Search

Citrus fruit quality is defined as the combination of physical and chemical traits; some of which may change during the ripening phase, e.g., acidity and sugar content. A clear understanding of their genetic control would be very helpful for marker-assisted breeding programs especially with regard to the juvenile phase and some reproductive features that hamper the selection of improved hybrids. A genetic study was thus performed on the heredity of quality traits and QTL detection based on segregation in a progeny generated from a cross between clementine cv “Commun” (Citrus × reticulata cv clementine) and mandarin cv “Willow leaf” (C. reticulata Blanco). Parental and consensus genetic linkage maps were constructed using 645 SNP and SSR markers. These maps were represented by 10 linkage groups in clementine and 12 linkage groups in mandarin, representing 75% and 58% respectively of the previously published clementine reference map. A total of 16 traits, including fruit mass, equatorial diameter, juice percentage, total soluble solids, acidity, pH, glucose, fructose, sucrose, and citric and malic acid concentrations were evaluated at three maturation dates. High variations indicating transgressive segregation were found for all traits, with normal or close to normal distributions. QTL analysis performed using the multiple QTL model allowed the detection of 34 QTLs on the three maps. QTLs were distributed in different linkage groups and generally detected at only one date of the ripening phase. The percentage of total variation explained ranged from 12 to 37% per QTL. Major QTLs (R2 ≥ 30%) were detected for equatorial diameter, glucose, and fructose (expressed in percentage dry matter) on linkage groups 8 and 9. Co-localization of QTLs controlling correlated and uncorrelated traits were mainly found on linkage groups 2, 4, 8, and 9, particularly between fruit mass and acidity.

The peel essential oil (PEO) of sweet orange is used for flavoring liquors or foods and in the perfumery and cosmetics industry. The fruit maturity stage can modify the essential oil composition and aromatic properties, but little information is available on the evolution of PEO during the entire time set of fruit development. In this study, the yield, chemical composition and aromatic profile over the three phases of orange development were monitored. Four fruit traits (peel color, weight, acidity and sweetness) were recorded to characterize fruit development. Fruits of two sweet orange cultivars were sampled every two weeks from June to May of the next year. PEO was obtained by hydrodistillation and analyzed by gas chromatography coupled with a flame ionization detector (GC-FID). Compounds were identified with GC coupled with mass spectrometry (GC/MS). Ten expert panelists using the descriptor intensity method described the aromatic profile of PEO samples. The PEO composition was richer in oxygenated compounds at early fruit development stages, with an aromatic profile presenting greener notes. During fruit growth (Phases I and II), limonene's proportion increased considerably as a few aliphatic aldehydes brought the characteristic of orange aroma. During fruit maturation (from November to March), the PEO composition and aromatic profile were relatively stable. Later, some modifications were observed. Regardless of the fruit development stage, the two sweet oranges presented distinct PEO compositions and aromatic profiles. These results constitute a temporal reference for the chemical and aromatic evolution of sweet orange PEO in the fruit development process under Mediterranean conditions. During the first two phases of fruit development, many changes occur in the PEO composition and aroma, suggesting that their exploitation could create new products.

The great genetic and phenotypic diversities characterize the mandarin species (Citrus reticulata). The present study aimed to evaluate a part of this diversity with molecular markers and through the composition of leaf (LEO) and fruit peel (PEO) essential oils. Seventy-two cultivars were chosen for this study to represent some wild and cultivated mandarins growing in the same orchards. The essential oils were analyzed via gas chromatography (retention indices) and via gas chromatography coupled with mass spectrometry. The low similarity of ‘Tachibana’ and ‘Korail tachibana’ with mandarins and other species suggested that they were pure mandarins but were not involved in the genesis of the cultivated forms. This distinction was also evident at the aromatic level with specific compounds or unusual proportions, such as δ-3-carene in PEO or β-phellandrene at 24.9% in ‘Korail tachibana’ LEO. ‘Kunembo’ and ‘Ben di gang ju’ were genetically and chemically identical, with a high proportion of myrcene (>20%) in their LEO. In general, the genetic diversity of SSR markers was higher than the chemical diversity. From the 72 accessions, 54 genotypes were identified, with only 8 aromatic profiles in PEO and 9 in LEO. This diversity of essential oils of mandarins offers new perspectives for the research and validation of new aromatic properties for food and cosmetic purposes.

