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Variability in net ecosystem exchange from hourly to inter-annual time scales at adjacent pine and hardwood forests: a wavelet analysis.

Orthonormal wavelet transformation (OWT) is a computationally efficient technique for quantifying underlying frequencies in nonstationary and gap-infested time series, such as eddy-covariance-measured net ecosystem exchange of CO2 (NEE). We employed OWT to analyze the frequency characteristics of synchronously measured and modeled NEE at adjacent pine (PP) and hardwood (HW) ecosystems. Wavelet cospectral analysis showed that NEE at PP was more correlated to light and vapor pressure deficit at the daily time scale, and NEE at HW was more correlated to leaf area index (LAI) and temperature, especially soil temperature, at seasonal time scales. Models were required to disentangle the impacts of environmental drivers on the components of NEE, ecosystem carbon assimilation (Ac) and ecosystem respiration (RE). Sensitivity analyses revealed that using air temperature rather than soil temperature in RE models improved the modeled wavelet spectral frequency response on time scales longer than 1 day at both ecosystems. Including LAI improved RE model fit on seasonal time scales at HW, and incorporating parameter variability improved the RE model response at annual time scales at both ecosystems. Resolving variability in canopy conductance, rather than leaf-internal CO2, was more important for modeling Ac at both ecosystems. The PP ecosystem was more sensitive to hydrologic variables that regulate canopy conductance: vapor pressure deficit on weekly time scales and soil moisture on seasonal to interannual time scales. The HW ecosystem was sensitive to water limitation on weekly time scales. A combination of intrinsic drought sensitivity and non-conservative water use at PP was the basis for this response. At both ecosystems, incorporating variability in LAI was required for an accurate spectral representation of modeled NEE. However, nonlinearities imposed by canopy light attenuation were of little importance to spectral fit. The OWT revealed similarities and differences in the scale-wise control of NEE by vegetation with implications for model simplification and improvement.

Carbon Dioxide↗

Relationship between stem CO2 efflux, stem sap velocity and xylem CO2 concentration in young loblolly pine trees.

We measured diel patterns of stem surface CO2 efflux (Es, micromol m(-2) s(-1)), sap velocity (vs, mm s(-1)) and xylem CO2 concentration ([CO2]) (Xs, %) in 8-year-old loblolly pine trees during the spring to determine how vs and Xs influence Es. All trees showed a strong diel hysteresis between Es and stem temperature, where at a given temperature, Es was lower during the day than at night. Diel variations in temperature-independent Es were correlated with vs (R2= 0.54), such that at maximum vs, Es was reduced between 18 and 40%. However, this correlation may not represent a cause-and-effect relationship. In a subset of trees, vs was artificially reduced by progressively removing the tree canopy. Reducing vs to near zero had no effect on Es and did not change the diel hysteretic response to temperature. Diel Xs tended to decrease with vs and increase with Es, however, in defoliated trees, large increases in Xs, when vs approximately 0, had no effect on Es. We conclude that at this time of the year, Es is driven primarily by respiration of cambium and phloem tissues and that sap flow and xylem transport of CO2 had no direct influence on Es.

Carbon↗

Soil bacterial diversity in a loblolly pine plantation: influence of ectomycorrhizas and fertilization.

We studied the effect of ectomycorrhizas and fertilization on soil microbial communities associated with roots of 10-year-old loblolly pine. Ectomycorrhizas were identified using a combination of community terminal restriction fragment profiling and matching of individual terminal restriction fragments to those produced from ectomycorrhizal clones and sequences recovered from roots and sporocarps. Differences between bacterial communities were initially determined using cluster analysis on community terminal restriction fragment profiles and through subsequent recovery of 16S rDNA clones. Analysis of bacterial clones revealed that terminal restriction fragment length was often shared between taxonomically dissimilar bacterial types. Consequently, we could not reliably infer the identity of peaks in the bacterial community profile with some exceptions, notably chloroplast rDNA that generated an approximate peak size of 80.2 bp. Fertilization increased the frequency of a Piloderma-like ectomycorrhiza. However, we did not detect clear effects of fertilization or the presence of viable ectomycorrhizas on bacterial communities. Bacterial communities seemed to be determined largely by the carbon and nitrogen content of soil. These results suggest that important soil microbial groups respond differently to soil conditions and management practices, with ectomycorrhizal communities reflecting past nutrient conditions and bacterial communities reflecting current environmental conditions of soil microsites.

Agriculture↗

Impacts of fine root turnover on forest NPP and soil C sequestration potential.

