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Proton and copper (Cu) binding to soil and lignite-based humic acid (HA) was investigated by combining X-ray absorption fine structure (XAFS) spectroscopy, isothermal titration calorimetry (ITC), and nonideal-competitive-adsorption (NICA) modeling. NICA model calculations and XAFS results showed that bidentate and monodentate complexation occurred for Cu binding to HA. The site-type-specific thermodynamic parameters obtained by combining ITC measurements and NICA calculations revealed that copper binding to deprotonated carboxylic-type sites was entropically driven and that to deprotonated phenolic-type sites was driven by entropy and enthalpy. Copper binding to HA largely depended on the site-type and coordination environment, but the thermodynamic binding mechanisms for Cu binding to the specific site-types were similar for the different HAs studied. By comparing the site-type-specific thermodynamic parameters of HA-Cu complexation with those of low molar mass organic acids, the Cu coordination could be further specified. Bidentate carboxylic-Cu complexes made the dominating contributions to Cu binding to HA. The present study not only yields molecular-scale mechanisms of ion binding to carboxylic- and phenolic-type sites of HA but also provides the new insight that the universal nature of site-type-specific thermodynamic data enables quantitative estimation of the binding structures of heavy metal ions to humic substances.

Enzymes in the soil are vital for assessing heavy metal soil pollution. Although the presence of heavy metals is thought to change the soil enzyme system, the distribution of enzyme activities in heavy metal polluted-soil is still unknown. For the first time, using soil zymography, we analyzed the distribution of enzyme activities of alfalfa rhizosphere and soil surface in the metal-contaminated soil. The results showed that the growth of alfalfa was significantly inhibited, and an impact that was most pronounced in seedling biomass and chlorophyll content. Catalase activity (CAT) in alfalfa decreased with increasing heavy metal concentrations, while malondialdehyde (MDA) content continually increased. The distribution of enzyme activities showed that both phosphatase and β-glucosidase activities were associated with the roots and were rarely distributed throughout the soil. In addition, the total hotspot areas of enzyme activities were the highest in extremely heavy pollution soil. The hotspot areas of phosphatase were 3.4%, 1.5% and 7.1% under none, moderate and extremely heavy pollution treatment, respectively, but increased from 0.1% to 0.9% for β-glucosidase with the increasing pollution levels. Compared with the traditional method of enzyme activities, zymography can directly and accurately reflect the distribution and extent of enzyme activity in heavy metals polluted soil. The results provide an efficient research method for exploring the interaction between enzyme activities and plant rhizosphere.

Changes in plants and soils during natural succession have been evaluated, but little is known about the effects of succession on the activities of soil microbes and their interactions with soil erodibility. We conducted a field study on the Chinese Loess Plateau, typical of this semiarid area, to determine the effect of secondary succession on the stability of soil structure against erosion and on the composition of soil fungal communities. Characteristics of plant, soil, and fungal communities were assessed across a 30-year chronosequence of grassland developed from abandoned cropland. The diversity and composition of the fungal communities were determined using high-throughput sequencing of the internal transcribed spacer. Six grasslands were selected to represent different successional age-classes: 0 (cropland), 5, 10, 15, 20, and 30 y. Short-term decreases (initial 5 y) in the amounts of soil organic carbon, total nitrogen, available phosphorus, and fungal biomass and in fungal diversity had returned to original levels (i.e. cropland) within 15 y and were much higher after continued succession. Abandoning cropland for succession caused the soil erodibility (K) decrease and the aboveground coverage, soil nutrient levels, content of larger (> 5 mm) water-stable aggregate, mean aggregate weight diameter, and diversity of the fungal communities improvement including arbuscular mycorrhizas, ectomycorrhizas (EMF), and saprotrophs. The fungal communities were dominated by Ascomycota, Zygomycota, Basidiomycota, and Glomeromycota during the succession. The successional patterns of the plant and fungal communities were similar, although distinct fungal communities were not observed in the two initial stages, suggesting that fungal succession may develop more slowly than plant succession. Plant-roots biomass, EMF, and soil organic-carbon content accounted for most of the variation of soil erodibility (28.6, 19.5, and 11.8%, respectively), indicating their importance in shaping soil structure to prevent erosion. Our results demonstrated that abandoning cropland for natural succession could decrease soil erodibility and increase fungal diversity. EMF plays an important role in soil stability against erosion in the Loess Plateau. Abandoning cropland for natural succession should be recommended for alleviating soil erosion and improving the degraded soils in this area. This article is protected by copyright. All rights reserved.

