Volume 7 - 2016 | https://doi.org/10.3389/fpls.2016.01664 Many arable lands have accumulated large reserves of residual phosphorus (P) and a relatively large proportion of soil P is less available for uptake by plants Root released organic anions are widely documented as a key physiological strategy to enhance P availability while limited information has been generated on the contribution of rhizosphere organic anions to P utilization by crops grown in agricultural soils that are low in available P and high in extractable Ca We studied the role of rhizosphere organic anions in P uptake from residual P in four common crops Triticum aestivum and Brassica napus in low- and high-P availability agricultural soils from long-term fertilization field trials in a mini-rhizotron experiment with four replications Malate was generally the dominant organic anion More rhizosphere citrate was detected in low P soils than in high P soil napus showed 74–103% increase of malate in low P loam sativa had the greatest rhizosphere citrate concentration in all soils (5.3–15.2 μmol g−1 root DW) sativa also showed the highest level of root colonization by arbuscular mycorrhizal fungi (AMF; 36 and 40%) the greatest root mass ratio (0.51 and 0.66) in the low-P clay loam and loam respectively and the greatest total P uptake (5.92 mg P/mini-rhizotron) in the low-P loam napus had 15–44% more rhizosphere acid phosphatase (APase) activity ~0.1–0.4 units lower rhizosphere pH than other species the greatest increase in rhizosphere water-soluble P in the low-P soils and the greatest total P uptake in the low-P clay loam Shoot P content was mainly explained by rhizosphere APase activity water-soluble P and pH within low P soils across species P uptake was mainly linked to rhizosphere water soluble P The effects of rhizosphere organic anions varied among species and they appeared to play minor roles in improving P availability and uptake Improving the utilization of residual P accumulated in agricultural soils would help to reduce P fertilizer application and environmental stress studies on how to mobilize the less-available residual P and improve P-acquisition efficiency are needed The above reports suggest that further study is necessary to elucidate the role of root exudates in mobilizing plant less-available P Solanum tuberosum and Brassica napus) to investigate the contribution of root-exuded organic anions to improving P uptake in agricultural soils low in P availability For these four crops grown in three soils obtained from long-term fertilization field plots in Norway we addressed three hypotheses: (1) Low P availability will stimulate plant roots to release more organic anions and APase to rhizosphere soil; (2) the amounts of rhizosphere organic anions and APase will have positive correlations with rhizosphere plant-available P fractions and P uptake by plants in low P soils; (3) different crops will show differences in root released organic anions and APase in terms of using residual P from agricultural soils The ultimate goal of this study is to increase understanding of the contribution of root-released organic anions and APases to P uptake in low P availability agricultural soils in common crops The soils for the rhizotron experiment were collected from the plow layer (0–20 cm) of a clay loam and a loam of two long-term fertilization trials in southeastern Norway (Kristoffersen and Riley, 2005) 36% sand) was collected from field plots at Ås soil ALP) kg P ha−1 year−1 as single superphosphate since 1966 Both P treatments received 100 kg N ha−1 year−1 as calcium nitrate and 100 kg K ha−1 year−1 as potassium chloride 52% sand) was collected from a field in Møystad which had only received nitrogen (N) and potassium (K) fertilizers since 1922 (100 kg N ha−1 year−1 as calcium nitrate and 120 kg K ha−1 year−1 as potassium chloride) PAL-values below 30 mg kg−1 are considered low whilst those above 140 mg kg−1 are considered very high This standard was used in the present study The pH of soils AHP and ALP was adjusted from 5.4 and 5.0 respectively to ~6.5 by adding moderate amounts of CaCO3before use No P fertilizers were applied to the soils but all other basic nutrients were provided as follows and MgCl2 powders were mixed thoroughly with the soils assuring a homogeneous distribution of nutrients in the soils and then applied to each pot to ensure that plant growth was not limited by these micronutrients Canola (B. napus cv. MARIE), wheat (T. aestivum cv. AINO), oat (A. sativa cv. BELINDA), and micropropagated seedlings of potato (S. tuberosum cv. PIMPERNEL) were grown in 20 cm × 20 cm × 1 cm mini-rhizotrons consisting of Plexiglas plates (James et al., 1985) The experiment was conducted in a greenhouse with 18°C/15°C day/night temperature with a 16 h photoperiod at a light intensity of 200 ± 20 μmol m−2 s−1 and 50–75% relative humidity After filling each mini-rhizotron with ~0.5 kg fertilized homogenized soils water was added to achieve a soil moisture level of 25% (w/w) Mini-rhizotrons were wrapped in black plastic bags to avoid light exposure Five surface-sterilized seeds were sown in each mini-rhizotron and three uniform seedlings were kept in each mini-rhizotron after germination three tissue culture-derived seedlings of around 10 cm height were used There were four replicates and one mini-rhizotron without plant for each soil was set as control-unplanted bulk soil Deionized water (15–40 mL) was given daily to keep the soil surface moist the plants were irrigated according to weight loss and the position of the mini-rhizotrons was changed randomly every week Three plants in each mini-rhizotron were harvested 5 weeks after germination and no potato root tuber was produced during the 5-week experiment. At harvest, the intact plants were carefully removed from the mini-rhizotrons and divided into shoots and roots. The roots were first shaken slightly to remove excess soil. The soil remaining attached to the roots was defined as rhizosphere soil (e.g., Veneklaas et al., 2003) about 30 g rhizosphere fresh soil was carefully sampled using tweezers and spoons The rhizosphere soil was divided into two groups one air-dried for analysis of rhizosphere soil pH and another stored at −20°C for microbial P immobilization and soil enzyme activity measurements The collected rhizosphere extracts were immediately frozen and stored at −20°C until analysis with liquid chromatography triple quadrupole mass spectrometry (LC–MS/MS The root systems were then washed thoroughly to remove remaining soil and sub-sampled for AMF detection The remaining extract containing the rhizosphere soil in the container was centrifuged at 4000 rpm and the supernatant transferred to a new container The container with soil was then placed in a 65°C oven for 2 weeks The rhizosphere soil inside the container was then weighed Before analysis with LC-MS/MS, 910 μL extract was taken out of each sample to a separate vial, 50 μL deuterium-labeled succinic acid (0.2 μg) were added to be used as an internal standard (IS), and each vial was acidified with 40 μL concentrated formic acid. The LC-MS/MS analysis was performed as described previously (Wang et al., 2015) The concentrations were determined by comparison with their standard concentration measurements and further calculated based on the root dry weight or rhizosphere soil dry weight ten 1-cm pieces were randomly selected and five fields of vision were examined in each 1-cm root section at 100x microscopy; thus 50 fields of vision were examined for each sample The colonization percentage was calculated as the ratio of the colonized sections to the total sections examined Identification of AMF was based on observations of arbuscules and roots where only intraradical or extraradical hyphae or vesicles were observed were defined as non-AMF Shoots and roots were dried at 65°C for 48 h, and dry weight (DW) was measured. Root mass ratio was calculated as the ratio of root DW to the total plant DW. Shoot and root P concentrations were determined by inductively coupled plasma atomic emission spectroscopy (AtomComp 1100, Thermo Jarrell-Ash, MA, USA) according to Ogner et al. (1999) after digestion in a mixture of 65% (v/v) HNO3/72% (v/v) HClO4 (5: 1 Shoot and root P contents were calculated by P concentrations × shoot or root DW R software (version 3.2.3) was used for data analyses Two-way ANOVAs were used to study main effects of soil and their interaction on all parameters involved in this study followed by pairwise Tukey's honest significant difference tests for multiple comparisons along with the minimum significant difference (MSD) at p < 0.05 (presented in figure captions) Simple linear regressions were used to estimate the relationships among response variables linear regressions were made across species linear regressions were made across either similar soil properties (AHP and ALP) or similar P availability (ALP and B) No P was applied in our system. In order to confirm that P was the limiting factor for plant growth in low P soils, we calculated the N:P ratio in aboveground plant tissues (Table 2 and Figure 1). The N:P ratio varied from 6.9 to 10.5, 15.3 to 25.8, and 13.5 to 21.6 when soils AHP, ALP, and B were used as growth medium, respectively. According to van Duivenbooden et al. (1995) the N:P ratio of agricultural crops is in general between 6 and 8 and plants can be diagnosed as P limited when the N:P ratio is above 14 our soils ALP and B were P deficient and soil AHP was P sufficient Significance of a two-way ANOVA analysis for measured parameters N:P ratios of the aboveground tissues of Brassica napus and Solanum tuberosum grown in soils AHP (white bars) There was a significant interaction between species and soils for N:P ratio (p < 0.001 Shoot and total dry biomass varied significantly among soils and species (Table 2 and Figures 2A,B). All crops showed significant decrease of both shoot and total biomass in soil ALP, compared with soil AHP; however, crops grown in soil B showed almost the same total biomass as those grown in soil AHP, except T. aestivum (Figure 2) Dicots had similar shoot biomass in soil AHP and soil B while monocots had lower shoot biomass in soil B napus accumulated the most shoot biomass in all soil types (average 2.81 g tuberosum accumulated the least shoot biomass (average 1.27 g sativa had similar amounts of shoot biomass in all soils and (C) root mass ratio (dry matter basis) of Brassica napus There was a significant interaction between species and soils for shoot dry weight (p < 0.001 MSD 0.05 = 0.48 g) and root mass ratio (p < 0.01 Different root mass ratio (root DW: total DW) patterns were observed in the four crops (Figure 2C) napus showed a significant increase of root mass ratio in soil ALP and soil B aestivum did not show any significant differences in root mass ratios between soils and S tuberosum showed a decrease of root mass ratio by 19% when plants were grown in soil ALP compared with AHP but showed about the same value when plants were grown in soil B and soil AHP Shoot and root P concentrations varied significantly, and were unsurprisingly highest when plants were grown in soil AHP for all crops (Table 2 and Figures 3A,B) tuberosum had the highest shoot and root P concentrations napus had the lowest shoot and root P concentrations in soils ALP and B the shoot P concentrations in soil B were equal (S whereas the root P concentrations in soil B tended to be higher than in soil ALP with the exception of S and (D) P content ratio between soils ALP/AHP for shoot P (white bars) There was a significant interaction between species and soils for shoot P concentrations (p < 0.01 and root P concentrations (p < 0.001 and (C) rhizosphere total organic anion concentrations of Brassica napus and B (gray bars) based on root dry weight There was a significant interaction between species and soils for rhizosphere malate concentrations (p < 0.05 MSD 0.05 = 11.47 μmol g−1 DW roots) In soils AHP and ALP (with the same soil texture but different P availability) a significant negative linear correlation was found between the amount of citrate in the rhizosphere and the shoot P concentration for B a positive correlation was found between the amount of rhizosphere malate and shoot P concentration for S The pH of the rhizosphere varied significantly among species and soils (Table 2 and Figure 5) All species generally had a rhizosphere water extract pH between 5.5 and 6.3 aestivum had