Genotypic-dependent effects of N fertilizer, glutathione, silicon, zinc, and selenium on proteomic profiles, amino acid contents, and quality of rice genotypes with contrasting grain Cd accumulation.

Soil heavy metal (HM) contamination has posed a serious problem for safe food production. For restricting the translocation of HM into grain, many proteins were regulated to involve in the process. To identify these proteins, 2D-based proteomic analysis was carried out using different rice genotypes with distinct Cd accumulation in grains and as affected by an alleviating regulator (AR) in field experiments. AR application improved grain quality, with increased contents in Glu, Cys, His, Pro, and protein. Twenty-six low-grain HM accumulation-associated protein species were identified and categorized as physiological functions via two-dimensional gel electrophoresis (2DE) and mass spectrometry. Among these proteins, 8, 9, and 9 proteins exhibited higher accumulation, lower accumulation, and unchanged accumulation, respectively, in Xiushui817 (low accumulator) vs R8097 (high accumulator) under control conditions but showed differential accumulation patterns after AR application. These proteins included sucrose synthase 3, alanine aminotransferase, glutelin, cupin family protein, and zinc finger CCCH domain-containing protein 32. The differential expression of these protein species might contribute to decreased HM accumulation in grain via decreasing the protein accumulation which had high affinity to HM or regulating energy metabolism and signal transduction. Our findings provide valuable insights into the mechanisms of low-grain HM accumulation in rice and possible utilization of candidate protein species in developing low-grain HM accumulation genotypes.

Abundance and diversity of tetracycline resistance genes in soils adjacent to representative swine feedlots in China.

Tetracyclines are commonly used antibiotics in the swine industry for disease treatment and growth promotion. Tetracycline resistance was determined in soils sampled from farmlands in the vicinity of nine swine farms located in three cities in China. Totally, 15 tetracycline resistance (tet) genes were commonly detected in soil samples, including seven efflux pump genes (tetA, tetC, tetE, tetG, tetK, tetL, tetA/P), seven ribosomal protection proteins (RPPs) genes (tetM, tetO, tetQ, tetS, tetT, tetW, tetB/P), and one enzymatic modification gene (tetX). The quantitative real-time PCR was further used to quantify five RPPs genes (tetM, tetO, tetQ, tetW, tetT) and 16S rRNA gene abundances. The concentrations of total tetracyclines (5 typical tetracyclines and 10 of their degradation products) in these soils were measured using liquid chromatography-electrospray tandem mass spectrometry (LC-MS/MS) and were found to range from 5.4 to 377.8 µg·kg(-1) dry soil. Bivariate correlation analysis confirmed that absolute tet gene copies (sum of tetM, tetO, tetQ, tetW genes) were strongly correlated with the concentrations of tetracycline residues (r(2) = 0.45, P < 0.05), ambient bacterial 16S-rRNA gene copies in each soil sample (r(2) = 0.65, P < 0.01), and organic matter in soil (r(2) = 0.46, P < 0.05), respectively. Finally, the phylogenetic analysis on tetM combined with culture-independent molecular techniques revealed at least five genotypes of tetM in nine soil samples.

The effect of straw-returning on antimony and arsenic volatilization from paddy soil and accumulation in rice grains.

Pollution by antimony (Sb) and arsenic (As) in soil can pose a great threat to human health. Straw-returning is widely applied to paddy fields for improving and remediating soil. A pot experiment was conducted to investigate the effect of straw-returning on Sb and As transformation and translocation in a soil-rice system. In this study, Sb and As co-contaminated soil was thoroughly mixed with different proportions (0, 0.5, 1, and 2%) of straw and used for growing rice plants through the entire growing stage in a pot experiment and 4 weeks in a microcosm experiment. The straw application significantly increased Sb and As mobility. The concentrations of total Sb and As in soil-pore water increased after the application of straw in most growing stages. The Sb volatilization in the pot and microcosm experiments was also stimulated by straw application. With the high dose of straw application (2%), the concentration of Sb in brown grain was reduced by 72% compared with the control, but As concentrations increased by around 77%. These findings provide a new perspective in that straw-returning could affect the behavior of both Sb and As in soil and reduce the Sb accumulation in brown grain and some guidance in the use of straw-returning in Sb-contaminated paddy soil.

