Genome sequences and population genomics reveal climatic adaptation and genomic divergence between two closely related sweetgum species.

Understanding the genetic basis of population divergence and adaptation is an important goal in population genetics and evolutionary biology. However, the relative roles of demographic history, gene flow, and/or selective regime in driving genomic divergence, climatic adaptation, and speciation in non-model tree species are not yet fully understood. To address this issue, we generated whole-genome resequencing data of Liquidambar formosana and L. acalycina, which are broadly sympatric but altitudinally segregated in the Tertiary relict forests of subtropical China. We integrated genomic and environmental data to investigate the demographic history, genomic divergence, and climatic adaptation of these two sister species. We inferred a scenario of allopatric species divergence during the late Miocene, followed by secondary contact during the Holocene. We identified multiple genomic islands of elevated divergence that mainly evolved through divergence hitchhiking and recombination rate variation, likely fostered by long-term refugial isolation and recent differential introgression in low-recombination genomic regions. We also found some candidate genes with divergent selection signatures potentially involved in climatic adaptation and reproductive isolation. Our results contribute to a better understanding of how late Tertiary/Quaternary climatic change influenced speciation, genomic divergence, climatic adaptation, and introgressive hybridization in East Asia's Tertiary relict flora. In addition, they should facilitate future evolutionary, conservation genomics, and molecular breeding studies in Liquidambar, a genus of important medicinal and ornamental values.

Evaluation of the Potential Therapeutic Properties of Liquidambar orientalis Oil.

Liquidambar orientalis Mill., commonly called the Anatolian sweetgum or Sigla tree, is endemic to southwestern Turkey. It has been historically significant in traditional medicine. In our research, we delved into the therapeutic attributes of its oil, emphasizing its antioxidant, antimicrobial, and antitumor properties. The primary chemical constituent of the gum is styrene, accounting for 78.5 %. The gum demonstrated antioxidant capabilities in several assays, including in 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), cupric reducing antioxidant capacity (CUPRAC) and ferric reducing antioxidant power (FRAP). It displayed bactericidal actions against various gram-positive bacteria, such as Staphylococcus aureus, and gram-negative strains, including Escherichia coli. Additionally, the oil showcased potent antitumor effects against breast (MDA-MB-231), lung (A549), and prostate (PC3) cancer cell lines. These effects were found to be both time- and dose-dependent. L. orientalis Mill. oil showed the best antitumor activity against breast, lung, and prostate cancer cell lines after the 24 h and 48 h treatment. Its oil might induce autophagy in the PC3 prostate cancer cell line, whereas its cytotoxicity against MDA-MB-231 and A549 cancer cell lines might not be correlated with autophagy or apoptosis pathways. In conclusion, the oil from the Sigla tree offers promising therapeutic potential and warrants further exploration.

Evaluation of the effects of three natural products and a hemostatic agent on wound healing: an experimental study.

BACKGROUND: People have used many natural materials such as plant leaves, roots, liquids derived from plants, and animal products to treat wounds throughout history. It can be said that the research on wound care in recent years have focused on traditional and natural products again. This study aimed to investigate the effects of sweetgum oil, propolis, silk protein, and Ankaferd Blood Stopper (ABS) on wound healing in an experimental excisional wound model. METHODS: : Including 36 Balb/c inbreed mice in the study were divided equally into four groups. Two circular excisional wounds were created on the dorsal skin of mice under anesthesia using a punch biopsy device. The wounds of the first group of mice were topically dressed with sweetgum oil, the second group mice with propolis, the third group mice with silk protein, and the fourth group mice with ABS daily. Tissue samples were taken from the wounds of mice on the 7th and 14th day of wound formation, and histological examinations were performed. On the 14th day, the wounds created in all mice were healed, and the experiment was terminated. RESULTS: Mice in the silk protein group had faster wound healing. There was no statistical difference between the groups in immunohistochemical examinations. In the ABS group, the findings of the inflammatory process were more prominent. DISCUSSION: In conclusions, propolis, sweetgum oil, silk protein, and ABS positively affect different parameters in wound healing and support wound healing.