the highest rhizosphere pH (6.1) while S There was a significant interaction between species and soils for rhizosphere pH (p < 0.001 Dashed lines indicate the soil pH values in unplanted mini-rhizotrons The plant-available P (determined as PAL) in the rhizosphere increased slightly in soil B but decreased in soil ALP in all species, compared with bulk soils (Figure 6A) aestivum rhizosphere increased slightly in soil AHP (A) Rhizosphere AL-extractable P and (B) rhizosphere water soluble P of Brassica napus as well as changes compared to unplanted bulk soils (black diamonds) There was a significant interaction between species and soils for change in PAL (p < 0.05 and change in rhizosphere water soluble P (p < 0.01 There was a significant interaction between species and soils for acid phosphatase activities (p < 0.001 MSD 0.05 = 153 nmol MUF g−1 DM−1h−1) Phosphorus content immobilized by microbial biomass (Pmic) of bulk soil There was a significant interaction between species and soils for Pmic (p < 0.01 For mycorrhizal plant species, non-AMF were more abundant than AMF in roots (Table 3) Neither non-AMF nor AMF were influenced by soil P availability for any of the three species The percentage of root length colonized differed among the plant species in all soils sativa had the highest colonization of both AMF (32–40%) and non-AMF (58–75%) and colonization was lowest in S tuberosum (22–36% of non-AMF and no AMF were detected) The percentage of root length colonized by arbuscular mycorrhizal fungi (AMF) and non-arbuscular mycorrhizal fungi (non-AMF) in mycorrhizal species In within-soil analyses, significant correlations were found in low P availability soils ALP and B (Table 4) Plant shoot P content showed strong correlations with rhizosphere water-soluble P Total P uptake had weak positive correlations with rhizosphere APase and water-soluble P in soil ALP shoot P content and total P content showed strong positive correlations with root mass ratio in soil B Positive correlations between rhizosphere citrate concentration and root P content were found in soil ALP and soil B and rhizosphere malate correlated weakly with root P content in soil ALP rhizosphere water-soluble P showed strong correlations with rhizosphere APase and pH A weak correlation between rhizosphere malate concentration and water-soluble P was also found in soil B Rhizosphere APase correlated negatively with rhizosphere pH Correlations within low P availability soils ALP and B across species within each species across all three soils plant P uptake had a strong positive correlation with rhizosphere WSP for all species and root mass ratio explained the WSP and P uptake in B APase and pH while rhizosphere APase was linked to root mass ratio P mobilization and uptake was mainly explained by rhizosphere citrate and pH plant P uptake was mostly explained by rhizosphere APase and rhizosphere WSP (data not shown) for all species Rhizosphere pH also correlated significantly with rhizosphere APase and WSP for all species rhizosphere APase showed significant correlation with root mass ratio in A The only significant correlation between total P uptake and root mass ratio was found in A The results of this experiment suggested that rhizosphere organic anions made a minor contribution to P mobilization and uptake for the studied crops in the studied low P clay loam and loam soils Different plant species may have different growth potential in a mini-rhizotron and different needs for P to support their development and pH were likely to affect P availability and uptake rhizosphere WSP appears to have contributed greatly to P uptake in both high and low P availability soils possibly due to greater rhizosphere APase activities and lower rhizosphere pH sativa was another crop that could use P efficiently in our study possibly due to its greater root mass ratio and higher percentage of root colonizing AMF sativa had a larger proportion of root length colonized by AMF than T The implications of these findings and other points of interest are discussed below it is very hard to extract all the rhizosphere organic anions and to perform accurate analytical determination not all the rhizosphere soil in this study was taken to extract organic anions due to other soil measurements our data probably underestimated organic anion concentrations but may still reflect the relative differences between the plant species in soils AHP and ALP (soils with the same soil texture but different P availabilities) tuberosum had negative correlations with rhizosphere citrate concentration a single trait like larger amounts of rhizosphere organic anions did not result in improved P uptake and other factors such as root morphology (which we did not investigate) and pH may play key roles hypothesis (2) that rhizosphere organic anions correlate with P availability and uptake was not supported our hypothesis (2) that APase correlates with P availability and uptake and hypothesis (3) were supported Further study is needed to prove whether APase can hydrolyze soil organic P raised the rhizosphere pH by 0.2–0.7 pH units and was associated with higher rhizosphere Colwell P (bicarbonate-extractable P) which had a higher coverage percent of root colonizing AMF had higher rhizosphere pH and lower APase in low P soils than the non-mycorrhizal plant B we could not reach any conclusions due to lack of non-mycorrhizal control treatments and enough non-mycorrhizal species It would be necessary to carry out new experiments that include proper non-mycorrhizal control to reveal the mechanisms involved We found significant correlations of pH with WSP and APase within or across species in low P soils Moreover, it has been reported that phosphorus adsorption by soil was enhanced with an increase in clay content in suspension (Syers et al., 1973; Ullah et al., 1983) since with increasing clay content there is often increased content of Fe- and Al-(hydro)oxides which are important constituents for P sorption A lower content of clay in soil B suggests that this soil might adsorb less P (lower P sorption capacity) and organic anions than soil ALP and that plants may therefore grow better in soil B than in soil ALP Another explanation is that more sand in