Rice nitrogen use efficiency does not link to ammonia volatilization in paddy fields.

Ammonia (NH3) volatilization is a major pathway of nitrogen (N) losses from paddy fields, and could be potentially mitigated by cultivation of high nitrogen use efficiency (high-NUE) rice cultivars. However, the relationship between NUE and NH3 volatilization has not been validated under field conditions. A field experiment was conducted to evaluate the impact of four rice cultivars with different NUE [Wuyunjing 23 (W23), Zhendao 11 (Z11), Wuyujing 3 (W3), and Aoyusi 386 (A386)] on NH3 volatilization, as well as the related mechanisms. Two high-NUE rice cultivars W23 and Z11 was not more effective in reducing total NH3 volatilization from the paddy field compared to cultivar A386 with the lowest NUE. Cultivar A386 had 12.7-17.8% and 35.7-54.1% lower NH3 volatilization than other three rice cultivars at tillering fertilization stage (TFS) and panicle fertilization stage (PFS), respectively, mainly due to its greater shoot N accumulation, root biomass and volume at TFS and its greater shoot biomass, leaf area index and shoot N accumulation at PFS. There was no significant difference in NH3 volatilization among W23, Z11 and W3 at TFS. However, premature senescence phenomenon at later growth stages of A386 eventually led to its lowest NUE among the four rice cultivars. Our results suggest that NUE of rice does not link to NH3 volatilization from paddy fields. In order to make high-NUE rice cultivars also effective in mitigating NH3 volatilization, future breeding works should aim to improve N uptake capability and canopy structure at early tillering and panicle development stages while prevent premature senescence of rice plants to maintain high yields.

The characterization of arsenic biotransformation microbes in paddy soil after straw biochar and straw amendments.

Straw biochar and straw application to paddy soil dramatically altered arsenic (As) biogeochemical cycling in soil-rice system, but it remains unknown how As biotransformation microbes (ABMs) contribute to these processes. In this study, rice pot experiments combining terminal restriction fragment length polymorphism (T-RFLP) analysis and clone library were performed to characterize ABMs. Through linear discriminant analysis (LDA) effect size (LEfSe) and correlation analysis, results revealed that arrA-harbouring iron-reducing bacteria (e.g., Geobacter and Shewanella) and arsC-harbouring Gammaproteobacteria (e.g., fermentative hydrogen-producing and lignin-degrading microorganisms) potentially mediated arsenate [As(V)] reduction under biochar and straw amendments, respectively. Methanogens and sulfate-reducing bacteria (SRB) carrying arsM gene might regulate methylated As concentration in soil-rice system. Network analysis demonstrated that the association among ABMs in rhizosphere was significantly stronger than that in bulk soil. Arsenite [As(III)] methylators carrying arsM gene exhibited much stronger co-occurrence pattern with arsC-harbouring As(V) reducers than with arrA-harbouring As(V) reducers. This study would broaden our insights for the dramatic variation of As biogeochemical cycling in soil-rice system after straw biochar and straw amendments through the activities of ABMs, which could contribute to the safe rice production and high rice yield in As-contaminated fields.

Microbiota in non-flooded and flooded rice culms.

Rice plants are the habitat for large and diverse populations of microbes, which play important roles on rice health and productivity. However, the response of microbiome on rice culm to water flooding is poorly understood. In this study, the bacterial community on non-flooded (RSA) and flooded (RSB) rice culms was investigated through 16S rRNA gene sequencing. The results showed that RSA and RSB had significantly distinct bacterial communities. In RSA, Gammaproteobacteria and Pantoea were the most abundant class (57%), genus (37.06%), respectively, while in RSB, the most abundant phylum and genus was Firmicutes (54%) and Bacillus (52.63%), respectively. Compared with RSA, the abundance of 27 genera significantly increased and 21 genera significantly decreased in RSB, and some remarkably changed species, such as Aeromonas, Bacillus were identified, which are sensitive to non-flooded or flooded conditions. In addition, rare operational taxonomic units (OTUs) was much more than abundant OTUs in all samples, and RSB had significantly higher bacterial richness than RSA due to having more rare taxa. Our study would advance the insights into the microbiome of rice culms and its response to flooding, which would help to identify potential beneficial bacteria for improving crop health and sustainable productivity in agroecosystems.