Resolution of traumatic mydriasis and accommodative dysfunction eight years after sweetgum ball ocular injury.

Purpose: To present a case of traumatic mydriasis (MD) and accommodative dysfunction (AD) secondary to a sweetgum ball ocular injury that resolved 8 years after the inciting trauma. Observations: A 6-year-old female presented with left eye ocular trauma after being hit with a sweetgum ball. Sweetgum balls are the small, spiky fallen fruits of the American Sweetgum tree (Liquidambar styraciflua). Due to their size and shape, children often use them as projectiles during play. On presentation, the patient had a partial thickness corneal laceration, traumatic mydriasis (TM), and accommodative dysfunction (AD). Her corneal laceration was repaired. Her TM and AD persisted. She was treated with bifocal spectacles and patching. At her 7-year follow-up visit, her TM and AD showed minimal signs of improvement. Eight years post-injury, her TM and AD had both improved significantly. Conclusion and Importance: Sweetgum balls, when used as projectiles, pose a risk of serious ocular injury. Pupillary and accommodative function in TM may improve much later than previously appreciated. Young age may contribute to parasympathetic neuroregeneration. Patching may have prevented amblyopia in this case, allowing her left eye to achieve its full visual potential once her pupillary and accommodative function returned.

Comparative Transcriptomic, Anatomical and Phytohormone Analyses Provide New Insights Into Hormone-Mediated Tetraploid Dwarfing in Hybrid Sweetgum (Liquidambar styraciflua × L. formosana).

Polyploid breeding is an effective approach to improve plant biomass and quality. Both fast growth and dwarf types of in vitro or ex vitro plants are produced after polyploidization. However, little is known regarding the dwarf type mechanism in polyploids grown in vitro. In this study, the morphological and cytological characteristics were measured in tetraploid and diploid hybrid sweetgum (Liquidambar styraciflua × L. formosana) with the same genetic background. RNA sequencing (RNA-Seq) was used to analyse shoot and root variations between tetraploid and diploid plants; important metabolites were validated. The results showed that the shoot and root lengths were significantly shorter in tetraploids than in diploids after 25 d of culture. Most tetraploid root cells were wider and more irregular, and the length of the meristematic zone was shorter, while tetraploid cells were significantly larger than diploid cells. Differentially expressed genes (DEGs) were significantly enriched in the plant growth and organ elongation pathways, such as plant hormone biosynthesis and signal transduction, sugar and starch metabolism, and cell cycles. Hormone biosynthesis and signal transduction genes, such as YUCCA, TAA1, GH3, SAUR, CPS, KO, KAO, GA20ox, GA3ox, BAS1 and CYCD3, which help to regulate organ elongation, were generally downregulated. The auxin, gibberellin, and brassinolide (BL) contents in roots and stems were significantly lower in tetraploids than in diploids, which may greatly contribute to slow growth in the roots and stems of tetraploid regenerated plants. Exogenous gibberellic acid (GA3) and indole-3-acetic acid (IAA), which induced plant cell elongation, could significantly promote growth in the stems and roots of tetraploids. In summary, comparative transcriptomics and metabolite analysis showed that the slow growth of regenerated tetraploid hybrid sweetgum was strongly related to auxin and gibberellin deficiency. Our findings provide insights into the molecular mechanisms that underlie dwarfism in allopolyploid hybrid sweetgum.

Forest stand and canopy development unaltered by 12 years of CO2 enrichment.