Estimates of forest net primary production (NPP) demand accurate estimates of root production and turnover. We assessed root turnover with the use of an isotope tracer in two forest free-air carbon dioxide enrichment experiments. Growth at elevated carbon dioxide did not accelerate root turnover in either the pine or the hardwood forest. Turnover of fine root carbon varied from 1.2 to 9 years, depending on root diameter and dominant tree species. These long turnover times suggest that root production and turnover in forests have been overestimated and that sequestration of anthropogenic atmospheric carbon in forest soils may be lower than currently estimated.

Atmosphere↗

Differential expression of pine and Cronartium quercuum f. sp. fusiforme genes in fusiform rust galls.

Cronartium quercuum f. sp. fusiforme is the causative agent of fusiform rust disease of southern pines in the United States. This disease is characterized by the formation of woody branch and stem galls. Differential display was used to identify pine genes whose expression is altered by C. quercuum f. sp. fusiforme infection and to identify C. quercuum f. sp. fusiforme genes that are expressed in fusiform rust galls. Six pine cDNAs that appeared to be differentially expressed in galled and healthy stems and 13 C. quercuum f. sp. fusiforme cDNAs expressed in galled tissues were identified. A probe that hybridizes specifically to C. quercuum f. sp. fusiforme 18S rRNA was used to estimate that 14% of the total RNA in fusiform rust galls was from C. quercuum f. sp. fusiforme. This finding was used to calibrate gene expression levels in galls when comparing them to expression levels in uninfected pines or in isolated C. quercuum f. sp. fusiforme cultures. According to Northern analysis and reverse transcriptase PCR analysis, all six of the pine clones were expressed at lower levels in galls than in healthy tissues. Seven of the nine C. quercuum f. sp. fusiforme clones that were assayed were expressed at higher levels in galls than in axenic culture. A number of the cDNAs encode proteins that are similar to those that play roles in plant development, plant defense, or fungal stress responses.

Basidiomycota↗

Fungal community analysis by large-scale sequencing of environmental samples.

Fungi are an important and diverse component of soil communities, but these communities have proven difficult to study in conventional biotic surveys. We evaluated soil fungal diversity at two sites in a temperate forest using direct isolation of small-subunit and internal transcribed spacer (ITS) rRNA genes by PCR and high-throughput sequencing of cloned fragments. We identified 412 sequence types from 863 fungal ITS sequences, as well as 112 ITS sequences from other eukaryotic microorganisms. Equal proportions of Basidiomycota and Ascomycota sequences were present in both the ITS and small-subunit libraries, while members of other fungal phyla were recovered at much lower frequencies. Many sequences closely matched sequences from mycorrhizal, plant-pathogenic, and saprophytic fungi. Compositional differences were observed among samples from different soil depths, with mycorrhizal species predominating deeper in the soil profile and saprophytic species predominating in the litter layer. Richness was consistently lowest in the deepest soil horizon samples. Comparable levels of fungal richness have been observed following traditional specimen-based collecting and culturing surveys, but only after much more extensive sampling. The high rate at which new sequence types were recovered even after sampling 863 fungal ITS sequences and the dominance of fungi in our libraries relative to other eukaryotes suggest that the abundance and diversity of fungi in forest soils may be much higher than previously hypothesized. All sequences were deposited in GenBank, with accession numbers AY 969316 to AY 970290 for the ITS sequences and AY 969135 to AY 969315 for the SSU sequences.

Computational Biology↗

The monosaccharide transporter gene family in land plants is ancient and shows differential subfamily expression and expansion across lineages.

BACKGROUND: In plants, tandem, segmental and whole-genome duplications are prevalent, resulting in large numbers of duplicate loci. Recent studies suggest that duplicate genes diverge predominantly through the partitioning of expression and that breadth of gene expression is related to the rate of gene duplication and protein sequence evolution.Here, we utilize expressed sequence tag (EST) data to study gene duplication and expression patterns in the monosaccharide transporter (MST) gene family across the land plants. In Arabidopsis, there are 53 MST genes that form seven distinct subfamilies. We created profile hidden Markov models of each subfamily and searched EST databases representing diverse land plant lineages to address the following questions: 1) Are homologs of each Arabidopsis subfamily present in the earliest land plants? 2) Do expression patterns among subfamilies and individual genes within subfamilies differ across lineages? 3) Has gene duplication within each lineage resulted in lineage-specific expansion patterns? We also looked for correlations between relative EST database representation in Arabidopsis and similarity to orthologs in early lineages. RESULTS: Homologs of all seven MST subfamilies were present in land plants at least 400 million years ago. Subfamily expression levels vary across lineages with greater relative expression of the STP, ERD6-like, INT and PLT subfamilies in the vascular plants. In the large EST databases of the moss, gymnosperm, monocot and eudicot lineages, EST contig construction reveals that MST subfamilies have experienced lineage-specific expansions. Large subfamily expansions appear to be due to multiple gene duplications arising from single ancestral genes. In Arabidopsis, one or a few genes within most subfamilies have much higher EST database representation than others. Most highly represented (broadly expressed) genes in Arabidopsis have best match orthologs in early divergent lineages. CONCLUSION: The seven subfamilies of the Arabidopsis MST gene family are ancient in land plants and show differential subfamily expression and lineage-specific subfamily expansions. Patterns of gene expression in Arabidopsis and correlation of highly represented genes with best match homologs in early lineages suggests that broadly expressed genes are often highly conserved, and that most genes have more limited expression.

