V. De Micco

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.

Intra-annual density fluctuations in tree rings: how, when, where, and why?

Intra-annual density fluctuations (IADFs) in tree rings are generally considered structural anomalies caused by deviations from the “normal course” of xylogenesis during the growing season. This definition is based on the bias that, under “normal conditions”, cambial activity stops once a year. Each tree ring can thus be dated to one calendar year, which is one of the principles of dendrochronology. The formation of IADFs can be triggered directly by environmental changes, especially in precipitation and temperature, that affect cambial activity and cell differentiation. It can also be the result of limited photosynthesis, due to defoliation induced by biotic or abiotic constraints.Often indicated with alternative terms, IADFs were first described in the 1930s, and recently reported for many trees and shrubs from different ecosystems throughout the world, particularly for Mediterranean species. Different types of IADFs have been detected; their formation and structural properties depend on many factors including tree genotype, age, size, rooting depth, habitat, soil, climate, photosynthetic activity, and allocation strategies. Whether IADFs affect the adaptive capability of plants remains, however, unclear.We provide an overview of the main anatomical features of IADFs and their occurrence in tree rings from various environments and climatic regimes. We propose a simplified way of classifying them and discuss the hypotheses about their functional role and the factors triggering their formation. To understand the ecological role of IADFs better, we recommend a multidisciplinary approach, involving wood anatomy, dendroecology, and stable isotopes, which has already been applied for Mediterranean species. We conclude by considering that IADFs appear to be the “rule” rather than “anomalies” in some ecosystems where they help plants cope with fluctuating environmental conditions. Moreover, their anatomical structure represents a valuable proxy of past climatic conditions at a sub-seasonal resolution and may be relevant to adapt hydraulic functioning of living trees to changing climatic conditions.

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.

Fire influence on Pinus halepensis: wood responses close and far from the scars

Tree rings provide information about environmental change through recording stress events, such as fires, that can affect their growth. The aim of this study was to investigate wood growth reactions in Pinus halepensis Mill. trees subjected to wildfires, by analysing anatomical traits and carbon and oxygen isotope composition. The study area was Southern France where two sites were selected: one subjected to fires in the last 50 years, the other characterised by comparable environmental conditions although not affected by fire events (control site). We analysed whether wood growth depends on the tangential distance between developing xylem cells and the limit where the cambium was directly damaged by fire. In the burnt site, thick wood sections, including fire-scar, were taken from surviving plants. Digital photo-micrographs were analysed to measure early- and latewood width, wood density, and tracheid size. Anatomical and isotopic traits were analysed in two series of tree rings (5 rings before and 5 after the fire) selected at different positions along the circumference (close or far from the scar). Anatomical and isotopic traits were quantified also on tree rings of the same years from cored trees growing at the control site. Results showed different wood reaction tendencies depending on the distance from the scar. The comparison between plants from the two sites allowed to exclude possible climate interference.Our results are discussed in terms of two kinds of growth reactions: the local need to promptly compartmentalise the scarred cambial zone and sapwood after fire, and the general growth perturbations due to tree reaction to crown scorch during fire. Anatomical results, combined with dendrochronological and isotopic analysis, could provide an efficient way to distinguish between direct growth reactions due to heat-related damage on cambium and indirect outcomes related to defoliation.

Leaf Anatomy and Photochemical Behaviour of Solanum lycopersicum L. Plants from Seeds Irradiated with Low-LET Ionising Radiation

Plants can be exposed to ionising radiation not only in Space but also on Earth, due to specific technological applications or after nuclear disasters. The response of plants to ionising radiation depends on radiation quality/quantity and/or plant characteristics. In this paper, we analyse some growth traits, leaf anatomy, and ecophysiological features of plants of Solanum lycopersicum L. “Microtom” grown from seeds irradiated with increasing doses of X-rays (0.3, 10, 20, 50, and 100 Gy). Both juvenile and compound leaves from plants developed from irradiated and control seeds were analysed through light and epifluorescence microscopy. Digital image analysis allowed quantifying anatomical parameters to detect the occurrence of signs of structural damage. Fluorescence parameters and total photosynthetic pigment content were analysed to evaluate the functioning of the photosynthetic machinery. Radiation did not affect percentage and rate of seed germination. Plants from irradiated seeds accomplished the crop cycle and showed a more compact habitus. Dose-depended tendencies of variations occurred in phenolic content, while other leaf anatomical parameters did not show distinct trends after irradiation. The sporadic perturbations of leaf structure, observed during the vegetative phase, after high levels of radiation were not so severe as to induce any significant alterations in photosynthetic efficiency.

