Giovanna Aronne

Variations of vessel diameter and δ13C in false rings of Arbutus unedo L. reflect different environmental conditions.

Woody species in Mediterranean ecosystems form intra-annual density fluctuations (IADFs) in tree rings in response to changes in environmental conditions, especially water availability. Dendrochronology, quantitative wood anatomy and high-resolution isotopic analysis (using a laser ablation technique) were used to characterize IADFs in Arbutus unedo shrubs grown on two sites with different water availability on the island of Elba (Italy). Our findings show that IADF characterization can provide information about the relationship between environmental factors and tree growth at the seasonal level. At the more xeric site, IADFs mainly located in the early and middle parts of the annual ring, showed a decrease in vessel size and an increase in δ(13) C as a result of drought deficit. Opposite trends were found at the more mesic site, with IADFs located at the end of the ring and associated with a lower δ(13) C. Moreover, at the first site, IADFs are induced by drought deficit, while at the second site IADFs are linked with the regrowth in the last part of the growing season triggered by favourable wet conditions. This combined approach is a promising way for dating problematic wood samples and interpreting the phenomena that trigger the formation of IADFs in the Mediterranean environment.

Variations of Wood Anatomy and δ13C Within-Tree Rings of Coastal Pinus Pinaster Showing Intra-Annual Density Fluctuations

We investigated the variation of wood anatomical characteristics and carbon isotopic composition of tree rings showing intra-annual density fluctuations (IADFs) in plants of Pinus pinaster Ait. growing at a coastal plantation in Tuscany (Italy). IADFs are regions of the tree ring where wood density changes abruptly due to a sudden change of environmental conditions, particularly of water availability. Dendrochronological analyses allowed dating of the rings and four regions were considered in each tree ring: earlywood, IADF, late-earlywood and latewood. Although IADF commonly has been classified as latewood-like tissue in the literature, we found differences in anatomical characteristics and carbon isotopic composition between tracheids of the two regions. The lumen area of tracheids in IADF was significantly larger than in latewood, while still smaller than in earlywood and late-earlywood. Latewood and IADF had a greater proportion of narrow tracheids than both earlywood and late-earlywood. Although latewood and IADF were characterized by tracheids with lumina lengthened in the tangential direction, while earlywood tracheids were elongated in the radial direction, some differences were found also between latewood and IADF. Moreover, IADF tracheids had a higher 13C/12C ratio than any other region and showed isotopic values significantly different from the latewood. The quantification of anatomical features of tracheids within rings was useful to discriminate between latewood and IADFs, as well as helpful for the identification of tree-ring boundaries. The overall interpretation of dendrochronological, wood anatomical and carbon isotopic data seems to be a promising approach for the dating and the ecological interpretation of tree rings in Mediterranean ecosystems and for gaining climatic information with intra-annual resolution.

Wood anatomy and hydraulic architecture of stems and twigs of some Mediterranean trees and shrubs along a mesic-xeric gradient

Woody species populating the major Mediterranean ecosystems in the world are characterized by different levels of adaptation to the seasonal Mediterranean climate conditions. Many species of these ecosystems show wood features that allow high efficiency of transport when water is available, while maintaining hydraulic safety during drought periods. This study focuses on the anatomy of juvenile and mature wood of some species representative of continuous sequences of Mediterranean vegetation formations according to gradients of water availability, from xeric to relatively mesic: Cistus monspeliensis L., Rhamnus alaternus L., Myrtus communis L., Pistacia lentiscus L., Olea europaea L., Quercus ilex L., Fraxinus ornus L. and Ostrya carpinifolia L. Twigwood collected in Southern Italy was anatomically compared with the stemwood of the same species represented in the reference slide collection of the National Herbarium of the Netherlands (Lw). The “hydraulic distance” between the wood of main stems and twigs was estimated on the basis of suites of anatomical features related to water efficiency/safety. Although some attributes (i.e. porosity and type of imperforate tracheary elements) were similar in young twigs and older rings, other traits (i.e. vessel frequency and size) evidenced the different hydraulic properties of twig and stemwood. The difference between juvenile and mature structures was large in the species of the mesic end of the gradient while it was relatively small in those more xeric. This tendency is in agreement with the habit gradient from medium-sized trees to small evergreen/drought deciduous shrubs according to decreasing water availability in Mediterranean vegetation types.

