High-speed identification system for fresh tea leaves based on phenotypic characteristics utilizing an improved genetic algorithm.

BACKGROUND: High-quality tea requires leaves of similar size and tenderness. The grade of the fresh leaves determines the quality of the tea. The automated classification of fresh tea leaves improves resource utilization and reduces manual picking costs. The present study proposes a method based on an improved genetic algorithm for identifying fresh tea leaves in high-speed parabolic motion using the phenotypic characteristics of the leaves. During parabolic flight, light is transmitted through the tea leaves, and six types of fresh tea leaves can be quickly identified by a camera. RESULTS: The influence of combinations of morphology, color, and custom corner-point morphological features on the classification results were investigated, and the necessary dimensionality of the model was tested. After feature selection and combination, the classification performance of the Naive Bayes, k-nearest neighbor, and support vector machine algorithms were compared. The recognition time of Naive Bayes was the shortest, whereas the accuracy of support vector machine had the best classification accuracy at approximately 97%. The support vector machine algorithm with only three feature dimensions (equivalent diameter, circularity, and skeleton endpoints) can meet production requirements with an accuracy rate reaching 92.5%. The proposed algorithm was tested by using the Swedish leaf and Flavia data sets, on which it achieved accuracies of 99.57% and 99.44%, respectively, demonstrating the flexibility and efficiency of the recognition scheme detailed in the present study. CONCLUSION: This research provides an efficient tea leaves recognition system that can be applied to production lines to reduce manual picking costs. © 2022 Society of Chemical Industry.

Molecular and Biochemical Characterization of Jasmonic Acid Carboxyl Methyltransferase Involved in Aroma Compound Production of Methyl Jasmonate during Black Tea Processing.

Methyl jasmonate (MeJA), a volatile organic compound, is a principal flowery aromatic compound in tea. During the processing of black tea, MeJA is produced by jasmonic acid carboxyl methyltransferase (JMT) of the jasmonic acid (JA) substrate, forming a specific floral fragrance. CsJMT was cloned from tea leaves; the three-dimensional structure of CsJMT was predicted. Enzyme activity was identified, and protein purification was investigated. Site-directed deletions revealed that N-10, S-22, and Q-25 residues in the beginning amino acids played a key functional role in enzyme activity. The expression patterns of CsJMT in tea organs differed; the highest expression of CsJMT was observed in the fermentation process of black tea. These results aid in further understanding the synthesis of MeJA during black tea processing, which is crucial for improving black tea quality using specific fragrances and could be applied to the aromatic compound regulation and tea breeding improvement in further studies.

Metabolite and Transcriptome Profiling on Xanthine Alkaloids-Fed Tea Plant (Camellia sinensis) Shoot Tips and Roots Reveal the Complex Metabolic Network for Caffeine Biosynthesis and Degradation.

While caffeine is one of the most important bioactive metabolites for tea as the most consumed non-alcohol beverage, its biosynthesis and catabolism in tea plants are still not fully understood. Here, we integrated purine alkaloid profiling and transcriptome analysis on shoot tips and roots fed with caffeine, theophylline, or theobromine to gain further understanding of caffeine biosynthesis and degradation. Shoot tips and roots easily took up and accumulated high concentrations of alkaloids, but roots showed much faster caffeine and theophylline degradation rates than shoot tips, which only degraded theophylline significantly but almost did not degrade caffeine. Clearly feedback inhibition on caffeine synthesis or inter-conversion between caffeine, theophylline, and theobromine, and 3-methylxanthine had been observed in alkaloids-fed shoot tips and roots, and these were also evidenced by significant repression of TCS and MXMT genes critical for caffeine biosynthesis. Among these responsively repressed genes, two highly expressed genes TCS-4 and TCS-8 were characterized for their enzyme activity. While we failed to detect TCS-4 activity, TCS-8 displayed N-methyltransferase activities towards multiple substrates, supporting the complex metabolic network in caffeine biosynthesis in tea plants since at least 13 TCS-like N-methyltransferase genes may function redundantly. This study provides new insight into complex metabolic networks of purine alkaloids in tea plants.

