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Basal rot is a widespread onion disease caused by fungi belonging to Fusarium genus. The disease is known to incur onion yield loss worldwide. This study aims to identify and determine the pathogenicity of Fusarium species associated with basal rot of onions in Burkina Faso. Thirteen Fusarium isolates from seven regions of Burkina Faso were identified based on morphological observations and molecular diagnosis and their pathogenicity was assessed in laboratory and Greenhouse. Results showed that, the 13 isolates belonged to Fusarium falciforme, Fusarium acutatum and Fusarium oxysporum species. F. falciforme, F. acutatum and F. oxysporum were pathogenic on onion seeds, seedlings and bulbs. F. falciforme caused lower seedling damping-off rate compared to that caused by F. acutatum and F. oxysporum. All these species of Fusarium genus induced onion bulbs rots. The results of this study confirmed the presence of these species in Burkina Faso and that they are pathogenic on onion. Therefore, it would be useful to suggest an implement sustainable management approach of these pathogens.

Additional file 3: Figure S3. Characterization of the 12 sampled key developmental stages throughout the Arabica somatic embryogenesis (SE) process at morphological level.

Additional file 1: Figure S1. The availability of large-scale protocols for coffee somatic embryogenesis (SE) guaranteed reliability and development synchronization at each developmental stage as well as biological efficiency.

Additional file 2: Figure S2. Metabolic pathways and hormone dynamics during the four main developmental phase switches.

RATIONALE - Arabusta coffee was created by crossing Coffea arabica L. and C. canephora Pierre ex-Froehner after doubling the chromosome number of C. canephora. For breeding studies and coffee quality improvement, clones have been planted in French Guiana. As green bean metabolite content largely influences the coffee cup quality, biochemical and sensory analyses of Robusta and Arabusta were done on green beans and compared as part of Breedcafs H2020 project. METHODS - Arabusta cherries were harvested on 6 clones in 4 farms in Apatou and in Combi research station, and those from Robusta in Combi research station. They were wet processed before grinding. Phenolic compounds and alkaloids were quantified by HPLC (Campa et al, 2017), sucrose by enzymatic analysis (Megazyme kit, Ireland), diterpenes by NMR (Portaluri et al, 2020) and VOC by headspace SPME-GC/ MS (Marie et al, 2020). Lipids were extracted (Folch et al, 1957) and quantified by cgfb (Bordeaux). Sensory analyses followed the protocol guidelines of the Specialty Coffee Association (SCA) for C. arabica (Marie et al, 2020).We used Statistica software for analyses. RESULTS - Compared to Robusta, Arabusta had a lower caffeine content and higher trigonelline, sucrose and fatty acid contents. Arabusta evidenced both 16-O-methylcafestol and kahweol, known markers for Robusta and Arabica species, respectively. 17 of the 25 non-volatile and 23 of the 47 volatile compounds showed significant differences (p≤0.05) between Arabusta and Robusta. Except 5-caffeoylquinate and coumaroylquinate, chlorogenic acids accumulated more in Robusta. A PCA based on volatile compound content separated samples in three groups: Robusta samples, most of the Arabusta from Apatou (richer in pyrazine, toluene, 2-propanone) and those from Combi associated with 3 from Apatou (richer in D-limonene, DMSO, methyl-butanoic acid, and IBMP). A sensory analysis based on 9 sensory parameters indicated that 3 samples received a final score higher than 80. A PCA confirmed that 2 Arabusta samples were particularly different from all the others. CONCLUSIONS & PERSPECTIVES - With higher contents in sucrose, trigonelline, diterpenes, fatty acids and D-limonene and a lower content in caffeine, the composition of the Arabusta green beans seems significantly different from Robusta. The sensory analysis supports the hypothesis that some Arabusta clones possess the potential to produce quality coffee. However, it is necessary to continue the research to confirm these results. This research was funded by BREEDCAFS project, supported by the European Commission under the Horizon 2020 – Research and Innovative Program, H2020-SFS-2016-2, grant agreement number: 727934.

