Journal of Tea Science

Cloning and Response Analysis of the CsMDHAR Gene Under the Abiotic Stress in Camellia sinensis

LIN Shijia, LI Hui, LIU Hao, TENG Ruimin, LIU Jingyu, WANG Shuang, ZHUANG Jing

Journal of Tea Science. 2019, 39(5): 495-505. doi:10.13305/j.cnki.jts.2019.05.001

Abstract ( 639 )

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In this study, a MDHAR gene (CsMDHAR) was cloned from Camellia sinensis cv. ‘Longjing 43’ based on the transcriptome data of tea plant. Sequence analysis shows that the open reading frame length of CsMDHAR was 1β305βbp, encoding 434 amino acids with a molecular weight of approximately 47.21βkDa and the theoretical isoelectric point of 5.99. CsMDHAR was a hydrophilic protein, including two unordered regions and 32 phosphorylation sites. CsMDHAR belonged to Pyr-redox-2 super-family containing a highly conserved region called FAD domain, and mainly composed of α-helix and random coil. PlantCARE and PLACE database prediction analysis suggest that there were many cis-elements related to light, hormones and stress resistance in the 1β000βbp upstream region of CsMDHAR gene. The expression profiles of CsMDHAR, CsAO and CsAPX in tea cv ‘Longjing 43’ and ‘Yingshuang’ under high temperature, low temperature, drought, and salt treatments were detected by qRT-PCR. The results indicate that the expression profiles of CsMDHAR, CsAO and CsAPX were suppressed under 4℃, and there were no significantly differences in ‘Longjing 43’ and ‘Yingshuang’. However, the expression profiles of CsMDHAR gene were upregulated under 38℃ or 200βg·L^-1 PEG treatments in ‘Longjing 43’, with the highest 2.5 and 5 times of the control at 8βh and 2βh, respectively. In addition, the expression trends of CsAO and CsAPX in both cultivars were similar under NaCl (200βmmol·L^-1) treatment, but the variation ranges were different, which might be related to the different stress response in tea plant.

Transcriptome Analysis of Root Induced by Aluminum in Tea Plants (Camellia sinensis)

HUANG Danjuan, TAN Rongrong, CHEN Xun, WANG Hongjuan, GONG Ziming, WANG Youping, MAO Yingxin

Journal of Tea Science. 2019, 39(5): 506-520. doi:10.13305/j.cnki.jts.2019.05.002

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The aim of this study was to investigate the gene regulation network and expression pattern of the response to aluminum (Al) in tea plants, and to identify the key candidate genes for understanding molecular mechanism of Al tolerance in tea plants. The roots’ antioxidant enzyme activities and Al content of Fuding Dabaicha cultivar were detected under 0βmmol·L^-1, 0.2βmmol·L^-1, 1βmmol·L^-1, 2βmmol·L^-1 and 4βmmol·L^-1 Al³⁺ concentrations for 7βd. The total RNA of roots under 0βmmol·L^-1 (R0), 1βmmol·L^-1 (R1) and 4βmmol·L^-1 (R4) Al³⁺ concentrations were extracted for high-through transcriptome sequencing by Illumina Hiseq Xten platform. The results showed that with the increase of Al concentration, POD activity decreased while APX activity increased gradually. SOD activity reached the highest peak at the Al concentration of 1βmmol·L^-1. However, CAT activity showed no significant difference among five treatments. The Al content first increased and then decreased with the increase of Al³⁺ concentration, and reached the highest peak at the Al³⁺ concentration of 1βmmol·L^-1.The DEGs of R1 VS R0, R4 VS R0, R4 VS R1 were 1β894, 2β439 and 1β384 respectively with 733 (1β161), 846 (1β593) and 628 (756) DEGs significantly up-regulated (down-regulated). GO enrichment analysis shows that the most enrichment biological pathway of three samples were all stimulus responses. In terms of molecular function and cell components, R1 VS R0 and R4 VS R0 were mostly enriched in nucleic acid binding transcription factor activity and cell periphery, while R4 vs R1 were mostly enriched in redox enzyme activity and membrane. KEGG enrichment analysis illustrates that they were significantly enriched in 29, 41, and 19 pathways, respectively, including transcription factors, transporters, plant-pathogen interactions, and phenylpropanoid biosynthesis pathways. It was found that genes involved in physiological processes such as reactive oxygen metabolism, organic acids or metal transporters, transcription factors and cell wall structure modification were up-regulated or inhibited after Al induction, suggesting that these genes were closely related to the molecular mechanism of Al tolerance in tea plants.

