Mature Needles Research Articles

Responses of needle terpene concentrations and characteristics of resin canals to different warming treatments in Scots pine and Norway spruce seedlings grown in a field experiment

We studied responses of needle terpene concentrations and resin canal characteristics to warming in Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) seedlings grown in a controlled field set-up in eastern Finland. Warming was simulated during the growing seasons using infrared heaters that increased air temperature in study 1 (2016–2019) by 1°C and in study 2 (2019–2020) by 2°C and 4°C, compared to ambient conditions. Terpenes were sampled in study 1 from non-matured current year and matured previous year needles in June 2019, and study 2 from mature current-year needles in August 2020. In study 1, we also studied resin canal anatomy. We found that 1°C elevation of temperature caused two-fold increase in concentrations of total terpenes, oxygenated monoterpenes and sesquiterpenes of non-matured current year needles of Norway spruce. Further, it caused 1.3-1.8-fold increases in sesquiterpene concentrations both in unmatured and matured needles of Scots pine. It also decreased resin canal diameter of mature needles in Norway spruce. In study 2, the stronger warming treatments did not affect terpene concentrations of matured current-year needles in either species. Based on our findings, even minor elevation of temperature may affect terpene concentrations of non-mature needles in boreal conifers.

Comparative transcriptomics of Pinus massoniana organs provides insights on terpene biosynthesis regulation.

Terpenoids are the most important natural products collected from conifer species. However, the molecular mechanisms and core factors underlying terpenoid biosynthesis in Pinus massoniana remain unclear. To clarify these mechanisms, this study aimed to identify potential genes that might participate in the terpenoid biosynthesis of P. massoniana. In this study, single molecule real-time (SMRT) sequencing and expression analysis were used to confirm the expression patterns of genes involved in the cones, immature needles, mature needles, immature branches, and mature branches of P. massoniana. A total of 31,331 lncRNAs and 71,240 mRNAs were identified from these organs, and the greatest number of differentially expressed genes (DEGs) was discovered between needles and branches. Weighted gene coexpression network analysis (WGCNA) classified all expressed genes into nine typical modules with 11 kinds of transcription factors (TFs), namely, AP2-ERF, ARF, AUX-IAA, C2H2, Dof, F-box, SBP, WRKY, bHLH, bZIP, and GRAS, and seven kinds of functional genes, namely, ABC transporter, cellulose synthase (CesA), leucine-rich repeats (LRR), cytochrome P450 (CYT P450), pathogenesis-related protein (PR), terpene synthase (TPS), and chlorophyllase enzyme. A molecular network was constructed for hub genes, TFs, and functional genes in three modules. The potential function of eight candidate genes, including PmbHLH2, PmERF1, PmRGA, PmGAI, PmbZIP1, PmLOB1, PmMADS1, and PmMYB1, was validated through correlation analysis between terpenoid contents and expression levels, subcellular localization, and transcriptional activation activity, which provides us with probable regulators of terpenoid biosynthesis in conifers.

Variación fitoquímica entre procedencias de oyamel (Abies religiosa [Kunth] Schltdl. & Cham.) en un gradiente altitudinal

Introduction: A species’ populations differ across its distribution due to selection pressure from the climate gradient. Objective: To describe the phytochemical variation among 14 tree provenances in an altitudinal gradient of Abies religiosa (Kunth) Schltdl. & Cham. to determine if there are associations of chemical profiles among populations, expressed under a single environment. Materials and methods: Seeds were collected from 165 trees from an altitudinal transect from 2 850 to 3 540 m in Cerro San Andrés, Municipio Libre de Hidalgo, Michoacán, Mexico. The plant was grown in a common garden trial; at 28 months, mature needles were collected per tree and analyzed by gas-mass chromatography. Data were analyzed with diversity indices, ANOVA and multivariate analysis (principal components, correspondences and cluster). Results and discussion: Thirty-two compounds were identified, most of them terpenes. The populations showed differences in presence/absence of terpenes and in their concentration; above 3 350 m, diversity was lower, but with higher concentration. Three altitude groups were found based on the chemical profile of the populations: low = 2 850 to 3 300 m and 3 400 m; moderate = 3 350, 3 450 and 3 500 m; and high= 3 540 m. Conclusions: The diversity and concentration of terpenes in A. religiosa varied depending on the altitude of populations. There are three chemical profiles that should be considered in the implementation of germplasm collection programs for reforestation and restoration