Micro-NIR spectrometer for quality assessment of tea: Comparison of local and global models.

Near-infrared (NIR) spectroscopy is an effective tool for analyzing components relevant to tea quality, especially catechins and caffeine. In this study, we predicted catechins and caffeine content in green and black tea, the main consumed tea types worldwide, by using a micro-NIR spectrometer connected to a smartphone. Local models were established separately for green and black tea samples, and these samples were combined to create global models. Different spectral preprocessing methods were combined with linear partial-least squares regression and nonlinear support vector machine regression (SVR) to obtain accurate models. Standard normal variate (SNV)-based SNV-SVR models exhibited accurate predictive performance for both catechins and caffeine. For the prediction of quality components of tea, the global models obtained results comparable to those of the local models. The optimal global models for catechins and caffeine were SNV-SVR and particle swarm optimization (PSO)-simplified SNV-PSO-SVR, which achieved the best predictive performance with correlation coefficients in prediction (Rp) of 0.98 and 0.93, root mean square errors in prediction of 9.83 and 2.71, and residual predictive deviations of 4.44 and 2.60, respectively. Therefore, the proposed low-price, compact, and portable micro-NIR spectrometer connected to smartphones is an effective tool for analyzing tea quality.

Rapid detection of quality index of postharvest fresh tea leaves using hyperspectral imaging.

BACKGROUND: The quality of fresh tea leaves after harvest determines, to some extent, the quality and price of commercial tea. A fast and accurate method to evaluate the quality of fresh tea leaves is required. RESULTS: In this study, the potential of hyperspectral imaging in the range of 328-1115 nm for the rapid prediction of moisture, total nitrogen, crude fiber contents, and quality index value was investigated. Ninety samples of eight tea-leaf varieties and two picking standards were tested. Quantitative partial least squares regression (PLSR) models were established using a full spectrum, whereas multiple linear regression (MLR) models were developed using characteristic wavelengths selected by a successive projections algorithm (SPA) and competitive adaptive reweighted sampling. The results showed that the optimal SPA-MLR models for moisture, total nitrogen, crude fiber contents, and quality index value yielded optimal performance with coefficients of determination for prediction (R2 p) of 0.9357, 0.8543, 0.8188, 0.9168; root mean square error of 0.3437, 0.1097, 0.3795, 1.0358; and residual prediction deviation of 4.00, 2.56, 2.31, and 3.51, respectively. CONCLUSION: The results suggested that the hyperspectral imaging technique coupled with chemometrics was a promising tool for the rapid and nondestructive measurement of tea-leaf quality, and had the potential to develop multispectral imaging systems for future online detection of tea-leaf quality. © 2020 Society of Chemical Industry.

Tea plant genomics: achievements, challenges and perspectives.

Tea is among the world's most widely consumed non-alcoholic beverages and possesses enormous economic, health, and cultural values. It is produced from the cured leaves of tea plants, which are important evergreen crops globally cultivated in over 50 countries. Along with recent innovations and advances in biotechnologies, great progress in tea plant genomics and genetics has been achieved, which has facilitated our understanding of the molecular mechanisms of tea quality and the evolution of the tea plant genome. In this review, we briefly summarize the achievements of the past two decades, which primarily include diverse genome and transcriptome sequencing projects, gene discovery and regulation studies, investigation of the epigenetics and noncoding RNAs, origin and domestication, phylogenetics and germplasm utilization of tea plant as well as newly developed tools/platforms. We also present perspectives and possible challenges for future functional genomic studies that will contribute to the acceleration of breeding programs in tea plants.

Qualitative and quantitative diagnosis of nitrogen nutrition of tea plants under field condition using hyperspectral imaging coupled with chemometrics.