Photoperiod length induces in temperate plants major changes in growth rates, morphology and metabolism with, for example, modifications in the partitioning of photosynthates to avoid starvation at the end of long nights. However, this has never been studied for a tropical perennial species adapted to grow in a natural photoperiod close to 12 h/12 h all year long. We grew Coffea arabica L., an understorey perennial evergreen tropical species in its natural 12 h/12 h and in a short 8 h/16 h photoperiod, and we investigated its responses at the physiological, metabolic and transcriptomic levels. The expression pattern of rhythmic genes, including core clock genes, was affected by changes in photoperiod. Overall, we identified 2859 rhythmic genes, of which 89% were also rhythmic in Arabidopsis thaliana L. Under short-days, plant growth was reduced, and leaves were thinner with lower chlorophyll content. In addition, secondary metabolism was also affected with chlorogenic acid and epicatechin levels decreasing, and in agreement, the genes involved in lignin synthesis were overexpressed and those involved in the flavanol pathway were underexpressed. Our results show that the 8 h/16 h photoperiod induces drastic changes in morphology, metabolites and gene expression, and the responses for gene expression are similar to those observed in the temperate annual A. thaliana species. Short photoperiod induces drastic changes in gene expression, metabolites and leaf structure, some of these responses being similar to those observed in A. thaliana.

RATIONALE - The circadian clock (CC) is a critical regulator to optimize the plant physiology and metabolism to the correct time of the day, providing plants with the ability to anticipate daily and seasonal environmental changes. Plant transcriptome is massively modulated by the CC which is an integrator of many environmental cues. Through several experiments we show here how the CC of Arabica is modulated by the environment and how the genetic background can also modulate the CC with huge impact on primary metabolism. METHODS RNA sequencing was systematically used to study gene expression over 24h in combination with metabolomical and physiological analysis. Then we have integrated information of multiple circadian RNA-seq diurnal time-course in order to estimate the fraction of the Arabica transcriptome that is circadian regulated. Comparison of all the rhythmic genes identified in coffee genome with those of Arabidopsis thaliana revealed they are identical at almost 90%. RESULTS - We present how photoperiod, full-moon light, and thermos-cycles, led to changes in transcript abundance of thousand of genes during the diurnal cycle altered the CC amplitude which in cascade modify the expression of thousand of rythmic genes. Moreover, we will explain how the genetic background can change the CC and the implication in terms of primary metabolism, physiological behaviour and disease resistance. In this presentation we pay special attention to three major genes of the core clock for which we recall how important they are for the regulation of the transcriptome and in fine on the development of the plant and the major agronomic traits. CONCLUSIONS & PERSPECTIVES - Variations in the environment and/or genetic background have a major impact on the rhythmicity and daily amplitude of expression of a large number of Arabica coffee genes. The same trends as in model plants (notably Arabidopsis) are found here. The study of variations in the expression of the main core clock genes is fundamental and needs to be refined. A major question that needs to be addressed is: can we find allelic variation in circadian genes in the Arabica species and can we use it to breed news varieties with higher performances.