Determination of the Volatile Organic Compounds (VOCs) in Mature and Immature Foliage of Five Species of Pinaceae by Gas Chromatography–Mass Spectrometry (GC-MS) with Principal Component Analysis (PCA) and Cluster Analysis (CA)

Conifer needles are rich in resources and have essential ecological and biological functions. However, comparative analysis of the volatile components of immature and mature needles of the family Pinaceae has not been reported to date. In this study, headspace solid-phase microextraction coupled with gas chromatography–mass spectrometry (GC-MS) was used to identify the volatile organic compounds (VOCs) in the immature and mature needles of five Pinaceae species. Altogether, 51 VOCs were identified, 31 of which reportedly show biological activity. The identified compounds include alcohols (6), terpenes (35), esters (7), and others (3). Terpenes were the major metabolites (78.38% to 97.18%) in the needles of the conifers. The needles were divided into four categories based on principal component analysis (PCA) and cluster analysis (CA). This is the first study to show a detailed examination of the differences in VOCs between immature and mature conifer needles. Furthermore, immature and mature needles of the five species were distinguished based on their specific VOCs. Our findings provide a reference for the utilization of conifer needles as valuable renewable resources.

Fungal diversity within the phyllosphere of Pinus massoniana and the possible involvement of phyllospheric fungi in litter decomposition

Fungi play key roles in forest ecosystems and help to shape the forest’s diverse functions. However, little is known about the diversity of phyllospheric fungi or their possible relationships with fungal communities residing in different micro-environments of Pinus massoniana forests. We investigated seven different sample types: mature needles (NM), dead needles (ND), needles falling as litter (L), fermenting needles (F), humus (H), top soil (0–20 cm) (TS), and secondary soil (20–40 cm) (SS). These seven fungal communities were examined and compared with ITS amplicons using a high-throughput sequencing technique. A total of 1213 fungal operational taxonomic units (OTUs) were obtained at a 97% sequence similarity level. Distinct fungal communities were associated with different sample types. A greater number of OTUs were present in both NM and F samples than those shared by both NM and TS samples, indicating that phyllospheric fungi may play crucial roles in litter decomposition. Sixty OTUs (the core microbiome) were found in all sample types, and they may probably play different ecological roles in different sample types. These findings extend our knowledge of the fungal diversity of the phyllosphere and its possible interactions with fungal communities found in distinct forest micro-habitats.

1,6-Dehydropinidine Is an Abundant Compound in Picea abies (Pinaceae) Sprouts and 1,6-Dehydropinidine Fraction Shows Antibacterial Activity against Streptococcus equi Subsp. equi.

Knowledge about the defensive chemistry of coniferous trees has increased in recent years regarding a number of alkaloid compounds; in addition to phenolics and terpenes. Here, we show that Norway spruce (Picea abies (L.) H. Karst.), an important boreal zone tree species; accumulates 1,6-dehydropinidine (2-methyl-6-(2-propenyl)-1,6-piperideine) in its needles and bark. We reanalyzed previously published GC-MS data to obtain a full picture of 1,6-dehydropinidine in P. abies. 1,6-dehydropinidine appeared to especially accumulate in developing spring shoots. We used solid-phase partitioning to collect the alkaloid fraction of the sprouts and thin-layer chromatography to purify 1,6-dehydropinidine. The antibacterial properties of the 1,6-dehydropinidine fraction were tested using a broth microdilution method; with Streptococcus equi subsp. equi as a model organism. Based on our results 1,6-dehydropinidine is common in alkaloid extractions from P. abies (0.4 ± 0.03 mg g−1 dw in mature needles) and it is especially abundant in young spruce shoots (2.7 ± 0.5 mg g−1 dw). Moreover; 1,6-dehydropinidine extracted from P. abies sprouts showed mild antibacterial potential against Streptococcus equi subsp. equi (MIC 55 µg mL−1). The antibacterial activity of a plant compound thought of as an intermediate rather than an end-product of biosynthesis calls for more detailed studies regarding the biological function of these coniferous alkaloids

Production of constitutive and induced secondary metabolites is coordinated with growth and storage in Norway spruce saplings.

