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It is known that selenium (Se) enhances plant growth and arsenic (As) accumulation in As-hyperaccumulator Pteris vittata, but the associated mechanisms are unclear. In this study, P. vittata was exposed to 50 μM arsenate (AsV) under hydroponics plus 25 or 50 μM foliar selenate. After 3-weeks of growth, the plant biomass, As and Se contents, As speciation, malondialdehyde (MDA) and glutathione (GSH and GSSG) levels, and important genes related to As-metabolism in P. vittata were determined. Foliar-Se increased plant biomass by 17 - 30 %, possibly due to 9.1 - 19 % reduction in MDA content compared to the As control. Further, foliar-Se enhanced the As contents by 1.9-3.5 folds and increased arsenite (AsIII) contents by 64 - 136 % in the fronds. The increased AsV reduction to AsIII was attributed to 60 - 131 % increase in glutathione peroxidase activity, which mediates GSH oxidation to GSSG (8.8 -29 % increase) in the fronds. Further, foliar-Se increased the expression of AsIII antiporters PvACR3;1-3;3 by 1.6 - 2.1 folds but had no impact on phosphate transporters PvPht1 or arsenate reductases PvHAC1/2. Our results indicate that foliar-Se effectively enhances plant growth and arsenic accumulation by promoting the GSH-GSSG cycle and upregulating gene expression of AsIII antiporters, which are responsible for AsIII translocation from the roots to fronds and AsIII sequestration into the fronds. The data indicate that foliar-Se can effectively improve phytoremediation efficiency of P. vittata in As-contaminated soils., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Pteris vittata is the first-reported arsenic (As) hyperaccumulator, which has been applied to phytoremediation of As-contaminated soil. PvACR3, a key arsenite (AsIII) antiporter, plays an important role in As hyperaccumulation in P. vittata . However, its functions in plants are not fully understood. In this study, the PvACR3 gene was heterologously expressed in tobacco, driven by its native promoter ( Pro PvACR3 ). After growing at 5 μM AsIII or 10 μM AsV in hydroponics for 1-5 days, PvACR3 -expression enhanced the As levels in leaves by 66.4-113 and 51.8-101%, without impacting the As contents in the roots or stems. When cultivated in As-contaminated soil, PvACR3 -expressed transgenic plants accumulated 47.9-85.5% greater As in the leaves than wild-type plants. In addition, PvACR3 -expression increased the As resistance in transgenic tobacco, showing that enhanced leaf As levels are not detrimental to its overall As tolerance. PvACR3 was mainly expressed in tobacco leaf veins and was likely to unload AsIII from the vein xylem vessels to the mesophyll cells, thus elevating the leaf As levels. This work demonstrates that heterologously expressing PvACR3 under its native promoter specifically enhances leaf As accumulation in tobacco, which helps to reveal the As-hyperaccumulation mechanism in P. vittata and to enhance the As accumulation in plant leaves for phytoremediation.

The first-known As-hyperaccumulator Pteris vittata is efficient in As uptake and translocation, which can be used for phytoremediation of As-contaminated soils. However, the underlying mechanisms of As-enhanced plant growth are unknown. We used untargeted metabolomics to investigate the potential metabolites and associated metabolic pathways regulating As-enhanced plant growth in P. vittata. After 60 days of growth in an MS-agar medium containing 15 mg kg -1 As, P. vittata biomass was 33-34 % greater than the no-As control. Similarly, the As contents in P. vittata roots and fronds were 272 and 1300 mg kg -1 , considerably greater than the no-As control. Univariate and multivariate analyses based on electrospray ionization indicate that As exposure changed the expression of 1604 and 1248 metabolites in positive and negative modes. By comparing with the no-As control, As exposure significantly changed the expression of 14 metabolites including abscisic acid, d-glucose, raffinose, stachyose, chitobiose, xylitol, gibberellic acids, castasterone, citric acid, riboflavin-5-phosphate, ubiquinone, ubiquinol, UDP-glucose, and GDP-glucose. These metabolites are involved in phytohormone synthesis, energy metabolism, and sugar metabolism and may all potentially contribute to regulating As-enhanced plant growth in P. vittata. Our data provide clues to understanding the metabolic regulations of As-enhanced plant growth in P. vittata, which helps to enhance its phytoremediation efficiency of As-contaminated soils., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Sparingly-soluble phosphate rock (PR), a raw material for P-fertilizer production, can be effectively utilized by the As-hyperaccumulator Pteris vittata but not most plants. In this study, we investigated the associated mechanisms by measuring dissolved organic carbon (DOC) and acid phosphatase in the rhizosphere, and nutrient uptake and gene expression related to the As metabolism in P. vittata . The plants were grown in a soil containing 200 mg kg -1 As and/or 1.5% PR for 30 days. Compared to the As treatment, the P. vittata biomass was increased by 33% to 4.6 g plant -1 in the As+PR treatment, corresponding to 27% decrease in its frond oxidative stress as measured by malondialdehyde. Due to PR-enhanced DOC production in the rhizosphere, the Ca, P, and As contents in P. vittata fronds were increased by 17% to 9.7 g kg -1 , 29% to 5.0 g kg -1 , and 57% to 1045 mg kg -1 in the As+PR treatment, thereby supporting its better growth. Besides, PR-induced rhizosphere pH increase from 5.0 to 6.9 promoted greater P uptake by P. vittata probably via upregulating low-affinity P transporters PvPTB1;1/1;2 by 3.7-4.1 folds. Consequently, 29% lower available-P induced the 3.3-fold upregulation of high-affinity P transporter PvPht1;3 in the As+PR treatment, which was probably responsible for the 58% decrease in available-As content in the rhizosphere. Consistent with the enhanced As translocation and sequestration, arsenite antiporters PvACR3/3;3 were upregulated by 1.8-4.4 folds in the As+PR than As treatment. In short, sparingly-soluble PR enhanced the Ca, P, and As availability in P. vittata rhizosphere and improved their uptake via upregulating genes related to As metabolism, suggesting its potential application for improving phytoremediation in As-contaminated soils.

Arsenic (As) contamination in soil poses a potential threat to human health via crop uptake. As-hyperaccumulator Pteris vittata serves as a model plant to study As uptake and associated mechanisms. This study focuses on a novel P/AsV transport system mediated by low-affinity phosphate transporter-B 1 family (PTB1) in P. vittata . Here, we identified two plasma-membrane-localized PTB1 genes, PvPTB1;1/1;2 , in vascular plants for the first time, which were 4.4-40-fold greater in expression in P. vittata than in other Pteris ferns. Functional complementation of a yeast P-uptake mutant and enhanced P accumulation in transgenic Arabidopsis thaliana confirmed their role in P uptake. Moreover, the expression of PvPTB1;1/1;2 facilitated the transport and accumulation of As in both yeast and A. thaliana shoots, demonstrating a comparable AsV uptake capacity. Microdissection-qPCR analysis and single-cell transcriptome analysis collectively suggest that PvPTB1;1/1;2 are specifically expressed in the epidermal cells of P. vittata roots. PTB1 may play a pivotal role in efficient P recycling during phytate secretion and hydrolysis in P. vittata roots. In summary, the dual P transport mechanisms consisting of high-affinity Pht1 and low-affinity PTB1 may have contributed to the efficient P/As uptake in P. vittata , thereby contributing to efficient phytoremediation for As-contaminated soils.

Phytate, the principal P storage in plant seeds, is also an important organic P in soils, but it is unavailable for plant uptake. However, the As-hyperaccumulator Pteris vittata can effectively utilize soluble Na-phytate, while its ability to utilize insoluble Ca/Fe-phytate is unclear. Here, we investigated phytate uptake and the underlying mechanisms based on the phytase activity, nutrient uptake, and expression of genes involved in As metabolisms. P. vittata plants were cultivated hydroponically in 0.2-strength Hoagland nutrient solution containing 50 μM As and 0.2 mM Na/Ca/Fe-phytate, with 0.2 mM soluble-P as the control. As the sole P source, all three phytates supported P. vittata growth, with its biomass being 3.2-4.1 g plant -1 and Ca/Fe-phytate being 19-29% more effective than Na-phytate. Phytate supplied soluble P to P. vittata probably via phytase hydrolysis, which was supported by 0.4-0.7 nmol P min -1 g -1 root fresh weight day -1 phytase activity in its root exudates, with 29-545 μM phytate-P being released into the growth media. Besides, compared to Na-phytate, Ca/Fe-phytate enhanced the As contents by 102-140% to 657-781 mg kg -1 in P. vittata roots and by 43-86% to 1109-1447 mg kg -1 in the fronds, which was accompanied by 21-108% increase in Ca and Fe uptake. The increased plant As is probably attributed to 1.3-2.6 fold upregulation of P transporters PvPht1;3/4 for root As uptake, and 1.8-4.3 fold upregulation of arsenite antiporters PvACR3/3;1/3;3 for As translocation to and As sequestration into the fronds. This is the first report to show that, besides soluble Na-phytate, P. vittata can also effectively utilize insoluble Ca/Fe-phytate as the sole P source, which sheds light onto improving its application in phytoremediation of As-contaminated sites.

