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Tea plant (Camellia sinensis L.) is capable of accumulating a large amount of fluorine (F) in leaves without showing toxicity symptoms and thus offers a good model for exploring F tolerance mechanisms. Here, gas chromatography time-of-flight mass spectrometry (GC-TOF–MS) was used to investigate metabolic changes in leaves of tea seedlings under control (0 mM), low F (0.2 mM) and high F (0.8 mM) conditions. Differentially changed metabolites such as galacturonic acid, lactose, fructose, malic acid, alanine were identified by the comparison among the three F treatment groups. A pathway map depicted based on the KEGG database reflected the involvement of pectin biosynthesis metabolism in F stress response. The gene expression and enzyme activity of key enzymes involved in pectin biosynthesis pathway and the content of pectic polysaccharides were increased by exogenous F treatments, indicating the promotion effect of F on the pectin biosynthesis. Pectin was also immunochemically stained in vivo using monoclonal antibody (2F4), which confirmed the increment. The increased pectin might contribute to combining the exogenous F in tea leaves. This research provided some novel insights into further research on F detoxification of plants.

The tea plant [Camellia sinensis (L.) O. Kuntze] is a species with a high concentration of fluorine in its leaves, especially in the mature leaves. The physiological mechanisms for fluorine absorption and accumulation have been well studied, but the related molecular mechanisms are poorly understood in the tea plant. In this study, transcriptome analysis by RNA-Seq following exposure to 16 mg/L of fluorine for 0, 3, 6, and 24 h was performed to identify the candidate genes involved in the transmembrane transportation of fluorine. More than 1.23 billion high-quality reads were generated, and 259.84 million unigenes were assembled de novo, with 518 216 of them being annotated in the seven databases used. Meanwhile, a large number of transporters, transcription factors, and heat-shock proteins with differential expression in response to high levels of fluorine (P ≤ 0.05) were identified. Comparative transcriptome analysis showed that the uptake of fluorine is related to photosynthesis, plant hormone signal transduction, and glutathione metabolism. Further systematic analysis of nitrate and potassium transporter genes revealed that many of these genes regulate fluorine transportation in roots and leaves. Gene expression and fluorine content analysis in different cultivars revealed CsNRT1/PTR 3.1 and CsPT 8 as the key genes regulating the transmembrane transportation of fluorine in the tea plant.

Volatile components in fresh leaves are involved in the regulation of many stress responses, such as insect damage, fungal infection and high temperature. However, the potential function of volatile components in hyperosmotic response is largely unknown. Here, we found that 7-day hyperosmotic treatment specifically led to the accumulation of (Z)-3-hexen-1-ol, (E)-2-hexenal and methyl salicylate. Transcriptome and qRT-PCR analyses suggested the activation of linolenic acid degradation and methyl salicylate processes. Importantly, exogenous (Z)-3-hexen-1-ol pretreatment dramatically enhanced the hyperosmotic stress tolerance of tea plants and decreased stomatal conductance, whereas (E)-2-hexenal and methyl salicylate pretreatments did not exhibit such a function. qRT-PCR analysis revealed that exogenous ABA induced the expressions of related enzyme genes, and (Z)-3-hexen-1-ol could up-regulate the expressions of many DREB and RD genes. Moreover, exogenous (Z)-3-hexen-1-ol tremendously induced the expressions of specific LOX and ADH genes within 24 h. Taken together, hyperosmotic stress induced (Z)-3-hexen-1-ol accumulation in tea plant via the activation of most LOX, HPL and ADH genes, while (Z)-3-hexen-1-ol could dramatically enhance the hyperosmotic stress tolerance via the decrease of stomatal conductance and MDA, accumulation of ABA and proline, activation of DREB and RD gene expressions, and probably positive feedback regulation of LOXs and ADHs. KEY MESSAGE: Hyperosmotic stress induced (Z)-3-hexen-1-ol accumulation in Camellia sinensis via the up-regulation of most LOX, HPL and ADH genes, while (Z)-3-hexen-1-ol could dramatically enhance the hyperosmotic stress tolerance via the decrease of stomatal conductance, accumulation of proline, activation of DREB and RD gene expressions, and probably positive feedback regulation of LOXs and ADHs.

The freshness and color quality of postharvest tea leaves can be markedly prolonged and retained by proper preservation measures. Here, we investigated the dynamic changes of chlorophyll and its derivatives in postharvest tea leaves under different low-temperature treatments using natural withering as a control. Chlorophyll decomposition was found closely related with chlorophyllide, pheophorbide, and pheophytin. Low-temperature withering could slow chlorophyll degradation in postharvest tea leaves via significant inhibition on the enzyme activity and gene expression of Mg-dechelatase, chlorophyllase, and pheophorbide a oxygenase. At the initial stage of withering, a significant increase was observed in the chlorophyll content, expression of chlorophyll-synthesis-related enzymes (such as glutamyl-tRNA synthetase, etc.), and chlorophyll synthase activity in newly picked tea leaves. Moreover, an obvious decrease was found in the content of l-glutamate as the foremost precursor substance of chlorophyll synthesis. Hence, our findings revealed that the chlorophyll synthesis reaction was induced by the light-dehydration-stress in the initial withering of tea leaves. This study provides a theoretical basis for exploring preservation technology in actual green tea production.

Recently, more and more attention has been given to a semiconductor oxide-based surface-enhanced Raman spectroscopy substrate for its great stability and biocompatibility. However, its poor SERS sensitivity limits the applications of semiconductor oxide SERS substrates. In this paper, we provide a facile reduction method to modulate oxygen vacancy concentrations in oxide SERS substrates. Using MoO2 as an example, the resonance coupling as well as charge transfer between the semiconductor oxide SERS substrate and the target molecules were promoted for the reason of artificial oxygen vacancy embodied in the Raman signals being improved. By using the TEM, SEM, and XPS measurements, we confirmed that we successfully prepared defective MoO2- x with a polycrystalline surface. MoO2- x modulated oxygen vacancy treated with 6 wt % Li shows a very high detection sensitivity of 10-8 M (4.79 ug/L) for R6G, and the intensity of the Raman signal was highly enhanced. Because of the existence of defective energy levels, resonance coupling, as well as charge transfer between semiconductor and molecules, was obviously promoted. More importantly, the method of modulating oxygen vacancy can be widely used in semiconductor oxide materials for its chemical enhancement capacity can be promoted by artificial oxygen vacancy.

