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Dielectric relaxation of water and water-plasticized biomolecules in relation to cellular water organization, cytoplasmic viscosity, and desiccation tolerance in recalcitrant seed tissues.

To understand the relationship between the organization of cellular water, molecular interactions, and desiccation tolerance, dielectric behaviors of water and water-plasticized biomolecules in red oak (Quercus rubra) seeds were studied during dehydration. The thermally stimulated current study showed three dielectric dispersions: (a) the relaxation of loosely-bound water and small polar groups, (b) the relaxation of tightly-bound water, carbohydrate chains, large polar groups of macromolecules, and (c) the "freezing in" of molecular mobility (glassy state). Seven discrete hydration levels (water contents of 1.40, 0.55, 0.41, 0.31, 0.21, 0.13, and 0.08 g/g dry weight, corresponding to -1.5, -8, -11, -14, -24, -74, and -195 MPa, respectively) were identified according to the changes in thermodynamic and dielectric properties of water and water-plasticized biomolecules during dehydration. The implications of intracellular water organization for desiccation tolerance were discussed. Cytoplasmic viscosity increased exponentially at water content < 0.40 g/g dry weight, which was correlated with the great relaxation slowdown of water-plasticized biomolecules, supporting a role for viscosity in metabolic shutdown during dehydration.

Biopolymers↗

Isoprene increases thermotolerance of fosmidomycin-fed leaves.

Isoprene is synthesized and emitted in large amounts by a number of plant species, especially oak (Quercus sp.) and aspen (Populus sp.) trees. It has been suggested that isoprene improves thermotolerance by helping photosynthesis cope with high temperature. However, the evidence for the thermotolerance hypothesis is indirect and one of three methods used to support this hypothesis has recently been called into question. More direct evidence required new methods of controlling endogenous isoprene. An inhibitor of the deoxyxylulose 5-phosphate pathway, the alternative pathway to the mevalonic acid pathway and the pathway by which isoprene is made, is now available. Fosmidomycin eliminates isoprene emission without affecting photosynthesis for several hours after feeding to detached leaves. Photosynthesis of fosmidomycin-fed leaves recovered less following a 2-min high-temperature treatment at 46 degrees C than did photosynthesis of leaves fed water or fosmidomycin-fed leaves in air supplemented with isoprene. Photosynthesis of Phaseolus vulgaris leaves, which do not make isoprene, exhibited increased thermotolerance when isoprene was supplied in the airstream flowing over the leaf. Other short-chain alkenes also improved thermotolerance, whereas alkanes reduced thermotolerance. It is concluded that thermotolerance of photosynthesis is a substantial benefit to plants that make isoprene and that this benefit explains why plants make isoprene. The effect may be a general hydrocarbon effect and related to the double bonds in the isoprene molecule.

Acclimatization↗

Production of isoprene by leaf tissue.

Isoprene production by Hamamelis virginiana L. and Quercus borealis Michx. leaves was studied. When ambient CO(2) concentrations were maintained with bicarbonate buffers, the rate of isoprene production at 125 microliters per liter of CO(2) was approximately four times that at 250 microliters per liter of CO(2). Isoprene production was drastically inhibited by 97% O(2). Dichlorodimethylphenylurea (0.1 mm), NaHSO(3) (10 mm), and alpha-hydroxy-2-pyridinemethanesulfonic acid (10 mm) inhibited isoprene production but increased the compensation point of the tissue. Isonicotinic acid hydrazide neither inhibited isoprene emission nor increased the compensation point of the tissue significantly. Inhibition of isoprene production does not seem to correlate with stomatal resistance. Isoprene was labeled by intermediates of the glycolate pathway, and similarities are noted between the biosynthesis of isoprene and that of beta-carotene.

Journal Article↗

Units of freezing of deep supercooled water in woody xylem.