Prebiotics may stimulate beneficial gut microorganisms. However, it remains unclear whether they can lower the oral bioavailability of early life arsenic (As) exposure via regulating gut microbiota and altering As biotransformation along the gastrointestinal (GI) tract. In this study, weanling mice were exposed to arsenate (iAs V ) via diet (7.5 μg As g -1 ) amended with fructooligosaccharides (FOS), galactooligosaccharides (GOS), and inulin individually at 1% and 5% (w/w). Compared to As exposure control mice, As concentrations in mouse blood, liver, and kidneys and As urinary excretion factor (UEF) were reduced by 43.7%-74.1% when treated with 5% GOS. The decrease corresponded to a significant proliferation of Akkermansia and Psychrobacter , reduced percentage of inorganic arsenite (iAs III ) and iAs V by 47.4% and 65.4%, and increased proportion of DMA V in intestinal contents by 101% in the guts of mice treated with 5% GOS compared to the As control group. In contrast, FOS and inulin either at l% or 5% did not reduce As concentration in mouse blood, liver, and kidneys or As UEF. These results suggest that GOS supplementation may be a gut microbiota-regulating approach to lower early life As exposure via stimulating the growth of Akkermansia and Psychrobacter and enhancing As methylation in the GI tract.

In contaminated soils, arsenic (As) often co-exists with copper (Cu). However, its effects on As accumulation and the related mechanisms in As-hyperaccumulator Pteris vittata remain unclear. In this study, P. vittata plants were exposed to 50 µM As and/or 50 µM Cu under hydroponics to investigate the effects of Cu on plant growth and As accumulation, as well as gene expression related to arsenic uptake (P transporters), reduction (arsenate reductases), and translocation and sequestration (arsenite antiporters). After 14 d of growth and compared to the As treatment, the As concentration in P. vittata fronds increased by 1.4-times from 793 to 1131 mg·kg -1 and its biomass increased by 1.2-fold from 18.0 to 21.1 g·plant -1 in the As+Cu treatment. Copper-enhanced As accumulation was probably due to upregulated gene expressions related to As-metabolisms including As uptake (1.9-fold in P transporter PvPht1;3), translocation (2.1-2.4 fold in arsenite antiporters PvACR3/3;2) and sequestration (1.5-2.0 fold in arsenite antiporters PvACR3;1/3;3). Our results suggest that moderate amount of Cu can help to increase the As accumulation efficiency in P. vittata, which has implication in its application in phytoremedation in As and Cu co-contaminated soils., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Arsenic (As) is easily accumulated in wild Boletus. However, the accurate health risks and adverse effects of As on humans were largely unknown. In this study, we analyzed the total concentration, bioavailability, and speciation of As in dried wild boletus from some typical high geochemical background areas using an in vitro digestion/Caco-2 model. The health risk assessment, enterotoxicity, and risk prevention strategy after consumption of As-contaminated wild Boletus were further investigated. The results showed that the average concentration of As was 3.41-95.87 mg/kg dw, being 1.29-56.3 folds of the Chinese food safety standard limit. DMA and MMA were the dominant chemical forms in raw and cooked boletus, while their total (3.76-281 mg/kg) and bioaccessible (0.69-153 mg/kg) concentrations decreased to 0.05-9.27 mg/kg and 0.01-2.38 mg/kg after cooking. The EDI value of total As was higher than the WHO/FAO limit value, while the bioaccessible or bioavailable EDI suggested no health risks. However, the intestinal extracts of raw wild boletus triggered cytotoxicity, inflammation, cell apoptosis, and DNA damage in Caco-2 cells, indicating existing health risk assessment models based on total, bioaccessible, or bioavailable As may be not accurate enough. Given that, the bioavailability, species, and cytotoxicity should be systematically considered in accurate risk assessment. In addition, cooking mitigated the enterotoxicity along with decreasing the total and bioavailable DMA and MMA in wild boletus, suggesting that cooking could be a simple and effective way to decrease the health risks of consumption of As-contaminated wild boletus., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Soil contamination by arsenic (As) poses potential health risks to humans. As-hyperaccumulator P. vittata has been used in As-contaminated soils for phytoremediation. Clarifying the mechanisms of its As-hyperaccumulation is critical to enhance its efficiency in phytoremediation. Here, based on transcriptome analysis, we determined the concentration-dependent patterns of As-related gene families by comparing As-hyperaccumulator P. vittata and non-hyperaccumulator P. ensiformis after exposing to 20 µM arsenate (AsV). As expected, arsenic induced more stress in P. ensiformis than P. vittata. Based on gene ontology, differences in transporter activity are probably responsible for their differential As accumulation. Though As exposure induced expression of phosphate transporter PvPht1;4 for AsV absorption in both plants, stronger AsV reduction, AsIII transport, and AsIII-GSH complexation were found in P. ensiformis roots. Unlike P. ensiformis, As metabolism processes occurred mainly in P. vittata fronds. Notably, tonoplast-localized ACR3s were only present in P. vittata, making it more effective in sequestrating AsIII into frond vacuoles. Further, vesicle As transformation via PvGAPC1 (glyceraldehyde 3-phosphate dehydrogenase), PvOCT4 (organic cation transporter 4), and PvGSTF1 (glutathione S-transferase) contributed little to As-hyperaccumulation. This study provides information on critical genes responsible for As-hyperaccumulation by P. vittata, which can be applied to construct As-hyperaccumulating plants by genetic engineering to enhance their phytoremediation efficiency in As-contaminated soils., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Few studies have investigated the long-term effect of exposure to arsenic (As), lead (Pb), and cadmium (Cd) via drinking water at the provisional guideline values on gut microflora. In this study, male and female mice were exposed to water As, Pb, or Cd at 10, 10, or 5 μg L -1 for 6 months. At the end of the exposure, the net weight gain of male mice exposed to As and Pb (9.91 ± 1.35 and 11.2 ± 1.50 g) was significantly ( p < 0.05) lower compared to unexposed control mice (14.1 ± 3.24 g), while this was not observed for female mice. Relative abundance of Akkermansia , a protective gut bacterium against intestinal inflammation, was reduced from 29.7% to 3.20%, 4.83%, and 17.0% after As, Pb, and Cd exposure in male mice, which likely caused chronic intestinal inflammation, as suggested by 2.81- to 9.60-fold higher mRNA levels of pro-inflammatory factors in ileal enterocytes of male mice. These results indicate that long-term exposure to drinking water As, Pb, and Cd at concentrations equivalent to the China provisional guideline values can cause loss of protective bacteria and lead to chronic intestinal inflammation, thereby affecting body weight gain in male mice.

