Non-invasive microelectrode cadmium flux measurements reveal the spatial characteristics and real-time kinetics of cadmium transport in hyperaccumulator and nonhyperaccumulator ecotypes of Sedum alfredii

This study aims to determine the spatial characteristics and real-time kinetics of cadmium transport in hyperaccumulator (HE) and non hyperaccumulator (NHE) ecotypes of Sedum alfredii using a non-invasive Cd-selective microelectrode. Compared with the NHE S. alfredii, the HE S. alfredii showed a higher Cd influx in the root apical region and root hair cells, as well as a significantly higher Cd efflux in the leaf petiole after root pre-treatment with cadmium chloride (CdCl₂). Thus, HE S. alfredii has a higher capability for the translocation of absorbed Cd to the shoot. Moreover, the mesophyll tissues, isolated mesophyll protoplasts, and intact vacuoles from HE S. alfredii exhibited an instantaneous influx of Cd in response to CdCl₂ treatment with mean rates that are markedly higher than those from NHE S. alfredii. Therefore, the hyper-accumulating trait of HE S. alfredii is characterized by the rapid Cd uptake in specific root regions, including the apical region and root hair cells, as well as by the rapid root-to-shoot translocation and the highly efficient Cd-permeable transport system in the plasma membrane and mesophyll cell tonoplast. We suggest that the non-invasive Cd-selective microelectrode is an excellent method with a high degree of spatial resolution for the study of Cd transport at the tissue, cellular, and sub-cellular levels in plants.     

Elevated CO2 improves root growth and cadmium accumulation in the hyperaccumulator Sedum alfredii

Aims

This study examined the effect of elevated CO₂ on plant growth, root morphology and Cd accumulation in S. alfredii, and assessed the possibility of using elevated CO₂ as fertilizer to enhance phytoremediation efficiency of Cd-contaminated soil by S. alfredii.

Methods

Both soil pot culture and hydroponic experiments were carried out to characterize plant biomass, root morphological parameters, and cadmium uptake in S. alfredii grown under ambient (350 μL L^?1) or elevated (800 μL L^?1) CO₂.

Results

Elevated CO₂ prompted the growth of S. alfredii, shoot and root biomass were increased by 24.6–36.7% and 35.0–52.1%, respectively, as compared with plants grown in ambient CO₂. After 10 days growth in medium containing 50 μM Cd under elevated CO₂, the development of lateral roots and root hairs were stimulated, additionally, root length, surface area, root volume and tip number were increased significantly, especially for the finest diameter roots. The total Cd uptake per pot was significantly greater under elevated CO₂ than under ambient CO₂. After 60 d growth, Cd phytoextraction efficiency was increased significantly in the elevated CO₂ treatment.

Conclusions

Results suggested that the use of elevated CO₂ may be a useful way to improve phytoremediation efficiency of Cd-contaminated soil by S. alfredii.     

Root foraging for zinc and cadmium requirement in the Zn/Cd hyperaccumulator plant Sedum alfredii

Positive root response to metals may enhance metal accumulation for greater requirement in hyperaccumulators. The effects of spatially heterogeneous Zn/Cd addition on root allocation, metal accumulation, and growth of the Zn/Cd hyperaccumulator Sedum alfredii were assessed in a pot experiment. Young shoots of S. alfredii were grown with or without supplied Zn/Cd. Two concentrations were used of each metal, and each metal concentration had one homogeneous and two heterogeneous treatments. Growth increased by 1.6–3.2 times with the increasing overall dose of Zn/Cd addition, and shoot biomass was positively correlated with shoot Zn/Cd concentration (P?<?0.001). In all heterogeneous treatments, the plants consistently allocated approximately 90% of root biomass to the metal-enriched patches, and shoot Zn/Cd contents were greater than or similar to those in the homogeneous treatment at each metal concentration. Plants in the control treatment showed symptoms of Zn deficiency, although their shoots had Zn concentrations 100-fold higher than the critical deficiency value for most plants. We conclude that S. alfredii has evolved root foraging mechanisms associated with its greater requirements for Zn/Cd. These results could have important implications both for phytoremediation and for investigation of positive role of Cd in higher plants.     

