Stimulation of root acid phosphatase by phosphorus deficiency is regulated by ethylene in Medicago falcata

Plants have developed numerous strategies to cope with phosphorus (P) deficiency resulting from low availability in soils. Evolution of ethylene and up-regulation of root secreted acid phosphatase activity are common for plants in response to P deficiency. To determine the role of ethylene in response of plants to P deficiency, we investigated the effects of ethylene precursor (1-amino cyclopropane-1-carboxylic acid, ACC) and ethylene synthesis antagonists (aminoethoxyvinylglycine AVG, cobalt, Co²⁺) on P concentrations in roots and shoots of Medicago falcata seedlings grown in P-sufficient (500 μM H₂PO₄⁻) and P-deficient (5 μM H₂PO₄⁻) solution. After transferring M. falcata seedlings from P-sufficient to P-deficient solution for 2 days, root P concentration was significantly reduced. The reduction in root P concentration was reversed by AVG and Co²⁺, and a similar reduction in root P concentration of seedlings exposed to P-sufficient solution was observed by ACC. Expression of high-affinity phosphate transporters (MfPT1, MfPT5) was enhanced by P-deficiency and this process was reversed by AVG and Co²⁺. There was a marked increase in activity of root acid phosphatase (APase) and expression of gene encoding APase (MfPAP1) under P-deficient conditions, and the increase in APAse activity and expression of MfPAP1 was inhibited by AVG and Co²⁺. APase activity and expression of MfPAP1 expression in seedlings grown in P-sufficient solution were enhanced by ACC. Root and shoot P concentrations were increased when organic phosphorus was added to the P-deficient solution, and the increase in P concentration was significantly inhibited by AVG and Co²⁺. These results indicate that ethylene plays an important role in modulation of P acquisition by possibly mobilizing organic P via up-regulating root APase activity and high-affinity phosphate transporters.     

Soil salinity and drought alter wood density and vulnerability to xylem cavitation of baldcypress (Taxodium distichum (L.) Rich.) seedlings

We investigated the role of hydraulic conductivity, wood density, and xylem cavitation in the response of baldcypress (Taxodium distichum) seedlings to increased soil salinity and drought. One-year-old, greenhouse-grown seedlings were irrigated daily with a 100 mM (≈6‰) salt solution or once per week with fresh water (drought). Controls were irrigated daily with fresh water. Gas exchange rates of stressed plants were reduced by approximately 50% (salt) and 70% (drought), resulting in a 50–60% reduction in diameter growth for both treatments. Stem-specific hydraulic conductivity (K_{S native}) of stressed plants was 33% (salt) and 66% (drought) lower than controls and we observed a strong positive correlation between K_{S native} and gas exchange. In addition, we found a strong relationship between CO₂ assimilation rate (A) and the soil-to-leaf hydraulic conductance (k_L). The relationship was identical for all treatments, suggesting that our moderate salt stress (as well as drought) did not affect the photosynthetic biochemistry of leaves, but rather reduced A via stomatal closure. Lower K_{S native} of stressed plants was associated with increased wood density and greater resistance to xylem cavitation. Xylem pressures causing 50% loss of hydraulic conductivity (P₅₀) were ?2.88 ± 0.07 MPa (drought), ?2.50 ± 0.08 MPa (salt) and ?2.01 ± 0.04 MPa (controls). P₅₀s were strongly correlated with wood density (r = ?0.71, P < 0.01) and K_{S native} (r = 0.74, P < 0.01). These findings support the hypothesis that there is a significant trade-off between a plant's cavitation resistance and its hydraulic efficiency. The results of the present study indicate that stressed plants partitioned their biomass in a way that strengthened their xylem and reduced vulnerability to xylem cavitation. Hence, these seedlings could be better suited to be planted in environments with elevated soil salinity. For most parameters (especially P₅₀), drought had an even more pronounced effect than salinity. This is important as nurseries could produce “stress-acclimated” seedlings simply by reducing irrigation amounts and would not have to contaminate the soils in their nursery beds with salt applications.     