Soil drought caused by climatic change and/or poor land management in arid and semi-arid regions are seldom recognised due to a lack of comparative data on soil moisture (SM) in soil profiles. This lack of information endangers the sustainability of these fragile ecosystems. The current study assessed spatial-temporal variations of soil drought, as indicated by dried soil layers (DSL), at a watershed scale, and tested the hypothesis that soil drought in deep profiles is controlled by the combined effects of meteorological processes, land use, and topography. We measured SM to a depth of 500 cm on 20 occasions at 73 locations from 2013 to 2016 at a small watershed on the Chinese Loess Plateau (CLP). We also collected data on possible environmental factors including meteorological variables, land use, topographical elements, and soil properties. The DSLs occurred at > 90% of the sampling sites within the watershed, and the spatially and temporally averaged DSL formation depth (DSLFD), DSL thickness (DSLT) and soil water content within the DSL (DSL-SWC) were 125 cm, 257 cm, and 10.4%, respectively. This suggests that 51.4% of the 500-cm-profile is drying out below 125 cm. The DSLFD, DSLT and DSL-SWC demonstrated a moderate degree of variability (20%

Variations in soil microbial metabolism currently represent one of the greatest areas of uncertainty with regard to soil nutrient cycles and the control of terrestrial carbon (C) and nitrogen (N) loss and are poorly understood in agricultural ecosystems with intensive farming practices. In this study, extracellular enzymatic stoichiometry models and quantitative PCR techniques were used to examine microbial metabolic limitation and its relationship with N-cycling gene expression in semi-arid agricultural ecosystems considering four N fertilization levels (N 0, N 100, N 250, and N 400 kg N ha−1) and two agronomic strategies (film mulching and no mulching). Film mulching increased microbial C limitation (reflecting microbial C metabolism size; 0.189 of the total effects), while very small effects on microbial phosphorus (P) limitation were observed (-0.007 of the total effects). N fertilization increased the microbial demand for P (microbial P limitation; 0.504 of the total effects). Increased microbial C metabolism was mainly attributed to increased soil moisture content after film mulching, which enhanced microbial decomposition of organic C (high C-acquiring enzyme activities). Changes in nutrient stoichiometry and the increase in N availability due to N fertilization were largely responsible for increased microbial P limitation. Furthermore, microbial P limitation negatively affected the abundance of AOA amoA, AOB amoA (involved in nitrification), nirK, nirS, nosZ (involved in denitrification) genes, strongly inhibiting nitrification and denitrification potential (-0.743 and -0.761 of the total effects, respectively). The present results suggest that agricultural ecosystems with film mulching are conducive to organic residue decomposition, while appropriate P limitation under N fertilization could reduce the loss of N due to nitrification and denitrification in soil. This study highlights the importance of elemental stoichiometry-driven microbial metabolic variation in understanding soil nutrient cycles and optimizing agricultural practices.

Article history:Received 29 November 2013Received in revised form 11 August 2014Accepted 12 August 2014Available online 22 August 2014Editor: J. FeinKeywords:EPSAdsorptionCu(II)ATR-FTIRITCXAFS Biosorption mechanisms of Cu(II) by extracellular polymeric substances (EPS) from Bacillus subtilis were inves-tigated using a combination of batch experiments, attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy, isothermal titration calorimetry (ITC), and X-ray absorption fine structure (XAFS) spectros-copy.Athreediscretesitenon-electrostaticmodelfitthepotentiometrictitrationdatabest,withthepK a valuesof4.12 ± 0.12, 6.60 ± 0.06, and 9.09 ± 0.03, and site concentrations of 4.81 ± 0.62 × 10 −3 , 2.16 ± 0.14 × 10 ,and2.87±0.21×10 −3 molpergramdrymassofEPS,respectively.TheATR–FTIRresultsconfirmedthepresenceofthe functional groups with above pK a valueswithin the EPSmolecules, and indicatedthat Cu(II) binding ontothe EPS involves either phosphoryl or carboxyl sites, or both. The calculated enthalpies and entropies of Cu(II)adsorption onto EPS suggest that Cu(II) binds with anionic oxygen-bearing ligands and forms inner-spherecomplexes with the EPS functional groups. The XAFS results were consistent with inner-sphere binding ofCu(II) by carboxyl sites with 2.03 C atoms at distance of 2.96 A in the second shell, which further suggests thatthe carboxyl groups are the dominant sites for Cu(II) adsorption by EPS at pH 5.0, and that a five-memberedchelate ring structure is the most likely binding environment for Cu(II) bound to EPS. The molecular bindingmechanisms obtained in this study will add fundamental knowledge of understanding the fate of heavy metalsin natural environments.© 2014 Published by Elsevier B.V.