International audience; Ichang lemon is a citrus fruit whose rind gives off a delicious and much appreciated fragrance and flavor. The volatile components of the fruit peel of Ichang lemon were investigated by GC-MS and GC-O (AEDA method). Simultaneously, its genetic origin was identified by using diagnostic SNP markers specific to ancestral species and multiallelic SSR and InDel markers. Ichang lemon combines three ancestral genomes (Citrus maxima, Citrus ichangensis, and Citrus reticulata) and may be a pummelo x Yuzu hybrid. Although the major compounds of the Ichang lemon aromatic profile were present in Citrus junos, a few pummelo-specific compounds were also detected, such as indole and nootkatone, in agreement with its maternal lineage. 3-Methyl-3-sulfanylbutyl acetate, reported to occur in passion fruit and brewed coffee, was identified by GC-MS, GC-QTOF-MS, and GC-FTIR for the first time in citrus. This odor-active compound has a sulfurous, tropical fruity, green note.

The Papeda Citrus subgenus includes several species belonging to two genetically distinct groups, containing mostly little-exploited wild forms of citrus. However, little is known about the potentially large and novel aromatic diversity contained in these wild citruses. In this study, we characterized and compared the essential oils obtained from peels and leaves from representatives of both Papeda groups, and three related hybrids. Using a combination of GC, GC-MS, and 13C-NMR spectrometry, we identified a total of 60 compounds in peel oils (PO), and 76 compounds in leaf oils (LO). Limonene was the major component in almost all citrus PO, except for C. micrantha and C. hystrix, where β-pinene dominated (around 35%). LO composition was more variable, with different major compounds among almost all samples, except for two citrus pairs: C. micrantha/C. hystrix and two accessions of C. ichangensis. In hybrid relatives, the profiles were largely consistent with their Citrus/Papeda parental lineage. This high chemical diversity, not only among the sections of the subgenus Papeda, but also between species and even at the intraspecific level, suggests that Papeda may be an important source of aroma diversity for future experimental crosses with field crop species.

Gene banks have been established to conserve the genetic diversity of crop species. Large germplasm collections lead to management problems (space, maintenance costs, etc.), especially in collections involving species with recalcitrant seeds that must be maintained as growing plants. Core collections (CCs) are thus developed to reduce the size of large germplasm collections while keeping the maximum variability. This also facilitates fine phenotypic evaluation. In this study, several software packages (DARwin, PowerMarker and MSTRAT) and methods (Max length subtree, M strategy, simulated annealing and MinSD) were compared to define a mandarin (Citrus reticulata) CC. One hundred and sixty-seven accessions were sampled from two germplasm collections, which were genotyped with 50 SSR, 24 InDel and 68 single nucleotide polymorphism markers. All the CC obtained were tested for the maintenance of the genetic variability parameters (Ho and He) of the initial collection, the level of linkage disequilibrium (LD) and the phenotypic diversity retention. The Max length subtree function from DARWin seemed to be the most appropriate method for establishing a CC in C. reticulata. It maintained 96.82% of the allelic richness and 17.96% of the size of the initial collection with only 30 accessions. Besides it did not increase the LD (r2 value) of the initial collection and retained the vast majority of the phenotypic variability. However, a CC with 70 accessions would be more helpful for genetic association studies. (Resume d'auteur)

Sour oranges (Citrus aurantium L.) are well known in the processing and cosmetics industries for the aromatic properties of their essential oils. Intercultivar genetic and aromatic diversity is not well documented. The objective of this study was to evaluate the impact of morphological selection and genetic mechanisms of varietal diversification (mutation or hybridization) on the aromatic and odor variability of sour orange essential oils. Forty-five sour orange accessions from INRAE-CIRAD citrus Biological Resources Center (France) were assessed for ten simple sequence repeat (SSR) and 54 single nucleotide polymorphism (SNP) markers, nine morphochemical fruit traits and with the aromatic components of leaf and peel essential oils. Thirty-nine sour oranges displayed no intercultivar molecular polymorphism and six genotypes originated from interspecific hybridizations involving sour orange, citron, pummelo or mandarin. The peel essential oil (PEO) diversity was low, in accordance with the genetic diversity. The predominance of limonene (&gt, 90%) prevents any possible correlation to be made between the composition and the variation in sensory profiles detected by panelists. Few compounds in the leaf essential oil (LEO), such as linalool, linalyl acetate, α-terpineol and geraniol were significantly different across sour oranges varieties. The morphological fruit attributes mainly used in varietal selection differed highly across the main genetically identical group of sour orange accessions. These results confirm that mutation can generate variability in aromatic compounds and aromas and that their exploitation requires an improvement in characterization processes.