Discrimination against 15N among recombinant inbred lines of Phaseolus vulgaris L. contrasting in phosphorus use efficiency for nitrogen fixation

Although isotopic discrimination processes during nitrogen (N) transformations influence the outcome of ¹⁵N based quantification of N₂ fixation in legumes, little attention has been given to the effects of genotypic variability and environmental constraints such as phosphorus (P) deficiency, on discrimination against ¹⁵N during N₂ fixation. In this study, six Phaseolus vulgaris recombinant inbred lines (RILs), i.e. RILs 115, 104, 34 (P deficiency tolerant) and 147, 83, 70 (P deficiency sensitive), were inoculated with Rhizobium tropici CIAT899, and hydroaeroponically grown with P-sufficient (250 μmol P plant⁻¹ week⁻¹) versus P-deficient (75 μmol P plant⁻¹ week⁻¹) supply. Two harvests were done at 15 (before nodule functioning) and 42 (flowering stage) days after transplanting. Nodulation, plant biomass, P and N contents, and the ratios of ¹⁵N over total N content (¹⁵N/Nt) for shoots, roots and nodules were determined. The results showed lower ¹⁵N/Nt in shoots than in roots, both being much lower than in nodules. P deficiency caused a larger decrease in ¹⁵N/Nt in shoots (−0.18%) than in nodules (−0.11%) for all of the genotypes, and the decrease in shoots was greatest for RILs 34 (−0.33%) and 104 (−0.25%). Nodule ¹⁵N/Nt was significantly related to both the quantity of N₂ fixed (R² = 0.96***) and the P content of nodules (R² = 0.66*). We conclude that the discrimination against ¹⁵N in the legume N₂-fixing symbiosis of common bean with R. tropici CIAT899 is affected by P nutrition and plant genotype, and that the ¹⁵N/Nt in nodules may be used to screen for genotypic variation in P use efficiency for N₂ fixation.     

The physiological mechanism of enhanced oxidizing capacity of rice (Oryza sativa L.) roots induced by phosphorus deficiency

Plants show various responses to phosphorus (P) deficiency. Root oxidizing capacity enhancement is one of adaptive mechanisms for rice (Oryza sativa L.) to P deficiency. However, it remains unclear how P deficiency enhances the root oxidizing capacity. In this study, rice seedlings were treated in P-deficient nutrient solution for different periods. Variations of reactive oxygen species (ROS), antioxidant enzyme activity, root lignin content, root porosity, root oxygen release, total oxidative substances and root structural changes in rice roots in response to P-sufficient and P-deficient treatments were investigated. Results indicated that P deficiency induced the production of H₂O₂ and O ₂ ^·? in roots significantly, which reached their maximum after 1- to 2-day P-deficient treatment. Interestingly, the endogenous total oxidative substances kept stable in rice roots. P deficiency increased the activities of peroxidase and superoxide dismutase by 89.5 and 51.8 % after 4-day P-deficient treatment, respectively. Moreover, one-day P deficiency elevated lignin accumulation. Root porosity of rice seedling under 2-day P-deficient treatment was 19.8 % higher than that under P-sufficient treatment. P deficiency also enhanced the release of both O₂ and total oxidative substances after 1- to 4-day P deficiency. In addition, results from electronic microscopy indicated that the thickness of root cell wall tended to increase after 2-day P-deficient treatment. Taken together, our results suggested that P-deficiency-induced enhancement of root oxidizing capacity in rice roots was probably associated with ROS production, antioxidant enzyme activity increment in root tissues, and the release of O₂ and oxidative substances from root inside to rhizosphere.     

De novo arginine biosynthesis in leaves of phosphorus-deficient citrus and poncirus species

Young, fully expanded leaves from 7-month-old P-deficient citrus rootstock seedlings had levels of nonprotein arginine that were 10- to 50-fold greater than those from P-sufficient control plants. Arginine content of the protein fraction increased 2- to 4-fold in P-deficient leaves. Total arginine content, which averaged 72 ± 6 micromoles per gram dry weight of P-sufficient leaf tissue (mean ± se, n = the four rootstocks) was 207, 308, 241, and 178 micromoles in P-deficient leaves from Citrus limon cv rough lemon, Poncirus trifoliata × C. sinensis cv Carrizo citrange and cv Troyer citrange, and P. trifoliata cv Australian trifoliate orange, respectively. For each rootstock, the accumulation of arginine paralleled an increase in the activity of the pathway for the de novo biosynthesis of arginine. The ratio of the nanomoles NaH¹⁴CO₃ incorporated into the combined pool of arginine plus urea per gram fresh weight intact leaf tissue during a 3-hour labeling period for P-deficient to P-sufficient plants was 91:34, 49:11, 35:11, and 52:41, respectively. When P-deficient plants were supplied with P, incorporation of NaH¹⁴CO₃ into arginine plus urea was reduced to the level observed for the P-sufficient control plants of the same age and arginine ceased to accumulate. Arginase and arginine decarboxylase activity were either unaffected or slightly increased during phosphorus deficiency. Taken together, these results provide strong evidence that arginine accumulation during phosphorus deficiency is due to increased activity of the de novo arginine biosynthetic pathway.     

