Jean-Paul Charpentier

Natural wood colouring process in Juglans sp. (J. nigra, J. regia and hybrid J. nigra 23 ×J. regia) depends on native phenolic compounds accumulated in the transition zone between sapwood and heartwood

Abstract Radial distribution of soluble phenolics was investigated at different heights in stems of Juglans nigra, J. regia and hybrids J. nigra 23 × J. regia. Four major phenolic compounds were studied: hydrojuglone glucoside (HJG), quercitrin (QUER) and two unknown compounds characterized as two ellagic acid derivatives E1 and E2. HJG and E1 content increased gradually in the sapwood, peaked in the sapwood-heartwood transition zone, and decreased drastically in the heartwood. QUER was accumulated preferentially around the transition zone, and its content was relatively low in the outer part of the sapwood and in the inner part of the heartwood. E2 content was low in the sapwood and increased in the heartwood. The heartwood formation was marked by the accumulation of new soluble compounds. The relationship between wood extractives and wood colour were evaluated and discussed. HJG was considered to be a major precursor of heartwood colour providing chromophores through hydrolysis (deglucosylation), oxidation and polymerization processes.

Expression of antisense chalcone synthase RNA in transgenic hybrid walnut microcuttings. Effect on flavonoid content and rooting ability

Walnut somatic embryos (Juglans nigra × Juglans regia) were transformed with a vector containing a neomycin phosphotransferase II, a β-glucuronidase and an antisense chalcone synthase (chs) gene. This antisense construct included a 400 bp cDNA fragment of a walnut chs gene under the control of the duplicated CaMV-35S promoter. Molecular, biochemical and biological characterizations were performed both on transformed embryos propagated by secondary somatic embryogenesis and on microshoots developed by in vitro culture of embryonic epicotyls from somatic embryos. Thirteen transformed lines with the vector containing the antisense chs gene, one line with only the gus and nptII genes and one untransformed line were maintained in tissue culture. Six of the antisense lines were shown to be flavonoid-deficient. They exhibited a strongly reduced expression of chs genes, very low chalcone synthase activity and no detectable amounts of quercitrin, myricitrin, flavane-3-ols and proanthocyanidins in stems. Rooting tests showed that decreased flavonoid content in stems of antisense chs transformed lines was associated with enhanced adventitious root formation. Free auxin and conjugated auxin contents were determined during the latter phase of the micropropagation, and no variations were detected between control and antisense chs transformed lines. The in vitro plants developed a large basal callus and apical necrosis upon auxinic induction and the transformed lines highly deficient in flavonoids were more sensitive to exogenous application of indolebutyric acid (IBA).

Comprehensive genetic dissection of wood properties in a widely-grown tropical tree: Eucalyptus

BackgroundEucalyptus is an important genus in industrial plantations throughout the world and is grown for use as timber, pulp, paper and charcoal. Several breeding programmes have been launched worldwide to concomitantly improve growth performance and wood properties (WPs). In this study, an interspecific cross between Eucalyptus urophylla and E. grandis was used to identify major genomic regions (Quantitative Trait Loci, QTL) controlling the variability of WPs.ResultsLinkage maps were generated for both parent species. A total of 117 QTLs were detected for a series of wood and end-use related traits, including chemical, technological, physical, mechanical and anatomical properties. The QTLs were mainly clustered into five linkage groups. In terms of distribution of QTL effects, our result agrees with the typical L-shape reported in most QTL studies, i.e. most WP QTLs had limited effects and only a few (13) had major effects (phenotypic variance explained > 15%). The co-locations of QTLs for different WPs as well as QTLs and candidate genes are discussed in terms of phenotypic correlations between traits, and of the function of the candidate genes. The major wood property QTL harbours a gene encoding a Cinnamoyl CoA reductase (CCR), a structural enzyme of the monolignol-specific biosynthesis pathway.ConclusionsGiven the number of traits analysed, this study provides a comprehensive understanding of the genetic architecture of wood properties in this Eucalyptus full-sib pedigree. At the dawn of Eucalyptus genome sequence, it will provide a framework to identify the nature of genes underlying these important quantitative traits.