芘对黑麦草根系几种低分子量有机分泌物的影响

植物根系释放分泌物与有机污染物的植物修复机制密切相关,研究具有有机污染物修复潜力植物在污染胁迫条件下的根系分泌物特征有助于揭示其修复机制.以多环芳烃修复研究中常用的黑麦草为材料(Lolium perenneL.)为材料,在营养液栽培方式下研究了在芘胁迫处理下,黑麦草根系几种低分子量有机物分泌特征.结果表明,黑麦草对芘具有较强的耐受能力,芘胁迫处理下,生物量无显著变化.黑麦草根系分泌的低分子量有机酸主要为草酸.3、6 mg/L和9 mg/L芘胁迫处理下,低分子量有机酸的组成无明显变化,但含量随芘胁迫处理浓度上升而显著增加(P＜0.05);总糖分泌量随着芘胁迫处理浓度升高而呈现先略微上升后下降的趋势,相对高峰值出现在芘胁迫处理浓度3 mg/L,但差异不显著;氨基酸分泌总量随着芘胁迫处理浓度的增加而显著增多,芘胁迫浓度在3、6 mg/L和9 mg/L时,根系氨基酸的分泌总量分别是空白的1.37、2.02倍和2.65倍,但根系分泌的氨基酸组成无明显变化,19种常见氨基酸分泌的数量变化情况却不相同,分泌量总体均随着芘胁迫处理浓度的提高而增加,其中苏氨酸、丝氨酸、脯氨酸、甘氨酸、丙氨酸、亮氨酸、组氨酸和鸟氨酸的分泌量显著增多,差异显著(P＜0.05).     

Identification of root exudates from the Pb-accumulator Sedum alfredii under Pb stresses and assessment of their roles

The Pb-accumulator Sedum alfredii is a good phytoremediation material, and widely used in the phytoremediation research of soils contaminated with Pb. The root exudates from it may be playing a significant role in the process of phytoremediation. In this study, the metabonomics method which based on gas chromatography–mass spectrometry (GC–MS) and pattern recognition analysis was used to identify the remarkable root exudates from S. alfredii under different Pb stresses, including exposure concentrations (0, 10, 50, 200 and 1000?µmol/L) and times (4 and 8 days). And batch extraction experiments were used to verify the roles of these remarkable root exudates. According to the results, 11 metabolites were considered as the remarkable metabolites. Oxalic acid, galactonic acid and glyceric acid can remove Pb in soil, and the removal effect was: oxalic acid?>?galactonic acid?>?glyceric acid. Xylose, glucose and maltose have no removal effect for Pb in soil.     

Root exudates from sunflower (Helianthus annuus L.) show a strong adsorption ability toward Cd(II)

Abstract

Cd(II) adsorption of root exudates from sunflower (Helianthus annuus L.) seedling was investigated by Cd ion-selective electrode, Fourier Transform Infrared spectroscopy, and fluorescence spectroscopy. Root exudates from Helianthus annuus L. had strong adsorption ability toward Cd(II). The adsorption process was pH-dependent and the maximum adsorption capacity, 150.8 mg g^?1, was observed at pH 7.0. Root exudates had pK _a1 at 4.7 for carboxyl and pK _a2 at 9.2 for phenolic, and amino groups. The aliphatic and aromatic (C?H) groups, amide III group, and the C (=O)?O and sulfonate groups were responsible for Cd(II) adsorption. The excitation emission matrix fluorescence spectroscopy showed protein-like substances participated in Cd adsorption and formed strong complexes, with conditional stability constants of 4.70 and 4.32, which is a little lower than that determined by potentiometric methods, 5.13. The strong Cd complexing ability of root exudates implies that root exudates may significantly affect mobility, toxicity, and phytoavailability of Cd. Cd binding of root exudates may be attributed to its interaction with the proteins, polysaccharides, and phenolic compounds in root exudates.     

