Grandis Trees Research Articles

Organ-specific rest and quiescence in response to chilling and forcing in subtropical Torreya grandis trees

Organ-specific rest and quiescence in response to chilling and forcing in subtropical Torreya grandis trees

Integrated Metabolomics, Transcriptome and Functional Analysis Reveal Key Genes Are Involved in Tree Age-Induced Amino Acid Accumulation in Torreya grandis Nuts

Torreya grandis is native Chinese tree species of economic significance, renowned for its long lifespan and the rich nutritional value of its nuts. In this study, we analyzed the morphological characteristics, metabolites, associated gene expressions, and regulatory mechanism in nuts from young (10 years old) and old (1000 years old) T. grandis trees. We observed that the length, width, and weight of nuts from older trees were considerably greater than those from younger trees. Metabolomic analysis revealed that the concentrations of 18 amino acids and derivatives (including histidine and serine) in nuts from older trees were markedly higher than those in nuts from younger trees. Transcriptome and metabolomic correlation analysis identified 16 genes, including TgPK (pyruvate kinase), TgGAPDH (glyceraldehyde 3-phosphate dehydrogenase), and others, which exhibit higher expression levels in older trees compared to younger trees, as confirmed by qRT-PCR. These genes are associated with the biosynthesis of histidine, glutamic acid, tryptophan, and serine. Transient expression of TgPK in tobacco led to increased pyruvate kinase activity and amino acid content (histidine, tryptophan, and serine). Additionally, dual-luciferase assays and yeast one-hybrid results demonstrated that TgWRKY21 positively regulates TgPK expression by directly binding to the TgPK promoter. These findings not only demonstrate the nutritional differences between nuts from young and old trees but also offer fresh insights into the development of nutritional sources and functional components based on nuts from old trees, enriching our understanding of the potential benefits of utilizing nuts from older trees.

Early and long-distance uptake by Eucalyptus grandis of N, K and Ca tracers injected down to a depth of 7 m

Eucalyptus plantations are commonly fertilized at planting to sustain high biomass production over successive rotations in nutrient-poor soils. However, the dynamics of nutrient uptake by tree roots in the early stages of stand development, depending on the lateral distance from the trees and the depth in the soil, are poorly documented. We assessed the potential uptake of N, K and Ca, from 6 to 18 months after planting, by Eucalyptus grandis trees depending on: i) the lateral distance from the tracer injection point and ii) the depth in the soil. At 6 months after planting, we injected 15NO3–, Rb+, Sr2+ and Cs+ tracers at depths of 1, 3, 5, and 7 m, respectively. Recently expanded leaves were then sampled monthly on 16 target E. grandis trees located at lateral distances ranging from 0.75 to 11.1 m from the injection points, and on 4 more distant E. grandis control trees; 15N contents (atom %) and concentrations of Rb, Cs and Sr in the leaves were determined at each sampling date, and compared to those of control trees to detect the dates of tracer uptake for each individual. A LOESS model was applied to account for the temporal trend and inter-tree variability among the control trees. Tracer uptake by a target tree was detected if the measurements exceeded the upper limit of the prediction interval (α = 0.005) generated by the model. Only the 6 target trees located within 2.0 m from the injection point took up the 15NO3– tracer. However, Rb+, Sr2+ and Cs+ tracers were taken up to lateral distances of approx. 11 m from the injection point. Tracer uptake occurred as early as 9, 7, 8 and 9 months after planting for 15NO3–, Rb+, Sr2+ and Cs+, respectively. Four trees located within 2.8 m from the injection point took up the 3 cation tracers, injected between the depths of 3 m and 7 m. All but one of the target trees took up at least one cation tracer. The potential of Eucalyptus trees to take up nutrients early, at long distance and at depth, can be promoted by silvicultural practices enhancing rapid root development after planting, as subsoiling in commercial plantations and large planting holes in smallholder plantations. Our results also suggest that fertilization could be applied only once at planting in Eucalyptus stands established on deep tropical soils, with low risks of nutrient losses by deep drainage.

