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Revista de la ciencia del suelo y nutrición vegetal

versión On-line ISSN 0718-2791

R.C. Suelo Nutr. Veg. v.8 n.especial Temuco  2008 


J. Soil Sci. Plant Nutr. v8 no especial 2008 (211-234)



Session 3. Soil-Root-Microbe Interaction & Their Effects on Biophysical Transformation, Fate, and Toxicity of Metal and Metalloids

Effect of Biosolid Incorporation on Cu, Cr, Ni, Pb and Zn Extractability in Mollisol and Inceptisol Soils of Central Chile

I. Ahumada1*. M. Ávila1, E . Contreras1, L. Asear1, P. Richter1 and M.A. Carrasco2

1 Universidad de Chile, Facultad de Ciencias Químicas y Farmacéuticas, Departamento de Química Inorgánica y Analítica, Casilla 233, Santiago, Chile. 2Universidad de Chile, Facultad de Ciencias Agronómicas, Departamento de Ingeniería y Suelos. Casilla 1004 Santiago, Chile. *E-mail:

Keywords: Biosolid; wastewater; mollisol and inceptisol soils.

In Santiago city almost 100% of the urban wastewater will be treated by the end of 2010, which implies the generation of large volumes of biosolids that will need to be disposed to avoid an environmental problem. An interesting alternative to face the accumulation of these residues is their application in agriculture as soil amendment and as a complement of the soil fertilization. This could be an attractive option of disposal and recycling of these residues. Biosolid application to soil assumes a significant supply of nutrients and micronutrients essential to plants. Nevertheless, it has been found that biosolid application to soil incorporates trace metals and other organic toxic compounds that may interact with organic matter or other soil components. It is, thus, necessary to learn how biosolid incorporation affects agricultural soils in relation to availability of trace metals such as Cu, Cr, Ni, Pb, and Zn. The purpose of this study was to determine the effect of biosolid amendment on Cu, Cr, Ni, Pb and Zn extractability from the soil components through the BCR (Community Bureau of Reference) procedure, which considers three sequential extraction to obtain an acid-soluble fraction (metal soluble, exchangeable and bound carbonates), a reducible fraction (metal associate with iron/manganese oxides), an oxidizable fraction (metal bound to organic matter and as sulphides) and residual fraction. The study included agricultural soils of the Mollisol and Inceptisol order. Soil samples were collected from the surface soil layer (0- 20 cm) at six different sites close to the city of Santiago in the Metropolitan Region. These were amended with biosolids at rates of 0, 15, 30, 45 and 60 Mg ha_1 and incubated for 60 days under controlled conditions of moisture and temperature. Among the metals determined in biosolids and soils, Zn, Cu and Cr showed the highest concentration. Total trace metal contents found were below the limits allowed by the Chilean regulation for both samples. Biosolid fractionation showed that Ni and Zn acid-soluble fraction was predominant and the incubation process increased this fraction for both metals, which confirms the greater availability of these metals in biosolids. In soils generally, without considering the residual fraction, the metals were found predominantly as reducible and oxidizable form and in the Zn case the first fraction was important too. Biosolid application to agricultural soils had effect on trace metal distribution in some of them, with an important increase in Zn availability in all the soils and Cu in some of them, while Cr, Ni and Pb showed no significant variations in most of them. A two-level factorial design was satisfactorily applied to assess the effect of biosolid application rate and the incubation time.

Acknowledgements: This research was supported by FONDECYT, Project 1080357.


Use of Sewage Sludge and Mussel Shells in Mine Soils Recovery

M.L. Andrade*. E.F. Covelo and F.A. Vega

Departamento de Biología Vegetal y Ciencia del Suelo. Universidad de Vigo, España *E-mail:

Keywords: Minesoils; recovery; amendments.

Opencast mines cause serious environmental impact like the destruction of natural soils, the extraction of important volumes of materials, and produce a large amount of waste because the ore is a small fraction of the total volume of the mined material. This causes the formation of new soils (Anthropic Regosols) on the accumulated wastes of the mine with low contents of nutrients and organic matter and unfavourable texture and structure. High levels of trace elements and the acidic drainage due to the oxidation of sulphide are also frequently common characteristics of the most mine tailings. The scarce vegetation leads to acidification of adjacent soils by leaching the mine spoils, giving rise to high environmental contamination. The extraction of metal ores causes generally a multielemental contamination of the environment In addition; the natural metal content of the soils will be increased if materials with heavy metals are added. The restoration of soil function and mining soil quality is essential to long-term ecosystem stability. In Touro (Corana, NW Spain, 42°52'34" N, 8°20'40" W) there were two opencast mines, Arinteiro and Bama, from which were extracted chalcopyrite and pyrrotine between the years 1973 and 1988. The copper extraction stopped in 1988 and, since then, materials for road construction were extracted. The mine spoils cover a surface of 760 ha, and were formed of great piles of coarse rock fragments with strong slopes. In addition they did not begin to be recovered until two years ago. The only measures of the recovery made were the addition of sewage sludge, like source of organic matter, and mussel shells, like liming material. This study was conducted to evaluate, at four tailings from this opencast mine, the effects of sewage sludge and mussel shells on chemical characteristics and heavy metal content of minesoils. The superficial horizons of eight soils were sampled. Four come from the depleted copper mine of Touro treated with sewage sludge and mussel shells during two years and four soils without treatment. The main chemical limitations of untreated mine soils are their acidity (pH from 3.62 to 4.78), low organic matter content (from 1.12 to 12 mg kg-1 ), low CEC (from 1.83 to 9.47 cmol(+) kg-1 ), and Ca+ and Mg+ contents, and imbalance between exchange bases. The used amendments modified many of these characteristics. Thus, the treatment increased soil pH (varies from 7.8 to 8.1), organic matter content, that vary between 61 and 77 mg kg-1, also it increased considerably the CEC (from 60 to 156 cmol(+) kg-1 ), as a result of high increases of exchangeable Ca and Mn contents (from 51 to 137 and from 5.94 to 6.02 respectively). Nevertheless it continues the imbalances between exchange bases. All the soils contain high concentrations of Cu, Pb, Ni, and Zn. The Cu, Cr, and Ni levels surpass the maximum limits established in some reference guides. The used amendments increase the Cd, Cu, and Pb availability and diminish the total content. Nevertheless Cr, Ni, and Zn availability diminish and total content increase. It is probably due to the formation of Cu, Cd and Pb soluble organic complexes, and to Cr, Ni and Zn fixation when the pH increases until basic values. Therefore it is necessary to modify the dose of the amendments to reduce organic matter content and pH, and to balance the exchangeable cation content. Also it is advisable to add substances that adsorb heavy metals, like clay minerals, and fertilizers to ameliorate the mine soils. These used amendments, and the proposed ones, can improve the soil properties, and enhance soil functions and quality.


Bacterial Community Structure Associated with the Rhizosphere of Cirsium arvense in an Italian Arsenic-Contaminated Soil

V. Andreoni*. C. Romagnoli, K. Tunesi, R. Zanchi, S. Bachate, M. Colombo, L. Cavalca and

E. Canzi

Department of Food Science Technology and Microbiology, University of Milan, Milan, Italy.


Keywords: Alphaproteobacteria; rhizosphere; arsenic-contaminated soil.

The rhizosphere bacterial community of a spontaneous plant (Cirsium arvensé) inhabiting an Italian soil located in Tuscany Region contaminated with arsenic (As tot= 214 mg/Kg) was characterized by cultivation-independent and -dependent methods. FISH analysis showed that the dominant bacteria present in bulk and in rhizosphere fraction of soil belonged to Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria and Pseudomonas. Both soil fractions contained high percentage of arsenate and arsenite resistant bacteria. Most of the culturable As-resistant isolates were affiliated to the Alphaproteobacteria. Some of the isolated strains possessed genetic determinants for As-resistance. Genes coding for arsenite efflux pump (arsB and acr3p) and arsenate reductase (arsC) were PCR amplified from both As (V) and As(III) resistant bacteria. Several aerobic microorganisms that are potentially involved in As-cycling were isolated from the enrichment cultures. When grown in a defined liquid medium containing an organic carbon source, a group of the As (III) resistant bacteria transformed efficiently arsenite to arsenate. Among them there were Sinorhizobium sp., and Ancylobacter sp. which rapidly grew chemolithoautotrophically. The microbial oxidation of As (III) to As (V) impact the mobility and speciation of arsenic in rhizosphere where plants could uptake arsenate via phosphate transporter. Moreover, As (III) oxidation is the basis for bioremediation of As(III) polluted systems, because As(V) can be immobilized onto strong adsorbents.

Speciation of Manganese in Oxisols of the Mining Area of Moanda (Gabon, Africa), and Its Possible Impact on Growth of Maize (Zea mays) and Cassia (Chamaecrista rotundifolia), and Heavy Metal Bioaccumulation, under Greenhouse Conditions

F. Andreux1*, C. Boupassia1'3'4, J. Lévéque2, A. Edou-Minko3 and P. Nziengui4

1 UMR Microbiologie des Sols et de VEnvironnement INRA - Université de Bourgogne 6, Dijon, France. 2UMR Bio géo sciences CNRS - Université de Bourgogne 6, Dijon, France. 3Université des Sciences et Techniques de Masuku - Unite de Recherche en Sciences de la Terre et de VEnvironnement (URESTE) Faculté des Sciences Franceville, Gabon. "Compagnie Miniére de l'Ogooué (COMILOG), Cellule Assurance-Qualité et Environnement - Moanda, Gabon. *E-mail: or

Keywords: Manganese; oxisoil; maize.