苯并[α]芘对不同修复潜力羊茅属植物的根系分泌物中几种低分子量有机物的影响

根系分泌物是植物与土壤间进行物质交换和信息传递的重要载体, 是植物响应外界胁迫的重要途径, 也是构成根际微生态特征的关键因素。根系分泌物与有机污染物的植物修复密切相关, 研究胁迫条件下不同修复潜力植物间根系分泌物的释放特征有助于揭示植物修复的内在机制。该文借助根际袋土培试验研究了苯并[α]芘(BaP)胁迫下5种羊茅属(Festuca)植物根系不同生长期(30-70天)几种低分子量有机物的分泌特征。结果表明: 1) BaP浓度在10.25-161.74 mg·kg^-1范围内时, 待试植物能有效地促进土壤中BaP的去除, 其修复潜力依次为苇状羊茅(F. arundinacea) > 草原羊茅(F. chelungkiangnica) ≥ 毛稃羊茅(F. rubra subsp. arctica) ≥ 贫芒羊茅(F. sinomutica) > 细芒羊茅(F. stapfii)。2) BaP胁迫增强了植物根系对可溶性糖的分泌: 随着胁迫强度的增大、胁迫期的延长, 其分泌量变化呈“先升后降”趋势。3) BaP胁迫促进了植物根系低分子量有机酸的释放, 植物的修复潜力越大, 有机酸高峰值出现时的胁迫浓度越高; 组成成分较稳定, 草酸、乙酸、乳酸和苹果酸为主要组分(>97.34%), 在修复潜力较强植物的根系分泌物中检测出微量的反丁烯二酸。4) BaP胁迫对氨基酸种类影响不大, 但对分泌量影响较大。其中, 苏氨酸、丝氨酸、甘氨酸、丙氨酸的分泌量随BaP胁迫强度的增强而剧增; 脯氨酸、羟脯氨酸和天冬氨酸近乎以加和效应甚至协同效应的形式参与植物对BaP胁迫的应激反应: 参与应激组分的分泌量随胁迫强度的增强而剧增, 植物的修复潜力越强, 参与的组分越多。可见BaP胁迫下, 5种羊茅属植物根系分泌物中几种低分子量有机物的释放特征与植物自身的修复潜力有关: 修复潜力越强, 释放量越多且成分也越复杂, 并呈现出较强的环境适应性及生理可塑性。     

Soil nitrogen and microbial biomass carbon dynamics in native forests and derived agricultural land uses in a humid tropical climate of India

Sedum alfredii, a cadmium (Cd) and zinc (Zn) hyperaccumulator at a mine located in Qu Zhou City, Zhejiang Province, China, can accumulate Cd and Zn exceeding 1,000 and 10,000 mg kg^?1, respectively in its shoot (dry weight) when growing under metal-contaminated habitats. Several strains of bacteria were isolated from the rhizosphere of S. alfredii thriving in different Pb/Zn mines in Hunan Province and Zhejiang Province, China, which can resist high levels of heavy metals. Among the different strains isolated, Burkholderia cepacia showed the highest ability in mobilizing Cd and Zn as well as resisting high concentrations of soluble Zn (500 mg L^?1). The soluble Zn concentration in the medium increased from 13 to 72 and 99% (p?<?0.001) after bacterial inoculation in the medium supplemented with insoluble zinc oxide and zinc carbonate, respectively, while pH dropped from 7 to 2.93. The soluble Cd concentration was also increased from 8 to 96% (p?<?0.001), and pH decreased from 7 to 2.65. Short-chain organic acids were also analyzed and the results indicated that oxalic acid, tartaric acid, formic acid and acetic acid had a significant correlation (p?<?0.001) with the concentrations of Cd and Zn being mobilized during the assay. The present results implicated that certain bacteria associated with metal hyperaccumulators could contribute significantly in mobilizing heavy metals, which would enhance the phytoextraction process.     

