The lineage-specific evolution of the oleosin family in Theaceae.

Oleosins play essential roles in stabilization of lipid droplets (LDs) and seed oil production. However, evolution of this gene family has not been reported in Theaceae, a large plant family that contains many important tea and oil tea species. In this study, a total of 65 oleosin genes were identified in nine genome-sequenced Theaceae species. Among these genomes, the gene number of oleosin showed significant difference, with Camellia sinensis var. sinensis cv. Shuchazao and Camellia lanceoleosa displayed more oleosin numbers than other species. Phylogenetic analyses revealed that Theaceae oleosin genes were classified into three clades (U, SL, SH) respectively. Proteins within the same clade had similar gene structure and motif composition. Segmental duplication was the primary driving force for the evolution of oleosin genes in Shuchazao (SCZ), Huangdan (HD), C.lanceoleosa (Cla), and wild tea (DASZ). Synteny analysis showed that most oleosin genes displayed inter-species synteny among tea and oil tea species. Expression analysis demonstrated that oleosin genes were specifically expressed in seed and kernel of Huangdan (HD) and C.lanceoleosa. Moreover, expression divergence was observed in paralogous pairs and ∼1-2 oleosin genes in each clade have become activate. This study leads to a comprehensive understanding of evolution of oleosin family in Theaceae, and provides a rich resource to further address the functions of oleosin in tea and oil tea species.

Discovery and Biochemical Characterization of N-methyltransferase Genes Involved in Purine Alkaloid Biosynthetic Pathway of Camellia gymnogyna Hung T.Chang (Theaceae) from Dayao Mountain.

In the present study, purine alkaloid analysis and transcriptome of Camellia gymnogyna Hung T. Chang (Theaceae) from Dayao Mountain were performed by high-performance liquid chromatography (HPLC) and RNA-Seq, respectively. The results showed that the major purine alkaloids accumulated in Camellia gymnogyna Hung T. Chang (Theaceae) were theobromine together with a small amount of theacrine and caffeine. Through polymerase chain reaction (PCR), three types of cDNA encoding N-methyltransferases were isolated from the leaves of Camellia gymnogyna Hung T. Chang (Theaceae) and designated GCS1, GCS2, and GCS3. We subsequently expressed GCS1, GCS2, and GCS3 in Escherichia coli and incubated lysates of the bacterial cells with a variety of xanthine substrates in the presence of S-adenosyl-L-methionine as the methyl donor. We found that the recombinant GCS1 proteins catalyzed 1,3,7-trimethyluric acid to produce theacrine, the recombinant GCS3 proteins catalyzed 7-methylxanthine to produce theobromine, while the recombinant GCS2 proteins did not catalyze any xanthine derivatives. Simultaneous analysis of the expressions of GCS1, GCS2, GCS3, and a caffeine synthase gene (TCS1) in Camellia gymnogyna Hung T. Chang (Theaceae) and other tea plants provided a reference for further research on the functions of these genes.

Functional characteristics of phenolic compounds accumulated in young leaves of two subtropical forest tree species of different successional stages.

The abundance of phenolic compounds (including anthocyanins) in leaves is associated with photosynthetic performance, but the regulatory mechanism is unclear. Schima superba Gardn. et Champ. and Cryptocarya concinna Hance., which exhibit distinct anthocyanin accumulation patterns, are dominant tree species in the early- and late-successional stages, respectively, of subtropical forests in China. RNA-seq and analyses of phenolic concentrations, antioxidant capacity and photosynthetic characteristics were performed on young and mature leaves of these two species under contrasting light conditions. The high-light-acclimated young leaves of S. superba and C. concinna and low-light-acclimated young leaves of C. concinna were red. These red leaves had higher ratios of electron transport rate to gross photosynthesis (ETR:Pgross) and total antioxidant capacity to chlorophyll (TAC:Chl) than did the green leaves, regardless of light conditions. In addition, the red leaves had a higher expression level of the UDP-glucose:flavonoid 3-O-glucosyltransferase (UFGT) gene than did the green leaves, irrespective of light conditions. Total antioxidant capacity was positively correlated with flavonoid content in C. concinna leaves and with total phenolic content in leaves of both species under both high and low light. Consistent with the measurements of photosynthetic performance and flavonoids:Chl ratio, photosynthesis-related genes were extensively downregulated and flavonoid-pathway-related genes were extensively upregulated in young leaves relative to mature leaves. Under high and low light, both non-photochemical quenching and TAC:Chl, which serve as different types of photoprotective tools, were enhanced in young leaves of S. superba, whereas only TAC:Chl was enhanced in young leaves of C. concinna. Our results indicate that the biosynthesis of phenolic compounds in young leaves is likely enhanced by an imbalance between photosynthetic electron supply and demand and that flavonoids play a larger role in meditating photoprotection in late-successional species than in early-successional ones.