BACKGROUND: Rapid and accurate diagnosis of nitrogen (N) status in field crops is of great significance for site-specific N fertilizer management. This study aimed to evaluate the potential of hyperspectral imaging coupled with chemometrics for the qualitative and quantitative diagnosis of N status in tea plants under field conditions. RESULTS: Hyperspectral data from mature leaves of tea plants with different N application rates were preprocessed by standard normal variate (SNV). Partial least squares discriminative analysis (PLS-DA) and least squares-support vector machines (LS-SVM) were used for the classification of different N status. Furthermore, partial least squares regression (PLSR) was used for the prediction of N content. The results showed that the LS-SVM model yielded better performance with correct classification rates of 82% and 92% in prediction sets for the diagnosis of different N application rates and N status, respectively. The PLSR model for leaf N content (LNC) showed excellent performance, with correlation coefficients of 0.924, root mean square error of 0.209, and residual predictive deviation of 2.686 in the prediction set. In addition, the important wavebands of the PLSR model were interpreted based on regression coefficients. CONCLUSION: Overall, our results suggest that the hyperspectral imaging technique can be an effective and accurate tool for qualitative and quantitative diagnosis of N status in tea plants. © 2019 Society of Chemical Industry.

Enantiomeric Trimethylallantoin Monomers, Dimers, and Trimethyltriuret: Evidence for an Alternative Catabolic Pathway of Caffeine in Tea Plant.

Racemic trimethylallantoin monomer (1), mesomeric and racemic trimethylallantoin dimers (2 and 3), were isolated from tea. Two pairs of optically pure enantiomers (1a, 1b and 3a, 3b) were separated by chiral column from the two racemes (1 and 3). Their structures were elucidated by a combination of extensive spectroscopic techniques, single-crystal X-ray diffraction, and experimental and calculated electronic circular dichroism. A novel caffeine catabolic pathway was proposed based on the caffeine stable isotopic tracer experiments.

Fabricating an Acetylcholinesterase Modulated UCNPs-Cu2+ Fluorescence Biosensor for Ultrasensitive Detection of Organophosphorus Pesticides-Diazinon in Food.

In this study, a highly sensitive upconversion fluorescence (FL) biosensor was developed for the detection of organophosphorus pesticides (OPs) based on an acetylcholinesterase (AChE) modulated FL "off-on-off" strategy. The luminescence of synthesized UCNPs could be quenched strongly by Cu2+ due to an energy transfer effect. Upon addition of AChE and acetylthiocholine (ATCh), the enzymatic hydrolysate (thiocholine) could seize Cu2+ from UCNPs-Cu2+ mixture, resulting in the quenched FL triggered on. OPs could irreversibly impede the activity of AChE, which caused the formation of thiocholine to decrease, thus, reduced the recovery of FL. Under the optimum conditions, a linear detection range from 0.1 to 50 ng/mL was achieved for the representative OPs (diazinon) with LOD of 0.05 ng/mL. Furthermore, the ability of the biosensor to detect OPs was also confirmed in adulterated environmental and agricultural samples. In validation analysis, the proposed sensor showed satisfactory results ( p > 0.05) with GC-MS.

Tea Plant Information Archive: a comprehensive genomics and bioinformatics platform for tea plant.

Tea is the world's widely consumed nonalcohol beverage with essential economic and health benefits. Confronted with the increasing large-scale omics-data set particularly the genome sequence released in tea plant, the construction of a comprehensive knowledgebase is urgently needed to facilitate the utilization of these data sets towards molecular breeding. We hereby present the first integrative and specially designed web-accessible database, Tea Plant Information Archive (TPIA; http://tpia.teaplant.org). The current release of TPIA employs the comprehensively annotated tea plant genome as framework and incorporates with abundant well-organized transcriptomes, gene expressions (across species, tissues and stresses), orthologs and characteristic metabolites determining tea quality. It also hosts massive transcription factors, polymorphic simple sequence repeats, single nucleotide polymorphisms, correlations, manually curated functional genes and globally collected germplasm information. A variety of versatile analytic tools (e.g. JBrowse, blast, enrichment analysis, etc.) are established helping users to perform further comparative, evolutionary and functional analysis. We show a case application of TPIA that provides novel and interesting insights into the phytochemical content variation of section Thea of genus Camellia under a well-resolved phylogenetic framework. The constructed knowledgebase of tea plant will serve as a central gateway for global tea community to better understand the tea plant biology that largely benefits the whole tea industry.