Effects of silicon on morphology,ultrastructure and exudates of rice root under heavy metal stress

Soil contamination with toxic heavy metals (such as Cd or Zn) is becoming a serious problem worldwide because of the rapid development of social economy. Silicon plays a substantial role in alleviating heavy metal toxicity in crop plants. In this study, two rice varieties, Feng-Hua-Zhan and Hua-Hang-Si-Miao, were chosen to determine the effects of Si application on root morphological traits, cell structure and exudates of rice roots under Cd and/or Zn stress. Single or combined applications of Cd and Zn resulted in significant reduction of total root length, root surface area, root volume, average root diameter and root activity. However, 1.5 mM Si addition reversed these negative effects. Transmission electron microscopy observations showed that rice root cortex cells were heavily damaged under Cd and/or Zn stress for both two varieties, whereas Si addition resulted in improved cell structure integrity. In addition, lower levels of oxalic, acetic, tartaric, maleic and fumaric acids in root exudates were observed for Feng-Hua-Zhan under Cd and/or Zn stress, but addition of Si increased the acid levels. For Hua-Hang-Si-Miao, heavy metal treatments significantly reduced oxalic and fumaric acid levels and increased acetic, tartaric and maleic acid levels, whereas Si treatment showed opposite results. The above results indicated that Si could ameliorate the toxicity of heavy metals (Cd and Zn) for rice which resulted in improving root traits, cell structure and influencing root exudates.     

不同品种(系)牧草镉修复能力综合评价

该研究选用3种高丹草、2种苏丹草、2种杂交狼尾草、一年生黑麦草和多年生黑麦草共9个牧草品系，采用盆栽试验，设置0(CK)、10(Cd10)、20 mg/kg(Cd20)镉胁迫处理，考察不同品系的生长指标、镉含量和富集指标，运用极差化法和主成分分析建立综合评价模型，通过计算各牧草在模型中的综合得分，筛选出最适宜实地修复的牧草，探究不同品种和不同品系牧草的镉修复能力差异，建立Cd污染土壤植物修复能力评价体系，发掘兼顾富集量和后处理的镉污染土壤植物修复资源。结果表明：(1)镉胁迫会不同程度降低牧草的株高和生物量，其中江苏苏丹草在Cd10和Cd20中的耐胁迫能力均较强；(2)在Cd10和Cd20处理下，一年生和多年生黑麦草均具有较高的地上部和地下部Cd含量，高丹草的3个品系在Cd含量和Cd积累量两方面存在较大差异；(3)在Cd10和Cd20处理下，高丹草和黑麦草的富集指标表现优异，且富集指标在高丹草品系间存在较大差异；(4)2个镉质量分数处理下综合评价得分最高的均是一年生黑麦草，其次为高丹草。研究表明，参试牧草对镉胁迫均有较强的耐受性，但耐受能力和富集能力在品系间存在明显差异，并以高丹草品系间差异最大；一年生黑麦草能耐受20 mg/kg以下镉胁迫，且生物量较小，对镉有较强的浓缩效应，可作为镉污染土壤实地修复的可靠材料；综合评价模型可作为镉污染修复植物资源筛选的可靠方法。     

Organic Materials Could Improve the Phytoremediation Efficiency of Soil Potentially Hazardous Metal by <i>Sedum alfredii</i> Hance

Soil potentially hazardous metal (PHM) is continually attracting public attention worldwide, due to its highly toxic properties and potentially huge damage to human being through food chain. Phytoremediation is an effective and eco-friendly way in remediation technology. A pot experiment was carried out to investigate the effect of different organic materials (biogas residue (BR), mushroom residue (MR), and bamboo-shoot shell (BS)) application on phytoremediation of two PHM-contaminated soils (Fuyang soil as ‘heavily-polluted soil’ and Wenzhou soil as ‘moderately-polluted soil’, respectively) by Sedum alfrecdii Hance. The results indicated: 1) for moderately-polluted soil, the 5% BR treatment had the strongest activation to Cu and Zn, for heavily-polluted soil, 1% BS treatment had the highest activation effect for Cu, Zn, Pb and Cd. 2) the above-ground biomass of Sedum alfredii Hance increased with the addition rate of organic materials. 3) for Cd uptake of Sedum alfredii Hance in moderately-polluted soil, only 1% BS treatment had a better accumulation effect, compared to the control, for Zn element, MR treatments were weaker than the control, while other treatments were better than the control, of which 5% BR, 1% BS and 5% BS accumulated more Zn element by 39.6%, 32.6% and 23.8%, respectively; in heavily-polluted soil, the treatments of 5% BS, 1% BR and 5% BR accumulated more Cd than the control by 12.9%, 12.8% and 6.2%, respectively, the treatments with organic materials addition promoted Zn accumulation in shoots of Sedum alfredii Hance, and the best treatment was 5% BS. Therefore, an appropriate application rate of BS and BR could improve the remediation efficiency for Zn/Cd contaminated soils by Sedum alfredii Hance.     