NH4H2PO4-extractable arsenic provides a reliable predictor for arsenic accumulation and speciation in pepper fruits (Capsicum annum L.).

Dietary arsenic (As) intake from food is of great concern, and developing a reliable model capable of predicting As concentrations in plant edible parts is desirable. In this study, pot experiments were performed with 16 Chinese upland soils spiked with arsenate [As(V)] to develop a predictive model for As concentrations in pepper fruits (Capsicum annum L.). Our results showed that after three months' aging, concentrations of bioavailable As (extracted by 0.05 M NH4H2PO4) in various soils varied widely, depending on soil total As concentrations and soil properties such as soil pH and amorphous iron (Fe) contents. Furthermore, both the bioconcentration factor (BCF, denoted as the ratio of fruit As to soil As) and total As concentrations in pepper fruits were largely determined by concentrations of bioavailable As, which explained 27% and 69% variations in the BCF and fruit As concentrations, respectively. Apart from bioavailable As, soil pH and Fe contents were another two important factors influencing As accumulation in pepper fruits. Taking the three factors into account, concentrations of fruit As can be well predicted using a stepwise multiple linear regression (SMLR) analysis (R2 = 0.80, RMSE = 0.17). Arsenic species in soils and edible parts were also analyzed. Although As(V) predominated in soils (>96%), As in pepper fruits presented as As(V) (46%) and arsenite [As(III)] (39%) with small amount of methylated As (<15%). Aggregated boosted tree (ABT) analysis revealed that inorganic As concentrations in pepper fruits were determined by concentrations of bioavailable As, phosphorus (P) and Fe in soils. In contrast to inorganic As, methylated As concentrations were not correlated with those factors in soils. Taken together, this study established an empirical model for predicting As concentrations in pepper fruits. The predictive model can be used for establishing the As threshold in fruit vegetable farming soils.

Turning pig manure into biochar can effectively mitigate antibiotic resistance genes as organic fertilizer.

The composting of fresh manure is an effective way to inactivate pathogens and reduce the levels of antibiotics and some antibiotic resistance genes (ARGs) prior to its application on agricultural land as organic fertilizer. However, some ARGs could still exist and even be enriched after composting. This study investigated whether converting composted pig manure into biochar could reduce the dissemination of ARGs into the soil in comparison with a compost amendment. We performed a pot experiment using pakchoi (Brassica chinensis), with two pig manure-based composts and the biochar derived from composted pig manure, as organic fertilizers. The distributions of the antibiotic resistome, mobile genetic elements (MGEs) and bacterial community composition in soils during cultivation were evaluated by high-throughput qPCR and Illumina sequencing. The total ARGs and MGEs abundance in the biochar-treated soils were significantly lower than those in the compost-amended soils during cultivation. The total ARGs abundance in the biochar-amended soils was similar to that in the control soils during cultivation. Thus, the dissemination of ARGs from animal waste to the environment can be effectively mitigated by converting manure into biochar.

Genotypic and environmental variation in chromium, cadmium and lead concentrations in rice.

Genotypic and environmental variation in Cr, Cd and Pb concentrations of rice grains and the interaction between these metals were investigated by using 138 rice genotypes grown in three contaminated soils. There were significant genotypic differences in the three heavy metal concentrations of rice grains, with the absolute difference among 138 genotypes in grain Cr, Cd and Pb concentrations being 24.5-, 9.1- and 23.8-folds, respectively, under the slightly contaminated soil (containing 4.61mgkg(-1) Cr, 1.09mgkg(-1) Cd and Pb 28.28mgkg(-1), respectively). A highly significant interaction occurred between genotype and environment (soil type) in the heavy metal concentrations of rice grains. Cr concentration in rice grains was not correlated with Cd and Pb concentration. However, there was a significant correlation between Cd and Pb in slightly and highly contaminated soils. The results suggest the possibility to develop the rice cultivars with low Cd and Pb concentrations in grain.

[Effects of Straw Incorporation on Cadmium Accumulation and Subcellular Distribution in Rice].