The behaviors of antimony (Sb) and arsenic (As) in plants are different, though they are chemical analogs. Here, we examined the Sb uptake and speciation in two As-hyperaccumulators P. vittata and P. cretica, which were exposed to 0.5 or 5 mg L -1 antimonate (SbV) or antimonite (SbIII) under hydroponics for 7 d. Both plants grew better under Sb exposure, especially for P. cretica. The biomass of P. cretica roots increased by 29-46% after exposing to SbV, possibly due to increased S. Further, the Sb content in P. vittata was 17-93% greater than P. cretica, with 2-3 times more SbIII than SbV in both plants and > 92% Sb being concentrated in the roots, showing limited translocation. Under SbV exposure, SbV was dominant in P. vittata roots at 86-94%, while SbIII was predominant in P. cretica roots at 36-95%. P. cretica's stronger reducing ability than P. vittata may be due to arsenate reductases HAC1 and ACR2, which were upregulated in both plants. In short, while effective in Sb accumulation, it is mostly concentrated in the roots for both plants. The differences in their accumulation and speciation may help to better understand Sb behaviors in other plants., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Microplastics exposure is a new human health crisis. Although progress in understanding health effects of microplastic exposure has been made, microplastic impacts on absorption of co-exposure toxic pollutants such as arsenic (As), i.e., oral bioavailability, remain unclear. Microplastic ingestion may interfere As biotransformation, gut microbiota, and/or gut metabolites, thereby affecting As oral bioavailability. Here, mice were exposed to arsenate (6 μg As g -1 ) alone and in combination with polyethylene particles of 30 and 200 μm (PE-30 and PE-200 having surface area of 2.17 × 10 3 and 3.23 × 10 2  cm 2  g -1 ) in diet (2, 20, and 200 μg PE g -1 ) to determine the influence of microplastic co-ingestion on arsenic (As) oral bioavailability. By determining the percentage of cumulative As consumption recovered in urine of mice, As oral bioavailability increased significantly (P < 0.05) from 72.0 ± 5.41% to 89.7 ± 6.33% with PE-30 at 200 μg PE g -1 rather than with PE-200 at 2, 20, and 200 μg PE g -1 (58.5 ± 19.0%, 72.3 ± 6.28%, and 69.2 ± 17.8%). Both PE-30 and PE-200 exerted limited effects on pre- and post-absorption As biotransformation in intestinal content, intestine tissue, feces, and urine. They affected gut microbiota dose-dependently, with lower exposure concentrations having more pronounced effects. Consistent with the PE-30-specific As oral bioavailability increase, PE exposure significantly up-regulated gut metabolite expression, and PE-30 exerted greater effects than PE-200, suggesting that gut metabolite changes may contribute to As oral bioavailability increase. This was supported by 1.58-4.07-fold higher As solubility in the presence of up-regulated metabolites (e.g., amino acid derivatives, organic acids, and pyrimidines and purines) in the intestinal tract assessed by an in vitro assay. Our results suggested that microplastic exposure especially smaller particles may exacerbate the oral bioavailability of As, providing a new angle to understand health effects of microplastics., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Arsenic-hyperaccumulator Pteris vittata is efficient in taking up arsenate (AsV) and arsenite (AsIII), however, their impacts on P. vittata growth and nutrient uptake remain unclear. The uptake of AsV and AsIII, their influences on nutrient uptake and plant biomass, and As speciation were investigated in P. vittata after exposing to 5 or 50 μM AsV or AsIII for 12 d under hydroponics. The results show that AsV uptake in P. vittata was 1.2 times more efficient than AsIII, corresponding to 1.7-2.1 fold greater biomass than the control at 50 μM As. While AsV was dominant in the roots at ∼60%, AsIII was more dominant in the fronds at ∼70% in all treatments. Macronutrients P, K, Ca, and S were increased by 118-185% at 50 μM As, with greater uptake of micronutrients Fe, Mn, Cu, and Zn at 5 μM As. Further, positive correlations between P. vittata biomass and its As contents (r = 0.97), and P. vittata biomass and its S, Mg, P, or Ca contents (r = 0.70-0.98) were observed. Our results suggest that its increased nutrient uptake probably enhanced P. vittata growth under As exposure., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Early-life arsenic (As) exposure is a particular health concern. However, it is unknown if As ingested early in life is more readily absorbed from the gastrointestinal (GI) tract, i.e., higher in oral bioavailability. Here, weanling (3-week) and adult (6-week-old) female mice were exposed to arsenate in the diet (10 μg g -1 ) over a 3-week period with As oral bioavailability estimated using As urinary excretion as the bioavailability endpoint. The As urinary excretion factor was 1.54-fold higher in weanling mice compared to adult mice (82.2 ± 7.29 versus 53.1 ± 3.73%), while weanling mice also showed 2.28-, 1.50-, 1.48-, and 1.89-fold higher As concentration in small intestine tissue, blood, liver, and kidneys, demonstrating significantly higher As oral bioavailability of early-life exposure. Compared to adult mice, weanling mice significantly differed in gut microbiota, but the difference did not lead to remarkable differences in As biotransformation in the GI tract or tissue and in overall gut metabolite composition. Although the expression of several metabolites (e.g., atrolactic acid, hydroxyphenyllactic acid, and xanthine) was up-regulated in weanling mice, they had limited ability to elevate As solubility in the intestinal tract. Compared to adult mice, the intestinal barrier function and intestinal expression of phosphate transporters responsible for arsenate absorption were similar in weanling mice. However, the small intestine of weanling mice was characterized by more defined intestinal villi with greater length and smaller width, providing a greater surface area for As to be absorbed across the GI barrier. The results highlight that early-life As exposure can be more readily absorbed, advancing the understanding of its health risk.

Phytate as a root exudate is rare in plants as it mainly serves as a P storage in the seeds; however, As-hyperaccumulator Pteris vittata effectively secretes phytate and utilizes phytate-P, especially under As exposure. This study investigated the effects of As on its phytate and phytase exudation and the impacts of As and/or phytate on each other's uptake in P. vittata through two hydroponic experiments. Under 10-100 μM arsenate (AsV), the exudation of phytate and phytase by P. vittata was increased by 50-72% to 20.4-23.4 μmol h -1 g -1 and by 28-104% to 18.6-29.5 nmol h -1 plant -1 , but they were undetected in non-hyperaccumulator Pteris ensiformis at 10 μM AsV. Furthermore, compared to 500 μM phytate, the phytate concentration in the growth media was reduced by 69% to 155 μM, whereas the P and As contents in P. vittata fronds and roots were enhanced by 68-134% and 44-81% to 2423-2954 and 82-407 mg kg -1 under 500 μM phytate plus 50 μM AsV. The increased P/As uptake in P. vittata was probably attributed to 3.0-4.5-fold increase in expressions of P transporters PvPht1;3-1;4 . Besides, under As exposure, plant P may be converted to phytate in P. vittata roots, thereby increasing phytate's contents by 84% to 840 mg kg -1 . Overall, our results suggest that As-induced phytate/phytase exudation and phytate-P uptake stimulate its growth and As hyperaccumulation by P. vittata .

Background: Elevating dietary calcium (Ca) intake can reduce metal(loid)oral bioavailability. However, the ability of a range of Ca minerals to reduce oral bioavailability of lead (Pb), cadmium (Cd), and arsenic (As) from indoor dust remains unclear., Objectives: This study evaluated the ability of Ca minerals to reduce Pb, Cd, and As oral bioavailability from indoor dust and associated mechanisms., Methods: A mouse bioassay was conducted to assess Pb, Cd, and As relative bioavailability (RBA) in three indoor dust samples, which were amended into mouse chow without and with addition of CaHPO 4 ,  CaCO 3 , Ca gluconate, Ca lactate, Ca aspartate, and Ca citrate at 200 - 5,000 μ g / g   Ca . The mRNA expression of Ca and phosphate (P) transporters involved in transcellular Pb, Cd and As transport in the duodenum of mice was quantified using real-time polymerase chain reaction. Serum 1,25-Dihydroxyvitamin D3 [ 1,25 ( OH ) 2 D 3 ], parathyroid hormone (PTH), and renal CYP27B1 activity controlling 1,25 ( OH ) 2 D 3 synthesis were measured using ELISA kits. Metal(loid) speciation in the feces of mice was characterized using X-ray absorption near-edge structure (XANES) spectroscopy., Results: In general, mice exposed to each of the Ca minerals exhibited lower Pb-, Cd-, and As-RBA for three dusts. However, RBAs with the different Ca minerals varied. Among minerals, mice fed dietary CaHPO 4 did not exhibit lower duodenal mRNA expression of Ca transporters but did have the lowest Pb and Cd oral bioavailability at the highest Ca concentration ( 5,000 μ g / g   Ca ; 51%-95% and 52%-74% lower in comparison with the control). Lead phosphate precipitates (e.g., chloropyromorphite) were observed in feces of mice fed dietary CaHPO 4 . In comparison, mice fed organic Ca minerals (Ca gluconate, Ca lactate, Ca aspartate, and Ca citrate) had lower duodenal mRNA expression of Ca transporters, but Pb and Cd oral bioavailability was higher than in mice fed CaHPO 4 . In terms of As, mice fed Ca aspartate exhibited the lowest As oral bioavailability at the highest Ca concentration ( 5,000 μ g / g   Ca ; 41%-72% lower) and the lowest duodenal expression of P transporter (88% lower). The presence of aspartate was not associated with higher As solubility in the intestine., Discussion: Our study used a mouse model of exposure to household dust with various concentrations and species of Ca to determine whether different Ca minerals can reduce bioavailability of Pb, Cd, and As in mice and elucidate the mechanism(s) involved. This study can contribute to the practical application of optimal Ca minerals to protect humans from Pb, Cd, and As coexposure in the environment. https://doi.org/10.1289/EHP11730.

Selenate enhances arsenic (As) accumulation in As-hyperaccumulator Pteris vittata , but the associated molecular mechanisms are unclear. Here, we investigated the mechanisms of selenate-induced arsenic accumulation by exposing P. vittata to 50 μM arsenate (AsV 50 ) and 1.25 (Se 1.25 ) or 5 μM (Se 5 ) selenate in hydroponics. After 2 weeks, plant biomass, plant As and Se contents, As speciation in plant and growth media, and important genes related to As detoxification in P. vittata were determined. These genes included P transporters PvPht1;3 and PvPht1;4 (AsV uptake), arsenate reductases PvHAC1 and PvHAC2 (AsV reduction), and arsenite (AsIII) antiporters PvACR3 and PvACR3;2 (AsIII translocation) in the roots, and AsIII antiporters PvACR3;1 and PvACR3;3 (AsIII sequestration) in the fronds. The results show that Se 1.25 was more effective than Se 5 in increasing As accumulation in both P. vittata roots and fronds, which increased by 27 and 153% to 353 and 506 mg kg -1 . The As speciation analyses show that selenate increased the AsIII levels in P. vittata , with 124-282% more AsIII being translocated into the fronds. The qPCR analyses indicate that Se 1.25 upregulated the gene expression of PvHAC1 by 1.2-fold, and PvACR3 and PvACR3;2 by 1.0- to 2.5-fold in the roots, and PvACR3;1 and PvACR3;3 by 0.6- to 1.1-fold in the fronds under AsV 50 treatment. Though arsenate enhanced gene expression of P transporters PvPht1;3 and PvPht1;4 , selenate had little effect. Our results indicate that selenate effectively increased As accumulation in P. vittata , mostly by increasing reduction of AsV to AsIII in the roots, AsIII translocation from the roots to fronds, and AsIII sequestration into the vacuoles in the fronds. The results suggest that selenate may be used to enhance phytoremediation of As-contaminated soils using P. vittata .