Canopy structure-the size and distribution of tree crowns and the spatial and temporal distribution of leaves within them-exerts dominant control over primary productivity, transpiration and energy exchange. Stand structure-the spatial arrangement of trees in the forest (height, basal area and spacing)-has a strong influence on forest growth, allocation and resource use. Forest response to elevated atmospheric CO2 is likely to be dependent on the canopy and stand structure. Here, we investigated elevated CO2 effects on the forest structure of a Liquidambar styraciflua L. stand in a free-air CO2 enrichment experiment, considering leaves, tree crowns, forest canopy and stand structure. During the 12-year experiment, the trees increased in height by 5 m and basal area increased by 37%. Basal area distribution among trees shifted from a relatively narrow distribution to a much broader one, but there was little evidence of a CO2 effect on height growth or basal area distribution. The differentiation into crown classes over time led to an increase in the number of unproductive intermediate and suppressed trees and to a greater concentration of stand basal area in the largest trees. A whole-tree harvest at the end of the experiment permitted detailed analysis of canopy structure. There was little effect of CO2 enrichment on the relative leaf area distribution within tree crowns and there was little change from 1998 to 2009. Leaf characteristics (leaf mass per unit area and nitrogen content) varied with crown depth; any effects of elevated CO2 were much smaller than the variation within the crown and were consistent throughout the crown. In this young, even-aged, monoculture plantation forest, there was little evidence that elevated CO2 accelerated tree and stand development, and there were remarkably small changes in canopy structure. Questions remain as to whether a more diverse, mixed species forest would respond similarly.

Global Transcriptome and Coexpression Network Analyses Reveal New Insights Into Somatic Embryogenesis in Hybrid Sweetgum (Liquidambar styraciflua × Liquidambar formosana).

Somatic embryogenesis (SE) is a process of somatic cells that dedifferentiate to totipotent embryonic stem cells and generate embryos in vitro. Despite recent scientific headway in deciphering the difficulties of somatic embryogenesis, the overall picture of key genes, pathways, and co-expression networks regulating SE is still fragmented. Therefore, deciphering the molecular basis of somatic embryogenesis of hybrid sweetgum remains pertinent. In the present study, we analyzed the transcriptome profiles and gene expression regulation changes via RNA sequencing from three distinct developmental stages of hybrid sweetgum: non-embryogenic callus (NEC), embryogenic callus (EC), and redifferentiation. Comparative transcriptome analysis showed that 19,957 genes were differentially expressed in ten pairwise comparisons of SE. Among these, plant hormone signaling-related genes, especially the auxin and cytokinin signaling components, were significantly enriched in NEC and EC early. The K-means method was used to identify multiple transcription factors, including HB-WOX, B3-ARF, AP2/ERF, and GRFs (growth regulating factors). These transcription factors showed distinct stage- or tissue-specific expression patterns mirroring each of the 12 superclusters to which they belonged. For example, the WOX transcription factor family was expressed only at NEC and EC stages, ARF transcription factor was expressed in EC early, and GRFs was expressed in late SE. It was noteworthy that the AP2/ERF transcription factor family was expressed during the whole SE process, but almost not in roots, stems and leaves. A weighted gene co-expression network analysis (WGCNA) was used in conjunction with the gene expression profiles to recognize the genes and modules that may associate with specific tissues and stages. We constructed co-expression networks and revealed 22 gene modules. Four of these modules with properties relating to embryonic potential, early somatic embryogenesis, and somatic embryo development, as well as some hub genes, were identified for further functional studied. Through a combination analysis of WGCNA and K-means, SE-related genes including AUX22, ABI3, ARF3, ARF5, AIL1, AIL5, AGL15, WOX11, WOX9, IAA29, BBM1, MYB36, LEA6, SMR4 and others were obtained, indicating that these genes play an important role in the processes underlying the progression from EC to somatic embryos (SEs) morphogenesis. The transcriptome information provided here will form the foundation for future research on genetic transformation and epigenetic control of plant embryogenesis at a molecular level. In follow-up studies, these data could be used to construct a regulatory network for SE; Key genes obtained from coexpression network analysis at each critical stage of somatic embryo can be considered as potential candidate genes to verify these networks.

Light alters the impacts of nitrogen and foliar pathogens on the performance of early successional tree seedlings.