Influence of knots on the adhesion of wood from young Eucalyptus grandis plantations

ABSTRACT The structural use of wood from young trees requires gluing smaller timber pieces to form large elements. In young Eucalyptus timber of structural dimensions, knots do not significantly diminish the mechanical properties, yet their influence on wood adhesion has not been widely studied. Therefore, the aim of the study was to assess the influence of knots on the bond quality of face-glued wood from young E. grandis. Wood from seven-year-old E. grandis trees was tested in shear on the bond-line (polyurethane adhesive) using paired samples of knot-containing and clear wood samples. Knot features did not correlate with shear strength or wood failure percentage. Knot-containing samples had a lower percentage of wood failure (5.8%), but the same shear strength as clear wood samples. Below a wood density of 650 kg m−3, clear and knot-containing wood behaved similarly, but above this value, knots influenced wood adhesion negatively. By simulating glulam beams with high-density wood on the outer lamellas, it was possible to show that shear stress on those bond-lines is lower than the minimum shear strength observed on dense knot samples. Hence, lower adhesion caused by knots on denser wood probably does not represent a significant problem for glued laminated products from young Eucalyptus grandis trees.

Field Drying for Enhancing Biomass Quality of Eucalyptus Logs and Trees in Florida, USA

Field drying Eucalyptus biomass can minimize the storage requirements, transportation costs, and/or the cost associated with biomass drying for biochar and other products. In central Florida, 12 6-year-old and 12 8-year-old Eucalyptus amplifolia and Eucalyptus grandis trees were field dried over two seasons, with two months of field drying in each season, and two tree forms (logs, whole tree). The whole tree average initial moisture content (MCod) on a dry basis ranged between 115 and 121% in E. amplifolia and 116 and 119% in E. grandis. The season and duration of field drying affected drying, but species, tree size, and tree form did not. In southern Florida, 19 3+-year-old coppice stems of an E. grandis × Eucalyptus urophylla hybrid clone (EH1) were field dried over two months of one season, with and without tree tops. The whole tree initial MCod ranged between 119 and 138%. The duration of field drying affected drying, but tree size and tree tops did not. Both studies suggest that field drying can effectively reduce wood MC and thus enhance E. amplifolia, E. grandis, and EH1 biomass quality.

Calonectria queenslandica: Causal Agent of Eucalyptus Leaf Blight in Southern China.

Calonectria leaf blight caused by Calonectria spp. is among the most serious diseases affecting the health and sustainability of Eucalyptus plantations in southern China. Recent outbreaks of this disease in GuangDong Province prompted a need to identify the species involved. Typical symptoms of Calonectria leaf blight were observed on 2-year-old Eucalyptus urophylla × E. grandis trees in a plantation in the ZhaoQing region. In total, 38 Calonectria isolates were collected from 32 diseased trees. All isolates were identified using DNA sequence analyses of the translation elongation factor 1-α (tef1), β-tubulin (tub2), calmodulin (cmdA), and histone H3 (his3) gene regions. Phylogenetic analyses revealed that Calonectria queenslandica was the dominant species, accounting for 81.6% of the isolates collected. Other species isolated included C. pseudoreteaudii (10.5%), C. reteaudii (5.3%), and C. aconidialis (2.6%). This is the first report of C. queenslandica in China and all isolates had identical sequences in all four gene regions. PCR amplification using primers targeting the MAT1-1-1 and MAT1-2-1 genes in all C. queenslandica isolates revealed that only the MAT1-2 idiomorph was present. The results suggest that C. queenslandica was introduced into the sampled area with very limited genetic diversity. Pathogenicity tests were conducted on two Eucalyptus genotypes widely planted in the GuangDong Province using isolates representing all species collected. The results showed that these species could all cause disease but the predominance of C. queenslandica on infected trees suggests that it is the major driver of the disease problem studied. Different Eucalyptus genotypes used in the pathogenicity tests differed in susceptibility to infection by the Calonectria spp. tested, providing opportunities to avoid leaf blight by deploying disease-tolerant planting stock.