Background and Aims The origin of limes and lemons has been a source of conflicting taxonomic opinions. Biochemical studies, numerical taxonomy and recent molecular studies suggested that cultivated Citrus species result from interspecific hybridization between four basic taxa (C. reticulata, C. maxima, C. medica and C. micrantha). However, the origin of most lemons and limes remains controversial or unknown. The aim of this study was to perform extended analyses of the diversity, genetic structure and origin of limes and lemons. Methods The study was based on 133 Citrus accessions. It combined maternal phylogeny studies based on mitochondrial and chloroplastic markers, and nuclear structure analysis based on the evaluation of ploidy level and the use of 123 markers, including 73 basic taxa diagnostic single nucleotide polymorphism (SNP) and indel markers. Key Results The lime and lemon horticultural group appears to be highly polymorphic, with diploid, triploid and tetraploid varieties, and to result from many independent reticulation events which defined the sub-groups. Maternal phylogeny involves four cytoplasmic types out of the six encountered in the Citrus genus. All lime and lemon accessions were highly heterozygous, with interspecific admixture of two, three and even the four ancestral taxa genomes. Molecular polymorphism between varieties of the same sub-group was very low. Conclusions Citrus medica contributed to all limes and lemons and was the direct male parent for the main sub-groups in combination with C. micrantha or close papeda species (for C. aurata, C. excelsa, C. macrophylla and C. aurantifolia – 'Mexican' lime types of Tanaka's taxa), C. reticulata (for C. limonia, C. karna and C. jambhiri varieties of Tanaka's taxa, including popular citrus rootstocks such as 'Rangpur' lime, 'Volkamer' and 'Rough' lemons), C. aurantium (for C. limetta and C. limon – yellow lemon types – varieties of Tanaka's taxa) or the C. maxima × C. reticulata hybrid (for C. limettioides – 'Palestine sweet' lime types – and C. meyeri). Among triploid limes, C. latifolia accessions ('Tahiti' and 'Persian' lime types) result from the fertilization of a haploid ovule of C. limon by a diploid gamete of C. aurantifolia, while C. aurantifolia triploid accessions ('Tanepao' lime types and 'Madagascar' lemon) probably result from an interspecific backcross (a diploid ovule of C. aurantifolia fertilized by C. medica). As limes and lemons were vegetatively propagated (apomixis, horticultural practices) the intra-sub-group phenotypic diversity results from asexual variations. (Resume d'auteur)

BACKGROUND The physiological roles of organic acids in fruit cells are not fully understood, especially in citrus, whereas the decline in titratable acidity during ripening shown by many citrus fruits is due to the utilization of citric acid. We induced carbohydrate depletion by removing source leaves at two key periods in mandarin development (early and full citric acid accumulation). Then, we assessed the resulting changes in the short term (within 48 h) and long term (several weeks until ripening). RESULTS Control mature fruits were characterized by elevated fresh weight, large diameters and high quantities of malic acid, citric acid and sucrose. At the same stage, fruits subjected to early or late defoliation had higher glucose, fructose, citric acid concentrations and lower sucrose concentrations. They differed only in their malic acid concentrations, which were higher in early defoliation fruits and similar in late defoliation fruits when compared to control fruits. Finally, fruits subjected to late defoliation were characterized by high proline and γ-aminobutyric acid concentrations, and low fructose and glucose concentrations. CONCLUSION We have shown that short- and long-term carbohydrate limitation modifies sugar and organic acid metabolism during mandarin fruit growth. © 2015 Society of Chemical Industry

The chemical composition of the essential oil (EO), microwave extract (ME) and hydrolate extract (HE) from the same batch of leaves of Fortunella japonica, was investigated: by combination of chromatographic (GC, CC) and spectroscopic techniques (GC-MS, 13C NMR). F. japonica essential oil and extracts are complex mixtures of 28-60 compounds being mainly oxygenated sesquiterpenes. The EO composition was dominated by germacrene D (14.9%), β-elemol (9.1%), cis-guai-6-en-10β-ol (6.3%), β-eudesmol (5.5%), and δ-elemene (5.2%). Limonene was the unique monoterpene identified at appreciable amount (7.1%). The extract obtained by microwave assisted hydrodistillation contained as main components: β-elemol (12.4%), germacrene D (9.9%), cis-guai-6-en- 10β-ol (9.0%), β-eudesmol (8.2%), germacra-l(l0),5-dien-4α-ol (7.1%) and α-eudesmol (6.4%). Finally, the highest content of oxygenated sesquiterpenes (near 92%) was found in the hydrolate extract displaying cryptomeridiol (23.3%, but totally absent in the EO and ME), β-eudesmol (20.6%) and α-eudesmol (10.7%). Combined analysis by chromatographic and spectroscopic techniques appeared useful for identification of various sesquiterpenols bearing a tertiary alcohol function.