Light limitation is a major driver of succession and an important determinant of the performance of shade-intolerant tree seedlings. Shade intolerance may result from a resource allocation strategy characterized by rapid growth and high metabolic costs, which may make shade-intolerant species particularly sensitive to nutrient limitation and pathogen pressure. In this study, we evaluated the degree to which nitrogen availability and fungal pathogen pressure interact to influence plant performance across different light environments. To test this, we manipulated nitrogen availability (high, low) and access by foliar fungal pathogens (sprayed with fungicide, unsprayed) to seedlings of the shade-intolerant tree, Liquidambar styraciflua, growing at low and high light availability, from forest understory to adjacent old field. Foliar fungal damage varied with light and nitrogen availability; in low light, increasing nitrogen availability tripled foliar damage, suggesting that increased nutrient availability in low light makes plants more susceptible to disease. Despite higher foliar damage under low light, spraying fungicide to exclude pathogens promoted 14% greater plant height only under high light conditions. Thus, although nitrogen availability and pathogen pressure each influenced aspects of plant performance, these effects were context dependent and overwhelmed by light limitation. This suggests that failure of shade-intolerant species to invade closed-canopy forest can be explained by light limitation alone.

The Essential Oil Composition and Antimicrobial Activity of Liquidambar formosana Oleoresin.

The oleoresin essential oils of Liquidambar formosana have potential therapeutic benefits. However, current research on L. formosana oleoresin essential oil is still in its early stages, and its chemotypic characterization is undefined. For better leveraging of plant resources and application of the essential oil, we collected 25 L. formosana oleoresin essential oil samples of individual trees from different geographical areas of Southern China. The essential oils were obtained by hydrodistillation and analyzed by gas chromatography-mass spectrometry (GC-MS) and gas chromatography-flame ionization detection (GC-FID). The major components of the essential oils were (E)-caryophyllene (3.3%-64.4%), α-pinene (0.6%-34.5%), ß-pinene (0.6%-26.0%), camphene (0.3%-17.3%), and limonene (0.2%-7.9%). A chiral GC-MS analysis was carried out on the essential oil samples and (-)-α-Pinene, (-)-ß-pinene, (-)-camphene, and (-)-limonene were the dominant enantiomers in L. formosana essential oil. The chemical categories of L. formosana oleoresin essential oils were clarified by agglomerative hierarchical cluster analysis (AHC) and principal component analysis (PCA). The multivariate analyses demonstrated that a total of four chemical groups can be delineated for L. formosana. The L. formosana essential oils were screened for antimicrobial activity against a panel of potentially pathogenic bacteria and fungi and showed promising antimicrobial activities with minimum inhibitory concentration (MIC) ≤ 625 µg/mL. These results highlight the economic value of L. formosana oleoresin essential oil, the importance of L. formosana sustainability, and the potential therapeutic benefits of its oleoresin essential oils.

Acanthotomicus sp. (Coleoptera: Curculionidae: Scolytinae), a New Destructive Insect Pest of North American Sweetgum Liquidambar styraciflua in China.

A previously unknown bark beetle species, Acanthotomicus sp., has emerged as a lethal pest of American sweetgum (Liquidambar styraciflua) in China. Our survey of nursery records from around Shanghai suggests that American sweetgum have been under heavy attack since at least 2013, resulting in the death of > 10,000 trees. Mass attacks of the apparently sweetgum-specific Acanthotomicus sp. can be diagnosed by accumulation of resinous exudates on the trunk, wilted foliage, and eventual numerous exit holes of the new generation. A Chinese native sweetgum Liquidambar formosana can also be colonized by Acanthotomicus sp. This pest is of concern not only as a killer of sweetgum in the Chinese nursery trade but also as a potentially destructive invasive pest of sweetgum in North America. This discovery suggests that global preinvasion assessment of pests is warranted.

Expected Timber-Based Economic Impacts of a Wood-Boring Beetle (Acanthotomicus Sp.) That Kills American Sweetgum.