Wood density prediction using near-infrared hyperspectral imaging for early selection of Eucalyptus grandis trees

Wood density prediction using near-infrared hyperspectral imaging for early selection of Eucalyptus grandis trees

Comparison of growth and few wood quality parameters of 24–25-year-old Tectona grandis (teak) trees raised under three agroforestry practices

Comparison of growth and few wood quality parameters of 24–25-year-old Tectona grandis (teak) trees raised under three agroforestry practices

First Report of Powdery Mildew in Eucalyptus Caused by Erysiphe neolycopersici in China.

Eucalyptus is one of the most fast-growing and widely planted hardwood trees in the tropical and subtropical regions (Grattapaglia and Kirst, 2008). In December 2021, powdery mildew diseases were observed on the Eucalyptus urophylla, E. urophylla × E. grandis, E. grandis × E. urophylla, and E. grandis trees growing in the Eucalyptus garden of the Guangxi University campus in Nanning (108°22'E, 22°48'N) of Guangxi Zhuang Autonomous Region, where is the main plantation area for Eucalyptus. The spread of this disease would bring potential challenges on the Eucalyptus plantation management in this region of China. The early symptoms of this disease in Eucalyptus were that the irregular white spots with surface-attached powder was observed on the leaves. At the late stages, this symptom was diffused to the whole leaves and even petioles and stems. It would finally cause significant defoliation, but barely lead to plant death in Eucalyptus. Microscopic observation showed that the mycelium was straight or flexuous, hyaline, thin-walled, septate, branched, and 3-7 μm wide (n = 50; average 4.86 μm). The appressorium was lobed and attached to one end of the mycelium alone, or paired attached to both ends of the mycelium. The conidiophore was straight or flexuous, unbranched, 54-100 × 6-10 μm (n = 40; average 75.47 μm × 8.22 μm). One to 3 conidium were connate on the conidiophores. Foot-cells were straight or flexuous at base, 5-8 μm wide (n = 40; average 6.53 μm). The conidium were ellipsoid or oval, and the size was 38-56 × 12-21 μm (n = 70; average 44.92 μm × 15.69 μm). The lobed or rod-shaped bud tube was produced at the conidium. According to the morphology, the fungus was identified as Erysiphe neolycopersici (Hsiao, et al. 2022). For the molecular characterization of the isolate, the sequences of the internal transcribed spacer (ITS), the 18S and 28S large subunit ribosomal DNA (SSU and LSU) (Scholin et al. 1994 , White et al. 1990), were sequenced and deposited in GenBank (OM422667, OM424285 and ON514159). The phylogenetic analysis showed that the ITS sequence showed 100% identity with sequences of E. neolycopersici (MW082786, MT370492, and JQ972700). The 28S rDNA sequence had the highest identity (99.69%) with that of E. neolycopersici (LC371327, LC371320, and OM368490). The SSU sequence had the highest identity (99.72%) with that of E. neolycopersici (LC516961). The pathogenicity test of the fungus was repeated thrice following the Koch's postulates. The diseased leaves were gently rubbed against 3 to 4 healthy mature leaves of more than five E. grandis seedlings (two-month-old). The inoculated and control plants were then cultured in the greenhouse (25 ℃, 16-h light/8-h dark and 70% humidity). Similar disease symptoms were observed on the inoculated leaves, but not on the control leaves seven days after inoculation. The isolates from three independent experiments were morphologically and genetically identical with the original isolate. As far as we know, this study is the first report of powdery mildew disease in Eucalyptus caused by E. neolycopersici in China.