Legume–rhizobium symbiosis plays an important role in agriculture and ecological restoration. However, the regulatory mechanisms of rhizobium in alleviating heavy metal stress through the biochemical response of plant-soil system is limited. In this study, alfalfa was inoculated with a copper (Cu)-resistant rhizobium, and its effect on plant growth and the spatial distribution of phosphatase in the rhizosphere under Cu stress was assessed. Our results showed that rhizobium inoculation alleviated Cu-induced growth inhibition, and increased the nitrogen and phosphorus content in alfalfa tissues. Moreover, inoculated plants had a higher Cu uptake than non-inoculated plants, with a much higher increase in the roots than in the shoots; thus, inoculation with rhizobium was shown to decrease the transfer coefficient and promote Cu phytostabilization. The zymograms illustrated that the distribution of phosphatase activities was associated with the presence of roots. Compared with the non-inoculated treatment, the rhizobium inoculation increased the hotspot areas of phosphatase by 26.1% and 39.3% at the Cu 0 and Cu 800 treatments, respectively. In addition, the available phosphorus in the soil showed negative correlations with soil phosphatase activity (p

Long non-coding RNAs (lncRNAs) were found to widely exist in eukaryotes and play important roles in most biological processes. Trees belonging to Populus spp. are economically important because of their fast-growing characteristics and wide use for wood, pulp, paper, and fuel. However, the prevalence of leaf rust disease caused by Melampsora spp. on some species of Populus severely affects their growth and decreases wood production. Therefore, the identification and characterization of lncRNAs involved in the defense of Melampsora spp. would not only help us to understand plant-pathogen interactions but also provide genetic elements for producing disease-resistant poplars. In the present study, an RNA-Seq transcriptome analysis was performed for poplar leaves in response to early infection by M. larici-populina. The hybrid variety “NL895” (P. × euramericana) was used as plant material. A total of 3994 lncRNAs were identified by mining the RNA-Seq data. The identified lncRNAs had lower abundance, fewer exons, and shorter lengths when compared to protein-coding genes. In addition, 53 lncRNAs were differentially expressed between treatments and controls. Two of these 53 lncRNAs were predicted to compete to be the target of miRNAs. Moreover, the differentially expressed lncRNAs were found to be preferentially located in close proximity to the protein-coding genes they co-expressed. In this study, we were able to show that lncRNAs may play important roles in plant-pathogen interactions in the poplar. This study also improves our understanding of how plants deploy their defense system when encountering biotic stresses.

Grapevine is an important fruit crop that has undergone a long history of evolution. Analysis of the whole genome sequence of grapevine has revealed presence of an early palaeo-hexaploid along with three complements. Thus, gene duplication and genome expansion are common in this genome. In this study, we identified 17,922 duplicated genes in the whole grapevine genome. Among these, 2,039; 628; 1,428; 722; and 2,942 were identified respectively as produced by genome-wide, tandem, proximal, retrotransposed, and DNA-based transposed duplications. Analyses of the evolutionary patterns for different types of duplication using non-synonymous and synonymous substitution rates uncovered a series of underlying rules. Thereafter, all the grapevine genes were classified into families, and the contributions of different types of duplication to the expansion of large families were revealed. No duplication type was solely responsible for the formation of any large gene family, but some families showed enrichment of a special type of duplication. On the basis of this study, we believe that uncovering the underlying rules for gene duplications, expansions of gene families, and their evolutionary styles will contribute significantly to a comprehensive understanding of the features of the grapevine genome.