American sweetgum trees (Liquidambar styraciflua L. [Altingiaceae]) in China are being killed by a newly discovered wood-boring beetle "sweetgum inscriber" (Acanthotomicus sp.). It has not been detected in the United States yet, but given the extent of trade with Asian countries, eventual arrival of this beetle is a serious concern. The American sweetgum is one of the main hardwood species in the southern United States, and provides several economic and ecological benefits to society. We present the first economic analysis of the potential damage from sweetgum inscriber (SI) to timber-based land values in the southern United States. We modeled economic impacts for a range of feasible SI arrival rates that reflect policy interventions: 1) no efforts to prevent arrival (scenario A, once every 14 and 25 yr), 2) partial prevention by complying with ISPM 15 standards (scenario B, once every 33 and 100 yr), and 3) total prevention of arrival (scenario C, zero transmission of SI). Our results indicated much lower land values for sweetgum plantations without the prevention on SI establishment (scenario A, US$1,843-US$4,383 ha-1) compared with partial prevention (scenario B, US$5,426-US$8,050 ha-1) and total eradication of SI (scenario C, US$9,825). Across the region, upper bound timber-based economic losses to plantation owners is US$151.9 million (US$4.6 million annually)-an estimate that can help inform policy decisions.

Elevated atmospheric carbon dioxide increases soil carbon.

The general lack of significant changes in mineral soil C stocks during CO2 -enrichment experiments has cast doubt on predictions that increased soil C can partially offset rising atmospheric CO2 concentrations. Here, we show, through meta-analysis techniques, that these experiments collectively exhibited a 5.6% increase in soil C over 2-9 years, at a median rate of 19 g C m-2 yr-1 . We also measured C accrual in deciduous forest and grassland soils, at rates exceeding 40 g C m-2 yr-1 for 5-8 years, because both systems responded to CO2 enrichment with large increases in root production. Even though native C stocks were relatively large, over half of the accrued C at both sites was incorporated into microaggregates, which protect C and increase its longevity. Our data, in combination with the meta-analysis, demonstrate the potential for mineral soils in diverse temperate ecosystems to store additional C in response to CO2 enrichment.

Sweetgum: An ancient source of beneficial compounds with modern benefits.

Sweetgum trees are large, deciduous trees found in Asia and North America. Sweetgum trees are important resources for medicinal and other beneficial compounds. Many of the medicinal properties of sweetgum are derived from the resinous sap that exudes when the outer bark of the tree has been damaged. The sap, known as storax, has been used for centuries to treat common ailments such as skin problems, coughs, and ulcers. More recently, storax has proven to be a strong antimicrobial agent even against multidrug resistant bacteria such as methicillin-resistant Staphylococcus aureus. In addition to the sap, the leaves, bark, and seeds of sweetgum also possess beneficial compounds such as shikimic acid, a precursor to the production of oseltamivir phosphate, the active ingredient in Tamiflu®-an antiviral drug effective against several influenza viruses. Other extracts derived from sweetgum trees have shown potential as antioxidants, anti-inflammatory agents, and chemopreventive agents. The compounds found in the extracts derived from sweetgum sap suppress hypertension in mice. Extracts from sweetgum seeds have anticonvulsant effects, which may make them suitable in the treatment of epilepsy. In addition to the potential medicinal uses of sweetgum extracts, the extracts of the sap possess antifungal activity against various phytopathogenic fungi and have been effective treatments for reducing nematodes and the yellow mosquito, Aedes aegypti, populations thus highlighting the potential of these extracts as environment-friendly pesticides and antifungal agents. The list of value-added products derived from sweetgum trees can be increased by continued research of this abundantly occurring tree.

Chemical and anatomical changes in Liquidambar styraciflua L. xylem after long term exposure to elevated CO2.

The anatomical and chemical characteristics of sweetgum were studied after 11 years of elevated CO2 (544 ppm, ambient at 391 ppm) exposure. Anatomically, branch xylem cells were larger for elevated CO2 trees, and the cell wall thickness was thinner. Chemically, elevated CO2 exposure did not impact the structural components of the stem wood, but non-structural components were significantly affected. Principal component analysis (PCA) was employed to detect differences between the CO2 treatments by considering numerous structural and chemical variables, as well as tree size, and data from previously published sources (i.e., root biomass, production and turnover). The PCA results indicated a clear separation between trees exposed to ambient and elevated CO2 conditions. Correlation loadings plots of the PCA revealed that stem structural components, ash, Ca, Mg, total phenolics, root biomass, production and turnover were the major responses that contribute to the separation between the elevated and ambient CO2 treated trees.