soil B makes it more porous which probably gives better aeration of the soil and thereby more oxygen to the roots Further studies in long-term field experiments under varied agricultural soil textures might help to clarify this Through this experiment using four common crops and three agricultural soils we found that plant P uptake may be linked to rhizosphere WSP Rhizosphere organic anions appear to play a minor role in improving P uptake We conclude that our hypothesis (1) that low P availability soils will stimulate plant roots to release more organic anions and phosphatase enzymes to rhizosphere soil was not supported Our hypothesis (2) that the amounts of rhizosphere APase will have positive correlations with rhizosphere plant-available P fractions and P uptake by plants in low P soils was supported but this was not the case for rhizosphere organic anions WSP can be used to study P mobilization and assess soil P availability Hypothesis (3) that different crops will show differing root released organic anions and APase in terms of using residual P from agricultural soils was supported sativa are good candidates to study P utilization The results and information generated in this study are valuable for understanding P mobilization and P uptake in low P agricultural soils and for future effective utilization of P and improving the productivity of the studied crops and YW made contributions to the design of the study MH and EK made a contribution to analysis of soil enzyme activities and Pmic All authors participated in preparing the manuscript This study was supported by the strategic institute program on “Opportunities for sustainable use of phosphorus in food production” at the Norwegian Institute of Bioeconomy Research The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest The reviewer IJ and handling Editor declared their shared affiliation and the handling Editor states that the process nevertheless met the standards of a fair and objective review Many thanks to Hans Lambers (University of Western Australia) for valuable suggestions concerning the study and Torfinn Torp (NIBIO) for their valuable help with seed collection Thanks also should go to Paula Gruner and Pascal Nassal (University of Hohenheim) for help with Pmic/MUF analysis The Supplementary Material for this article can be found online at: http://journal.frontiersin.org/article/10.3389/fpls.2016.01664/full#supplementary-material (A) Shoot P content and (B) root P content of Brassica napus Avena sativa and Solanum tuberosum grown in soils AHP (stippled bars) and (C) rhizosphere total organic anion concentrations for Brassica napus and B (gray bars) based on rhizosphere soil dry weight and (C) N-acetyl-β-glucosaminidase activity of bulk soil for Brassica napus (A) Ratio of acid phosphatase activity to Pmic (B) ratio of acid phosphatase activity to β-glucosidase activity (C) ratio of acid phosphatase activity to N-acetyl-β-glucosaminidase activity and (D) ratio of acid phosphatase activity to β-xylosidase activity of bulk soil Interactions between light intensity and phosphorus nutrition affect the phosphate-mining capacity of white lupin (Lupinus albus L.) Untersuchungen über die chemische Bodenanalyse als Grundlage für die Beurteilung des Nährstoffzustandes der Böden Chemische Extraktionsmethoden zur Phosphor-und Kaliumbestimmung Kungliga Lantbrukshögskolans Annaler 26 “Advances and perspectives to improve the phosphorus availability in cropping systems for agroecological phosphorus management.” Adv Root-released organic acids and phosphorus uptake of two barley cultivars in laboratory and field experiments Characterization of transgenic Trifolium subterraneum L which expresses phyA and releases extracellular phytase: growth and P nutrition in laboratory media and soil Optimization of hydrolytic and oxidative enzyme methods for ecosystem studies Microplate-scale fluorometric soil enzyme assays as tools to assess soil quality in a long-term agricultural field experiment An evaluation of techniques for measuring vesicular arbuscular mycorrhizal infection in roots CrossRef Full Text | Google Scholar Plant-induced changes in the rhizosphere of rape (Brassica napus var pH change and the increase in P concentration in the soil solution Google Scholar soil phosphate fractions and phosphatase activity Google Scholar The effect of rhizosphere phosphorus status on the pH phosphatase activity and depletion of soil phosphorus fractions in the rhizosphere and on the cation-anion balance in the plants Bioavailability of soil inorganic P in the rhizosphere as affected by root-induced chemical changes: a review CrossRef Full Text | Google Scholar CrossRef Full Text | Google Scholar A root observation and sampling chamber for pot studies CrossRef Full Text | Google Scholar Organic acids in the rhizosphere–a critical review CrossRef Full Text | Google Scholar Structural and functional diversity of soil microbial community in the rhizosphere of maize CrossRef Full Text Plant assemblage composition and soil P concentration differentially affect communities of AM and total fungi in a semi-arid grassland Measurement of soil microbial biomass phosphorus by an anion exchange membrane method CrossRef Full Text | Google Scholar Effects of soil compaction and moisture regime on the root and shoot growth and phosphorus uptake of barley plants growing on soils with varying phosphorus status New PRecommendations for Grass and Cereals in Norwegian Agriculture Leaf manganese accumulation and phosphorus-acquisition efficiency Plant nutrient-acquisition strategies change with soil age Root structure and functioning for efficient acquisition of phosphorus: matching morphological and physiological traits Shift from complementarity to facilitation on P uptake by intercropped wheat neighboring with faba bean when available soil P is depleted Improved phosphorus acquisition by tobacco through transgenic expression of mitochondrial malate dehydrogenase from Penicillium oxalicum PubMed Abstract | CrossRef Full Text Root phenes for enhanced soil exploration and phosphorus acquisition: tools for future crops Estimation of the phosphorus sorption capacity of acidic soils in Ireland CrossRef Full Text | Google Scholar