The composition of an oil sample isolated from leaves of Fortunella japonica (Thunb.) Swingle was investigated by combination of chromatographic and spectroscopic techniques. Among the compounds present at appreciable amounts, a special attention was brought to sesquiterpenes bearing a tertiary alcohol function and particularly to germacra-1(10),5-dien-4α-ol. Indeed, identification of diastereoisomers α and β of germacra-1(10),5-dien-4-ol has been the subject of controversy in the literature. We report here for the first time 13C NMR data of the α isomer. The criteria for differentiation of both diastereoisomers are discussed. In parallel, leaf oils from four other Fortunella species (F. crassifolia, F. hindsii, F. margarita and F. obovata) have been analyzed by GC(RI) and 13C NMR. The main components of the investigated Fortunella leaf oils were β-elemol (8.7-30.9%), germacrene D (2.1–16.3%), β-eudesmol (4.6–13.1%) and Δ9-10-eremophilen-11-ol (1.4–11.2%). The occurrence of germacra-1(10),5-dien-4α-ol (1.1–7.7%) was observed in the five Fortunella leaf oils. Copyright © 2015 John Wiley & Sons, Ltd.

International audience; Citrus reticulata is considered one of the four main ancestral taxa of cultivated citrus and is widely associated with the mandarin horticultural group. However, several studies suggest introgressions of other taxa and the precise structure of the mandarin group is not known. To analyse the structuration of mandarin germplasm and the potential interspecific introgressions, 208 accessions were genotyped by the KASPar method, with sixty-seven single nucleotide polymorphism (SNP) markers mined in 27 nuclear genes and in clementine BAC-ends. These SNP markers were employed for the study of 208 genotypes comprised of 169 ` mandarin-like' genotypes and 39 genotypes belonging to ancestral taxa. Structure software was used to identify groups within the mandarin germplasm. Six mandarin groups were identified, some of them in agreement with the species defined by Tanaka. Many mandarins were located between these mandarin groups, indicating a high level of admixture between the identified parental groups. New insights in the organization of mandarin germplasm and its structure are presented in this work, which will be useful for better management of citrus germplasm collections.

International audience; Despite considerable differences in morphology, the genera representing 'true citrus fruit trees' are sexually compatible, but their phylogenetic relationships remain unclear. Most of the important commercial species of Citrus are believed to be of interspecific origin. By studying SNP and indel polymorphisms of 27 nuclear genes on 32 genotypes of Citrus and relatives of Citrus, the phylogenetic relationship between 'true citrus fruit trees' was clarified. Additionally, four mitochondrial (mtDNA) and five chloroplastic (cpDNA) SSRs markers were analysed for the citrus species to specify the maternal phylogeny of Citrus secondary species. A total of 16238 kb of DNA was sequenced for each genotype, and 1156 SNPs and 58 indels were identified. Nuclear phylogenetic analysis revealed that C. reticulata, Poncirus, and Fortunella form a clade that is clearly differentiated from the clade that includes three other basic taxa of cultivated citrus (C. maxima, C. medica and C. micrantha). Clymenia is included in a third strong clade with Eremocitrus and Microcitrus. The nuclear phylogeny of Citrus and its sexually compatible relatives was consistent with the geographic origins of these genera. The origin of Citrus secondary species was analysed both at nuclear and cytoplasmic level and the conclusions mostly agree with previous hypotheses. The SNPs and indels identified at the nuclear level from a relatively large discovery panel will be useful for systematic characterisation of citrus germplasm, both at intra-and inter-specific level.

The availability of a saturated genetic map of Clementine was identified by the International Citrus Genome Consortium as an essential prerequisite to assist the assembly of the reference whole genome sequence based on a Clementine derived haploid. The primary goals of the present study were to establish a Clementine reference map, and to perform comparative mapping with pummelo and sweet orange. Five parental genetic maps were established with SNPs, SSRs and InDels. A medium density reference map (961 markers for 1084.1 cM) of Clementine was established and used by the ICGC to facilitate the chromosome assembly of the haploid genome sequence. Comparative mapping with pummelo and sweet orange revealed that the linear order of markers was highly conserved. Reasonable inferences of most citrus genomes should be obtained by mapping next-generation sequencing data against the haploid reference genome sequence. Skewed segregations were frequent and higher in the male than female Clementine potentially leading to false interpretation of the genetic determinism of phenotypic traits. The mapping data confirmed that Clementine arose from hybridization between 'Mediterranean' mandarin and sweet orange and identified nine recombination break points for the sweet orange gamete that contributed to the Clementine genome. Introgression of pummelo genome fragments were identified in heterozygosity in each chromosome. Moreover, it appeared that the genome of the haploid Clementine used to establish the citrus reference genome sequence was inherited primarily from the 'Mediterranean' mandarin. The usefulness of this genetic map, anchored in the reference whole genome sequence, is discussed. Citrus Research and Development Foundation 108O568 Ministerio de Ciencia e Innovación: AGL2008-00596-MCI, 2008/121, IPT-010000-2010-43, PSE-060000-2009-8, AGL2007-65437-C04-01/AGR European Commission Citrus Research and Development Foundation itl-bio-03-10122 015453 This work was principally funded by the French ANR CITRUSSEQ project. The European Commission, under the FP6-2003-INCO-DEV-2 project CIBEWU (n°015453), the Spanish Ministerio de Ciencia e Innovación grants, AGL2007-65437-C04-01/AGR and AGL2008-00596-MCI, the Spanish PSE-060000-2009-8 and IPT-010000-2010-43 projects, the Prometeo project 2008/121 Generalidad Valenciana, the Turkish TUBITAK Project No: 108O568; the California Citrus Research Board and UC Discovery grant itl-bio-03-10122 and the Florida Citrus Research and Development Foundation (CRDF), grants #67 and 71 also contributed to the work.

