Non-invasive Micro-test Technology Research Articles

A newly identified algicidal bacterium of Pseudomonas fragi YB2: Algicidal compounds and effects

Harmful algal bloom (HAB) is an unresolved existing problem worldwide. Here, we reported a novel algicidal bacterium, Pseudomonas fragi YB2, capable of lysing multiple algal species. To Chlorella vulgaris, YB2 exhibited a maximum algicidal rate of 95.02 % at 120 h. The uniqueness of YB2 lies in its ability to self-produce three algicidal compounds: 2-methyl-1, 3-cyclohexanedione (2-MECHD), N-phenyl-2-naphthylamine, and cyclo (Pro-Leu). The algicidal properties of 2-MECHD have not been previously reported. YB2 significantly affected the chloroplast and mitochondrion, thus decreasing in chlorophyll a by 4.74 times for 120 h and succinate dehydrogenase activity by 103 times for 36 h. These physiological damages disrupted reactive oxygen species and Ca2+ homeostasis at the cellular level, increasing cytosolic superoxide dismutase (23 %), catalase (35 %), and Ca2+ influx. Additionally, the disruption of Ca2+ homeostasis rarely reported in algicidal bacteria–algae interaction was observed using the non-invasive micro-test technology. We proposed a putative algicidal mechanism based on the algicidal outcomes and physiological algicidal effects and explored the potential of YB2 through an algicidal simulation test. Overall, this study is the first to report the algicidal bacterium P. fragi and identify a novel algicidal compound, 2-MECHD, providing new insights and a potent microbial resource for the biocontrol of HAB.

Plant cadmium resistance 2.7 from Brassica napus (BnPCR2.7) improves copper and cadmium tolerance

Copper (Cu) and cadmium (Cd) are highly phytotoxic heavy metals that are widespread contaminants in soil. Plants are efficient at taking up heavy metals, which adversely impacts human health. Therefore, it is important to decrease the accumulation of Cu/Cd in plants to reduce human exposure from the food web. Here, we determined the function of a rapeseed (Brassica napus) Cu/Cd transporter, plant cadmium resistance protein 2.7 (BnPCR2.7), in enhancing Cu/Cd tolerance in seedlings and decreasing the accumulation of Cu/Cd in seeds. A subcellular localization analysis revealed that BnPCR2.7 is localized at the plasma membrane (PM). CRISPR/Cas9-mediated BnPCR2.7 knockout lines and knockdown lines had increased sensitivity to high Cu/Cd than wild-type (WT) plants. In contrast, overexpression of BnPCR2.7 enhanced the adaptation to high Cu/Cd than in WT. Additionally, the Cu/Cd content in the roots of overexpression lines was significantly lower than those of the WT, while the contents in the stems increased. A non-invasive micro-test technology (NMT) assay showed that overexpression plants promoted the efflux of Cu and Cd from the roots. A field trial of rapeseed grown in soil contaminated with Cu or Cd showed that overexpression plants grew and developed better than the WT with higher yields and less Cu/Cd that accumulated in the seeds, while knockout and knockdown lines were contrary to these results. Additionally, when grown in soils contaminated by both Cu and Cd, the content of these heavy metals decreased by 12–20 % and 20–30 %, respectively, in seeds of overexpression lines. Collectively, BnPCR2.7 may promotes resistance to Cu/Cd by efflux pathways. Significantly, it is a candidate genetic resource to increase the resistance to heavy metals and reduce the accumulation of Cu/Cd in rapeseed.