A set of soils with high manganese (Mn) content was collected in the manganese mining area of Moanda, in the South West of the Republic of Gabon. The prevailing mineral substratum is an ampelite from mid-Precambrian and the soils are developed on the 3-5 m thick overlaying material, which is composed of ferruginous pisolites, the main weathering product of the ampelite. Assays with either locally cultivated species (maize, peanut, manioc.) or native legume grasses (Pueraria phaseoloides, Crotalaria retusa, Chamaecrista rotundifolia...) were run, both in the field and in the greenhouse, with the objective of optimizing the vegetation re-growth on the mine landscape. In the present work, a greenhouse experiment with maize (Zea Mais) and round-leaf cassia (Chamaecrista rotundifolia), grown on two soil materials, was carried out. The test material (S-Mn) was a composite of surface soils with high Mn content, developed on pisolitic deposits. The blank material (S-Bl) was a composite of surface soils with low Mn content, developed on weathered bedrock of the foothill, near Moanda. In soils composing the S-Mn material, the total Mn content was frequently higher than that of Al or Fe (100-300 %o of soil weight, oxide basis; exchangeable Mn ranged from 0.2 to 1.5 cmol (+) kg-1 and occupied from 100 to 650 %o of the effective cation exchange capacity, concurrently with Al ions. When soil samples were submitted to ammonium oxalate (Ox) and citrate-bicarbonate-dithionite (CBD) reagents, Mn was highly soluble (700-900 %o) in both reagents, Fe was moderately soluble in Ox (100-200 %o) and CBD (400-600 %o), and Al was much less extractable. The S-Mn material did not significantly modify the germination rate of both seeds, although the growth was frequently slower and the 60 day-old seedlings showed more yellowing and necroses on S-Mn than on S-Bl, especially for C. retundifolia. The accumulation of Mn in stems and roots of both plants was about 8 times higher on S-Mn than on S-Bl. Other metals (Cu, Co and Pb) followed the same tendency, but with much less differences and only in the roots of one or both plants. For most nutrients and other metals (Fe, Cr and Ni), bioaccumulation could be lower on S-Mn than on S-Bl. These results suggest that, in the peculiar type of Oxisol tested in our study, biological activity was still high and able to sustain the nutrition of the seedlings. The high dissolution of Mn by organic acids seemed to be responsible for the main disturbance in plant growth and metal mobility, especially in the absence of any fertilization.

Arbuscular Mycorrhizal Fungal Communities Associated to Four Metallophytes in a Mediterranean Ecosystem Affected by Copper Mining

P. Cornejo1*, E. Sieverding2 and F. Borie1

1Departamento de Ciencias Químicas, Universidad de La Frontera, Temuco, Chile;2Institute of Plant Production and Agroecology in the Tropics and subtropics, University ofHohenheim, Stuttgart, Germany. *E-mail: pcornejo@ufro.c¡

Keywords: Arbuscular mycorrhizal; metallophytes; copper mining.

The arbuscular mycorrhizal fungi (AMF) form mutualistic symbiosis with most of terrestrial plants, which obtain a benefit from a greater nutrient uptake and an improved tolerance to abiotic stresses, including the heavy metal (HM) presence in the soil. The HMA diversity can be affected by high levels of HM, producing structural changes in the plant communities. In central Chile several ecosystems have been affected by the deposition of particulate matter and wastes originated in diverse mining activities, which have accelerated strong processes of loss of plant cover and diversity. Nowadays, these communities are only composed by some species able to tolerate high HM levels, specifically Cu, Zn and As. The aim of this work was to study the AMF diversity in an ecosystem affected by mining activities, with the view to obtain native AMF ecotypes to use in bioremediation programs using endemic metallophytes from Central Chile. The communities of AMF associated to four plants (Argemone subfusiformis, Baccharis linearis, Oenothera affinis and Polypogon viridis) from a Mediterranean scrubland near the Ventanas smelter (Puchuncavi Valley, Valparaiso, Chile) presenting a strong Cu pollution and soil acidity, were analyzed. Spores collected from rhizospheric soil were morphologically characterized as well as by the use of PCR-TTGE analysis of RNA-SSU. The results showed a strong effect of the plant specie on the size and structures of the fungal communities, being the population size more than 600 spores in 100 g of soil in O. affinis rhizospheric soil, whereas in the other plant species the total spore community size was 3-6 times smaller. A high specificity was found between O. affinis and an AMF ecotype (Acaulospora spl, probably a not previously described AMF specie), registering low specific richness (S'=2) and high dominancy (λ= 0.97). The other metallophytes presented AMF communities with high ecotypes richness (between 3.7 and 6 ecotypes) and homogeneity (H' between 0.86 and 1.51). Acaulospora spl was also found in the AMF communities associated to the other plant species analyzed, suggesting a greater adaptation to the stress conditions by high levels of Cu in the soil, in addition to a strong effect of the plant species on the fungal community structure. Moreover, there was a high specificity between Acaulospora spl and O. affinis, that could be related with a probable Cu bioaccumulator potential in this plant specie recently described, making possible its use in phytoremediation processes in ecosystems affected by mining activities in Central Chile.

Acknowledgments: This study was supported by Fondecyt 3070052 Grant.

Mycorrhizal Fungal Propagules in a Mediterranean Ecosystem Polluted by a Copper Smelter in Central Chile

P. Cornejo1*. S. Meier1,1. González2, A. Neaman2 and F. Borie1

1Departamento de Ciencias Químicas, Universidad de La Frontera, Casilla 54-D, Temuco, Chile. 2Área de Medio Ambiente, Facultad de Agronomía, Pontificia Universidad Católica de Valparaíso, Casilla 4-D, Quillota, Chile. *E-mail:

Keywords: Mycorrhizal fungal; mediterranean ecosystem; copper smelter

Arbuscular mycorrhizal fungi (AMF) are present in the most of terrestrial ecosystems, contributing to the establishment and nutrition of plants through mutualistic symbiosis. In heavy metal (HM) polluted ecosystems, the arbuscular mycorrhizal (AM) symbiosis plays a fundamental role, increasing the presence and dissemination of metallophytes able to tolerate high levels of HM. In Chile the role that plays the AMF in ecosystems extremely affected by mining activities is still unknown, which is not related with the high number of mining deposits and ecosystems contaminated by mining practices. Based on the above mentioned, the objective of this study was to quantify the density of AMF propagules in HM polluted soils and to analyze the relation with the Cu pollution level. To carry out this study, rhizospheric soil of four metallophytes species present in a Mediterranean ecosystem influenced by the activities of the Ventanas smelter (ENAMI) at the Puchuncavi Valley (Valparaiso, Central Chile) was collected. The selected plants were Argemone subfusiformis, Baccharis linearis, Oenothera affinis and Polypogon viridis. The levels of available Cu fluctuated between 18.8 and 326 mg kg-1 in the rhizospheric soil of O. affinis and P. viridis, respectively. On the other hand, the density of mycorrhizal mycelium were found between 1.2 and 4.5 m g-1 in B. linearis and P. viridis, respectively, being this parameter highly correlated with the available Cu (r=0,83; p<0,01). The density of AMF spores was scarce in all the plant species (between 100 and 200 AMF spores in 100 g of soil), except in O. afflnis, that showed densities near the 600 AMF spores in 100 g of soil. The mycorrhizal colonization fluctuated between 33.9% in B. linearis and 57.9% in A. subfusiformis. The rhizospheric soil of P. viridis presented the highest value for the acid P-ase activity (2.6 μg PNF g h-1 ); whereas O. afflnis presented the higher value for dehidrogenase activity (11 |_ig INTF g-1 h-1 ), 2 to 11 folds higher than in the other soils. These results are the first reported for AMF propagules densities in highly HM polluted environments in Central Chile, and suggest an important role of this kind of soil fungi in the establishment of some endemic metallophytes under this type of environmental stress. Particularly, P. viridis presents the highest mycelium density associated with high levels of P-ase activity (and available P), which could allow its growth in extreme conditions and that could support its inclusion in further phytoremediation programs. Moreover, this study gives new evidence on the role of AMF in soils polluted by mining activities, and suggests a strong influence of the plant in the development of the different AMF propagules in HM polluted soils.

Acknowledgments: Hiis study was supported by Fondecyt 3070052 Grant.


Bioavailability of As and P in the Rhizosphere of Plants Inoculated with Arbuscular Mycorrhizal Fungi

V. Cozzolino*. M. Pigna, V. Di Meo and A. Violante

Department of Soil, Plant, Environmental Sciences and Animal Production, University of Naples, Federico II, Via Universitá 100, Portici, Italy. *E-mail address:

Keywords: Glomus intraradices; Lactuca sativa; arsenic.