Effects of controlled irrigation on water potential,nitrogen uptake and biomass production in Dalbergia sissoo seedlings

Biomass production, pattern of nodulation, nutrient uptake, net photosynthetic rate (P_n), leaf temperature (T_leaf), leaf nitrate reductase (NR) activity and free proline of Dalbergia sissoo seedlings planted in containers with 120 kg soil were studied under different water stress levels to assess the productive potential of the species in dry areas. Seedlings were irrigated at 20 mm (W₁), 14 mm (W₂), 10 mm (W₃), 8 mm (W₄) throughout the experimental period to maintain the respective treatment up to the lowest soil water content of 7.43%, 5.64%, 4.30% and 3.23%, respectively. There was a treatment (W₅) in which seedling were irrigated once to −0.03 MPa and left without re-irrigation. Decreased irrigation level resulted in lowering of leaf water potential (LWP), net photosynthetic rate (P_n), total number of root nodules and nodule dry mass and nitrogen uptake in the seedling. P_n, leaf nitrate reductase (NR) activity and seedling biomass were highest in W₁ indicating a positive relations of NR activity with CO₂ assimilation and biomass production. The decrease in P_n, leaf NR activity and LWP was sharp at W₃ onwards. Monthly changes in the values of P_n, T_leaf and NR activity indicate environmental effect on these physiological variables. Proline was detected only in the seedlings of W₃, W₄ and W₅ treatments after February and was highest in the seedlings of W₅ treatment. The study suggests that severe water deficit adversely affect physiological and biochemical processes that resulted in reduced growth, nutrient uptake and biomass productivity in D. sissoo seedlings. Re-irrigation above W₃ level is recommended for this species.     

Immobilization of glycolate oxidase from Medicago falcata on magnetic nanoparticles for application in biosynthesis of glyoxylic acid

Glycolate oxidase was isolated from Medicago falcata Linn. after a screening from 13 kinds of C₃ plant leaves, with higher specific activity than the enzyme from spinach. The M. falcata glycolate oxidase (MFGO) was partially purified and then immobilized onto hydrothermally synthesized magnetic nanoparticles via physical adsorption. The magnetic nanoparticles were characterized with scanning electron microscope (SEM), transmission electron microscopy (TEM) and Fourier transform infrared (FT-IR) spectroscopy. The maximum load of MFGO was 56 mg/g support and the activity recovery was 45%. Immobilization of MFGO onto magnetic nanoparticles enhanced the enzyme stability, and the optimum temperature was significantly increased from 15 °C to 30 °C. The immobilized biocatalyst was successfully used in a batch reactor for repeated oxidization of glycolic acid to synthesize glyoxylic acid, retaining ca. 70% of its initial activity after 4 cycles of reaction at 30 °C for nearly 70 h, and its half-life was calculated to be 117 h.     

Effects of shade treatments on the photosynthetic capacity,chlorophyll fluorescence,and chlorophyll content of Tetrastigma hemsleyanum Diels et Gilg

Tetrastigma hemsleyanum Diels et Gilg was grown under full sunlight and moderate and high levels of shade for one month to evaluate its photosynthetic and chlorophyll fluorescence response to different light conditions. The results showed that T. hemsleyanum attained greatest leaf size and P_n when cultivated with 67% shade. Leaves of seedlings grown with 90% shade were the smallest. Leaf color of plants grown under full sunlight and 50% shade was yellowish-green. The P_n value increased rapidly as PPFD increased to 200 μmol m^?2 s^?1 and then increased slowly to a maximum, followed by a slow decrease as PPFD was increased to 1000 μmol m^?2 s^?1. P_n was highest for the 67% shade treatment and the LSP for this shade treatment was 600 μmol m^?2 s^?1. Full sunlight and 50% shade treatments resulted in significant reduction of ETR and qP and increased NPQ. Chl a, Chl b and total chlorophyll content increased and Chl a/b values decreased with increased shading. Results showed that light intensity greater than that of 50% shade depressed photosynthetic activity and T. hemsleyanum growth. Irradiance less than that of 75% shade limited carbon assimilation and led to decreased plant growth. Approximately 67% shade is suggested to be the optimum light irradiance condition for T. hemsleyanum cultivation.     