Arabidopsis↗

Phenotypic plasticity of fine root growth increases plant productivity in pine seedlings.

BACKGROUND: The plastic response of fine roots to a changing environment is suggested to affect the growth and form of a plant. Here we show that the plasticity of fine root growth may increase plant productivity based on an experiment using young seedlings (14-week old) of loblolly pine. We use two contrasting pine ecotypes, "mesic" and "xeric", to investigate the adaptive significance of such a plastic response. RESULTS: The partitioning of biomass to fine roots is observed to reduce with increased nutrient availability. For the "mesic" ecotype, increased stem biomass as a consequence of more nutrients may be primarily due to reduced fine-root biomass partitioning. For the "xeric" ecotype, the favorable influence of the plasticity of fine root growth on stem growth results from increased allocation of biomass to foliage and decreased allocation to fine roots. An evolutionary genetic analysis indicates that the plasticity of fine root growth is inducible, whereas the plasticity of foliage is constitutive. CONCLUSIONS: Results promise to enhance a fundamental understanding of evolutionary changes of tree architecture under domestication and to design sound silvicultural and breeding measures for improving plant productivity.

Adaptation, Physiological↗

Genetic transformation of conifers and its application in forest biotechnology.

Genetic modification of conifers through gene transfer technology is now an important field in forest biotechnology. Two basic methodologies, particle bombardment and Agrobacterium-mediated transformation, have been used on conifers. The use of particle bombardment has produced stable transgenic plants in Picea abies, P. glauca, P. mariana, and Pinus radiata. Transgenic plants have been produced from Larix decidua, Picea abies, P. glauca, P. mariana, Pinus strobus, P. taeda, and P. radiata via Agrobacterium-mediated transformation. Agrobacterium-mediated transformation has advantages over particle bombardment such as a simpler integration pattern and a limited rearrangement in the introduced DNA. At present, genetic transformation of conifers has been directed toward improving growth rate, wood properties and quality, pest resistance, stress tolerance, and herbicide resistance, which will drive forestry to enter a new era of productivity and quality.

Agrobacterium tumefaciens↗

Heat-shock response of Pinus and Picea seedlings.

The effect of a short period of elevated temperature, or heat shock, on protein synthesis was investigated in 2-day-old seedlings of jack pine (Pinus banksiana Lamb.), loblolly pine (P. taeda L.), lodgepole pine (P. contorta Dougl.), black spruce (Picea mariana (Mill.) BSP), and white spruce (P. glauca (Moench) Voss.). In all species, heat shock led to increased [(35)S]methionine incorporation into heat-shock proteins (hsp's) with molecular masses of 83 and 72 kDA. Heat shock also induced synthesis of several low molecular weight proteins that were absent from control seedlings. The low molecular weight hsp's produced by pine seedlings had molecular masses of 27, 24.6, 20.5 and 17.5 kDa, whereas those produced by spruce seedlings had molecular masses of 27.2, 19.8, 18.4, 17.2 and 16.0 kDa. All of the low molecular weight hsp's showed isoelectric variants. Heat shock led to increased [(35)S]methionine incorporation into a group of low molecular weight hsp's that were also present in control seedlings.

Journal Article↗

Gene silencing of cinnamyl alcohol dehydrogenase in Pinus radiata callus cultures.