Potassium fertilization enhances xylem plasticity and growth recovery of Eucalyptus grandis trees in response to drastic changes in water availability

Most Eucalyptus plantations in Brazil are located in regions currently experiencing periods of water shortage, where fertilizers such as potassium (K) are intensively used to achieve high productivity. In a changing environment, improving our understanding of the effects of K nutrition on tree growth rates and xylem plasticity across tree ages as a function of climate is crucial for forest management. The objective of this study was to gain insight into the interactive effects of K fertilization, water availability, and tree age on stem growth and changes in xylem traits of E. grandis trees over a full rotation. Changes in growth and xylem traits in response to a severe drought that occurred in 2014 were also studied. An experimental plantation of E. grandis trees was established in São Paulo, Brazil and monitored over a full 6-year rotation period. The experiment consisted of a split-plot design with two water supply regimes (37 % throughfall reduction vs undisturbed throughfall) and two fertilization regimes (K supply vs control). Basal area increment (BAI) was monitored using band dendrometers at 14-day intervals from 1 to 6 years after planting. Xylem traits including wood density (WD), cell wall fraction (CWF), cell density (CDen), theoretical xylem specific hydraulic conductivity (Kst), vessel diameter (VD), vessel density (VDen), vessel area (VA), and the vulnerability index (VI) were measured in 6-month periods (rainy and dry seasons) throughout the rotation. K fertilization increased BAI by a factor of 3 in plots where throughfall was not disturbed but led to an interruption of tree growth during the severe drought in 2014. The growth patterns showed a positive effect of K for growth recovery following drought. Throughfall reduction affected growth only in K-fertilized trees and up to 30 months of age. K fertilization also increased changes in xylem traits in response to drought in E. grandis trees. The main changes occurred in vessel traits, with a 53 % decrease in Kst during the drought but recovery was rapid after the first rainfall events. High plasticity was also observed for WD, CWF, CDen, and VI.

Using Tectona Grandis Biomass to Produce Valuable Adsorbents for Pesticide Removal from Liquid Effluent.

This work presents a first approach concerning the valorization of Tectona Grandis tree by-products, from East Timor through their transformation into high activated carbon (AC) by chemical activation with KOH and K2CO3. The better ACs, Teak-KOH-1-1-700 and Teak-K2CO3-1-2-700, presented a high ABET (995 and 1132 m2·g−1) and micropore volume (0.43 and 0.5 cm3·g−1), respectively. Both ACs were tested on the removal of four pesticides, from the liquid phase. Both ACs performed better than existing commercial types, presenting a maximum adsorption capacity of 1.88, 1.67, 1.10 and 0.89 mmol·g−1, for 4-chloro-2-methylphenoxyacetic acid, 2,4-dichlorophenoxyacetic acid, diuron and atrazine, respectively. Pesticide adsorption from diluted and concentrated solutions confirms that diffusion is the limiting factor. The possibility of implementing a production unit for ACs in East Timor is very promising for that country. It presents an opportunity for job creation, biomass waste reduction and a contribution to environmental sustainability, thereby following the principles of a circular economy.

Fungal Communities of Eucalyptus grandis Leaves Are Influenced by the Insect Pest Leptocybe invasa.

Fungal communities in above-ground tree tissues are hyperdiverse and are influenced by biotic interactions with other organisms living in or on these tissues. These biotic interactions are, however, still poorly understood. In this study, we aimed to understand how insect-associated gall formation on Eucalyptus foliage correlates with the diversity of foliar fungal communities in surrounding healthy leaf tissue, as well as the co-occurrence patterns among the members of the fungal community. We used ITS metabarcoding to characterise the foliar fungal communities of 179 individual E. grandis trees. These trees were assigned to infestation levels of the wasp Leptocybe invasa (Eulophidae: Hymenoptera), which causes gall formation on shoot tips and leaves of its host. Fungal community networks were calculated using a Pearson correlation coefficient. The composition and diversity of fungal communities were influenced by the severity of L. invasa infestations. We identified potential Eucalyptus pathogens with high sequence abundance at all disease severity levels, but network analysis indicated that the co-occurrence of potential pathogens between no to mild and medium to heavy infestation differed significantly. A better understanding of microbial interactions, especially the role of pathogens, can be useful for controlling disease- and beneficial host-associated microbial communities.