The glycosides are presumed to influence the quality of green tea but the molecular mechanism behind remains unclear. To elucidate the contribution of glycosides to the flavor formation of green tea, changes of both glycosidically bound non-volatiles (GBNVs) and glycosidically bound volatiles (GBVs) during the manufacturing of green tea were investigated using a modification-specific metabolomics method. A total of 64 glycosides (47 GBNVs and 17 GBVs) were identified and their contents mainly changed during the pan firing and drying stages of green tea manufacturing. Notably, the contents of GBVs significantly increased by 1.12-4.46-fold during pan firing. Correlation analysis showed that the GBVs contents were negatively related to the contents of volatiles and glucose. Model experiments revealed that enzymatic synthesis contributed to the increase in the content of GBVs during the pan firing. This comprehensive study on the glycosides changes revealed the molecular bases for GBVs increments during the pan firing.

Histone methylation plays an important regulatory role in the drought response of many plants, but its regulatory mechanism in the drought response of the tea plant remains poorly understood. Here, drought stress was shown to induce lower relative water content and significantly downregulate the methylations of histone H3K4 in the tea plant. Based on our previous analysis of the SET Domain Group (SDG) gene family, the full-length coding sequence (CDS) of CsSDG36 was cloned from the tea cultivar ‘Fuding Dabaicha’. Bioinformatics analysis showed that the open reading frame (ORF) of the CsSDG36 gene was 3138 bp, encoding 1045 amino acids and containing the conserved structural domains of PWWP, PHD, SET and PostSET. The CsSDG36 protein showed a close relationship to AtATX4 of the TRX subfamily, with a molecular weight of 118,249.89 Da, and a theoretical isoelectric point of 8.87, belonging to a hydrophilic protein without a transmembrane domain, probably located on the nucleus. The expression of CsSDG36 was not detected in the wild type, while it was clearly detected in the over-expression lines of Arabidopsis. Compared with the wild type, the over-expression lines exhibited lower hyperosmotic resistance by accelerating plant water loss, increasing reactive oxygen species (ROS) pressure, and increasing leaf stomatal density. RNA-seq analysis suggested that the CsSDG36 overexpression caused the differential expression of genes related to chromatin assembly, microtubule assembly, and leaf stomatal development pathways. qRT-PCR analysis revealed the significant down-regulation of stomatal development-related genes (BASL, SBT1.2(SDD1), EPF2, TCX3, CHAL, TMM, SPCH, ERL1, and EPFL9) in the overexpression lines. This study provides a novel sight on the function of histone methyltransferase CsSDG36 under drought stress.

The identification of aroma composition and key odorants contributing to aroma characteristics of white tea is urgently needed, owing to white tea&rsquo, s charming flavors and significant health benefits. In this study, a total of 238 volatile components were identified in the three subtypes of white teas using headspace solid-phase microextraction (HS-SPME) combined with comprehensive two-dimensional gas chromatography&ndash, time-of-flight mass spectrometry (GC &times, GC&ndash, TOFMS). The multivariate statistical analysis demonstrated that the contents of 103 volatile compounds showed extremely significant differences, of which 44 compounds presented higher contents in Baihaoyinzhen and Baimudan, while the other 59 compounds exhibited higher contents in Shoumei. The sensory evaluation experiment carried out by gas chromatography&ndash, olfactometry/mass spectrometry (GC&ndash, O/MS) revealed 44 aroma-active compounds, of which 25 compounds were identified, including 9 alcohols, 6 aldehydes, 5 ketones, and 5 other compounds. These odorants mostly presented green, fresh, floral, fruity, or sweet odors. Multivariate analyses of chemical characterization and sensory evaluation results showed that high proportions of alcohols and aldehydes form the basis of green and fresh aroma characteristic of white teas, and phenylethyl alcohol, &gamma, Nonalactone, trans-&beta, ionone, trans-linalool oxide (furanoid), &alpha, ionone, and cis-3-hexenyl butyrate were considered as the key odorants accounting for the different aroma characteristics of the three subtypes of white tea. The results will contribute to in-depth understand chemical and sensory markers associated with different subtypes of white tea, and provide a solid foundation for tea aroma quality control and improvement.

The vigor of tea plants (Camellia sinensis) and tea quality are strongly influenced by the abundance and forms of nitrogen, principally NO3&minus, NH4+, and amino acids. Mechanisms to access different nitrogen sources and the regulatory cues remain largely elusive in tea plants. A transcriptome analysis was performed to categorize differentially expressed genes (DEGs) in roots and young leaves during the early response to four nitrogen treatments. Relative to the continuously nitrogen-replete control, the three nitrogen-deprived and resupplied treatments shared 237 DEGs in the shoots and 21 DEGs in the root. Gene-ontology characterization revealed that transcripts encoding genes predicted to participate in nitrogen uptake, assimilation, and translocation were among the most differentially expressed after exposure to the different nitrogen regimes. Because of its high transcript level regardless of nitrogen condition, a putative amino acid transporter, TEA020444/CsCAT9.1, was further characterized in Arabidopsis and found to mediate the acquisition of a broad spectrum of amino acids, suggesting a role in amino acid uptake, transport, and deposition in sinks as an internal reservoir. Our results enhance our understanding of nitrogen-regulated transcript level patterns in tea plants and pinpoint candidate genes that function in nitrogen transport and metabolism, allowing tea plants to adjust to variable nitrogen environments.

Wild teas are valuable genetic resources for studying domestication and breeding. Here we report the assembly of a high-quality chromosome-scale reference genome for an ancient tea tree. The further RNA sequencing of 217 diverse tea accessions clarifies the pedigree of tea cultivars and reveals key contributors in the breeding of Chinese tea. Candidate genes associated with flavonoid biosynthesis are identified by genome-wide association study. Specifically, diverse allelic function of CsANR, CsF3’5’H and CsMYB5 is verified by transient overexpression and enzymatic assays, providing comprehensive insights into the biosynthesis of catechins, the most important bioactive compounds in tea plants. The inconspicuous differentiation between ancient trees and cultivars at both genetic and metabolic levels implies that tea may not have undergone long-term artificial directional selection in terms of flavor-related metabolites. These genomic resources provide evolutionary insight into tea plants and lay the foundation for better understanding the biosynthesis of beneficial natural compounds., Wild teas are considered as valuable resource for studying domestication and breeding. Here, Zhang et al. report genome of wild tea DASZ and transcriptome of 217 accessions, which clarify pedigree of Chinese tea cultivars and show tea may not have undergone long-term artificial directional selection on flavor-related metabolites.