temperature and media on acid and alkaline phosphatase activity in “clinical” and “nonclinical” isolates of Bordetella bronchiseptica A modified single solution method for the determination of phosphate in natural waters CrossRef Full Text | Google Scholar Moderating mycorrhizas: arbuscular mycorrhizas modify rhizosphere chemistry and maintain plant phosphorus status within narrow boundaries The Chemical Analysis Program of the Norwegian Forest Research Institute 2000 Ås: Norwegian Forest Research Institute Google Scholar Root carboxylate exudation capacity under phosphorus stress does not improve grain yield in green gram Variation in morphological and physiological parameters in herbaceous perennial legumes in response to phosphorus supply Physiological and morphological adaptations of herbaceous perennial legumes allow differential access to sources of varyingly soluble phosphate Carboxylate composition of root exudates does not relate consistently to a crop species' ability to use phosphorus from aluminium Elevated phosphorus impedes manganese acquisition by barley plants Plant mechanisms to optimise access to soil phosphorus CrossRef Full Text | Google Scholar Exudation of carboxylates in Australian Proteaceae: chemical composition Carbon trading for phosphorus gain: the balance between rhizosphere carboxylates and arbuscular mycorrhizal symbiosis in plant phosphorus acquisition Function and mechanism of organic anion exudation from plant roots Can citrate efflux from roots improve phosphorus uptake by plants Testing the hypothesis with near-isogenic lines of wheat Šarapatka Effect of pH and phosphate supply on acid phosphatase activity in cereal roots Google Scholar Phosphorus supplying capacity of heavily fertilized soils II Dry matter yield of successive crops and phosphorus uptake at different temperatures CrossRef Full Text | Google Scholar Phosphorus fractionation and sorption in P-enriched soils of Norway Roles of arbuscular mycorrhizas in plant phosphorus nutrition: interactions between pathways of phosphorus uptake in arbuscular mycorrhizal roots have important implications for understanding and manipulating plant phosphorus acquisition Google Scholar Mycorrhizal fungi can dominate phosphate supply to plants irrespective of growth responses Phosphate sorption by soils evaluated by the Langmuir adsorption equation doi: 10.2136/sssaj1973.03615995003700030015x CrossRef Full Text | Google Scholar Comparative efficiency of acid phosphatase originated from plant and fungal sources doi: 10.1002/1522-2624(200106)164:3<279::AID-JPLN279>3.0.CO;2-L The influence of soil pH and texture on the adsorption of phosphorus by soils Google Scholar Influence of different trap solutions on the determination of root exudates in Lupinus albus L CrossRef Full Text | Google Scholar Phosphorus acquisition and use: critical adaptations by plants for securing a nonrenewable resource CrossRef Full Text | Google Scholar van Duivenbooden phosphorus and potassium relations in five major cereals reviewed in respect to fertilizer recommendations using simulation modelling Chickpea and white lupin rhizosphere carboxylates vary with soil properties and enhance phosphorus uptake a simple staining technique for arbuscular-mycorrhizal fungi Contrasting responses of root morphology and root-exuded organic acids to low phosphorus availability in three important food crops with divergent root traits Dual effects of transgenic Brassica napus overexpressing CS gene on tolerances to aluminum toxicity and phosphorus deficiency Localization of acid phosphatase activities in the roots of white lupin plants grown under phosphorus-deficient conditions Watts-Williams How important is the mycorrhizal pathway for plant Zn uptake CrossRef Full Text | Google Scholar Carbon and phosphorus exchange may enable cooperation between an arbuscular mycorrhizal fungus and a phosphate-solubilizing bacterium Kandeler E and Clarke N (2016) Rhizosphere Organic Anions Play a Minor Role in Improving Crop Species' Ability to Take Up Residual Phosphorus (P) in Agricultural Soils Low in P Availability Received: 29 May 2016; Accepted: 21 October 2016; Published: 07 November 2016 Copyright © 2016 Wang, Krogstad, Clarke, Hallama, Øgaard, Eich-Greatorex, Kandeler and Clarke. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY) distribution or reproduction in other forums is permitted provided the original author(s) or licensor are credited and that the original publication in this journal is cited in accordance with accepted academic practice distribution or reproduction is permitted which does not comply with these terms *Correspondence: Tore Krogstad, dG9yZS5rcm9nc3RhZEBubWJ1Lm5v Nicholas Clarke, bmljaG9sYXMuY2xhcmtlQG5pYmlvLm5v Disclaimer: All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher 94% of researchers rate our articles as excellent or goodLearn more about the work of our research integrity team to safeguard the quality of each article we publish Metrics details Phosphorus (P) accumulators used for phytoremediation vary in their potential to acquire P from different high P regimes Growth and P accumulation in Polygonum hydropiper were both dependent on an increasing level of IHP (1–8 mM P) and on a prolonged growth period (3-9 weeks) and those of the mining ecotype (ME) were higher than the non-mining ecotype (NME) stem and leaf of both ecotypes were significantly greater in IHP relative to other organic P (Po) sources (G1P but lower than those in inorganic P (Pi) treatment (KH2PO4) P accumulation in the ME exceeded the NME from different P regimes The ME demonstrated higher root activity compared to the NME grown in various P sources Acid phosphatase (Apase) and phytase activities in root extracts of both ecotypes grown in IHP were comparable to that in Pi Higher secreted Apase and phytase activities were detected in the ME treated with different P sources relative to the NME the ME demonstrates higher P-uptake efficiency and it is a potential material for phytoextraction from P contaminated areas Repeated and substantial application of animal manure to farmland will increase risks of accumulation of Po in soils and P-pollution due to P runoff and leaching more attention should be paid to