The low temperature exotherms (LTE) of 1-year-old twigs of Haralson apple (Malus pumila Mill.), shagbark hickory (Carya ovata [Mill.] K. Koch), green ash (Fraxinus pennsylvanica Marsh), honey locust (Gleditsia triacanthos L.), American chestnut (Castanea dentata [Marsh] Borkh.), and red oak (Quercus rubra L.) were determined by differential thermal analysis (DTA). In one type of experiment freezing during a DTA experiment was halted for up to 2.5 hours after part of the supercooled water had frozen at temperatures between -25 and -42 C. Upon resumption of cooling the freezing started within 2 C of the stopping temperature. In a second type of experiment living and dead cells were microscopically observed in the same ray after partial freezing in the DTA apparatus. In another experiment, the LTE persisted even after tangential and radial sectioning of the twig to 0.13 millimeters. In a final experiment the LTE of a single multiseriate ray of red oak had the same shape as the LTE of wood with many uniseriate rays.These experiments confirm that the deep supercooled water in woody xylem or pith freezes in numerous independent events over a span of as much as 20 C. The units which freeze in an event are single cells or small groups of cells. Ice grows very slowly if at all from these units, and water moves very slowly from unfrozen cells to frozen ones. Deep supercooling of ray parenchyma does not require an intact ray.

Journal Article↗

Fractionation of Carbon Isotopes during Biogenesis of Atmospheric Isoprene.

The stable carbon isotope composition of isoprene emitted from leaves of red oak (Quercus rubra L.) was measured. Isoprene was depleted in (13)C relative to carbon recently fixed by photosynthesis. The difference in isotope composition between recently fixed carbon and emitted isoprene was independent of the isotopic composition of the source CO(2). beta-Carotene, an isoprenoid plant constituent, was depleted in (13)C relative to whole leaf carbon to the same degree as isoprene, but fatty acids were more depleted. Isoprene emitted from leaves fed abscisic acid was much less depleted in (13)C than was isoprene emitted from unstressed leaves. We conclude that isoprene is made from an isoprenoid precursor that is derived from acetyl-CoA made from recent photosynthate. The carbon isotope composition of isoprene in the atmosphere is likely to be slightly more negative (less (13)C) than C(3) plant material but when plants are stressed the isotopic composition could vary.

Journal Article↗

Two-generation analysis of pollen flow across a landscape. I. Male gamete heterogeneity among females.

Gene flow is a key factor in the spatial genetic structure in spatially distributed species. Evolutionary biologists interested in microevolutionary processess and conservation biologists interested in the impact of landscape change require a method that measures the real time process of gene movement. We present a novel two-generation (parent-offspring) approach to the study of genetic structure (TwoGener) that allows us to quantify heterogeneity among the male gamete pools sampled by maternal trees scattered across the landscape and to estimate mean pollination distance and effective neighborhood size. First, we describe the model's elements: genetic distance matrices to estimate intergametic distances, molecular analysis of variance to determine whether pollen profiles differ among mothers, and optimal sampling considerations. Second, we evaluate the model's effectiveness by simulating spatially distributed populations. Spatial heterogeneity in male gametes can be estimated by phiFT, a male gametic analogue of Wright's F(ST) and an inverse function of mean pollination distance. We illustrate TwoGener in cases where the male gamete can be categorically or ambiguously determined. This approach does not require the high level of genetic resolution needed by parentage analysis, but the ambiguous case is vulnerable to bias in the absence of adequate genetic resolution. Finally, we apply TwoGener to an empirical study of Quercus alba in Missouri Ozark forests. We find that phiFT = 0.06, translating into about eight effective pollen donors per female and an effective pollination neighborhood as a circle of radius about 17 m. Effective pollen movement in Q. alba is more restricted than previously realized, even though pollen is capable of moving large distances. This case study illustrates that, with a modest investment in field survey and laboratory analysis, the TwoGener approach permits inferences about landscape-level gene movements.

Genetic Variation↗

Composting is an effective treatment option for sanitization of Phytophthora ramorum-infected plant material.