Binding of lead (Pb) to soil fulvic acid (JGFA), soil humic acids (JGHA, JLHA), and lignite-based humic acid (PAHA) was investigated through binding isotherms and XAFS. Pb binding to humic substances (HS) increased with increasing pH and decreasing ionic strength. The NICA-Donnan model described Pb binding to the HS satisfactorily. The comparison of the model parameters showed substantial differences in median Pb affinity constants among JGFA, PAHA, and the soil HAs. Milne's "generic" parameters did not provide an adequate prediction for the soil samples. The Pb binding prediction with generic parameters for the soil HAs was improved significantly by using the value n(Pb1) = 0.92 instead of the generic value n(Pb1) = 0.60. The n(Pb1)/n(H1) ratios obtained were relatively high, indicating monodentate Pb binding to the carboxylic-type groups. The nPb2/nH2 ratios depended somewhat on the method of optimization, but the values were distinctly lower than the n(Pb1)/nH1 ratios, especially when the optimization was based on Pb bound vs log [Pb(2+)]. These low values indicate bidentate binding to the phenolic-type groups at high Pb concentration. The NICA-Donnan model does not consider bidentate binding of Pb to a carboxylic- and a phenolic-type group. The EXAFS results at high Pb loading testified that Pb was bound in bidentate complexes of one carboxylic and one phenolic group (salicylate-type) or two phenolic groups (catechol-type) in ortho position.

Ethylene (ET) is a gaseous plant hormone that plays essential roles in biotic and abiotic stress responses in plants. However, the role of ET in cold tolerance varies in different species. This study revealed that low temperature promotes the release of ET in grapevine. The treatment of exogenous 1-aminocyclopropane-1-carboxylate increased the cold tolerance of grapevine. By contrast, the application of the ET biosynthesis inhibitor aminoethoxyvinylglycine reduced the cold tolerance of grapevine. This finding suggested that ET positively affected cold stress responses in grapevine. The expression of VaERF057, an ET signaling downstream gene, was strongly induced by low temperature. The overexpression of VaERF057 also enhanced the cold tolerance of Arabidopsis. Under cold treatment, malondialdehyde content was lower and superoxide dismutase, peroxidase, and catalase activities were higher in transgenic lines than in wild-type plants. RNA-Seq results showed that 32 stress-related genes, such as CBF1-3, were upregulated in VaERF057-overexpressing transgenic line. Yeast one-hybrid results further demonstrated that VaERF057 specifically binds to GCC-box and DRE motifs. Thus, VaERF057 may directly regulate the expression of its target stress-responsive genes by interacting with a GCC-box or a DRE element. Our work confirmed that ET positively regulates cold tolerance in grapevine by modulating the expression of VaERF057.

The adsorption to goethite of extracellular polymeric substances (EPS) isolated from Pseudomonas putida was investigated using batch adsorption experiments, electrophoretic mobility (EM) measurements, Fourier transform infrared (FTIR) spectroscopy, isothermal titration calorimetry (ITC) and X-ray absorption fine structure (XAFS) spectroscopy. The adsorption of EPS decreased the point of zero charge of goethite from 7.6 to 3.2, suggesting the formation of negatively charged inner-sphere surface complexes. The adsorption isotherms of EPS on goethite conformed to the Langmuir equation. The adsorption energy constant (K) of EPS on goethite was in the sequence of EPS phosphate-containing moieties > nitrogen-containing moieties > carbon-containing moieties, indicating those containing phosphate were the most strongly adsorbed. FTIR showed ligand exchange of phosphate groups of EPS with surface hydroxyls on goethite. Extended X-ray absorption fine structure (EXAFS) and X-ray absorption near edge structure (XANES) spectroscopy results further demonstrated phosphate groups of EPS can form monodentate inner-sphere complexes at lower pH 3.0, while form bidentate inner-sphere complexes at higher pH 9.0. The oxidation state of iron in goethite was not changed after the reaction with EPS at different pH values. The information obtained in this study is of fundamental significance for the understanding of the interaction mechanisms between bacteria and minerals in soil and aquatic environments.