Phosphorus removal from agricultural runoff by constructed wetland

A free-water surface wetland covering an area of 2800 m² was operated from March 2002 to June 2004 for agricultural runoff treatment in the Dianchi Valley in China. In the wetland were grown Zizania Caduciflora Turez Hand-mazt and Phragmites australis (Cav.) Trin.ex Steud. The instantaneous inflow rate was measured and the integrated flux was recorded by an ultrasonic flow instrument all year round. The average inflow rate, hydraulic loading rate (HLR) and hydraulic retention time (HRT) were kept at 242 m³ d^?1, 12.7 cm d^?1 and 2.0 d, respectively. The annual average total phosphorus (TP) in the inflow was 0.87 mg L^?1, and the corresponding removal efficiency was calculated to be 59.0%. Biannual plant uptake and removal by harvesting and seed transport was the main pathway for TP removal, while the influent TP load was 12.9 g m^?2 year^?1. Hydraulic retention time had a significant positive correlation with the removal of P (r² = 0.88). Water temperature, inflow phosphorus load, inflow and hydraulic load rates were positively correlated with the removal of P. Inflow phosphorus concentrations were negatively correlated with the removal of P. It is shown that the free-water surface wetland was an effective and economical system for agricultural runoff treatment in lake regions.     

Role of Ethylene on de Novo Shoot Regeneration from Cotyledonary Explants of Brassica campestris ssp. pekinensis (Lour) Olsson in Vitro

The promotive effect of AgNO₃ and aminoethoxyvinylglycine (AVG) on in vitro shoot regeneration from cotyledons of Brassica campestris ssp. pekinensis in relation to endogenous 1-amino-cyclopropane-1-carboxylic acid (ACC) synthase, ACC, and ethylene production was investigated. AgNO₃ enhanced ACC synthase activity and ACC accumulation, which reached a maximum after 3 to 7 days of culture. ACC accumulation was concomitant with increased emanation of ethylene which peaked after 14 days. In contrast, AVG was inhibitory to endogenous ACC synthase activity and reduced ACC and ethylene production. The promotive effect of AVG on shoot regeneration was reversed by 2-chloroethylphosphonic acid at 50 micromolar or higher concentrations, whereas explants grown on AgNO₃ medium were less affected by 2-chloroethylphosphonic acid. The distinctive effect of AgNO₃ and AVG on endogenous ACC synthase, ACC, and ethylene production and its possible mechanisms are discussed.     

Production of rhoifolin and tiliroside from callus cultures of Chorisia chodatii and Chorisia speciosa

The current work aims to stimulate the production of rhoifolin and tiliroside as two valuable phytochemicals from Chorisia chodatii Hassl. and Chorisia speciosa A. St.-Hil. callus cultures. A comparison between three explants from the in vitro germinated seedlings of both species for callus induction and accumulation of both flavonoids was carried out. Highly efficient calluses were induced from the leaves, stems and roots of C. chodatii seedlings on Gamborg’s B5 (B5) and Murashige and Skoog (MS) media containing 2.0 mg/l β-naphthalene acetic acid (NAA) and 0.5 mg/l 6-benzyladenin (BA) or kinetin (Kn), while those of C. speciosa seedlings efficiently produced calluses on both media supplemented with 0.5 or 1.0 mg/l NAA and 0.5 mg/l BA. Besides, the highest contents of rhoifolin (1.927 mg/g DW) and tiliroside (1.776 mg/g DW) from C. speciosa cultures were obtained from the calluses of seedlings’ roots and stems maintained on B5 medium containing 1.0 mg/l NAA and 0.5 mg/l BA, respectively. On the other hand, the maximum rhoifolin content (0.555 mg/g DW) from C. chodatii cultures was obtained from the calluses of seedlings’ stems grown on B5 medium supplemented with 2.0 mg/l NAA and 0.5 mg/l BA, whereas the highest tiliroside content (0.547 mg/g DW) was provided by the root explants on B5 medium containing 2.0 mg/l NAA and 0.5 mg/l Kn. Both flavonoids were bioaccumulated in greater amounts than the wild and cultivated intact plants, which provides a promising tool for their future commercial production under a controlled environment, independent of climate and soil conditions.     