Identification and Bioinformatic Analysis of Pectin Acetylesterases from Tea Plant

LIU Yanli, MA Linlong, CAO Dan, JIN Xiaofang, FENG Lin, GONG Ziming

Journal of Tea Science. 2019, 39(5): 521-529. doi:10.13305/j.cnki.jts.2019.05.003

Abstract ( 1107 )

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Tea plant is a fluoride hyperaccumulator with high fluoride in leaves, which has many health benefits to human body. Tea pectin acetylesterase (PAE) may be related to fluoride accumulation and detoxification by regulating pectin deacytelation. However, few reports on tea PAEs were available. In the study, the identification and analysis of characteristic, evolutionary and subcellular localization of CsPAEs were performed by bioinformatics method based on tea genome database and three next-generation sequencing data of Camellia sinensis var. sinensis cv ‘Shuchazao’. The results indicate that 12 CsPAEs belonged to five members, including PAE5, PAE8, PAE9, PAE10 and PAE12. Nine conserved motifs including CLDG, PxYH, GGGWC, GS, NWN, rYCDg, GCSAG, NaAYDSWQ and HCQ were found in CsPAEs; CsPAEs were more closely related to PAEs of Vitis vinifera and Theobroma cacao according to evolutionary relationship; Amino acid length, molecular weight and theoretical isoelectric point of 12 CsPAEs varies from 326-515, 36.7-56.9βkDa and 5.1-8.9, respectively. Except that CsPAE5 and CsPAE5-1 were predicated to be localized in the mitochondrion, and CsPAE10-1 might be present in cellular martix. The rest CsPAEs were predicated to be localized in the cell wall. CsPAE5, CsPAE5-1, CsPAE12 and CsPAE12-1 were transmembrane proteins. The results provided a foundation for functional analysis of CsPAEs involved in fluoride accumulation and detoxification.

Study on Physiological Characteristics of Leaves with Different Colors of ‘Huangjinya’

FAN Yangen, ZHAO Xiuxiu, WANG Hanyue, TIAN Yueyue, XIANG Qinzeng, ZHANG Lixia

Journal of Tea Science. 2019, 39(5): 530-536. doi:10.13305/j.cnki.jts.2019.05.004

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In order to screen suitable materials for studying the molecular mechanism of ‘Huangjinya’, the spring shoots of ‘Huangjinya’ were treated with double shading (average light intensity of 10βklx), single shading (average light intensity of 40βklx) and non-shading. And three new shoots, light green (H_1W), light yellow (H_4W) and bright yellow (Hs), were obtained. At the same time, the second leaf under bud was selected as material with ‘Fudingdabai’ green shoot (CK_f) as control. The content of hydrogen peroxide, oxygen-and histochemical localization, antioxidant enzyme activity, photosynthetic response curve, chlorophyll fluorescence and other physiological indicators of leaves were measured and the structure of chloroplast membrane system was observed. The results show that The H₂O₂, O₂^-contents, Fv/Fm value and chloroplast membrane system of light yellow leaf H_4Wwere similar to those of light green leaf H_1W, which were in a stress-free state. At the same time, the photosynthetic and physiological characteristics of light yellow leaf H_4W and bright yellow leaf Hs were similar. There were no significant differences in chlorophyll fluorescence parameters and photosynthetic indexes such as ΦPSⅡ, qP, light saturation point and light compensation point, and the characteristics of light response curve were similar. Because of the above physiological characteristics of light yellow leaf H_4W, it could be used as the research material to explore the molecular mechanism of yellowing of golden bud tea varieties.

