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Ericoid mycorrhizal fungi are common root associates of a Mediterranean ectomycorrhizal plant (Quercus ilex).

Mycorrhiza samples of neighbouring Quercus ilex and Erica arborea plants collected in a postcutting habitat were processed to see whether plants differing in mycorrhizal status harbour the same root endophytes. Three experiments were performed in parallel: (i) isolation, identification and molecular characterization of fungi from surface-sterilized roots of both plant species; (ii) re-inoculation of fungal isolates on axenic E. arborea and Q. ilex seedlings; (iii) direct inoculation of field-collected Q. ilex ectomycorrhizas onto E. arborea seedlings. About 70 and 150 fungal isolates were obtained from roots of Q. ilex and E. arborea, respectively. Among them, Oidiodendron species and five cultural morphotypes of sterile isolates formed typical ericoid mycorrhizas on E. arborea in vitro. Fungi with such mycorrhizal ability were derived from both host plants. Isolates belonging to one of these morphotypes (sd9) also exhibited an unusual pattern of colonization, with an additional extracellular hyphal net. Ericoid mycorrhizas were also readily obtained by direct inoculation of E. arborea seedlings with Q. ilex ectomycorrhizal tips. Polymerase chain-restriction fragment length polymorphism and random amplified polymorphic DNA analyses of the shared sterile morphotypes demonstrate, in the case of sd9, the occurrence of the same genet on the two host plants. These results indicate that ericoid mycorrhizal fungi associate with ectomycorrhizal roots, and the ecological significance of this finding is discussed.

DNA, Ribosomal Spacer↗

Cytostatic and cytocidal effects of mistletoe (Viscum album L.) quercus extract Iscador.

Mistletoe (Viscum album) extract (Iscador) is used either alone or in combination with chemotherapy or radiotherapy in the treatment of tumor patients. In this study the effects of Viscum album Quercus (VA Qu) extract (1000 ng lectin and 6 microg viscotoxin/ml) in various doses were investigated in an in vitro model with tumor cells: three multiple myeloma (MM) cell lines (OPM-2, RPMI-8226, U-266) and three B cell lymphoma cell lines (U-698, DOHH-2, WSU-1). None of the three B lymphoma cell lines and none of the three multiple myeloma cell lines produced interleukin (IL)-6 spontaneously or after treatment with VA Qu extract. All three multiple myeloma cell lines expressed surface IL-6R and gp130, the B cell lymphomas expressed only gp130. Viscum album/Qu extract markedly downregulated the membrane expression of IL-6R and gp130 in RPMI-8226 (down to 29% and 32%) and the expression of gp130 in WSU-1 (down to 22%). There was a marked reduction of viable cells of RPMI-8226 (down to 28%) and WSU-1 (down to 8 %) at 100 microg/10(6) cells /ml. There was a clear relationship between the inhibition of proliferation and viability: the percentage reductions of the viable cells at 48 and 72 h were similar to those of proliferation at 24 and 48 h, respectively. This means that firstly the proliferation of the tumor cell is inhibited and then afterwards these cells die by apoptosis or necrosis. The inhibitory effect of VA Qu on the proliferation can be termed cytostatic, on the survival cytocidal. VA Qu was more effective in cells having a high proliferation rate than in those with a low proliferation rate. The effective dose range lay between 25 and 100 microg/10(6) cells/ml (5-20 ng lectin/10(6) cells/ml) for all parameters.

Antineoplastic Agents, Phytogenic↗

alpha-Glycosidase inhibitory activity of hexagalloylglucose from the galls of Quercus infectoria.

Hexagalloylglucose (3-O-digalloyl-1,2,4,6-tetra-O-galloyl-beta-D- glucose), which was isolated from the methanol extract of the galls of Quercus infectoria, significantly inhibited alpha-glycosidases such as sucrase, maltase and isomaltase. Its inhibitory activity was comparable to acarbose being used as a hypoglycemic agent, while the inhibitory activity on alpha-amylase was approximately 10 times lower than that of acarbose. The results indicate that, when compared to acarbose, hexagalloylglucose might reduce the side effects by reducing inhibition of alpha-amylase.