Morpho-Anatomical Traits for Plant Adaptation to Drought

Plant resistance to drought relies on adaptive strategies based on the timing of phenophases and on the presence of structural traits mainly related to: (1) increase of water uptake and storage; (2) reduction of water loss during dry periods; and (3) mechanical reinforcement of tissues to prevent wilting that may lead to irreversible collapse and damage of cells. In this chapter, after a few evolutionary considerations, we focus on the adaptive value of the main phenological, morpho- logical and anatomical properties. We report the common existence of such traits in both desert and semiarid environments, especially in Mediterranean-type ecosys- tems. All morpho-anatomical characteristics are interpreted considering that plant resistance to drought also depends on the ability to respond to multiple stressors. We conclude that various combinations of anatomical features can contribute in different degrees to the adaptive capacity of plants to drought.

Drought impact on water use efficiency and intra‐annual density fluctuations in Erica arborea on Elba (Italy)

Erica arborea (L) is a widespread Mediterranean species, able to cope with water stress and colonize semiarid environments. The eco-physiological plasticity of this species was evaluated by studying plants growing at two sites with different soil moistures on the island of Elba (Italy), through dendrochronological, wood-anatomical analyses and stable isotopes measurements. Intra-annual density fluctuations (IADFs) were abundant in tree rings, and were identified as the key parameter to understand site-specific plant responses to water stress. Our findings showed that the formation of IADFs is mainly related to the high temperature, precipitation patterns and probably to soil water availability, which differs at the selected study sites. The recorded increase in the (13) C-derived intrinsic water use efficiency at the IADFs level was linked to reduced water loss rather than to increasing C assimilation. The variation in vessel size and the different absolute values of δ(18) O among trees growing at the two study sites underlined possible differences in stomatal control of water loss and possible differences in sources of water uptake. This approach not only helped monitor seasonal environmental differences through tree-ring width, but also added valuable information on E. arborea responses to drought and their ecological implications for Mediterranean vegetation dynamics.

Discrete versus continuous analysis of anatomical and δ13C variability in tree rings with intra-annual density fluctuations

Intra-annual density fluctuations (IADFs) are anomalies of tree rings where wood density is abruptly altered after sudden changes in environmental conditions. Their characterisation can provide information about the relationship between environmental factors and eco-physiological processes during tree growth. This paper reports about the variability of anatomical traits and stable carbon isotopic composition along tree rings as resulting from the application of two different methodological approaches: (a) the separation of each ring into different regions (earlywood, latewood and IADF) and the comparison of anatomical and isotopic parameters measured in those specific sectors and (b) the analysis of such features in continuum along ring width. Moreover, different parameters of vessels (i.e. ecd—equivalent circle diameter, elongation, sphericity and convexity of vessel lumen) were considered to identify those more appropriate for the representation of intra-annual anatomical variations. The analysis was conducted on Arbutus unedo L. growing on the Elba Island (Italy); tree rings of this species form IADFs with features clearly responsive to the environmental conditions experienced during plant growth. Results showed that the first approach, although more suitable to obtain data for subsequent statistical comparisons and for the calculation of correlations with environmental parameters, suffers from elements of subjectivity due to the size and position of the selected tree-ring regions. The in continuum method allows a clearer identification of the variation of tree-ring properties along ring width. Regarding anatomical parameters, shape indexes were not suitable indicators of intra-annual variability. The overall analysis suggested that using both methodologies in synergy helps to gain complete information and avoid misleading interpretations of IADFs in tree rings.

Seasonal dimorphism in wood anatomy of the Mediterranean Cistus incanus L. subsp. incanus

In Mediterranean-type ecosystems, seasonal dimorphism is an adaptive strategy aimed to save water by developing brachyblasts with xeromorphic summer leaves as opposed to dolichoblasts with more mesomorphic winter leaves. The aim of this study was to analyse the anatomical properties of 1-year-old twigs of Cistus incanus subsp. incanus, a seasonally dimorphic shrub, to highlight properties allowing its adaptation to the Mediterranean environment. A more specific purpose was to verify the occurrence of seasonal dimorphism in wood anatomy in order to understand: (a) whether and to what extent the traits of efficiency/safety of water transport are expressed in brachyblasts and dolichoblasts, and (b) the effects on the formation of growth ring boundaries in wood. Our overall analysis showed that anatomical features of branches in C. incanus are designed to: (a) protect from desiccation by developing thick cuticle and suberized epidermal and sub-epidermal layers; (b) defend the plant from predators by accumulating phenolics; and (c) regulate water transport through the development of specific wood anatomy, according to the season, thus optimising properties of efficiency/safety. Regarding the latter point, our results indicated that brachyblast wood is safer than dolichoblast wood insofar as it has narrower and more frequent vessels; measurement of other specific anatomical traits, such as vessel wall thickness, suggested that brachyblast wood has a higher resistance to implosion due to drought-induced embolism. Finally, peculiar anatomy of brachyblast and dolichoblast wood results in the formation of so-called false rings. Hence, wood rings in C. incanus should be considered to be “seasonal” rather than “annual”.