Tea plant (Camellia sinensis) may hyperaccumulate fluorine (F) in its leaves, which may cause fluorosis in tea consumers. Recent studies have implied that exogenous calcium (Ca) may reduce F in tea leaves, although our mechanistic understanding of this phenomenon remains limited. Here, the effects of exogenous Ca on the physiological, biochemical and ionic homeostasis of tea leaves were investigated in the presence and absence of F. Elevated levels of malondialdehyde (MDA) and impaired cellular ultrastructure indicated that exogenous F induced stress in tea plants subjected to deficient Ca (0.01, 0.05 mM) and extremely excessive Ca (10 mM) treatments. Additionally, more F were accumulated in leaves compared to the control when tea plants were treated with 0.5 mM Ca. The lowest levels of MDA and F were observed at an optimal level of 5 mM Ca. F increased the levels of caffeine, polyphenols, and catechins, but decreased the content of soluble sugars and gallic acid when the level of Ca was within 5 mM. Moreover, based on a multivariate analysis on ionic composition, the Ca-regulated disorder in the homeostasis of B, Al, Cu, and Zn was strongly correlated with the accumulation of F. Our results demonstrate that within a range of concentrations, exogenous Ca was able to reduce F content and enhance F tolerance in tea leaves. These effects of exogenous Ca on F tolerance may be related to ionic homeostasis.

This study investigated the effects of conventional hot-air drying, microwave drying, far-infrared drying, halogen lamp drying and halogen lamp-microwave combination drying on sensory and chemical quality of black tea. Sensory evaluation revealed that the black tea dried with halogen lamp-microwave and microwave got more uniform black color, fresher taste and higher sweet aroma. Total quality score was in the order: halogen lamp-microwave dried tea (89.3) > microwave dried tea (88.2) > halogen lamp dried tea (86.4) > far-infrared dried tea (85.2) > hot-air dried tea (83.1). The highest contents of polyphenols, catechins and theaflavins were found when black tea dried with microwave. Far-infrared drying and hot-air drying could remarkably increase the contents of amino acids and soluble sugars. The volatiles of black tea were identified with gas chromatography-mass spectrometry. Microwave dried black tea got the maximum contents of volatile compounds, followed by halogen lamp-microwave dried tea, far-infrared dried tea, halogen lamp dried tea and hot-air dried tea. Overall, microwave drying and halogen lamp-microwave drying could ameliorate the drying process of black tea.

As one of the most popular beverages globally, tea has enormous economic, cultural, and medicinal importance that necessitates a comprehensive metabolomics study of this species. In this study, a large-scale targeted metabolomics analysis on two types of leaf tissues of nine tea cultivars from five representative geographical origins within China was carried out using the liquid chromatography-mass spectrometry technique. RNA-seq-based transcriptomic analysis was in parallel conducted on the same samples, and gene expression and metabolic differentiation between tissues as well as between the multiple tea cultivars were investigated. The data obtained provide an accessible resource for further studies of naturally occurring metabolic variation of tea plants, which will aid in thoroughly interpreting the underlying genetic and molecular mechanisms of biosynthesis of specialized metabolites in this critical species. Candidate genes including a transcription factor (CsMYB5-like), which were highly correlated with both the content of flavonoids and the expression level of genes participating in the phenylpropanoid and flavonoid biosynthesis pathway, were identified as potential targets for quality improvement of tea.

The tea plant (Camellia sinensis (L.) O. Kuntze) is indigenous to China, where its wild ancestors are broadly distributed in Southwest China. As an aluminum (Al) accumulator, tea plant is very tolerant to Al and accumulates Al at high levelin the leaves. Here an Al tolerant transcription factor of CsSTOP1 was characterized and assumed to regulate multiple genes critical for Al tolerance. The transcriptional regulations by STOP1-like proteins were conserved and conferred the ability to survive in acid soil. Furthermore, a 9-bp deletion was found in five varieties of assamica subspecies and CsSTOP1Mkdy-OE Arabidopsis lines showed more tolerant to Al than CsSTOP1JM1-OE lines, which might be the natural selection of the genetic variation for the tea plant’s adaptation to acidic soil. Given the CsSTOP1Mkdy allele more tolerant to Al and tea plant gradually spreading from the original center of Southwest China, this present study suggests that CsSTOP1 is labelled as an ‘adaptive’ trait that increases tea plant fitness in a particular environmental context of rhizotoxicity Al toxicity in acid soil. The qPCR result suggests the 9-bp deletion is not responsible for transcriptional activity while this deletion may affect the transcriptional regulation level.

In this paper, the flower-shaped silver nanoparticle was used as substrate to investigate the SERS effect of pesticide molecules under different treatment conditions. Due to the variation of molecular structure and molecular interactions with pH value and solvent type, the combination mode of molecules and metal surface and electron transfer also changes, resulting in different SERS effects with a certain regularity. It was demonstrated that the optimum pH values for test conditions are in the range of 6.66–11.11 due to the auxiliary of H + ions and undestroyed complete functional groups of ethion. Meanwhile, the ethion in the acetone was the best in the three solvents (ultrapure water, absolute ethanol and acetone). Under optimal conditions, ethion can be detected with a minimum detectable concentration of 10 −10 M. The regression model obtained through principal component analysis (PCA) can be used for rapid and precise detection of the concentration of ethion, suggesting the importance of condition optimization for SERS test. Therefore, the solvent environment can be effectively improved to promote non-resonance enhancement of chemical bonding between adsorbate and metal substrate and resonance enhancement of light-induced charge transfer in molecular-metal system by excitation light. The solvent environment has significant effect on the SERS sensor from chemical enhancement.