Arsenic (As) is ubiquitous in the environment and it is highly toxic to most biological systems. Soil unproductiveness, due to build up of arsenic (As) concentrations in soils from human activities is an environmental concern. Elevated soil As concentrations generally decrease the ability of the soil to support an economically viable crop on contaminated sites, but phytotoxicity of As is often reached prior to accumulation levels that would be toxic to wildlife or to humans ingesting the plants. Understanding plant uptake metabolism of As is indispensable for the development of technologies that alleviate As toxicity in the crop plants. Arbuscular mycorrhizal fungi (AMF) can form symbiotic relationships with the vast majority of land plants, and are known to benefit the phosphorus (P) of the host plants by increasing P acquisition. Due to the physicochemical similarity between phosphate and arsenate, AMF are likely to have a strong influence on arsenate uptake and resistance. The fate of As in the rhizosphere and its interaction with P, in presence of AMF originating from different environment, has not yet been fully explored. In this study, we examined the role of AMF on As uptake in lettuce plants (Lactuca sativa cv. Trocadero) growing in a As-polluted soil collected from Tuscany, Italy. In particular, we tested the effectiveness of native AMF or inoculation with a commercial inoculum (colonized by Glomus intraradices) with and without P fertilizer on the growth and uptake of As by lettuce and their effects on As and P fractions in rhizosphere and bulk soil. The soil under investigations was a clay loam, alkaline (pH 8.2), containing 51 mg kg-1 total CaC03 7.5 mg kg-1 P (Olsen) and 250 mg kg-1 total As. Greenhouse pot experiments (six replicates each) were established in a randomized block design. Plants were grown for three months, with and without a commercial inoculum (M+, M-) and two levels of P application (P- at 0 kg P ha-1 and P+ at 90 kg P ha-1). At harvest, soil (bulk and rhizo soil) and plant (root and leaf) samples were collected. Commercial inoculum and P application increased plant biomass enormously and the highest plant growth was in M+P+ treatment. Leaves and root As concentrations were slightly reduced by inoculation with commercial inoculum where P was added. On the contrary, M+P- treatments showed the highest As root concentrations. Root and leaves As and P concentrations in M-P- plants was lower compared with M+P- treatment. Root and leaves P concentrations in M+P+ was higher compared with M-P+ treatment. Analyses of rhizo and bulk soils at the harvest indicated that commercial inoculum plus P fertilization did not increase more available As fractions in the rhizosphere, but evidenced a P soil depletion respect to plants M-P+. Commercial inoculum protected its host plants from toxicity of excessive As through P nutrition by absorbing more P from the soil.

Modelling of Heavy Metal Sorption and Retention by Antropic Regosols

E.F. Covelo*. F.A. Vega and M.L. Andrade

Departamento de Biología Vegetal y Ciencia del Suelo. Facultad de Ciencias, Universidad de Vigo, Lagoas, Marcosende, Vigo 36310, Spain. *E-mail:

Keywords. Sorption; Desorption; Modelling.

Many mine soils are chemically, physically, and biologically unstable and deficient. They are sometimes amended with sewage sludge and ashes but often contain heavy metals that increase the already high mine soils heavy metal contents. Cd, Cr, Cu, Ni, Pb, and Zn in mutual competition were added to five mine soils (Galicia, Spain). The studied mine soils (all Anthropic Regosols) are located at the former copper mine at Touro (Spain), from which road material is currently extracted, and at the lignite mine at Meirama. Ten solutions of mixtures of Cd, Cr, Cu, Ni, Pb, and Zn nitrates (between 10 and 400 mg L-1) were prepared to obtain adsorption isotherms. Twelve grams soil samples were treated with 200 mL of solution, shaken for 24 hours at 25°C. After centrifugation, metal in solution was determined by ICP-OES. Desorption experiments were performed using the pellets resulting from adsorption experiments. Each sample was treated with 200 mL of an acetic acid (0.02M) and sodium acetate (0.02M) solution, buffered at pH 4.5 and shaken for 24 hours at 25°C. All of the experiments were performed in triplicate. Soil capacities for heavy metal sorption and retention were determined by means of distribution coefficients. In previous papers Kdxsp proposed by Kaplan was adapted in order to evaluate the total sorption and retention capacity of all heavy metals that contain the added solution to soils. By means of Kdzsp medium values and data of soil properties it was made a multiple linear regression analysis in order to predict the heavy metal sorption and retention capacities of mine soils. This analysis was made by backward stepwise. This predictive model could be applied to soils that present their characteristics within the interval established by the studied minesoils properties. The selected variables to elaborate these predictive equations were the contents of humified organic matter (HOM) and Fe, Mn and Al oxides (mg kg-1 ), the cationic exchange capacity (cmol kg-1 ) and the proportion in soils of sand, silt and clay. The obtained equations are:

Sorption:Kd∑sp = -0.32sand -1.09 clay-1.92 CECe + l. 66 FeOx-2.37 MnOx +52.09.

R2 = 0.99.

Retention: Kd∑sp = -0.56sand-5.68clay-1.21 HMO-15.50CECe +14.35FeOx + 232.12.

R2 = 0.98.

The properties of studied soils that influence their sorption capacity and therefore they will influence the capacity of soils with similar characteristics are the sand and clay proportion, the cationic exchange capacity and the Fe and Mn oxides content (mg kg-1 ). The variables that allows predict the heavy metals retention capacity of similar minesoils are the percentage of sand and clay, the cationic exchange capacity and the contents of humified organic mater and Fe oxides (mg kg-1 ). Therefore the content of Mn oxides more influences the heavy metal sorption capacity of minesoils whereas after desorption experiments the minesoils humified organic matter influences their retention capacity. We represent the empirical data of heavy metals sorption and retention capacities as opposed to the predicted ones by means of multiple linear regression. They were obtained two correlation coefficients: Sorption = 0.99 and Retention = 0.975.

Assessment of Heavy Metals in Selected Vegetables from Harichand Area in District Charsadda, Pakistan

M. Ibrahim1* and I.U. Mohammadzai2

1Postgraduate Programme in Biochemical Toxicology, Department of Chemistry, CCNE, Federal University of Santa Maria, CEP 97105-900, Camobi, Santa Maria, RS, Brazil. 2Institute of Chemical Sciences University Of Peshawar, Pakistan. *E-mail:

Keywords: Trace metals, Pollution, Bioavailabihty, Pakistan, WHO.

Consumption of vegetables and fish contaminated with the heavy metals Fe, Zn, Cu, Ni, Cr, Pb and Mn is the most likely route for human exposure in Harichand area in district Charsadda, Pakistan. The present study was undertaken to investigate the bioavailabihty of heavy metal in vegetables and to assess the extent of heavy metal contamination of vegetation due to irrigation with canal water on agricultural land of Harichand District Charsadda. -Digestion of samples (2 g each) was carried out using 10 ml nitric acid, according to the procedure used for soil samples (Lark, 2002) using Hitachi Zeeman Japan Z-8000, Atomic Absorption Spectrophotometer equipped with standard hallow cathode lamps as radiation source and air acetylene flame were used for absorption measurement of elements Cr, Cu, Ni, Zn, Cd, Pb and Fe. The present results demonstrate that the general order of decreasing metals concentration in the vegetables was Fe>Zn> Mn>Cr>Cu>Ni>Pb and the decreasing order of metal contents in the soil sample was Fe>Mn>Zn>Cr>Ni>Cu>Pb. -The present results demonstrated that the results show the presence of the heavy metals in vegetables, below the WHO standards.


Indian J. Environ. Health (2002) 44: 164-167.


Effect of the Aluminum (Al) Toxicity on the Lipid Peroxidation, and Antioxidant Activity in Blueberry Vaccinium corymbosum L.

C. Inostroza-Blancheteau1*, M. Reyes-Díaz2, M. Alberdi3 and M.L. Mora3

1Programa de Doctorado en Ciencias de Recursos Naturales, Universidad de La Frontera, Casilla 54- D, Temuco-Chüe. 2Instituto de Agroindustria, Universidad de La Frontera, Casilla 54-D, Temuco-Chile. 3Departamento de Ciencias Químicas, Universidad de La Frontera, Casilla 54-D, Temuco-Chile. *E-mail:

Keywords: Blueberry; aluminium phytotoxity; antioxidant activity.

In South-Central Chile where acid soils are predominant, blueberry {Vaccinium corymbosum L.) is an important agronomic activity. The acidity of these soils increases the aluminium phytotoxity (Al3+), with a concomitant decrease of the productivity and strongly yields reductions. Some alternative for the decrease of the concentration of exchangeable Al3+ is the application of calcium amendments. However, this application is only conditioned to the first centimeters of soil. Other alternative is the searching of genome of plants with aluminium (Al) resistance. Currently, did not exit studies involved with the identification of germoplasm of blueberry with Al resistance characteristics. In this work the Al response of three cultivars: cv. Brigitta, cv. Legacy and cv. Bluegold (the most important cultivars cultivated in Chile) was evaluated in a nutritive solution with different Al concentration (0, 25, 50, 100 y 200 µM Al3+) at different times (0-20 days). The Al resistance level was estimated by the lipid peroxidation throughout the TBARS method in roots and by the antioxidant activity in roots and leaves using the DPPH method. The results showed that the three cultivars had a differential Al resistance (P< 0.05), being cv. Legacy the most resistance cultivar (5 to 10 mmol MDA g-1 FW). Cultivar Brigitta and cv. Bluegold showed higher lipid peroxidation (up to 83 mmol MDA g-1) and therefore the greater Al sensitivity. These results are associated with a lower decrease of the antioxidant activity in roots with respect to the initial values in cv. Legacy (34%) following by cv. Brigitta (40%) and cv. Bluegold (58%). Interestly, cv. Legacy showed initially a higher antioxidant activity (7.4 mg TE/gFW) in leaves than the other cultivars (2.4 mg TE/gFW), suggesting a constitutively larger antioxidant activity of cv. Legacy. This high antioxidant activity and its low lipid peroxidation suggest a higher Al resistance in cv. Legacy. We concluded that in the long term cv. Legacy showed the best behavior to Al stress, differently as showed previously in short term experiments performed by our group.