Growth of the fungus Paecilomyces lilacinus with n-hexadecane in submerged and solid-state cultures and recovery of hydrophobin proteins

The filamentous fungus Paecilomyces lilacinus was grown on n-hexadecane in submerged (SmC) and solid-state (SSC) cultures. The maximum CO₂ production rate in SmC (V_max = 11.7 mg CO₂ L_g⁻¹ day⁻¹) was three times lower than in SSC (V_max = 40.4 mg CO₂ L_g⁻¹ day⁻¹). The P. lilacinus hydrophobin (PLHYD) yield from the SSC was 1.3 mg PLHYD g protein⁻¹, but in SmC, this protein was not detected. The PLHYD showed a critical micelle concentration of 0.45 mg mL⁻¹. In addition, the PLHYD modified the hydrophobicity of Teflon from 130.1 ± 2° to 47 ± 2°, forming porous structures with some filaments <1 μm and globular aggregates <0.25 μm diameter. The interfacial studies of this PLHYD could be the basis for the use of the protein to modify surfaces and to stabilize compounds in emulsions.     

Influence of phosphorus nutrition on growth and metabolism of Duo grass (Duo festulolium)

Use of suitable plants that can extract and concentrate excess P from contaminated soil serves as an attractive method of phytoremediation. Plants vary in their potential to assimilate different organic and inorganic P-substrates. In this study, the response of Duo grass (Duo festulolium) to variable rates of soil-applied potassium dihydrogen phosphate (KH₂PO₄) on biomass yield and P uptake were studied. Duo grown for 5 weeks in soil with 2.5, 5 and 7.5 g KH₂PO₄ kg^?1 soil showed a significantly higher biomass and shoot P content of 8.3, 11.4 and 12.3 g P kg^?1 dry weight respectively compared to plants that received no soil added P. Also, the ability of Duo to metabolize different forms of P-substrates was determined by growing them in sterile Hoagland's agar media with different organic and inorganic P-substrates, viz. KH₂PO₄, glucose-1-phosphate (G1P), inositiol hexaphosphate (IHP), adenosine triphosphate (ATP) and adenosine monophosphate (AMP) for 2 weeks. Plants on agar media with different P-substrates also showed enhanced biomass yield and shoot P relative to no P control and the P uptake was in the order of ATP > KH₂PO₄ > G1P > IHP = AMP > no P control. The activities of both phytase (E.C.3.1.3.26) and acid phosphatases (E.C.3.1.3.2) were higher in all the P received plants than the control. Duo grass is capable of extracting P from the soil and also from the agar media and thus it can serve as possible candidate for phytoextraction of high P-soil.     

Mechanism of photosynthetic response in Microcystis aeruginosa PCC7806 to low inorganic phosphorus

Photosynthetic response of Microcystis aeruginosa PCC7806 to different concentrations of phosphorus supply was studied so as to elucidate if the declining process of Microcystis bloom under freshwater ecosystem is related to soluble reactive phosphorus (SRP) decrease in water volume. Growth rate of M. aeruginosa PCC7806 was significantly reduced under P-deficient conditions, and its photosynthetic activity in terms of rETR_max (maximum electron transport rate) decreased significantly after 48 h growth, while it kept elevating and reached to a relative stable value when supplied with rich phosphorus of 0.6 mg/L. With the increasing actinic irradiance along the rapid light curves of M. aeruginosa PCC7806 cultured under low-phosphorus level, qP (photochemical quenching) and rETR (relative electron transport rate) decreased greatly, and the increase in qN (non-photochemical quenching) and Φ_PS (actual photochemical efficiency of PSII) was obviously inhibited. The affinity of M. aeruginosa PCC7806 to inorganic carbon was reduced evidently in 0.02 mg/L P compared with in 0.6 mg/L P. When P was reduced from 0.6 to 0.02 mg/L, the decreasing rate of rETR_max (77%) was significantly greater than that of photosynthetic carbon assimilation (22%), which indicated that down-regulation of CO₂ affinity caused by P-deficiency was, but not the only reason that resulted in the decline of photosynthetic efficiency. Instantaneous low-temperature significantly limited rETR_max under rich-P condition but had no effect on it when P was insufficient, and 1% ethanol could enhance rETR_max at low-P level but did not influence it at rich-P level. These two results proved that the decrease in thylakoid membrane fluidity caused by P-deficiency was another important reason that results in the decline of photosynthetic efficiency of M. aruginosa PCC7806.     

