Transcriptome analysis of gibberellins and abscisic acid during the flooding response in Fokienia hodginsii.

Flooding is one of the main abiotic stresses suffered by plants. Plants respond to flooding stress through regulating their morphological structure, endogenous hormone biosynthesis, and genetic signaling transduction. We previously found that Fokienia hodginsii varieties originating from Gutian exhibited typical flooding tolerance traits compared to three other provenances (Yongzhou, Sanming, Nanping), expressed as increased height, longer diameter at breast height (DBH), and smaller branch angle. Herein, the changes in endogenous gibberellins (GA) and abscisic acid (ABA) contents were measured under flooding stress in F. hodginsii, and ABA was found to decrease, whereas GA increased with time. Furthermore, the GA and ABA contents of the varieties originating from Gutian and the three other provenances were measured, and the results indicated that F. hodginsii from Gutian could respond more rapidly to flooding stress. The transcriptomes of the varieties originating from Gutian and the other three provenances were compared using RNA sequencing to explore the underlying genetic mechanisms of the flood-resistant phenotypes in F. hodginsii. The results indicated that two flood-stress response genes (TRINITY_DN142_c0_g2 and TRINITY_DN7657_c0_g1) were highly related to both the ABA and GA response in F. hodginsii.

Development and technical application of SSR-based individual identification system for Chamaecyparis taiwanensis against illegal logging convictions.

Chamaecyparis taiwanensis is an endemic plant suffering illegal logging in Taiwan for its high economic value. Lack of direct evidence to correlate stump and timber remains a hurdle for law enforcement. In this report, 23 polymorphic Genomic Simple Sequence Repeat (gSSR) and 12 Expressed Sequence Tag (EST)-SSR markers were developed and their transferability was assessed. The individual identification system built from selected non-linkage 30 SSR markers has a combined probability of identity as 5.596 × 10-12 equivalents to identifying an individual in a population of up to 18 million C. taiwanensis with 99.99% confidence level. We also applied the system in an actual criminal case by selecting 19 of these markers to correlate illegally felled timbers and victim trees. Our data demonstrate that molecular signals from three timbers hit with three victim trees with confidence level more than 99.99%. This is the first example of successfully applying SSR in C. taiwanensis as a court evidence for law enforcement. The identification system adapted advanced molecular technology and exhibits its great potential for natural resource management on C. taiwanensis.

Calculation procedures to estimate fine root production rates in forests using two-dimensional fine root data obtained by the net sheet method.

Several recent studies have used the net sheet method to estimate fine root production rates in forest ecosystems, wherein net sheets are inserted into the soil and fine roots growing through them are observed. Although this method has advantages in terms of its easy handling and low cost, there are uncertainties in the estimates per unit soil volume or unit stand area, because the net sheet is a two-dimensional material. Therefore, this study aimed to establish calculation procedures for estimating fine root production rates from two-dimensional fine root data on net sheets. This study was conducted in a hinoki cypress (Chamaecyparis obtusa (Sieb. & Zucc.) Endl.) stand in western Japan. We estimated fine root production rates in length and volume from the number (RN) and cross-sectional area (RCSA) densities, respectively, for fine roots crossing the net sheets, which were then converted to dry mass values. For these calculations, we used empirical regression equations or theoretical equations between the RN or RCSA densities on the vertical walls of soil pits and fine root densities in length or volume, respectively, in the soil, wherein the theoretical equations assumed random orientation of the growing fine roots. The estimates of mean fine root (diameter <1 mm) production rates were ∼80-100 g m-2 year-1 using the empirically obtained regression equations, whereas those from the theoretical equations were ∼40-50 g m-2 year-1. The difference in the estimates was attributed to larger slope values of the empirical regression equations than those of the theoretical equations, suggesting that fine root orientation was not random in our study site. In light of these results, we concluded that fine root production rates were successfully estimated from two-dimensional fine root data on the net sheets using these calculation procedures, with the empirical regression equations reflecting fine root orientation in the study site.

Physiological and morphological acclimation to height in cupressoid leaves of 100-year-old Chamaecyparis obtusa.