Potassium alleviates fluoride accumulation and enhances fluoride tolerance in Camellia sinensis

Tea plants [Camellia sinensis (L.) O. Kuntze] are hyperaccumulators of fluoride (F), but excessive F will harm the photosynthesis and metabolic activities of tea plants. Potassium (K) is one of the essential macronutrients of tea plants, which can enhance the photosynthesis of tea plants and their stress resistance. However, there are fewer studies on K in the accumulation of F in tea plants. In this study, we set three K treatment concentrations of 0 mM, 5 mM, 10 mM and finally found that when the tea plants were treated with F, exogenously applied 5 mM K treatment increased chlorophyll content, photosynthesis, and antioxidant enzyme activity, and reduced the extent of cellular damage, where starch particles content was reduced and the interstices of the chloroplast grana layer were more compact. After K treatment, the F content of new shoots decreased, and the flow rate of absorption of K by roots under F treatment was significantly increased through the Non-invasive Micro-test Technology (NMT), and 5 mM K treatment could effectively stabilize the membrane potential of roots. In addition, after K treatment, the relative expression of the F exporter CsFEX in tea plants was significantly upregulated, while the stress resistance gene expression of CsHAKs in tea plants was downregulated. Above all, appropriate K application could effectively alleviate the hyperaccumulation and stress effects of F on tea plants.

A plasma membrane-localized transporter remobilizes aleurone layer magnesium for seed germination in rice.

The aleurone layer in cereal grains acts as a major reservoir of essential mineral nutrients, significantly influencing seed germination. However, the molecular mechanism underlying the redistribution of nutrients from the aleurone layer in the germinating seed is still not well understood. Here, in rice, we identified a plasma membrane (PM) localized magnesium transporter, MAGNESIUM RELEASE TRANSPORTER 3 (MGR3), is critical for seed germination. OsMGR3 is predominantly expressed in the aleurone layer cells of endosperm, facilitating magnesium remobilization during germination. Non-invasive Micro-test Technology assay data demonstrated that the loss-of-function of OsMGR3 restrained magnesium efflux from the aleurone layer. In the embryo/endosperm grafting experiment, we observed that the mutation of OsMGR3 in the aleurone layer suppressed the growth and differentiation of the embryo during germination. Furthermore, magnesium fluorescence imaging revealed the osmgr3 mutant seeds showed impaired exportation of aleurone layer-stored magnesium to the embryo, consequently delaying germination. Importantly, we discovered that disrupting OsMGR3 could inhibit pre-harvest sprouting without affecting rice yield and quality. Therefore, the magnesium efflux transporter OsMGR3 in the aleurone layer represents a promising genetic target for future agronomic trait improvement.

Physiological response and molecular mechanism of Quercus variabilis under cadmium stress

Heavy metal pollution is a global environmental problem, and Quercus variabilis has a stronger tolerance to Cd stress than do other species. We aimed to explore the physiological response and molecular mechanisms of Q. variabilis to Cd stress. In this study, the antioxidant enzyme activities of leaves were determined, while the photosynthetic parameters of leaves were measured using Handy PEA, and ion fluxes and DEGs in the roots were investigated using noninvasive microtest technology (NMT) and RNA sequencing techniques, respectively. Cd stress at different concentrations and for different durations affected the uptake patterns of Cd2+ and H+ by Q. variabilis and affected the photosynthetic efficiency of leaves. Moreover, there was a positive relationship between antioxidant enzyme (CAT and POD) activity and Cd concentration. Transcriptome analysis revealed that many genes, including genes related to the cell wall, glutathione metabolism, ion uptake and transport, were significantly upregulated in response to cadmium stress in Q. variabilis roots. WGCNA showed that these DEGs could be divided into eight modules. The turquoise and blue modules exhibited the strongest correlations, and the most significantly enriched pathways were the phytohormone signaling pathway and the phenylpropanoid biosynthesis pathway, respectively. These findings suggest that Q. variabilis can bolster plant tolerance by modulating signal transduction and increasing the synthesis of compounds, such as lignin, under Cd stress. In summary, Q. variabilis can adapt to Cd stress by increasing the activity of antioxidant enzymes, and regulating the fluxes of Cd2+ and H+ ions and the expression of Cd stress-related genes.