Effects of nutrients on arsenic accumulation by arsenic hyperaccumulator Pteris vittata L.

This research investigated the effects of various nutrients on arsenic (As) removal by arsenic hyperaccumulator Pteris vittata L. in a Hoagland nutrient solution (HNS). The treatments included different concentrations of Ca and K in 20% strength of HNS, different strengths of HNS (10, 20 and 30%), different strengths of HNS (10 and 20%) with and without CaCO₃, and different concentrations of Ca, K, NO₃, NH₄, and P in 20% strength of HNS. The plants were grown in nutrient solution containing 1 mg As L^?1 for 4 weeks except the Ca/K experiment where the plants were grown in nutrient solution containing 10 or 50 mg As L^?1 for 1 week. Adding up to 4 mM Ca or 3 mM K to 20% strength HNS significantly (P < 0.05) increased plant arsenic accumulation when the solution contained 10 mg As L^?1. Plant arsenic removal was reduced with increasing Ca and K concentrations at 50 mg As L^?1. Lower strength of HNS (10%) resulted in the greatest plant arsenic removal (79%) due to lower competition of P with As for plant uptake. Addition of CaCO₃ to 20% strength of HNS significantly increased arsenic removal by P. vittata. Among the nutrients tested, NO₃ and CaCO₃ were beneficial to plant arsenic removal while NH₄, P and Cl had adverse effects. This experiment demonstrated that it is possible to optimize plant arsenic removal by adjusting nutrients in the growth medium.     

Synthesis of theophylline derivatives and study of their activity as antagonists at adenosine receptors

The synthesis of oligo(ethylene glycol)-alkene substituted theophyllines in positions 7 and/or 8 is described. The binding activity at adenosine receptors of selected derivatives was studied. Compound 2 showed high affinity for human A_2B receptor (K_i = 4.16 nM) with a selectivity K_iA2A/K_iA2B of 24.1, and a solubility in water of 1 mM. The alkenyl substituent in some of the theophylline derivatives allows for covalent attachment of them onto hydrogen-terminated silicon substrate surfaces via hydrosilylation. Alternatively, an azido group was incorporated to an oligo(ethylene glycol)theophylline derivative as an anchor for tethering the molecules on ethynyl presenting surfaces via click reaction.     

Physiological responses of Ostreopsis ovata to changes in N and P availability and temperature increase

Ostreopsis ovata is a benthic dinoflagellate that produces palytoxin and ovatoxins. Blooms of O. ovata causing human health problems and mortality of benthic fauna have been reported from many tropical and temperate marine waters. In the present study we examined the combined effects of temperature and different nutrient conditions on the biochemical composition, growth, toxicity and carbohydrate production of an O. ovata strain originating from the Tyrrhenian Sea. O. ovata cultures with N:P ratios of 1.6, 16 and 160 (N deficient, NP sufficient and P deficient, respectively) were grown at 20 °C and 30 °C. Biomass accumulation, growth rates, cell volumes, biochemical composition, cell toxicity and carbohydrate production in each treatment were studied. Results indicated that under nutrient sufficiency O. ovata biomass accumulation increased significantly compared to N and P deficiency and also that N limitation severely affected growth. The highest growth rates were recorded at 30 °C. Cellular contents and the atomic ratios of C, N and P were higher in the cells grown at 20 °C than in those grown at 30 °C. O. ovata cell volumes increased at 20 °C. N deficiency significantly increased cell toxicity. Toxicity per cell was higher at 20 °C, but per carbon was highest at 30 °C. The highest carbohydrate production was found in conditions of N deficiency and at the lower temperature.Our study suggests that temperature increases due to global warming and nutrient enrichment of coastal waters stimulate the proliferation of O. ovata, particularly for the strains that have become adapted to warm temperate waters.     