Effect of Fluoride and Aluminum Interaction on the Chlorophyll Fluorescence Characteristics of Tea Leaves

ZHONG Qiusheng, LIN Zhenghe, HAO Zhilong, CHEN Changsong, CHEN Zhihui, YOU Xiaomei, SHAN Ruiyang

Journal of Tea Science. 2019, 39(5): 537-546. doi:10.13305/j.cnki.jts.2019.05.005

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To study the effect of fluoride and aluminum intertreatment on the chlorophyll fluorescence characteristics of tea leaves, different concentrations of fluorine and aluminum were used and the fluorescence parameters were analyzed by modulated chlorophyll fluorescent instrument. The results found that: the actual photosynthetic efficiency Y(Ⅱ), the maximum (F_m), primary maximum photochemical efficiency of PSⅡ such as F_v/F_m and coefficient of photochemical quenching such as qP and qL of tea seedlings decreased with the increase of fluorine concentration without aluminum. But the non-photochemical quenching coefficients (NPQ), F_o, non-photochemical quenching coefficient (NPQ and qN) and PSII regulated energy dissipation increased. At low aluminium levels, F_m, F_v/F_m, Y(Ⅱ), NPQ and qN decreased with the increase of fluorine concentration, while qP and qL increased. There were no significant differences between the F_o, Y(NO) and Y(NPQ) values at this time. Under high aluminium treatment, with the increase of fluorine treatment concentration, the F_o gradually increased, Y(Ⅱ), F_v/F_m, qP decreased, Y(NPQ) and NPQ increased significantly under low fluorine concentration (100βmg·L^-1). In short, for tea trees under fluorine stress, aluminum could reduce the damage to tea plants, especially under low aluminum level. The improving effects of photosynthetic efficiency were more obvious.

Optimization of Performance Parameters of Black Tea Withering Machine Based on CFD and RSM

AN Ting, JIANG Yongwen, LIANG Gaozhen, HU Bin, DONG Chunwang

Journal of Tea Science. 2019, 39(5): 547-554. doi:10.13305/j.cnki.jts.2019.05.006

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In order to ascertain the influence of the structural parameters of the withering machine on withering performance, the numerical simulation analysis of withering environment was carried out by the fluid mechanics software. The temperature-integrated index after the dimensionless was used as the evaluation index of the withering performance and the response surface methodology (RSM) was used to optimize the three factors (the withering layer height x₁, the withering machine and the withering room space x₂, and the gas position x₃) that affect withering quality. The result shows that the order of importance of each factor on the withering quality was as follows: gas path position, withering machine layer height, space distance. The best set for temperature field performance of the black tea withering machine were x₁=10βcm, x₂=10βcm, x₃ =149.6βcm. The model’s R²=0.968β3, adjR²=0.927β5, the standard deviation was 0.061. The theoretical and actual values of the optimal scheme were: 0.951β9 and 0.909β6, respectively. The CFD and RSM fusion analysis method was suitable for optimizing the performance parameters of the black tea withering machine.

Effects of Frost-free Refrigerator and Direct-cooled Ice Bar on Storage Quality of Longjing Green Tea

WANG Jinjin, YUAN Haibo, TAO Ruitao, HUA Jinjie, DENG Yuliang, DONG Chunwang, JIANG Yongwen, WANG Jiyun

Journal of Tea Science. 2019, 39(5): 555-566. doi:10.13305/j.cnki.jts.2019.05.007

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The effects of frost-free refrigerator and direct-cooled ice bar on the storage quality of longjing green tea were explored. Teas were stored in the cold storage room and freezer room of the frost-free refrigerator and direct-cooled ice bar respectively, and the storage effects of the different storage conditions on green tea were compared by measuring the contents of biochemical components and sensory quality evaluation when the teas were stored for 90, 180, 270 and 360 days. The results show that the storage effect on biochemical components in freezer room was better than that in cold storage room during the whole storage process and the freezer room of frost-free refrigerator could keep the lower water content, with the tea water content for 180 days significantly lower than that of the direct-cooled ice bar (P<0.05). The freezer room of direct-cooled ice bar was conducive to the maintenances of chlorophyll, vitamin C, caffeine, and the contents of vitamin C and caffeine when the tea was stored for 180 days. During the storage period of 180 days, the retention rates of amino acids, tea polyphenols, catechins were higher in the freezer room of the frost-free refrigerator (they were all around 95%). But the retention rates were higher in the freezing chamber of straight colder bar during the later stage of storage, and were significantly higher than those of the frost-free refrigerator (P<0.05). Frost-free refrigerator was conducive to maintaining the taste and aroma of green tea. While direct-cooled ice bar was conducive to maintaining the appearance of green tea, suggesting the effects of storage conditions on the organoleptic properties of tea are different. Orthogonal partial least squares discriminant analysis (PLS-DA) showed that the quality indexes of liquor color, taste, aroma, caffeine and vitamin C were significantly different under different storage conditions. Combined with the characteristics of two devices, it can be seen that frost-free refrigerator is more suitable for short-term storage of tea, while direct-cooled ice bar is better for the long-term storage of tea.