Synergistic Effects of Exogenous Nutrient Ions on the Real-Time Cadmium Extraction by an Accumulator

Bidens tripartita L. is a cadmium (Cd) accumulator. However, the real-time influx or efflux of Cd2+ around its root apex has not yet been performed. The object of this experiment was to compare the roles of added ions in solution on dynamic Cd extraction by B. tripartita root tip. Quartz sand was used to grow the seedling of B. tripartite. The Cd concentrations of all samples were determined by using ICP-OES after digestion. The Cd2+ influx around the root apex was measured in vivo, i.e., using non-invasive micro-test technology (NMT). The results showed that the Cd2+ influx was found to be decreased by 35.9%, 43.7%, 20.6%, and 57.5% under 10 μM Cd combined with high content Ca2+, Mg2+, Fe3+, or K+ (16 mM, 8 mM, 0.5 mM, 18 mM, respectively), compared to that under 10 μM Cd stress. But Cd treatments with low content ions with 0.05 mM Fe3+ or 0.5 mM S increased the Cd2+ influx in roots by 20.5% and 34.6%, respectively. It was also found that Cd treatment with high concentrations of Ca2+ or K+ increased the shoot biomass of B. tripartita seedlings. Chl a and b contents were significantly decreased in the Cd treatments with low concentrations of Fe3+ or S compared to those under Cd stress alone, and the dehydrogenase activity of the roots decreased in the treatment of Cd with 0.05 mM Fe3+ or 0.5 mM S. Our results indicate that the addition of 0.05 mM Fe3+ or 0.5 mM S promoted Cd2+ influx and Cd uptake by B. tripartita. Unlike traditional measurement, the Cd2+ movements of three-dimensional space around the B. tripartita root tip had been performed by NMT. It was suggested that the effects of S and Fe3+ on the remediation potential of B. tripartita need to be further researched in the future. The results of this study provided a real-time and micro-dynamic theoretical basis for phytoremediation mechanisms.

The fluoride exporter (CsFEX) regulates fluoride uptake/accumulation in Camellia sinensis under different pH

Fluoride (F) can be absorbed from the environment and hyperaccumulate in leaves of Camellia sinensis without exhibiting any toxic symptoms. Fluoride exporter in C. sinensis (CsFEX) could transport F to extracellular environment to alleviate F accumulation and F toxicity, but its functional mechanism remains unclear. Here, combining with pH condition of C. sinensis growth, the characteristics of CsFEX and mechanism of F detoxification were further explored. The results showed that F accumulation was influenced by various pH, and pH 4.5 and 6.5 had a greater impact on the F accumulation of C. sinensis. Through Non-invasive Micro-test Technology (NMT) detection, it was found that F uptake/accumulation of C. sinensis and Arabidopsis thaliana might be affected by pH through changing the transmembrane electrochemical proton gradient of roots. Furthermore, diverse expression patterns of CsFEX were induced by F treatment under different pH, which was basically up-regulated in response to high F accumulation, indicating that CsFEX was likely to participate in the process of F accumulation in C. sinensis and this process might be regulated by pH. Additionally, CsFEX functioned in the mitigation of F sensitivity and accumulation strengthened by lower pH in Escherichia coli and A. thaliana. Moreover, the changes of H+ flux and potential gradient caused by F were relieved as well in transgenic lines, also suggesting that CsFEX might play an important role in the process of F accumulation. Above all, F uptake/accumulation were alleviated in E. coli and A. thaliana by CsFEX through exporting F-, especially at lower pH, implying that CsFEX might regulate F accumulation in C. sinensis.

A method for separating tonoplast from wheat

Vacuoles account for 90% of plant cell volume and play important roles in maintaining osmotic pressure, storing metabolites and lysosomes, compartmentalizing harmful ions, and storing and reusing minerals. These functions closely relay on the ion channels and transporters located on the tonoplast. The separation of intact vacuoles from plant cells is the key technology utilized in the study of tonoplast-located ion channels and transporters. However, the current vacuole separation methods are available for Arabidopsis and some other dicotyledons but are lacking for monocot crops. In this study, we established a new method for the vacuole separation from wheat mesophyll cells and investigated the transmembrane proton flux of tonoplasts with non-invasive micro-test technology (NMT). Moreover, our study provides a technology for the study of vacuole functions in monocot crops.