A mechanistic understanding of how trees balance the trade-offs between growth, storage and defense is limited but crucial for predicting tree responses to abiotic and biotic stresses. Here we investigated how trees allocate storage of non-structural carbohydrates (NSC) to growth and constitutive and induced secondary metabolites (SM). We exposed Norway spruce (Picea abies) saplings to 5 weeks of complete darkness to induce light and/or carbon limitation and then applied methyl jasmonate (MeJA) to simulate biotic attack. We measured changes in biomass, NSC (sum of soluble sugars and starches), and constitutive and induced SM (sum of phenolic compounds and terpenoids) in current-year developing and previous-year mature needles and branches, as well as volatiles emitted from the canopy. Under darkness, NSC storage was preferentially used for constitutive biosynthesis of monoterpenes rather than biosynthesis of stilbenes and growth of developing organs, while SM stored in mature organs cannot be remobilized and recycled. Furthermore, MeJA-induced production of SM was constrained by low NSC availability in developing organs but not in mature organs grown in the dark. Emissions of volatiles were suppressed in the dark but after 1 h of re-illumination, emissions of both constitutive and induced monoterpene hydrocarbons recovered rapidly, whereas emissions of linalool and sesquiterpene produced via de novo synthesis did not recover. Our results highlight that light and/or carbon limitation may constrain constitutive and JA-induced biosynthesis of SM in coordination with growth, NSC storage and mobilization.

Seasonal changes in eastern hemlock (Tsuga canadensis) foliar chemistry

Eastern hemlock (Tsuga canadensis (L.) Carrière) is an eastern North American conifer threatened by the invasive hemlock woolly adelgid (Adelges tsugae Annand). Changes in foliar terpenes and phenolics were evaluated in new (current-year growth) and mature (1-year-old growth) hemlock needles during the growing season and into plant dormancy. From April through September, foliar concentrations of nonvolatile soluble phenolics, condensed tannins, lignin, mono- and sesquiterpenes, α-pinene, camphene, isobornyl acetate, and diterpene resin were quantified. After September, additional analyses of metabolites that continued to differ significantly between new and mature foliage were carried out. Total soluble phenolic and condensed tannin concentrations in new foliage remained low relative to those of mature foliage throughout the growing season and converged in December. Lignin concentration in new foliage converged with that of mature foliage by July. Concentrations of α-pinene, camphene, isobornyl acetate, and diterpene resin in new foliage converged with those of mature foliage within 1 month of budbreak. The convergence of terpene concentrations in new and mature foliage suggests that these metabolites may play a role in herbivore defense during the peak growing season. Conversely, soluble phenolics, including condensed tannins, may defend foliage from herbivory outside of the spring growth period.

Using unmanned aerial vehicle‐based multispectral, RGB and thermal imagery for phenotyping of forest genetic trials: A case study inPinus halepensis

The assessment of genetic differentiation in functional traits is fundamental towards understanding the adaptive characteristics of forest species. While traditional phenotyping techniques are costly and time‐consuming, remote sensing data derived from cameras mounted on unmanned aerial vehicles (UAVs) provide potentially valid high‐throughput information for assessing morphophysiological differences among tree populations. In this work, we test for genetic variation in vegetation indices (VIs) and canopy temperature among populations ofPinus halepensisas proxies for canopy architecture, leaf area, photosynthetic pigments, photosynthetic efficiency and water use. The interpopulation associations between vegetation properties and above‐ground growth (stem volume) were also assessed. Three flights (July 2016, November 2016 and May 2017) were performed in a genetic trial consisting of 56 populations covering a large part of the species range. Multispectral (visible and near infrared wavelengths), RGB (red, green, blue) and thermal images were used to estimate canopy temperature and vegetation cover (VC) and derive several VIs. Differences among populations emerged consistently across flights for VC and VIs related to leaf area, indicating genetic divergence in crown architecture. Population differences in indices related to photosynthetic pigments emerged only in May 2017 and were probably related to a contrasting phenology of needle development. Conversely, the low population differentiation for the same indices in July 2016 and November 2016 suggested weak interpopulation variation in the photosynthetic machinery of mature needles ofP. halepensis. Population differences in canopy temperature found in July 2016 were indicative of variation in stomatal regulation under drought stress. Stem volume correlated with indices related to leaf area (positively) and with canopy temperature (negatively), indicating a strong influence of canopy properties and stomatal conductance on above‐ground growth at the population level. Specifically, a combination of VIs and canopy temperature accounted for about 60% of population variability in stem volume of adult trees. This is the first study to propose UAV remote sensing as an effective tool for screening genetic variation in morphophysiological traits of adult forest trees.