Stochastic changes affect Solanum wild species following autopolyploidization

Polyploidy is very common within angiosperms, and several studies are in progress to ascertain the effects of early polyploidization at the molecular, physiological, and phenotypic level. Extensive studies are available only in synthetic allopolyploids. By contrast, less is known about the consequences of autopolyploidization. The current study aimed to assess the occurrence and extent of genetic, epigenetic, and anatomical changes occurring after oryzaline-induced polyploidization of Solanum commersonii Dunal and Solanum bulbocastanum Dunal, two diploid (2n=2×=24) potato species widely used in breeding programmes. Microsatellite analysis showed no polymorphisms between synthetic tetraploids and diploid progenitors. By contrast, analysis of DNA methylation levels indicated that subtle alterations at CG and CHG sites were present in tetraploids of both species. However, no change occurred concurrently in all tetraploids analysed with respect to their diploid parent, revealing a stochastic trend in the changes observed. The morpho-anatomical consequences of polyploidization were studied in leaf main veins and stomata. With only a few exceptions, analyses showed no clear superiority of tetraploids in terms of leaf thickness and area, vessel number, lumen size and vessel wall thickness, stomata pore length and width, guard cell width, and stomatal density compared with their diploid progenitors. These results are consistent with the hypothesis that there are no traits systematically associated with autopolyploidy.

Combined histochemistry and autofluorescence for identifying lignin distribution in cell walls

Histological staining methods commonly used for detecting cellulose and lignin in cell walls were combined with epifluorescence microscopy to visualize differences in lignification between and within cellular elements. We tested our approach on sections of one-year-old branches of Fraxinus ornus L., Myrtus communis L., Olea europaea L., Pistacia lentiscus L. and Rhamnus alaternus L., containing both normal and tension wood. Sections were subjected to various staining techniques, viz. safranin O, safranin O/fast green FCF, and alcoholic solutions of safranin O/astra blue, according to the commonly accepted protocols. Stained and unstained sections were compared using both light and epifluorescence microscopy. Safranin O with or without counterstaining hid the strong fluorescence of vessel walls, cell corners and middle lamellae allowing the secondary wall fibers to fluoresce more clearly. Epifluorescence microscopy applied to stained sections showed more cell wall details than autofluorescence of unstained sections or white light microscopy of counterstained sections. This simple approach proved reliable and valuable for detecting differences in lignification in thick sections without the need for costly equipment.

Xylem Development and Cell Wall Changes of Soybean Seedlings Grown in Space

BACKGROUND AND AIMS Plants growing in altered gravity conditions encounter changes in vascular development and cell wall deposition. The aim of this study was to investigate xylem anatomy and arrangement of cellulose microfibrils in vessel walls of different organs of soybean seedlings grown in Space. METHODS Seeds germinated and seedlings grew for 5 d in Space during the Foton-M2 mission. The environmental conditions, other than gravity, of the ground control repeated those experienced in orbit. The seedlings developed in space were compared with those of the control test on the basis of numerous anatomical and ultrastructural parameters such as number of veins, size and shape of vessel lumens, thickness of cell walls and deposition of cellulose microfibrils. KEY RESULTS Observations made with light, fluorescence and transmission electron microscopy, together with the quantification of the structural features through digital image analysis, showed that the alterations due to microgravity do not occur at the same level in the various organs of soybean seedlings. The modifications induced by microgravity or by the indirect effect of space-flight conditions, became conspicuous only in developing vessels at the ultrastructural level. The results suggested that the orientation of microfibrils and their assembly in developing vessels are perturbed by microgravity at the beginning of wall deposition, while they are still able to orient and arrange in thicker and ordered structures at later stages of secondary wall deposition. CONCLUSIONS The process of proper cell-wall building, although not prevented, is perturbed in Space at the early stage of development. This would explain the almost unaltered anatomy of mature structures, accompanied by a slower growth observed in seedlings grown in Space than on Earth.