Stomatal uptake of O3 in a Schima superba plantation in subtropical China derived from sap flow measurements.

Canopy stomatal ozone (O3) flux (Fst,O3) in a plantation of Schima superba, an ecologically and economically important evergreen pioneer tree species in subtropical China, was quantified based on sap flow measurements during a 2-year period. Mean Fst,O3 and accumulated Fst,O3 (AFst0) were significantly higher in wet seasons from April to September (4.62 nmol m(-2) s(-1) and 35.37 mmol m(-2), respectively) than in dry seasons from October to March (3.90 nmol m(-2) s(-1) and 24.15 mmol m(-1), respectively), yet comparable between the 2 years of the experiment, being 4.23 nmol m(-2) s(-1) and 58.23 mmol m(-2) in April 2013-March 2014 and 4.29 nmol m(-2) s(-1) and 60.80 mmol m(-2) in April 2014-March 2015, respectively. At the diurnal scale, Fst,O3 generally peaked in the early to middle afternoon hours (13:00-15:00), while the maximum stomatal conductance (Gst,O3) typically occurred in the middle to late morning hours (09:00-11:00). Monthly integrated AFst0 reached the maximum in July, although accumulated O3 exposure (SUM0) was highest in October. Seasonally or yearly, the accumulated O3 doses, either exposure-based or flux-based, notably exceeded the currently adopted critical thresholds for the protection of forest trees. These results, on the one hand, demonstrated the decoupling between the stomatal uptake of O3 and its environmental exposure level; on the other hand, indicated the potential O3 risk for S. superba in the experimental site. Therefore, the present study endorses the use of sap flow measurements as a feasible tool for estimating Fst,O3, and the transition from the exposure-based toward flux-based metrics for assessing O3 risk for forest trees. Further studies are urgently needed to relate stomatal O3 uptake doses with tree growth reductions for an improved understanding of O3 effects on trees under natural conditions.

Proteomic analysis reveals differences in tolerance to acid rain in two broad-leaf tree species, Liquidambar formosana and Schima superba.

Acid rain (AR) is a serious environmental issue inducing harmful impacts on plant growth and development. It has been reported that Liquidambar formosana, considered as an AR-sensitive tree species, was largely injured by AR, compared with Schima superba, an AR-tolerant tree species. To clarify the different responses of these two species to AR, a comparative proteomic analysis was conducted in this study. More than 1000 protein spots were reproducibly detected on two-dimensional electrophoresis gels. Among them, 74 protein spots from L. formosana gels and 34 protein spots from S. superba gels showed significant changes in their abundances under AR stress. In both L. formosana and S. superba, the majority proteins with more than 2 fold changes were involved in photosynthesis and energy production, followed by material metabolism, stress and defense, transcription, post-translational and modification, and signal transduction. In contrast with L. formosana, no hormone response-related protein was found in S. superba. Moreover, the changes of proteins involved in photosynthesis, starch synthesis, and translation were distinctly different between L. formosana and S. superba. Protein expression analysis of three proteins (ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit, ascorbate peroxidase and glutathione-S-transferase) by Western blot was well correlated with the results of proteomics. In conclusion, our study provides new insights into AR stress responses in woody plants and clarifies the differences in strategies to cope with AR between L. formosana and S. superba.

Photosynthetic and antioxidant responses of Liquidambar formosana and Schima superba seedlings to sulfuric-rich and nitric-rich simulated acid rain.

