Elevated Ozone Reduces the Quality of Tea Leaves but May Improve the Resistance of Tea Plants.

Tropospheric ozone (O3) pollution can affect plant nutritional quality and secondary metabolites by altering plant biochemistry and physiology, which may lead to unpredictable effects on crop quality and resistance to pests and diseases. Here, we investigated the effects of O3 (ambient air, Am; ambient air +80 ppb of O3, EO3) on the quality compounds and chemical defenses of a widely cultivated tea variety in China (Camellia sinensis cv. 'Baiye 1 Hao') using open-top chamber (OTC). We found that elevated O3 increased the ratio of total polyphenols to free amino acids while decreasing the value of the catechin quality index, indicating a reduction in leaf quality for green tea. Specifically, elevated O3 reduced concentrations of amino acids and caffeine but shows no impact on the concentrations of total polyphenols in tea leaves. Within individual catechins, elevated O3 increased the concentrations of ester catechins but not non-ester catechins, resulting in a slight increase in total catechins. Moreover, elevated O3 increased the emission of biogenic volatile organic compounds involved in plant defense against herbivores and parasites, including green leaf volatiles, aromatics, and terpenes. Additionally, concentrations of main chemical defenses, represented as condensed tannins and lignin, in tea leaves also increased in response to elevated O3. In conclusion, our results suggest that elevated ground-level O3 may reduce the quality of tea leaves but could potentially enhance the resistance of tea plants to biotic stresses.

Increasing global precipitation whiplash due to anthropogenic greenhouse gas emissions.

Precipitation whiplash, including abrupt shifts between wet and dry extremes, can cause large adverse impacts on human and natural systems. Here we quantify observed and projected changes in characteristics of sub-seasonal precipitation whiplash and investigate the role of individual anthropogenic influences on these changes. Results show that the occurrence frequency of global precipitation whiplash is projected to be 2.56 ± 0.16 times higher than in 1979-2019 by the end of the 21st Century, with increasingly rapid and intense transitions between two extremes. The most dramatic increases of whiplash show in the polar and monsoon regions. Changes in precipitation whiplash show a much higher percentage change than precipitation totals. In historical simulations, anthropogenic greenhouse gas (GHG) and aerosol emissions have increased and decreased precipitation whiplash occurrences, respectively. By 2079, anthropogenic GHGs are projected to increase 55 ± 4% of the occurrences risk of precipitation whiplash, which is driven by shifts in circulation patterns conducive to precipitation extremes.

Melatonin-mediated regulation of anthocyanin biosynthesis and antioxidant defense confer tolerance to arsenic stress in Camellia sinensis L.

Arsenic is a toxic metalloid for both animals and plants. The signaling molecule melatonin can enhance abiotic stress tolerance, but the effects of As and melatonin on tea plants and the mechanisms of resilience remain unclear. Here we report that excess As causes severe oxidative stress in tea leaves as revealed by significantly reduced maximal photochemical efficiency of photosystem-II, and increased reactive oxygen species accumulation and lipid peroxidation. However, exogenous melatonin application alleviated the As phytotoxicity and increased the anthocyanin content upto 69.4 % by selectively upregulating the expression of its biosynthetic genes such as CsCHS and CsANS. Comparison of As tolerance between two tea genotypes differing in basal levels of anthocyanin revealed that a tea cultivar with increased anthocyanin content, Zijuan (ZJ), showed enhanced tolerance to As stress compared with Longjing 43 (LJ43) that contained relatively low levels of anthocyanin. Interestingly, exogenous anthocyanin also enhanced As tolerance in LJ43, but exogenous melatonin did not improve As tolerance in ZJ genotype. Analysis of As content in tea leaves revealed that melatonin significantly reduced As content in LJ43 but not in ZJ, suggesting that melatonin-enhanced tolerance to As stress is largely dependent on the basal levels of anthocyanin in tea plants.

Exogenous melatonin improves tea quality under moderate high temperatures by increasing epigallocatechin-3-gallate and theanine biosynthesis in Camellia sinensis L.