Surface Enhanced Raman Scattering (SERS) spectroscopy technology is widely used in materials analysis, environmental monitoring, biomedical, food security and other fields. Flower-shaped silver nanoparticles have been successfully synthesized by a simple aqueous phase silver nitrate reduction by ascorbic acid in the presence of polyvinylpyrrolidone (PVP) surfactant. The nanoparticles diameters were adjusted from 450 to 1000nm with surface protrusions up to 10-25nm. The flower-shaped silver nanostructures obtained were used as stable SERS substrates with high SERS activity for detecting Rhodamine 6G (R6G), at a concentration of only 10-9mol/L, where the SERS signal is still clear. SERS spectroscopy of four different food colorants (e.g. food blue, tartrazine, sunset yellow, acid red) were analysed and the characteristic bands were identified. An improved principle component analysis (PCA) was used for four different food colorants detection, at concentrations down to about 10-8mol/L. Thus, the LOD of food blue, tartrazine, sunset yellow and acid red are 79.285μg/L, 5.3436μg/L, 45.238μg/L and 50.244μg/L, respectively.

Flower-shaped silver nanoparticles have been successfully synthesized by a simple aqueous phase silver nitrate reduction by ascorbic acid in the presence of polyvinylpyrrolidone (PVP) surfactant. The nanoparticles diameters were adjusted from 450 to 1000nm with surface protrusions up to 10-25nm. The growth direction of silver nuclei is controlled by their degree of coating by PVP. The flower-shaped silver nanostructures obtained were used as stable Surface Enhanced Raman Scattering (SERS) substrates with high SERS activity for detecting Rhodamine 6G (R6G), at a concentration of only 10-9M, where the SERS signal is still clear. SERS spectra of the dye carmine was analysed and the characteristic bands were identified. An improved principle component analysis (PCA) was used for carmine detection, at concentrations down to 10-8M. The characteristic peaks of the carmine (1019, 1360, and 1573cm-1) remained at 10-8M. This indicated that the minimum detection limit of AgNP-based substrate for carmine is about 10-8M.

The objective of the present study was to evaluate the effect of steeping temperature on the biological activities of green tea, including the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging capacity, α-glucosidase and α-amylase inhibitory activities, and glucose uptake inhibitory activity in Caco-2 cells. Results showed that, with increasing extraction temperature, the polyphenol content increased, which contributed to enhance antioxidant activity and inhibitory effects on α-glucosidase and α-amylase. Green tea steeped at 100°C showed the highest DPPH radical-scavenging activity and inhibitory effects on α-glucosidase and α-amylase activities with EC50 or IC50 values of 6.15μg/mL, 0.09mg/mL, and 6.31mg/mL, respectively. However, the inhibitory potential on glucose uptake did not show an upward trend with increasing extraction temperature. Green tea steeped at 60°C had significantly stronger glucose uptake inhibitory activity (p

A novel disease characterized by small brown-black spots (1 to 2 mm in diameter) on tender tea leaves (Camellia sinensis) has been observed in many regions of Hubei Province, China, which severely affects the yield and quality of tea. Tea leaf samples with typical symptoms were collected from three major tea-cultivation regions of Hubei, and were subjected to pathogen isolation for etiological analysis. As a result, 34 Pestalotiopsis isolates were obtained from 20 samples, and they were identified as Pestalotiopsis theae (14 isolates), P. camelliae (12), and P. clavispora (8), determined by morphologies and phylogenetic analysis based on internal transcribed spacer, and partial β-tubulin and translation elongation factor 1-alpha genes. Pathogenicity tests on detached tea leaves showed that no matter what mycelial discs or conidium suspensions were used, inoculation of the Pestalotiopsis fungi could result in small brown-black spots (1 to 2 mm in diameter) on wounded leaves, similar to those observed in the field in the sizes and colors. It also revealed that only P. theae had pathogenicity on unwounded tea leaves, and P. theae and P. clavispora showed significantly higher virulence than P. camelliae. Inoculation test with conidium suspension on intact tea leaves in the field further confirmed that P. theae as the pathogen of brown-black spots. Reisolation of the pathogens from diseased leaves confirmed that the symptom was caused by the inoculation of Pestalotiopsis fungi. The P. theae isolates responsible for brown-black spots were also compared with those for tea gray blight disease in growth rate, pathogenicity, and molecular characteristics in parallel. To our knowledge, this is the first report that the Pestalotiopsis fungi cause brown-black spot disease on tender tea leaves. The results provide important implications for the prevention and management of this economically important disease.

A total of 299,113 unigenes were generated and 15,817 DEGs were identified. We identified candidate genes associated with the regulation of catechins biosynthesis during leaf development in tea plant. The tea plant (Camellia sinensis (L.) O. Kuntze) is one of the most economically significant crops worldwide because of its positive effects on human health. The health benefits of tea are mainly attributed to catechins, which are the predominant polyphenols that accumulate in tea. Catechins are products of the phenylpropanoid and flavonoid biosynthetic pathways. Although catechins were identified in tea leaves long ago, the molecular mechanisms regulating catechins biosynthesis remain unclear. To identify candidate genes involved in catechins biosynthesis, we analyzed the transcriptomes of tea leaves during five different leaf stages of development using RNA-seq. Approximately 809 million high-quality reads were obtained, trimmed, and assembled into 299,113 unigenes with an average length of 565 bp. A total of 15,817 unigenes were differentially expressed during the different stages of leaf development. These differentially expressed genes were enriched in a variety of processes such as the regulation of the cell cycle, starch and sucrose metabolism, photosynthesis, phenylpropanoid biosynthesis, phenylalanine metabolism, and flavonoid biosynthesis. Based on their annotations, 51 of these differentially expressed unigenes are involved in phenylpropanoid and flavonoid biosynthesis. Furthermore, transcription factors such as MYB, bHLH and MADS, which may involve in the regulation of catechins biosynthesis, were identified through co-expression analysis of transcription factors and structural genes. Real-time PCR analysis of candidate genes indicated a good correlation with the transcriptome data. These findings increase our understanding of the molecular mechanisms regulating catechins biosynthesis in the tea plant.