Cadmium (Cd) is classified as a Group-1 human carcinogen and rice consumption constitutes a major source of dietary intake of Cd for populations whose staple food is rice. Straw incorporation is widely performed in Cd-contaminated paddy fields, which may significantly affect the bioavailability of Cd in soil and the distribution of Cd in rice plants, consequently altering Cd accumulation in rice grains. In this study, both pot and field trials were conducted to investigate the effects of different amounts of straw incorporation (0.0%, 1.0%, 2.5%, and 5.0%) on Cd sub-cellular distribution in rice plants and Cd accumulation in rice grains. The results showed that Cd was mainly sequestered in cell wall, accounting for 86%-95% and 30%-51% of total cadmium in root and shoot cells, respectively. In shoot cells, about 35%-61% of Cd was distributed in cellular soluble fractions. When rice straw was incorporated at 1.0% and 2.5% levels, Cd sequestration in the cell wall significantly increased and Cd translocation from roots to shoots significantly decreased. However, when rice straw was incorporated at the 5% level, Cd sequestration in root cell walls significantly decreased and Cd translocation from roots to shoots significantly increased at the tillering stage. At the filling stage, 5% rice straw incorporation still significantly increased Cd sequestration in root cell walls and Cd translocation from roots to shoots did not significantly change. The rice straw and rape straw used for the field trail contained high concentrations of Cd (0.49 and 0.67 mg·kg-1, respectively). Rape straw incorporation alone or together with lime did not significantly affect Cd accumulation in brown rice or rice straw. Rice straw incorporation alone did not significantly affect Cd accumulation in brown rice or rice straw, while incorporation with lime significantly decreased Cd accumulation in both brown rice and rice straw. Biochar application can also significantly reduce Cd accumulation in rice and when biochar was added together with lime, the reduction in Cd accumulation in rice was more significant. Therefore, at Cd-contaminated paddy fields, rice straw or rape straw is not suggested to be returned directly; incorporation with lime would be better for reducing Cd accumulation in rice grains. The results of this study will provide theoretical and practical guidance for the safe production of rice and for straw recycling at Cd-polluted paddy fields.

Genotypic and environmental variation in cadmium, chromium, arsenic, nickel, and lead concentrations in rice grains.

Genotypic and environmental variation in Cd, Cr, As, Ni and Pb concentrations of grains, and the relationships between these heavy metals and Fe, Zn were investigated using 9 rice genotypes grown in 6 locations for two successive years. Significant genotypic variation was detected in the five heavy metal concentrations in grains, indicating the possibility to reduce the concentration of these heavy metals in grains through breeding approach. The environmental effect varied with metal, with Pb and Ni having greater variation than the other three metals. There was significant genotype-environment (location) interaction of the concentrations of all five heavy metals in grains, suggesting the importance of cultivar choice in producing rice with low heavy metal concentrations in grains for a given location. Correlation analysis showed that Cd and As, Cr and Ni, and As and Pb concentrations in rice grains were closely associated, and that Ni concentration in grains was negatively correlated with Zn concentration.

Genotypic and environmental variation in cadmium, chromium, lead and copper in rice and approaches for reducing the accumulation.

The field scale trials revealed significant genotypic and environmental differences in grain heavy metal (HM) concentrations of 158 newly developed rice varieties grown in twelve locations of Zhejiang province of China. Grain Pb and Cd contents in 5.3% and 0.4% samples, respectively, were above the maximum permissible concentration (MPC); none of samples had Cr/Cu exceeding MPC. Stepwise multiple linear regression analysis estimated soil HM critical levels for safe rice production. Low grain HM accumulation cultivars such as Xiushui817, Jiayou08-1 and Chunyou689 were recommended as suitable cultivars for planting in slight/medium HM contaminated soils. The alleviating regulator (AR) of (NH4)2SO4 as N fertilizer coupled with foliar spray of a mixture containing glutathione (GSH), Si, Zn and Se significantly decreased grain Cd, Cr, Cu and Pb concentrations grown in HM contaminated fields with no effect on yield, indicating a promising measurement for further reducing grain HM content to guarantee safe food production.

Effects of pH, Fe, and Cd on the uptake of Fe(2+) and Cd (2+) by rice.