Increasing Ca2+ accumulation in salt glands under salt stress increases stronger selective secretion of Na+ in Plumbago auriculata tetraploids.

Under salt stress, recretohalophyte Plumbago auriculata tetraploids enhance salt tolerance by increasing selective secretion of Na+ compared with that in diploids, although the mechanism is unclear. Using non-invasive micro-test technology, the effect of salt gland Ca2+ content on Na+ and K+ secretion were investigated in diploid and tetraploid P. auriculata under salt stress. Salt gland Ca2+ content and secretion rates of Na+ and K+ were higher in tetraploids than in diploids under salt stress. Addition of exogenous Ca2+ increased the Ca2+ content of the salt gland in diploids and is accompanied by an increase in the rate of Na+ and K+ secretion. With addition of a Ca2+ channel inhibitor, diploid salt glands retained large amounts of Ca2+, leading to higher Ca2+ content and Na+ secretion rate than those of tetraploids. Inhibiting H2O2 generation and H+-ATPase activity altered Na+ and K+ secretion rates in diploids and tetraploids under salt stress, indicating involvement in regulating Na+ and K+ secretion. Our results indicate that the increased Na+ secretion rate of salt gland in tetraploids under salt stress was associated with elevated Ca2+ content in salt gland.

Potassium transporter OsHAK9 regulates seed germination under salt stress by preventing gibberellin degradation through mediating OsGA2ox7 in rice.

Soil salinity has a major impact on rice seed germination, severely limiting rice production. Herein, a rice germination defective mutant under salt stress (gdss) was identified by using chemical mutagenesis. The GDSS gene was detected via MutMap and shown to encode potassium transporter OsHAK9. Phenotypic analysis of complementation and mutant lines demonstrated that OsHAK9 was an essential regulator responsible for seed germination under salt stress. OsHAK9 is highly expressed in germinating seed embryos. Ion contents and non-invasive micro-test technology results showed that OsHAK9 restricted K+ efflux in salt-exposed germinating seeds for the balance of K+/Na+. Disruption of OsHAK9 significantly reduced gibberellin 4 (GA4) levels, and the germination defective phenotype of oshak9a was partly rescued by exogenous GA3 treatment under salt stress. RNA sequencing (RNA-seq) and real-time quantitative polymerase chain reaction analysis demonstrated that the disruption of OsHAK9 improved the GA-deactivated gene OsGA2ox7 expression in germinating seeds under salt stress, and the expression of OsGA2ox7 was significantly inhibited by salt stress. Null mutants of OsGA2ox7 created using clustered, regularly interspaced, short palindromic repeat (CRISPR)/CRISPR-associated nuclease 9 approach displayed a dramatically increased seed germination ability under salt stress. Overall, our results highlight that OsHAK9 regulates seed germination performance under salt stress involving preventing GA degradation by mediating OsGA2ox7, which provides a novel clue about the relationship between GA and OsHAKs in rice.

Synergistic effect of silicon availability and salinity on metal adsorption in a common estuarine diatom

Increasing nitrogen and phosphorus discharge and decreasing sediment input have made silicon (Si) a limiting element for diatoms in estuaries. Disturbances in nutrient structure and salinity fluctuation can greatly affect metal uptake by estuarine diatoms. However, the combined effects of Si and salinity on metal accumulation in these diatoms have not been evaluated. In this study, we aimed to investigate how salinity and Si availability combine to influence the adsorption of metals by a widely distributed diatom Phaeodactylum tricornutum. Our data indicate that replete Si and low salinity in seawater can enhance cadmium and copper adsorption onto the diatom surface. At the single-cell level, surface potential was a dominant factor determining metal adsorption, while surface roughness also contributed to the higher metal loading capacity at lower salinities. Using a combination of non-invasive micro-test technology, atomic force microscopy, X-ray photoelectron spectroscopy, and Fourier transform infrared spectroscopy, we demonstrate that the diversity and abundance of the functional groups embedded in diatom cell walls vary with salinity and Si supply. This results in a change in the cell surface potential and transient metal influx. Our study provides novel mechanisms to explain the highly variable metal adsorption capacity of a model estuarine diatom.