The tea plant ( Camellia sinensis L. O. Kuntze) is a high aluminum (Al) tolerant and accumulator species. Candidate genes related to Al tolerance in tea plants were assembled based on de novo transcriptome analysis. The homologs implied some common and distinct Al-tolerant mechanism between tea plants and rice, Arabidopsis and buckwheat. In addition to high Al tolerance, the tea plant exhibits good performance exposure to a proper Al level, and accumulates high Al in the leaves without any toxicity symptom. Therefore, Al was considered as a hyperaccumulator and beneficial element for tea plants. However, the whole-genome molecular mechanisms accounting for Al-tolerance and accumulation remain unknown in tea plants. In this study, transcriptome analysis by RNA-Seq following a gradient Al-level exposure was assessed to further reveal candidate genes involved. Totally more than 468 million high-quality reads were generated and 213,699 unigenes were de novo assembled, among which 8922 unigenes were all annotated in the seven databases used. A large number of transporters, transcription factors, cytochrome P450, ubiquitin ligase, organic acid biosynthesis, heat shock proteins differentially expressed in response to high Al (P ≤ 0.05) were identified, which were most likely ideal candidates involved in the Al tolerance or accumulation. Furthermore, a few of the candidate Al-responsive genes related to Al sequestration, cell wall modification and organic acid excretion have been well elucidated as was already found in Arabidopsis, rice, and buckwheat. Thus, some consistent Al-tolerance mechanisms across the species are indicated. In conclusion, the transcriptome data provided useful insights of promising candidates for further characterizing the functions of genes involved in Al tolerance and accumulation in tea plants.

A series of incubation systems of pure (−)-Epigallocatechin gallate (EGCG), ascorbic acid (AA) and dehydroascorbic acid (DHAA) at 80 °C were performed to investigated the effect and mechanism of AA on the stability of EGCG. Results shows the dual function of AA, protect action at low concentration and promoting degradation at high concentration, and the critical concentration is about 10 mmol/L. The protective properties of AA due to the reversible reaction from AA to DHAA inhibiting oxidation pathway of EGCG to EGCG quinone or other activated intermediates, and both AA and DHAA can inhibit the hydrolysis of EGCG. The properties of promoting degradation is mainly due to the fact that DHAA, the oxidation product of AA, can react with EGCG to generate some ascorbyl adducts of EGCG. This result is helpful to control the stability of catechins and further clarify the complex interaction on healthy between EGCG and AA.

The influence mechanism of different withering methods (CK, indoor natural spreading; LTD, low-temperature plus dark; LTY, low-temperature plus yellow-light; LTCD, low-temperature plus CO2) on non-volatile compounds in postharvest tea leaves was investigated by UHPLC-Q-TOF/MS-based non-targeted metabolomic and transcriptomic analyses. Compared with CK, low-temperature withering could slow down polyphenol oxidation by inhibiting polyphenol oxidase activity and keeping the expression of genes for flavanol synthesis. After withering, the proteinaceous amino acid content increased significantly, especially for LTCD and LTY, mainly due to increased peptidase activity and up-regulation of genes involved in the biosynthesis of valine, leucine, aspartic acid, glutamic acid, phenylalanine, and proline. Moreover, LTCD and LTY enhanced the synthesis of γ-aminobutyric acid and metabolism of phenylalanine-methyl salicylate and tryptophan-indole, respectively. Meanwhile, the transformation of theobromine to caffeine was accelerated under low-temperature withering. This research provides ageneticmetabolicbasis for the application of low-temperature withering to actual green tea processing.

Surface-enhanced Raman spectroscopy (SERS) has the potential to detect pesticide residues in agricultural products. However, some systemic pesticides, such as chlorpyrifos, can enter the plant tissue, and not just stay on the surface. Consequently, many SERS studies halted at practical application because of its complexity. In this work, SERS technology was used to detect chlorpyrifos residues in tea products at the semiquantitative level. A simple pretreatment method effectively avoided interference of other fluorescent substances, and all major peaks could be distinguished on the basis of a novel substrate. A principal component analysis algorithm was applied to form a regression model, and a nanogram detection limit was obtained. Furthermore, chlorpyrifos residues in the same tea products were also measured by gas chromatography–mass spectrometry, and the results show a small range of errors. From the comparative study of the two detection methods, the results suggest the great promise of SERS technology for rapid inspection of agricultural products.

Volatile compounds are important contributors to dark tea quality. However, the aroma formation during the key processing pile-fermentation remains unclear. Herein, a total of 46 compounds including 17 alcohols, nine ketones, eight alkenes, five aldehydes, four esters, one heterocyclic oxygen, one alkane, and one pyranoid were identified by GC-MS. At the end of pile-fermentation, alcohols and aldehydes decreased by 21.2% and 12.9% respectively, while ketones and alkenes increased by 40.3% and 11.5% respectively, esters doubled compared to the initial period. PLS-DA analysis demonstrated that the characteristic aroma components were mainly ionone, linalool and its oxides (floral, citrus-like, fruity), safranal (woody, herbal and spice), β-cyclocitral, β-terpinene, limonene (fruity), trans-2-hexenyl hexanoate (fragrance) and roses oxide (rose). Importantly, Pearson correlation analysis demonstrated microbial communities and volatile compounds were highly correlated (|r|>0.7, p

A kind of graphene oxide/silver (GO/Ag) nanocomposites with high Surface enhanced Raman scattering (SERS) activity were fabricated via a facile and green liquid phase reduction method. The synthesized materials were characterized in detail using various microscopic and spectroscopic techniques. In this method, the GO sheets worked as a holder which makes silver nanoparticles (AgNPs) aggregate to a particular morphology, and under a suitable dosage of silver ions, well-dispersed AgNPs on the surface of GO were obtained, which could generate more “hot spots” of SERS. Moreover, SERS technique based on the obtained GO/Ag nanocomposites was used as an effective way to detect l -Theanine. The detection limit was estimated to be as low as 10−7 M, and a multivariate linear regression model for the concentration of l -Theanine was established. The optimal fitting equation is Y = 5.6765 + 0.0307X1458 − 0.0267X1251.