Cupressoid (scale-like) leaves are morphologically and functionally intermediate between stems and leaves. While past studies on height acclimation of cupressoid leaves have focused on acclimation to the vertical light gradient, the relationship between morphology and hydraulic function remains unexplored. Here, we compared physiological and morphological characteristics between treetop and lower-crown leaves of 100-year-old Chamaecyparis obtusa Endl. trees (~27 m tall) to investigate whether height-acclimation compensates for hydraulic constraints. We found that physiological acclimation of leaves was determined by light, which drove the vertical gradient of evaporative demand, while leaf morphology and anatomy were determined by height. Compared with lower-crown leaves, treetop leaves were physiologically acclimated to water stress. Leaf hydraulic conductance was not affected by height, and this contributed to higher photosynthetic rates of treetop leaves. Treetop leaves had higher leaf area density and greater leaf mass per area, which increase light interception but could also decrease hydraulic efficiency. We inferred that transfusion tissue flanking the leaf vein, which was more developed in the treetop leaves, contributes to water-stress acclimation and maintenance of leaf hydraulic conductance by facilitating osmotic adjustment of leaf water potential and efficient water transport from xylem to mesophyll. Our findings may represent anatomical adaptation that compensates for hydraulic constraints on physiological function with increasing height.

Sticky Traps Baited with Synthetic Aggregation Pheromone Predict Fruit Orchard Infestations of Plautia stali (Hemiptera: Pentatomidae).

The brown-winged green bug, Plautia stali Scott, mainly reproduces on Japanese cedar or cypress cones in Japanese plantation forests during summer and autumn. It often depletes its food sources in forest habitats and moves to cultivated crops in large numbers. To establish an easy method for assessing the risk of fruit orchard infestation by P. stali, we conducted a 3-yr field survey that monitored the attraction of bugs to the synthetic P. stali aggregation pheromone using a sticky trap. We used a morphological indicator, variable body size depending on food intake, to estimate the nutritional status in nymphs, which showed that nymphs attracted to the synthetic pheromone were starving. Comparisons between increasing changes in the number of stylet sheaths left on the cones by P. stali and the number of trapped nymphs show that monitoring nymphs with the pheromone-baited sticky trap is useful for inferring conditions regarding food resources in forest habitats. The trend toward trapping second instars can provide a timely overview of resource competition for cones. Trapping middle-to-late (third-fifth) instars is a warning that the cones are finally depleted and that there is a high probability that adults will leave the forests and invade the orchards. In addition, trends in trapping adults suggest that there is a potential risk of orchard infestation by the pest and predict the intensity and period of the invasion. The pheromone-baited sticky trap is an easy but useful survey tool for predicting P. stali orchard infestations.

Intraspecific variation in fine root respiration and morphology in response to in situ soil nitrogen fertility in a 100-year-old Chamaecyparis obtusa forest.

Soil N fertility has an effect on belowground C allocation, but the physiological and morphological responses of individual fine root segments to variations in N availability under field conditions are still unclear. In this study, the direction and magnitude of the physiological and morphological function of fine roots in response to variable in situ soil N fertility in a forest site were determined. We measured the specific root respiration (Rr) rate, N concentration and morphology of fine root segments with 1-3 branching orders in a 100-year-old coniferous forest of Chamaecyparis obtusa. Higher soil N fertility induced higher Rr rates, root N concentration, and specific root length (SRL), and lower root tissue density (RTD). In all fertility levels, the Rr rates were significantly correlated positively with root N and SRL and negatively with RTD. The regression slopes of respiration with root N and RTD were significantly higher along the soil N fertility gradient. Although no differences in the slopes of Rr and SRL relationship were found across the levels, there were significant shifts in the intercept along the common slope. These results suggest that a contrasting pattern in intraspecific relationships between specific Rr and N, RTD, and SRL exists among soils with different N fertility. Consequently, substantial increases in soil N fertility would exert positive effects on organ-scale root performance by covarying the Rr, root N, and morphology for their potential nutrient and water uptake.

Determination of Seed Soundness in Conifers Cryptomeria japonica and Chamaecyparis obtusa Using Narrow-Multiband Spectral Imaging in the Short-Wavelength Infrared Range.