AIMS: To determine the effects of heat and composting treatments on the viability of the plant pathogen Phytophthora ramorum grown on both artificial and various natural substrates. METHODS AND RESULTS: Phytophthora ramorum was grown on V8 agar, inoculated on bay laurel leaves (Umbellularia californica) and on woody tissues of coast live oak (Quercus agrifolia). Effects on growth, viability and survival were measured as a result of treatment in ovens and compost piles. Direct plating onto PARP medium and pear-baiting techniques were used to determine post-treatment viability. No P. ramorum was recovered at the end of the composting process, regardless of the isolation technique used. By using a PCR assay designed to detect the DNA of P. ramorum, we were able to conclude the pathogen was absent from mature compost and not merely suppressed or dormant. CONCLUSIONS: Some heat and composting treatments eliminate P. ramorum to lower than detectable levels on all substrates tested. SIGNIFICANCE AND IMPACT OF THE STUDY: Composting is an effective treatment option for sanitization of P. ramorum-infected plant material. Assaying for pathogen viability in compost requires a direct test capable of differentiating between pathogen suppression and pathogen elimination.

Pest Control, Biological↗

Honeydew of aphids as a source of sugar for Phlebotomus ariasi.

Wild-caught Phlebotomus ariasi Tonnoir, starved until their sugar meals had been digested, were caged for 24 h with plants or plants infested with aphids and then tested for fructose, a constituent of plant sap. No evidence was found that the flies took sap directly from nine types of plants present in their habitat. About two-thirds of flies caged with oak (Quercus ilex L.) infested with an aphid [Lachnus roboris (L.)] and about a fifth kept with leaves of the french bean (Phaseolus vulgaris L.) infested with an aphid took honeydew. Experiments with three other aphid species gave negative results. Of sandflies caught in an oak tree infested with aphids, half of the females and three-quarters of the males contained fructose. Tests on females caught in a house and grouped according to state of engorgement showed that, having taken blood, the females stop taking sugar until the bloodmeal is completely digested. It is predicted that honeydews are important in the development of Leishmania infantum Nicolle in the gut of P. ariasi.

Animals↗

Honeydew sugars in wild-caught Phlebotomus ariasi detected by high performance liquid chromatography (HPLC) and gas chromatography (GC).

Phlebotomus ariasi Tonnoir sandflies were caught in light traps hung in oak trees and in a house in the Cévennes focus of leishmaniasis in the South of France. The flies were cryopreserved either immediately on removal from the traps, or after starvation for 6-7 days, or after 6-7 days starvation followed by exposure to oak infested with the aphid genera Lachnus or Thelaxes. After transportation to the laboratory, the sandflies were thawed and aqueous extracts of the crushed flies were analysed for their carbohydrate content using high performance liquid chromatography (HPLC) and gas chromatography (GC). Starved female sandflies lacked significant amounts of any saccharides. Four types of sugar, melezitose and its hydrolysis products turanose, glucose and fructose, were observed in flies which had been starved previously and then placed with Lachnus infested oak. The results also indicate the presence of hydrolysis products of melezitose: (a) in flies previously starved and placed with Thelaxes infested oak, (b) in P.ariasi cryopreserved direct from the light traps hung in oak trees infested with Lachnus and Thelaxes, and (c) flies caught in a house. Unidentifiably small quantities of a trisaccharide were also detected in the latter groups of flies. In previous tests, sugars were detected in P.ariasi after their exposure to aphid-infested oak (Quercus ilex L.), but not when P.ariasi females were exposed to washed oak leaves without aphids. The results indicate that P.ariasi feed on melezitose and/or turanose, the main local source of which is aphid honeydew. A better understanding of sugar meal sources of sandflies using HPLC and GC techniques will assist in our understanding of sandfly/Leishmania relationships, parasite transmission and epidemiology.

Animals↗

Native oak chloroplasts reveal an ancient divide across Europe.