Flower-like silver nanoparticles, with diameters of 450–500 nm and protrusions on the surface up to 10–15 nm, have been successfully synthesized via liquid phase reduction method, by using ascorbic acid as a reductant and polyvinyl pyrrolidone (PVP) as a surfactant. It is found that the proper coating degree of PVP on silver nuclei leads to anisotropic growth of the silver nuclei resulting in the formation of flower-like structure with more protrusions. Furthermore, when the obtained flower-like silver nanostructure is used as Surface Enhanced Raman Scattering(SERS) substrates, SERS signal is still clearly even if the concentration of R6G solution is reduced down to 10 −8 M. The SERS enhancement mechanism of flower-like silver nanostructures in SERS spectroscopy has been further discussed. SERS technology based on the obtained flower-like silver nanostructure substrates can be also used as an effective way to detect caffeine. In this work, it is found that the average intensity of SERS signals at 807 cm −1 (N C H), 958 cm −1 (Pyrimidine ring) and 1327 cm −1 (Imidazole trigonal ring) linear decreased with the −log 10 C (C is the concentration of caffeine), and the coefficient of determination (R 2 ) is as high as 0.975. This work provides a new way for the quantitative detection of caffeine based on SERS spectroscopy.

Self-incompatibility (SI) prevents inbreeding depression in angiosperms. Camellia sinensis is an important cash crop, but breeding improvements and genetic studies of the plant are hindered by SI. However, the SI mechanism in C. sinensis remains unclear. In this study, a putative S-RNase gene (KU852488) was cloned from C. sinensis . The full-length cDNA of CsS-RNase is 1121 bp, which encodes 238 amino acids. It shares the closest relationship with an S-RNase gene (ADA67883.1), which was cloned from a self-incompatibility Citrus reticulata cultivar ‘Wuzishatangju’. The expression level of CsS-RNase in the styles were 3–259 (‘Fuding Dabaicha’) and 5.6–119 (‘Zhongcha108’) times higher than the other tissues, for example petals, pollen grains, filaments and buds. And its expression rose in self-pollinated styles with 24 h earlier than cross-pollinated styles. The genotypes of CsS-RNase in 10 cultivars and one breeding line of C. sinensis were analyzed. Totally, 11 polymorphic amino acid residues were identified. A single nucleotide polymorphism (SNP) marker of CsS-RNase was developed. Finally, the CsS-RNase was mapped onto a reference genetic linkage map of tea plant.

Histone methylation mediated by SET Domain Group (SDG) proteins plays an important role in plant response to various abiotic stresses, regulating the expressions of many stress-inducible genes. However, little is known about the SDG genes in Camellia sinensis. In this study, we identified 46 CsSDGs by genome-wide search with the conserved SET domain. Phylogenetic tree, domain composition and gene structure analyses revealed that 46 CsSDGs were classified into five groups, including E(z), ASH1, TRX, SU(VAR) and class VI/VII genes. Furthermore, In silico and qRT-PCR analyses suggested the diverse expression patterns of the 46 CsSDGs in different tissues and in response to various abiotic stresses. To reveal the vital role of CsSDGs in plant responses to abiotic stresses, CsSDG43 was chosen for the functional analysis in hyperosmosis in Arabidopsis. Importantly, CsSDG43 significantly weakened the hyperosmotic tolerance by increasing water loss, reactive oxygen species (ROS) pressure and AHG3 expression in the overexpression lines compared with the wild type of Arabidopsis.

Tea plant is capable of hyper-accumulating fluoride (F) in leaves, suggesting drinking tea may cause excessive F intake in our body and threaten the health. This study investigated the changes in the structure, composition, and F content in the leaf cell wall of the tea (Camellia sinensis) under different F conditions to demonstrate the role of cell wall in F enrichment in tea plants. The cell wall was shown as the main part for F accumulation (67%-92%), with most of F distributed in the pectin fraction (56%-71%). With increasing F concentration, a significant increase (p 0.05) was observed in the F content of cell wall and its components, the level of cell wall metal ions (i.e. Cu, Mg, Zn, Al, Ca, Ba, Mn), as well as the content of total cell wall materials, cellulose, and pectin. Meanwhile, the level of Cu, Mg, Zn, pectin, and cellulose was significantly positively correlated with the F content in the leaf cell wall. F addition was shown to increase the fluorescence intensity of LM19 and 2F4 antibody-labeled low-methylesterified homogalacturonans (HGs), while decrease LM20-labeled high-methylesterified HGs, coupled with an increase in the activity and gene expression of pectin methyl esterases (PMEs) in tea leaves. All these results suggest that F addition can increase pectin content and demethylesterification, leading to increased absorption of metal cations and chelation of F in the cell wall through the action of metal ions.

Nitrogen (N) is a macroelement with an indispensable role in the growth and development of plants, and tea plant (Camellia sinensis) is an evergreen perennial woody species with young shoots for harvest. During senescence or upon N stress, autophagy has been shown to be induced in leaves, involving a variety of autophagy-related genes (ATGs), which have not been characterized in tea plant yet. In this study, a genome-wide survey in tea plant genome identified a total of 80 Camellia Sinensis autophagy-related genes, CsATGs. The expression of CsATG8s in the tea plant showed an obvious increase from S1 (stage 1) to S4 (stage 4), especially for CsATG8e. The expression levels of AtATGs (Arabidopsis thaliana) and genes involved in N transport and assimilation were greatly improved in CsATG8e-overexpressed Arabidopsis. Compared with wild type, the overexpression plants showed earlier bolting, an increase in amino N content, as well as a decrease in biomass and the levels of N, phosphorus and potassium. However, the N level was found significantly higher in APER (aerial part excluding rosette) in the overexpression plants relative to wild type. All these results demonstrated a convincing function of CsATG8e in N remobilization and plant development, indicating CsATG8e as a potential gene for modifying plant nutrient utilization.

Although tea catechins in green tea and green tea beverages must be stable to deliver good sensory quality and healthy benefits, they are always unstable during processing and storage. Ascorbic acid (AA) is often used to protect catechins in green tea beverages, and AA is easily oxidized to form dehydroascorbic acid (DHAA). However, the function of DHAA on the stability of catechins is not clear. The objective of this study was to determine the effects of DHAA on the stability of catechins and clarify the mechanism of effects by conducting a series of experiments that incubate DHAA with epigallocatechin gallate (EGCG) or catechins. Results showed that DHAA had a dual function on EGCG stability, protecting its stability by inhibiting hydrolysis and promoting EGCG consumption by forming ascorbyl adducts. DHAA also reacted with (&minus, )-epicatechin (EC), (&minus, )-epicatechin gallate (ECG), and (&minus, )-epigallocatechin (EGC) to form ascorbyl adducts, which destabilized them. After 9 h of reaction with DHAA, the depletion rates of EGCG, ECG, EC, and EGC were 30.08%, 22.78%, 21.45%, and 13.55%, respectively. The ability of DHAA to promote catechins depletion went from high to low: EGCG, ECG, EGC, and EC. The results are important for the processing and storage of tea and tea beverages, as well as the general exploration of synergistic functions of AA and catechins.