Arsenic (As) is toxic and ubiquitous in the environment, posing a growing threat to human health. As-hyperaccumulator Pteris vittata has been used for phytoremediation of As-contaminated soil. Symbiosis with arbuscular mycorrhizal fungi (AMF) enhances As accumulation by P. vittata , which is different from As inhibition in typical plants. In this study, P. vittata seedlings inoculated with or without AMF were cultivated in As-contaminated soils for 2 months. AMF-root symbiosis enhanced plant growth, with 64.5% greater As contents in the fronds. After exposure to AsV for 2 h, the arsenate (AsV) and arsenite (AsIII) contents in AMF-roots increased by 1.8- and 3.6-fold, suggesting more efficient As uptake by P. vittata with AMF-roots. Plants take up and transport AsV via phosphate transporters (Phts). Here, for the first time, we identified a novel mycorrhiza-specific Pht transporter, PvPht1;6, from P. vittata . The transcripts of PvPht1;6 were strongly induced in AMF-roots, which were localized to the plasma membrane of arbuscule-containing cells. By complementing a yeast mutant lacking 5-Phts, we confirmed PvPht1;6's transport activity for both P and AsV. In contrast to typical AMF-inducible phosphate transporter LePT4 from tomato, PvPht1;6 showed greater AsV transport capacity. The results suggest that PvPht1;6 is probably critical for AsV transport at the periarbuscular membrane of P. vittata root cells, revealing the underlying mechanism of efficient As accumulation in P. vittata with AMF-roots.

Plants often up-regulate gene expression of P transporters under P deficiency, but down-regulate them under arsenic stress. Different from other P transporters, PvPht1;3 and PvPht1;4 expressions in As-hyperaccumulator Pteris vittata are up-regulated under P deficiency and As stress, showing strong transport capacity for both As and P. This study examined the mechanisms behind As-induced up-regulation of P transporters in P. vittata after exposing to 10-50 µM arsenate (AsV) for 14 d under hydroponics, with non-hyperaccumulator P. ensiformis as a control. Under As stress, P. vittata was more efficient in taking up both As and P than P. ensiformis, showing 48-84% more P content in the fronds and roots, leading to 18-79% greater biomass. Though As enhanced the P uptake by P. vittata, the inorganic P was reduced by 25-64% from 140-347 to 65-126 mg kg -1 . It is likely that, under As stress, more P was utilized by P. vittata to counter As toxicity, causing reduction in inorganic P. This was supported by As-induced conversion of inorganic P to phytate in P. vittata, with phytate-P being increased by 26-75% from 239-713 to 418-1221 mg kg -1 , maintaining internal low P levels. Under As-induced low inorganic-P conditions, the expressions of P transporters PvPht1;3 and PvPht1;4 were up-regulated by 1.4- and 2.7-fold in the roots, helping greater As and P uptake by P. vittata. Clearly, As-induced overexpression of P transporters in P. vittata roots plays a critical role in taking up both As and P, thereby increasing its efficiency in As-hyperaccumulation from contaminated media., (Copyright © 2022 Elsevier B.V. All rights reserved.)