Pompia is a citrus fruit endemic of Sardinia, Italy, with an essential oil profile showing outstanding anti-inflammatory and anti-microbic properties. Despite its remarkable pharmaceutical potential, little taxonomic and genetic information is available for this species. We applied flow cytometry and classical cytogenetic techniques to assess the DNA content and to reconstruct the karyotype of several Pompia accessions. Molecular data from plastid DNA barcoding and nuclear DNA sequencing were used to study the genetic distance between Pompia and other citrus species. Flow cytometric estimates of DNA content and somatic chromosome counts suggest that Pompia is a regular diploid Citrus species. DNA polymorphisms of nuclear and chloroplast markers allowed us to investigate the genetic relationships between Pompia accessions and other Citrus species. Based on DNA polymorphism data we propose that Pompia is a very recent interspecific hybrid generated by a cross between C. aurantium (as seed bearer) and C. medica (as pollen donor). Our findings pave the way for further and more specific investigations of local Pompia germplasm resources that may help the preservation and valorisation of this valuable citrus fruit tree.

International audience; To understand the genotypic variation of citrus to mild salt stress, a proteomic approach has been carried out in parallel on two citrus genotypes ('Cleopatra' and 'Willow leaf' mandarins), which differ for Na+ and Cl- accumulation, and their cognate autotetraploids (4x). Using two-dimensional electrophoresis approximately 910 protein spots were reproducibly detected in control and salt-stressed leaves of all genotypes. Among them, 44 protein spots showing significant variations at least in one genotype were subjected to mass spectrometry analysis for identification. Salt-responsive proteins were involved in several functions, including photosynthetic processes, ROS scavenging, stress defence, and signalling. Genotype factors affect the salt-responsive pattern, especially that of carbon metabolism. The no ion accumulator 'Cleopatra' mandarin genotype showed the highest number of salt-responsive proteins, and up-regulation of Calvin cycle-related proteins. Conversely the ion accumulator 'Willow leaf' mandarin showed high levels of several photorespiration-related enzymes. A common set of proteins (twelve spots) displayed higher levels in salt-stressed leaves of 2x and 4x 'Cleopatra' and 4x 'Willow leaf' mandarin. Interestingly, antioxidant enzymes and heat shock proteins showed higher constitutive levels in 4x 'Cleopatra' mandarin and 4x 'Willow leaf' mandarin compared with the cognate 2x genotype. This work provides for the first time information on the effect of 8 weeks of salt stress on citrus genotypes contrasting for ion accumulation and their cognate autotetraploids. Results underline that genetic factors have a predominant effect on the salt response, although a common stress response independent from genotype was also found. (C) 2013 Elsevier GmbH. All rights reserved.

Trees; International audience; Citrus plants originate from southeastern Asia, in a large area with various climates characterized by a broad range of temperatures. Some species have been diversified in temperate climates, others in subtropical climates. Temperature is assumed to be a key factor in citrus species adaptation and diversification of basic cellular functions. In a field experiment, the tolerance of the three fundamental Citrus species C. medica L., C. reticulata Blanco and C. maxima (Burm.) Merr., and Fortunella japonica (Thunb.) Swing. to photooxidative stress caused by seasonal climatic changes was evaluated on adult trees by measuring net photosynthesis (Pnet), stomatal conductance (Gs), maximum photosynthesis (Pmax) and chlorophyll fluorescence (Fv/Fm). In addition, seasonal changes in oxidative status, antioxidant enzymes (superoxide dismutase, catalase, ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase and glutathione reductase) and antioxidant metabolites (ascorbate and glutathione) were monitored. Mandarin and pummelo appeared to be the most tolerant, showing the lowest down-regulation of photosynthetic parameters, and the lowest accumulation of oxidized compounds associated with efficiency of their antioxidant system. Kumquat showed intermediate behaviour, with a large diminution of photosynthetic parameters and marked accumulation of hydrogen peroxide, whereas the malondialdehyde content remained low, with a strong induction of glutathione synthesis. Finally, citron appeared to be the most sensitive genotype with a marked decrease in photosynthetic performance, the largest accumulation of oxidative parameters, insufficient induction of antioxidant enzymes and down-regulation of ascorbate and glutathione synthesis.