Acknowledgments. This project was supported by Bicentenario-Conicyt PSD-26 and Fondecyt 1080372.


Copper and Zinc Content in Oenothera afflnis at Increasing Cu Levels as Affected by Different Ecotypes of Arbuscular Mycorrhizal Fungi

S. Meier, A. Seguel, F. Borie and P. Cornejo*

Departamento de Ciencias Químicas, Universidad de La Frontera, Casilla 54-D, Temuco, Chile. *E-mail: pcornejo@ufro. cl

Keywords: Copper; Oenothera afflnis; arbuscular mycorrhizal.

Metallophytes can grow in soils with high heavy metals (HM) levels, as those that have received the deposit of wastes of mining activities. In Chile, copper (Cu) mining takes place in a vast geographic area, affecting several natural ecosystems. However, knowledge of metallophytes and its ability to stabilize or accumulate HM is scarce, and less known is the role of arbuscular mycorrhizal fungi (AMF) under these conditions. The AMF has several mechanisms allowing the establishment of plants in soil HM polluted, promoting phytostabilization and/or phytoextraction processes. The aim of this study was to analyze the effect of mycorrhizal symbiosis and the type of AMF on the Cu and Zn uptake and compartmentalization in Oenothera afflnis at increasing soil Cu levels to establish its potential use in phytoremediation. For this, seedlings of O. afflnis were grown in: i) rhizospheric soil of O. afflnis with the native AMF inoculum (+M); ii) soil inoculated with Glomus claroideum (GC), and iii) sterile soil (-M). The growing substrate was sterile soil from a Mediterranean ecosystem near Ventanas smelter (Puchuncavi Valley, Valparaiso, Central Chile), which presented 178 mg kg-1 of available DTPA-Cu. This soil, was used to form seven increasing Cu levels, which consisted of 0, without soil (a mixture of sand and sepiolite); 1, soil diluted with sand and sepiolite (1:1:1); 2, soil undiluted; 3, soil with 75 mg Cu kg-1 (as CuS04 solution); 4, soil with 150 mg Cu kg-1 ; 5, soil with 225 mg Cu kg-1 and; 6, soil with 300 mg Cu kg-1 . Plants grew for 12 weeks and subsequently the contents of Cu and Zn in shoot and root, as well as the DTPA-available content of both HM in the soil, were quantified. The Cu concentration in shoot increased in all mycorrhizal treatments until level 3 (from 20 to 50-55 mg Cu kg-1 in level 0 and level 3, respectively), and decreased at higher soil Cu levels (levels 4 to 6) up to 30-40 mg Cu kg-1 . In root was observed high Cu concentrations, especially in the +M treatments (up to 748 mg Cu kg-1 in level 6), significantly different to the other treatments (up to 326 and 603 mg Cu kg-1 in GC level 5 and -M level 6, respectively). The Zn was also preferentially concentrated in roots, being observed in this case that treatment -M presented a higher Zn concentrations (up to 313 mg Zn kg-1 in level 6) compared to the other mycorrhizal treatments (up to 192 and 174 mg Zn k-1 in +M level 3 and GC level 3, respectively). The bioaccumulation of Cu was higher in the root compared to the shoot (up to 39 and 88 mg Cu kg in the treatment +M level 6, for the shoot and root, respectively); while the Zn was preferentially accumulated in shoot (up to 56 mg Zn kg-1in +M level 2 for the shoot and up to 30 mg Zn kg-1 in -M level 1 for the root). Cu and Zn contents in both organs showed independence of the levels reached by both HM in the soil. These results suggest complexes mechanisms of HM homeostasis by O. afflnis, in which the mycorrhizal association would favor the Cu transport and accumulation in the root, but could prevent the Zn accumulation in this organ. In addition, these results suggest a strong potential of the symbiosis between O. afflnis and some ecotypes of HMA natives from soils polluted by high Cu levels to promote phytostabilization processes.

Acknowledgments: This study was supported by Fondecyt 3070052 grant.

Effect of High Biosolid Rates on Copper Mobility in Contaminated Soils

J. Mendoza* and T. Garrido

Facultad de Ciencias Químicas y Farmacéuticas. Universidad de Chile, Casilla 233, Santiago, Chile. *E-mail: jmendoza@ciq. uchile. cl

Keywords: Copper; DOM; contaminated soils.

A strategy for heavy-metal stabilization in mining wastes and soils contaminated with mine wastes is the use of organic amendments, biosolids among them, in order to decrease concentration of metals in solution, leaching to deeper layers, and drifting by the wind. This practice, in turn, provides nutrients and organic matter favouring the growth of vegetation 1,2. In the present study, the effect is assessed of a biosolid applied at rates of 100 ton/ha to copper-contaminated soils. The study included two soils (0-20 cm depth) near the cities of Rancagua (Clay loam) and Santiago (Loam) in the central valley of Chile, with Cu total contents of 1046 and 475 ppm, respectively. The biosolid was obtained from a household wastewater treatment plant. Chemical analysis of both substrates was carried out with standardized methods for soils. For mobility studies, glass chromatographic columns were used, packed with 45 g of soil. The columns, previously moistened, were eluted with 10-4 M CaCl2 at a flow rate of 0.1 mL/min, by means of a peristaltic pump. The eluate was collected in 300-drop fractions by means of a fraction collector. The biosolid was added at the top of the column. In the eluted fractions, determinations were done of pH, total Cu by FAAS, free Cu by ISE and organic carbon. Each column was prepared in duplicate and was eluted for about 48 h to obtain from 15 to 20 pore volumes (pv). In both soils, with no sludge application, an increase of eluted Cu was observed about the first 5 pv, decreasing then to undetectable values about 15 pv. With this volume, the accumulated total was 207 and 82 µg Cu for Loam and Clay loam soils, respectively. Rapid elution of dissolved organic matter (DOM) from the soil was observed, but its peak decreased faster than Cu peak and before 10 pv. Sludge application implied mobilization of a remarkable amount of copper and organic matter compared with control soils, even though the elution pattern of both analytes showed similar characteristics. The amount of eluted copper after 15 pv was 1137 and 148 µg, values which are 5.5 and 1.8 times higher than control for Loam and Clay loam soils, respectively. On the other hand, organic matter now mainly provided by sludge, showed increases of 4.3 and 2.6 times compared with control for the same soils, respectively. Application of mass balance determined that the Cu mobilized from soils by DOM corresponds to 881 and 16.4 µg, for Loam and Clay loam, respectively, which is about 9.4% and 3.8% of available Cu. These results would indicate that the mobile fraction of the biosolid organic matter has a significant effect on Cu mobility in soils contaminated by this metal. Acknowledgments: This study was supported by FONDECYT, project N° 1080632

Keywords: Copper; DOM; contaminated soils.


J. Kumpiene, A. Lagerkvist and C. Maurice (2008). Waste Management 28: 215-225. E. Madejon, A. Perez de Mora, E. Felipe, P. Burgos and F. Cabrera. (2006). Environmental Pollution 139: 40-52.


Effect of Aluminium (Al3+ ) Toxicity on Total Soluble Sugars in Blueberry Cultivars (Vaccinium corymbosum L.)

R. Millaleo1*, M. Reyes-Díaz2, M. Alberdi3 and M.L. Mora3

1Doctorado en Ciencias de Recursos Naturales, Facultad de Ingeniería, Ciencias y Administración, Universidad de La Frontera, Casilla 54-D, Temuco, Chile; 2Instituto de Agroindustria, Universidad de La Frontera;sDepartamento de Ciencias Químicas, Universidad de La Frontera. *E-mail: rayenmillaleo@gmail. com

Keywords: Aluminum toxicity; total soluble sugar; Vaccinium corymbosum.

The aluminum toxicity (Al3+) is the most important factor that inhibits the plants growth in acid soils, because it accumulates in the roots cells causing the inhibition of the cell elongation. The carbohydrates are direct products of the photosynthetic activity and they constitute an energy source and metabolites, which represent significant percentages of the dry weight of woody plants. The objective of this work was to determine the total soluble sugars (AST) content in three cultivars of blueberry (Vaccinium corymbosum L.) in Hoagland solution at different times (0, 7, 14 and 20 days) and aluminium (Al) content (0, 25, 50, 100 and 200 uM). The three cultivars showed different AST levels in leaves and roots under Al stress. Generally, cv. Legacy showed a decrease with Al treatments at different times in leaves, but a highest decrease (62%) was observed in cv. Brigitta at 200 uM of Al treatment compared with the control ones. In roots, cv. Legacy maintained the higher AST concentration with respect of the others cultivars under aluminium stress. Differently, cv. Bluegold did not change the AST concentration in leaves and roots. The role of AST in aluminium stress is unclear, since both high and low levels of AST can be involved in the improvement to the oxidative stress depending of species. We concluded that cv. Legacy showed a better acclimation under Al stress, existing an increase in the AST concentration in roots at different the times, maintaining the AST levels in leaves.