Global warming has multifarious effects on crop growth and productivity. Nonetheless, the effects of moderate-high temperatures and melatonin on tea yield and quality remain unclear. In this study, we found that melatonin, a universal growth stimulatory molecule, not only promotes photosynthesis and biomass accumulation in tea plants (Camellia sinensis L.) but also improves tea quality under sub high temperature (SHT). SHT increased the dry biomass and photosynthesis by 40.8% and 28.1%, respectively, and exogenous melatonin caused a further improvement. Moreover, SHT increased the total polyphenol concentrations and decreased the free amino acid concentrations, leading to a significant increase (68.2%) in polyphenol to free amino acid ratio. However, melatonin decreased the polyphenol to free amino acid ratio by delicately improving the concentrations of polyphenols and amino acids. Consistent with the total polyphenol, melatonin increased the concentrations of (-)-catechin, (-)-gallocatechin (GC), and (-)-epigallocatechin-3-gallate (EGCG) in tea leaves. The qRT-PCR analysis revealed that melatonin increased the transcript levels of catechins biosynthesis genes, such as CsCHS, CsCH1, CsF3H, CsDFR, CsANS, CsLAR, and CsANR under SHT. Meanwhile, the theanine concentration was decreased by SHT, which was attributed to the attenuated expression of CsGS, CsGOGAT, CsGDH, and CsTS1. Nonetheless, melatonin significantly increased those transcripts and the content of theanine under SHT. Melatonin also increased the caffeine content by inducing the expression of CsTIDH, CssAMS, and CsTCS1. These results suggest that melatonin could positively alter tea growth and quality by modulating the photosynthesis and biosynthesis of polyphenols, amino acids, and caffeine in tea leaves under SHT.

Evaluation of 2 Rat Models for Sepsis Developed by Improved Cecal Ligation/Puncture or Feces Intraperitoneal-Injection.

BACKGROUND The aim of this study was to evaluate the clinical characteristics of 2 rat models of sepsis for improved cecal ligation/puncture (CLP) and feces intraperitoneal-injection (FIP), including systemic inflammation, organ dysfunction, and blood coagulation. MATERIAL AND METHODS Sixty-two male SD rats were randomly divided into 3 groups: a normal control group (NC, n=6), a CLP group (n=28), and a FIP group (n=28). Ten rats each in the CLP and FIP groups were observed for 72-h mortality rate. The remaining 18 rats in each group were divided into 3 subgroups (n=6) according to their post-operation period (6, 12, and 24 h). Abdominal arterial blood was collected to determine the lactic acid (Lac) concentration, prothrombin time (PT), active partial prothrombin time (APTT), plasmic interleukin-6 (IL-6) level, and cardiac troponin (cTnI) level. The intestines, lung, and heart were collected for pathological examination. RESULTS The 72-h mortality rates in the CLP and FIP groups were 60% and 100%, respectively. The Lac level in both groups was significantly elevated at 6, 12, and 24 h after modeling. Compared with the NC group, PT in the CLP and FIP groups was prolonged at 12 and 24 h, and APTT was significantly prolonged at 6 h. IL-6 levels in the CLP and FIP groups peaked at 6 h. The cTnI level in the FIP group was significantly higher at 12 h after modeling compared with the NC group. The intestines, lung, and heart were pathologically damaged at 6 h, and this damage worsened over time. CONCLUSIONS Both modeling methods induced sepsis in rats and closely mimicked the clinical conditions, but FIP was easier to establish and was more suitable for standardization.

[Analysis of prognostic risk factors of bloodstream infection in intensive care unit patients].