To study whether differential responses occur in photosynthesis and antioxidant system for seedlings of Liquidambar formosana, an acid rain (AR)-sensitive tree species and Schima superba, an AR-tolerant tree species treated with three types of pH 3.0 simulated AR (SiAR) including sulfuric-rich (S-SiAR), nitric-rich (N-SiAR), sulfate and nitrate mixed (SN-SiAR), we investigated the changes of leaf necrosis, chlorophyll content, soluble protein and proline content, photosynthesis and chlorophyll fluorescence characteristics, reactive oxygen species production, membrane lipid peroxidation, small molecular antioxidant content, antioxidant enzyme activities and related protein expressions. Our results showed that SiAR significantly caused leaf necrosis, inhibited photosynthesis, induced superoxide radical and hydrogen peroxide generation, aggravated membrane lipid peroxidation, changed antioxidant enzyme activities, modified related protein expressions such as Cu/Zn superoxide dismutase (SOD), l-ascorbate peroxidase (APX, EC 1. 11. 1. 11), glutathione S transferase (GST, EC 2. 5. 1. 18) and Rubisco large subunit (RuBISCO LSU), altered non-protein thiols (NPT) and glutathione (GSH) content in leaves of L. formosana and S. superba. Taken together, we concluded that the damages caused by SiAR in L. formosana were more severe and suffered from more negative impacts than in S. superba. S-SiAR induced more serious damages for the plants than did SN-SiAR and N-SiAR.

[Effects of nitrogen and phosphorus fertilization on litterfall production and nutrient dynamics in a Schima superba forest in Zhejiang Province of East China].

In 2011, an investigation was made on the litterfall production and nutrient dynamics in a phosphorus-limited subtropical evergreen broadleaved forest dominated by Schima superba in Tiantong, Zhejiang Province of East China under different levels of nitrogen (N) and phosphorous (P) fertilization. After the N and P fertilization, the annual litterfall production ranged from 6.82 to 8.30 t x hm(-2) x a(-1), with three peaks in May or June, August, and October, respectively. Under fertilization, the annual average N and P concentrations of the litterfall (except the annual average N concentration under P fertilization) increased, and had a seasonal variation. There were no significant differences in the nitrogen return amount (60.05-71.47 kg x hm(-2) x a(-1)) and phosphorus return amount (2.94-3.93 kg x hm(-2) x a(-1)) of the litterfall among different treatments. As compared to the control, the N:P ratio of the litterfall under fertilization was higher at the beginning of the experiment (spring, 2011), but lower in the winter, 2011, indicating that nitrogen and phosphorus fertilization could alter the original nitrogen and phosphorus limited conditions of the studied forest ecosystem.

Case report. Efficacy of Hoodia for weight loss: is there evidence to support the efficacy claims?

Increasing rates of adult obesity and its negative health consequences are likely to become an increasing burden to the Canadian health care system. Consumers are looking for treatment options and often try the natural health products that are heavily promoted as safe, fast and effective. In this case report, MH, a 57-year-old overweight female wanted advice regarding whether she should use the natural product Hoodia to help her attain her weight loss goals. A literature search was conducted using Medline, EMBASE, the Cochrane Library, Natural Medicines Comprehensive Database and IPA from inception to March 2009. The internet, files of the authors and bibliographies of articles were searched for additional references. No published, peer-reviewed randomized controlled trials examining efficacy of Hoodia were found. Unpublished data from two small trials reported promising results with no adverse events. However, this leaves many unanswered questions regarding the use of Hoodia for weight loss such as the appropriate dose and duration, short and long term safety and use in patients with concomitant diseases. Literature suggests that some commercial products may not actually contain Hoodia at all. Additionally, Hoodia is not yet listed in the Canadian Licensed Natural Health Products Database meaning products sold in Canada may not meet Canadian regulatory standards. Upon discussing this information, MH decided not to use Hoodia, and other evidence-based recommendations were discussed.

[Effects of tree diameter at breast height and soil moisture on transpiration of Schima superba based on sap flow pattern and normalization].

The eigenvalues of continuous sap flow pattern, i. e. , skewness and kurtosis, were used to investigate the water usage of Schima superba with different diameter at breast height (DBH), and the method of normalization was firstly applied to eliminate the effects of strong affecting factor (photosynthetic active radiation, PAR) to explore the possible relationship between weak affecting factor (soil moisture) and sap flow. Generally, the trees with larger DBH had smaller skewness of sap flux density and later-appeared but larger peak values, suggesting that much more water was transpired, and the larger trees showed smaller skewness and later-appeared larger peak values in wet season than in dry season, suggesting that more water was transpired in wet season. On the other hand, smaller trees had lesser differences in the skewness between dry and wet seasons, suggesting that there was no significant difference in the transpiration between the two seasons. The relationship between individual tree's transpiration and soil moisture was significant and positive after the two parameters being normalized with PAR peak values. When the soil moisture content was higher, the transpiration of the trees with larger DBH was steadily increasing with soil moisture, while that of the trees with moderate or smaller DBH had opposite trend, presumably due to their transpiration and water absorption were approached to the limit.