Acknowledgments: This study was supported by Bicentenario-Conicyt PSD-26 and FONDECYT N°1080372.

Arsenic and Cadmium Uptake by Field-Grown Corn (Zea Maiz L.): Modeling and Characterization

M. Molina1*. M. Escudey2, W. Chen3 and A.C. Chang3

1Departamento de Ciencias Vegetales, Facultad de Agronomía e Ingeniería Forestal, Pontificia Universidad Católica de Chile, Santiago, Chile. 2Departamento de Química de ¡os Materiales, Facultad de Química y Biología, Universidad de Santiago de Chile, Santiago, Chile.3Department of Environmental Sciences, University of California at Riverside, CA 92521, USA. *E-mail:

Keywords: Arsenic; cadmium; corn.

Potentially hazardous trace elements may be inadvertently introduced into cropland soils through applications of fertilizers and other soil amendments. In agricultural systems, much of the trace element removal from the soil will depend on the ability of crops to uptake these elements and translocate them from roots to above-ground biomass. Through plant uptake,the elements accumulated in soils may be incorporated into the food chain to adversely affect consumers of the harvests. To characterize the risks, the trace element absorption process of plants must be quantified. We conducted a two seasons-long field experiment where grain corn was grown from seed to harvest. The data from soil and plant analyses was used to characterize the absorption of arsenic (As) and cadmium (Cd) in terms of plant uptake factors (PUF= element concentration in plant/element concentration in soil solution ratio), and of mean adsorption rates throughout the growing season. The cultivated soil was a clay loam Mollisol (Mixed, thermic, Ultic Haploxerolls), representing the soils used for intensive corn production in central Chile. This soil has been monocultivated with corn over a 15 year period, receiving high-dose application of P fertilizers. Soil samples were taken before sowing, and seven composite plant samples were taken in triplicate along each season. Soil and plant samples were analyzed to determinate the soil solution concentrations and total concentration of the elements, respectively. Total uptake of As and Cd by corn was 0.52±0.02 g ha-1 and 0.43±0.02 g ha-1, respectively. During the growing season the PUF decreased exponentially with time (R2>0.941), indicating a mass dilution effect for both elements. The PUF decrease by approximately 3.5 times for As and 3.0 times for Cd from the early growth stages to the time close to harvest. This behavior has been attributed to a greater competition between roots and/or to the presence of less active roots as the plant develops to maturity. The concentration of As in plants was slightly higher than the Cd concentrations. On the other hand, the As concentration in soil solution was higher than Cd concentration. The Cd PUF was more than 6 times higher than the As PUF, indicating a higher bioavailability of soil Cd for corn plants. The uptake rate at a specific sampling date (mean uptake rate) followed a sigmoidal pattern (R2>0.917). The maximum uptake rate were 3.9 x 10-3 g ha day-1 and 3.0 x 10-3 g ha day-1 for As and Cd, respectively. The maximum mean uptake rate was reached close to Rl stage (silking) for both trace elements and after that remained constant while the cumulative above-ground biomass was still increasing.

Acknowledgments: FONDECYT 1070116. Mauricio Molina would like to thank PUC-MECESUP (0210) Doctoral scholarship and PUC-VRAID international exchange.

Phytostabilization of Contaminated Soils in the Puchuncavi Valley, Central Chile

V. Muena, I. González and A. Neaman*

Facultad de Agronomía, Pontificia Universidad Católica de Valparaíso, Quillota, Chile

*E-mail: alexander. neaman@ucv. cl

Keywords: Oenothera affinis; phytostabilization; metal contaminated soil.

Mediterranean ecosystems of the Puchuncavi valley (central Chile) have been exposed to massive contamination with sulfur dioxide and metal-rich particles due to emissions from the Ventanas copper smelter. The soils in the areas near the smelter are now sparsely-vegetated, severely-eroded, acid, and metal-contaminated. Phytostabilization is defined as use of plants to stabilize contaminated soil and reduce the movement of toxic elements from the soil to the environment. The objective of the study was to determine the effectiveness of lime and fertilizers for establishment and growth of Oenothera affinis, a local species that is tolerant to metals. We carried out a two-year field experiment. The studied topsoils exhibited pH in the range of 4.0-4.5 (in salt extracts). Spatial distribution of total Cu concentrations was very heterogeneous in the studied topsoils, in the range of 310-750 mg/kg. The field plot was divided into 5 sub-plots of about 50 m2 each. Two controls and three treatments were used: (1) calcium hydroxide [Ca(OH)2], (2) nitrogen, phosphorus, and potassium fertilizers (NPK), and (3) Ca(OH)2 + NPK. Oenothera affinis was propagated and transplanted into experimental plots. At the end of the first life cycle (after 6 months of growth), plants at each sub-plot were harvested, and shoot biomass was determined. During the second year of the experiment, natural growth of Oenothera affinis seedlings was monitored. Shoot biomass production and natural seedlings establishment were considerably higher in the case of the Ca(OH)2 treatment in comparison to the control. In contrast, the NPK treatment did not produce any effect. The effect of Ca(OH)2 + NPK was similar to that of Ca(OH)2 alone. The data demonstrate a possibility of revegetation of contaminated soils of Puchuncavi valley by Oenothera affinis using lime, a low-cost agricultural amendment. Acknowledgment: This study is funded by the FONDECYT projects 1050403 and 1085005.


Effects of a Long-term Mercury Contamination on Soil Biological Properties as Assessed by Combined: Chemical, Biochemical, Ecotoxicological, and Molecular Methods

P. Ruggiero1*, R. Terzano1, A. Santoro1, M. Colombo2, C. Romagnosi2, V. Andreoni2, M. Rao3, P. Perucci4 and C. Casucci4

'Dip. di Biología e Chimica Agro-forestale ed Ambiéntale, Universitá di Bari, Italy. 2Dip. di Scienze e Tecnologie Alimentan e Microbiologiche, Universitá di Milano, Italy. 3Dip. di Sci. del Suolo, delta Pianta e dellAmb., Universitá di Napoli Federico II, Italy. 4Dip. di Scienze Ambientali e Produzioni Vegetali, Universitá Pol. delle Marche, Ancona, Italy. *E-Mail:

Keywords: Mercury contamination; heterotrophic bacteria; Glycirrhiza glabra.

Mercury (Hg) is a non-essential heavy metal that can pose serious risks to human beings and living organisms. However, its bioavailability and toxic effects are largely influenced by speciation. In the last years, combined chemical, biological and ecotoxicological studies on contaminated soils have increased importance to predict metal fate in the environment. The results obtained with these different approaches can be then used and compared to better categorize potential risk. In this research, different soil samples characterized by a long-term Hg-contamination were collected at two depths (0-10 and 40-50 cm) nearby an industrial area in the South of Italy ("Val Basento", Basilicata, Italy). These samples were studied for Hg total content, fractionation, phytotoxicity and influence on the bacterial community. Hg contamination ranged from 1 to 50 mg kg-1 and most of it was speciated in scarcely soluble forms (eg., HgS, Hg2Cl2, Hg), as assessed by sequential extractions and thermal desorption experiments. Hg in soil is known to have a deleterious effect on the number of bacteria, microbial biomass and activities, and diversity within bacterial populations. Anyway, in these soils, the biochemical quality indexes investigated (biomass, different enzyme activities) apparently were not influenced by the different degrees of Hg contamination. Differently, the addition of Hg in soluble form (HgCl2) showed a marked effect on these indexes. In addition, a quantitative approach based on enumeration of viable heterotrophic bacteria (VH) and mercury resistant bacteria (Hg ) was used to analyze the Microbe Interaction & Their Effects on Biophysical Transformation, Fate, and Toxicity of Metal and Metalloids structure of the bacterial communities of the soils and to follow population's dynamics to the addition of HgCl2. Again, the number of VH in soils as well as the functional diversity investigated by colony development on solid TSA diluted medium, were not related to the Hg content of soils. The investigated soils exhibited low percentage of Hg resistant bacteria ranging from less than 0.001% to 0.25 % and mercury addition induced an increase in the number of Hg bacteria only in some studied soils, suggesting that in a soil, bacteria are not exposed to the same level of the toxicant. Gram-positive and Gram-negative Hg resistant bacteria capable to grow in a minimal medium containing HgCl2 were also isolated and in some of them the presence of MerA and MerB genes was detected. Differently from bacteria, some influence of Hg concentration on seeds germination and root elongation was observed for Lepidium sativum L. and Lycopersicum esculentum L.. Probably, chelating agents exuded by plants can mobilize Hg thus increasing its bioavailability for root uptake. As a confirmation of this, spontaneous flora (Glycirrhiza glabra L.) collected in the investigated area showed non negligible amounts of Hg in the root system. In conclusion, the combination of the data obtained in this study showed that, despite a quite high Hg concentration, most of the Hg in these long-term contaminated soils was scarcely mobile and available and did not significantly influence the soil bacterial community. However, particular attention should be paid to Hg potential remobilisation that could be naturally favoured by the activity of plant roots.