Flavonoid composition and concentration were investigated in 12 different tissues of 'Ti-1' lotus (Nelumbo nucifera) by high performance liquid chromatography equipped with photodiode array detection tandem electrospray ionization mass spectrometry (HPLC-DAD-ESI-MS(n)). A total of 20 flavonoids belonging to six groups (myricetin, quercetin, kaempferol, isohamnetin, diosmetin derivatives) were separated and identified. Myricetin 3-O-galactoside, myricetin 3-O-glucuronide, isorhamnetin 3-O-glucuronide and free aglycone diometin (3',5,7-trihydroxy-4'-methoxyflavone) were first reported in lotus. Flavonoid composition varied largely with tissue type, and diverse compounds (5-15) were found in leaf and flower stalks, flower pistils, seed coats and embryos. Flower tissues including flower petals, stamens, pistils, and, especially, reproductive tissue fruit coats had more flavonoid compounds (15-17) than leaves (12), while no flavonoids were detectable in seed kernels. The flavonoid content of seed embryos was high, 730.95 mg 100g(-1) DW (dry weight). As regards the other tissues, mature leaf pulp (771.79 mg 100 g(-1) FW (fresh weight)) and young leaves (650.67 mg 100 g(-1) FW) had higher total flavonoid amount than flower stamens (359.45 mg 100 g(-1) FW) and flower petals (342.97 mg 100g(-1) FW), while leaf stalks, flower stalks and seed coats had much less total flavonoid (less than 40 mg 100 g(-1) FW).

The extraction protocol of flavonoids from lotus (Nelumbo nucifera) leaves was optimized through an orthogonal design. The solvent was the most important factor comparing solvent, solvent:tissue ratio, extraction time, and temperature. The highest yield of flavonoids was achieved with 70% methanol-water and a solvent:tissue ratio of 30:1 at 4 °C for 36 h. The optimized analytical method for HPLC was a multi-step gradient elution using 0.5% formic acid (A) and CH₃CN containing 0.1% formic acid (B), at a flow rate of 0.6 mL/min. Using this optimized method, thirteen flavonoids were simultaneously separated and identified by high performance liquid chromatography coupled with photodiode array detection/electrospray ionization mass spectrometry (HPLC/DAD/ESI-MS(n)). Five of the bioactive compounds are reported in lotus leaves for the first time. The flavonoid content of the leaves of three representative cultivars was assessed under the optimized extraction and HPLC analytical conditions, and the seed-producing cultivar 'Baijianlian' had the highest flavonoid content compared with rhizome-producing 'Zhimahuoulian' and wild floral cultivar 'Honglian'.

Cyanobacteria are promising biosorbent for heavy metals in bioremediation. Although sequestration of metals by cyanobacteria is known, the actual mechanisms and ligands involved are not very well understood. The binding characteristics of Cu(II) and Cd(II) by the cyanobacterium Spirulina platensis were investigated using a combination of chemical modifications, batch adsorption experiments, Fourier transform infrared (FTIR) spectroscopy and X-ray absorption fine structure (XAFS) spectroscopy. A significant increase in Cu(II) and Cd(II) binding was observed in the range of pH 3.5-5.0. Dramatical decrease in adsorption of Cu(II) and Cd(II) was observed after methanol esterification of the nonliving cells demonstrating that carboxyl functional groups play an important role in the binding of metals by S. platensis. The desorption rate of Cu(II) and Cd(II) from S. platensis surface was 72.7-80.7% and 53.7-58.0% by EDTA and NH(4)NO(3), respectively, indicating that ion exchange and complexation are the dominating mechanisms for Cu(II) and Cd(II) adsorption. XAFS analysis provided further evidence on the inner-sphere complexation of Cu by carboxyl ligands and showed that Cu is complexed by two 5-membered chelate rings on S. platensis surface.

The effect of extracellular polymeric substances (EPS) of Gram-positive Bacillus subtilis and Gram-negative Pseudomonas putida on Cu(II) adsorption was investigated using a combination of batch adsorption, potentiometric titrations, Fourier transform infrared spectroscopy. Both the potentiometric titrations and the Cu(II) adsorption experiments indicated that the presence of EPS in a biomass sample significantly enhance Cu(II) adsorption capacity. Surface complexation modeling showed that the pKa values for the three functional groups (carboxyl, phosphate and hydroxyl) were very similar for untreated and EPS-free cells, indicating no qualitative difference in composition. However, site concentrations on the untreated cell surface were found to be significantly higher than those on the EPS-free cell surface. Infrared analysis provided supporting evidence and demonstrated that carboxyl and phosphate groups are responsible for Cu(II) adsorption on the native and EPS-free cells.