Caffeine Content and Related Gene Expression: Novel Insight into Caffeine Metabolism in Camellia Plants Containing Low, Normal, and High Caffeine Concentrations.

Caffeine is a crucial secondary metabolic product in tea plants. Although the presence of caffeine in tea plants has been identified, the molecular mechanisms regulating relevant caffeine metabolism remain unclear. For the elucidation of the caffeine biosynthesis and catabolism in Camellia plants, fresh, germinated leaves from four Camellia plants with low (2), normal (1), and high (1) caffeine concentrations, namely, low-caffeine tea 1 (LCT1, Camellia crassicolumna), low-caffeine tea 2 (LCT2, C. crassicolumna), Shuchazao (SCZ, C. sinensis), and Yunkang 43 (YK43, C. sinensis) were used in this research. Transcriptome and purine alkaloids analyses of these Camellia leaves were performed using RNA-Seq and liquid chromatography-mass spectrometry (LC-MS). Moreover, 15N-caffeine tracing was performed to determine the metabolic fate of caffeine in leaves of these plants. Caffeine content was correlated with related gene expression levels, and a quantitative real-time (qRT) PCR analysis of specific genes showed a consistent tendency with the obtained transcriptomic analysis. On the basis of the results of stable isotope-labeled tracer experiments, we discovered a degradation pathway of caffeine to theobromine. These findings could assist researchers in understanding the caffeine-related mechanisms in Camellia plants containing low, normal, and high caffeine content and be applied to caffeine regulation and breeding improvement in future research.

Using stable isotope signatures to delineate the geographic point-of-origin of Keemun black tea.

BACKGROUND: Confirmation of food labeling that claims production in a small geographic region is critical to traceability, quality control and brand protection. In the current study, isotope ratio mass spectrometry (IRMS) was used to generate profiles of δ13 C and δ15 N to determine if the stable isotope signatures of Keemun black tea differ within the three counties that claim production. Other factors (cultivar type, leaf maturity and manufacturing process) were considered for their potential effects. RESULTS: Both cultivar type and leaf maturity have remarkable impact on the δ15 N values of tea leaves, and that the cultivar influenced the δ13 C values. Keemun black tea from Qimen county could be easily discriminated from samples from Dongzhi and Guichi counties based on δ15 N signatures. The k-NN model was cross-validated with an accuracy of 91.6%. Environmental factors and/or genotype seem to be the major reasons for δ15 N differences in Keemun black tea from the selected regions. CONCLUSION: This article provides a potential effective method to delineate the geographic point-of-origin of Keemun black tea based on δ15 N signatures. © 2018 Society of Chemical Industry.

Significantly increased amino acid accumulation in a novel albino branch of the tea plant (Camellia sinensis).

MAIN CONCLUSION: A normal tea plant with one albino branch was discovered. RNA sequencing, albinism phenotype and ultrastructural observations provided a valuable understanding of the albino mechanism in tea plants. Tea plants with a specific color (white or yellow) have been studied extensively. A normal tea plant (Camellia sinensis cv. quntizhong) with one albino branch was discovered in a local tea plantation in Huangshan, Anhui, China. The pure albino leaves on this special branch had accumulated a fairly high content of amino acids, especially theanine (45.31 mg/g DW), and had a low concentration of polyphenols and an extremely low chlorophyll (Chl) content compared with control leaves. Ultrastructural observation of an albino leaf revealed no chloroplasts, whereas it was viable in the control leaf. RNA sequencing and differentially expressed gene (DEG) analysis were performed on the albino leaves and on control leaves from a normal green branch. The related genes involved in theanine and polyphenol biosynthesis were also investigated in this study. DEG expression patterns in Chl biosynthesis or degradation, carotenoid biosynthesis or degradation, chloroplast development, and biosynthesis were influenced in the albino leaves. Chloroplast deletion in albino leaves had probably destroyed the balance of carbon and nitrogen metabolism, leading to a high accumulation of free amino acids and a low concentration of polyphenols in the albino leaves. The obtained results can provide insight into the mechanism underlying this special albino branch phenotype, and are a valuable contribution toward understanding the albino mechanism in tea plants.