Resume Apparu lors d'un croisement naturel entre un mandarinier et un oranger a la fin du XIXe siecle, la clementine s'est ensuite diversifiee par selection clonale de mutants somatiques. Distinguer les differentes varietes clonales etant pratiquement impossible, le developpement d'outils moleculaires d'identification varietale s'est donc avere necessaire. Nous avons etudie trois cultivars, representant des groupes de maturite des fruits tres distincts. Nous avons recherche des criteres d'identification au niveau phenotypique (caracteristiques organoleptiques, morphologie des feuilles) et au niveau de l'ADN (isozymes, RAPD et ISSR). La diversite phenotypique observee est relativement elevee et contraste avec le faible polymorphisme moleculaire. En effet, seul le cultivar « Guerdane » presente des profils d'empreintes genetiques differents de ceux des deux autres cultivars. La frequence des modifications genetiques serait donc variable d'un cultivar a un autre. Les marqueurs moleculaires specifiques du cultivar « Guerdane » s'ajoutent aux marqueurs phenotypiques et etendent les possibilites d'identification aux jeunes plants en pepiniere. Pour citer cet article : B. Chahidi et al., C. R. Biologies 331 (2008).

Essential oils of 113 Citrus clementina (Commun) × Citrus deliciosa (Willow Leaf) hybrids were obtained from leaves collected on trees located in the same orchard. Their chemical composition was investigated by capillary GC, GC–MS and 13C-NMR and the results were submitted to statistical analysis. Three chemical compositions were characterized. The crossing between clementine and mandarin produces mainly leaf oils characterized by a composition similar to those of the parents (sabinene/linalool or methyl N-methylanthranylate). However, several samples exhibited an original composition (γ-terpinene/linalool). Copyright © 2008 John Wiley & Sons, Ltd.

The volatile compounds from leaves and peels of an interspecific citrus somatic hybrid, Citrus aurantifolia (Christm.) Swing. + Citrus paradisi Macfayden, obtained by fusion of protoplasts from lime, Citrus aurantifolia (cv. Mexican Lime) and grapefruit, Citrus paradisi (cv. Star Ruby), were extracted by pentane: ether (1 : 1) from liquid nitrogen ball-milled leaves and flavedo and examined by GC-MS in comparison to those of its parents. The hybrid quantitatively retained the ability of the lime parent to synthesize in its leaves the major monoterpene aldehydes (neral, geranial) the monoterpene alcohols (nerol, geraniol), and their acetates, and also the capacity of the grapefruit parent to produce a sesquiterpene aldehyde ([beta]-sinensal) in its leaves and nootkatone in its peel. Conversely, synthesis of most sesquiterpene hydrocarbons and long chain aliphatic aldehydes, which are present in the lime parent leaves and peel, was strongly inhibited in the hybrid, as in the grapefruit parent. In comparison to its parents, the hybrid overproduced citronellal in its leaves and [alpha]-sinensal and [beta]-sinensal in its peel. Based on these results, the future prospects for a better understanding of the inheritance mechanisms with regards to aroma biosynthesis in citrus leaves and peels are discussed. (Resume d'auteur)

The chemical composition of the essential oil (EO), microwave extract (ME) and hydrolate extract (HE) from the same batch of leaves of Fortunella japonica, was investigated by combination of chromatographic (GC, CC) and spectroscopic techniques (GC-MS, 13C NMR). F. japonica essential oil and extracts are complex mixtures of 28–60 compounds being mainly oxygenated sesquiterpenes. The EO composition was dominated by germacrene D (14.9%), β-elemol (9.1%), cis-guai-6-en-10p-ol (6.3%), β-eudesmol (5.5%), and δ-elemene (5.2%). Limonene was the unique monoterpene identified at appreciable amount (7.1%). The extract obtained by microwave assisted hydrodistillation contained as main components: β-elemol (12.4%), germacrene D (9.9%), cis-guai-6-en-10β-ol (9.0%), β-eudesmol (8.2%), germacra-1(10),5-dien-4α-ol (7.1%) and α-eudesmol (6.4%). Finally, the highest content of oxygenated sesquiterpenes (near 92%) was found in the hydrolate extract displaying cryptomeridiol (23.3%, but totally absent in the EO and ME), β-eudesmol (20.6%) and α-eudesmol (10.7%). Combined analysis by chromatographic and spectroscopic techniques appeared useful for identification of various sesquiterpenols bearing a tertiary alcohol function.