Sensitivity of stomatal and canopy conductance to elevated CO2 concentration - interacting variables and perspectives of scale.

• The hydrological response of forests to rising CO2 is a critical biotic feedback in the study of global climate change. Few studies, however, have investigated this highly dynamic response at relevant temporal and spatial scales. • A combination of leaf and whole-tree measurements and stand-level extrapolations were used to assess how stomatal conductance, canopy transpiration and conductance, and evapotranspiration might be affected by future, higher CO2 concentrations. • Midday measurements of stomatal conductance for leaves sampled in a 12-yr-old sweetgum (Liquidambar styraciflua) stand exposed to free-air CO2 enrichment were up to 44% lower at elevated than at ambient CO2 concentrations, whereas canopy conductance, averaged over the growing season, was only 14% lower in stands exposed to CO2 enrichment. The magnitude of this response was dependent on vapor pressure deficit and soil water potential. Annual estimates of evapotranspiration showed relatively small reductions due to atmospheric CO2 enrichment. • These data illustrate that the hydrological response of a closed-canopy plantation to elevated CO2 depends on the temporal and spatial scale of observation. They emphasize the importance of interacting variables and confirm that integration of measurements over space and time reduce what, at the leaf level, might otherwise appear to be a large and significant response.

Interactions between drought and elevated CO2 on growth and gas exchange of seedlings of three deciduous tree species.

Interactions between elevated atmospheric CO2 and drought on growth and gas exchange of American sycamore (Platanus occidentalis L.), sweetgum (Liquidambar styraciflua L.) and sugar maple (Acer saccharum Marsh.) were investigated using 1-yr-old seedlings, planted in 8 1 pots and grown in four open-top chambers, containing either ambient air or ambient air enriched with 300 µmol mol-1 CO2 . Two soil moisture regimes were included within each chamber: a 'well-watered' treatment with plants watered daily and a 'drought' treatment in which plants were subjected to a series of drought cycles. Duration and depth of the drought cycles were determined by soil matric potential. Mean soil water potential at rewatering for the water-stressed seedlings under ambient CO2 for sugar maple, sweetgum and sycamore was -0.5, -0.7 and -1.8 MPa, respectively, compared with > -0.1 MPa for all well-watered plants. Elevated CO2 increased relative growth rate of well-watered sugar maple by 181%, resulting in a 4.3-fold increase in total plant dry weight after 81 d, compared with 1.4 and 1.6-fold increases for sweetgum and sycamore, respectively, after 69 d. Although elevated CO2 increased net CO2 assimilation rate of sugar maple by 115%, there was a 10-fold increase in leaf area which contributed to the growth response. Although drought did not eliminate a growth response of sugar maple to elevated CO2 it greatly reduced the elevated CO2 -induced enhancement of relative growth rate. In contrast, relative growth rates of sweetgum and sycamore were not significantly increased by elevated CO2 . Drought, under elevated CO2 , reduced leaf area of all three species to a greater extent than it reduced net CO2 assimilation rate. The response ranged from no effect in sugar maple to a 40 % reduction in sycamore, with sweetgum exhibiting an intermediate response. Results indicate that drought may alter the growth response, gas exchange and water relations of tree species growing in an elevated CO2 atmosphere. Under high nutrient and water availability, sugar maple may benefit the most (of the three species studied) from a CO2 - enriched atmosphere, but productivity gains will be limited if frequent drought is prevalent.

Fine-root respiration in a loblolly pine and sweetgum forest growing in elevated CO2.