Guanine deaminase provides evidence of the increased caffeine content during the piling process of pu'erh tea.

Wet piling is a key process for producing pu'erh tea because various components change under the action of microorganisms. Among these components, caffeine content is increased. Evidence has indicated a salvage pathway for caffeine biosynthesis in microbes, in which xanthine is methylated in the order of N-3 → N-1 → N-7. In addition, guanine can be used to synthesize xanthine through guanine deaminase (EC: 3.5.4.3). In this study, we investigated the variation in caffeine content during piling fermentation with supplementary guanine, 15N-labeled guanine and xanthine. We cloned the guanine deaminase gene (GUD1) from Saccharomyces cerevisiae (one dominant strain in piling fermentation). The results revealed that [15N]xanthine could be synthesized from [15N]guanine, and [15N]caffeine was also detected during piling with supplementary [15N]xanthine. Furthermore, ScGUD1 could catalyze the conversion of guanine to xanthine, which is likely to be methylated for caffeine synthesis under microorganism action. The obtained results revealed the mechanism underlying the increased caffeine content during piling of pu'erh tea.

Metabolic Flux Enhancement and Transcriptomic Analysis Displayed the Changes of Catechins Following Long-Term Pruning in Tea Trees ( Camellia sinensis).

The tea tree is a perennial woody plant, and pruning is one of the most crucial cultivation measurements for tea plantation management. To date, the relationship between long-term pruning and metabolic flux enhancement in tea trees has not been studied. In this research, 11-year-old pruned tea trees from four different cultivars were randomly selected for transcriptome analysis and characteristic secondary metabolite analysis together with controls. The findings revealed that epigallocatechin gallate (EGCG) accumulation in pruned tea trees was significantly higher than that in unpruned tea trees. SCPL1A expression (encoding a class of serine carboxypeptidase), which has been reported to have a catalytic ability during EGCG biosynthesis, together with LAR, encoding leucoanthocyanidin reductase, was upregulated in the pruned tea trees. Moreover, metabolic flux enhancement and transcriptome analysis revealed low EGCG accumulation in the leaves of unpruned tea trees. Because of the bitter and astringent taste of EGCG, these results provide a certain understanding to the lower bitterness and astringency in teas from "ancient tea trees", growing in the wild with no trimming, than teas produced from pruned plantation trees.

Corrigendum: (-)-Epigallocatechin-3-Gallate Ameliorates Atherosclerosis and Modulates Hepatic Lipid Metabolic Gene Expression in Apolipoprotein E Knockout Mice: Involvement of TTC39B.

[This corrects the article on p. 195 in vol. 9, PMID: 29593532.].

Enantioselective Bromocyclization of Tryptamines Induced by Chiral Co(III)-Complex-Templated Brønsted Acids under an Air Atmosphere.

The chiral Co(III)-complex-templated Brønsted acids were found to be efficient bifunctional phase-transfer catalysts for the highly enantioselective bromocyclization of protected tryptamines with readily available N-bromosuccinimide (NBS) under an air atmosphere. The 3-bromohexahydropyrrolo[2,3- b]indoles, which are key building blocks of cyclotryptamine alkaloids, were thus obtained in up to 95% yield and 93.5:6.5 er.

(-)-Epigallocatechin-3-Gallate Ameliorates Atherosclerosis and Modulates Hepatic Lipid Metabolic Gene Expression in Apolipoprotein E Knockout Mice: Involvement of TTC39B.