Dietary Intake Risk Assessment of Pesticide Residues in Tea in Qiandongnan Prefecture

WANG Wei, YAN Renyu, LAN Jiyu, HE Jie, PAN Chengdan, HU Yi

Journal of Tea Science. 2019, 39(5): 567-575. doi:10.13305/j.cnki.jts.2019.05.008

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In order to understand the quality and safety of tea in Qiandongnan, the pesticide residues in tea were determined, and the risk factors of pesticide residues detected in tea were analyzed according to dietary exposure assessment and FSA (Food Standards Agency) risk ranking system. The results show that the detection rate of pesticide residues in tea was 54.05%, with 22 kinds of pesticides detected. The detection rate of single pesticide ranged from 0.90% to 31.53%. According to the maximum residue limit of pesticides in foods (GB 2763—2016), the qualified rate is 97.30%. According to the EU standard limit, the qualified rate is 87.39%. The chronic intake risk (% ADI) of the detected pesticides and acute intake risk (%ARfD) were 0.0037%-1.1780% and 0.03%-41.42%, which are acceptable. Based on the pesticide risk ranking, the pesticides at low, medium and high risk levels are 45.45%, 31.82% and 22.73%, with the highest risk of Carbofuran. Therefore, the risk of pesticide residues in this study was within an acceptable range, indicating their quality and safety were good.

Determination of 79 Pesticide Residues in Tea by Solid Phase Extraction with GC-MS/MS and LC-MS/MS

ZENG Yan, LANG Hong, YANG Qiaohui, LI Xiaxue, SHI Chaoting

Journal of Tea Science. 2019, 39(5): 576-586. doi:10.13305/j.cnki.jts.2019.05.009

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In order to detect various pesticide residues in tea, a reliable, rapid and high throughput detection method was established: a solid phase extraction was developed for the determination of 79 pesticide residues in tea by gas chromatography-tandem mass spectrometry (GC-MS/MS) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). These samples were extracted by 20βmL acetonitrile. 10βmL extracts were cleaned by Carbon/NH₂ column with 20βmL acetonitrile + toluene (3∶1) and then mixed with 5βmL acetonitrile, then these procdssed samples were detected by GC-MS/MS and LC-MS/MS, respectively. The results show that the concentrations of 79 pesticide residues had good linearity relationships in the range of 0.01-0.40βmg·L^-1, with the correlation coefficients (R²) above 0.995, the recoveries of high, low and medium levels were ranged from 67.3%-130.8%, the relative standard deviations (RSDs) were less than 15%, the limits of quantification (LOQs) of 98.7 % pesticides were less than or equal to 0.01βmg·kg^-1, multiple types of tea were detected, and the results were proved to be qualified. Consequently, the method can simultaneously detect 5 kinds of pesticides, including organophosphorus, organochlorine, pyrethroids, carbamates and organic heterocyclic pesticides, which would be useful to determine multi-pesticide residues in high numbers of tea samples.

Weeds Control Effect and Residues of Several Herbicides in Tea Gardens

GAO Wanjun, ZHANG Yongzhi, TONG Mengmeng, MA Huiqin, QIAN Shanshan, WANG Tianyu, LI Yeyun, WU Huiping, HOU Ruyan

Journal of Tea Science. 2019, 39(5): 587-594. doi:10.13305/j.cnki.jts.2019.05.010

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The weeds control effect of several herbicides (glyphosate, glufosinate and combination of glyphosate-acetochlor and glyphosate-auxiliaries) and the herbicides residues in the soil of treated plots in the tea gardens were compared. The results show that the glufosinate had a faster control effect on the weeds. The coverage of weeds decreased to 9.15% on the 7^th treatment day. The control efficiency of glufosinate to the Geranium carolinianum L., Sonchus asper (L.) Hill., Conyza canadensis (L.) Cronq．reached more than 95% on the 14^th day. Glufosinate showed a faster degradation (the dissipation ratio was 94% on the 30^th day) in the soil than the glyphosate (the residue concentration was 0.01βmg·kg^-1 on the 58^th treatment day). Therefore, glufosinate might be an alternative herbicide for weed control in tea gardens. Acetochlor, as a closed herbicide, had the potential of inhibiting the emergence of gramineous weeds and faster dissipation rate in the soil (the dissipation ratio was 97% on the 21^th day), and therefore, it was recommended to control gramineous or the other weeds.