Distribution and biosynthesis of theanine in Theaceae plants.

The theanine content of the leaves of 27 species or varieties of Theaceae plants was investigated. Theanine was present in 21 species or varieties, but in much lower amounts (<0.2 mumol/g fresh weight) than the quantity detected in Camellia sinensis var. sinensis. The major free amino acids in leaves of four species belonging to the genera Schima and Eurya, were glutamic acid, aspartic acid, glutamine, asparagine, alanine and proline and content of these amino acids is similar to or higher than theanine. Accumulation of free amino acids in these plants was generally lower than in C. sinensis var. sinensis. The biosynthetic activity of theanine, assessed by the incorporation of radioactivity from [(14)C]ethylamine, was detected in seedlings of two species of Schima. The theanine biosynthetic activity in roots was higher than that of leaves.

[Provenance difference in stable carbon isotope discrimination of Schima superba].

The difference in leaf stable carbon isotope discrimination (delta) of 18 representative Schima superba provenances was investigated based on three provenance trails located at Jian' ou of Fujian Province and Chun' an and Qingyuan of Zhejiang Province, and the geographic variation pattern and the effects of trial site and provenances growth rate were studied. Significant differences in leaf delta were observed among the provenaces, and the differences in leaf delta value between the highest and lowest provenances reached 6.9%, 3.0%, and 3.7% in 3 experimental sites, respectively. A classic latitudinal clinical variation pattern was found, because there were significant correlations between the leaf delta value and the latitude of seed sources in all the 3 sites while no significant correlations were observed between the leaf delta value and the longitude of the seed sources. The southern provenances showed higher leaf delta value than the northern provenances, indicating that the water use efficiency (WUE) was lower in southern provenances. The leaf delta value of different provenances was demonstrated to be greatly affected by the environment of trial sites. The leaf delta value increased significantly with the improvement of site environment and with the increase of annual rainfall. Significant positive correlations were observed between the leaf delta value and the growth traits including tree height, DBH, total number of lateral branches, and length of the strongest lateral branch, which indicated that the provenances with higher growth rate and denser crown had larger leaf delta value. Two and four superior provenances with high growth rate and low leaf delta value (or high WUE) were selected for Jian' ou of Fujian and Chun' an of Zhejiang, respectively.

[Effects of soil moisture condition and phosphorus supply on the seedlings growth and phosphorus efficiency of Schima superba provenances].

Taking four representative Schima superba provenances from Longquan in Zhejiang, Jian'ou and Youxi in Fujian, and Ji'an in Jiangxi as test materials, and with the S. wallichii from Guangxi as the contrast, a pot experiment was conducted to study their seedlings growth and physiological responses under effects of different soil moisture condition and phosphorus supply. Significant differences were observed in the seedlings growth, root morphological parameters, and phosphorus absorption efficiency among the provenances under different soil moisture condition and phosphorus supply. The provenances from Jian'ou in Fujian and Longquan in Zhejiang exhibited larger growth increment, more developed root, and higher phosphorus absorption efficiency, compared with those from Youxi in Fujian and Ji'an in Jiangxi, while the S. wallichii from Guangxi Province remained its original characteristics of fast growing and drought and infertility resistance. Soil moisture condition and phosphorous supply had significant effects on the growth and development of S. superba. The seedlings growth, dry matter accumulation, root parameters, and phosphorus absorption efficiency were 18.5%-105.6% higher under appropriate soil moisture condition than under drought stress, and 37.5%-286.2% higher under high phosphorus than under low phosphorus supply. However, under drought stress and low phosphorus supply, S. superba seedlings allocated more photosynthetic products to their roots, and had higher phosphorus use efficiency, which could be an important physiological mechanism of the adaptation to drought stress and low phosphorus supply. Relative to the significant independent effects of provenance, watering level, and phosphorus supply, no significant interactive effects of the three factors were observed.