Effects of auxin-transport-inhibitor and defoliation on wood formation in locally-heated Abies homolepis

ABSTRACTTo understand the precise process of wood formation, it is necessary to identify the factors that regulate cambial activity and development of cambial derivatives. Here, we investigated the combined effects of localized-heating and auxin on cambial reactivation and the formation of earlywood tracheids in seedlings of the evergreen coniferAbies homolepisin winter. Three treatments were applied, namely heating (artificial increase in temperature 20–22 °C), heating-plus-auxin transport inhibitor N-(1-naphthyl) phthalamic acid (NPA) and heating-plus-defoliation (removal of needles and buds), with an approximate control, for investigations of cambial activity by light microscopy. After one week of heating, cambial reactivation occurred in the heating, heating-plus-NPA and heating-plus-defoliation treatments. In untreated controls, cambial reactivation occurred later than in heated stems. Earlywood tracheids were formed after three and six weeks of heating in the heating and heating-plus-NPA treatments, respectively. No tracheids were formed after eight weeks of heating in heated-defoliated seedlings. Numbers of new tracheids were reduced in heated stems by NPA. Our results suggest that an increase in the temperature of the stem is one of the most important limiting factors in cambial reactivation, which is independent of needles and buds and of the polar transport of auxin from apical sources. However, after cambial reactivation, initiation and continuous formation of earlywood tracheids require basipetally transported auxin and other endogenous factors originating in mature needles and buds.

Quality and quantity of piperidine alkaloids in needles and bark of Scots pine (Pinus sylvestris) seedlings

The Northern boreal forest zone is dominated by two coniferous species that synthesize piperidine alkaloids: Scots pine (Pinus sylvestris) and Norway spruce (Picea abies). These compounds are known to have antifeedant properties. We have earlier shown that Norway spruce has a diverse alkaloid chemistry, but reports from P. sylvestris are few, and no quantitative analysis has been conducted so far. Here we have studied 2-year-old seedlings of P. sylvestris to reveal possible differences in alkaloid chemistry compared to P. abies. Alkaloids were extracted from bark and mature needles by solid-phase partitioning and analysed by GC–MS. We detected only four individual compounds from P. sylvestris samples, confirming earlier assumptions that the species lacks large parts of the biosynthesis pathway of coniferous alkaloids. Euphococcinine, also present in mature P. abies needles, is the sole end-product of alkaloid biosynthesis in P. sylvestris, although a compound tentatively identified as an isomer of euphococcinine was also detected. The two other alkaloid compounds detected are also encountered in P. abies, but only in juvenile plant parts, such as developing needles and stems. Concentrations of all alkaloids were extremely low, with totals amounting to about 25% of the amount found in P. abies. It is also notable that concentrations in three out of ten seedlings were under the detection limit, in bark or in both plant parts, whereas in P. abies individuals lacking alkaloids are virtually non-existent. The compound composition, concentrations and absence of alkaloids in some individuals emphasize the difference between these two major boreal zone species, and this calls for further studies on the ecological significance of these compounds.

Pest categorisation of Gilpinia hercyniae.