Occurrence and Damage of Overwintering Populations of Ricania sublimbata Jacobi in Xinyang Tea-producing Area

JIN Yinli, MA Quanchao, ZHANG Fangmei, QIAO Li, GENG Shubao, YIN Jian

Journal of Tea Science. 2019, 39(5): 595-601. doi:10.13305/j.cnki.jts.2019.05.011

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In the present study, not only the scarring damage caused by overwintering eggs of Ricania sublimbata Jacobi in tea gardens with intensive or extensive management, but also the scale of tea branch chosen by R. sublimbata for oviposition, the length of scar and number of overwintering eggs in each scar were investigated. The results indicate that the damage rates in tea gardens with extensive management (9.86%, 13.79%) were significantly higher than those in intensively managed tea gardens. Adult R. sublimbata preferred to lay overwintering eggs in tea branches, with scale diameter of about 0.20-0.30βcm. Moreover, the average distance from the top of oviposition scar to tip of the tea shoot was 7.22βcm. The mean length of oviposition scar was 1.35βcm. The results also indicate that there was a significantly positive linear relationship between the length of oviposition scar (x) and number of overwintering eggs in the scar (y) and the relationship could be expressed as: y=17.83x+1.12 (F=3β652, df=1β145, R²=0.96, P<2.2×10^-16). It was suggested to cut off the tea branches with oviposition scar from early October to next March, and improve management of tea garden so as to control the regional damage of this pest. This study provided a theoretical basis for the prediction and ecological control of overwintering population of R. sublimbata in Xinyang tea-producing area, which was of great significance for reducing the use of pesticides and improving yield and quality of Xinyang Maojian tea.

An Empirical Analysis of the Trade and Influencing Factors of China's Black Tea Export to Countries along the Belt and Road

XU Yongmei, SHI Yunfeng

Journal of Tea Science. 2019, 39(5): 602-610. doi:10.13305/j.cnki.jts.2019.05.012

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China is the origin of tea and the first black tea export country in the world. Tea, particularly black tea was one of the most important trade commodities on the Chinese ancient Silk Road. Here, the trade profile of China's black tea export to countries along the belt was first described and the main factors affecting the black tea export was analyzed. The expanded trade gravity model was applied to analyze the influencing factors of China's black tea export to countries along the belt. Empirical analysis shows that the average export price of Chinese black tea, the growth of the distance between China and the importing countries of black tea, and the exchange rate of RMB were significantly but negatively correlated with Chinese black tea export. The population growth of the importing countries and the GDP growth of China and the importing countries had a significant and positive impact on China's black tea export.

Analysis of Fatty Acid Compositions and Contents in Oolong Tea from Fujian Province

GUO Li, LYU Haipeng, CHEN Mingjie, ZHANG Yue, BA Yichen, GUO Yaling, LIN Zhi

Journal of Tea Science. 2019, 39(5): 611-618. doi:10.13305/j.cnki.jts.2019.05.013

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Fatty acids (FAs) are the key precursors for aroma formation of Oolong tea. Eighteen Minbei Oolong tea cultivars (raw and re-baked tea), and three Minnan Oolong tea cultivars with faint scent were selected as the experiment materials. The fatty acid methyl esters were prepared by the method of sulfuric acid-methanol and analyzed by gas chromatography-mass spectrometry. The results show that 11 fatty acids were detected in the Oolong teas from Fujian province, and 16C- and 18C- fatty acids were the most abundant. The total fatty acid contents and unsaturation degree from Minbei Oolong teas were lower than those of Minnan Oolong teas. The contents of nine FAs including C16:0, C18:2, C18:3, and so on, showed significant variation among different cultivars, and Queshe showed the highest C18:3 and polyunsaturated fatty acids (PUSFAs). The effect of re-baking on fatty acids in Minbei Oolong tea was related with their number of carbon atoms and unsaturation. Docosanoic acid and tetracosanoic acid degraded completely, and C18:3 content decreased more than others during re-baking process. Thus, FA contents and compositions were affected by locations, cultivars and processing technology.

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