Cultivar And Environmental Effects And Regulation Of Heavy Metal Accumulation In Rice

Soil heavy metal (HM) pollution poses a severe detrimental influence on environment, safety food production and human health. HMs are absorbed by plants and accumulated in edible parts, thus pose a potential threat to human health via food chain. Rice (Oryza sativa), as a staple food cereal in the world, is the major source of HM intakes for human. It is a cost-effective approach to realize safe food production, via breeding low-HM-accumulation cultivars, regulating HM uptake and translocation, and reducing grain HM accumulation, especially in slight or medium HM polluted soils for sustainable agriculture. Screening low-HM-accumulation cultivars and understanding the underlying mechanisms related to HM accumulation and tolerance are the basis to breed low-HM-accumulation cultivars. Accordingly, a series of experiments were conducted based on the evaluation of present situation of soil HM pollution in different main rice production areas in Zhejiang province, to identify low-HM-accumulation cultivars; develop forecast index system to predict grain HM concentration; develop alleviating regulator (AR) to reduce grain HM accumulation and the corresponding mechanisms. Meanwhile, the Cd-tolerant and grain low-Cd-accumulation related proteins were also identified. The main results were summarized as follows:1. Cultivar and environmental variation in heavy metal concentration in rice grainsField scale trial was conducted to evaluate cultivar and environmental differences in grain HM concentrations of144newly developed rice grown in twelve locations of Zhejiang province, China, and as affected by soil properties. Significant cultivar and environmental differences were observed in grain HM concentration. Grain Pb and Cd contents in5.3%and0.4%samples respectively were above maximum permissible concentration (MPC), none of samples had Cr/Cu exceeding MPC. There was higher grain Cd concentration in hybrid rice than that in conventional cultivars. The stepwise multiple linear regression analysis showed the multiple effects of soil pH and EDTA-extractable heavy metal contents on grain HM content, and the critical levels of soil EDTA-Cd, Cr and Pb for safe rice production were estimated as0.200,0.372and22.093mg kg-1, respectively. Cultivars Xiushui817, Jiayou08-1and Chunyou689showed consistently low grain HM concentrations in different locations, thus recommended as suitable cultivars for planting in slight/medium HM contaminated soils.2. Effect of combined application of cadmium, chromium, lead and copper on grain, leaf and stem heavy metal contents at different stagesPot experiments were carried out to investigate the cultivar differences in yield, and grain Cd, Cr concentration and the relationship with leaf and stem at different growth stages under combined application of Cd, Cr, Pb and Cu according to orthogonal design. Yield was significantly decreased under higher Pb and Cd treatment in both cultivars, and Xiushui817was affected more. The yield was not affect with increasing soil Cu level. Only grain Cd concentration showed external Cd level dependant increase in both cultivars. Meanwhile, Zheda821has higher grain Cd content than Xiushui817. Soil Pb, Cd, Cr and Cu content significantly affect grain Cd, Cr and Cu concentration. Similar trends were also observed in leaves and stems at harvest stage. Besides root Cd concentration, Cr and Cu content was also increased with increasing soil Cr and Cu level, respectively. Grain Cd and Cr concentrations were strongly correlated with both leaves and stems at different growth stages except leaf Cr at filling and stem Cr at seedling stages. Regression equations between grain and leaf/stem-Cd/Cr at different stages were established and the critical levels of leaf/stem Cd and Cr for safe rice production were also estimated.3. Cultivar differences of root proteomic expression in response to Cd toxicity and as affected by silicon applicationHydroponic experiments were performed using Zheda821and Xiushui817to evaluate cultivar differences of root proteomic expression in response to5μM Cd stress in presence of silicon (Si) by TMT-based quantitative comparative proteomics. The results showed that Zheda821exhibited higher Cd tolerance than Xiushui817. Although root Cd concentration was higher