Enzyme Inhibitors↗

Different sources of reduced carbon contribute to form three classes of terpenoid emitted by Quercus ilex L. leaves.

Quercus ilex L. leaves emit terpenes but do not have specialized structures for terpene storage. We exploited this unique feature to investigate terpene biosynthesis in intact leaves of Q. ilex. Light induction allowed us to distinguish three classes of terpenes: (i) a rapidly induced class including alpha-pinene; (ii) a more slowly induced class, including cis-beta-ocimene; and (iii) the most slowly induced class, including 3-methyl-3-buten-1-ol. Using 13C, we found that alpha-pinene and cis-beta-ocimene were labeled quickly and almost completely while there was a delay before label appeared in linalool and 3-methyl-3-buten-1-ol. The acetyl group of 3-methyl-3-buten-1-yl acetate was labeled quickly but label was limited to 20% of the moiety. It is suggested that the ocimene class of monoterpenes is made from one or more terpenes of the alpha-pinene class and that both classes are made entirely from reduced carbon pools inside the chloroplasts. Linalool and 3-methyl-3-buten-1-ol are made from a different pool of reduced carbon, possibly in nonphotosynthetic plastids. The acetyl group of the 3-methyl-3-buten-1-yl acetate is derived mostly from carbon that does not participate in photosynthetic reactions. Low humidity and prolonged exposure to light favored ocimenes emission and induced linalool emission. This may indicate conversion between terpene classes.

Journal Article↗

Effect of tannin-rich leaves of oak (Quercus incana) on various microbial enzyme activities of the bovine rumen.

1. The objective of the present experiment was to study the effects of oak (Quercus incana) leaves rich in tannins on various enzyme activities of the bovine rumen. 2. The procedure employed was incubation of tannin-rich, very-low-tannin or virtually tannin-free leaves in nylon-gauze bags in the rumen, and determination of enzyme activities in microbes tightly bound to the solid matrix and in microbes loosely plus tightly attached to the solid matrix. 3. The activities of urease (EC 3.5.1.5), carboxymethylcellulose, glutamate dehydrogenase (EC 1.4.1.2) and alanine aminotransferase (glutamic-pyruvic transaminase) (EC 2.6.1.2) were significantly lower in the tannin-rich group, whereas the activities of glutamate ammonia ligase (glutamine synthetase) (EC 6.3.1.2; both gamma-glutamyltransferase (EC 2.3.2.2) and the forward reaction) were higher in the tannin-rich group. These changes were more marked in micro-organisms tightly bound to the solid matrix than in the more complex microbial compartment. 4. The protein, DNA and RNA contents, and protein: RNA ratio, were significantly lower in the tannin-rich group, whereas no difference was observed for protein: DNA between the groups. 5. Effects of tannin-containing extracts of oak leaves on various rumen enzymes in vitro showed a trend similar to that observed in nylon-gauze bags, suggesting that the changes observed in various compartments were due to the tannins of oak leaves.

Alanine Transaminase↗

Histology of Organogenic and Embryogenic Responses in Cotyledons of Somatic Embryos of Quercus Suber L.

In cork oak (Quercus suber L.), recurrent embryogenesis is produced in vitro through autoembryony without exogenous plant growth regulators (PGRs); secondary embryos appear on the embryo axis but seldom on cotyledons. Focusing mainly on the histological origin of neoformations, we investigated the influence of the embryo axis and exogenous PGRs on the embryogenic potential of somatic embryo cotyledons. Isolated cotyledons of somatic embryos became necrotic when cultured on PGR-free medium but gave secondary embryos when cultured on media containing benzyladenine and naphthaleneacetic acid. Cotyledons of cork oak somatic embryos are competent to give embryogenic responses. Isolated cotyledons without a petiole showed a lower percentage of embryogenic response than did those with a petiole. In petioles, somatic embryos arose from inner parenchyma tissues following a multicellular budding pattern. Joined to the embryo axis, cotyledons did not show morphogenic responses when cultured on PGR-free medium but revealed budlike and phylloid formations when cultured on medium with PGRs. The different morphogenic behavior displayed by somatic cotyledons indicates an influence of the embryo axis and indicates a relationship between organogenic and embryogenic regeneration pathways.