Ethyl vinyl ketone activates oxidative and calcium burst and CML8-ACA8 participates in calcium recovery in Arabidopsis leaves

Plants produce ethyl vinyl ketone (evk) in response to biotic stress, but the evk's identification and downstream defense response remain unclear. In this paper, it is predicted by docking for the first time that evk can be recognized by RBOH protein and assist the electron transfer of RBOHD/RBOHF by binding to its FAD or NADPH binding site. Surface plasmon resonance (SPR) binding assay shows that evk indeed bind to RBOHD. Here, we show that evk treatment increased H2O2 and intracellular calcium concentrations in Arabidopsis thaliana mesophyll cells, as observed by confocal laser scanning microscopy and non-invasive micro-test technology, and that H2O2 signaling functioned upstream of Ca2+ signaling. Yeast two-hybrid, firefly luciferase complementation imaging, and in vitro pull-down assays demonstrated that the ACA8 (AUTOINHIBITED Ca2+-ATPASE, ISOFORM 8)–CML8 (CALMODULIN-LIKE 8) interaction promoted Ca2+ efflux to return Ca2+ levels to the resting state.

Aluminum decreases cadmium accumulation by down-regulating the expression of cadmium-related genes in wheat

Aluminum (Al) and cadmium (Cd) often coexist in the acidic soils of southern China, but their interactive effects remain unclear. In this study, this was examined in Al-resistant (Atlas 66) and Al-sensitive (Scout 66) wheat cultivars. The results showed that Al significantly alleviated the root growth of Al-tolerant Atlas 66 under Cd stress but had no effect on the root growth of Al-sensitive Scout 66 or the shoot growth of both cultivars. However, Al substantially decreased Cd accumulation in the roots and shoots of Atlas 66 and Scout 66. This could be attributed to the decreased uptake of Cd by the roots rather than the changes in Cd distribution in various organs or the Cd binding and adsorption of the cell wall. Using non-invasive micro-test technology, we further confirmed that Al and Cd co-exposure significantly inhibited the net Cd2+ influx into the roots of Atlas 66 and Scout 66 compared to Cd-only stress. Furthermore, the higher Cd2+ influx into roots should be responsible for the high Cd accumulation in the roots and shoots of Atlas66 than in those of Scout66 under Al and Cd co-exposure, which may be due to the higher Al accumulation in the roots of Al-sensitive Scout 66 that resulted in the larger decrease of negative charges on root surfaces. Finally, the effects of Al on the expression of Cd-related genes responsible for Cd uptake and translocation in wheat roots were investigated. The results have suggested that Al significantly downregulated the expression of TaNramp5 in Atlas 66 and the expression of TaNramp5, TaIRT1 and TaHMA2 in Scout66.

Differences in Cadmium Uptake and Accumulation in Seedlings of Wheat Varieties with Low- and High-Grain Cadmium Accumulation under Different Drought Stresses.

