Miren K. Duñabeitia

Differential responses of three fungal species to environmental factors and their role in the mycorrhization of Pinus radiata D. Don

Three ectomycorrhizal (ECM) isolates of Rhizopogon luteolus, R. roseolus and Scleroderma citrinum were found to differ markedly in their in vitro tolerance to adverse conditions limiting fungal growth, i.e. water availability, pH and heavy metal pollution. S. citrinum was the most sensitive, R. luteolus intermediate and R. roseolus the most tolerant species. Pinus radiata D. Don seedlings were inoculated in the laboratory and in a containerised seedling nursery with spore suspensions of the three ECM species. Colonisation percentage was considerably lower under nursery conditions, probably due to competition by native fungi. The effects of nursery ECM inoculation on seedling growth depended on the fungal species. Only R. roseolus-colonised plants showed a significantly higher shoot growth than non-mycorrhizal plants. All three fungi induced significantly higher root dry weights relative to control plants. Despite the low mycorrhizal colonisation, mycorrhization with all three species improved the physiological status of nursery-grown seedlings, e.g. enhanced root enzyme activity, shoot nutrient and pigment content, net photosynthesis rate and water use efficiency. Of the three fungal species, R. roseolus was the most effective; this species was also the most adaptable and showed the greatest range of tolerance to adverse environmental conditions in pure culture. It is, therefore, proposed as a promising fungal species for ECM inoculation of P. radiata in the nursery.

Role of Photoprotective Systems of Holm-Oak (Quercus ilex) in the Adaptation to Winter Conditions

Summary Holm-oak (Quercus ilex L.) is a Mediterranean evergreen sclerophyllous species adapted to a wide range of environmental conditions and consequently its leaves must be able to withstand variable stresses. Changes in pigment and antioxidant composition were assessed in relation to chilling stress, light environment and leaf aging. Photosynthetic capacity during winter was two-fold higher than in spring. However, the photochemical efficiency of PSII was reduced in sun leaves probably because of an increased rate of thermal energy dissipation related to the superior zeaxanthin concentration. Lutein, neoxanthin, β-carotene and the xanthophyll cycle pool increased during winter especially in sun leaves, while lutein epoxide and α-carotene decreased markedly. The main antioxidant accumulated during winter was ascorbate, while glutathione and α-tocopherol were not seasonally affected. Catalase increased in spring as a consequence of higher photorespiration rates, while GR and MDHAR activities were higher in sun leaves during winter. The high MDHAR activity indicates an important role for this enzyme in the adaptation to environmental factors associated to winter. The present study demonstrates that holm-oak is able to acclimate to a variety of environmental stress factors by inducing dramatic changes in their photosynthetic characteristics, pigment composition and antioxidant defences.

Field performance of Pinus radiata D. Don produced in nursery with different types of containers

We have studied the development in nurseries of containerized Pinus radiata produced with different container systems in order to choose the most suitable system for producing well-balanced plants with an optimal root system. At the end of the production period, significant differences were found in morphological responses among the seedlings to the various container characteristics. Seedlings grown in containers that permitted lateral air puning presented less growth and lower biomass production. However, root deformations were more frequent and severe in plants produced in closed-wall containers. Field performance was likewise mainly affected by container type and plant growth rate, as faster grown plants showed more problems of stability than plants with a balanced root and stem development.

Ectomycorrhizae synthesized between Pinus radiata and eight fungi associated with Pinus spp.

The capability of several fungal species, collected in the field in association with species of the genus Pinus, to form ectomycorrhizae with Pinus radiata have been determinated in vitro and in con- tainers. Hebeloma longicaudum, Russula sanguinea, Tricholoma albobruneum, Tuber borchii and Xerocomus badius have formed ectomycorrhizae with Pinus ra- diata by pure culture synthesis. These species and other three, Boletus pinophilus, Gautieria othii and Scleroderma polyrhizum, have formed ectomycorrhizae in open containers by methods of vegetative mycelial inoculation and spore inoculation. These eight sym- bioses with Pinus radiata have not been previously described.

Spatial and temporal dynamics of the colonization of Pinus radiata by Fusarium circinatum, of conidiophora development in the pith and of traumatic resin duct formation

· Fusarium circinatum causes pitch canker disease in a wide range of pine trees, including Pinus radiata, with devastating economic consequences. · To assess the spatial and temporal dynamics of growth of this pathogen in radiata pine, we examined the process of infection using both real-time PCR to quantify fungal biomass inside the plant host, and confocal microscopy using a green fluorescent protein (GFP)-tagged strain of F. circinatum. · Pathogen growth exhibited three distinct phases: an initial exponential increase in fungal biomass, concomitant with pathogen colonization of the cortex and phloem; a slowdown in fungal growth coincident with sporulating hyphae deep within the host; and stabilization of the fungal biomass when the first wilting symptoms appeared. The number of resin ducts in the xylem was found to increase in response to infection and the fungus grew inside both constitutive and traumatic resin ducts. · These results indicate that conidiation may contribute to the spatial or temporal dissemination of the pathogen. Moreover, the present findings raise the intriguing possibility that the generation of traumatic resin ducts may be of more benefit to the fungus than to the plant.