Effect of Native and Exogenous Mycorrhizal Fungi on the Growth of Oenothera Affinis and Glomalin at Increasing Copper Levels

A. Seguel, S. Meier, F. Borie and P. Cornejo*

Departamento de Ciencias Químicas, Universidad de La Frontera, Casilla 54-D, Temuco, Chile. *E-mail: pcornejo@ufro. cl

Keywords: Mycorrhizal fungi; glomalin; copper

Pollution of natural ecosystems by mining activities is associated with the discharge of heavy metals (HM) enriched wastes. In these conditions of environmental stress, the arbuscular mycorrhizal fungi (AMF) may promote the establishment and growth of plants through the formation of arbuscular mycorrhizas. In addition, the glomalin, a glycoprotein produced by AMF and released to the soil in large amounts, can sequester and immobilize HM in the soil, improving the restoration of a plant cover through bioremediation technologies. The aim of this study was to analyze the effect of the mycorrhizal symbiosis and the type of AMF on the growth of Oenothera affinis and the accumulation of glomalin at increasing soil Cu levels. For this, seedlings of O. affinis were grown in: i) rhizospheric soil of O. Affinis with the native AMF inoculum (+M); ii) soil inoculated with Glomus claroideum (GC), and iii) sterile soil (-M). The growing substrate was sterile soil from a Mediterranean ecosystem near Ventanas smelter (Puchuncavi Valley, Valparaiso, Central Chile), which presented 178 mg kg-1 of available DTPA-Cu. This soil, was used to form seven increasing Cu levels, which consisted of 0, without soil (a mixture of sand and sepiolite); 1, soil diluted with sand and sepiolite (1:1:1); 2, soil undiluted; 3, soil with 75 mg Cu kg-1 (as CuS04 solution); 4, soil with 150 mg Cu kg-1; 5, soil with 225 mg Cu kg-1 and; 6, soil with 300 mg Cu kg-1. The glomalin was determined as glomalin-related soil protein (GRSP), obtained as total (T-GRSP) and easily extractable (EE-GRSP) fractions and quantified by Bradford assay. The results showed a strong decrease in shoot biomass production in all the AMF treatments used until level 3 (from 1.7-1.4 g pot-1 at level 0 to 0.5-0.6 g pot-1 in level 3). However, levels cupper n°4 showed an increase in shoot biomass, which was higher in the treatments +M, reaching up to 1.1 g pot-1 at level 6. Similar results were observed in root biomass production, but in -M was not observed an increase in root biomass at level 4 on higher (between 0.3-0.4 g pot-1). At the other hand, the glomalin production was higher in all the Cu levels in the GC treatments (amounts of 25-30 mg T-GRSP g-1 soil), compared to + M and -M (up to 25 mg g-1 soil). At 0 levels of-M and +M both GRSP fractions represent 30-50% of the GRSP observed in the other levels, but all levels exhibit a similar GRSP amount in GC. These results suggest a greater adaptation of native AMF from the Cu polluted soil to high exogenous levels of this element, contributing to promote the O. affinis growth even at extreme Cu concentrations in the soil. In addition, higher levels of GRSP in GC suggest that their release to the soil could be one of the more important mechanisms by which this unadapted fungus can tolerate high Cu concentrations. However, further studies are needed to explain the divergent behavior observed at moderate and extreme Cu concentrations, and what are the fungi into the natural communities that provide the greatest benefits for its further use in bioremediation programs.

Acknowledgment: This study was supported by Fondecyt grant 3070052. .


Determination of Carbon Sequestration and Nitrate Leaching Under Different Irrigation and Dairy Manure Levels in Wheat (Triticum aestivum L.) on Typic Calciagrids

K. Shahzad*

Institute of Soil and Environmental Sciences, Pakistan. *E-mail:

Keywords: Carbon sequestration; nitrate leaching; farmyard manure.

Nitrogen (N) losses from agriculture are negatively impacting groundwater, air, and surface water quality. New tools are needed to quickly asses these losses and provide nutrient managers and conservationists with effective tools to assess the effects of current and alternative management practices on N loss pathways. A field experiment was conducted to study the effect of different levels of irrigation and Farmyard manure on nitrate leaching in wheat (Triticum aestivum L). Recommended doze of NPK Fertilizers was applied. One third dose of N and full doses of P and K was applied at the time of sowing and the remaining dose of N will be applied at second and third irrigation. Farmyard manure was applied (0, 150 and 300 kg ha-1) in combination with different irrigation and nitrogen levels. A control of nitrogen and FYM was included for comparison. First treatment regarding Irrigation was applied after 30, 80 and 120 days. Second and third treatments of irrigation are applied after 30, 60, 90,125 and 21, 42, 60, 80, 95, 115 days simultaneously. Samples for nitrate analysis at depth of 0-5, 5-10, 10-30, 30-60, 60-90 cm was taken out at initial and final stages of experiment. Carbon sequestration is analyzed at the end of experiment. The data obtained will be analyzed statistically by using Fisher's analysis of variance technique and difference among treatment means will be compared by using least significant difference test (LSD) at 5% probability level.


Propericiazine as a Reagent for the Spectrophotometric Determination of Osmium

A. Thimme Gowda*

Department of Chemistry, A.V.K. College for Women, Hassan 573 201, Karnataka, India *E-mail: ankapuratgowda@rediffmail. com

Keywords: Propericiazine; osmium; synthetic ores.

Propericiazine (PPC) is proposed as a new sensitive and selective reagent for the spectrophotometric determination of microgram amounts of osmium. PPC forms an orange red coloured species with osmium (VIII) or osmium (VI) instantaneously at room temperature in 6M hydrochloric acid medium. The orange red species exhibits maximum absorbance at 511 nm. Beer's law is valid over the concentration range 0.1-8.0 ppm for osmium (VIII) and 0.2-10 ppm for osmium (VI). Sandell's sensitivity of the reaction is 9.4 ng cm-2 for osmium (VIII) and 13.5 ng cm-2 for osmium (VI). The effects of time, temperature, acidity, order of addition of reagents, reagent concentration and diverse ions are investigated. The application of the proposed method in the determination of osmium content in synthetic ores and minerals has been explored.


Competitive Heavy Metal Sorption by Different Soils

F.A. Vega*. E.F. Covelo and M.L. Andrade

Departamento de Biología Vegetal y Ciencia del Suelo. Facultad de Ciencias. Universidad de Vigo. Lagoas. Marcosende. Vigo 36310. Spain. Tel: +34986812630. *E-mail:

Keywords. Sorption, Metals, Soil.

Heavy metals in the environment are posing significant threats to human health and the environment. In recent years, increased anthropogenic inputs of heavy metals in terrestrial environments have caused considerable concern relative to their impact on groundwater contamination. Soils are an important sink for these metals due to soils high metal retention capacities. Heavy metals are consistently, and simultaneously, added to soils in increasing quantities in the form of fertilizers, pesticides, livestock manures, sewage sludge, and industrial emissions. Many of these metals show moderate mobility under relatively acid soil conditions (pH 5-7) because of increased solubility and formation of soluble complexes with organic ligands. Two soils from Galicia (Spain) were selected. S-l is developed in a serpentine-amphibolite area. Their pH is 6.9; the organic matter content is 52.59 mg kg-1 and the CECe is 25.93 cmol kg-1. The parent matter of S-2 is gneis, their pH is 5; the organic matter content is 67.59 mg kg-1 and the CECe is 2.13 cmol kg. Twelve solutions of mixtures of Cd, Cu and Pb nitrates (between 0.1 y 3 mmoles L-1) were prepared to obtain sorption isotherms. Six grams of soil samples were treated with 100 mL of solution, shaken for 24 hours at 25°C. After centrifugation, metal in solution was determined by ICP-OES. In this solution also they were analyzed the cations of the cationic exchange capacity (Al, Ca, K, Mg and Na). All of the experiments were performed in triplicate. The Cu and Pb sorption isotherms are curves tipe H in both soils. The obtained Cd isotherm in S-l shows an initial slope similar to the one of an H curve, but Cd sorbed amounts diminish in the most concentrated equilibrium solutions. The Cd sorption isotherm of S-2 shows an irregular curve and it is not similar to the ones established in the bibliography. In both soils Cd is the metal sorbed in least amounts, so that Cu and Pb compete favourably for sorption binding sites. S-1 sorbs highest heavy metals amounts than S-2. This fact is probably due to S-l has a pH higher than the one of S-2. In addition the CECe of S-l is higher than the one of S-2 and this factor also influences the heavy metal sorption capacity. Attending to the highest experimental sorbed amounts of each one of the studied metals, S-l sorbs higher Cu amounts than Pb whereas S-2 sorbs higher Pb amounts than Cu. The amounts of each one exchangeable cations released in the sorption experiments were studied and compared with the amounts of sorbed heavy metals. Mg is the exchangeable cation that is in highest amounts in the sorption equilibrium solution of S-l whereas Al is the exchangeable cation in highest proportions in the sorption equilibrium solution of S-2. Mg and Al are respectively the most aboundant exchangeable cations in the CECe of the soils S-l and S-2. So that, Pb and Cu are compete for sorption binding sites. The higher or lesser Cu and Pb sorbed amounts will depend mainly on soils pH and on their cationic exchange capacity.