Mechanisms of primordium formation during adventitious root development from walnut cotyledon explants

Abstract. In walnut (Juglans regia L.), an otherwise difficult-to-root species, explants of cotyledons have been shown to generate complete roots in the absence of exogenous growth regulators. In the present study, this process of root formation was shown to follow a pattern of adventitious, rather than primary or lateral, ontogeny: (i) the arrangement of vascular bundles in the region of root formation was of the petiole type; (ii) a typical root primordium was formed at the side of the procambium within a meristematic ring of actively dividing cells located around each vascular bundle; (iii) the developing root apical meristem was connected in a lateral way with the vascular bundle of the petiole. This adventitious root formation occurred in three main stages of cell division, primordium formation and organization of apical meristem. These stages were characterized by expression of LATERAL ROOT PRIMORDIUM-1 and CHALCONE SYNTHASE genes, which were found to be sequentially expressed during the formation of the primordium. Activation of genes related to root cell differentiation started at the early stage of primordium formation prior to organization of the root apical meristem. The systematic development of adventitious root primordia at a precise site gave indications on the positional and biochemical cues that are necessary for adventitious root formation.

Modifications of hybrid Walnut (Juglans nigra 23 x Juglans regia) wood colour and phenolic composition under various steaming conditions

Summary The effects of steaming were studied on Walnut wood from a hybrid (Juglans nigra 23 × J. regia) tree in terms of wood colour and phenolic composition. Wood samples were subjected to treatments at 75, 100 and 125°C for 4, 8, 16 and 24 hours. Colour changes were measured in the CIELCh colour system and phenolics were characterized and quantified by means of HPLC. Steaming mainly resulted in a darkening of wood tissues. Colour changes increased as temperature increased from 75 to 125°C. Lightness L* gradually decreased from 0 to 24 h at 75°C, whereas with higher temperatures, most of the darkening occurred within the first 4 hours of exposure. Contrast in colour, between sapwood and heartwood in particular, could be reduced by steaming at 125°C. The best steaming treatment, leading to a colour of steamed sapwood close to that of natural heartwood, was at 100°C/16 h. Hydrojuglone glucoside (HJG), ellagic and gallic acid derivatives (E1, E2 and G respectively) were gradually degraded by steaming, whereas a new flavonol (F) and oxidation products (OP) increased. Relationships between wood colour and phenolic content are discussed.

Histological investigation of walnut cotyledon fragments for a better understanding of in vitro adventitious root initiation

SummaryRapid formation of adventitious roots by walnut cotyledon fragments in vitro was traced by light microscopy. It was shown that this plant model is characterized by two major developmental processes: a) confined elongation of the cotyledon petiole caused by a limited number of cell divisions and b) formation of a morphogenetic zone around each initially wounded vascular bundle within 36 h after detachment of the embryonic axis. During the first phase of development, granular storage protein bodies dissolved, and starch grains were deposited mainly in the distal portion of the cotyledon fragments. Rapidly, new globular protein bodies were formed, and phenolic inclusions accumulated in the vacuoles of epidermal and subepidermal cells and of individual cells close to the vascular bundles. Each adventitious root was found to be in continuity with a single vascular bundle of the cotyledon petiole. A short auxin treatment suppressed the formation of large roots and induced numerous tiny rootlets dispersed all over the surface of the cotyledons.

Durability of five native Argentine wood species of the genera Prosopis and Acacia decayed by rot fungi and its relationship with extractive content.

The natural durability of four Argentinean species of Prosopis and one of Acacia was evaluated in laboratory tests, according to European standards, using three brown rot and one white rot fungi. These tests were complemented by assessing the wood chemical composition. All the species were from moderately slightly durable to very durable (classes 4–1), and in all cases the heartwood was the most resistant to fungal attack. Chemical extractives content (organic, aqueous, tannic and phenolic) was higher in the heartwood. However, species durability was not related to extractive contents nor with wood density. Instead, it is possible that extractives could contribute to natural durability in different ways, including the effects related to the antioxidant properties of some of them.