Po in potential environmental P-pollution issues these P accumulators have defects like; relatively low DW yield Polygonum hydropiper represents a worthy candidate to remediate excess P because of its great attributes of being able to grow in both terrestrial and aquatic areas and high potentials of P uptake and P removal The previous studies provide a sound theoretical basis for evolving a P-phytoextraction strategy in the ME and NME grown in sole Po conditions it is necessary to achieve a thorough understanding of the pattern of P nutrition in the two ecotypes using Po sources we supposed that the ME and the NME may differ in P uptake from Po and their physiological responses to Po supply remained different three different experiments were performed to compare the differences between the ME and the NME in: 1) tolerance by determining biomass under high levels of P or different growth periods; 2) P uptake ability by analyzing tissue P accumulation; 3) physiological responses by determining root activity extracted Apase and phytase activities and secreted Apase and phytase activities from the roots to assess the utilization of Po when the two ecotypes of P hydropiper were grown in a range of media supplied with different P sources Biomass (a) and P accumulation (b) in the whole plant of P hydropiper grown under hydroponic media containing 1–8 mM P supplied as IHP for 5 weeks Values represent mean ± standard error of four replicates The histograms with different small letters are statistically different (p < 0.05) among the various Po concentrations and * represents significant difference (p < 0.05) between the two ecotypes P. hydropiper accumulated different P amounts in the whole plant when grown in different levels of Po added as IHP (Fig. 1b) Whole plant P accumulation of the ME significantly increased at 4 mM a continued slowdown of whole plant P accumulation was observed in the NME with the increasing Po concentrations Whole plant P accumulation in the ME was significantly greater relative to that of the NME The ME demonstrated whole plant P accumulation in a relatively narrow range of 9.39–12.84 mg plant−1 while the NME’s whole plant P accumulation ranged from 3.16–9.27 mg plant−1 As shown in Table 1 biomass of both ecotypes significantly increased with the increasing growth periods The greatest increment of biomass was observed from 5 to 7 weeks in both ecotypes The ME showed significantly greater biomass in root stem and leaf relative to the NME at 7 weeks and there were no obvious differences in the biomass between the two ecotypes in the other growth periods This suggests that both ecotypes are able to obtain P from high concentrations of IHP P accumulations in the two ecotypes differed greatly among the different growth periods (Table 2) P accumulations of both ecotypes were in the order of stem>leaf>root Stem and leaf P accumulations of both ecotypes significantly increased with prolonged growth periods Stem P accumulation in the ME seedlings increased in response to increasing IHP and it reached 31.07 and 40.94 mg plant−1 at 7 and 9 weeks The same pattern was noticed in leaf P accumulation the ME demonstrated significantly higher P accumulations in the stem and leaf compared to the NME The growth of the mining ecotype (left) and the non-mining ecotype (right) of P hydropiper grown under perlite media containing 3 mM P supplied either as G1P (a) In both ecotypes, P accumulations were dependent on both P sources and ecotypes (Table 4) The ME accumulated P in the roots from various P sources ranging from 0.50–2.47 mg plant−1 which was significantly greater than root P accumulation of the NME in G1P and AMP The seedlings exhibited a similar pattern with a highest stem P accumulation of 29.76 mg plant−1 for the ME and 22.82 mg plant−1 for the NME from Pi media the ME showed a significantly higher stem P accumulation of 1.30–1.93 times compared to the NME Leaf P accumulation from IHP source was comparable to P amount accumulated from Pi source The accumulations from IHP and Pi media were significantly greater than from the other Po sources in both ecotypes No significant difference was noticed in accumulations of leaves in the ME and NME even in Pi media hydropiper grown under perlite media containing 3 mM P supplied either as G1P Values represent mean ± standard error of six replicates The histograms with different small letters are significantly different (p < 0.05) among P sources * indicates significantly different (p < 0.05) between ecotypes Activities of APase (a) and phytase (b) in root extracts of P Secreted APase (a) and phytase (b) activities of P Whole plant P accumulation in the ME did not decrease with increasing Po levels hydropiper might be effectively used for IHP removal from eutrophic water with different degrees of pollution an interesting result observed in the two ecotypes was that higher biomass and P content was accumulated in the ME seedlings supplied with any P level or P source greater shoot P accumulation in the ME was observed in high P media indicating the ME was more efficient relative to the NME and other P accumulators to uptake and remove P and it is a promising species for phytoremediation of P polluted areas the harvest time also affected the capacity of P removal by P accumulators from real eutrophic water or high P soils It will significantly improve phytoremediation efficiency by harvesting seedlings at a growth period with the maximum ability for P uptake P accumulation was dependent on the growth period and reached a maximal value in 9 weeks seedlings was just in flowering stage of 9-week growth and an advisable measure involving harvesting of P hydropiper in flowering might be effective in decreasing excess P levels integrated attributes are the key decisive factors to heighten P assimilation in a plant it was also observed that extracted and secreted Apase activity of the ME was significantly higher than that of the NME as a result of more appreciable tissue biomass and P uptake high extracellular activities of Apase and phytase from roots not just high levels of intracellular Apase and phytase activities are key stimulators to P utilization and uptake of P Modified Hoagland’s salts mixture without