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Intra-specific variation in xylem cavitation in interior live oak (Quercus wislizenii A. DC.).

Xylem cavitation induced by water stress reduces plant hydraulic conductance and can indicate the habitat a species evolved in and its phylogenetic background. Species differ widely in cavitation resistance, but less is known about intra-specific variation. Cavitation resistance was assessed for field-collected adult and sapling size classes from three populations of interior live oak (Quercus wislizenii A. DC.) in California, USA. Root and stem cavitation resistance of two-year old seedlings from a greenhouse experiment was also measured. Cavitation resistance curves were determined by injecting air into the vascular system to induce cavitation and measuring the subsequent decline in hydraulic conductance. Based on the air-seeding hypothesis, the absolute value of the air pressures should be equivalent to the tensions that cause cavitation under dehydrating conditions. Conductance declined exponentially with applied pressure for both roots and stems. Comparisons between populations did not reveal significant differences despite good statistical power. The 50% loss in conductance point occurred between 1.0-1.6 MPa; conductance declined more slowly thereafter. Conductance was 21-30% of maximum at 4.0 MPa and 7-14% at 8.0 MPa. Saplings exhibited a nearly identical pattern compared with adults except at 4.0 MPa, where saplings exhibited slightly less cavitation (7%). Greenhouse seedling stems were more resistant compared with both field-collected adults and with seedling roots. The 50% loss in conductance point occurred at 0.83 and 2.6 MPa for seedling roots and stems, respectively. Seedling stems maintained conductance of 20.9% at 8.0 MPa while most roots were fully cavitated between 5.0-8.0 MPa.

Seasons↗

Effects of severe dehydration on leaf photosynthesis in Quercus petraea (Matt.) Liebl.: photosystem II efficiency, photochemical and nonphotochemical fluorescence quenching and electrolyte leakage.

Leaf disks of oak (Quercus petraea (Matt.) Liebl.) trees were subjected to rapid dehydration in air in the dark. Optimal photochemical efficiency of PS II (F(v)/F(M)), photochemical (q(P)) and nonphotochemical (q(NP)) quenchings of chlorophyll a fluorescence, and relative conductivity (C(r)) of leaf disk diffusate were measured in leaf disks with different water deficits (D). No effect of dehydration was detected before D reached values above 0.30. When D increased from 0.30 to 0.50, q(NP) increased without any change in q(P), which may indicate that thermal deexcitation of PS II increased, allowing reduced photochemical activity and maintenance of a large pool of oxidized primary acceptors (QA), although carbon reduction was impaired. Large changes in electron transport chain activity, leading to decreases in both q(P) and q(NP), appeared only in leaf disks subjected to severe water deficits (D > 0.60) and were correlated with a modification of membrane structure. However, stability of F(v)/F(M) indicated that the functional integrity of PS II was not altered until D reached values above 0.75. We conclude that the photosynthetic apparatus of Q. petraea is rather insensitive to leaf dehydration per se during drought under natural conditions.

Journal Article↗

Loss of hydraulic conductivity due to water stress in intact juveniles of Quercus rubra and Populus deltoides.

Despite many studies of the percent loss of hydraulic conductivity in excised branches, there is doubt as to whether cutting stems in air introduces unnatural embolism into the xylem at the cut surface. To address this question, hydraulic conductivity was measured in seedlings of northern red oak (Quercus rubra L.) and rooted scions of eastern cottonwood (Populus deltoides Bartr. ex Marsh.) that had been droughted in pots. Results indicate that in situ dehydration produced a very similar vulnerability curve (% loss of conductivity versus water potential) to those previously obtained by bench-top dehydration of excised branches of eastern cottonwood and red oak. In eastern cottonwood cuttings, conductivity loss increased sharply below water potentials of -1.0 MPa, with 100% loss of conductivity occurring by -2.0 MPa, whereas conductivity loss in red oak seedlings was more gradual, i.e., increasing below -1.5 MPa and sustaining 100% loss of conductivity by about -4.0 MPa.