Enhanced cadmium accumulation in maize roots—the impact of organic acids

Low molecular weight organic acids are important components of root exudates and therefore, knowledge regarding the mechanisms of cadmium (Cd) uptake and distribution within plants under the influence of organic acids, is necessary for a better understanding of Cd behavior in the plant–soil system. In this study, acetic and malic acids increased the uptake of Cd by maize (Zea mays L. cv. TY2) roots and enhanced Cd accumulation in shoots under hydroponic conditions. Concentration-dependent net Cd influx in the presence and absence of organic acids could be resolved into linear and saturable components. The saturable component followed Michaelis–Menten kinetics, which indicated that Cd uptake across the plasma membrane was transporter-mediated. While the K _m values were similar, the V _max values in the presence of acetic and malic acids were respectively 6.0 and 3.0 times that of the control. Zinc transporters were the most probable pathways for Cd accumulation. It was hypothesized that Cd(II)–organic acid complexes associated with the root zone, could decompose and liberate Cd²⁺ for subsequent absorption by maize roots; and that in the layer of the roots or within the root free space, depletion of Cd²⁺ was buffered by the presence of Cd(II)–organic acid complexes. Plant response to elevated Cd levels involved overproduction of organic acids in maize roots as a resistance mechanism to alleviate Cd toxicity.     

可降解螯合剂对镉胁迫下籽粒苋根系形态及生理生化特征的影响

采用盆栽试验研究了可降解螯合剂EDDS和NTA对镉胁迫下籽粒苋(Amaranthus hybridus L.)根系形态及生理生化特征的影响。结果表明:当螯合剂施入10 mg/kg的镉污染土壤后,籽粒苋根系生物量和总长等根系形态指标与对照无显著差异,过氧化物酶(POD)、过氧化氢酶(CAT)活性、谷胱甘肽(GSH)和可溶性蛋白含量显著上升。当螯合剂施入100 mg/kg的镉污染土壤后,籽粒苋根系生物量、总长、表面积、体积及侧根数比对照显著减少了12.30%—23.98%、17.01%—24.90%、41.87%—57.93%、16.46%—32.94%和23.48%—53.35%;EDDS的施入使籽粒苋根系POD、CAT活性、GSH和可溶性蛋白含量显著升高;而NTA施入后,根系中的POD活性比对照降低了4.12%—35.95%,并且CAT活性和可溶性蛋白含量在2 mmol/kg NTA处理下分别显著降低了14.66%—15.79%和26.81%—30.48%;EDDS和NTA施入后,GSH含量比对照显著升高了14.73%—65.65%和28.05%—84.10%。当镉处理浓度分别为10 mg/kg和100 mg/kg时,螯合剂的施入显著增强了籽粒苋根系对镉的吸收,比对照分别增加了40.76%—103.10%和15.03%—49.49%。因此,EDDS和NTA施入镉污染土壤后,通过影响籽粒苋根系形态和生理生化过程以响应重金属镉的胁迫。     

Ectomycorrhizal exudates and pre-exposure to elevated CO2 affects soil bacterial growth and community structure