Glacial refugia and postglacial migration are major factors responsible for the present patterns of genetic variation we see in natural populations. Traditionally postglacial history has been inferred from fossil data, but new molecular techniques permit historical information to be gleaned from present populations. The chloroplast tRNA(Leu1) intron contains regions which have been highly conserved over a billion years of chloroplast evolution. Surprisingly, in one of these regions which has remained invariant for all photosynthetic organisms so far studied, we have found intraspecific site polymorphism. This polymorphism occurs in two European oaks, Quercus robur and Q. petraea, indicating hybridisation and introgression between them. Two distinct chloroplast types occur and are distributed geographically as eastern and western forms suggesting that these oaks are each derived from at least two separate glacial refugia.

Base Sequence↗

Potential refugia in Taiwan revealed by the phylogeographical study of Castanopsis carlesii Hayata (Fagaceae).

In this study, we examined spatial patterns of chloroplast DNA (cpDNA) variation in a total of 30 populations of Castanopsis carlesii Hayata (Fagaceae), a subtropical and temperate tree species, including 201 individuals sampled throughout Taiwan. By sequencing two cpDNA fragments using universal primers (the trnL intron and the trnV-trnM intergenic spacer), we found a total of 1663 bp and 21 polymorphic sites. These gave rise to a total of 28 cpDNA haplotypes. The level of differentiation among the populations studied was relatively high (GST = 0.723). Two ancestral haplotypes are widely distributed. The Central Mountain Ridge (CMR) of Taiwan represents an insurmountable barrier to the east-west gene flow of C. carlesii. Among the populations studied, three separated populations, at Lienhuachih, Fushan and Lichia, have high nucleotide diversity. Estimates of NST-GST for populations on both sides of the CMR indicate that no phylogeographical structure exists. According to the genealogical tree, number of rare haplotype and population genetic divergence, this study suggests that two potential refugia existed during the last glaciation: the first refugium was located in a region to the north of Hsuehshan Range (HR) and west of the CMR; the second refugium was located in south, especially southeastern Taiwan. In fact, the second refugium happens to be the same as that reported for Quercus glauca. A 'star-like' genealogy is characteristic when all haplotypes rapidly coalesce and is a general outcome of population expansion. The neutrality test and mismatch distribution also suggest demographic expansion recovering from a bottleneck.

Base Sequence↗

Quantification of pollen solute release using pollen grain column chromatography.

The impact of pollen on the respiratory mucosa was modeled by studying the process by which solutes are eluted from pollen grains. Rye grass (Lolium perenne), short ragweed (Ambrosia artemisiifolia), and white oak (Quercus alba) pollens were packed between glass wool plugs in small columns. Water was pumped through the columns and the eluate solute yield was determined by measurement of the dry solute weight. Solute separation was rapid, and concentrations and osmolalities of the eluate decreased exponentially. Theoretical initial solute concentrations were 179 g/l for rye grass, 55 g/l for short ragweed and 349 g/l for oak pollen eluates. Theoretical initial osmolarities of the same eluates were 321 mOsm/kg for rye grass, 196 mOsm/kg for ragweed and 424 mOsm/kg for oak pollen. Sequential separation of allergens (Lol p I, Amb a I, Amb a V), enzymes and proteins was demonstrated by specific assays. These observations suggest that the complex stimulus produced immediately after pollen grain hydration at the respiratory mucosal fluid interface is much more intense than previously envisioned. Sequential separation of pollen components has important implications for the production of improved allergenic extracts.

Allergens↗

Antifungal activity of three spermidine conjugates.