As the staple food for Asian countries and with its ability in arsenic accumulation, rice consumption becomes a dominant pathway for As exposure to humans. Here, we collected 108 rice samples from local markets and online sources in 13 major rice-producing regions in China, and determined As contents, speciation and bioaccessibility in the samples. Total As contents were 25-327 μg kg -1 (averaging 120), showing regional differences, with Hunan province being greater than other provinces at 180 vs 110. In rice grains, inorganic As was the dominant species, being 39.9-88.5 (61.1 %), but all being within the Chinese standard at 200 μg kg -1 . Based on the modified physiologically-based extraction test (MPEBT), arsenic bioaccessibility in rice samples was 20.1-82.2 (52.3 %) in the gastric phase and 47.2-113 (81.2 %) in the intestinal phase. Strong positive correlation between total As and bioaccessible As suggested bioaccessible As was content-dependent. Based on the intestinal phase, the rice samples from northern region had lower As bioaccessibility than other regions (59.2 vs 83.2 %), and Japonica variety had lower As bioaccessibility than Indica variety (71.1 vs 83.1 %). This study suggests that rice from markets in China is safe, with their As contents and bioaccessibility showing regional and variety differences., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Contamination of drinking water by carcinogen arsenic (As) is of worldwide concern as its exposure poses potential threat to human health. As such, it is important to understand the mechanisms associated with As-induced toxicity to humans. The Nrf2/Keap1 signal pathway is one of the most important defense mechanisms in cells to counter oxidative stress; however, limited information is available regarding its role in countering As-induced stress in model animal mouse. In this study, we assessed the responses of Nrf2/keap1 pathway in mice after chronic exposure to As at environmentally-relevant concentrations of 10-200 μg L -1 for 30 days via drinking water. Our results indicate that chronic As exposure had limited effect on mouse growth. However, As induced oxidative stress to mice as indicated by increased content of malondialdehyde (MDA; 52-90%), an index of lipid peroxidation. Further, arsenic exposure reduced the activity of superoxide dismutase (SOD; 14-18%), an indication of reduced anti-oxidative activity. Besides, arsenic exposure increased MnSOD mRNA transcription by 25-66%, and decreased the mRNA transcriptions of Cu/ZnSOD by 72-83% and metallothionein by 16-75%, a cysteine-rich protein involved in metal detoxification. To counter arsenic toxicity, the expression of transcription factor for Nrf2 and Keap1 was increased by 2.8-8.9 and 0.2-8.1 fold in mice. To effectively reduce As-induced oxidative stress, the Nrf2/Keap1 transcription factor upregulated several downstream anti-oxidative genes, including heme oxygenase-1 (0.9-2.5 fold), glutamate-cysteine ligase catalytic subunit (0.6-1.7 fold), and NADH quinone dehydrogenase 1 (2.1-4.8 fold). This study shows the importance of Nrf2/Keap1 signaling pathway and associated anti-oxidative enzymes in countering As-toxicity in mice, possibly having implication for human health., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Antimony (Sb) and arsenic (As) are chemical analogs, but their behaviors in plants are different. To investigate the Sb uptake, translocation and speciation in As-hyperaccumulator P. cretica, a hydroponic experiment was conducted. In this study, P. cretica was exposed to 0.2-strength Hoagland nutrient solution, which contained 0.5 or 5 mg/L antimonite (SbIII) or antimonate (SbV). After 14 d exposure, P. cretica took up 1.4-2.8 times more SbIII than SbV. Since P. cretica was unable to translocate Sb, its roots accumulated >97% Sb with the highest at 7965 mg/kg. In both SbIII and SbV treatments, SbIII was the predominant species in P. cretica, with 90-100% and 46-100% SbIII in the roots. As the first barrier against Sb to enter plant cells, more Sb was accumulated in cell wall than cytosol or organelles. The results suggest that P. cretica may detoxify Sb by reducing SbV to SbIII and immobilizing it in root cell walls. Besides, the presence of SbIII significantly reduced the concentrations of dissolved organic C including organic acids in P. cretica root exudates. Further, increasing Sb levels promoted P accumulation in the plant, especially in the fronds, which may help P. cretica growth. The information from this study shed light on metabolic transformation of Sb in As-hyperaccumulators P. cretica, which helps to better understand Sb uptake and detoxification by plants., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Arsenic (As) and copper (Cu) are common co-contaminates in soils. However, their interactive effects on their accumulation and distribution in As-hyperaccumulator Pteris vittata are poorly understood. A hydroponic experiment was conducted with As being 0, 5, or 50 μM and Cu being 0.32, 3.2, or 32 μM to evaluate their phytotoxicity, accumulation, and distribution in P. vittata. In addition, As and Cu uptake kinetics were examined using the Michaelis-Menten kinetics model. Total As and Cu concentrations in P. vittata were up to 487 and 1355 mg kg -1 . About 39-81% of the As was in the fronds compared to 0.6-18% for Cu. At 50 μM As, increasing Cu concentration from 0.32 to 32 μM increased root As while decreasing frond As concentrations, with the translocation factor (ratio of As in fronds to roots) being reduced from 4.0 to 0.31. In contrast, As did not affect Cu accumulation in P. vittata. Michaelis constant K m value for As was higher than that of Cu (6.49-24.9 vs. 0.43-3.36), consistent with higher Cu uptake than As. Besides, Cu reduced root K but increased P levels in the roots, whereas As increased the K and P concentrations in the fronds. Our results suggest that P. vittata accumulated more Cu than As in the roots, contributing to its low As translocation. As such, high levels of Cu are likely to reduce As uptake by P. vittata during phytoremediation of As-contaminated sites., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The beneficial effects of selenium on As uptake and plant growth in As-hyperaccumulator Pteris vittata are known, but the associated mechanisms remain unclear. Here, we investigated the effects of selenate on arsenic accumulation by P. vittata under two arsenate levels. P. vittata plants were exposed to 13 (As 13 ) or 133 µM (As 133 ) arsenate and 5 µM selenate in 0.2-strength Hoagland solution. After 14 d of growth, plant biomass, Se and As content, As speciation, and malondialdehyde (MDA), glutathione reductase (GR), glutathione peroxidase (GPX), and glutathione (GSH and GSSG) levels were determined. The results show that selenate promoted P. vittata growth and increased As concentrations in the roots and fronds by 256% from 97 to 346 mg kg -1 and 142% from 213 to 514 mg kg -1 under As 13 treatment, and by 166% from 500 to 1332 mg kg -1 and 534% from 777 to 4928 mg kg -1 under As 133 treatment. In addition, selenate increased the glutathione content in P. vittata roots and fronds by 75-86% under As 13 treatment and 44-45% under As 133 treatment. Selenate also increased the GPX activity by 161-173%, and GR activity by 72-79% in P. vittata under As 13 and As 133 treatments. The HPLC-ICP-MS analysis indicated that selenate increased both AsIII and AsV levels in P. vittata, with AsIII/AsV ratio being lower in the roots and higher in the fronds, i.e., more AsIII was being translocated to the fronds. Taken together, our results suggest that, via GPX-GR mediated enhancement of GSH-GSSG cycle, selenate effectively increases plant growth and As uptake in P. vittata by improving AsV reduction in the roots and AsIII translocation from the roots to the fronds., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Developing P-efficient plants helps improve P uptake from soils with low-available P and reduce environmental damage by P runoff. Here, we investigated a novel root-specific phytase PvPHY1 from As-hyperaccumulator Pteris vittata, which can efficiently utilize phytate, a recalcitrant organic phosphorus in soil. Unlike other plants, expression of PvPHY1 in P. vittata was greater in the roots than the fronds. A pure phytase with considerable activity was obtained via prokaryotic expression. Expressing PvPHY1 in tobacco (PvPHY1-Ex) enhanced its growth (2.8 to 3.5-3.9 g per plant) and increased its P accumulation by 10-50% under low- and adequate-P conditions. Further, PvPHY1-Ex tobacco showed 25-32% lower intracellular phytate and 30-56% higher inorganic P in the roots, likely due to phytase-mediated hydrolysis of phytate. Decrease of phytate levels up-regulated phosphate transporter genes (NbPht1;1, NbPht1;2 and NbPht1;6), leading to greater P and As uptake. However, As translocation to the shoots was low, probably due to competition from increased inorganic P via phytate hydrolysis. As such, PvPHY1 facilitated P uptake from soils and phytate hydrolysis in plants, thereby promoting tobacco growth. Overall, PvPHY1 from P. vittata helps better understand the novel phytase to increase soil P utilization efficiency, thereby reducing P fertilizer requirements for crop production., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Understanding the distribution of hyperaccumulators helps to implement more efficient phytoremediation strategies of contaminated sites, however, limited information is available. Here, we investigated the geographical distribution of the first-known arsenic-hyperaccumulator Pteris vittata in China and the key factors under two climate change scenarios (SSP 1-2.6 and SSP 5-8.5) at two time points (2030 and 2070). Species distribution model (MaxEnt) was applied to examine P. vittata distribution based on 399 samples from field surveys and existing specimen records. Further, among 23 environmental factors, 11 variables were used in the MaxEnt model, including temperature, precipitation, elevation, soil property, and UV-B radiation. The results show that P. vittata can grow in ~23% of the regions in China. Specifically, it is mainly distributed in 11 provinces of southern China, including Hainan, Guangdong, Guangxi, Yunnan, Guizhou, Hunan, Hubei, Jiangxi, Fujian, Zhejiang, and Jiangsu. Besides, eastern Sichuan, and southern Henan, Shaanxi, and Anhui are suitable for P. vittata growth. Under two climate change scenarios, P. vittata distribution in China would decrease by ~5.76-7.46 × 10 4  km 2 in 2030 and ~3.22-4.68 × 10 4  km 2 in 2070, with southern Henan and most Jiangsu being unsuitable for P. vittata growth. Among the 11 environmental variables, the minimum temperature of coldest month (bio6) and temperature annual range (bio7) are the two key factors limiting P. vittata distribution. At bio6 <-5 °C and/or bio7 >33 °C, the regions are unsuitable for P. vittata growth. Based on the MaxEnt model, precipitation had limited effects, so P. vittata can probably survive under both dry and moist environments. This study helps guide phytoremediation of As-polluted soils using P. vittata and provides an example to evaluate habitat suitability of hyperaccumulators at international scales., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Arsenite (AsIII) antiporter ACR3 is crucial for arsenic (As) translocation and sequestration in As-hyperaccumulator Pteris vittata, which has potential for phytoremediation of As-contaminated soils. In this study, two new ACR3 genes PvACR3;2 and PvACR3;3 were cloned from P. vittata and studied in model organism yeast (Saccharomyces cerevisiae) and model plant tobacco (Nicotiana tabacum). Both ACR3s mediated AsIII efflux in yeast, decreasing its As accumulation and enhancing its As tolerance. In addition, PvACR3;2 and PvACR3;3 were expressed in tobacco plant. Localized on the plasma membrane, PvACR3;2 mediated both AsIII translocation to the shoots and AsIII efflux from the roots in tobacco, resulting in 203 - 258% increase in shoot As after exposing to 5 μM AsIII under hydroponics. In comparison, localized to the vacuolar membrane, PvACR3;3 sequestrated AsIII in tobacco root vacuoles, leading to 18 - 20% higher As in the roots and 15 - 36% lower As in the shoots. Further, based on qRT-PCR, both genes were mainly expressed in P. vittata fronds, indicating PvACR3;2 and PvACR3;3 may play roles in AsIII translocation and sequestration in the fronds. This study provides not only new insights into the functions of new ACR3 genes in P. vittata, but also important gene resources for manipulating As accumulation in plants for phytoremediation and food safety., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Animal bioassays have been developed to estimate oral relative bioavailability (RBA) of metals in soil, dust, or food for accurate health risk assessment. However, the comparability in RBA estimates from different labs remains largely unclear. Using 12 soil and soil-like standard reference materials (SRMs), this study investigated variability in lead (Pb) and arsenic (As) RBA estimates employing a mouse bioassay in 3 labs at Nanjing University, University of Jinan, and Shandong Normal University. Two performances of the bioassay at Nanjing University in 2019 and 2020 showed reproducible Pb and As RBA estimates, but increasing the number of mouse replicates in 2020 produced more precise RBA measurements. Although there were inter-lab variations in diet consumption rate and metal accumulation in mouse liver and kidneys following SRM ingestion due to differences in diet composition, bioassays at 3 labs in 2019 yielded overall similar Pb and As RBA estimates for the 12 SRMs with strong linear correlations between each 2 of the 3 labs for Pb (R 2  = 0.95-0.98 and slope = 0.85-1.02) and As RBA outcomes (R 2  = 0.46-0.86 and slope = 0.56-0.79). The consistency in RBA estimates was attributed to the relative nature of the final bioavailability outcome, which might overcome the inter-lab variation in diet consumption and metal uptake in mice. These results increased the confidence of use of mouse bioassays in bioavailability studies.