Cadmium (Cd) and drought, as abiotic stresses, have long been significant challenges for crop growth and agricultural production. However, there have been relatively few studies conducted on the effects of drought stress on Cd uptake, especially regarding the differences in Cd uptake characterization in varieties with varying Cd accumulation under different drought stress. To investigate the effects of drought conditions on Cd uptake by wheat in different genotypes under specific background levels of Cd pollution, we validated the differences in root absorption characteristics of low- (YM) and high-grain Cd accumulating wheat genotypes (XM) using non-invasive micro-test technology, and we conducted a hydroponic experiment on the Cd addition and different drought levels in a climate-controlled chamber. The biomass, root morphology, Cd uptake, and accumulation were determined under Cd (100 µmol L-1) and different drought levels of 0% (0 MPa), 5% (-0.100 Mpa), 10% (-0.200 Mpa), and 15% (-0.388 Mpa) simulated by polyethylene glycol (PEG-6000). We found that the simultaneous exposure to Cd and drought had a suppressive effect on the total root lengths, root surface areas, and root volumes of XM and YM, albeit with distinct patterns of variation. As the concentration of PEG-6000 increased, the Cd concentrations and the amount of Cd accumulated in the roots and shoots of XM and YM decreased. Specifically, the Cd concentration in the roots exhibited a reduction ranging from 12.51% to 66.90%, while the Cd concentration in the shoots experienced an even greater decrease of 50.46% to 80.57%. The PEG-6000 concentration was significantly negatively correlated (p < 0.001) with Cd concentration of roots and shoots and Cd accumulation in roots, shoots, and the whole plants and significantly negatively correlated (p < 0.05) with the total length, surface area, and volume of roots. This study confirms that drought stress (5% PEG-6000) can decrease the uptake and accumulation of Cd in wheat seedlings without significant inhibition of biomass, and the change of root morphology (root length) and the decrease of Cd concentration in roots may be the main direct pathways for achieving these effects under drought stress. This research provides a new perspective and idea for water management in Cd-contaminated farmland.

Salt-Stress-Induced Ion Transport Contributes to K+/Na+ Homeostasis in Roots of Ping’ou Hybrid Hazelnut

Soil salinity is a worldwide problem that adversely affects plant growth and development. Soil salinization in Xinjiang of China is very serious. Ping’ou hybrid hazelnut, as an important ecological and economic tree species, as well as a salt-tolerant plant, has been grown in Xinjiang for over 20 years. Understanding the salt-tolerance mechanism of Ping’ou hybrid hazelnut is of great significance for the breeding of salt-tolerant varieties and the rational utilization of salinized land. In this study, ‘Liaozhen 7’, a fine variety of Ping’ou hybrid hazelnut, was selected as test material, and seedlings were treated with 0 (control), 50, 100 and 200 mM NaCl. Subsequently, the pattern of NaCl-induced fluxes of Na+, K+ and H+ in the root meristematic zone and their response to ion transport inhibitors were studied using non-invasive micro-test technology (NMT). Different concentrations of NaCl stress significantly increased the Na+ concentration in roots, while K+ concentration decreased first and then increased with the increase of NaCl concentration. Meanwhile, NaCl stress induced a significant decline in K+/Na+ ratio. Control and 200 mM NaCl-induced Na+ and K+ fluxes in roots exhibited an outward efflux, whereas an inward flux was observed for H+. Under 200 mM NaCl stress, the average rates of net Na+ and K+ efflux, as well as H+ influx in roots were significantly increased, which were 11.6, 6.7 and 2.3 times higher than that of control, respectively. Furthermore, pharmacological experiments showed that 200 mM NaCl-induced Na+ efflux; H+ influx was significantly suppressed by amiloride, an inhibitor of plasma membrane (PM) Na+/H+ antiporter, and sodium vanadate, an inhibitor of PM H+-ATPase. Net Na+ efflux and H+ influx induced by NaCl decreased by 89.9% and 135.0%, respectively. The NaCl-induced Na+ efflux was mediated by a Na+/H+ antiporter using energy provided by PM H+-ATPase. The NaCl-induced K+ efflux was significantly restricted by tetraethylamine chloride, a K+ channel inhibitor, and promoted by sodium vanadate, which decreased by 111.2% and increased by 80.8%, respectively, indicating that K+ efflux was regulated by depolarization-activated outward-rectifying K+ channels and non-selective cation channels (NSCCs). In conclusion, NMT data revealed that NaCl stress up regulated the root Na+/H+ antiporter and H+ pump (an activity of PM Na+/H+ antiport system) of ‘Liaozhen 7’, which compelled the Na+/H+ exchange across the PM and restricted K+ loss via depolarization-activated K+ channels and NSCCs simultaneously, thereby maintaining the K+/Na+ homeostasis and higher salt tolerance.