Root structure of Rumex scutatus growing on slopes.

The effect of slope on root architecture and anatomy of Rumex scutatus L. was analysed in plants growing on slopes and on flat ground, on the flanks of Mt. Vesuvius. Roots of plants on steep slopes developed asymmetrically, with lateral roots growing upslope acting as tie-rods. In roots growing on flat ground, lignified xylem was limited to thin radial strips, surrounded by unlignified parenchyma cells filled with starch. A trend towards a larger lignified area was evident from plants growing on flat ground to those on steep slopes, as well as from taproots to tie-rod roots. The latter also showed larger vessels and a lower incidence of narrow vessels than taproots. Roots developing on steep slopes were characterised by high frequency of very-thick-walled, lignified fibres with long tips which accumulated both starch and phenolic compounds. Such fibres did not show lignified cell walls in roots growing on flat ground. Overall analysis showed that slope influences root architecture and anatomical traits, not only affecting mechanical properties but also water transport capacity.

Xylogenesis reveals the genesis and ecological signal of IADFs in Pinus pinea L. and Arbutus unedo L.

Background and Aims Mediterranean trees have patterns of cambial activity with one or more pauses per year, leading to intra-annual density fluctuations (IADFs) in tree rings. We analysed xylogenesis (January 2015-January 2016) in Pinus pinea L. and Arbutus unedo L., co-occurring at a site on Mt. Vesuvius (southern Italy), to identify the cambial productivity and timing of IADF formation. Methods Dendrochronological methods and quantitative wood anatomy were applied and enabled IADF identification and classification. Key Results We showed that cambium in P. pinea was productive throughout the calendar year. From January to March 2015, post-cambial (enlarging) earlywood-like tracheids were observed, which were similar to transition tracheids. The beginning of the tree ring was therefore not marked by a sharp boundary between latewood of the previous year and the new xylem produced. True earlywood tracheids were formed in April. L-IADFs were formed in autumn, with earlywood-like cells in latewood. In A. unedo, a double pause in cell production was observed, in summer and winter, leading to L-IADFs in autumn as well. Moreover, the formation of more than one IADF was observed in A. unedo. Conclusions Despite having completely different wood formation models and different life strategies, the production of earlywood, latewood and IADF cells was strongly controlled by climatic factors in the two species. Such cambial production patterns need to be taken into account in dendroecological studies to interpret climatic signals in wood from Mediterranean trees.

Anatomy, photochemical activity, and DNA polymorphism in leaves of dwarf tomato irradiated with X-rays

The response of higher plants to ionising radiation depends on factors related to both radiation properties and plant features including species, cultivar, age, and structural complexity of the target organ. Adult plants of dwarf tomato were irradiated with different doses of X-rays to investigate possible variations in leaf morpho-anatomical traits, photosynthetic efficiency, and genomic DNA. In order to assess if and how responses depend on leaf developmental stage, we analysed two types of leaves; nearly mature leaves (L1) and actively developing leaves (L2), whose lamina size corresponded to 70 and 25 %, respectively, of the lamina size of the fully expanded leaves. The results show that the X-rays prevented full lamina expansion of the L2 leaves at all doses and induced early death of tissue of plants irradiated with doses higher than 20 Gy. Most anatomical modifications were not clearly dose-dependent and the radiation-induced increase in phenolic compounds was irrespective of dose. At high doses of X-rays (50 and 100 Gy), photochemical efficiency decreased significantly in both leaf types, whereas total chlorophyll content significantly decreased only in the L2 leaves. The random amplification of polymorphic DNA data show that the X-rays induced mutagenic effects in the L2 leaves even at low doses despite the absence of severe phenotypic alterations. Genetic structure found in the population of samples corroborates the results of anatomical and eco-physiological analyses: the 20 Gy dose seems to mark the threshold dose above which genetic alterations, structural anomalies, and perturbations in the photosynthetic apparatus become significant, especially in the actively expanding leaves.