Sequential Effects of Acidic Precipitation and Drought on Water Relations of Pinus radiata Seedlings

Summary In this work we investigated the sequential effects of simulated acid rain and drought on water relations of radiata pine ( Pinus radiata D. Don) seedlings. Whole seedlings were firstly subjected during 1 month to acid rain, pH 3.0, applied 5 times a week at 3 mm precipitation equivalent per day. Afterwards, drought treatment was applied by withholding water for 20 days and subsequent rewatering for 6 days to analyze the sequential effect of acid precipitation and drought. Water status parameters were determined before the initiation of acid rain treatment and within 3 weeks after the end of treatment. Acid rain treatment in well-watered plants did not affect either water potential or relative water content, whereas a marked effect on electrolyte leakage from the needles and on instantaneous transpiration was recorded. Drought treatment had great effects on leaf water potential (-2.5 MPa), RWC (50 % diminution), membrane permeability (340 % increase) and transpiration rate (25 % inhibition). Interactions involving acid precipitation and drought led to much greater impacts on all of the parameters analyzed. We conclude that although acid rain had slight, if any, direct effects on the water relations of well-watered radiata pine, it altered the drought tolerance of this tree species. Under conditions of soil-water deficit, plants exposed previously to acid rain are more sensitive to drought and desiccation.

Effect of Photorespiratory C2 Acids on CO2 Assimilation, PS II Photochemistry and the Xanthophyll Cycle in Maize

The photorespiration cycle plays an important role in avoiding carbon drainage from the Calvin cycle and in protecting plants from photoinhibition. The role of photorespiration is frequently underestimated in C4 plants, since these are characterized by low photorespiration rates. The aim of this work was to study the relationship between CO2 assimilation, PS II photochemistry and the xanthophyll cycle when the photorespiratory cycle is disrupted in Zea mays L. To this end, the photorespiration inhibitor phosphinothricin (PPT) was applied individually or together with the photorespiratory C2 acids, glycolate and glyoxylate to maize leaves. Application of PPT alone led to the inhibition of CO2 assimilation. Moreover, feeding with glycolate or glyoxylate enhanced the effect of PPT on CO2 assimilation. Our results confirm that the avoidance of the accumulation of the photorespiratory metabolites glycolate, glyoxylate or phosphoglycolate, is of vital importance for coordinated functioning between the glycolate pathway and CO2 assimilation. Relatively early changes in PS II photochemistry also took place when the photorespiratory cycle was interrupted. Thus, fluorescence photochemical quenching (qP) was slightly reduced (10%) due to the application of PPT together with glycolate or glyoxylate. A decrease in the efficiency of excitation-energy capture by open PS II reaction centres (F′v/F′m) and an increase in thermal energy dissipation (non-photochemical quenching, NPQ) were also measured. These observations are consistent with a limitation of activity of the Calvin cycle and a subsequent lower demand for reduction equivalents. The increase in NPQ is discussed on the basis of changes in the xanthophyll cycle in maize, which seem to provide a limited protective role to avoid photoinhibition when the glycolate pathway is blocked. We conclude that C2 photorespiratory acids can act as physiological regulators between the photorespiratory pathway and the Calvin cycle in maize

15N NATURAL ABUNDANCE EVIDENCES A BETTER USE OF N SOURCES BY LATE NITROGEN APPLICATION IN BREAD WHEAT

This work explores whether the natural abundance of N isotopes technique could be used to understand the movement of N within the plant during vegetative and grain filling phases in wheat crop (Triticum aestivum L.) under different fertilizer management strategies. We focus on the effect of splitting the same N dose through a third late amendment at flag leaf stage (GS37) under humid Mediterranean conditions, where high spring precipitations can guarantee the incorporation of the lately applied N to the soil-plant system in an efficient way. The results are discussed in the context of agronomic parameters as N content, grain yield and quality, and show that further splitting the same N dose improves the wheat quality and induces a better nitrogen use efficiency. The nitrogen isotopic natural abundance technique shows that N remobilization is a discriminating process that leads to an impoverishment in 15N of senescent leaves and grain itself. This technique also reflects the more efficient use of N resources (fertilizer and native soil-N) when plants receive a late N amendment.