The Panel on Plant Health performed a pest categorisation of the Diprionid sawfly, Gilpinia hercyniae Hartig (Hymenoptera: Diprionidae), for the EU. G. hercyniae is a well‐defined and distinguishable species, native to Europe but also present in North America, Japan, Mongolia, Korea and Pakistan, and recognised as a pest of spruce (Picea spp.). The pest is distributed in 19 Member States (MSs) of the EU. It is a quarantine pest listed in Annex IIB of Council Directive 2000/29/EC. Protected zones are in place in Greece, Ireland and the United Kingdom (Northern Ireland, Isle of Man and Jersey). Plants for planting of Picea spp. and soil and litter associated with Picea spp. are considered as pathways for this pest, which is also able to disperse by flight. The prepupae overwinter inside cocoons in the litter or in the foliage. In spring, the adults, mostly females emerge and lay 35–60 eggs per female in mature needles. The larvae feed on the mature needles through five instars. There are 1–3 generations per year; some of the prepupae undergo prolonged diapause for more than 1 year. The impact on Picea abies (= excelsa) is minimal, because only the needles of the previous years are attacked; however, outbreaks have occurred on non‐native spruce, Picea glauca and Picea sitchensis. The pest is controlled everywhere by natural enemies, including nuclear polyedrosis viruses. The insects spread on plants for planting of Picea spp., with soil and litter associated with Picea spp., and by flight. The EU protected zones have a similar climate and similar host plants as the MS where G. hercyniae is established. All criteria assessed by EFSA for consideration as potential protected zone quarantine pest and as a potential regulated non‐quarantine pest were met.

Pest categorisation of Cephalcialariciphila.

The Panel on Plant health performed a pest categorisation of the larch web‐spinning sawfly Cephalcia lariciphila (Hymenoptera: Pamphiliidae) for the EU. The insect has been reported in 11 EU Member States (MSs). It is a quarantine pest listed in Annex IIB of Council Directive 2000/29/EC. Protected zones are in place in Ireland and the UK (Northern Ireland, Isle of Man and Jersey). C. lariciphila can feed on all species of the genus Larix. There have been reported outbreaks in the Czech Republic, Germany, the Netherlands and the UK (England and Wales) in plantations of European larch (Larix decidua) and Japanese larch (Larix kaempferi = Larix leptolepis). C. lariciphila is absent in the protected zones. The pest can enter the protected zones by human‐assisted spread or by natural spread from EU areas where the pest is present. Plants for planting are considered the most important pathway for the pest. The pest can establish in the protected zones because the climatic conditions are similar to those of the 11 MSs where C. lariciphila is established, and the pest's main host plants are present. The prepupae overwinter in the litter, the adults emerge during May–June, and each female lays 30–40 eggs in slits in mature needles. The larvae feed on the needles through four instars. There is one generation per year; some of the prepupae undergo prolonged diapause for more than 1 year. The impact where the pest occurs is mainly related to the loss of tree growth following defoliation, while tree mortality was locally observed only after repeated defoliation. However, impact is likely to be mitigated by local biological control agents. All criteria assessed by EFSA above for consideration as a potential protected zone quarantine pest and as a potential regulated non‐quarantine pest were met.

Variation in piperidine alkaloid chemistry of Norway spruce (Picea abies) foliage in diverse geographic origins grown in the same area

Volatile piperidine alkaloids are widespread compounds found in low amounts in coniferous species. Here, we investigated the variation of piperidine alkaloid chemistry of Norway spruce (Picea abies (L.) Karst.) in a provenance-type experiment. Volatile piperidine alkaloids were analysed from current-year mature needles of 12 geographical origins grown for 35 years at the same site. Results indicate that, despite long-lasting similar growth conditions, origins were still showing different alkaloid composition and concentrations, indicating solid genotype variation. The highest total piperidine alkaloid concentrations were detected in the northernmost origins, and a marked positive correlation between piperidine alkaloid concentrations and latitude was found. Positive correlation with latitude seems to be closely linked to negative correlation with annual temperature sum of origins. Interestingly, the changes in concentrations detected were not related to needle size and dilution effect. Regarding the possible herbivore-defense role of P. abies piperidine alkaloids, the high variation between origins could play a role for preventing the invasion of genotypes from different geographical areas.