Selection of Copper Resistant Bacteria in Copper-polluted Agricultural Soils from Central Chile

C. Yáftez*. M. González and M. Seeger

Laboratorio de Microbiología Molecular y Biotecnología Ambiental, Departamento de Química and Millennium Nucleus of Microbial Ecology and Environmental Microbiology and Biotechnology, Universidad Técnica Federico Santa María, Valparaíso, Chile. *E-mail:

Keywords: Copper Resistant Bacteria; cop A andpcoA genes; resistance mechanisms.

Copper mining is the most important economic activity in Chile. Significant increases in copper concentrations in soils have been reported in agricultural areas located in the vicinity of mining activities. Copper is an essential micronutrient to all organisms but it can be toxic to plants and soil organisms at elevated concentrations. Tight control of intracellular copper concentration is thus necessary. To meet this challenge, bacteria have developed copper homeostasis and resistance mechanisms. It has been generally assumed that soil bacteria are more sensitive to metal stress than other soil organisms. For that reason, they may represent a relevant indicator for copper pollution in agricultural soils. The aim of the present study was to assess the effects of high copper concentrations on soil bacterial communities and to study the bacterial tolerance to metal pollution. To accomplish this goal, we used culture-dependent (isolation of copper resistant strains) and culture-independent approaches (ribosomal RNA intergenic spacer analysis [RISA]; PCR detection of copper resistance genes) for the examination of the bacterial community structure. Composite soil samples were collected from five sampling sites, located at variable distances from copper smelters. Soils presented variable total copper concentrations, ranging from 130 to 580 mg/kg. Soils with higher total copper concentrations (from 470 to 580 mg/kg) were located near copper smelters. Analyses of RISA fingerprint patterns suggested a higher bacterial diversity in control soils (< 165 mg/kg) compared to soils with high copper concentration. Predominant RISA bands were observed in contaminated soils, probably related to bacterial groups selected by their metal resistance. Copper resistant bacteria were isolated from contaminated soils and presented a great metal tolerance (up to 275 mg/kg). Analysis by electron microscopy showed changes in the outer envelope and intracellular copper accumulation in some resistant strains. Detection of copA (copper transporting ATPase) and pcoA genes (multi-copper oxidase) in resistant strains suggests copper efflux and/or sequestration as resistance mechanisms. Our results suggest that soil bacterial communities respond to high copper concentration exposure by changes in microbial community structure and a selection of resistant strains. Future research will focus both on the evaluation of copper accumulation and the search of other genetic determinants involved in copper tolerance in resistant strains.

Acknowledgments: MILLENIUM P04/007-F and USM 130836 grants.


Competitive Adsorption of Copper and Lead on Volcanic Soil

E. Espinosa1*. M. Gutiérrez1, R. Calderón1, M. Cazanga1, C. Pizarro1, M. Escudey1, F. Garrido2 and M.T. García-González2

Universidad de Santiago de Chile, Facultad de Química y Biología. Av. L. Bernardo O'Higgins 3363, Estación Central, Casilla 40 - Correo 33, Santiago, Chile. Instituto de Ciencias Agrarias, Centro de Ciencias Medioambientales, Consejo Superior de Investigaciones Científicas, Serrano 115 dpdo, 28006 Madrid, España. *E-mail:

Keywords: Competitive adsorption; volcanic soil; heavy metals

Chilean soils are characterized by their heterogeneous components due to the various interactions of their parental materials; which is dependent on the climatic conditions along the country. Volcanic soils represent a 70 % of the total agronomic land. The heavy metals retention depends on the nature and relative abundance of mineralized and organic phases which according to the geochemical soil solutions; control the adsorption processes and also regulates the transport of contaminants to another environmental compartments. The metal ion characteristics as size, charge and hydrated ion size are another factors to be considered in the competitive interactions between them and with others solution species for specific position or adsorption site. Ever more, the different reactive phases may offer adsorption positions of varied bonding energy to the solution species. The competitive adsorptions processes are relevant because heavy metals presence as nature or contaminant elements can be found simultaneously in the soil systems. Bioavailability of heavy metals is the potential fractions constituted by a soluble and absorbed metal content by plant species. The metal Bioavailability depends on its solubility and the adsorption capacity the soil colloidal fractions. Unfertilized Ralun Soil (41° 32' S 73° 05' W), classified as Andisol, was studied the more relevant chemical characteristic are: high content of organic matter, high cationic interchange capacity and high water retention content. The soil was characterized using the standard methods. Monocomponent are bicomponent adsorption isotherms was realized using bath experiment were studied by scanning electron microscopy (SEM) and x-ray diffractions were used to monitor Cu, Pb adsorptions on Ralun Soil and Microprobe analysis and microprobe profiles were made with a Link Isis energy dispersive spectrometry (EDS) micro analytical system on the SEM. From the studies realized can be appreciated that the adsorptions increment follows the sequence: Pb > Cu. The isotherm fits Freundlich and Langmuir models.

Acknowledgments: The authors thank FONDECYT N° 1070116, DICYT-USACH 010741CS, USACH-CSIC, Scholarships VRID and CONICYT, CSIC-Madrid. .


Selenium-Induced Amelioration of the Oxidative Stress Triggered by Aluminium in Ryegrass Plants

P. Cartes1* , A.A. Jara2 and M.L. Mora2

1instituto de Agroindustria, 2Departamento de Ciencias Químicas. Universidad de la Frontera, Temuco, Chile. *E-mail:

Keywords: Aluminium; Ryegrass; Selenium.

Aluminium (Al) toxicity is one of the major factors that limit the production and quality of pastures in Andisols of Southern Chile. Although Al is a non-transition metal, it has been suggested that Al induce oxidative stress in plants (i.e. Ezaki et al., 2000; Yamamoto et al., 2001). On the other hand, the protective role of selenium (Se) against oxidative damage in higher plants (i.e. Hartikainen et al., 2005; Cartes et al., 2005; Mora et al, 2008), suggest an enhancement of the plant antioxidant properties due to Se additions under Al stress conditions. In order to evaluate the ability of Se to ameliorate the Al- induced oxidative stress in higher plants, ryegrass (Lolium perenne cv. Nui) was cultivated in a nutrient solution containing Al (0 or 200 µM) and Se (0 to 10 µM, as Na2Se03) at pH 4.8. Al toxicity negatively affected the yield and the antioxidant system of ryegrass, and this effect was more apparent in the roots than in the shoots tissues. Aluminium addition of 0.2 mM decreased the dry matter (DM) production in about 17%, whereas Se additions up to 0.2 µM significantly increased the yield of plants treated with 0 and 0.2 mM Al. Aluminium- treated plants accumulated more Al in the roots than in the shoots, and a raise of Al concentration in the plant tissues was observed by effect of Se additions. Furthermore, Se concentration in roots and shoots was steadily increased at increasing Se supply levels, and the highest concentrations were registered in Al-stressed plants. In general, the addition of 0.2 mM Al activated the antioxidant enzymes superoxide dismutase (SOD), peroxidase (POD) and ascorbate peroxidase (APX) in roots and shoots. When supplied at rates up to 2 M, Se ameliorated the oxidative stress by decreasing the lipid peroxidation (TBARS), and the most noticeable reduction of damage of cellular membranes occurred plants supplied with Se and Al. The Se-dependent decrease in TBARS accumulation was accompanied by 50% decrease of the root SOD activity in Al- treated plants. This fact suggests that Se may counteract Al-induced oxidative stress by increasing the spontaneous dismutation of the superoxide anions produced under Al stress conditions. Furthermore, Al-treated plants showed thresholds for the reduction of lipid peroxidation at Se concentrations higher than those of plants supplied with only Se. This fact denotes the highest requirement of Se of Al-stressed plants to counteract the oxidative stress induced by Al toxicity.

Acknowledgements: PSD 26 CONICYT-UFRO andFONDECYT 1061262


B. Ezaki, R. Gardner, Y. Ezaki and H. Matsumoto. (2000). Plant Phys. 122: 657-665.

P. Cartes, L. Gianfreda and M.L. Mora (2005).Plant Soil. 276: 359-367.

H.J. Hartikainen. (2005). Trace Elem. Med. Biol. 18: 309-318.

M.L. Mora, L. Pinilla, A. Rosas and P. Cartes. (2008). Plant and Soil. 303: 139-149.

Y. Yamamoto, Y. Kobayashi andH. Matsumoto. (2001). Plant Phys. 125: 199-208.


Short-term Aluminium Exposure Induced Rapid Changes in Proline, Carbohydrates and Aluminum Concentration in XJgni molinae and Vaccinium corymbosum

M. Reyes-Diaz1*, Y. Rojas-Lillo2, C. Inostroza-Blancheteau3, M. Alberdi4 and M.L. Mora4

1instituto de Agroindustria, Universidad de La Frontera, Casilla 54-D. Temuco. 2Programa de Magíster en Ciencias de Recursos Naturales, Universidad de La Frontera, Temuco, Chile.3Programa de Doctorado en Ciencias de Recursos Naturales, Universidad de La Frontera, Chile. 4Departamento de Ciencias Químicas, Universidad de La Frontera, Casilla 54-D, Temuco, Chile. *E-mail: reyesm(a),

Keywords: Vaccinium corymbosum; Ugni molinae; aluminium.