OBJECTIVE: To explore the prognostic risk factors of bloodstream infection (BSI) in intensive care unit (ICU) patients, furthermore, to provide a reliable evidence for early warning and treatment for the critical patients with BSI. METHODS: A retrospective study was performed. The clinical data of patients with blood culture-positive admitted to ICU of the Affiliated Hospital of Guizhou Medical University from January 1 to September 30, 2019 were analyzed. The data including gender, age, proportion of basic diseases, acute physiology and chronic health evaluation II (APACHE II), the duration of mechanical ventilation after being diagnosed with BSI, length of stay in ICU, aggressive operation and blood product infusion after BSI, proportion of using continuous renal replacement therapy (CRRT) and continuous vasoactive agents for more than 72 hours after being diagnosed with BSI, and site of central venous catheterization were recorded. Meantime, the worst laboratory values within 72 hours after being diagnosed with BSI, information about pathogenic microorganism categories and distributions were collected and analyzed. The patients were divided into survival and death groups based on the the 28-day prognosis, and the differences of clinical data between the two groups were compared. Logistic regression analysis was used to select the independent prognostic risk factors of BSI. RESULTS: One hundred and fifty-four patients with positive results of BSI were enrolled, and the 156 strains of bacteria were detected, including 75 Gram positive (G+) bacteria strains, 70 Gram negative (G-) bacteria strains and 11 fungi strains among those BSI patients. The top five pathogenic microorganisms were Coagulase negative staphylococcus (35.9%), Klebsiella pneumonia (12.8%), Acinetobacter baumannii (9.0%), Enterococci (9.0%), and Escherichia coli (8.3%). There were 45 strains (60.0%) of multidrug-resistant strains in G+ bacteria and 40 strains of multidrug-resistant strains (57.1%) in G- bacterial strains, but in fungi there was only 1 strain (9.1%). There were 81 cases in survival group and 73 cases in death group according to 28-day prognosis. We found that there were no significant differences between the comparators in age, lenth of stay in ICU, duration for mechanical ventilation after being diagnosed with BSI, percentage of BSI with chronic obstructive pulmonary disease (COPD), hypertension, cardiovascular disease or chronic kidney dysfunction (all P > 0.05). In death group, the proportion of male was obviously lower than that of survival group [58.9% (43/73) vs. 75.3% (61/81), P < 0.05] and APACHE II score was significantly higher than that in survival group (27.1±7.0 vs. 19.5±6.7, P < 0.05). The mean arterial pressure (MAP) of death group on first 3 days of BSI was significantly lower than that of survival group [mmHg (1 mmHg = 0.133 kPa): 72.8±13.6 vs. 79.7±12.9, P < 0.05), in the death group, the proportion of patients complicated with diabetes, cancer [28.8% (29/73) vs. 12.3% (10/81), 19.2% (14/73) vs. 7.4% (6/81)], post-BSI CRRT and blood transfusion [39.7% (29/73) vs. 16.0% (13/81), 64.4% (47/73) vs. 46.9% (38/81)], and continuous use of asoactie drugs for ≥ 72 hours [37.0% (27/73) vs. 12.3% (10/81)] were significantly higher than those in the survival group (all P < 0.05). In death group, platelet count (PLT) was significantly decreased than that of survival group [×109/L: 124.93±98.21 vs. 181.15±116.39,P < 0.05], aspartate aminotransferase (AST) level was significantly higher than that of survival group [U/L: 75.40 (38.50, 140.95) vs. 56.20 (29.20, 85.70), P < 0.05], the rest of the laboratory indexes had no statistically significant differences between the two groups (all P > 0.05). The results of Logistic regression analysis showed that the APACHE II score [odds ratio (OR) = 1.279, 95% confidence interval (95%CI) was 1.158 to 1.412, P < 0.001], CRRT after BSI (OR = 3.522, 95%CI was 1.013 to 12.245, P = 0.048) were independent risk factors affecting the prognosis of patients with BSI, and MAP is a protective factor for prognosis (OR = 0.961, 95%CI was 0.927 to 0.996, P = 0.031). CONCLUSIONS: In our ICU, G+ bacteria are still dominant in bloodstream infection, G- bacteria take the second place. Besides, APACHE II score and CRRT after being diagnosed with BSI are the independent prognostic risk factors.

Isolation and functional analysis of squalene synthase gene in tea plant Camellia sinensis.