• The loss of carbon below-ground through respiration of fine roots may be modified by global change. Here we tested the hypothesis that a reduction in N concentration of tree fine-roots grown in an elevated atmospheric CO2 concentration would reduce maintenance respiration and that more energy would be used for root growth and N uptake. We partitioned total fine-root respiration (RT ) between maintenance (RM ), growth (RG ), and N uptake respiration (RN ) for loblolly pine (Pinus taeda) and sweetgum (Liquidambar styraciflua) forests exposed to elevated CO2 . • A substantial increase in fine-root production contributed to a 151% increase in RG for loblolly pine in elevated CO2 . Root specific RM for pine was 24% lower under elevated CO2 but when extrapolated to the entire forest, no treatment effect could be detected. • R G (< 10%) and RN (< 3%) were small components of RM in both forests. Maintenance respiration was the vast majority of RT , and contributed 92% and 86% of these totals at the pine and sweetgum forests, respectively. • The hypothesis was rejected because the majority of fine-root respiration was used for maintenance and was not reduced by changes in root N concentration in elevated CO2 . Because of its large contribution to RT and total soil CO2 efflux, changes in RM caused by warming may greatly alter carbon losses from forests to the atmosphere.

High-frequency induction of adventitious shoots from hypocotyl segments ofLiquidambar styracifiua L. by thidiazuron.

The effects of thidiazuron (TDZ) on adventitious bud and shoot formation from hypocotyl segments of sweetgum (Liquidambar styracifiua) were tested alone and in combination with 2,4-dichlorophenoxyacetic acid (2,4-D). The combination of 1 mg/1 TDZ with 0.01 mg/l 2,4-D resulted in the highest frequency of bud production. Lower concentrations of TDZ stimulated shoot production, generating the most shoots at 0.1 mg/1 TDZ with 0.01 mg/1 of 2,4-D. Inhibition of shoot elongation by TDZ was overcome by transferring shoot cultures to a shoot proliferation medium lacking TDZ or containing naphthaleneacetic acid and benzyladenine in addition to TDZ. Shoot production in liquid culture was significantly greater than that in solid culture. Comparisons of in vitro and ex vitro rooting of the adventitious shoots demonstrated that ex vitro rooting produced plants with faster growth rates and more extensive root systems.

Carbon dioxide stimulation of photosynthesis in Liquidambar styraciflua is not sustained during a 12-year field experiment.

Elevated atmospheric CO2 (eCO2) often increases photosynthetic CO2 assimilation (A) in field studies of temperate tree species. However, there is evidence that A may decline through time due to biochemical and morphological acclimation, and environmental constraints. Indeed, at the free-air CO2 enrichment (FACE) study in Oak Ridge, Tennessee, A was increased in 12-year-old sweetgum trees following 2 years of ∼40 % enhancement of CO2. A was re-assessed a decade later to determine if the initial enhancement of photosynthesis by eCO2 was sustained through time. Measurements were conducted at prevailing CO2 and temperature on detached, re-hydrated branches using a portable gas exchange system. Photosynthetic CO2 response curves (A versus the CO2 concentration in the intercellular air space (Ci); or A-Ci curves) were contrasted with earlier measurements using leaf photosynthesis model equations. Relationships between light-saturated photosynthesis (Asat), maximum electron transport rate (Jmax), maximum Rubisco activity (Vcmax), chlorophyll content and foliar nitrogen (N) were assessed. In 1999, Asat for eCO2 treatments was 15.4 ± 0.8 µmol m(-2) s(-1), 22 % higher than aCO2 treatments (P < 0.01). By 2009, Asat declined to <50 % of 1999 values, and there was no longer a significant effect of eCO2 (Asat = 6.9 or 5.7 ± 0.7 µmol m(-2) s(-1) for eCO2 or aCO2, respectively). In 1999, there was no treatment effect on area-based foliar N; however, by 2008, N content in eCO2 foliage was 17 % less than that in aCO2 foliage. Photosynthetic N-use efficiency (Asat : N) was greater in eCO2 in 1999 resulting in greater Asat despite similar N content, but the enhanced efficiency in eCO2 trees was lost as foliar N declined to sub-optimal levels. There was no treatment difference in the declining linear relationships between Jmax or Vcmax with declining N, or in the ratio of Jmax : Vcmax through time. Results suggest that the initial enhancement of photosynthesis to elevated CO2 will not be sustained through time if N becomes limited.