Three tri-substituted spermidines, di-p-coumaroyl-caffeoylspermidine, tri-caffeoylspermidine and tri-p-coumaroylspermidine, isolated from pollen of Quercus alba, were examined for antifungal activity. Both di-p-coumaroyl-caffeoylspermidine and tri-p-coumaroylspermidine reduced mycelial growth of the oat leaf stripe pathogen, Pyrenophora avenae and reduced powdery mildew (Blumeria graminis f. sp. hordei) infection of barley seedlings when applied as a post-inoculation treatment. When used as a pre-inoculation treatment, only di-p-coumaroyl-caffeoylspermidine reduced powdery mildew infection significantly. Growth of P. avenae in the presence of 100 microM di-p-coumaroyl-caffeoylspermidine reduced activity of S-adenosylmethionine decarboxylase (AdoMetDC), and led to a reduction in the incorporation of labelled ornithine into spermidine. The other two spermidine conjugates increased AdoMetDC activity and the flux label from ornithine into spermine in P. avenae significantly.

Antifungal Agents↗

Diversity of culturable bacterial populations associated to Tuber borchii ectomycorrhizas and their activity on T. borchii mycelial growth.

Isolation and physiological and molecular characterisation of culturable bacterial strains belonging to actinomycetes, pseudomonads and aerobic spore-forming bacteria were carried out on mycorrhizal root tips of Quercus robur var. peduncolata infected by Tuber borchii. Cellular density of the three bacterial groups in ectomycorrhizal root tips was estimated to be 1.3+/-0.11 x 10(6) cfu g(-1) dry weight for total heterotrophic bacteria and 1.08+/-0.6 x 10(5) (mean+/-S.E.), 1.3+/-0.3 x 10(5) and 1.4+/-0.2 x 10(5) cfu g(-1) dry weight for pseudomonads, actinomycetes and spore-forming bacteria respectively. Identification of pseudomonads by the Biolog system indicated, besides the most represented species Pseudomonas fluorescens (biotypes B, F and G), the occurrence of strains belonging to Pseudomonas corrugata. Amplified ribosomal DNA restriction analysis of actinomycetes and spore formers revealed at least three and six different groups of patterns, respectively. Many bacterial isolates were able to induce variations in growth rates of T. borchii mycelium; among these, 101 strains showed antifungal activity, whereas 17 isolates, belonging to spore formers, were able to increase mycelial growth up to 78% when compared to uninoculated mycelial growth. The potential role of these populations in the development and establishment of mycorrhizas is discussed.

Actinomycetales↗

Spironucleosis in Australian king parrots (Alisterus scapularis).

OBJECTIVE: To describe a syndrome of wasting, diarrhoea and mortality in Australian king parrots (Alisterus scapularis). DESIGN: Field observations and laboratory examinations. Procedure Pathological examinations were performed on 50 Australian king parrots with wasting and diarrhoea. Wet preparations of intestinal contents were examined by light microscopy. Tannins were extracted from acorns (Quercus sp) and tested for toxicity in mice. CLINICAL SIGNS AND EPIDEMIOLOGY: A syndrome of wasting, diarrhoea and mortality was observed in wild juvenile Australian king parrots in eastern Australia from 1984 to 2000. Sporadic cases and outbreaks of disease occurred from May to September in New South Wales, the Australian Capital Territory and Victoria. Outbreaks in the Australian Capital Territory in 1990 and 1991 were associated with parrots congregating to feed on acorns. Most affected birds failed to respond to treatment with dimetridazole and died 1 to 14 days after hospitalisation. Selected cases recovered following treatment with metronidazole. PATHOLOGY: Affected birds were emaciated, with faecal matting of feathers around the cloaca and yellow-green fluid, foamy intestinal contents. Abundant motile Spironucleus trophozoites were observed in wet preparations of faeces of clinically affected birds and intestinal contents of birds examined within 1 h of death. Protozoa were detected histologically in crypts of Lieberkühn in the intestine in association with exudation of mucus (catarrhal enteritis) or lymphoplasmacytic enteritis. Toxicology Tannin extracts from acorns induced periacinar hepatic necrosis in mice. CONCLUSION: Wasting, diarrhoea and mortality in wild juvenile Australian king parrots were associated with Spironucleus-like protozoa in the intestine. Acorns were not considered to be the cause of the syndrome.

Animals↗

Microbial degradation of lignocellulose: the lignin component.