This study focused on non-targeted metabolomics analysis of the relationship of the chemical composition, quality traits, bioactivity of black tea (Camellia sinensis L.) with five withering degrees (WD65, WD60, WD55, WD50, and WD45, corresponding to 65, 60, 55, 50, and 45% water content in withered leaves, respectively) and their effects on sensory quality traits. WD60 was shown as the optimal withering degree (WD) with the highest total quality score (89.7), followed by WD55 (88.7). With increasing WD, the content of most volatile components increased significantly, but with no significant difference between some substances below WD55. The total amount of free amino acids increased with increasing WD, in contrast to a decrease in the total amount of catechins, flavonoids/flavonoid glycosides, theaflavins and glycosidically bound volatiles (GBVs). Black tea with a WD between WD65 and WD60 was shown to have higher antioxidant activity and stronger inhibitory effect on α-amylase and α-glucosidase activity or intestinal glucose uptake. The overall data indicate that WD60 treatment not only benefits the formation of sensory quality traits, but also shows better bioactivity.

A stable substrate is extraordinarily important for the trace detection of Surface Enhanced Raman Scattering technique. We have designed in this paper a simple seed growth method to prepare Ag@ZrO2 core-shell nanoparticles as well as regulate the core-shell ratio by adjusted concentration of zirconium propoxides added. The shell thickness effect and spacing on SERS activities have been simulated in different finite time domains, which goes perfectly with those previous experiment results. With Ag@ZrO2 core-shell nanoparticles adopted as the SERS substrate and R6G as molecular probe, the detection range could reach as great as 10-8 M. The regression model, obtained through principal component analysis, is adopted for some rapid and precise detection; and the determination coefficient (R2) is going up to 0.9743, proving that the SERS substrate we have prepared has extremely high detection accuracy. To explore the stability of core-shell nanoparticles, we have taken both two different cases of strong acid solution as well as strong alkali solution for the core-shell nanoparticle etching, and this is how a SERS test could be carried out. These experimental results have indicated that the outer zirconia layer could keep the silver surface from oxidation and its stability is ensured, quite crucial for the applied SERS.

A surface enhanced Raman scattering (SERS) method is described for the determination of vanillin, methyl vanillin and ethyl vanillin at trace levels. Flower-like silver nanoparticles on a silicon wafer are used as the SERS substrate, and the analytes can be specifically and non-destructively recognized by their specific Raman bands. The molecules can be recognized rapidly by identifying the characteristic bands. The SERS spectra of vanillin (C8H8O3) were used as mid-contrast, and specific bands of methyl vanillin and ethyl vanillin (C9H10O3) were acquired at 775 cm−1, 1350 cm−1 and 1282 cm−1, 1382 cm−1, respectively. In addition, by using an improved principal component analysis (PCA) algorithm, the organic molecule can be quantitatively determined. Dissolved in water, vanillin, methyl vanillin and ethyl vanillin still can be detected at a concentration of 10−8 M, at which their characteristic Raman peaks are still visible. The method was successfully applied to the determination of vanillin in milk powder products.

Colletotrichum, the causative agent of anthracnose, is an important pathogen that invades the tea plant (Camellia sinensis). In this study, 38 isolates were obtained from the diseased leaves of tea plants collected in different areas of Zhejiang Province, China. A combination of multigene (ITS, ACT, GAPDH, TUB2, CAL, and GS) and morphology analyses showed that the 38 strains belonged to two different species, namely, C. camelliae (CC), and C. fructicola (CF). Pathogenicity tests revealed that CC was more invasive than CF. In vitro inoculation experiments demonstrated that CC formed acervuli at 72 hpi and developed appressoria on wound edges, but CF did not develop these structures. Under treatment with catechins and caffeine, the growth inhibition rates of CF were remarkably higher than those of CC, indicating that the nonpathogenic species CF was more vulnerable to catechins and caffeine. Growth condition testing indicated that CF grew at a wide temperature range of 15–35°C and that the optimum temperature for CC growth was 25°C. Growth of both CC and CF did not differ between acidic and weakly alkaline environments (pH 5–8), but the growth of CC was significantly reduced at pH values of 9 and 10. Furthermore, the PacC/RIM101 gene, which associated with pathogenicity, was identified from CC and CF genomes, and its expression was suppressed in the hyphae of both species under pH value of 5 and 10, and much lower expression level was detected in CC than that in CF at pH 6. These results indicated that temperature has more important effect than pH for the growth of two Colletotrichum species. In conclusion, the inhibition by secondary metabolite is an important reason why the pathogenicity by CC and CF are different to tea plant, although the environmental factors including pH and temperature effect the growth of two Colletotrichum species.

Withering is an indispensable process for improving flavors in green, black and white teas during their manufacturing. The effects of the withering process on the formation of tea flavors were investigated using transcriptome and metabolite profiling in withered tea leaves. A total of 3268, 23,282 and 25,185 differentially expressed genes (DEGs) were identified at 3 h (68%, water content), 12 h (61%) and 24 h (48%) of the withering process, respectively. The DEGs, involved in flavonoid biosynthesis were significantly downregulated, which could be correlated with the reduction of catechins. Enhancement of terpenoids and alpha-linolenic acid metabolism could trigger an increase in the total content and number of volatiles. The increase in free amino acid-content could be related to 261 DEGs. Our study suggests that dehydration stress during withering induced significant changes in the gene transcription and content of the tea flavor compounds, which promoted the special flavors in various teas.

The formation of and dynamic changes in aroma during white tea processing have not previously been systematically investigated. In this study, advanced comprehensive two-dimensional gas chromatography-time-of-flight mass spectrometry was employed to investigate the mechanism of white tea aroma formation. A total of 172 volatiles were identified and mainly comprising endogenous volatiles, which displayed diverse change trends during the withering period. In this process, free aroma precursor amino acids and glycosidically bound volatiles (GBVs) were found to contribute to the formation of white tea aroma, with the differential expression of aroma-related key genes accounting for various accumulation of endogenous volatiles and GBVs. In addition, the drying was also shown to play an important role in the formation of white tea aroma. Our study provides the first characterization of white tea aroma formation and establishes a theoretical basis for quality control during white tea processing operations.