Equilibrium adsorption experiments, isothermal titration calorimetry and potentiometric titration techniques were employed to investigate the adsorption of Cu(II) by extracellular polymeric substances (EPS) extracted from Pseudomonas putida X4, minerals (montmorillonite and goethite) and their composites. Compared with predicted values of Cu(II) adsorption on composites, the measured values of Cu(II) on EPS-montmorillonite composite increased, however, those on EPS-goethite composite decreased. Potentiometric titration results also showed that more surface sites were observed on EPS-montmorillonite composite and less reactive sites were found on EPS-goethite composite. The adsorption of Cu(II) on EPS molecules and their composites with minerals was an endothermic reaction, while that on minerals was exothermic. The positive values of enthalpy change (Delta H) and entropy change (DeltaS) for Cu(II) adsorption on EPS and mineral-EPS composites indicated that Cu(II) mainly interacts with carboxyl and phosphoryl groups as inner-sphere complexes on EPS molecules and their composites with minerals.

In this study, Fourier transform infrared spectroscopy, potentiometric titration, along with sorption experiments using chemically modified bacteria were conducted to compare the behavior of Gram-positive Bacillus thuringiensis and Gram-negative Escherichia coli as sorbents for Cu 2+ and Cd 2+ ions. The IR spectra showed that there were obvious changes connected with the–COOH groups for Cu(II)- and Cd(II)-loaded B. thuringiensis . A three site non-electrostatic model provides an excellent fit to the titration curves of both E. coli and B. thuringiensis with the first corresponding p K a values of 4.16 ± 0.18 and 3.30 ± 0.24, respectively, implying that B. thuringiensis contains more carboxyl groups than E. coli . Chemical modification and metal sorption experiments further confirmed that carboxyl groups may play a more important role in the binding Cd(II) and Cu(II) for B. thuringiensis than for E. coli , which could be attributed to the higher concentration of carboxyl sites of B. thuringiensis than E. coli . These results show that the main chemical functional group responsible for the binding of metal ions on B. thuringiensis and E. coli is obviously different because of the different composition and structure of bacterial cell walls. The concentrations of the chemical functional groups on bacterial cell walls are regarded to govern their role in the binding of metal ions and affect the affinity between the bacterial cells and metal ions.

A GIS approach and HJ-1B images were employed to determine the effect of landscape structure on aerosol optical depth (AOD) patterns. Landscape metrics, fractal analysis and contribution analysis were proposed to quantitatively illustrate the impact of land use on AOD patterns. The high correlation between the mean AOD and landscape metrics indicates that both the landscape composition and spatial structure affect the AOD pattern. Additionally, the fractal analysis demonstrated that the densities of built-up areas and bare land decreased from the high AOD centers to the outer boundary, but those of water and forest increased. These results reveal that the built-up area is the main positive contributor to air pollution, followed by bare land. Although bare land had a high AOD, it made a limited contribution to regional air pollution due to its small spatial extent. The contribution analysis further elucidated that built-up areas and bare land can increase air pollution more strongly in spring than in autumn, whereas forest and water have a completely opposite effect. Based on fractal and contribution analyses, the different effects of cropland are ascribed to the greater vegetation coverage from farming activity in spring than in autumn. The opposite effect of cropland on air pollution reveals that green coverage and human activity also influence AOD patterns. Given that serious concerns have been raised regarding the effects of built-up areas, bare land and agricultural air pollutant emissions, this study will add fundamental knowledge of the understanding of the key factors influencing urban air quality.

The methylation of mercury (Hg) is of great concern as methylmercury (MeHg), the most toxic species, is produced. This study examined the possibilities of tert-butyl compounds (tert-butyl alcohol (TBA) and tert-butyl hydroperoxide (TBH)) and other alcohols serving as methyl donors for Hg photo-methylation under light irradiation. The yield of MeHg varied among the methyl donors, and it was also significantly influenced by salinity and pH. MeHg could be generated in the presence of TBH under visible light irradiation. The hydroxyl radical (OH) was found to promote MeHg production at low levels, but degrade MeHg in excess. The photo-production of MeHg was tentatively proposed via the complexation of Hg and methyl donors, the formation of an intermediate (O(Hg)C(CH3)3), and the intramolecular methyl transfer from methyl donors to Hg. This study implicates photoreactions between Hg and organic pollutants in understanding the fate and transformation of Hg in the aquatic environment.