A new procedure was developed for the study of lignin biodegradation by pure or mixed cultures of microorganisms. Natural lignocelluloses were prepared containing C in primarily their lignin components by feeding plants l-[U-C]phenylalanine through their cut stems. Lignin degradation was observed in numerous soils by monitoring evolution of CO(2) from [C]lignin-labeled oak (Quercus albus), maple (Acer rubrum), and cattail (Typha latifola). An organism (Thermonospora fusca ATCC 27730) that is known to degrade cellulose but not lignin was shown to grow on lignocellulose in the presence of [C]lignocelluloses without evolution of CO(2). A known lignin degrader (a white-rot fungus, Polyporus versicolor) was shown to readily evolve CO(2) from damp C-labeled cattail and C-labeled maple.

Journal Article↗

Effects of cadmium on aquatic hyphomycetes.

Two kinds of experiments, sporulation and growth experiments, were carried out to demonstrate the effect of cadmium on aquatic hyphomycetes. Oak (Quercus petraea L.) leaves were exposed in a hard-water stream (Lüssel, Swiss Jura) and a soft-water stream (Ibach, Black Forest) for 2 months. In the laboratory, fungal sporulation on the leaves in stream water enriched with cadmium (as CdCl(2)) was studied. A measurable effect was found when the cadmium concentration exceeded 0.1 ppm (0.1 mg/liter). Concentrations higher than 100 ppm inhibited conidium production completely. This toxic effect of cadmium was species dependent and much higher in soft water (water with low concentrations of calcium and magnesium) than in hard water. Growth experiments with Alatospora acuminata Ingold, Clavariopsis aquatica De Wildeman, Flagellospora curvula Ingold, Heliscus lugdunensis Saccardo and Therry, and Tetracladium marchalianum De Wildeman showed the same pattern of cadmium sensitivity as that seen in the sporulation experiments. Mycelial growth was less sensitive to cadmium than was fungal sporulation. High concentrations of competing cations (e.g., calcium and zinc) or potential ligands could reduce cadmium toxicity. Calcium content seems to be the most important factor responsible for the different sensitivity of aquatic hyphomycetes in hard and soft water.

Journal Article↗

Effects of ozone and sulfur dioxide on phyllosphere fungi from three tree species.

Short-term effects of ozone (O(3)) on phyllosphere fungi were studied by examining fungal populations from leaves of giant sequoia (Sequoiadendron giganteum (Lindl.) Buchholz) and California black oak (Quercus kelloggii Newb.). Chronic effects of both O(3) and sulfur dioxide (SO(2)) were studied by isolating fungi from leaves of mature Valencia orange (Citrus sinensis L.) trees. In this chronic-exposure experiment, mature orange trees were fumigated in open-top chambers at the University of California, Riverside, for 4 years with filtered air, ambient air plus filtered air (1:1), ambient air, or filtered air plus SO(2) at 9.3 parts per hundred million. Populations of Alternaria alternata (Fr.) Keissler and Cladosporium cladosporioides (Fres.) de Vries, two of the four most common fungi isolated from orange leaves, were significantly reduced by chronic exposure to ambient air. In the short-term experiments, seedlings of giant sequoia or California black oak were fumigated in open-top chambers in Sequoia National Park for 9 to 11 weeks with filtered air, ambient air, or ambient air plus O(3). These short-term fumigations did not significantly affect the numbers of phyllosphere fungi. Exposure of Valencia orange trees to SO(2) at 9.3 parts per hundred million for 4 years reduced the number of phyllosphere fungi isolated by 75% compared with the number from the filtered-air treatment and reduced the Simpson diversity index value from 3.3 to 2.5. A significant chamber effect was evident since leaves of giant sequoia and California black oak located outside of chambers had more phyllosphere fungi than did seedlings within chambers. Results suggest that chronic exposure to ambient ozone or SO(2) in polluted areas can affect phyllosphere fungal communities, while short-term exposures may not significantly disturb phyllosphere fungi.

Journal Article↗