New foliage growth is a significant, unaccounted source for volatiles in boreal evergreen forests

Abstract. Estimates of volatile organic compound (VOC) emissions from forests are based on the assumption that foliage has a steady emission potential over its lifetime, and that emissions are mainly modified by short-term variations in light and temperature. However, in many field studies this has been challenged, and high emissions and atmospheric concentrations have been measured during periods of low biological activity, such as in springtime. We conducted measurements during three years, using an online gas-exchange monitoring system to observe volatile organic emissions from a mature (1 year-old) and a growing Scots pine shoot. The emission rates of organic vapors from vegetative buds of Scots pine during the dehardening and rapid shoot growth stages were one to two orders of magnitude higher than those from mature foliage; this difference decreased and finally disappeared when the new shoot was maturing in late summer. On average, the springtime monoterpene emission rate of the bud was about 500 times higher than that of the mature needles; during the most intensive needle elongation period, the monoterpene emission rate of the growing needles was 3.5 higher than that of the mature needles, and in September the monoterpene emission rate of the same years' needles was even lower (50%) than that of the previous years' needles. For other measured compounds (methanol, acetone and methylbutenol) the values were of the same order of magnitude, except before bud break in spring, when the emission rates of buds for those compounds were on average about 20–30 times higher than that of mature needles. During spring and early summer the buds and growing shoots are a strong source of several VOCs, and if they are not accounted for in emission modeling a significant proportion of the emissions – from a few percent to even half of the annual cumulative emissions – will remain concealed. The diurnal emission pattern of growing shoots differed from the diurnal cycle in temperature as well as from the diurnal emission pattern of mature shoots, which may be related to processes involved in shoot or needle elongation. Our findings imply that global estimations of monoterpene emission rates from forests are in need of revision, and that the physiological state of the plants should be taken into account when emissions of the reactive gases such as monoterpenes are estimated.

Survival and growth of eastern larch, balsam fir, and black spruce six years after winter browning in northeastern Ontario, Canada

In spring 2007, young conifers suffered extensive needle and bud injury near Hearst, Kapuskasing, and Cochrane in northeastern Ontario, Canada. Analysis of weather data revealed that the damage was likely caused by earlier than normal loss of cold hardiness during higher than normal temperatures in late April and early May. The dehardened trees were damaged in the cold days that followed. We report the survival and growth of eastern larch (Larix laricina), balsam fir (Abies balsamea), and black spruce (Picea mariana) in the affected area based on field assessments and stem analysis of affected trees following the event. Total survival through six growing seasons following damage was 98% in eastern larch but only about 70% in balsam fir and black spruce. The effect of the damage on tree growth was greater and longer lasting for diameter than height. Both survival and growth reductions were associated with high levels of mature needle loss and bud damage. Implications of these findings for forest management are discussed in light of expected climate change.

Inter population genetic diversity analysis using ISSR markers in Pinus roxburghii (Sarg.) from Indian provenances

Inter population genetic diversity analysis was undertaken between four Pinus roxburghii populations of India using ISSR markers. A total of 72 accessions of genomic DNA from mature needles from four distantly located populations (two native/natural from sub-tropical forests and two provenance trial plantations in plains) were isolated and subjected to ISSR analysis. The percent band polymorphism was 100%, observed number of alleles (na*-2.00), effective number of allele (ne*-1.30), Nei’s Gene diversity/Heterozygosity (h*-0.19) and Shanon’s information index (I*- 0.31). For these 4 populations upon inter-population genetic diversity analysis, Gene flow (Nm*-1.39) was obtained. Unweighted paired group method with arithmetic mean for similarity index and genetic distances were calculated using Jaccard’s coefficient. The highest genetic distance of 0.0904 was obtained between native and provenance trial plantation. Likewise, for similarity index the clusters in the dendrogram were found separated at 0.3 to 0.4 similarity values, indicating robust separations. Such type of diversity analysis will be helpful in raising sustainable plantation forestry programs in plains of India for this species. Diversity analysis data using distinctly located populations of P. roxburghii has been done for the first time in Indian provenances. Key words: Inter simple sequence repeat (ISSR), genetic diversity, inter-population diversity analysis, Nei’s gene diversity, Shannon’s information index.