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In vitro shoot proliferation determined by explant orientation of juvenile and mature Quercus rubra L.

Shoot cultures of Quercus rubra (L.) were established from both juvenile and adult plant material. Initial explants from epicormic shoots formed on the basal zone of the trunks had a greater capacity for in vitro establishment than explants from crown branches. The growth of vigorous axillary shoots was obtained by culturing decapitated shoots horizontally on Woody Plant Medium supplemented with 0.2 mg l(-1) of 6-benzylaminopurine. After 3 weeks of culture the shoots were transferred to fresh medium for two more weeks, giving a 5-week multiplication cycle. Efficient shoot production was achieved by combining three treatments favoring the growth of lateral buds: excision of the apex, horizontal culture and cytokinin treatment. The addition of indoleacetic acid or indolebutyric acid to the multiplication medium did not improve shoot proliferation rates, and naphthaleneacetic acid was detrimental. Recycling the same explant for several successive subcultures improved the efficiency of the propagation procedure. Using the optimal multiplication procedures, nine clones (six of juvenile origin and three from adult trees) were tested in vitro and it was found that genotype and age affected performance.

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Seasonal patterns of light-saturated photosynthesis and leaf conductance for mature and seedling Quercus rubra L. foliage: differential sensitivity to ozone exposure.

Extrapolation of the effects of ozone on seedlings to large trees and forest stands is a common objective of current assessment activities, but few studies have examined whether seedlings are useful surrogates for understanding how mature trees respond to ozone. This two-year study utilized a replicated open-top chamber facility to test the effects of subambient, ambient and twice ambient ozone concentrations on light-saturated net photosynthesis (P(max)) and leaf conductance (g(l)) of leaves from mature trees and genetically related seedlings of northern red oak (Quercus rubra L.). Gas exchange measurements were collected four times during the 1992 and 1993 growing seasons. Both P(max) and g(l) of all foliage followed normal seasonal patterns of ontogeny, but mature tree foliage had greater P(max) and g(l) than seedling foliage at physiological maturity. At the end of the growing season, P(max) and g(l) of the mature tree foliage exposed to ambient ( approximately 80-100 ppm-h) and twice ambient ( approximately 150-190 ppm-h) exposures of ozone were reduced 25 and 50%, respectively, compared with the values for foliage in the subambient ozone treatment ( approximately 35 ppm-h). In seedling leaves, P(max) and g(l) were less affected by ozone exposure than in mature leaves. Extrapolations of the results of seedling exposure studies to foliar responses of mature forests without considering differences in foliar anatomy and stomatal response between juvenile and mature foliage may introduce large errors into projections of the response of mature trees to ozone.

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Influence of water relations on Quercus cerris-Hypoxylon mediterraneum interaction: a model of drought-induced susceptibility to a weakness parasite.

The association between water stress and susceptibility of Quercus cerris to the fungal parasite Hypoxylon mediterraneum was studied in field-grown trees and greenhouse-grown seedlings. Susceptibility of Q. cerris to the fungus, expressed as extension of wood discolorations, increased in field-grown trees during drought and increased in greenhouse-grown seedlings in response to limited water supply. In both seedlings and trees, spread of the fungus in the vascular system was higher when leaf water potentials were low than when leaf water potentials were high, and was significantly correlated to loss of hydraulic conductivity of xylem. The presence of the fungus in the xylem suggests that H. mediterraneum could use embolized vessels to spread in the host.

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Site-specific water relations and stomatal response of Quercus ilex in a Mediterranean watershed.