The rhizosphere plays an important role in altering cadmium (Cd) solubility in paddy soils and Cd accumulation in rice. However, more studies are needed to elucidate the mechanism controlling rice Cd solubility and bioavailability under different rhizosphere conditions to explain the discrepancy of previous studies. A rice culture with nutrient solution and vermiculite was conducted to assess the effects of pH, Eh, and iron (Fe) concentration on Cd, Fe fractions on the vermiculite/root surface and their uptake by rice. The solution pH was set from 4.5 to 7.5, with additions of Fe (30 and 50 mg L(-1)) and Cd (0.5 and 0.9 mg L(-1)). At pH 5.5, the Eh in the rice rhizosphere was higher whereas transpiration, Cd(2+), and Fe(2+) adsorption on the vermiculite/root surface and accumulation in rice were lower than the other pH treatments. Cadmium addition had no impact on pH and Eh in rice rhizosphere while Fe addition decreased pH and increased Eh significantly. Compared with control, Fe addition resulted in the decrease of rhizosphere Cd, Fe solubility and bioavailability. Higher redox potential in the rice rhizosphere resulted in the decline of transpiration, Cd, and Fe accumulation in the rice tissues, suggesting that the transfer of two elements from soil to rice was depressed when the rhizosphere was more oxidized.

Modulation of exogenous glutathione in phytochelatins and photosynthetic performance against cd stress in the two rice genotypes differing in Cd tolerance.

Greenhouse hydroponic experiments were conducted using Cd-sensitive (Xiushui63) and tolerant (Bing97252) rice genotypes to evaluate genotypic differences in response of photosynthesis and phytochelatins to Cd toxicity in the presence of exogenous glutathione (GSH). Plant height, chlorophyll content, net photosynthetic rate (Pn), and biomass decreased in 5 and 50 µM Cd treatments, and Cd-sensitive genotype showed more severe reduction than the tolerant one. Cadmium stress caused decrease in maximal photochemical efficiency of PSII (Fv/Fm) and effective PSII quantum yield [Y(II)] and increase in quantum yield of regulated energy dissipation [Y(NPQ)], with changes in Cd-sensitive genotype being more evident. Cadmium-induced phytochelatins (PCs), GSH, and cysteine accumulation was observed in roots of both genotypes, with markedly higher level in PCs and GSH on day 5 in Bing97252 compared with that measured in Xiushui63. Exogenous GSH significantly alleviated growth inhibition in Xiushui63 under 5 µM Cd and in both genotypes in 50 µM Cd. External GSH significantly increased chlorophyll content, Pn, Fv/Fm, and Y(II) of plants exposed to Cd, but decreased Y(NPQ) and the coefficient of non-photochemical quenching (qN). GSH addition significantly increased root GSH content in plants under Cd exposure (except day 5 of 50 µM Cd) and induced up-regulation in PCs of 5 µM-Cd-treated Bing97252 throughout the 15-day and Xiushui63 of 5-day exposure. The results suggest that genotypic difference in the tolerance to Cd stress was positively linked to the capacity in elevation of GSH and PCs, and that alleviation of Cd toxicity by GSH is related to significant improvement in chlorophyll content, photosynthetic performance, and root GSH levels.

[Diversity of plant in Jiaxing Shijiuyang ecological wetland for drinking water during operation].

The Shijiuyang ecological wetland for drinking water of Jiaxing City, Zhejiang Province is one of the biggest constructed wetlands for water resource protection in China. To ensure a deep understanding of the present status of the wetland vegetation of Shijiuyang ecological wetland which has been run for 2.5 years and provide support for the vegetation management of ecological wetland, systematic investigation was carried out by using plot method and quadrat method in October to November, 2010. The species composition, dynamics of plant diversity and the biomass production during operation were analyzed. Altogether 70 species belonging to 28 families and 62 genera were recorded. Among them, there were 26 wetland plants, 20 mesophytes, 14 emergent, 4 submerged, 6 floating ones. Compared with the preliminary stage, the species numbers of wetland plants increased significantly from 15 species to 70 species. The spatial pattern of riparian species diversity was examined by adopting the Simpson index and Shannon-Wiener index as species diversity indices. The results showed that the riparian species diversity was higher in the west of the Beijiaohe river (Simpson index = 0.468 3, Shannon-Wiener index = 0.835 2) than that in the south of the Dongsheng Road (Simpson index = 0.357 6, Shannon-Wiener index = 0.660 4). The analyses of quantitative characteristics of wetland vegetation showed that the plants in the root-channel purification zone in the south of the Dongsheng Road grew better than those in the west of the Beijiaohe river. With regard to the riparian vegetation, the riparian plants in the west of the Beijiaohe river were more abundant. The mean biomass production (dry weight) in the root-channel purification zone was 1.73 kg x m(-2) and the total area was 9.12 x 10(4) m2, so the total biomass production was estimated to be 157.8 t. In the same way, the mean riparian vegetation biomass production(dry weight) was 0.83 kg x m(-2) and the total vegetation area was 3.75 x 10(4) m2, so the total riparian vegetation biomass production(dry weight) was estimated as 31.1 t.