Tea contains high quantities and diverse types of triterpenoids, particularly in the form of saponins. However, little is yet known about the molecular basis of triterpenoid biosynthesis in tea plant. Here we report on isolation and functional analysis of squalene synthase (SQS) gene from tea plant (Camellia sinensis var. sinensis), which controls the biosynthesis of triterpenoids precursor. First, a full-length cDNA of squalene synthase, designated CsSQS, was isolated from tea plant. The protein is highly homologous to SQSs from other plants. Using CsSQS-reporter assays, CsSQS was demonstrated to be endoplasmic reticulum membrane-bound. The coding region of CsSQS excluding transmemberane sequence was expressed in Escherichia coli. Recombinant CsSQS catalyzed the formation of squalene using farnesyl-pyrophosphate (FPP) as substrate with NADPH and Mg2+. In tea plant leaves, CsSQS expression was significantly induced by both herbivore and mechanical damages. Consistent with the stronger induction of CsSQS expression by mechanical damage than herbivory, tea plants injured mechanically released squalene as a volatile compound, which however was not detected from herbivore-damaged tea plants. Furthermore, it was found that the flowers of another tea plant cultivar Camellia sinensis var. assamica contain higher concentrations of squalene than the cultivar sinensis, indicating variations among tea plant varieties. With the identification and molecular characterization of squalene synthase in tea plant, next, we can ask the questions about the roles of squalene as a volatile product as well as a precursor for triterpenoids, which may promote product development from diverse tea materials and mining of excellent tea germplasm resources.

Cloning of a new glutathione peroxidase gene from tea plant (Camellia sinensis) and expression analysis under biotic and abiotic stresses.

BACKGROUND: Tea plant, Camellia sinensis (L.) O. Kuntze, a well-known heavy metal hyperaccumulator, possesses a powerful tolerance to heavy metals. The heavy metal stresses lead to reactive oxygen species (ROS) production, and high concentration of ROS is harmful to plants. The glutathione peroxidase gene has positive function to damage induced by ROS. To understand the mechanism of tolerance to deferent stresses in tea plant, a new glutathione peroxidase gene of tea plant was cloned and its expression pattern was analyzed under abiotic and biotic stresses. RESULTS: A novel cDNA encoding glutathione peroxidase of tea plant (Camellia sinensis) was isolated by rapid amplification of cDNA ends (RACE) method and designated as CsGPX2 (GenBank Accession No. JQ247186). This full-length sequence was 917 nucleotides including a 510 bp open reading frame (ORF), which encoded a polypeptide of 169 amino acids. The deduced amino acid sequence showed high homology with glutathione peroxidases of angiosperms and contained the characteristic conserved motifs of ILAFPCNQF and FTVKD, the highest level of similarity was 85% to a glutathione peroxidase from Ricinus communis (Accession NO. XP_002509790.1). Tissue expression pattern analysis indicated that CsGPX2 expressed similarly in root, stem, leaf and flower of tea plant. The CsGPX2 gene showed strong responses to most abiotic stresses including salinity, heavy metal toxicity, drought, heat, plant hormones, but could not be induced by biotic treatment. CONCLUSIONS: The result suggested that CsGPX2 had potential function in protecting tea plant from peroxidative damage induced by some abiotic stresses.

[Preparation of a monoclonal antibody against polyhedrin of Ectropis obliqua nucleopolyhedrovirus].

To develop a method based on immunoreactions for detection of Ectropis obliqua Nucleopolyhedrovirus (EoNPV), the polyhedra of the virus were purified and used to immunize the mouse BALB/c. The spleen cells from the immunized mice were then fused with the myeloma cell line Sp2/0. A hybridoma cell line which can stably secrete the monoclonal antibody against EoNPV was achieved by using indirect ELISA screening and cloning methods, and was named as 7D3. Meanwhile, the polyhedrin gene was cloned from EoNPV and expressed in E. coli. Western blotting analysis showed that the monoclonal antibody prepared from 7D3 could specifically react with the recombinant polyhedrin. An indirect ELISA method based on this monoclonal antibody for detecting EoNPV in infected tea looper was developed.