Rice consumption is a major dietary source of human exposure to arsenic (As), with As bioavailability being an important factor influencing its health risk. In this study, the As bioaccessibility was measured in 11 rice grains (140-335 μg As kg -1 ), which were compared to As relative bioavailability previously measured based on a mouse bioassay (Li et al., 2017). Using modified physiologically-based extraction test for rice (MPBET), As bioaccessibility in raw rice samples (44-88% in the gastric phase and 47-102% in the intestinal phase) was similar to those in cooked rice (42-73% and 43-99%). Arsenic bioaccessibility in rice was generally higher in the intestinal phase than in the gastric phase, with Fe and Ca concentrations in rice being negatively correlated with As bioaccessibility in the gastric phase (R 2  = 0.47-0.49). In addition, for cooked rice, strong positive correlation was observed between bioaccessible As and inorganic As (R 2  = 0.63-0.72), suggesting inorganic As in rice was easier to dissolve than organic As in gastrointestinal digestive fluids. Due to limited variation in As bioaccessibility and As bioavailability among the 11 samples, a weak correlation was observed between them (R 2  = 0.01-0.03); however, As bioaccessibility values measured by the gastric phase (GP) of the MPBET agreed with As bioavailability values based on a mouse bioassay, suggesting the potential of the MPBET GP to predict As bioavailability in rice. Future work is needed to ascertain the robustness of the MPBET GP in predicting As bioavailability in rice using additional samples., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Biochar is widely used as an amendment to optimize the composting process. In this study, we firstly investigated the effects of biochar amendment on methylation and volatilization of arsenic (As), and the microbial communities during manure composting. Biochar amendment was found to increase the concentrations of monomethylarsonic acid (MMA) and dimethylarsinic acid (DMA) during mesophilic (days 0-10) and early thermophilic (days 11-15) phases, and promote As volatilization during the maturing phase (days 60-80) of composting. In addition, the abundances of As(V) reductase (arsC) and As(III) S-adenosyl-L-methionine methyltransferase (arsM) genes were higher in the biochar treatment than that in the control. Moreover, biochar amendment influenced the microbial communities by promoting As methylation and volatilization via Ensifer and Sphingobium carrying arsC genes, and Rhodopseudomonas and Pseudomonas carrying arsM genes. This study emphasized the considerable role of biochar on methylation and volatilization of As during manure composting and provided an overall characterization of the community compositions of arsC and arsM genes during manure composting. It will broaden our insights in As biogeochemical cycle during manure composting with biochar amendment, which will facilitate the regulation of As during manure composting and its application in agricultural soil., (Copyright © 2021. Published by Elsevier B.V.)

Oral bioavailability of arsenic (As) determines levels of As exposure via ingestion of As-contaminated soil, however, the role of gut microbiota in As bioavailability has not evaluated in vivo although some in vitro studies have investigated this. Here, we made a comparison in As relative bioavailability (RBA) estimates for a contaminated soil (3913 mg As kg -1 ) using a mouse model with and without penicillin perturbing gut microbiota and metabolites. Compared to soil exposure alone (2% w/w soil in diets), addition of penicillin (100 or 1000 mg kg -1 ) reduced probiotic Lactobacillus and sulfate-reducing bacteria Desulfovibrio, enriched penicillin-resistant Enterobacter and Bacteroides, and decreased amino acid concentrations in ileum. With perturbed gut microbiota and metabolic profile, penicillin and soil co-exposed mice accumulated 2.81-3.81-fold less As in kidneys, excreted 1.02-1.35-fold less As in urine, and showed lower As-RBA (25.7-29.0%) compared to mice receiving diets amended with soil alone (56 ± 9.63%). One mechanism accounted for this is the decreased concentrations of amino acids arising from the gut microbiota shift which resulted in elevated iron (Fe) and As co-precipitation, leading to reduced As solubilization in the intestine. Another mechanism was conversion of bioavailable inorganic As to less bioavailable monomethylarsonic acid (MMA V ) and dimethylarsinic acid (DMA V ) by the antibiotic perturbed microflora. Based on in vivo mouse model, we demonstrated the important role of gut microbiota and gut metabolites in participating soil As solubilization and speciation transformation then affecting As oral bioavailability. Results are useful to better understand the role of gut bacteria in affecting As metabolism and the health risks of As-contaminated soils., (Copyright © 2021 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Global warming severely hinders both rice (Oryza sativa L.) quality and yield by increasing arsenic (As) bioavailability in paddy soils. However, details regarding As biotransformation and migration in the rice-soil system at elevated temperatures remain unclear. This study investigated the effects of increasing temperature on As behavior and translocation in rice grown in As-contaminated paddy soil at two temperature treatments (33 °C warmer temperature and 28 °C as control). The results showed that increasing temperature from 28 °C to 33 °C significantly favored total As, arsenite (As(III)) and arsenate (As(Ⅴ)) release into the soil pore-water. This increase in As bioavailability resulted in significantly higher As(III) accumulation in the whole grains at warmer treatment relative to the control. Moreover, the results suggest that increasing temperature to 33 °C promoted As(III) migration from the roots to the whole grains. Furthermore, the As(V)-reducing Xanthomonadales order and Alcaligenaceae family, and As(V) reductase-encoding arsC gene were enriched in the rhizosphere soils incubated at 33 °C. This suggests that the increase in As bioavailability in that treatment was due to enhanced As(V) reductive dissolution into the soil pore-water. Overall, this study provides new insights on how warmer future temperatures will exacerbate As accumulation in rice grains., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Arsenic (As) contamination in soils is of great concerns due to its toxicity to plants. As an analogue, phosphorus plays an important role in protecting plants from As toxicity. In this study, we identified a new phosphate transporter 2 (PHT2), PvPht2;1, from As-hyperaccumulator Pteris vittata and analyzed its functions in As and P transport in a yeast mutant, and model plant Arabidopsis thalian . PvPht2;1 contained 12 transmembrane domains, sharing high identity with PHT2 genes in diverse plants. Further, independent of external P or As levels, PvPht2;1 was mainly expressed in P. vittata fronds with the expression being 3-4 folds higher than that in the roots and rhizomes. Localized to the chloroplasts based on GFP-fused PvPht2;1 in model plant tobacco, PvPht2;1 functioned as a low-affinity P transporter. Under As exposure, PvPht2;1 yeast transformants showed comparable growth with the control while high-affinity P transporter PvPht1;3 transformants showed better growth, suggesting that PvPht2;1 transported P but slower than PvPht1;3 transporter. Expressing PvPht2;1 in A. thaliana increased its shoot P concentration without influencing its As accumulation. Further, the chloroplasts' P content in transgenic A. thaliana increased by 37-59% than wild-type (WT) plants. Under As exposure, the photosynthesis of PvPht2;1 -expressing A. thaliana remained stable but that of WT plants decreased. The data indicate that, under As stress, expressing PvPht2;1 in A. thaliana enhanced its P transport to the chloroplasts and protected its photosynthesis. In short, highly expressed in the fronds and not impacted by As exposure, chloroplast-located PvPht2;1 may have protected As-hyperaccumulator P. vittata from As toxicity by efficiently transporting only P to its chloroplasts.

Bioavailability and speciation of arsenic (As) are impacted by fertilization and bacteria in the rice rhizosphere. In this study, we investigated the effects of long-term manure application on As bioavailability, microbial community structure, and functional genes in a rice paddy field. The results showed that manure application did not affect total As in the soil but increased soluble As forms by 19%, increasing arsenite (As(III)) accumulation in rice grains and roots by 34 and 64% compared to a control. A real-time quantitative polymerase chain reaction (qPCR) and high-throughput sequencing analysis demonstrated that manure application increased the relative abundance of Rhizobium , Burkholderia , Sphingobium , and Sphingomonas containing arsenate reductase genes ( arsC ) in the rhizosphere soil, consistent with the 529% increase in arsC , which may have promoted arsenate (As(V)) reduction and increased As availability in pore water. In addition, manure application significantly altered the iron (Fe)-plaque microbial community structure and diversity. The microbes, particularly, Bradyrhizobium , Burkholderia , and Ralstonia , were mostly associated with As, Fe, and sulfur (S) cycles. This result was consistent with changes in the functional genes related to As, Fe, and S transformation. Although manure application promoted As(V) reduction ( arsC) in Fe-plaque by 682%, it inhibited Fe and S reduction by decreasing Fe III reduction bacteria (Geobacteraceae) and the sulfate-reducing gene ( dsrA ) abundance. Further, manure application changed the composition of the microbial community that contained the arsC gene. In short, caution needs to be excised even in the soil with a low As concentration as manure application increased As(III) accumulation in rice grains.

Arsenic-hyperaccumulator Pteris vittata is efficient in As absorption, reduction, and translocation. But the molecular mechanisms and locations of arsenate (AsV) reduction in P. vittata are still unclear. Here, we identified two new arsenate reductase genes from P. vittata, PvHAC1 and PvHAC2. Two PvHAC genes encoded a rhodanase-like protein, which were localized in the cytoplasm and nucleus. Both recombinant Escherichia coli strains and transgenic Arabidopsis thaliana lines showed arsenate reductase ability after expressing PvHAC genes. Further, expressing PvHAC2 enhanced As tolerance and reduced As accumulation in A. thaliana shoots under AsV exposure. Based on expression pattern analysis, PvHAC1 and PvHAC2 were predominantly expressed in the rhizomes and fronds of P. vittata. Different from those of HAC homologous genes in non-hyperaccumulators, little PvHAC was expressed in the roots. Besides, PvHAC1 expression was strongly upregulated under AsV exposure but not AsIII. The data suggest that arsenate reductase PvHAC1 in the rhizomes coupled with arsenate reductase PvHAC2 in the fronds of P. vittata played a critical role in As-hyperaccumulation by P. vittata, which helps to further improve its utility in phytoremediation of As-contaminated soils., (Published by Elsevier B.V.)