Non-Invasive Micro-Test Technology and Reciprocal Grafting Provide Direct Evidence of Contrasting Na+ Transport Strategies between Cucurbita moschata and Cucurbita maxima

Cucurbita species are widely used as rootstocks for cucumber, watermelon, and other plants for the restriction of toxic Na+ transport from root to shoot. Previous studies have found distinct salt tolerance strategies between Cucurbita moschata and Cucurbita maxima; the former accumulates a large amount of Na+ in the root, while the latter accumulates Na+ in the shoot. To further study the mechanism of Na+ transport in plants, four reciprocal grafts were made between MB (C. moschata) and JHL (C. maxima), denoted as MB/MB, JHL/JHL, MB/JHL, and JHL/MB (scion/rootstock). The results showed that using MB as the rootstock effectively reduced the accumulation of Na+ in the scion. Conversely, JHL rootstock tended to transfer more Na+ to the scion. To clarify this phenomenon, the velocities of Na+ flows on the root surface, stem, and vein of grafting combinations were measured using non-invasive micro-test technology. Compared with the seedlings using JHL as rootstock, the grafted combination with MB as rootstock had a higher root Na+ efflux and lower Na+ fluxes in the stem and vein. qRT-PCR analyses revealed the critical roles of salt overly sensitive 1 and high-affinity potassium as components of the mechanism enabling Na+ exclusion from the root and Na+ unloading from the stem xylem. Compared with the seedlings using MB as rootstocks, the JHL-grafted plants showed more rapid stomatal closure and decreased transpiration rate in the first three hours after salt stress but maintained a higher level under prolonged salt treatment (120 h). The tissue tolerances of JHL and MB were assessed using the isolated leaves under NaCl to exclude the influence of the root and stem. The results showed that the salinity inflicted more serious damage to MB leaves than to JHL leaves. qRT-PCR analyses indicated that the intracellular Na+/H+ transporter in the leaf vein was involved in this process. All these findings indicated that C. moschata and C. maxima adopted different strategies for regulating Na+ transport, and grafting can be used as a tool to create more salt-tolerant plants.

Cadmium uptake and membrane transport in roots of hyperaccumulator Amaranthus hypochondriacus L.

Hyperaccumulator Amaranthus hypochondriacus L. has huge potential in the remediation of cadmium (Cd)-contaminated soils and is necessary to understand the mechanism of Cd uptake by the roots. In this study, the mechanism of Cd uptake into the root of A. hypochondriacus was investigated using the non-invasive micro-test technology (NMT) by analyzing the rate of Cd2+ fluxes at different regions of the root tip; also we assessed the impact of different channel blockers and inhibitors on the Cd accumulation in the roots, the real-time Cd2+ fluxes, and the distribution of Cd along the roots. The results showed that the Cd2+ influx was greater near the root tip (within 100 μm of the tip). All the inhibitors, ion-channel blockers, and metal cations had different degrees of inhibition on the absorption of Cd in the roots of A. hypochondriacus. The net Cd2+ flux in the roots was significantly decreased by the Ca2+ channel blockers lanthanum chloride (LaCl3) by up to 96% and verapamil by up to 93%; as for the K+ channel blocker tetraethylammonium (TEA), it also caused a 68%-reduction on the net Cd2+ flux in the roots. Therefore, we infer that the uptake by A. hypochondriacus roots is mainly through the Ca2+ channels. The Cd absorption mechanism appears to be related to the synthesis of plasma membrane P-type ATPase and phytochelatin (PC), which is reflected by the inhibition of Ca2+ upon addition of inorganic metal cations. In conclusion, access of Cd ions into the roots of A. hypochondriacus is achieved through various ion channels, with the most important being the Ca2+ channel. This study will further enhance the literature regarding Cd uptake and pathways of membrane transport in roots of Cd hyperaccumulators.