monopotassium phosphate (KH2PO4) as described by Ye et al.8 was used as basal nutrient medium There were three pot experiments as follows: the first hydroponic experiment was carried out to determine the effect of increasing levels of Po (1 6 and 8 mM) added as IHP [Sigma] on the growth and P uptake of P Four replicates were performed for each treatment the pots were painted outside with a black varnish Two healthy and uniform plants were transferred in each pot with 5 L of modified Hoagland’s solution The seedlings were fixed by the sponge and hard cystosepiment to keep the shoots above cystosepiment The cystosepiment has two apertures of 2 cm and its thick is 2 cm The nutrient media (pH 5.8) were replaced every 5 days Plants were harvested after 5 weeks of growth with sunlight Plants were gently removed from the cystosepiments samples were washed with tap water and distilled water respectively and blotted with absorbing paper the second hydroponic experiment was performed in a greenhouse using barrels of 3.5 L filled with basal media with 3 mM P supplied as IHP The pretreatment of the corresponding items was the same as the above mentioned Four healthy seedlings of similar size were transferred to 1:2 Hoagland’s solution Seedlings of 10 d old were then transplanted into nutrient solution with 3 mM P added as IHP Experimental treatment was repeated using three replicates for each harvest Barrels were randomized by a complete block design This experiment was performed with sunlight The harvested plants were treated as above and divided into root the third experiment was conducted to investigate the effect of various Po sources on P accumulation and physiological characteristics of P Perlite was selected as the immobilizing matrix Basal nutrient solution medium (pH 5.8) was used by adding 3 mM P supplemented either as α-D-glucose 1-phosphate disodium salt (G1P) adenosine 3′:5′ cyclic monophosphate sodium salt (AMP) adenosine-5′-triphosphate disodium salt (ATP) The control was 3 mM P added as KH2PO4 (Pi) hydropiper were first transferred to 1:2 Hoagland’s solution for preculture of 10 days and then transplanted respectively in each barrel (3.5 L) containing 0.3 kg perlite and 2 L of the medium Further addition of the medium was 300 mL every 3 or 4 days for each pot water management was performed by the weight method This greenhouse experimental design was a completely randomized design and each treatment was replicated six times They were treated as above and divided into root The reaction was maintained at 37 °C and incubated for 2 h 2 mL of 1 M sulphuric acid solution was added to stop reaction A control was determined by adding 2 mL of 1 M sulphuric acid solution first The other operating steps in the control test were done according to the above procedures all roots were taken out from the triangular flask and dried using absorbent paper The dried roots were homogenized with a mortar and pestle in 2 mL acetic ether and the supernatants were transferred into a 10 mL volumetric flask The extraction process was repeated 2–3 times until the supernatant was colourless The absorbance of the colored solution was determined spectrophotometrically at 485 nm when TTC contacts with live cells of roots it will be reduced by dehydrogenase enzymes into triphenyltetrazolium formazan (TTF) the colorless root will turn to red root and shades of red in the roots are positively correlated with root activity Root activity was measured from the release of TTF and defined as TTF μg g−1 FW h−1 Roots from different P regimes were separated after being washed thoroughly then frozen in liquid nitrogen and stored at −80 °C Fresh tissues of 0.3 g were chilled on ice and homogenized with a mortar and pestle in 5 mL of 15 mM 2-morpholinoethanesulfonic acid monohydrate (MES) buffer (0.5 mM CaCl2·H2O The extracts were centrifuged at 4 °C (10,000 rpm 20 min) and the supernatants were used to analyze activities of Apase and phytase The assay for APase activity was performed in 3 mL liquid containing 2 mL of 10 mM pNPP and 1 mL enzyme extract The components were mixed and Apase activity was determined after 30 min incubation at 37 °C followed by the addition of 2 mL 0.25 M NaOH The reaction for analysis of phytase activity was initiated by the addition of 2 mL of 15 mM MES buffer (pH 5.5) to an assay mixture containing 1 mL enzyme extract and 1 mL of 2 mM IHP 2 mL of ice-cold 20% (w/v) trichloroacetic acid was added to terminate the reaction after incubation at 37 °C for 60 min APase and phytase activities were determined from the release of p-nitrophenol (pNP) and soluble Pi spectrophotometrically using a UV-VIS spectrophotometer at 412 nm and 882 nm APase activity was defined as pNP μg g−1 FW min−1 Phytase activity was expressed as mU g−1 root FW where 1 U releases 1 μmol of soluble Pi min−1 Five week-old seedlings grown in the various P media were harvested and the roots were washed with sterile deionized water and wiped The roots were incubated for 2 h in 30 mL of 15 mM MES buffer (pH 5.5) containing 10 mM pNPP for APase or 2 mM IHP for phytase roots were also incubated in buffer in the absence of IHP as a control to account for the possible P efflux from roots The roots were washed with distilled water after the incubation and the FW was recorded The secreted APase and phytase activities were analyzed as described above Statistical analyses were conducted by the DPS 11.0 software package using variance analysis Differences at significant level of p < 0.05 were estimated using LSD Graphical work was accomplished by Origin 8.0 Different plant ecotypes responded differently to P media The ME was more tolerant to high levels of Po (IHP) and therefore demonstrated higher biomass and P accumulation Superior capability in growth and P uptake was also observed in the ME from various Po sources ATP and AMP reduced the biomass and P accumulation in both ecotypes grown in perlite media relative to IHP and Pi APase and phytase activities in root extracts and secretions were observed in the ME compared with the NME enhanced root productions and secretions of Apase and phytase may be responsible for increased mineralization of various Po sources to release available P for ME seedlings growth