Rice plant is efficient in arsenic (As) accumulation due to enhanced soil As release under flooded condition and its effective As uptake. Therefore, rice plant can be used to remove bioavailable As from paddy soil. In this study, the depleting dynamics of soil As with rice growth was mapped two-dimensionally with the zirconium-oxide diffusive gradients in thin films (DGT) technique. Further, the key biochemistry promoting soil As bioavailability was studied to better understand the underlying processes. Results revealed that the average DGT-As encompassing the root zone decreased steeply from 331 in the seedling stage to 136 in the heading & flowering stage and further to 118 μg l -1 at harvest, which was 26% lower than that of the control at 160 μg l -1 . During this process, rhizosphere porewater As developed a dynamic profile similar to Fe and dissolved organic carbon, with the diversity of arrA gene peaking at heading & flowering stage. The data support soil As release from microbial reduction of Fe hydroxides fueled by root exudation as carbon source. Arsenic was mainly accumulated in the roots, accounting for up to 95% of total As in rice plants. Removal of rice roots resulted in ~19% lower DGT-As in post-harvest soil compared to without removing the roots. As a result, a sharp decline in As accumulation in rice plants was obtained in the second planting after removing one crop of rice roots. The results highlight that rice, as a paddy-adapted plant, is effective in As uptake in the roots, and thereby removing rice roots efficiently depletes bioavailable As from paddy soils., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Arsenic (As) and cadmium (Cd) are ubiquitous in the environment and they are both toxic to humans. When present in soils, they can enter food chain, thereby threatening human health. Water spinach (Ipomoea aquatica) is an important leafy vegetable, which is widely consumed in Asian countries. However, it is efficient in taking up As and Cd from soils and accumulating them in the edible parts. Therefore, it is of significance to reduce its As and Cd content, especially in contaminated soil. In this study, pot experiments were conducted to investigate the ability of As-hyperaccumulator Pteris vittata in reducing As and Cd uptake by water spinach under different phosphorus treatments. P. vittata was grown for 60 d in a contaminated-soil amended with P fertilizer (+P) or phosphate rock (+PR), followed by water spinach cultivation for another 30 d. Plant biomass, As and Cd contents in plants and soils, and soil pH were analyzed. We found that, P. vittata coupled with PR decreased the As concentration in water spinach shoots by 42%, probably due to As uptake by P. vittata. Moreover, P. vittata decreased the Cd accumulation in water spinach by 24-44%, probably due to pH increase of 0.47-0.61 after P. vittata cultivation. Taking together, the results showed that P. vittata coupled with PR decreased the As and Cd content in water spinach, which is of significance for improving food safety and protecting human health., (Published by Elsevier Ltd.)

Behavior of trace elements in flooded/lowland rice soils is controlled by root-zone iron oxidation. Insoluble iron species bind/capture toxic elements, i.e., arsenic. However, it was recently observed that within this territory of arsenic immobilization lies a zone of prolific iron release, accompanied by a significant flux of arsenic in close proximity to rice root apices. Questions still remain on how common this phenomenon is and whether the chemical imaging approaches or soils/cultivars used influence this event. Here, three types of ultrathin/high-resolution diffusive gradient in thin films (DGT) substrates were integrated with oxygen planar optodes in a multilayer system, providing two-dimensional mapping of solute fluxes. The three DGT approaches revealed a consistent/overlapping spatial distribution with localized flux maxima for arsenic, which occurred in all experiments, concomitant with iron mobilization. Soil/porewater microsampling within the rhizosphere revealed no significant elevation in the solid phase's total iron and arsenic concentrations between aerobic and anaerobic zones. Contrary to arsenic, phosphorus bioavailability was shown to decrease in the arsenic/iron flux maxima. Rice roots, in addition to their role in nutrient acquisition, also perform a key sensory function. Flux maxima represent a significant departure from the chemical conditions of the bulk/field environment, but our observations of a complete rhizosphere reveal a mixed mode of root-soil interactions.

Organoarsenicals have been used in poultry production for years, however, studies focused on roxarsone (ROX), with little attention to p-arsanilic acid (ASA). We assessed arsenic (As) concentration and speciation in chicken meat collected from 10 cities in China. The geometric mean for total As in 249 paired raw and cooked samples was 4.85 and 7.27 μg kg -1 fw, respectively. Among 81 paired raw and cooked samples, ASA and ROX were detected in >90% samples, suggesting the prevalence of organoarsenical use in China. ASA contributed the most (45% on average) to total As in cooked samples, followed by As(V), DMA, As(III), and ROX (7.2-22%). ASA was found to contribute more to total As in chicken meat compared to ROX for the first time. Arsenic in chicken meat showed considerable geographic variation, with higher inorganic arsenic (iAs) being detected from cities with higher ROX and ASA, indicating that organoarsenical use increased iAs concentration in chicken meat. When health risk was estimated, dietary exposure to iAs would result in an increase of 3.2 bladder and lung cancer cases per 100,000 adults. The result supports the removal of organoarsenicals in poultry production from Chinese market and further supports its removal from the global markets., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Arsenic-hyperaccumulator Pteris vittata is efficient in As uptake, probably through phosphate transporters (Pht). Here, for the first time, we cloned a new PvPht1;4 gene from P. vittata and investigated its role in arsenate (AsV) uptake and transport in yeast and transgenic tobacco plants. On the basis of quantitative real-time polymerase chain reaction (qRT-PCR), PvPht1;4 was abundantly expressed in P. vittata fronds and roots, with its transcripts in the roots being induced by both P deficiency and As exposure. PvPht1;4 was localized to the plasma membrane, which complemented a yeast-mutant defective in P uptake and showed higher P transport affinity than PvPht1;3. Under AsV exposure, PvPht1;4 yeast transformants showed comparable tolerance as PvPht1;3, but higher As accumulation than PvPht1;2 transformants, indicating that PvPht1;4 had considerable AsV and P transport activity. However, in soil and hydroponic experiments, PvPht1;4 expressing tobacco lines accumulated 26-44 and 37-55% lower As in the shoots than wild type plants, with lower root-to-shoot As translocation. In the roots of PvPht1;4 lines, higher glutathione (GSH) contents and expression levels of GSH synthetase gene NtGSH2 were observed. In addition, the transcripts of AsIII-GSH transporter NtABCC1 in PvPht1;4 lines were upregulated. The data suggested that PvPht1;4 lines probably detoxified As by reducing AsV to AsIII, which was then complexed with GSH and stored in the root vacuoles, thereby reducing As translocation in transgenic tobacco. Given its strong AsV transport capacity, expression of PvPht1;4 provides a new molecular approach to reduce As accumulation in plant shoots.

Soils are often co-contaminated with As, Pb, and Cd. To what extent ingested metal(loid)s interact with each other in the gastrointestinal tract and influence their RBA (relative bioavailability) is largely unknown. Three soils predominantly contaminated with As (MS, mining/smelting impacted), Pb (WR, wire rope production impacted), and Cd (EP, enamel pottery production impacted) were administered to mice individually or in binary and tertiary combinations with sodium arsenate, Cd chloride, and/or Pb acetate. In binary combinations, ∼10-fold higher Pb addition decreased As-RBA in MS (26.0 ± 6.28% to 17.1 ± 1.08%), while ∼10-fold higher As addition decreased Pb-RBA in WR (61.3 ± 2.41% to 28.8 ± 5.45%). This was possibly due to the formation of insoluble Pb arsenate in mouse intestinal tract, as indicated by the formation of precipitates when As and Pb co-occurred in water or simulated human gastrointestinal fluids. Due to competition for shared absorption transporters, ∼10- and 100-fold higher Pb addition decreased Cd-RBA in EP (95.8 ± 12.9% to 67.8 ± 12.8% and 62.8 ± 8.24%). Tertiary combinations showed that interactions between two metal(loid)s were affected by the presence of the third metal(loid). Our study suggests that As oxyanion could interact with Pb or Cd ions in the mouse gastrointestinal tract, and the interactions vary depending on concentration and solution characteristics.

Microbial arsenic transformation is important in As biogeochemical cycles in the environment. In this study, a new As-resistant bacterial strain Leclercia adecarboxylata As3-1 was isolated and its associated mechanisms in As resistance and detoxification were evaluated based on genome sequencing and gene annotations. After subjecting strain As3-1 to medium containing arsenate (AsV), AsV reduction occurred and an AsV-enhanced bacterial growth was observed. Strain As3-1 lacked arsenite (AsIII) oxidation ability and displayed lower AsIII resistance than AsV, probably due to its higher AsIII accumulation. Polymerase chain reaction and phylogenetic analysis showed that strain As3-1 harbored a typical AsV reductase gene (arsC) on the plasmids. Genome sequencing and gene annotations identified four operons phoUpstBACS, arsHRBC, arsCRDABC and ttrRSBCA, with 8 additional genes outside the operons that might have involved in As resistance and detoxification in strain As3-1. These included 5 arsC genes explaining why strain As3-1 tolerated high AsV concentrations. Besides ArsC, TtrB, TtrC and TtrA proteins could also be involved in AsV reduction and consequent energy acquisition for bacterial growth. Our data provided a new example of diverse As-regulating systems and AsV-enhanced growth without ArrA in bacteria. The information helps to understand the role of As in selecting microbial systems that can transform and utilize As., (Published by Elsevier B.V.)