A comparative survey of proteins from recalcitrant tissues of a non‐model gymnosperm, Douglas‐fir

Most research in plants and other organisms has, for the sake of convenience, focused on the use of model species to identify mechanisms that are conserved throughout the whole kingdom. Nevertheless, unique features and processes such as those related to plant cell wall and fiber formation, and to wood quality, sometimes need to be studied directly in the non-model organism of interest. Such organisms, like the economically and ecologically important gymnosperm Douglas-fir (Pseudotsuga menziesii), which is one of the crucial softwood timber species in Northern America, are often difficult to investigate. High phenolic, resin, and tannin contents in the woody tissues, as well as an incompletely sequenced genome, have contributed greatly to the species' recalcitrance for molecular biology investigations. In this study, we present a complete procedure detailing protein sample preparation, separation, and proteomic analysis based on cross-species identification of Douglas-fir. Proteins from the cambial zone, mature needles, and in vitro callus were extracted, purified, and separated via 1D and 2D SDS-PAGE. One-dimensional electrophoresis coupled with ESI-MS/MS was used for cross-species protein identification in order to evaluate the potential of this approach and reveal major differences in protein profiles among tested tissues. Identified proteins were functionally and developmentally compared. The likely contribution of these proteins to the properties of the cell wall and wood is indicated and discussed.

Defoliation negatively affects plant growth and the ectomycorrhizal community of Pinus pinaster in Spain

In this work, by artificially reproducing severe (75%) and moderate (25%) defoliation on maritime pines Pinus pinaster in NW Spain, we investigated, under natural conditions, the consequences of foliage loss on reproduction, abundance, diversity and richness of the fungal symbionts growing belowground and aboveground. The effect of defoliation on tree growth was also assessed. Mature needles were clipped during April 2007 and 2008. Root samples were collected in June-July 2007 and 2008. Collection of sporocarps was performed weekly from April 2007 to April 2009. Taxonomic identity of ectomycorrhizal fungi was assessed by using the internal transcribed spacer (ITS) regions of rDNA through the polymerase chain reaction (PCR) method, subsequent direct sequencing and BLAST search. Ectomycorrhizal colonization was significantly reduced (from 54 to 42%) in 2008 by 75% defoliation, accompanied with a decline in species richness and diversity. On the other hand, sporocarp abundance, richness and diversity were not affected by foliage loss. Some ECM fungal symbionts, which are assumed to have a higher carbon cost according to the morphotypes structure, were reduced due to severe (75%) defoliation. Furthermore, 75% foliage loss consistently depressed tree growth, which in turn affected the ectomycorrhizal growth pattern. Defoliation impact on ECM symbionts largely depends on the percentage of foliage removal and on the number of defoliation bouts. Severe defoliation (75%) in the short term (2 years) changed the composition of the ECM community likely because root biomass would be adjusted to lower levels in parallel with the depletion of the aboveground plant biomass, which probably promoted the competition among mycorrhizal types for host resources. The persistence of fungal biomass in mycorrhizal roots would be crucial for nutrient up-take and recovery from defoliation stress of the host plants.

Combined Proteomic and Transcriptomic Analysis Identifies Differentially Expressed Pathways Associated to Pinus radiata Needle Maturation

Needle differentiation is a very complex process that leads to the formation of a mature photosynthetic organ from pluripotent needle primordia. The proteome and transcriptome of immature and fully developed needles of Pinus radiata D. Don were compared to described changes in mRNA and protein species that characterize the needle maturation developmental process. A total of 856 protein spots were analyzed, defining a total of 280 spots as differential between developmental stages, from which 127 were confidently identified. A suppressive subtractive library (2048 clones, 274 non redundant contigs) was built, and 176 genes showed to be differentially expressed. The Joint data analysis of proteomic and transcriptomic results provided a broad overview of differentially expressed pathways associated with needle maturation and stress-related pathways. Proteins and genes related to energy metabolism pathways, photosynthesis, and oxidative phosphorylation were overexpressed in mature needles. Amino acid metabolism, transcription, and translation pathways were overexpressed in immature needles. Interestingly, stress related proteins were characteristic of immature tissues, a fact that may be linked to defense mechanisms and the higher growth rate and morphogenetic competence exhibited by these needles. Thus, this work provides an overview of the molecular changes affecting proteomes and transcriptomes during P. radiata needle maturation, having an integrative vision of the functioning and physiology of this process.