RATIONALE - Somatic embryogenesis (SE) is one of the most promising processes for large-scale dissemination of elite varieties. However, whatever the species considered, SE research still remains essentially empirical resulting in many drawbacks to fulfil market demands, especially due to an overall slow technical progress over the last 20 years. Knowledge about the molecular events involved in the key steps of the SE process is urgently needed to pilot the optimization of SE protocols. In this study, we took advantage of the latest metabolomics technologies and applied them to one of the most advanced and reliable large-scale SE processes, the one developed for coffee. METHODS - Sampling covered 15 key developmental stages. Five independent leaf introductions were carried out with more than 4,000 leaf explants and a total of 25 independent cell lines. All obtained cell lines were high-yielding and time-synchronized during embryo regeneration enabling a successful sampling. Primary metabolites, secondary metabolites and phytohormones were quantified using GC-MS, HPLC and UPLC- MS/MS respectively. A robust statistical method was used to identify metabolic pathway changes associated with the main developmental phases and phase switches. Histological analysis and cell imaging were also required to characterize developmental stages and associate metabolic profiles with cell structure organization. Lastly, comparing Arabica embryogenic and non-embryogenic calli enabled the identification of metabolic markers of the embryogenic capacity. RESULTS - Statistical analysis performed on 104 metabolites revealed that massive re-configuration of metabolic pathways induced SE. During initial dedifferentiation, a sharp decrease in phenolic compounds and caffeine levels was observed while auxins, cytokinins and ethylene levels were at their highest. Totipotency reached its highest expression during the callus stages when a shut-off in hormonal and metabolic pathways related to sugar and energetic substance hydrolysis was evidenced. Abscisic acid, leucine, maltotriose, myo-inositol, proline, tricarboxylic acid cycle metabolites and zeatin appeared as key metabolic markers of the embryogenic capacity. Combining metabolomics with multiphoton microscopy led to the identification of chlorogenic acids as markers of embryo redifferentiation. CONCLUSIONS & PERSPECTIVES - The present analysis shows that metabolite fingerprints are signatures of cell fate and represent a starting point for optimizing SE protocols in a rational way. These findings should be informative and useful to a wide range of plant species, offering unprecedented perspectives in plant micropropagation.

RATIONALE - Arabusta coffee is an interspecific hybrid derived from C. arabica x C. canephora, often used in coffee breeding to study both the gene transfer from Robusta and its possible use in coffee production. The plants have very different morphological characteristics, as well as different levels of fruit production when compared with the parental species. Arabusta coffee was also studied from a sensory point of view, but in general the literature on this hybrid is rather scarce and the knowledge on it is still fragmentary. From a morphoanatomical point of view, as far as we know, no investigations have been devoted to study Arabusta seed morphology and to disclose possible Arabica-like or Robusta-like traits in its cell structure and this highly stimulated the present work. METHODS - Seed samples of C. arabica L. (Costarica, 2019), C. canephora Pierre ex-Froehner (French Guyana, 2019) and F1 clones of Arabusta coffee (French Guyana, end of 2018) were harvested, properly processed and selected. Seeds were kept in a fixative solution for several days, then rinsed in tap water and cut at -20°C with a cryostat (Leica CM1520). Seed sections of 60 μ in thickness were observed by a Scanning Electron Microscope (Hitachi TM3030plus). Sections of 12 μ in thickness were staining in a Toluidine Blue O solution to highlight the main cell components. Measurements of cell wall area were performed on the electronic images by a Leica Software (Las X). RESULTS ( Arabusta is characterized by an elongated seed, slightly rounded, generally with a linear furrow. The endosperm cells are regular, characterized by a polygonal shape, with not frequent nodes present in their cell wall. Cell wall thickness is not significantly different from the parental species (5,7 ± 1,6 μ). However, the area occupied by the cell wall respect to the cell total area (28%) is more close to that measured in Arabica seeds (29%) than that in Robusta seeds (43%). Optical microscopy observations furtherly put in evidence this tissue aspect, showing similar cell size in Arabusta and Arabica seeds, and smaller cells for Robusta coffee. This feature is probably related to ploidy level (Arabica and Arabusta : 2n = 4x = 44, Robusta 2n = 2x = 22). No histochemical differences in the cell content were observed among the examined coffee species under the chosen experimental conditions. CONCLUSIONS & PERSPECTIVES - The measurements performed on cell wall area and measured diameter as well as cell content area and endosperm cell size show that the morphoanatomical characteristics of Arabusta seeds corresponds more to those of Arabica than Robusta seeds. Further studies are necessary to confirm this preliminary view.