Arsenic-hyperaccumulator Pteris vittata is efficient in As accumulation and has been used in phytoremediation of As-contaminated soils. Arsenate (AsV) is the predominant As species in aerobic soils and is taken up by plants via phosphate transporters (Pht) including P. vittata . In this work, we cloned the PvPht1;3 full length coding sequence from P. vittata and investigated its role in As accumulation by yeast and plants. PvPht1;3 complemented a yeast P uptake mutant strain and showed a stronger affinity and transport capacity to AsV than PvPht1;2. In transgenic tobacco, PvPht1;3 enhanced AsV absorption and translocation, increasing As accumulation in the shoots under both hydroponic and soil experiments. On the basis of the expression patterns via qRT-PCR, PvPht1;3 was strongly induced by P deficiency but not As exposure. To further understand its expression pattern, transgenic Arabidopsis thaliana and soybean expressing the GUS reporter gene, driven by PvPht1;3 promoter, were produced. The GUS staining showed that the reporter gene was mainly expressed in the stele cells, indicating that PvPht1;3 was expressed in stele cells and was likely involved in P/As translocation. Taken together, the data suggested that PvPht1;3 was a high-affinity AsV transporter and was probably responsible for efficient As translocation in P. vittata . Our results suggest that expressing PvPht1;3 enhances As translocation and accumulation in plants, thereby improving phytoremediation of As-contaminated soils.

Rice ( Oryza sativa ) is a major food crop in the world, feeding half of the world's population. However, rice is efficient in taking up toxic metalloid arsenic (As), adversely impacting human health. Among different As species, inorganic As is more toxic than organic As. Thus, it is important to decrease inorganic As in rice to reduce human exposure from the food chain. The arsenite (AsIII) antiporter gene PvACR 3;1 from As-hyperaccumulator Pteris vittata decreases shoot As accumulation when heterologously expressed in plants. In this study, three homozygous transgenic lines (L2, L4, and L7) of T3 generation were obtained after transforming PvACR 3;1 into rice. At 5 μM of AsIII, PvACR 3;1 transgenic rice accumulated 127%-205% higher As in the roots, with lower As translocation than wild type (WT) plants. In addition, at 20 μM of AsV, the transgenic rice showed similar results, indicating that expressing PvACR3;1 increased As retention in the roots from both AsIII and AsV. Furthermore, PvACR 3;1 transgenic rice plants were grown in As-contaminated soils under flooded conditions. PvACR3;1 decreased As accumulations in transgenic rice shoots by 72%-83% without impacting nutrient minerals (Mn, Zn, and Cu). In addition, not only total As in unhusked rice grain of PvACR 3;1 transgenic lines were decreased by 28%-39%, but also inorganic As was 26%-46% lower. Taken together, the results showed that expressing PvACR3;1 effectively decreased both total As and inorganic As in rice grain, which is of significance to breed low-As rice for food safety and human health.

The robustness of in vitro bioaccessibility assays to predict oral relative bioavailability (RBA) of multiple metals in contaminated soils requires validation using additional soil samples. In this study, 11 contaminated soils from mining/smelting areas were analyzed for As-, Cd-, and Pb-RBA using a mouse bioassay and metal bioaccessibility via the UBM gastric phase assay. Metal-RBA varied considerably among soils, with As-RBA (2.5-23%, mean 12%) being generally lower than Cd-and Pb-RBA (3.4-88 and 3.3-59%, mean 42 and 28%), due to higher proportions of As in the residual fractions. Metal-RBA generally decreased with increasing metal concentrations probably due to reduced labile metal fractions. In addition, strong negative correlations were observed between total Fe with As-, Cd-, and Pb-RBA (R 2  = 0.46-0.77), suggesting the role of Fe in controlling metal-RBA in soils. Like RBA, metal bioaccessibility by the UBM assay also varied among samples. However, strong in vivo-in vitro correlations (IVIVCs) were observed between metal-RBA and bioaccessibility (R 2  = 0.52-0.81). Further, there were little differences when As-, Cd-, and Pb-IVIVCs established using soils from this study and soils pooled from literature were compared, suggesting the robustness of the UBM assay to predict metal-RBA in contaminated soils., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The lack of efficient methods to treat As-rich biomass is a drawback for phytoremediation technology. In this study, we applied anaerobic digestion to reduce biomass and remove As from As-rich Pteris vittata biomass. P. vittata biomass including control (3.1 mg kg -1 As) and As-rich (2665 mg kg -1 As), together with positive and negative controls, was anaerobically digested at 35 °C for 35 d. Arsenic partitioning among gas, liquid and solid phases after anaerobic digestion was determined. Methane index potential assay was used to assess methane yields whereas liquid-displacement method was used to measure methane gas production. After 35 d, As partitioning in the liquid, solid and gas phases was 79, 30 and 1%, respectively. Besides, volatile solid was decreased from 91 to 12-17% total solid, while P. vittata biomass was decreased by 73-83%. Moreover, anaerobic digestion solubilized 76% As from P. vittata biomass, with 90% soluble As at 4.95 mg L -1 being recovered by As-Mg precipitation. Finally, methane production after 35 d was 197-212 L N CH 4 /kg volatile solid, showing slight As inhibition. Effective As removal from P. vittata biomass prior to disposal can improve the phytoremediation process., (Published by Elsevier Ltd.)

Improper disposal of arsenic-rich biomass and the lack of efficient methods to treat it may cause contamination in the environment. We developed an efficient method for arsenic (As) removal and biomass reduction of As-rich biomass of the As-hyperaccumulator Pteris vittata by coupling ethanol extraction with anaerobic digestion. This study assessed As partitioning among the three phases (gas, liquid and solid) after anaerobic digestion of P. vittata biomass. Biomass with and without As was first extracted with ethanol. Ethanol extraction removed ~93% As, with remaining As concentration at 197 mg kg -1 . The extracted biomass was then digested at 35 °C under anaerobic conditions for 35 d. Arsenic in the digested biomass was reduced by 89%, with remaining As concentration at 60 mg kg -1 . In addition, anaerobic digestion reduced the biomass by 64-71% and decreased the volatile solids content from 94 to 15-18%. Methane production was 145 and 160 L N CH 4 /kgVS after 35 d for As-rich and control biomass, respectively. As a final step, As concentration in anaerobic digestate supernatant was reduced to 0.26 mg L -1 by As-Mg precipitation. Overall, coupling ethanol extraction with anaerobic digestion decreased As concentration in P. vittata biomass from 2665 to 60 mg kg -1 , or by 98%. At this level (<100 mg As kg -1 ), P. vittata biomass can be considered a safe material based on USEPA regulations. Effective As removal from P. vittata biomass prior to disposal improves the phytoremediation process and lowers biomass transport and landfill disposal costs., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Arsenate (AsV) reduction in bacteria is essential to alleviate their arsenic (As) toxicity. We isolated a Bacillus strain PVR-YHB1-1 from the roots of As-hyperaccumulator Pteris vittata. The strain was efficient in reducing AsV to arsenite (AsIII), but the associated mechanisms were unclear. Here, we investigated its As resistance and reduction behaviors and associated genes at genome level. Results showed that the strain tolerated up to 20 mM AsV. When grown in 1 mM AsV, 96% AsV was reduced to AsIII in 48 h, with its AsV reduction ability being positively correlated to bacterial biomass. Two ars operons arsRacr3arsCDA and arsRKacr3arsC for As metabolisms were identified based on draft genome sequencing and gene annotations. Our data suggested that both operons might have attributed to efficient As resistance and AsV reduction in PVR-YHB1-1, providing clues to better understand As transformation in bacteria and their roles in As transformation in the environment., (Published by Elsevier Ltd.)

Mushrooms accumulate arsenic (As), yet As concentrations, speciation, and localization in cultivated mushrooms across a large geographic distribution are unknown. We characterized 141 samples of nine species from markets in nine capital cities in China, with samples of Lentinula edodes, Pleurotus ostreatus, and Agaricus bisporus being analyzed for As speciation and localization. Total As concentrations ranged from 0.01 to 8.31 mg kg -1 dw, with A. bisporus (0.27-2.79 mg kg -1 ) containing the most As followed by P. ostreatus and L. edodes (0.04-8.31 and 0.12-2.58 mg kg -1 ). However, As in A. bisporus was mostly organic including nontoxic arsenobetaine, while P. ostreatus and L. edodes contained mainly inorganic As (iAs). On the basis of in situ imaging using LA-ICP-MS, As in L. edodes was localized to the surface coat of the cap, while As in P. ostreatus was localized to the junction of the pileus and stipe. When As speciation and daily mushroom consumption (1.37 g d -1 dw) are considered, daily mushroom consumption may result in elevated iAs exposure, with increased bladder and lung cancer rates up to 387 cases per 100000. Our study showed that market mushrooms could be a health risk to the general public so its production should be monitored.