International audience; Different physiological behavior of a wide range of varieties and species belonging to the Citrus genus was analyzed when subjected to salt stress with the aim to seek new sources of tolerance that might be specie-specific. Our goal was to use physiological results obtained along a salt stress in order to clarify if it would be possible to associate them with the known citrus genetic diversity. For that purpose, we have selected 20 different genotypes representing the major species on the basis of the genetic diversity of Citrus genus complemented with one intergeneric hybrid Carrizo citrange (C. sinensis 9 P. trifoliata). A moderate salt stress of 75 mM of NaCl was applied for 12 weeks. For control plants, the main parameters contributing for more than 25 % to the diversity on the two axes of principal component analysis (PCA) were chlorophyll content, photosynthesis and Fv/ Fm under light. However, the dispersal of species and varieties on the PCA did not show any particular structure. Under salt stress condition, four parameters (leaf chloride content, leaf chlorophyll content, photosynthesis and stomatal conductance) contributed more specifically to the dispersion on PCA representation with more than 15 % of contribution for each parameter. Large differences were observed within citrus genus: mandarin and pummelo presented good tolerance to salt stress while citron was very sensitive. Furthermore, all secondary genotypes that presented good tolerance to salt tolerance shared mandarin or pummelo as female parent.

International audience; Fruit oil from five species of kumquat (Fortunella japonica, F. margarita, F. crassifolia, F. obovata, and F. hindsii) grown in the same pedoclimatic conditions have been analyzed by a combination of chromatographic and spectroscopic techniques. The compositions of the five fruit oils were strongly dominated by limonene (84.2-96.3%). Other components present with appreciable contents were myrcene (1.3-12.9%) and germacrene D (0.3-2.4%).

International audience; A dinucleotide-enriched genomic library was obtained from mandarin orange (Citrus reticulata Blanco). A subset of 101 positive clones was sequenced and primers were designed. The loci were screened for levels of variation using 26-29 wild mandarin oranges collected in Vietnam. Forty-three loci were polymorphic with the number of alleles ranging from two to 18. The observed heterozygosity (H-O) and expected heterozygosity (H-E) were from 0.03 to 0.96 and from 0.03 to 0.92, respectively.

Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; Citrus plants are currently facing biotic and abiotic stresses. Therefore, the characterization of molecular traits involved in the response mechanisms to stress could facilitate selection of resistant varieties. Although large cDNA microarray profiling has been generated in citrus tissues, the available protein expression data are scarce, in this study, to identify differentially expressed proteins in Citrus clementina leaves after infestation by the two-spotted spider mite Tetranychus urticae, a proteome comparison was undertaken using two-dimensional gel electrophoresis. The citrus leaf proteome profile was also compared with that of leaves treated over 0-72 h with methyl jasmonate, a compound playing a key role in the defense mechanisms of plants to insect/arthropod attack. Significant variations were observed for 110 protein spots after spider mite infestation and 67 protein spots after MeJA treatments. Of these, 50 proteins were successfully identified by liquid chromatography-mass spectrometry-tandem mass spectrometry. The majority constituted photosynthesis- and metabolism-related proteins. Five were oxidative stress associated enzymes, including phospholipid glutathione peroxidase, a salt stressed associated protein, ascorbate peroxidase and Mn-superoxide dismutase. Seven were defense-related proteins, such as the pathogenesis-related acidic chitinase, the protease inhibitor miraculin-like protein, and a lectin-like protein. This is the first report of differentially regulated proteins after T. urticae attack and exogenous MeJA application in citrus leaves. (C) 2010 Elsevier GmbH. All rights reserved.

International audience; Most of the studies on organic acids and sugars in citrus were performed during fruit maturation, and less is known before this stage of development. The aim of our study was to investigate acids and sugars in lemon, lime, and orange from fruit-set toward development. We chose to compare organic acid and sugar accumulation among acidic and acidless varieties within three species. We estimated the acidity by titrimetry and quantified the concentrations of seven organic acids and three sugars by reverse HPLC. During the first 50 days of development, quinic acid was the major organic acid whatever the variety. Afterward, citric acid predominated in acidic varieties, while in acidless, malic acid exceeded it. Fructose substituted citric acid in acidless and could be synthesized either from citric acid or directly from glucose. Our results provided the first complete report on sugar and organic acid accumulation during the early stages of fruit development in several citrus varieties.

Publication Inra prise en compte dans l'analyse bibliométrique des publications scientifiques mondiales sur les Fruits, les Légumes et la Pomme de terre. Période 2000-2012. http://prodinra.inra.fr/record/256699; International audience; Citrus fruits are complex sources of carotenoids with more than 100 kinds of pigments reported in this genus. To understand the origin of the diversity of carotenoid compositions of citrus fruit, 25 genotypes that belong to the 8 cultivated Citrus species were analyzed. Juice extracts of mature fruit were analyzed by high-performance liquid chromatography using a C-30 column. The 25 citrus genotypes presented different carotenoid profiles with 25 distinct compounds isolated. Statistical analyses revealed a strong impact of genotype on carotenoid compositions. Two kinds of classifications of genotypes were performed: on qualitative data and on quantitative data, respectively. The results showed that variability in carotenoid compositions was more interspecific than intraspecific. Two carotenoids, cis-violaxanthin and the beta-cryptoxanthin, strongly determined the classification on qualitative data, which was also in agreement with previous citrus variety classifications. These findings provide evidence that, as for other phenotypical traits, the general evolution of cultivated Citrus is the main factor of the organization of carotenoid diversity among citrus varieties. To the authors' knowledge this is the first study that links the diversity of carotenoid composition to the citrus genetic diversity. These results lead to the proposed major biosynthetic steps involved in the differential carotenoid accumulation. Possible regulation mechanisms are also discussed.