Corrective Interventions to End-Splitting and Surface Cracking in Kenya Grown <i>Eucalyptus grandis</i> Poles

This study determined the effect of three pole pre-drying handling techniques, on end-splitting and surface checking in Eucalyptus grandis poles from highland and low land areas in Kenya. A total of 144 Eucalyptus grandis trees were sampled from two sites; Kericho, representing the wet highlands and Londiani representing the drier lowlands regions of Kenya. Pole samples from both sites were subjected to the three pre-drying handling techniques for the first 30 days after felling and allowed to dry to the required moisture level under observations. The number of end splits and surface checks on each sample pole were counted and the length of the worst ones was measured in millimeters after every 15 days until all the poles reached 25% MC. The sap wood and heart wood ratios were determined from selected samples. Results showed that on the overall, poles from high land areas had the highest sapwood proportions and similarly had the highest number and the longest end splitting and surface checks. On the other hand, pre-drying techniques that allowed felled trees to dry slowly with their foliage intact for the first 30 days of felling produced the best quality poles, with fewer and shallower end splits and surface checks. The study recommended that felling and leaving poles to dry slowly with foliage intact be considered in reducing losses incurred as a result of wood stresses during drying of poles.

High growth recovery ability of Eucalyptus grandis trees following a 3-year period of 80% throughfall reduction

Eucalyptus plantations are already planted in - or expanding to - water-limited regions with a high risk of severe drought. In future drier and more variable climate including extreme events, the ability of trees to recover after severe droughts emerges as a crucial for the sustainability of forest plantations.An original experiment involving 80% reduction in throughfall was set up in Brazil to gain insight into the responses of Eucalyptus grandis trees to prolonged (3-year) extreme water deficit and the ability of this species to recover following water stress. Our study focused on the changes in basal area, stem radius, and total height measured by high-temporal resolution dendrometers and periodical surveys of trees affected by 80% throughfall reduction (treatment group) and in a control group. The differences in basal area, stem radius, and total height growth rate were compared between groups over (i) 37 months of 80% throughfall reduction and (ii) 31 months following the end of 80% throughfall reduction. The correlations among growth rates, stem radius fluctuations, and meteorological variables in each group were determined to gain insights into the trees’ responses to environmental conditions and stem water status.The 80% reduction in throughfall over 3 years significantly reduced tree growth rates by 73% in basal area and 95% in total height. However, under normal water availability following throughfall reduction, the basal area growth rate of water-stressed trees was 97% higher than that of the control trees while total height growth rate was only 8% higher. Despite the severe water stress, no tree mortality was observed. Trees recovered 51% of their basal area over the 31-month recovery period. In contrast, only 5% of total height was recovered.In the treatment group, rapid responses to variation in rainfall events were observed during the 80% throughfall reduction period. Also, correlations between stem radius fluctuations and vapor pressure deficit indicate increased transpiration following the end of throughfall reduction. These relationships indicate high conservation of the integrity of the xylem vascular system over the 3 years of 80% throughfall reduction, a key factor in the increased resilience of the trees. In the absence of tree mortality, our results suggest the 80% throughfall reduction had a severe impact on tree growth but demonstrate great recovery ability of Eucalyptus grandis trees in basal area growth following such a severe water deficit.

Spatial Patterns of Non-Structural Carbohydrates in Eucalyptus urophylla× E. grandis under Dry-Season Irrigation with Fertilization

Non-structural carbohydrates (NSC) affect tree growth and survival when photosynthesis is impacted by climate change, such as seasonal drought and extreme precipitation. Nevertheless, it is still unclear whether Eucalyptus suffers growth limitation under natural conditions and if trees recover under artificial cultivation. In present study, we conducted a field control experiment to compare the NSC storage in Eucalyptus urophylla × Eucalyptus grandis trees on fertilization and dry-season irrigation to determine the variations of NSC under drought stress. The results indicated total soluble sugar (TSS) was the primary existing form of NSC. In spatial patterns, NSC concentration showed gradient differences from source organ to sink organ, and finally accumulated in root. The TSS concentration showed a decreased trend with height except leaf, while the trend of starch concentration was contrast. Surprisingly, fertilization and dry-season irrigation had not changed the carbon distribution among all tissues but reduced the TSS concentration in most organs. The fast-growing E. urophylla × E. grandis will consume the assimilates and carbohydrates of storage organs, but maintains the NSC concentration at a certain threshold. Our results help to comprehend the NSC allocation and improve the productivity of E. urophylla × E. grandis plantations in seasonal arid areas.