Correction of moisture effects on near infrared calibration for the analysis of phenol content in eucalyptus wood extracts

Abstract• Methods based on near infrared spectroscopy used to assess wood properties are susceptible to variations in physical parameters (temperature, grain size, etc.). As wood is a hygroscopically sensitive material, we studied the effects of moisture on near infrared absorbance and calibration to accurately determine the application potential of this technique under routine.• A collection of Eucalyptus urophylla × E. grandis hybrid wood pieces were analysed to obtain reference calibration of polyphenol contents in wood extracts via NIR spectra acquired under constant moisture conditions. Other specimens from the same source were assessed to obtain spectra for eight moisture contents spanning a broad variation range. The effects of moisture on absorption and on estimates based on a reference model were analysed.• An increase in moisture content prompted a rise in near infrared absorption over the entire spectrum and for water O-H absorption bands. The polyphenol content estimates obtained by assessing specimens against the reference calibration at variable moisture contents revealed prediction bias. Five correction methods were then tested to enhance the robustness relative to moisture.• In-depth calibration and external parameter orthogonalization (EPO) were found to be the most efficient methods for offsetting this factor.Résumé• Les méthodes basées sur la spectroscopie proche infrarouge pour estimer les propriétés du bois sont sensibles aux variations des paramètres physiques (température, granulométrie…). Le bois étant un matériau hygroscopique sensible, l’influence de l’humidité sur l’absorbance et l’étalonnage proche infrarouge a été étudiée afin de mieux considérer les possibilités d’applications dans des conditions réelles.• Un étalonnage de référence de la quantité de polyphénols présents dans les extraits a été établi à partir de spectres d’une collection de bois d’hybrides d’Eucalyptus urophylla × E. grandis à humidité constante fixée. D’autres spectres ont été obtenus sur des échantillons de même provenance mais à 8 teneurs en eau couvrant une large plage de variation. L’influence de l’humidité sur l’absorbance proche infrarouge puis sur l’estimation par le modèle de référence a été analysée.• Une augmentation de l’humidité élève la ligne de base du spectre d’absorbance et également les régions d’absorbance caractéristiques de la liaison O-H de l’eau. Les estimations de la quantité de polyphénols à partir de l’étalonnage de référence sur les échantillons à humidité variable révèlent un biais sur les prédictions.• Parmi les cinq méthodes de correction testées pour rendre l’étalonnage robuste vis-à-vis de l’humidité, l’étalonnage exhaustif et l’EPO (External Parameter Orthogonalisation) s’avèrent être les plus efficaces et permettent de s’affranchir de ce facteur.

Histological and Biochemical Characterization of Adventitious Root Formation in Walnut Cotyledon Fragments

Studies conducted on the plant model Arabidopsis thaliana demonstrate that primary and lateral root meristem formation are controlled by different factors including the number, nature and environment of the original cells as well as hormonal and genetic controls (Aeschbacher et al., 1994, Celenza et al., 1995). The primary root meristem derives from one cell of the embryo and the hypophysis whereas the lateral root meristem rises from pericycle cells of primary root system. Adventitious root formation (ARF) is a mul- tistep developmental process in which competent cells are induced to form a root meristem (Mohnen, 1994). Thus, it can be hypothesized that adventitious root meristem (ARM) formation has its own characteristics. This specificity could reside in 1) the induction process since ARM can be formed from differenciated cells of variable origins (Chriqui, 1985) and 2) the ARM organization process which depends on the cellular neighbouring of the pre-meristematic cells.

Prediction of stilbene content from heartwood increment cores of Scots pine using near infrared spectroscopy methodology

High-throughput and non-destructive methods for quantifying the content of the stilbene compounds of Scots pine (Pinus sylvestris L.) heartwood are needed in the breeding for decay resistance of heartwood timber. In this study, near infrared (NIR) spectroscopy calibrations were developed for a large collection of solid heartwood increment core samples in order to predict the amount of the stilbene pinosylvin (PS), its monomethyl ether (PSM) and their sum (STB). The resulting models presented quite accurate predictions in an independent validation set with R2V values ranging between 0.79 and 0.91. The accuracy of the models strongly depended on the chemical being calibrated, with the lowest accuracy for PS, intermediate accuracy for PSM and highest accuracy for STB. The effect of collecting one, two or more (up to five) spectra per sample on the calibration models was studied and it was found that averaging multiple spectra yielded better accuracy as it may account for the heterogeneity of wood along the increment core within and between rings. Several statistical pretreatments of the spectra were tested and an automatic selection of wavenumbers prior to calibration. Without the automatic selection of wavenumbers, a first derivative of normalised spectra yielded the best accuracies, whereas after the automatic selection of wavenumbers, no particular statistical pretreatment appeared to yield better results than any other. Finally, the automatic selection of wavenumbers slightly improved the accuracy of the models for all traits. These results demonstrate the potential of NIR spectroscopy as a high-throughput and non-destructive phenotyping technique in tree breeding for the improvement of decay resistance in heartwood timber.