Background QTLs controlling individual sugars and acids (fructose, glucose, malic acid and tartaric acid) in grape berries have not yet been identified. The present study aimed to construct a high-density, high-quality genetic map of a winemaking grape cross with a complex parentage (V. vinifera × V. amurensis) × ((V. labrusca × V. riparia) × V. vinifera), using next-generation restriction site-associated DNA sequencing, and then to identify loci related to phenotypic variability over three years. Results In total, 1 826 SNP-based markers were developed. Of these, 621 markers were assembled into 19 linkage groups (LGs) for the maternal map, 696 for the paternal map, and 1 254 for the integrated map. Markers showed good linear agreement on most chromosomes between our genetic maps and the previously published V. vinifera reference sequence. However marker order was different in some chromosome regions, indicating both conservation and variation within the genome. Despite the identification of a range of QTLs controlling the traits of interest, these QTLs explained a relatively small percentage of the observed phenotypic variance. Although they exhibited a large degree of instability from year to year, QTLs were identified for all traits but tartaric acid and titratable acidity in the three years of the study; however only the QTLs for malic acid and β ratio (tartaric acid-to-malic acid ratio) were stable in two years. QTLs related to sugars were located within ten LGs (01, 02, 03, 04, 07, 09, 11, 14, 17, 18), and those related to acids within three LGs (06, 13, 18). Overlapping QTLs in LG14 were observed for fructose, glucose and total sugar. Malic acid, total acid and β ratio each had several QTLs in LG18, and malic acid also had a QTL in LG06. A set of 10 genes underlying these QTLs may be involved in determining the malic acid content of berries. Conclusion The genetic map constructed in this study is potentially a high-density, high-quality map, which could be used for QTL detection, genome comparison, and sequence assembly. It may also serve to broaden our understanding of the grape genome. Electronic supplementary material The online version of this article (doi:10.1186/s12870-015-0428-2) contains supplementary material, which is available to authorized users.

High and low resveratrol (Res) contents in two cultivars are correlated with the expression abundance of Myb14 , which could directly activate transcriptional expression of stilbene synthase gene ( STS ). Resveratrol (3,5,4′-trihydroxystilbene) is one of the natural polyphenols produced by secondary metabolism in some plants. Stilbene synthase (STS) is the key enzyme for the final step of precursor formation of resveratrol (Res) in grapevines. In this study, we found that Res contents in ripe berry skin were completely different in two grape cultivars, namely, ‘Z168’ (Vitis monticola × Vitis riparia) with high-Res and ‘Jingzaojing’ (Vitis vinifera) with low-Res. Moreover, the level of expression of STS gene was higher in the ripe berry skin of ‘Z168’ than in that of ‘Jingzaojing’. To further investigate the underlying mechanisms, we conducted a co-expression analysis through transcriptomic data. We confirmed that Myb14, an R2R3 Myb transcription factor, is the direct regulator of STS by binding to Box-L5 motif. Moreover, the expression pattern of Myb14 is associated with the variation of Res content. To test this prediction, we conducted a number of experiments in vivo and in vitro. The expression patterns of Myb14 and STS in grapevine leaves were identical under a series of stimulus. Myb14 showed higher expression in the ripe berry skin of ‘Z168’ than in that of ‘Jingzaojing’. Yeast one-hybrid assay indicated that grapevine Myb14 could interact with the promoter of STS in vitro, and the transient overexpression of Myb14 promoted the expression of STS. Furthermore, co-expressing 35S::Myb14 in transgenic Arabidopsis could activate GUS expression promoted by STS promoter. Thus, Myb14 is the direct activator of STS, and its expression pattern is associated with Res content variation in grapes.