International audience; Volatile compounds were extracted by a pentane/ether (1:1) mixture from the leaves of six citrus somatic allotetraploid hybrids resulting from various combinations of lime, lemon, citron, sweet orange, and grapefruit. Extracts were examined by gas chromatography-mass spectrometry (GC-MS) and compared with those of their respective parents. All hybrids having an acid citrus parent exhibit the same relative contents in hydrocarbons and oxygenated compounds as the acid citrus, while the (grapefruit + orange) hybrid behaves similarly to its two parents. When volatile compound contents (mu g g(-1)) are examined in detail, several behaviors are encountered in hybrids and seem to depend on the presence/absence of the considered parental compound and on the corresponding hybrid combination. Meanwhile, the sesquiterpene hydrocarbons are present in all hybrids at concentrations systematically lower than those of the highest parental producers. Statistical analyses show that hybrids exhibit hardly discriminable aromatic profiles, meaning that no strong dominance of one or the other parent was observed in hybrids with regards to the leaf volatile compound production.

Phlorin (3,5-dihydroxyphenyl beta-D-glucopyranoside), an orange peel marker, has been searched in 45 species and varieties of Citrus. The phlorin content was determined by high-pressure liquid chromatography in juices and aqueous peel extracts of these different fruits. The phlorin content in C. reticulata peel extract varies from 0 to 1012 mg L(-)(1) with a mean of 162 mg L(-)(1). On the contrary, phlorin was not found in mandarin and clementine juices except for mandarin "Commune" and "Beauty" (33 and 30 mg L(-)(1), respectively). In the 14 species of oranges and varieties, phlorin was detected in juices and peel extracts with a mean of 22 and 492 mg L(-)(1), respectively, while for grapefruits, means were 108 mg L(-)(1) in juices and 982 mg L(-)(1) for peel extracts. Tangors and tangelos which are hybrids (C. reticulata x C. sinensis and C. reticulata x C. paradisi, respectively) are characterized by the systematic presence of phlorin in peels (mean: 406 and 659 mg L(-)(1), respectively) while in juices its presence could be variable (0-73 mg L(-)(1)). These heterogeneity and values may be explained by the genetic variability of these hybrids and the phlorin content of their parentage group.

In France, cryopreservation is currently used for a limited number of forest tree species, but has been applied yet very little to horticultural or other agricultural species. The CRYOVEG project aims at (1) developing or optimizing cryopreservation techniques in a range of selected species; and (2) establishing a national scientific and technical network of plant biological resource centers (CRBs) using cryopreservation. The project has a network organization, with IRD/INRA Montpellier as the cryopreservation expertise centre and partners in continental France and in overseas departments in charge of genetic resource conservation for various species: INRA Petit Bourg, Guadeloupe (yam); INRA San Giuliano, Corsica (Citrus); INRA Bordeaux (Prunus); INRA Angers (apple and pear); INRA Montpellier (grapevine); INRA Ploudaniel (potato, Brassica); IRD La Reunion (coffee); CIRAD Roujol, Guadeloupe (sugarcane); and CIRAD La Reunion (vanilla, garlic). Deliverables will vary depending on the state-of-the-art of cryopreservation for each species: for potato, yam, sugarcane, garlic, Citrus, Brassica and coffee, optimized protocols will be tested on a range of representative accessions. For grapevine, Prunus, apple, pear and vanilla, operational protocols will be developed. The group will work as a think-tank on the integration of cryopreservation in the national plant genetic resource conservation strategy and an action plan for its implementation will be collectively designed. The project will work towards establishing a French research network on plant genetic resource cryopreservation, open to all partners interested in cryopreservation. The group will prepare research and development projects whose results will be disseminated through European plant genetic resource conservation networks e.g. ECP/ GR). Finally, network members will train French and foreign (from North and South countries) researchers and technicians in cryopreservation techniques. (Conflicts of interest: None declared. Source of funding: This project is supported by a French competitive grant (Call for Projects 2008 ''Biological Resource Centers") of the Consultative Committee for Biological Resources/Infrastructures in the Biology, Health and Agronomy sectors (CCRB/IBiSA).). (Texte integral)