Effects of acidic solutions on element dynamics in the monsoon evergreen broad-leaved forest at Dinghushan, China. Part 2: dynamics of Fe, Cu, Mn and Al.

BACKGROUND: Soil metal dynamics are affected by acid deposition. Little knowledge is available about the process in the lateritic soils under the monsoon forest in south China. METHODS: Samplings of Acmera acuminatissima, Cryptocarya concinna and Schima superba were grown from October, 2000 to July, 2002 in pots with a natural acid lateritic forest soil from Dinghushan. Pots were watered weekly with an acid solution (pH 3.05, 3.52, 4.00 or 4.40) or with tap water. Fe, Mn, Cu and Al were measured in soils, leachates and sapling leaves. RESULTS: Soil extractable Fe and leachate Al and Mn concentrations increased with a decreasing treatment pH. Soil reactive Al exhibited the opposite trend and decreased over time. The Ca/Al and Mg/ (Al+Mn) ratios did not decrease in the leaves of Schima superba, but decreased with a decreasing treatment pH for Cryptocaria concinna. Both ratios only decreased in the pH 3.05 treatment for Acmena CONCLUSIONS: Cu will not be toxic for plants since soil extractable Cu was not high and Fe will not be toxic either given that its root uptake was inhibited by Mn. Acid rains will lead to increased Mn and Al mobility in soil. Cryptocaria concinna will be the most sensible species to these changes (nutrient deficiency and direct Mn toxicity), while Schima superba should retain a good growth.

Simulated effects of acidic solutions on element dynamics in monsoon evergreen broad-leaved forest at Dinghushan, China. Part 1: dynamics of K, Na, Ca, Mg and P.

BACKGROUND: Acid deposition has become a concern in south China in recent years. This phenomenon has increased to a dramatic extent with the large use of cars and coal-fueled power plants. As a consequence, soils are becoming acidified and their element dynamics will change. A decrease in the nutrient availability will lead to slower plant growth and maybe to a change in the forest type with current species being replaced by new ones with less nutrient requirements. Because of these reasons, it is important to understand how the dynamics of elements will change and what mechanism is part of the process. This knowledge is important for modeling the acidification process and either finding ways to counter it or to predict its consequences. The primary purpose of this study was to provide information about how the dynamics of K, Na, Ca, Mg and P are affected by acid deposition in a typical forest in southern China. METHODS: Experimental soils and saplings were collected directly from the monsoon evergreen broad-leaved forest in Dinghushan. All saplings were transplanted individually into ceramic pots in August 2000 and placed in an open area near their origin site. Pot soils were treated weekly from October 2000 to July 2002 with an acidic solution at pH 3.05, pH 3.52, pH 4.00 or pH 4.40, or with tap water as a control. The concentrations of SO4(2-), NO3-, K+, Na+, Ca2+, Mg2+ and available P and the pH were measured in soil and leachate samples taken at different times. The sapling leaves were collected and their element concentrations were measured at the end of the experiment. RESULTS AND DISCUSSION: Concentrations of soil exchangeable Ca and Mg decreased quickly over time, although only Ca showed changes with the acidic solution treatment and soil exchangeable K was stable because of soil weathering. Leaching of K, Mg and Ca was dependent upon the treatment acidity. Soil available P decreased slowly without any correlation with the acidity of the treatment. All the NO3- added by the treatment was taken up by the plants, but the SO4(2-) added accumulated in the soil. Amongst the plant species, Schima superba was little affected by the treatment, the leaf P content was affected in Acmena acuminatissima plants and Cryptocarya concinna was the most susceptible species to soil acidification, with a marked decrease of, the leaf K, Ca and Mg concentrations when the treatment acidity increased. CONCLUSIONS: Simulated acid deposition affected the dynamics of K, Ca and Mg in the monsoon evergreen broad-leaved forest. The dynamics of Ca in the soil and of K, Mg and Ca in the soil leachates were affected by the acidic solution treatment. If such a soil acidification occurs, Cryptocarya concinna will be amongst the first affected species, but Schima superba will be able to sustain a good growth and mineral nutrition. RECOMMENDATIONS AND PERSPECTIVES: Acid deposition will lead to imbalance the nutrient elements in the evergreen broad-leaved forest because of accelerated leaching losses of soil exchangeable Ca and Mg. Measures should be developed to slow down soil acidification or nutrient decrease.