Regeneration of planted forests of Cryptomeria japonica (sugi) and Chamaecyparis obtuse (hinoki) is the pressing importance to the forest administration in Japan. Low seed germination rate of these species, however, has hampered low-cost production of their seedlings for reforestation. The primary cause of the low germinability has been attributed to highly frequent formation of anatomically unsound seeds, which are indistinguishable from sound germinable seeds by visible observation and other common criteria such as size and weight. To establish a method for sound seed selection in these species, hyperspectral imaging technique was used to identify a wavelength range where reflectance spectra differ clearly between sound and unsound seeds. In sound seeds of both species, reflectance in a narrow waveband centered at 1,730 nm, corresponding to a lipid absorption band in the short-wavelength infrared (SWIR) range, was greatly depressed relative to that in adjacent wavebands on either side. Such depression was absent or less prominent in unsound seeds. Based on these observations, a reflectance index SQI, abbreviated for seed quality index, was formulated using reflectance at three narrow SWIR wavebands so that it represents the extent of the depression. SQI calculated from seed area-averaged reflectance spectra and spatial distribution patterns of pixelwise SQI within each seed area were both proven as reliable criteria for sound seed selection. Enrichment of sound seeds was accompanied by an increase in germination rate of the seed lot. Thus, the methods described are readily applicable toward low-cost seedling production in combination with single seed sowing technology.

Relationships of tree height and diameter at breast height revisited: analyses of stem growth using 20-year data of an even-aged Chamaecyparis obtusa stand.

Stem diameter at breast height (DBH) and tree height (H) are commonly used measures of tree growth. We examined patterns of height growth and diameter growth along a stem using a 20-year record of an even-aged hinoki cypress (Chamaecyparis obtusa (Siebold & Zucc.) Endl.) stand. In the region of the stem below the crown (except for the butt swell), diameter growth rates (ΔD) at different heights tended to increase slightly from breast height upwards. This increasing trend was pronounced in suppressed trees, but not as much as the variation in ΔD among individual trees. Hence, ΔD below the crown can be regarded as generally being represented by the DBH growth rate (ΔDBH) of a tree. Accordingly, the growth rate of the stem cross-sectional area increased along the stem upwards in suppressed trees, but decreased in dominant trees. The stem diameter just below the crown base (D(CB)), the square of which is an index of the amount of leaves on a tree, was an important factor affecting ΔDBH. D(CB) also had a strong positive relationship with crown length. Hence, long-term changes in the D(CB) of a tree were associated with long-term changes in crown length, determined by the balance between the height growth rate (ΔH) and the rising rate of the crown base (ΔH(CB)). Within the crown, ΔD's were generally greater than the rates below the crown. Even dying trees (ΔD ≈ 0 below the crown) maintained ΔD > 0 within the crown and ΔH > 0 until about 5 years before death. This growth within the crown may be related to the need to produce new leaves to compensate for leaves lost owing to the longevity of the lower crown. These results explain the different time trajectories in DBH-H relationships among individual trees, and also the long-term changes in the DBH-H relationships. The view that a rise in the crown base is strongly related to leaf turnover helps to interpret DBH-H relationships.

A conifer ABI3-interacting protein plays important roles during key transitions of the plant life cycle.

ABI3 (for ABSCISIC ACID INSENSITIVE3), a transcription factor of the abscisic acid signal transduction pathway, plays a major role during seed development, dormancy inception, and dormancy maintenance. This protein appears to also function in meristematic and vegetative plant tissues and under certain stress conditions. We have isolated the ABI3 gene ortholog (CnABI3) from yellow cedar (Callitropsis nootkatensis) and found that it was functionally similar to other ABI3 genes of angiosperms. Here, we report that using a yeast (Saccharomyces cerevisiae) two-hybrid approach, we have identified another protein of yellow cedar (CnAIP2; for CnABI3 INTERACTING PROTEIN2) that physically interacts with CnABI3. Functional analyses revealed that CnAIP2 plays important roles during key transitions in the plant life cycle: (1) CnAIP2 impaired seed development and reduced seed dormancy; (2) CnAIP2 promoted root development, particularly the initiation of lateral roots, and the CnAIP2 gene promoter was exquisitely auxin sensitive; and (3) CnAIP2 promoted the transition from vegetative growth to reproductive initiation (i.e. flowering). The nature of the effects of CnAIP2 on these processes and other evidence place CnAIP2 in the category of a "global" regulator, whose actions are antagonistic to those of ABI3.