in Xiushui817than that in Zheda821, shoot Cd content showed no significant difference. Twenty-one Cd-tolerance/resistance-associated proteins in root were identified and categorized in stress and defense response, transport, metabolism and signal transduction, which constituted an integrated Cd tolerance related schematic diagram. There was thirty-five proteins were up-regulated in both cultivars after Cd stress. Meanwhile, we also identified twenty-five grain low-Cd-accumulation related proteins in Xiushui817, including vacuolar ATPase, vacuolar H+-pyrophosphatase, β-expansin precursor and UDP-glucose6-dehydrogenase which involved in cell wall synthesis, and FIFO-ATPase inhibitor protein, which might contribute to translocate Cd into vacuolar and further reduce Cd translocation into grain in Xiushui817.4. Comparative study of alleviating effects of GSH, Se and Zn under combined contamination of cadmium and chromium in rice (Oryza sativa)A hydroponic experiment was conducted to study the ameliorative effects of separate or combined application of exogenous glutathione (GSH), selenium (Se) and zinc (Zn) upon20μM cadmium (Cd) puls20(iM chromium (Cr) heavy metal stress (HM) in rice seedlings. The results showed that HM caused a marked reduction in seedling height, chlorophyll content (SPAD) and biomass, and activities of catalase (CAT) and ascorbate peroxidase (APX) in leaves and H+-ATPase in roots/leaves, and elevated superoxide dismutase (SOD) and guaiacol peroxidase (POD) activities in leaves with elevated malondialdehyde (MDA) accumulation both in leaves and roots, compared to control. The best mitigation effect was recorded in HM+GSH+Zn and HM+GSH (addition of GSH+Zn and GSH to HM solution), which greatly alleviated HM-induced growth inhibition and oxidative stress. Compared with HM alone, HM+GSH and HM+GSH+Zn markedly reduced Cr uptake and translocation but not affected Cd concentration; imporved H+-ATPase activity and Fe, Zn, Mn uptake and translocation, and repressed MDA accumulation. Meanwhile exogenous GSH and GSH+Zn counteracted HM-induced response of antioxidant enzymes, via suppressing HM-induced dramatic increase of root/leaf SOD and leaf POD activities, and elevating stress-depressed leaf APX and leaf/root CAT activities.5. Effect of different N fertilizers, reduced glutathione, Si, Zn and Se on heavy metal accumulation in rice grainsTwo years field experiments were carried out to investigated the role of (NH4)2SO4as N fertilizer coupled with foliar spray of a mixture containing glutathione (GSH), Si, Zn and Se (namely alleviating regulator, AR) on grain HM accumulation. Xiushui817and Jiayou08-1showed low grain Cd accumulation, which confirmed our previous study. The application of AR significantly decreased grain Cd, Cr, Cu and Pb concentrations both in high-and low HM accumulation cultivars grown in HM contaminated soils with no effect on yield, indicating a promising measurement for further reducing grain HM content to guarantee safe food production. In2011, leaf Cd, Cr and Cu, and stem Cd and Cu concentration at harvest stage were significantly decreased after application of AR practices, compared with control. A similar result were found in glumes, leaves and stems Cd, Cr and Cu concentrations, except stem Cu concentration which was markedly increased in2012.6. Effect of different N fertilizers, reduced glutathione, Si, Zn and Se on quality, amino acids and proteomic profiles in rice grainField experiment was performed to evaluate cultivar differences in grain protein expression, quality and amino acids content in R8097and Xiushui817and as affected by AR treatment. Results showed that contents of amino acids, amylose and protein in grains showed no or significant increase after AR treatment, compared with control. Thirty proteins were identified which were categorized mainly in metabolism, storage protein, energy, defense, detoxification, signal transduction, transcription. Compared with Xiushui817, seven proteins were up-regulated in R8097under control condition, but down or totally inhibited by AR application; two proteins were down-regulated in R8097but enhanced in AR treatment. Meanwhile,21proteins whose expression was up-regulated, down-regulated in both genotypes, or up-regulated in R8097and non-changed in Xiushui817after AR application were idetified compared with control.