Intraspecific variations in the water relations and stomatal response of Quercus ilex L. were analyzed under field conditions by comparing trees at two locations within a Mediterranean watershed (l'Avic, Catalonia, NE Spain). Distinct environmental gradients exist between the two sites (referred to as ridge top at 975 m and valley bottom at 700 m) with greater soil depth for water storage, reduced radiation, reduced wind and higher water vapor pressure deficits at the valley bottom than at the ridge top. Osmotic adjustment and changes in tissue elasticiity did not significantly increase drought resistance in the trees studied. The leaf-to-air vapor pressure difference (Deltaw) threshold for inducing stomatal closure was higher at the ridge top (15.6 kPa MPa(-1) +/- 0.5 SE) than at the valley bottom (9.8 kPa MPa(-1) +/- 1.0 SE). However, increases in Deltaw beyond the threshold were followed by greater reductions in leaf conductance of trees at the ridge top than at the valley bottom. At both sites, maximum leaf conductance was related to predawn shoot water potential which, in turn, was related to watershed stream flow. The effects of water deficits during the dry summer of 1989 were more severe in trees at the valley bottom than at the ridge top. During periods of high evaporative demand, site-specific differences in the control of water loss led to more conservative water use by trees at the ridge top and, thus, to even greater drought avoidance (higher predawn water potentials) in late summer.

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Relationship between CO(2)-dependent O(2) evolution and photosystem II activity in oak (Quercus petraea) trees grown in the field and in seedlings grown in ambient or elevated CO(2).

The light-response of the apparent quantum yield of photosynthetic O(2) evolution (Phi(O(2) )) under non-photorespiratory conditions was measured together with the photochemical efficiency of PS II (DeltaF/F(m)'), the photochemical efficiency of open PS II reaction centers (F(v)'/F(m)') and the photochemical fluorescence quenching (q(p)) of leaf disks punched from oak leaves of seedlings grown in ambient (350 micro mol mol(-1)) or elevated (700 micro mol mol(-1)) CO(2) in a greenhouse, and from sunlit leaves of mature oak trees (Quercus petraea (Matt.) Liebl.). There were marked differences between seedlings and trees. In seedlings, CO(2) concentration during growth did not modify the light response of photosynthesis or PS II activity. There was a single linear relationship between Phi(O(2) ) and DeltaF/F(m)' in seedling leaves that was independent of the CO(2) concentration imposed during growth. In contrast, this relationship was curvilinear in sunlit leaves of adult trees. In seedling leaves, the decrease in q(p) (i.e., the proportion of open PS II reaction centers) largely accounted for the decrease in DeltaF/F(m)', whereas the decrease in DeltaF/F(m)' in sunlit leaves of mature oak trees was dependent on both q(p) and F(v)'/F(m)'.

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Changes in respiration and chemical content during autumnal senescence of Populus tremuloides and Quercus rubra leaves.

Changes in respiration rate, chemical content and chemical concentration were measured in leaves of field-grown Populus tremuloides Michx. and Quercus rubra L. trees throughout the growing season and autumnal senescence. Chlorophyll, soluble sugar, N, P, K and Mg contents and concentrations all declined during leaf senescence, whereas Ca content and concentration increased. Leaf dry mass per area declined 24 and 35% in P. tremuloides and Q. rubra, respectively, during senescence. In leaves of both species, respiration rates peaked during leaf expansion in the spring and then declined, as a result of reduced cytochrome-mediated respiration, to reach relatively constant rates by midsummer. In senescing P. tremuloides leaves, respiration rates remained relatively constant until mid-October and then declined rapidly. In senescing Q. rubra leaves, respiration rates increased in late September, as a result of the appearance of residual respiration that could not be reduced by respiratory inhibitors, and then declined quickly in early November. No changes in alternative pathway respiratory activity were observed in leaves of either species during senescence until late autumn when rates declined. Because respiration rates were correlated with both leaf sugar and nitrogen content during leaf senescence, we conclude that respiration rates were maintained or increased during leaf senescence to supply energy for degradation and mobilization of chemical constituents.