Eyeing emergence: modified treatments for terminating dormancy of conifer seeds.

Many seeds of coniferous species display a deep primary dormancy at maturity and require several weeks of pretreatment to produce seed populations that germinate in a vigorous and timely manner. Facilitating an efficient transition from dormancy to germination by devising improved protocols for dormancy breakage is not only important to conifer seed research, aiding in the study of the dormancy process itself, but is also of interest and applicability to commercial forest nursery operations. In the forests of British Columbia, Canada, several conifer species are well-adapted to their environment, with seeds needing to experience long durations in the moist state at cool or fluctuating temperatures. These include yellow-cedar (Callitropsis nootkatensis), western white pine (Pinus monticola), and true fir species, such as Pacific silver fir and subalpine fir (Abies amabilis and A. lasiocarpa, respectively). In this chapter, we discuss the development of new dormancy-breaking protocols for the aforementioned species that centre on the balance of several key aspects: (1) reducing the time needed to terminate dormancy in the seed population; (2) synchronicity of germination; (3) ease of use; (4) cost-effectiveness; and (5) repeatability. Where possible, any new or modified protocol should be further tested in relationship to promoting rapid seedling growth in a forest nursery greenhouse setting and after planting at natural stands. Based on the five criteria listed above, very significant improvements compared to traditional dormancy-breaking methods have been achieved for the targeted conifer species. Where tested (e.g. yellow-cedar), the modified dormancy-breaking treatments result in vigorous growth in the greenhouse and after planting at natural stands.

Relationships among plants, soils and microbial communities along a hydrological gradient in the New Jersey Pinelands, USA.

BACKGROUND AND AIMS: Understanding the role of different components of hydrology in structuring wetland communities is not well developed. A sequence of adjacent wetlands located on a catenary sequence of soils and receiving the same sources and qualities of water is used to examine specifically the role of water-table median position and variability in affecting plant and microbial community composition and soil properties. METHODS: Two replicates of three types of wetland found adjacent to each other along a hydrological gradient in the New Jersey Pinelands (USA) were studied. Plant-community and water-table data were obtained within a 100-m(2) plot in each community (pine swamp, maple swamp and Atlantic-white-cedar swamp). Monthly soil samples from each plot were analysed for soil moisture, organic matter, extractable nitrogen fractions, N mineralization rate and microbial community composition. Multivariate ordination methods were used to compare patterns among sites within and between data sets. KEY RESULTS: The maple and pine wetlands were more similar to each other in plant community composition, soil properties and microbial community composition than either was to the cedar swamps. However, maple and pine wetlands differed from each other in water-table descriptors as much as they differed from the cedar swamps. All microbial communities were dominated by Gram-positive bacteria despite hydrologic differences among the sites. Water-table variability was as important as water-table level in affecting microbial communities. CONCLUSIONS: Water tables affect wetland communities through both median level and variability. Differentiation of both plant and microbial communities are not simple transforms of differences in water-table position, even when other hydrologic factors are kept constant. Rather, soil genesis, a result of both water-table position and geologic history, appears to be the main factor affecting plant and microbial community similarities.

Screening genes that change expression during compression wood formation in Chamaecyparis obtusa.

We screened cDNA fragments that change their expression during compression wood formation by fluorescent differential display (FDD) in five adult trees (Chamaecyparis obtusa (Siebold & Zucc.) Endl.) growing naturally at an angle to the vertical, and in two saplings, one vertical, the other inclined. We conducted anatomical observations and measurements of the released strain of growth stress on the five adult trees to confirm that they formed compression wood on the lower side of the inclined trunks. Based on sequencing results from selected cDNA fragments, we conducted homology searches of the GenBank database and designed specific primers for the 67 screened fragments. Using these primers and different saplings from those used for the FDD screening, we tested the expression levels of each fragment in normal, compression and opposite wood regions of saplings by semiquantitative reverse-transcription polymerase chain reaction. Twenty-four fragments showed reproducible expression patterns, indicating that these fragments changed their expression during compression wood formation. Some fragments showed differential expression between the apical and basal regions of the lower side of the inclined stem in the region of compression wood formation. Anatomical observations indicated more intense compression wood formation in the basal region than in the apical region of the stem, demonstrating a relationship between compression wood development and gene expression.