and P uptake It thus reveals that the ME has an efficient P uptake mechanism in response to Po sources and has attractive potential of extracting P from P-polluted area P accumulation and physiological responses to different high P regimes in Polygonum hydropiper for understanding a P-phytoremediation strategy Chemistry and dynamics of soil organic phosphorus in Phosphorus: Agriculture and the Environment (Eds Acid phosphomonoesterase and phytase activities of wheat (Triticum aestivum L.) roots and utilization of organic phosphorus substrates by seedlings grown in sterile culture Dynamics and availability of phosphorus in the rhizosphere of a temperate silvopastoral system phosphatase activity and P-nutrition of soybean as influenced by inoculation of Bacillus P accumulation potential of Polygonum hydropiper grown in high P media Response of root and sediment phosphatase activity to increased nutrients and salinity P accumulation and activities of phytase and acid phosphatases in two cultivars of annual ryegrass (Lolium multiflorum L.) Phosphorus fractions in poultry litter characterized by sequential fractionation coupled with phosphatase hydrolysis Investigation of the inorganic and organic phosphorus forms in animal manure Influence of phosphorus nutrition on growth and metabolism of Duo grass (Duo festulolium) Enhanced accumulation of phosphate by Lolium multiflorum cultivars grown in phosphate-enriched medium Characterization of phosphate accumulation in Lolium multiflorum for remediation of phosphorus-enriched soils Enhanced organic phosphorus assimilation promoting biomass and shoot P hyperaccumulations in Lolium multiflorum grown under sterile conditions Uptake and accumulation of phosphorus by dominant plant species growing in a phosphorus mining area P accumulation and physiological characteristics of two ecotypes of Polygonum hydropiper as affected by excess P supply P uptake and activities of acid phosphatase and phytase of Polygonum hydropiper Accumulation characteristics of and removal of nitrogen and phosphorus from livestock wastewater by Polygonum hydropiper soil P fractions and phosphatase activity as affected by swine manure Extracellular secretion of Aspergillus phytase from Arabidopsis roots enables plants to obtain phosphorus from phytate Engineering the root-soil interface via targeted expression of a synthetic phytase gene in trichoblasts Mechanisms for tolerance of very high tissue phosphorus concentrations in Ptilotus polystachyus Effects of phosphorus and potassium on forage nutritive value and quantity Growth characteristics and nutrient removal capability of plants in subsurface vertical flow constructed wetlands Chickpea facilitates phosphorus uptake by intercropped wheat from an organic phosphorus source truncatula phytase gene results in improved acquisition of organic phosphorus by Arabidopsis Transgenic expression of phytase and acid phosphatase genes in alfalfa (Medicago sativa) leads to improved phosphate uptake in natural soils Plant assimilation of phosphorus from an insoluble organic form is improved by addition of an organic anion producing Pseudomonas sp Improved phosphorus acquisition and biomass production in Arabidopsis by transgenic expression of a purple acid phosphatase gene from M Improving phosphorus acquisition of white clover Trifolium repens L by transgenic expression of plant-derived phytase and acid phosphatase genes Phosphorus acquisition from phytate depends on efficient bacterial grazing irrespective of the mycorrhizal status of Pinus pinaster Localization of the Bacillus subtilis beta-propeller phytase transcripts in nodulated roots of Phaseolus vulgaris supplied with phytate Effects of poultry manure amendment on phosphorus uptake by ryegrass soil phosphorus fractions and phosphatase activity Enhancement of nitrogen and phosphorus removal from eutrophic water by economic plant annual ryegrass (Lolium multiflorum) with ion implantation The growth and phosphorus utilisation of plants in sterile media when supplied with inositol hexaphosphate glucose 1-phosphate or inorganic phosphate Acquisition of phosphorus and nitrogen in the rhizosphere and plant growth promotion by microorganisms Limitations to the potential of transgenic Trifolium subterraneum L plants that exude phytase when grown in soils with a range of organic P content in Experiment Course of Plant Physiology (eds 30–31 (Sichuan Science and Technology Press 312–314 (Chinese Agricultural Science and Technology Press Download references This study was supported by the Sichuan Science and Technology Support Program (2013NZ0044) (2013NZ0029); the National Natural Science Foundation (31401377) (41271307) and Sichuan Provincial Education Department Key Project Program (14ZA0002) The authors also acknowledge the contribution of Craig Stapleton Guangdeng Chen and Barbara Lynn Hallanger in improving the construction and language of our manuscript carried out the majority of this research work (experiment design statistical analysis and writing this paper) experiment management and reviewed this manuscript The authors declare no competing financial interests Download citation Anyone you share the following link with will be able to read this content: a shareable link is not currently available for this article Sign up for the Nature Briefing newsletter — what matters in science The dates displayed for an article provide information on when various publication milestones were reached at the journal that has published the article activities on preceding journals at which the article was previously under consideration are not shown (for instance submission Ecotoxicology and Environmental SafetyCitation Excerpt :Previous studies indicated that long-term excessive application of pesticides will produce phytotoxicity once it exceeds the limits of non-target plants which will affect plants growth by decreasing biomass and chlorophyll content and so on (Kaya and Yigit The roots system could absorb and transport water and nutrients to ensure the growth of the plants (Lebrun et al. When the plant roots are exposed to excessive pesticides the physiological structure of the roots can be damaged All content on this site: Copyright © 2025 Elsevier B.V.