Selenium (Se) is a dietary essential trace element for humans and animals. Owing to Se being obtained from food or other source of supplementation, Se-enriched tea is a possible source of supplemental Se and thus has been drawing much attention. This study aimed to identify the variation of the Se concentrations among 14 elite Camellia sinensis cultivars against two-Se-levels supplementation at seedling stage. In the present study, the genetic diversity was assessed by 93 alleles revealed by 12 SSR primers in the C. sinensis accessions. Among the scored 93 alleles, 81 (87.10%) were polymorphic, and the average Nei's gene diversity ( He ) was 0.253 and average shannon information index ( I ) was 0.394, indicating a relatively high genetic variation. Significant variations were observed for the Se concentration among the accessions in both treatments ( P −1 and 0.357–0.537 mg kg −1 in control (0) and Se-supplied (4 mg kg −1 ) treatments respectively. The quality components content, tea polyphenols and amino acid, were sensitive to Se-supplement in a few of accessions, whereas soluble sugar and aqueous extract content appeared unchanged between the contrast treatments. In addition, the genetic variation among the tea plant cultivars was not well correlated with the Se concentration. Even then, the genetic variation in Se concentration across the 14 accessions indicated the possibility of utilizing tea plant accessions to develop optimized cultivars for Se-enriched tea. For the relative high Se concentration in the tea, tea plant is supposed to be a Se-enriched species. These results may enhance our understanding the genetic variation in tea plant.

Pesticide residues pose a great threat to human health, and it is an urgent matter to realize fast and accurate detection of pesticide. SERS (Surface Enhanced Raman Scattering), as a nondestructive detection technology, performs a prominent role in fast detection field due to the strong surface plasmon resonance from short range effect between analyte and nanoparticle. Therefore, in order to solve the incompatibility between organochlorine pesticides molecules and noble metal nanoparticles, this paper proposed a concept of "bridge" substances acting as an interconnect function role to achieve a binding model (object-binder-metal (OBM)) and developed a droplet concentration method to enhance Raman signals. Both combination mode of pesticide molecules to bridge molecules and energy transfer of SERS experiment may relate to the compound ring according to the changes of peaks based on surface plasmon resonance. The selectivity and stability of different bridge substances interacting with pesticides molecules were illumined via binding energy of these two substances obtained by DFT calculations. A droplet can capture nanoparticles and analytes, which is conducive to SERS performance. Chloride ions in the solution contribute to rearrangement of nanoparticles and can validly promote surface activation of Ag nanoparticles to improve energy transfer efficiency of plasma resonance, resulting in superior SERS effect.

This study aimed to figure out the optimum fermentation temperature by analyzing the major chemical constituents of black tea and its antioxidant activities, inhibitory activities against α-amylase, α-glucosidase and glucose uptake. Sensory evaluation revealed 28°C-fermented black tea showed best sensory quality with highest scores for aroma (6.5) and taste (29.8). With increasing fermentation temperature, polyphenols, polysaccharides, (−)-epigallocatechin, (+)-catechin, theaflavins and thearubigins decreased significantly, while amino acids, soluble sugars and volatiles increased when temperature increased up to 31 °C. Furthermore, black tea fermented at 28 °C showed strongest DPPH radical scavenging ability, anti-lipid peroxidation and inhibitory effect on α-glucosidase activity with IC50 value of 17.72 μg/mL, 1.49 mg/mL and 21.69 μg/mL respectively. While 25°C-fermented black tea showed highest inhibition rate of glucose uptake in Caco-2 cells. The results demonstrate that appropriately lowering fermentation temperature could improve both sensory quality and bioactivity of black tea.

Background Self-incompatibility (SI) is under genetic control and prevents inbreeding depression in angiosperms. SI mechanisms are quite complicated and still poorly understood in many plants. Tea (Camellia sinensis L.) belonging to the family of Theaceae, exhibits high levels of SI and high heterozygosity. Uncovering the molecular basis of SI of the tea plant may enhance breeding and simplify genomics research for the whole family. Results The growth of pollen tubes following selfing and crossing was observed using fluorescence microscopy. Self-pollen tubes grew slower than cross treatments from 24 h to 72 h after pollination. RNA-seq was employed to explore the molecular mechanisms of SI and to identify SI-related genes in C. sinensis. Self and cross-pollinated styles were collected at 24 h, 48 h and 72 h after pollination. Six RNA-seq libraries (SP24, SP48, SP72, CP24 CP48 and CP72; SP = self-pollinated, CP = cross-pollinated) were constructed and separately sequenced. In total, 299.327 million raw reads were generated. Following assembly, 63,762 unigenes were identified, and 27,264 (42.76 %) unigenes were annotated in five public databases: NR, KOG, KEGG, Swiss-Port and GO. To identify SI-related genes, the fragments per kb per million mapped reads (FPKM) values of each unigene were evaluated. Comparisons of CP24 vs. SP24, CP48 vs. SP48 and CP72 vs. SP72 revealed differential expression of 3,182, 3,575 and 3,709 genes, respectively. Consequently, several ubiquitin-mediated proteolysis, Ca2+ signaling, apoptosis and defense-associated genes were obtained. The temporal expression pattern of genes following CP and SP was analyzed; 6 peroxidase, 1 polyphenol oxidase and 7 salicylic acid biosynthetic process-related genes were identified. The RNA-seq data were validated by qRT-PCR of 15 unigenes. Finally, a unigene (CL25983Contig1) with strong homology to the S-RNase was analyzed. It was mainly expressed in styles, with dramatically higher expression in self-pollinated versus cross-pollinated tissues at 24 h post-pollination. Conclusions The present study reports the transcriptome of styles after cross- and self-pollination in tea and offers novel insights into the molecular mechanism behind SI in C. sinensis. We believe that this RNA-seq dataset will be useful for improvement in C. sinensis as well as other plants in the Theaceae family. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-2703-5) contains supplementary material, which is available to authorized users.