Growth strain in the trunk and branches of Chamaecyparis formosensis and its influence on tree form.

Distributions of growth strains in branches, straight trunks and basal sweeping trunks of Chamaecyparis formosensis Matsum. trees were measured with strain gauges. Microfibril angles (MFAs) of the S2 layer of the cell wall were measured by the iodine deposition method and their relationships with growth strain examined. The magnitude of the compressive stress on the lower side of trunks with a basal sweep was greater than that of the tensile stress at the surface of straight trunks. However, transverse compressive stress was similar around the trunk regardless of whether normal wood or compression wood was present. The released surface growth strains varied with MFA. At MFAs of 20-25 degrees , growth stress changed from tension to compression, and compressive stress increased dramatically in the compression wood region. Branches suffer bending stress due to self-loading. This stress is superimposed on the growth stress. Growth strains on the upper or lower sides of branches were larger than those in the trunks, suggesting that generation of growth stress on the lower sides of branches with extensive compression wood is affected by the gravitational bending stress due to self-loading. We conclude that branch form is affected by the interaction between the bending moment due to self-loading and that due to the asymmetric distribution of growth stress. Growth strain distribution in a branch differed depending on whether the branch was horizontal, upward bending or downward bending.

Dynamics of heterorhizic root systems: protoxylem groups within the fine-root system of Chamaecyparis obtusa.

To understand the physiology of fine-root functions in relation to soil organic sources, the heterogeneity of individual root functions within a fine-root system requires investigation. Here the heterogeneous dynamics within fine-root systems are reported. The fine roots of Chamaecyparis obtusa were sampled using a sequential ingrowth core method over 2 yr. After color categorization, roots were classified into protoxylem groups from anatomical observations. The root lengths with diarch and triarch groups fluctuated seasonally, whereas the tetrarch root length increased. The percentage of secondary root mortality to total mortality increased with increasing amounts of protoxylem. The carbon : nitrogen ratio indicated that the decomposability of primary roots might be greater than that of secondary roots. The position of diarch roots was mostly apical, whereas tetrarch roots tended to be distributed in basal positions within the root architecture. We demonstrate the heterogeneous dynamics within a fine-root system of C. obtusa. Fine-root heterogeneity should affect soil C dynamics. This heterogeneity is determined by the branching position within the root architecture.

Growth and transpiration of Japanese cedar (Cryptomeria japonica) and Hinoki cypress (Chamaecyparis obtusa) seedlings in response to soil water content.

To investigate the effects of soil water content on growth and transpiration of Japanese cedar (Cryptomeria japonica D. Don) and Hinoki cypress (Chamaecyparis obtusa (Siebold et Zucc.) Endl.), potted seedlings were grown in well-watered soil (wet treatment) or in drying soil (dry treatment) for 12 weeks. Seedlings in the wet treatment were watered once every 2 or 3 days, whereas seedlings in the dry treatment were watered when soil water content (Theta; m3 m(-3)) reached 0.30, equivalent to a soil matric potential of -0.06 MPa. From Weeks 7 to 12 after the onset of the treatments, seedling transpiration was measured by weighing the potted seedlings. After the last watering, changes in transpiration rate during soil drying were monitored intensely. The dry treatment restricted aboveground growth but increased biomass allocation to the roots in both species, resulting in no significant treatment difference in whole-plant biomass production. The species showed similar responses in relative growth rate (RGR), net assimilation rate (NAR) and shoot mass ratio (SMR) to the dry treatment. Although NAR did not change significantly in either C. japonica or C. obtusa as the soil dried, the two species responded differently to the dry treatment in terms of mean transpiration rate (E) and water-use efficiency (WUE), which are parameters that relate to NAR. In the dry treatment, both E and WUE of C. japonica were stable, whereas in C. obtusa, E decreased and WUE increased (E and WUE counterbalanced to maintain a constant NAR). Transpiration rates were lower in C. obtusa seedlings than in C. japonica seedlings, even in well-watered conditions. During soil drying, the transpiration rate decreased after Theta reached about 0.38 (-0.003 MPa) in C. obtusa and 0.32 (-0.028 MPa) in C. japonica. We conclude that C. obtusa has more water-saving characteristics than C. japonica, particularly when water supply is limited.