Background: Aberrant chronic inflammation and excess accumulation of lipids play a pivotal role in the occurrence and progression of atherosclerosis. (-)-Epigallocatechin-3-gallate (EGCG), the major catechins in green tea, displayed anti-atherosclerotic properties in vivo and in vitro. However, the effects and underlying mechanism of EGCG on atherosclerosis remain unclear. Methods: Male apolipoprotein E-knockout (ApoE-/-) mice (7 weeks old) fed with high-fat diet (HFD) were treated with normal saline or EGCG (40 mg/kg/d, i.g.) for 18 weeks. Atherosclerotic plaque and liver lipid accumulation were measured by Oil Red staining. Plasma lipids and cytokines were detected using commercial kits. The expression of protein and mRNA was analyzed by western blot and quantitative real-time reverse transcription-polymerase chain reaction, respectively. Results: EGCG administration markedly attenuated atherosclerotic plaque formation in HFD-fed ApoE-/- mice, which were accompanied by increased plasma interleukin-10 (IL-10) level and decreased plasma IL-6 and tumor necrosis factor-α (TNF-α) levels. In addition, EGCG modulated high-fat-induced dyslipidemia, evidencing by decreased total cholesterol (TC) and low-density lipoprotein levels and increased high-density lipoprotein level. Meanwhile, EGCG treatment alleviated high-fat-mediated liver lipid accumulation and decreased liver TC and triglyceride. Mechanistically, EGCG significantly modulated high-fat-induced hepatic tetratricopeptide repeat domain protein 39B (TTC39B) expression and its related genes (Lxrß, Abcg5, Abcg8, Abca1, Srebf1, Scd1, Scd2, Fas, Elovl5, Mylip) expression in liver from ApoE-/- mice. Notably, EGCG remarkably induced hepatic liver X receptor α (LXRα) and LXRß expression and inhibited both precursor and mature sterol regulatory element binding transcription factor-1 (SREBP-1) expression. Conclusion: Taken together, our data for the first time suggested that TTC39B was involved in EGCG-mediated anti-atherosclerotic effects through modulation of LXR/SREBP-1 pathway.

Functional Characterization of Salicylic Acid Carboxyl Methyltransferase from Camellia sinensis, Providing the Aroma Compound of Methyl Salicylate during the Withering Process of White Tea.

Methyl salicylate (MeSA) is one of the volatile organic compounds (VOCs) that releases floral scent and plays an important role in the sweet flowery aroma of tea. During the withering process for white tea producing, MeSA was generated by salicylic acid carboxyl methyltransferase (SAMT) with salicylic acid (SA), and the specific floral scent was formed. In this study, we first cloned a CsSAMT from tea leaves (GenBank accession no. MG459470) and used Escherichia coli and Saccharomyces cerevisiae to express the recombinant CsSAMT. The enzyme activity in prokaryotic and eukaryotic expression systems was identified, and the protein purification, substrate specificity, pH, and temperature optima were investigated. It was shown that CsSAMT located in the chloroplast, and the gene expression profiles were quite different in tea organs. The obtained results might give a new understanding for tea aroma formation, optimization, and regulation and have great significance for improving the specific quality of white tea.

Engineering an ABC Transporter for Enhancing Resistance to Caffeine in Saccharomyces cerevisiae.

In addressing caffeine toxicity to the producing cells, engineering a transporter that can move caffeine from cytoplasm across the cell membrane to the extracellular space, thus enhancing caffeine resistance and potentially increasing the yield in yeast, is important. An ABC-transporter bfr1 from Schizosaccharomyces pombe was cloned and transformed into S. cerevisiae, resulting in enhancing caffeine resistance. Afterward, a library of randomly mutagenized bfr1 mutants through error-prone PCR was generated. One mutant was identified with drastically increased caffeine resistance (15 mg/mL). Sequencing and structural analysis illustrated that many of the mutations occurred at the cytosolic domain. Site-directed mutagenesis of these mutations confirmed at least one amino acid that conferred enhancing caffeine resistance in the mutated bfr1. These data demonstrated engineering ABC-transporters can be an efficient way to reduce product toxicity in heterologous systems.