[Effect of iron plaque formation of root surface on as uptake by rice seedlings grown on different types of soils].

The objective of study was to investigate the effect of the amount of iron plaque of root surface on As uptake by rice seedlings grown on 14 types of soils. These results indicated that there were significant differences in the amounts of iron plaque (1.15-61.97 mg/g) formed on rice root surface among different soils. The amount of non-crystalloid Fe oxide in different submerged soils is one of main influencing factors. There was a significant positive correlation between As concentrations in iron plaque (0-1376 mg/kg) and the amount of iron plaque (r = 0.85, n=14, p < 0.05), but no significant correlation with the extractable As concentration of 14 types of soils. There were also significant positive correlation between As concentrations in shoot (0.400-12.98 mg/kg) and root (3.860-576.2 mg/kg) and the amounts of iron plaque respectively but not significant correlation with the extractable As concentrations of 14 types of soils (r = 0.88, n=14, p < 0.05; r = 0.91, n=14, p < 0.05). Moreover, there was a significant positive correlation between specific arsenic uptake (0.005-0.670 mg/g) and the amount of iron plaque (r = 0.91, n=14, p < 0.05). However, there was no significant correlation between shoot-As% and the amount of iron plaque. These results showed that the iron plaque on root surface was the pool of As in rhizosphere and increased As uptake but did not affect As transportation from rice root to shoot.

Resistance and resilience of Cu-polluted soil after Cu perturbation, tested by a wide range of soil microbial parameters.

Copper (Cu)-polluted and unpolluted soils were used to study the effect of initial pollution on soil biological resistance and resilience by measuring the responses to perturbation using different parameters. Microbial biomass carbon, substrate-induced respiration and copy numbers of 16S rRNA gene were grouped as general parameters, while potential ammonia oxidation rate and copy numbers of amoA gene were grouped as specific functions. In addition, to illustrate how initial pollution affects soil biological resistance and resilience following secondary perturbation, the microbial community structure, together with free Cu(2+) activities ([Cu(2+)]) in soil pore water and soil pH were also measured after secondary perturbation. Results showed that general parameters were more stable than specific ones. High [Cu(2+)] and low pH in soil pore water induced by Cu addition may lead to apparently low resistance and resilience, whereas the formation of a tolerant community after Cu pollution, secondary perturbation and Cu aging may contribute to resistance and resilience. Analysis of the phospholipid fatty acids profile showed that microbial community structure shifted along with the [Cu(2+)] gradient. The microbial community structure of the control soil was both resistant and resilient to 400 mg kg(-1) Cu perturbation, whereas other treatments were neither resistant nor resilient.

Mapping quantitative trait loci associated with arsenic accumulation in rice (Oryza sativa).

The quantitative trait loci (QTLs) associated with arsenic (As) accumulation in rice were mapped using a doubled haploid population established by anther culture of F1 plants from a cross between a Japonica cultivar CJ06 and an Indica cultivar TN1 (Oryza sativa). Four QTLs for arsenic (As) concentrations were detected in the map. At the seedling stage, one QTL was mapped on chromosome 2 for As concentrations in shoots with 24.4% phenotypic variance and one QTL for As concentrations in roots was detected on chromosome 3. At maturity, two QTLs for As concentrations in grains were found on chromosomes 6 and 8, with 26.3 and 35.2% phenotypic variance, respectively. No common loci were detected among these three traits. Interestingly, the QTL on chromosome 8 was found to be colocated for As concentrations in grain at maturity and shoot phosphorus (P) concentrations at seedling stage. These results provide an insight into the genetic basis of As uptake and accumulation in rice, and will be useful in identifying genes associated with As accumulation.