Conventional American cultivars of coffee are no longer adapted to global warming. Finding highly productive and stable cultivars in different environments without neglecting quality characteristics has become a priority for breeders. In this study, new Arabica F1 hybrids clones were compared to conventional American varieties in seven contrasting environments, for yield, rust incidence and volume of the canopy. The quality was assessed through size, weight of 100 beans, biochemical analysis (24 aroma precursors and 31 volatiles compounds) and sensory analysis. Conventional varieties were the least productive, producing 50% less than the best hybrid. The AMMI model analysis pointed out five hybrids as the most stable and productive. Two F1 hybrids clones, H1-Centroamericano and H16-Mundo Maya, were superior to the most planted American cultivar in Latin and Central America showing a high yield performance and stability performance. H1-Centroamerica and Starmaya contain more d-limonene than Caturra, while Starmaya contain more 3-methylbutanoic acid than the control. Those two latter volatiles compounds are linked with good cup quality in previous studies. In terms of sensory analysis, Starmaya and H1-Centroamericano scored better than control.

Conventional American cultivars of coffee are no longer adapted to global warming. Finding highly productive and stable cultivars in different environments without neglecting quality characteristics has become a priority for breeders. In this study, new Arabica F1 hybrids clones were compared to conventional American varieties in seven contrasting environments, for yield, rust incidence and volume of the canopy. The quality was assessed through size, weight of 100 beans, biochemical analysis (24 aroma precursors and 31 volatiles compounds) and sensory analysis. Conventional varieties were the least productive, producing 50% less than the best hybrid. The AMMI model analysis pointed out five hybrids as the most stable and productive. Two F1 hybrids clones, H1-Centroamericano and H16-Mundo Maya, were superior to the most planted American cultivar in Latin and Central America showing a high yield performance and stability performance. H1-Centroamerica and Starmaya contain more d-limonene than Caturra, while Starmaya contain more 3-methylbutanoic acid than the control. Those two latter volatiles compounds are linked with good cup quality in previous studies. In terms of sensory analysis, Starmaya and H1-Centroamericano scored better than control.

Culture under shade in AgroForestry Systems (AFS) is proposed to face the predicted negative effects of climate change on coffee plants. To decipher the molecular mechanisms involved in coffee tree adaptation to lower light, cultivars breaded for intensive full sun production systems have been grown under shade and full sun, in Mexico and Colombia. Leaves have been harvested for biochemical and transcriptomic comparative analyses. A common biochemical and transcriptomic response is observed in mature leaves of two years-old juvenile trees of C. arabicavar. Marsellesa cultivated in Mexico at different altitudes and of six-years old producing trees of eight different Arabica genotypes cultivated in Colombia. HPLC analysis showed a significant decrease in phenolic content under shade, especially glycosylated flavonoids involved in light protection. Transcriptomic analyses showed that 70% of the under-expressed genes under shade are heat shock proteins involved in temperature but also in various abiotic stress responses. Overall, our results show that Arabica culture under shade limits the biological costs for the plant, due to stresses. This energy saving opens new question: are the resources more allocated to primary metabolism, bean production and homeostasis? We are currently testing these genes and metabolites as markers to predict the adaptation to AFS and associated abiotic stresses.

Arabica coffee is a main economic crop throughout Central and South American highlands, where millions of people get primary outcome. Last coffee leaf rust (CLR) epidemic event in 2011, which impacted 53% of the coffee region areas, provoking losses up to USD 500 million, remembered the high vulnerability of the coffee sector to the fungus Hemileia vastatrix, causal agent of this disease and considered as the main restrictive illness for all coffee regions around the world. The main problem remains that the current commercial varieties reputed as "resistant", become sensitive to the CLR. During the last twenty years, coffee geneticist have created several intra-specific hybrids between commercial varieties and wild coffee accessions. Field observations confirmed that coffee hybrids are more productive and less impacted by the CLR even in regions where resistant materials become susceptible. Such field observations are in agreement with the general idea that hybrid plants present high homeostasis in terms of growth, yield and disease resistance. The aim of this research was to investigate the physiological response of coffee hybrids before and after inoculation with the CLR. Using controlled conditions, the metabolic response and physiological status of two hybrid genotypes were assessed under different stressful conditions involving three limiting factors: temperature, luminosity and nitrogen input. Hybrid behavior was compared to an inbred line tested under similar conditions. Results demonstrated that whatever the agronomic conditions and the temperature regimes, hybrids under test were less sensitive to CLR than the inbred line. Interestingly, the detailed transcriptomic analysis revealed that coffee hybrids seem to exhibit an altered circadian clock resulting in a higher photosynthetic efficiency together with an increased concentration of chlorophyll. Most of the over-expressed genes identified in the hybrids after CLR inoculation were associated to basal resistance mechanisms, while genes identified in the inbred line, were more related to abiotic stress responses. Overall our findings suggest that under stressful conditions, coffee hybrids are able to rapidly modify their energetic metabolism machinery leading to a more effective and rapid response to rust attack compared to the inbred line.