Aluminum (Al) toxicity is a major agronomic problem in acid soils. In southern Chile, soils derived from volcanic ash (Andisols) with an acid pH are frequent. Currently, the most preponderant fruticultural activity in this region is blueberry cultivation (Vaccinium corymbosum L). In this region also a native berry grows naturally (Ugni molinae L). This species seems to be adapted to the Andisol conditions, being of great interest to know its Al response mechanisms and to compare with those of cultivated species such as blueberry. Thus, our aim was to investigate at the short term the uptake of Al and its effect on proline and carbohydrate concentrations in both species. Brigitta, Legacy and Bluegold cultivars of V. corymbosum and four ecotypes of U. molinae (Toltén, T. Schmidt, Carahue and Curacautin) were grown in a greenhouse in hydroponic solutions containing different Al concentrations for 0-48 h and allowed to recover. Previous own results indicated that Toltén ecotype of U molinae was tolerant to Al toxicity, while Brigitta and Legacy cultivars were resistant and Bluegold cultivar was sensitive to this stress. The results showed that the Al accumulation was higher in roots than in leaves for both species, observing different patterns in Al uptake and transport to shoots. At the end of the Al treatment carbohydrate and proline concentrations in leaves decreased (19-48%) in Toltén and Carahue ecotypes of U. molinae, showing a posterior recovery. Contrarily, in leaves of cultivar Bluegold an increase of around 50% in carbohydrates and proline was found, maintaining similar values than the control by recovery. In U. molinae roots, carbohydrate concentrations practically did not varied during the treatments, whereas in Brigitta cultivar roots an increase (39%) was found at 24 h. No changes in proline concentrations of roots were found in cultivars and ecotypes, with exception of Bluegold cultivar and Carahue ecotype of U. molinae where an increase was observed. Statistically significant negative correlations between Al uptake and carbohydrates and proline were found in roots of Legacy cultivar and in leaves only with carbohydrates, whereas positive correlations between Al uptake and proline were found in roots of T. Schmidt and Curacautin ecotypes (P< 0.01). We concluded that the metabolites changes induced by Al did not correspond well with the degree of Al resistant of the investigated plants.

Acknowledgements: This study was supported by Bicentenario PSD-26 and Fondecyt 1080372.


Evaluation of Some Biological Activities in Four Landfill Municipal Soil from Southern Chile

M. Alvear1,2*.M. Arriagada2, P. Quezada2 and Y. Sandoval2

1Departamento de Ciencias Químicas, Universidad de La Frontera, Temuco, Chile;2 Asociación de Municipalidades del Valle Central de la Región de La Araucanía (AMVACEN). *E-mail:

Keywords: Microbial biomass, Fluorescein diacetate activity, Urease activity, Landfill soil

In landfill soils, where the pollution is usually going on for many years is very important know about soil quality status. Microbial biomass and some hydrolytic enzyme activities are important soil quality indicators. A study was conducted to determine the effect of municipal solid wastes (MSW) on four landfill soils from southern Chile considering four situation: with MSW freshly near and far the methane columns sites, also with two and four years without received MSW sites as well as four exotic forest soil near the each landfill as "control". Both Lautaros and Curacautins landfills received about 12.6 tons; Victoria's landfill received 6.1 tons and Perquenco's landfill received 2.5 tons annually. Overall microbial activity of soil was determined by fluorescein diacetate hydrolyzing activity (FDHA). Active cells carry out such hydrolysis by a combination of enzymes like lipases, proteases and esterase. Active microbial cells transport FDA inside the cell where it is hydrolyzed to polar fluorescein. When the storage capacity of the cells with respect to fluorescein exceeds, it is released out. The FDHA, urease and manganese peroxidase activities were measured spectrophotometrically. All analyses were made in triplicate for each field replicate. All analytical results were calculated on the basis of oven-dried (105°C) weigth. The results in all cases evaluated shown FDHA in landfill soils were significantly lower than four exotic forest soils due probably to lower organic matter and higher heavy metals content in landfill sites evaluated. Significant (P<0.05) variations existed in the metal concentrations among the landfill soils and were due to the heterogeneity in the waste materials at the respective sites over the years. Microbial biomass activity was lower and correlated with bioavailable metals. This indicated that it is the bioavailable form, which renders more toxicity compared to the total metals. The metals affected microbial biomass and their activities in combination with each other found that when metals were present in a mixture, they behaved synergistically or additively. The landfill soils also contained significantly amounts of total N and the urease activity was major in sites with fresh MSW. However, manganese peroxidase activity was higher in exotic forest soils than landfill soils. Our results are also in agreement with those reported by other investigators. In conclusion, present study, carried out on landfill sites revealed that the soils of these sites are contaminated with some heavy metals; more particularly the bioavailable forms detrimentally influenced the microbial biomass and enzyme activities in such soils.

Acknowledgments: The authors extend their gratitude to Municipalidades of Lautaro, Perquenco. Curacautin and Victoria for their cooperation in soil sampling.

Plant Growth Promoting Effect of Selected Autochthonous Microorganisms (bacteria, yeast and/or arbuscular mycorrhizal fungi) in a Heavy Metals Multicontaminated Soil

R. Azcón1*. M.C. Perálvarez1, A. Roldan2 and J.M. Barea1

1Departamento de Microbiología del Suelo y Sistemas Simbióticos, Estación Experimental del Zaidín (CSIC), Prof. Albareda 1, 18008 Granada, Spain. 2Centro de Edafología y Biología Aplicada del Segura (CSIC), Department of Soil and Water Conservation, P.O. Box 164, Campus de Espinardo, 30100Murcia, Spain. *E-mail:

Keywords: Heavy metals; autochthonous microorganisms; plant growth promoting.

As plant (clover) development was limited in a heavy metal multicontaminated soil the effect of selected autochthonous microorganisms as bacteria (B), yeast (Y) and arbuscular mycorrhizal (AM) fungi used either as single or dual inoculants, were assayed to ascertain whether these microbial treatment can benefit plant growth, nutrient uptake, and metal translocation . The inoculated microorganisms increased shoot biomass by 148% (B), 162% (Y) and 204% (AM), and root growth, particularly in dual associations, and decreased metals translocation from soil to shoots. Dual inoculations also produced the highest symbiotic (AM colonization and nodulation) rates. The plant growth effect and the metal sequestration or bioaccumulation abilities demonstrated by these microorganisms seem to be involved in such effects. The activity of microbial antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), glutattion reductase (GR) and ascorbate peroxidase (APX) known to play important roles in cell protection again abiotic stress, changed slightly in the culture medium of the bacteria or yeast in presence of metals. Such activities alleviate cellular oxidative damage, which is an important protective mechanism against heavy metal stress. These results are an indication of adaptation to high metal concentration of these autochthonous microorganisms. Thus, they may be considered as an index of microbial metal tolerance. Microbial inoculations showed a bioremediation effect since reduced shoot metal concentration and increased nutrient in plants allowing growth developments and helping plant recovery of the multicontaminated soil. Therefore, the tested mycorrhizosphere interaction could be used as an environmental biotechnology tool for improving plant developments in heavy metal-contaminated environments.


Exudation of Organic Acids in Ryegrass and White Clover in Response to Different Phosphorous and Manganese Doses

A. Rosas1, L. Pinilla2*, D. Menezes-Blackburn2 and M.L. Mora2

1Departamento de Suelos y Recursos Naturales, Facultad de Agronomía, Universidad de Concepción, P. O. Box 537, Chillan, Chile.2 Programa de Doctorado en Ciencias de Recursos Naturales, Universidad de La Frontera, Temuco, Chile. * Contact of authors: *E-mail:

Keywords: Organic Acids; ryegrass and white clover; manganese.

Soil chemical factors that limit root growth in acid soils reduce crop production, including Aluminium (Al) and manganese (Mn) toxicity, and also phosphorus (P) deficiency. Large areas of these grasslands are established in Southern Chile on acidic Andisols with both ryegrass (Lolium perenne L). The incidence of Mn toxicity depends of ambient factors, plant species, and accompanying fertilizer nutrients. Plants organic acid exudation is related to Al detoxification and P acquisition in the rhizosphere soil. This study investigated the effect of 5 levels of P (50 to 400 µM) on perennial ryegrass tolerance to excess Mn in 4 levels (50 to 355 µM). Ryegrass (cv. Nui) plants were cultivated for 30 days in nutrient solutions at pH 4.8, in a growth chamber with a light period of 16 h, temperature of 20°C. The plant response was evaluated through growth, and root exudation of 4 organic acid anions (citrate, malate, succinate and oxalate). Toxicity due to excess Mn supply in interaction with P deficiency resulted in significant reduction in plant dry weight. At highest rate of Mn supply (355 µM), the ryegrass plants showed a reduction of approximately 50 % in DW compared with treatments 2.4 µM of Mn. The decrease of the P supply from 96 to 44 µM for ryegrass caused a significant reduction in DW due to P deficiency. Addition of P had a beneficial effect in reducing the severity of Mn toxicity. Ryegrass plants exuded the highest quantity of organic acids, especially oxalic and citric acid, in conditions of deficiency of P and Mn toxicity, thus enhancing tolerance to Mn toxicity in this forage specie. Oxalic and malic acid exudation rates were 7 fold higher in this condition than in P and Mn sufficient plants. It was observed an increase in exudation of oxalate (more than 80%), malate (89 %), citrate (27 %) and succinate (33%) in comparison with the plant supplied with adequate P and Mn.

Acknowledgements: This work was supported by the International Cooperation FONDECYT project 7060093 and by the FONDECYT project 1061262 Grants.


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