Inter-annual effects of potassium/sodium fertilization and water deficit on wood quality of Eucalyptus grandis trees over a full rotation

In Brazil, most Eucalyptus plantations are located in regions experiencing periods of water shortage, where fertilizers such as potassium (K) are intensively used to achieve high productivity. Recently, sodium (Na) has also been considered a potential nutrient in K-deficient soils. K and Na supply can increase water stress in Eucalyptus grandis trees, which could negatively impact forest productivity over prolonged droughts. Wood properties are determinants of the quality and yield of products. They are important factors to consider when measuring the impacts of silvicultural practices and water deficit on forest productivity. However, alterations in wood properties due to interactions between K/Na fertilization, water availability, and stand age are not well documented.Through annual sampling of E. grandis trees throughout a complete rotation (6 years) in Brazil, we evaluated the interactive effects of K/Na fertilization, water availability, and stand age on stem volume, sapwood/heartwood ratio, wood density, fiber, and vessel features. We also evaluated the relationships between growth rate, wood density, and fiber and vessel features. The split-plot experimental design consisted of two water supply regimes (37% throughfall reduction versus undisturbed throughfall) and three fertilization regimes (K, Na, and control).Until six years of age, K and Na fertilization increased stem volume by 3 and 2-fold, respectively, and only K-fertilized trees affected stem volume by 37% throughfall reduction. Both K and Na detrimentally affected wood density and beneficially affected fiber length. The sapwood/heartwood ratio was highly and negatively related to the growth rate of trees. Wall thickness was not affected by either K or Na. As expected, K and Na affected vessel size due to an increase in basal area growth. However, the effects of Na were more pronounced than those of K. Significant interactions between fertilization and water availability suggest that, under water deficit, wood density will not be affected by K fertilization. In contrast, the effect of Na and control treatments were invariable regardless of water availability.The effects of fertilization on wood properties changed over time, with non-significant effects in the first year and stronger effects from two years of age of trees. The variations in wood density were strongly related to variations in the fiber wall thickness. High increases in basal area growth promoted by K and Na were not related to significant decreases in wood density, but they were related to increases in fiber length. These findings highlight the significant effects of K addition on stem volume without significant losses in wood quality, even under water stress over a complete rotation. The negative impacts observed on wood density and lower productivity in commercial E. grandis plantations with the use of Na as a substitute for K should be considered.

Relationship between vessel parameters and cleavage associated with checking in Eucalyptus grandis wood

In this work, the relationship between vessels parameters and the wood cleavage strength were studied to clarify the process of formation of this type of check, very common in Eucalyptus wood. The objective was to identify the relationship between the wood cleavage strength, the average area of the vessel, and the percentage area of vessels on the wood transverse surface. For this, two Eucalyptus grandis trees at 22 years old were felled and specimens for the cleavage test were produced to determine the wood cleavage strength. From these specimens, samples were taken to determine the average area of the vessel and the percentage area of vessels, aiming at adjusting mathematical models that explain the variation in the cleavage strength. The results showed that the higher the average area of the vessel and the percentage of area occupied by vessels in the wood, the lower its cleavage strength. The multiple linear regression model can estimate the cleavage strength as a function of the average area of the vessel, and the percentage area of the vessels