Xylem-derived Pinus radiata cell cultures, which can be induced to differentiate tracheary elements (TEs), were transformed with an RNAi construct designed to silence cinnamyl alcohol dehydrogenase (CAD), an enzyme involved in the biosynthesis of monolignols. Quantitative enzymatic CAD measurements revealed reduced CAD activity levels in most transclones generated. TEs from transclones with approximately 20% residual CAD activity did not release elevated levels of vanillin, which was derived from coniferyl-aldehyde through a mild alkali treatment. However, the activation of the phenylpropanoid pathway in transclones with approximately 20% residual CAD activity through the application of non-physiological concentrations of sucrose and l-phenylalanine produced phenotypic changes. The accumulation of metabolites such as dihydroconiferyl-alcohol (DHCA), which also accumulates in the P. taeda CAD mutant cad-n1, was observed. These results indicate that a substantial reduction in CAD activity is necessary for this enzyme to become a rate-limiting step in lignin biosynthesis in conifers such as P. radiata and confirm that transformable P. radiata callus cultures can be useful to investigate the function of xylogenesis-related genes in conifers.

Alcohol Oxidoreductases↗

Dependence of needle architecture and chemical composition on canopy light availability in three North American Pinus species with contrasting needle length.

Morphology and chemical composition of needles of shade-intolerant southern conifers (Pinus palustris Mill. (mean needle length +/- SD = 29.1 +/- 4.1 cm), P. taeda L. (12.3 +/- 2.9 cm) and P. virginiana Mill. (5.1 +/- 0.8 cm)) were studied to test the hypothesis that foliage acclimation potential to canopy light gradients is generally low for shade-intolerant species, and in particular, because of mechanical limitations, in species with longer needles. Plasticity for each needle variable was defined as the slope of the foliar characteristic versus irradiance relationship. A novel geometrical model for needle area and volume calculation was employed for the three-needled species P. palustris and P. taeda. Needle thickness (T) strongly increased, but width (W) was less variable with increasing daily integrated quantum flux density averaged over the season (Q(int)), resulting in changes in cross-sectional needle shape that were manifested in a positive relationship between the total to projected needle area ratio (A(T)/A(P)) and Q(int) in the three-needled species. In contrast, cross-sectional needle geometry was only slightly modified by irradiance in the two-needled conifer P. virginiana. Needle dry mass per unit total needle area (M(T)) was positively related to Q(int) in all species, leading to greater foliar nitrogen contents per unit area at higher irradiances. Separate examination of the components of M(T) (density (D) and the volume (V) to A(T) ratio; M(T) = DV/A(T)) indicated that the positive effect of light on M(T) resulted solely from increases in V/A(T), i.e., from increases in the thickness of foliage elements. Foliar chlorophyll content per unit mass increased with increasing Q(int), allowing an improvement in light-harvesting efficiency in low light. The variables characterizing needle material properties (D, the dry to fresh mass ratio, and needle carbon content per unit mass) were generally independent of Q(int), suggesting that needles were less stiff and had greater tip deflections under their own weight at lower irradiances because of smaller W and T. Comparisons with the literature revealed that plasticity in foliar characteristics tended to be lower in the studied shade- intolerant species than in shade-tolerant conifers, but plasticity among the investigated species was unaffected by needle length. However, we argue that, because of mechanical limitations, plastic changes in needle cross section in response to low irradiance may decrease rather than increase light-interception efficiency in long-needled species.

Biomass↗

Effects of anaerobic growth conditions on phosphorus tissue concentrations and absorption rates of southern pine seedlings.

A non-circulating, continuously flowing solution culture was used to examine the long- and short-term effects of anaerobic growth conditions on phosphorus uptake in 12-week-old seedlings of three pine species. Sand pine (Pinus clausa (Engelm.) Sarg.) and a drought-hardy loblolly pine (P. taeda L.) had the largest reductions in biomass after 8 weeks in anaerobic solution, whereas the more flood-tolerant pond pine (P. serotina Michx.) and wet-site loblolly pine seedlings were least affected. Anaerobic growth conditions reduced P concentrations in the shoot and increased P concentrations in the root. Short-term (32)P-uptake experiments were conducted with intact seedlings to determine absorption rates (influx) of inorganic phosphate (Pi) by aerobically and anaerobically grown pine seedlings. Influx of Pi was weakly correlated with shoot fresh weight, root fresh weight, root P and shoot P per gram fresh weight root, suggesting that internal P levels and biomass influenced Pi absorption. The highest Pi absorption rates were found in anaerobically grown seedlings under anaerobic (32)P-uptake conditions. Of these, sand pine had the highest absorption rate in 50 microM KH(2)PO(4) (0.96 micromol Pi g(-1) FW root h(-1)), and the wet-site loblolly pine exhibited the lowest rate (0.24 micromol Pi g(-1) FW root h(-1)). Aerobically grown seedlings had similar Pi absorption rates that were not significantly affected by O(2) concentration in the (32)P-uptake solution.