Practical method based on saturated hydraulic conductivity used to assess clogging in subsurface flow constructed wetlands

This paper presents a simple method for evaluating the degree of clogging of subsurface flow constructed wetlands based on saturated hydraulic conductivity measurements. The method was applied to two full-scale wetlands located inside the wastewater treatment plants of two small villages (2000 PE) in the province of Lleida, Catalonia, Spain. In addition, to gain an insight into the mechanisms that lead to clogging, other measurements and analyses were carried out including the quantification of accumulated solids and belowground plant biomass. X-ray diffraction analyses were carried out to evaluate the mineral composition of accumulated sludge and granular medium. Hydraulic conductivity measurements and samples for solids analyses were taken along two transects that spanned the length of each wetland. Patterns for hydraulic conductivity were the same in both wetlands: very low values from the inlet zone to the middle (<20 m/d), clearly higher from the middle to 4/5 of the length (600–800 m/d), and lower very near the outlet (40–70 m/d). These results indicate that the first half of the length of both wetlands is highly clogged. Total solids (TS) were generally higher near the inlet than the outlet (TS_inlet = 3–15 kg/m²; TS_outlet = 1–9 kg/m²). Belowground plant biomass values were variable and did not show a clear pattern. In both wetlands the mineral fraction of the solids represented more than 75% of TS in most of the samples. X-ray diffraction analyses showed that the mineral composition of the solids coincided with that of the granular medium (mostly calcite and quartz). The proposed method based on hydraulic conductivity measurements is straightforward to use, does not require costly devices and allows to successfully evaluate the degree of clogging.     

Schoolhouse wastewater purification in a LWA-filled hybrid constructed wetland in Estonia

This paper analyses the purification efficiency and mass removal of organic material, suspended solids, nitrogen and phosphorus in a hybrid constructed wetland (CW) system treating wastewater from a basic school in Paistu, Estonia. The CW consists of two subsurface flow filter beds using lightweight aggregates (LWA): a two-chamber vertical subsurface flow (VSSF) filter bed followed by a horizontal subsurface flow (HSSF) filter bed, with a total area of 432 m². This CW was constructed in summer 2002 by the Centre for Ecological Engineering in Tartu (CEET). Eighteen series of water samples (from 30.10.2003 to 15.10.2005) were undertaken. The analyses show the outstanding purification effect of the system: for BOD₇ the average purification efficiency is 91%; for total suspended solids (TSS)—78%, for total P—89%, for total N—63%, and for NH₄N—77%. The average outlet values for the above-listed parameters were 5.5, 7.0, 0.4, 19.2 and 9.1 mg L⁻¹, respectively. According to our results, the purification parameters meet the standards set by the Water Act of Estonia for wastewater treatment plants of 2000–9999 PE: 15, 25, and 1.5 mg L⁻¹ for BOD₇, TSS and total P, respectively. The results show that hybrid CW systems consisting of subsurface flow filter beds can work efficiently in conditions of changing hydraulic loading and relatively cold climate. We did not find significant differences between the removal efficiency, mass removal, and values of the first-order rate-constant k for most water quality indicators during the warm (May–October) and cold (November–April) periods. Locally produced LWA as a filter material in CWs has shown good hydraulic conductivity and phosphorus sorption capacity (k = 17.1 ± 12.4 m yr⁻¹). The Paistu CW, with its proper design and outstanding purification results, can be considered one of the best systems in Estonia.     

Phosphorus availability and rootstock affect copper-induced damage to the root ultra-structure of Citrus

The control of several citrus diseases requires continuous applications of fungicides containing copper (Cu) which favor to the accumulation of this metal in the soil. Therefore, the evaluation of how nutrient availability and rootstock interact with Cu toxicity in the citrus trees is required to maintain sustainability of fruit production in Cu-contaminated soils. Valencia orange trees on Sunki mandarin (SM) or Swingle citrumelo (SC) rootstock were grown in nutrient solutions combining adequate Cu (1.0 μmol L⁻¹), excess Cu (50.0 μmol L⁻¹), deficient phosphorus (P) (0.01 mmol L⁻¹) and sufficient P (0.5 mmol L⁻¹). The excess Cu reduced root and shoot growth, chlorophyll and relative water content in the leaves of the trees compared to those under adequate Cu supply. Furthermore, excess Cu caused severe damage to the root ultra-structure, characterized by the degeneration of the middle lamella and the presence of a thin and sinuous cell wall, as well as, starch accumulation in the plastids, disruption of the mitochondrial membranes and cellular plasmolysis. The damage caused by excess Cu in the cell wall and middle lamella on the root cells of SC was less severe than SM. Sufficient P supply improved the structure of the cell wall and middle lamella of trees subjected to excess Cu in comparison to P-deficient ones. Thus, the occurrence of more preserved cell wall and middle lamella supports the idea that sufficient P availability in the rooting medium and the use of SC rootstock might contribute to increase the ability of young citrus trees to cope with Cu toxicity.