the transformation were a strain of the bacterium Agrobacterium tumefaciens and one of Agrobacterium rhizogenes. The transformed clones were generally independent, presenting different integration patterns of the genetic construct. Four randomly distributed groups of five plants per transformed clone were planted along with 60 untransformed control trees. Over a 4-year period after plantation six releases of L. coffeella were performed. Mines on the leaves are the marks of larvae development and were counted on plants. A majority of the independent transformed clones harbouring the synthetic gene and transformed by the strain of A. tumefaciens displayed constantly much less mines than the control, therefore expressing a stable resistance. The need for complementary research is presented.

Genetic Architecture of Palm Oil Fatty Acid Composition in Cultivated Oil Palm (Elaeis guineensis Jacq.) Compared to Its Wild Relative E. oleifera (H.B.K) Cortés

We searched for quantitative trait loci (QTL) associated with the palm oil fatty acid composition of mature fruits of the oil palm E. guineensis Jacq. in comparison with its wild relative E. oleifera (H.B.K) Cortés. The oil palm cross LM2T x DA10D between two heterozygous parents was considered in our experiment as an intraspecific representative of E. guineensis. Its QTLs were compared to QTLs published for the same traits in an interspecific Elaeis pseudo-backcross used as an indirect representative of E. oleifera. Few correlations were found in E. guineensis between pulp fatty acid proportions and yield traits, allowing for the rather independent selection of both types of traits. Sixteen QTLs affecting palm oil fatty acid proportions and iodine value were identified in oil palm. The phenotypic variation explained by the detected QTLs was low to medium in E. guineensis, ranging between 10% and 36%. The explained cumulative variation was 29% for palmitic acid C16:0 (one QTL), 68% for stearic acid C18:0 (two QTLs), 50% for oleic acid C18:1 (three QTLs), 25% for linoleic acid C18:2 (one QTL), and 40% (two QTLs) for the iodine value. Good marker co-linearity was observed between the intraspecific and interspecific Simple Sequence Repeat (SSR) linkage maps. Specific QTL regions for several traits were found in each mapping population. Our comparative QTL results in both E. guineensis and interspecific materials strongly suggest that, apart from two common QTL zones, there are two specific QTL regions with major effects, which might be one in E. guineensis, the other in E. oleifera, which are independent of each other and harbor QTLs for several traits, indicating either pleiotropic effects or linkage. Using QTL maps connected by highly transferable SSR markers, our study established a good basis to decipher in the future such hypothesis at the Elaeis genus level.

Ancestral synteny shared between distantly-related plant species from the asterid (Coffea canephora and Solanum Sp.) and rosid (Vitis vinifera) clades

BackgroundCoffee trees (Rubiaceae) and tomato (Solanaceae) belong to the Asterid clade, while grapevine (Vitaceae) belongs to the Rosid clade. Coffee and tomato separated from grapevine 125 million years ago, while coffee and tomato diverged 83-89 million years ago. These long periods of divergent evolution should have permitted the genomes to reorganize significantly. So far, very few comparative mappings have been performed between very distantly related species belonging to different clades. We report the first multiple comparison between species from Asterid and Rosid clades, to examine both macro-and microsynteny relationships.ResultsThanks to a set of 867 COSII markers, macrosynteny was detected between coffee, tomato and grapevine. While coffee and tomato genomes share 318 orthologous markers and 27 conserved syntenic segments (CSSs), coffee and grapevine also share a similar number of syntenic markers and CSSs: 299 and 29 respectively. Despite large genome macrostructure reorganization, several large chromosome segments showed outstanding macrosynteny shedding new insights into chromosome evolution between Asterids and Rosids. We also analyzed a sequence of 174 kb containing the ovate gene, conserved in a syntenic block between coffee, tomato and grapevine that showed a high-level of microstructure conservation. A higher level of conservation was observed between coffee and grapevine, both woody and long life-cycle plants, than between coffee and tomato. Out of 16 coffee genes of this syntenic segment, 7 and 14 showed complete synteny between coffee and tomato or grapevine, respectively.ConclusionsThese results show that significant conservation is found between distantly related species from the Asterid (Coffea canephora and Solanum sp.) and Rosid (Vitis vinifera) clades, at the genome macrostructure and microstructure levels. At the ovate locus, conservation did not decline in relation to increasing phylogenetic distance, suggesting that the time factor alone does not explain divergences. Our results are considerably useful for syntenic studies between supposedly remote species for the isolation of important genes for agronomy.

Global analysis of Coffea canephora Pierre ex Froehner (Rubiaceae) from the Guineo-Congolese region reveals impacts from climatic refuges and migration effects

As the second species used for commercial coffee, evaluation of Coffea canephora Pierre ex Froehner population genetics is a challenging task for coffee breeding. This study examined the pattern of genetic variability and genetic relationships of cultivated and wild populations of C. canephora sampled across the Guineo-Congolese region of Africa and some improved populations maintained in field genebanks. A total of 293 individuals, sampled from 17 populations, were genotyped with 39 nuclear microsatellite markers. Genetic diversity and structure were investigated with both a model-based and a graphical approach; isolation by distance was also tested. Relationships between the diversity clusters are discussed with regard to differentiation due to several glacial refuges during the Last Glacial Maximum (LGM). High genetic diversity within C. canephora is confirmed with a mean number of alleles of 11.85 per marker, a mean gene diversity of 0.72 and a mean observed heterozygosity of 0.36. An overall structure of two main groups (Guinean and Congolese) subdivided in six subgroups (2 for the Guinean and 4 for the Congolese) was found, including one described for the first time in the Guinean group. A fine structure within the Guinean group was also newly detected. Genetic structure of C. canephora appears to be consistent with its geographic repartition at the continent scale. Structure of diversity was found in accordance with localizations of refuge zones during LGM and migration from this period. Results from this genetic structure study raise our capabilities to better manage and use the collections of genetics resources for breeding purposes. Those results will be used in future association studies to optimize the number of genotypes to be phenotyped.

An initial assessment of linkage disequilibrium (LD) in coffee trees: LD patterns in groups of Coffea canephora Pierre using microsatellite analysis.

BackgroundA reciprocal recurrent selection program has been under way for the Coffea canephora coffee tree for approximately thirty years in the Ivory Coast. Association genetics would help to speed up this program by more rapidly selecting zones of interest in the genome. However, prior to any such studies, the linkage disequilibrium (LD) needs to be assessed between the markers on the genome. These data are essential for guiding association studies.ResultsThis article describes the first results of an LD assessment in a coffee tree species. Guinean and Congolese breeding populations of C. canephora have been used for this work, with the goal of identifying ways of using these populations in association genetics. We identified changes in the LD along the genome within the different C. canephora diversity groups. In the different diversity groups studied, the LD was variable. Some diversity groups displayed disequilibria over long distances (up to 25 cM), whereas others had disequilibria not exceeding 1 cM. We also discovered a fine structure within the Guinean group.ConclusionsGiven these results, association studies can be used within the species C. canephora. The coffee recurrent selection scheme being implemented in the Ivory Coast can thus be optimized. Lastly, our results could be used to improve C. arabica because one of its parents is closely related to C. canephora.