Characteristics of Cd Uptake by the Roots of Bidens tripartita L. Under Salinity and pH Variations Assessed by Applying Non-invasive Micro-test Technology

Characteristics of Cd Uptake by the Roots of Bidens tripartita L. Under Salinity and pH Variations Assessed by Applying Non-invasive Micro-test Technology

Influencing Factors of Bidens pilosa L. Hyperaccumulating Cadmium Explored by the Real-Time Uptake of Cd2+ Influx around Root Apexes under Different Exogenous Nutrient Ion Levels.

Though Bidens pilosa L. has been confirmed to be a potential Cd hyperaccumulator, the accumulation mechanism is not yet clear. The dynamic and real-time uptake of Cd2+ influx by B. pilosa root apexes was determined using non-invasive micro-test technology (NMT), which partly explored the influencing factors of the Cd hyperaccumulation mechanism under the conditions of different exogenous nutrient ions. The results indicated that Cd2+ influxes at 300 μm around the root tips decreased under Cd treatments with 16 mM Ca2+, 8 mM Mg2+, 0.5 mM Fe2+, 8 mM SO42- or 18 mM K+ compared to single Cd treatments. The Cd treatments with a high concentration of nutrient ions showed an antagonistic effect on Cd2+ uptake. However, Cd treatments with 1 mM Ca2+, 0.5 mM Mg2+, 0.5 mM SO42- or 2 mM K+ had no effect on the Cd2+ influxes as compared with single Cd treatments. It is worth noting that the Cd treatment with 0.05 mM Fe2+ markedly increased Cd2+ influxes. The addition of 0.05 mM Fe2+ exhibited a synergistic effect on Cd uptake, which could be low concentration Fe2+ rarely involved in blocking Cd2+ influx and often forming an oxide membrane on the root surface to help the Cd uptake by B. pilosa. The results also showed that Cd treatments with high concentration of nutrient ions significantly increased the concentrations of chlorophyll and carotenoid in leaves and the root vigor of B. pilosa relative to single Cd treatments. Our research provides novel perspectives with respect to Cd uptake dynamic characteristics by B. pilosa roots under different exogenous nutrient ion levels, and shows that the addition of 0.05 mM Fe2+ could promote the phytoremediation efficiency for B. pilosa.

Drought priming mechanisms in wheat elucidated by in-situ determination of dynamic stomatal behavior.

Stomata play a critical role in balancing photosynthesis and transpiration, which are essential processes for plant growth, especially in response to abiotic stress. Drought priming has been shown to improve drought tolerance. Lots of studies have been done with the response of stomatal behavior to drought stress. However, how the stomatal dynamic movement in intact wheat plants response to drought priming process is not known. Here, a portable microscope was used to take microphotographs in order to in-stiu determination of stomatal behavior. Non-invasive micro-test technology was used for measurements of guard cell K+, H+ and Ca2+ fluxes. Surprisingly, the results found that primed plants close stomatal much faster under drought stress, and reopening the stomatal much quicker under recovery, in relation to non-primed plants. Compared with non-primed plants, primed plants showed higher accumulation of ABA and Ca2+ influx rate in guard cells under drought stress. Furthermore, genes encoding anion channels were higher expressed and K+ outward channels activated, leading to enhanced K+ efflux, resulting in faster stomatal closure in primed plants than non-primed plants. During recovery, both guard cell ABA and Ca2+ influx of primed plants were found to be significantly reducing K+ efflux and accelerating stomatal reopening. Collectively, a portable non-invasive stomatal observation of wheat found that priming promoted faster stomatal closure under drought stress and faster reopening during post-drought recovery in relation to non-primed plants, thereby enhancing overall drought tolerance.