Drying a Standing Teak Tree using a Solar Kiln Drying Method

Understanding the rate of drying of standing trees dried by using a solar kiln drying method could help in predicting the change of moisture content in the tree over a period of drying time. The purpose of this study was to observe the change of moisture content profile in a standing tree during drying. A standing Tectona grandis tree was selected for the experiment with a diameter at breast height of 29 cm in a mixed-species plantation. The tree was girdled with 20 cm width at 20cm above ground. For the experiment, the tree was wrapped in a transparent plastic sheet and a black plastic sheet was installed at the bottom with an angle of 39 °C facing Southwest to maximize solar radiation, with the method being modified from the solar kiln method, which is known as a greenhouse type, with 2 layers of plastic cover. The black and white sheets were placed on the inner side for insulation and for collecting heat from the sun’s radiation, while the outer layer was transparent, which allowed radiation to propagate unhindered in the black sheet. In this study, a sensor for temperature and relative humidity (DHT22) was attached inside the wrapped plastic and the data recorded every 6 min. The results showed that drying a standing teak tree from an initial average moisture content of 105 % to a constant point of 60 % took 80 days under the maximum of the collected temperature of 46 °C inside the wrapped plastic; this finding confirmed that the predicted model of recharge and discharge curve was likely accurate.

Nitrogen supply methods affect the root growth dynamics in Eucalyptus grandis

Sandy soils presenting low organic matter levels do not provide enough N to meet Eucalyptus sp. demands. Nitrogen fertilizers can be applied to soil surface, although N losses tend to happen and to decrease the amounts of this component used by plants. Alternatively, N can be applied to planting pits at seedling transplantation time or to small holes dug close to the plants. However, aspects such as the impact of different N supply modes applied to the soil on the growth parameters of root systems and the way these supply modes can affect morphological and physiological parameters of the shoot remain poorly known. The aim of the current study is to assess shoot and root growth in Eucalyptus grandis trees grown in soil subjected to different N supply modes. Eucalyptus grandis clones (GPC 23) were subjected to three N supply modes, namely: control (without N application) (C), N application to soil surface (SN) and N application to the pit (PN). Plant height, stem diameter, N concentration and chlorophyll a fluorescence in leaves, morphological parameters of the root system (total root surface area and mean root diameter), total length of active roots and total number of living and dead roots were analyzed. The PN supply mode has led to the highest mineral N levels (NH4+ and NO3−) in the soil, as well as increased the total length of active roots, the number of living roots and the root surface area in soil layers 20–40 cm and 40–60 cm. This process has enabled greater N uptake by plants, which, in its turn, has increased N concentrations in leaves, favored photosynthetic processes and resulted in taller plants presenting greater stem diameter. The SN supply method did not increase the mineral N in the soil, and it restricted the total length of active roots in soil layer (0–20 cm) and resulted in higher mean root diameter.

A cost-effective approach to DNA methylation detection by Methyl Sensitive DArT sequencing.

Several studies suggest the relation of DNA methylation to diseases in humans and important phenotypes in plants drawing attention to this epigenetic mark as an important source of variability. In the last decades, several methodologies were developed to assess the methylation state of a genome. However, there is still a lack of affordable and precise methods for genome wide analysis in large sample size studies. Methyl sensitive double digestion MS-DArT sequencing method emerges as a promising alternative for methylation profiling. We developed a computational pipeline for the identification of DNA methylation using MS-DArT-seq data and carried out a pilot study using the Eucalyptus grandis tree sequenced for the species reference genome. Using a statistic framework as in differential expression analysis, 72,515 genomic sites were investigated and 5,846 methylated sites identified, several tissue specific, distributed along the species 11 chromosomes. We highlight a bias towards identification of DNA methylation in genic regions and the identification of 2,783 genes and 842 transposons containing methylated sites. Comparison with WGBS, DNA sequencing after treatment with bisulfite, data demonstrated a precision rate higher than 95% for our approach. The availability of a reference genome is useful for determining the genomic context of methylated sites but not imperative, making this approach suitable for any species. Our approach provides a cost effective, broad and reliable examination of DNA methylation profile on MspI/HpaII restriction sites, is fully reproducible and the source code is available on GitHub (https://github.com/wendelljpereira/ms-dart-seq).