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Effects of thinning on soil and tree water relations, transpiration and growth in an oak forest (Quercus petraea (Matt.) Liebl.).

To quantify the effects of crown thinning on the water balance and growth of the stand and to analyze the ecophysiological modifications induced by canopy opening on individual tree water relations, we conducted a thinning experiment in a 43-year-old Quercus petraea stand by removing trees from the upper canopy level. Soil water content, rainfall interception, sap flow, leaf water potential and stomatal conductance were monitored for two seasons following thinning. Seasonal time courses of leaf area index (LAI) and girth increment were also measured. Predawn leaf water potential was significantly higher in trees in the thinned stand than in the closed stand, as a consequence of higher relative extractable water in the soil. The improvement in water availability in the thinned stand resulted from decreases in both interception and transpiration. From Year 1 to Year 2, an increase in transpiration was observed in the thinned stand without any modification in LAI, whereas changes in transpiration in the closed stand were accompanied by variations in LAI. The different behaviors of the closed and open canopies were interpreted in terms of coupling to the atmosphere. Thinning increased inter-tree variability in sap flow density, which was closely related to a leaf area competition index. Stomatal conductance varied little inside the crown and differences in stomatal conductance between the treatments appeared only during a water shortage and affected mainly the closed stand. Thinning enhanced tree growth as a result of a longer growing period due to the absence of summer drought and higher rates of growth. Suppressed and dominant trees benefited more from thinning than trees in the codominant classes.

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Diurnal changes in photoprotective mechanisms in leaves of cork oak (Quercus suber) during summer.

Daily variations in photoprotective mechanisms were studied in sun and shade leaves of 40-year-old cork oak (Quercus suber L.) trees during early summer in Portugal. Although trees were not severely water stressed because predawn leaf water potentials remained high, photosynthesis and stomatal conductance decreased at midday. The midday depression in gas exchange was not reversed by short-term exposure to "optimal" conditions of temperature, light and vapor pressure deficit. Chlorophyll a fluorescence, maximum photochemical yield of photosystem II and the quantum yield of noncyclic electron transport showed midday depressions, but recovered by the evening. Both short-term changes in the components of the xanthophyll cycle (reversible de-epoxidation of violaxanthin during the day) as well as long-term changes (higher xanthophyll content in sun compared with shade leaves) were detected and may play a role in the dissipation of excess energy at midday. Because the activities of enzymes of the antioxidant system, superoxide dismutase and ascorbate peroxidase, were high enough to cope with the increase in oxygen reactive species likely to arise under the stressful conditions of midday, we conclude that these enzymes may provide an additional mechanism for energy dissipation.

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Hay-scented fern (Dennstaedtia punctilobula (Michx.) Moore) interference with growth of northern red oak (Quercus rubra L.) seedlings.

We assessed the impacts of hay-scented fern (Dennstaedtia punctilobula (Michx.) Moore) and subsoil liming (CaO amendments) on root and shoot growth of greenhouse-grown, first-year, northern red oak (Quercus rubra L.) seedlings. Red oak seedlings and ferns were grown in reconstructed soil profiles of four common Pennsylvanian forest soils. When grown in the presence of hay-scented ferns, with or without subsoil liming, red oak seedlings had significantly reduced height growth, and foliar, stem and total root biomass. Fern foliar biomass was significantly reduced when ferns were grown with red oaks, but there was no significant difference in total belowground biomass of ferns. Belowground fern biomass was concentrated in the upper soil profile, whereas red oak roots showed a variety of distributions. In the presence of ferns, fine root branching in red oak was reduced in the organic horizons of three of the four soils tested. In both the presence and absence of ferns, root branching in red oak was also significantly and negatively correlated with the concentration of 0.01 M SrCl(2)-extractable aluminum in the mineral horizons (r(2) = 0.77). Subsoil liming generally improved root branching in red oaks. The presence of ferns significantly reduced ectomycorrhizal infection frequency in red oak. We conclude that hay-scented fern inhibited root branching and suppressed above- and belowground biomass accumulation of first-year northern red oak seedlings.

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