Journal Article↗

On the number of genes controlling the grass stage in longleaf pine.

The grass stage is an inherent and distinctive developmental trait of longleaf pine (Pinus palustris), in which height growth in the first few years after germination is suppressed. In operational forestry practice the grass stage extends for two to several years and often plays a role in planting failures and decisions to plant alternative species. Interspecies hybrids involving loblolly (P. taeda) and slash (P. elliottii var. elliottii) pines have been investigated as a means to produce planting stock with improved early height growth and to develop backcross populations for advanced generation breeding. We have reevaluated data from several interspecies populations, with the objective of estimating the number of genes contributing to the difference in first-year height growth between longleaf and loblolly pines. Estimates based on means and variances of parental and interspecies hybrid and backcross families suggest a minimum of 4 to 10 genes with standard errors less than half the estimates. These results suggest that the grass stage has evolved through the accumulation of alleles at several loci, each with small effects on various components of first-year height growth. Given the complexity of the grass-stage trait, tree breeders may need to combine genetic marker analysis with recurrent backcross breeding to efficiently develop longleaf pine planting stock for improved reforestation.

Analysis of Variance↗

Retroelements contribute to the excess low-copy-number DNA in pine.

Excess DNA in the single-copy component is rarely recognized as a contributor to the C-value paradox yet the single-copy component of the pine genome is reported to comprise over 3000 Mb of DNA, in large excess over the estimated 100 Mb required for gene expression. Two hypotheses regarding the factors that might contribute to the excess low-copy-number DNA were tested. The first hypothesis proposes that the excess low-copy kinetic component is actually overestimated by reassociation data analysis. To test this, a previously published C0t curve for Pinus strobus was reanalyzed using a new estimate of genome size based on laser flow cytometry. Part of the excess low-copy-number DNA in the pine genome could be attributed to the choice of parameters used in the analysis of the reassociation data. The second hypothesis holds that diverged retrotransposons contribute to the excess low-copy DNA. Sequences randomly sampled from single-copy and low-repetitive kinetic components of the P. taeda genome were characterized. Twelve of 46 fragments cloned from these fractions were found to show sequence similarity to retroelements: hence diverged retroelements contribute to the excess low-repetitive kinetic component in the pine genome. Similarity search was shown to be a conservative method for identifying retroelements, and thus the number of retroelements in the low-copy component was actually underestimated. Most of the retroelements in this fraction were nonfunctional. divergent from known retroelement families and previously reported only for flowering plants. Divergent retrotransposons are thus a major factor contributing to the expansion of the low-repetitive DNA component in higher plants.

Amino Acid Sequence↗

Survival and development of Lymantria monacha (Lepidoptera: Lymantriidae) on North American and introduced Eurasian tree species.

Lymantria monacha (L.) (Lepidoptera: Lymantriidae), the nun moth, is a Eurasian pest of conifers that has potential for accidental introduction into North America. To project the potential host range of this insect if introduced into North America, survival and development of L. monacha on 26 North American and eight introduced Eurasian tree species were examined. Seven conifer species (Abies concolor, Picea abies, P. glauca, P. pungens, Pinus sylvestris with male cones, P. menziesii variety glance, and Tsuga canadensis) and six broadleaf species (Betula populifolia, Malus x domestica, Prunus serotiaa, Quercus lobata, Q. rubra, and Q. velutina) were suitable for L. monacha survival and development. Eleven of the host species tested were rated as intermediate in suitability, four conifer species (Larix occidentalis, P. nigra, P. ponderosa, P. strobus, and Pseudotsuga menziesii variety menziesii) and six broadleaf species (Carpinus caroliniana, Carya ovata, Fagus grandifolia, Populus grandidentata, Q. alba, and Tilia cordata) and the remaining 10 species tested were rated as poor (Acer rubrum, A. platanoidies, A. saccharum, F. americana, Juniperus virginiana, Larix kaempferi, Liriodendron tulipfera, Morus alba, P. taeda, and P. deltoides). The phenological state of the trees had a major impact on establishment, survival, and development of L. monacha on many of the tree species tested. Several of the deciduous tree species that are suitable for L. monacha also are suitable for L. dispar (L.) and L. mathura Moore. Establishment of L. monacha in North America would be catastrophic because of the large number of economically important tree species on which it can survive and develop, and the ability of mated females to fly and colonize new areas.

Animals↗