Background WRKY transcription factors are one of the largest families of transcriptional regulators in plants. WRKY genes are not only found to play significant roles in biotic and abiotic stress response, but also regulate growth and development. Grapevine (Vitis vinifera) production is largely limited by stressful climate conditions such as cold stress and the role of WRKY genes in the survival of grapevine under these conditions remains unknown. Results We identified a total of 59 VvWRKYs from the V. vinifera genome, belonging to four subgroups according to conserved WRKY domains and zinc-finger structure. The majority of VvWRKYs were expressed in more than one tissue among the 7 tissues examined which included young leaves, mature leaves, tendril, stem apex, root, young fruits and ripe fruits. Publicly available microarray data suggested that a subset of VvWRKYs was activated in response to diverse stresses. Quantitative real-time PCR (qRT-PCR) results demonstrated that the expression levels of 36 VvWRKYs are changed following cold exposure. Comparative analysis was performed on data from publicly available microarray experiments, previous global transcriptome analysis studies, and qRT-PCR. We identified 15 VvWRKYs in at least two of these databases which may relate to cold stress. Among them, the transcription of three genes can be induced by exogenous ABA application, suggesting that they can be involved in an ABA-dependent signaling pathway in response to cold stress. Conclusions We identified 59 VvWRKYs from the V. vinifera genome and 15 of them showed cold stress-induced expression patterns. These genes represented candidate genes for future functional analysis of VvWRKYs involved in the low temperature-related signal pathways in grape.

KEY MESSAGE : Genome-wide identification of grapevine NAC domain genes and investigation of their chromosome locations, gene structures, duplication, evolution, phylogeny and expression profiles. Grapevine is a widely used fruit crop. NAC (NAM, ATAF1/2 and CUC2) domain genes are plant-specific transcription factors (TFs) that comprise a conserved NAM domain in the N-terminus. Members of this gene family have been reported to contribute to plant development. During this study, 74 NAC genes were identified from 12× assembled grapevine genomic sequences. The duplication patterns, genomic structures and phylogeny of these 74 grapevine NAC genes were investigated. To understand the roles of VvNAC during grapevine development, their expression profiles in different tissues including leaf, tendril, inflorescence, stem, root and veraison berry skin were tested using quantitative real-time PCR. Analysis revealed expression diversity of various VvNAC genes among different grapevine tissues. To identify candidate grapevine NAC genes with a role in response to stress, publicly available microarray data were obtained to calculate their expression change under abiotic and biotic treatments, with a number of VvNAC genes displaying up-regulation after stress induction. Therefore, this study has uncovered more knowledge relating to the gene structures, chromosome organizations, evolution, expression profiles and functions of VvNAC genes.

The role of extracellular polymeric substances (EPS) in Cd adsorption by Bacillus subtilis and Pseudomonas putida was investigated using a combination of batch adsorption experiments, potentiometric titrations, Fourier transform infrared spectroscopy (FTIR). An increased adsorption capacity of Cd was observed for untreated bacteria relative to that for EPS-free bacteria. Surface complexation modeling of titration data showed the similar pKa values of functional groups (carboxyl, phosphate and hydroxyl) between untreated and EPS-free bacteria. However, site concentrations on the untreated bacteria were found to be higher than those on the EPS-free bacteria. FTIR spectra also showed that no significant difference in peak positions was observed between untreated and EPS-free bacteria and carboxyl and phosphate groups were responsible for Cd adsorption on bacterial cells. The information obtained in this study is of fundamental significance for understanding the interaction mechanisms between heavy metals and biofilms in natural environments.

In order to have a better understanding of the interactions of heavy metals with bacteria and minerals in soil and associated environments, isothermal titration calorimetry (ITC), potentiometric titration and equilibrium sorption experiments were conducted to investigate the adsorption behavior of Cu(II) by Bacillus thuringiensis, Pseudomonas putida and their composites with minerals. The interaction of montmorillonite with bacteria increased the reactive sites and resulted in greater adsorption for Cu(II) on their composites, while decreased adsorption sites and capacities for Cu(II) were observed on goethite-bacteria composites. A gram-positive bacterium B. thuringiensis played a more important role than a gram-negative bacterium P. putida in determining the properties of the bacteria-minerals interfaces. The enthalpy changes (DeltaH(ads)) from endothermic (6.14 kJ mol(-1)) to slightly exothermic (-0.78 kJ mol(-1)) suggested that Cu(II) is complexed with the anionic oxygen ligands on the surface of bacteria-mineral composites. Large entropies (32.96-58.89 J mol(-1) K(-1)) of Cu(II) adsorption onto bacteria-mineral composites demonstrated the formation of inner-sphere complexes in the presence of bacteria. The thermodynamic data implied that Cu(II) mainly bound to the carboxyl and phosphoryl groups as inner-sphere complexes on bacteria and mineral-bacteria composites.