Ectomycorrhizal fungi produce low molecular weight organic compounds, supporting diverse microbial communities. To link mycorrhizal root exudation directly to bacterial responses, we used Scots pine exudates with (Suillus variegatus and Piloderma fallax) and without mycorrhiza as substrata for forest soil bacteria. Bacterial growth and vitality was monitored, and community composition determined using T-RFLP, cloning and sequencing. We investigated if the amount of organic acids in exudates explained bacterial growth, and whether bacterial communities were influenced by pre-exposure to elevated atmospheric CO₂. We demonstrated functional differences in bacterial growth rates related to CO₂. There was a shift in the bacterial community (e.g. Burkholderia sp. and gamma-proteobacteria) toward organisms better able to rapidly utilize exudates when pine microcosms were pre-exposed to elevated CO₂. Soil bacteria from all treatments tended to grow more abundantly and rapidly in exudates from Piloderma-colonized seedlings, suggesting that the organic acids and/or unidentified compounds present supported greater growth.     

Effects of inoculation of a plant growth promoting rhizobacterium <Emphasis Type="Italic">Burkholderia</Emphasis> sp. D54 on plant growth and metal uptake by a hyperaccumulator <Emphasis Type="Italic">Sedum alfredii</Emphasis> Hance grown on multiple metal contaminated soil

Batch experiments were designed to characterize a multiple metal resistant bacterium Burkholderia sp. D54 isolated from metal contaminated soils in the Dabaoshan Mine in South China, and a follow-up experiment was conducted to investigate the effects of inoculating the isolate on plant growth and metal uptake by Sedum alfredii Hance grown on soils collected from a heavily contaminated paddy field in Daxing County, Guangxi Zhuang Automounous Region, Southwest China. Our experiments showed that strain D54 produced indole acetic acid (IAA), siderophores, 1-aminocyclopropane-1-carboxylate (ACC) deaminase, and solubilizing inorganic phosphate and solubilized insoluble metal bearing minerals. Bacterial inoculation significantly enhanced S. alfredii biomass production, and increased both shoot and root Cd concentration, but induced little variation in root/shoot Pb concentration and shoot Zn concentration. Despite this, the total shoot and root uptake of Cd, Pb and Zn in S. alfredii inoculated with D54 increased greatly compared to the non-inoculated controls. It was concluded that inoculation with strain D54 could help S. alfredii grow better on metal contaminated soils, produce more biomass, and remove more metals from soil, which implies improved efficiency of phytoextraction from metal contaminated soil. The knowledge gained from the present experiments constitutes an important advancement in understanding of the interaction between plant growth-promoting bacteria and hyperaccumulators with regard to plant ability to grow and remove the multiple heavy metals from soils.     

Effects of low pH and aluminum stresses on common beans (Phaseolus vulgaris) differing in low-phosphorus and photoperiod responses

Using common beans differing greatly in the response to photoperiod and low-phosphorus (P) stress, we investigated their responses to acidity and aluminum (Al) toxicity and the relationship between Al tolerance and organic acid exudation under Al or low P stress. A genotype Ginshi was found to be sensitive to low pH treatment. When exposed to pH 4.5, serious curvature in the root tips of cv. Ginshi was observed; however, it was completely corrected by the application of 5 or 10 μmol/L AlCl₃; increasing calcium (Ca) could ameliorate Al toxicity, but it could not correct root curvature at pH 4.5. Common beans showed significant differences in both root growth and Al tolerance, and the varieties from the Andes were more tolerant to Al toxicity than those from the Mesoamerican origin. In the presence of 50 μmol/L AlCl₃, all the common bean genotypes exuded citrate, and a significant difference in the amounts of citrate was observed among genotypes. The genotypes originated in the Mesoamerica tended to release more citrate than other origins in the presence of Al. The P-inefficient genotype DOR364 exuded more citrate than the P-efficient genotype G19833 in the presence of 50 μmol/L AlCl₃, whereas no organic acids were detected in root exudates under low-P stress. A reduction of citrate exudation in the DOR364, but a slight increase of citrate exudation in the G19833, was observed under Al stress after they were exposed to 6-d P starvation. These results suggest that different low-P or Al tolerance in common beans might not be associated with organic acid exudation.