Nowadays coffee somatic embryogenesis (SE) can be considered as a model for woody perennial crops as its industrial application already allowed the dissemination of 50 millions of elite plants in the two cultivated species. The process works with high biological efficiency is observed at all developmental stages and somaclonal variation is mastered. Current research beneficiates from this regeneration model as well as from the progress of coffee genomics and biotechnologies, focuses on two distinct objectives. The recent advances of this research and their respective outputs are reviewed and discussed in this presentation. The first objective is to develop basic research based on-omics methodologies to decipher the fondamental mechanisms associated to cell reprogramming by focusing on two examples: i) Taking advantage of optimized SE protocols, we will unravel totipotency mechanisms and molecular events involved in the key developmental switches occurring during cell dedifferentiation/redifferentiation. A multi approach characterization through cell imaging, massive transcriptomic, epigenetic and metabolomic analyzes is applied at all the key developmental stages from the leaf explant to torpedo-shaped embryos (7 well characterized developmental stages). Big data integration and a system biology approach will provide a thorough knowledge of the expression of totipotency. ii) Coffee embryos acquire the capacity to photosynthetize very early in their developmental process (at the cotyledonary stage). Understanding the events leading to the acquisition of photo-autotrophy is also being studied by ecophysiological and molecular approaches applied to torpedo and cotyledonary-shaped embryo cultures under contrasted light, C02 and sugar conditions. Otherwise, the second objective is to provide - through technological approaches - powerful tools to accelerate the optimization of coffee breeding: i) The high-throughput molecular screening method has been recently developed by Nestle that allows to miniaturize the SE process and opens the way to study the biological effect of new active molecules on particular developmental switches' efficiency. It will also allow the development of miniaturized EMS (Ethyl Methyl Sulphate) mutant banks by simplifying their setting up and further management. ii) The genome editing of coffee genome by the Crispr/Cas9 technology recently described by using embryogenic tissues transformation gives us the possibility to rapidly and precisely manipulate plant genomes to introduce a trait of agronomic interest.

In a context where climate change is threatening coffee productivity, the management of coffee leaf rust is a challenging issue. Major resistant genes, which have been used for many years, are systematically being overcome by pathogens. Developing healthy plants, able to defend themselves and be productive even when attacked by the pathogen, should be part of a more sustainable alternative approach. We compared one hybrid (GPFA124), selected for its good health in various environments including a reduced rust incidence, and the cv. &lsquo, Caturra&rsquo, considered as a standard in terms of productivity and quality but highly susceptible to rust, for phenotypic variables and for the expression of genes involved in the circadian clock and in primary photosynthetic metabolism. The GPFA124 hybrid showed increased photosynthetic electron transport efficiency, better carbon partitioning, and higher chlorophyll content. A strong relationship exists between chlorophyll a fluorescence and the expression of genes related to the photosynthetic electron transport chain. We also showed an alteration of the amplitude of circadian clock genes in the clone. Our work also indicated that increased photosynthetic electron transport efficiency is related to the clone&rsquo, s better performance. Chlorophyll a fluorescence measurement is a good indicator of the coffee tree&rsquo, s physiological status for the breeder. We suggest a connection between the circadian clock and carbon metabolism in coffee tree.