Physiological Effects of Heavy Metals on the Moss Rhytidiadelphus squarrosus

Experiments on the effect of Cd, Cu, Ni, Pb and Zn, suppliedas single salt solutions between 10 µM and 0·1M, on the moss Rhytidiadelphus squarrosus showed little alterationto respiratory rates but reduced photosynthetic rates, and somemembrane damage as assessed by K leakage. Two distinct morphologicalforms of the moss showed different responses to supplied heavymetal. Storage of material, after 30 mins exposure to heavymetals, resulted in a further decrease in the photosyntheticrate. Expressing the photosynthetic decline relative to thetotal heavy metal recovered from the moss showed a similar pattern.Transfer of metal from extracellular exchange sites into theprotoplast was also demonstrated with storage after exposure.An approximately linear relationship was demonstrated betweenphotosynthetic decline and intracellular heavy-metal concentration,irrespective of the duration of exposure or morphological natureof the material used. Photosynthetic decline on storage is concludedto be a response to additional metal stress rather than a progressivedeterioration of the physiological process. Cadmium, copper, lead, nickel, zinc, photosynthesis, respiration, membrane damage, intracellular metals, Rhytidiadelphus squarrosus (Hedw.) Warnst     

Effects of copper and mineral nutrition on growth,copper accumulation and mineral element uptake in two Rumex japonicus populations from a copper mine and an uncontaminated field sites

Two Rumex japonicus populations, one from a copper mine and the other from an uncontaminated site, were studied in hydroponic experiments for the plant growth, copper accumulation and mineral nutrient content under excess copper and nutrient deficiency conditions. The tolerance indices of the contaminated population were significantly higher than that of the uncontaminated population, indicating the evolution of Cu resistance in the former. At control and low Cu treatment, there was no difference in Cu accumulation in roots between the two populations. At high Cu (100 μM) treatment, however, the contaminated population accumulated less Cu in roots than the uncontaminated one, suggesting the root exclusion mechanism existing in the former. The contaminated population was also more tolerant to general nutrient deficiency than the uncontaminated one. The results indicated that the contaminated population had evolved not only Cu-tolerance but also tolerance to low nutrient supply. Under Cu stress, the contaminated population had less change in nutrient composition than the uncontaminated one. The similar result was observed in general nutrient deficiency experiment. The results indicated that the mineral composition homeostasis under the stresses was important in metal tolerance and colonizing the Cu-enriched soils for the Cu-tolerant population. At high Cu (100 μM) treatment and general nutrient deficiency treatment, the contaminated population accumulated significantly lower copper and higher phosphorus in both roots and shoots than the uncontaminated one. This was not the case for other mineral nutrients Ca, Mg and Fe except for root Mg and root Fe at Cu treatment. The result suggested that the high Ca-metabolism in R. japonicus was uncorrelated with high Cu-tolerance and that P might play an important role in governing Cu bioaccumulation.     

Accumulation of heavy metals by aquatic mosses. 2: Rhynchostegium riparioides

A study was made of general ecology and metal accumulation in the widespread aquatic moss Rhynchostegium riparioides, (Hedw.) C. Jens. with a view to developing the use of this species as a monitor of heavy metal pollution. In order to establish a data bank for statistical analysis, samples of water and moss were taken within a 6-week period from 105 sites (10-m reaches) in Northern England from streams and rivers of diverse physical and chemical types. Analyses were made of 14 metals (Na, Mg, K, Ca, Cr, Mn, Fe, Co, Ni, Cu, Zn, Cd, Ba, Pb) in both 2-cm tips and whole plants. The same 14 metals were also measured in both total and filtrable water, together with 12 other variables. Samples of tips were easier to prepare for analysis, but had significantly (p < 0.001) lower concentrations of all metals except Na and K. Significant correlations (p < 0.001) between metal in moss and aqueous metal were found for ten metals (Na, K, Mg, Ca, Mn, Cu, Zn, Cd, Ba, Pb). Correlations between metals in moss and in water were in general similar for tips and whole plants, but much higher for tips with Na, Zn and Cd; the relationship was quite similar whether total or filtrable water was considered, with the exception of Ba where the correlation was much higher with the latter. A multiple regression was used to suggest which variables in water and/ or moss may influence accumulation of Co, Ni, Cu, Zn, Cd, Ba and Pb in the moss. For instance, the variables which had a very highly significant effect on Pb in tips were Pb, filtrable reactive phosphate and Zn in the water. A discussion is included of how the data may be used for monitoring purposes.     

Differential gene expression profiles associated with heavy metal tolerance in the soil insect Orchesella cincta

Field-selected tolerance to heavy metals has been reported for Orchesella cincta (Arthropoda: Collembola, springtails) populations occurring at metal-contaminated mining sites. Metal tolerance in O. cincta is correlated with heritable increase of excretion efficiency, decrease in cadmium-induced growth reduction, and over-expression of the single copy metallothionein (mt) gene. Although mt plays a major role in cadmium (Cd) detoxification an additional number of gene products may be altered in the tolerant phenotype that influence quantitative traits like excretion efficiency and growth. Using suppression subtractive hybridization analysis (SSH) genes were cloned that showed up- or down-regulation in response to Cd. This study reports about 192 SSH derived cDNA clones, of which 49 were confirmed to be differentially expressed after spot blot analysis. After sequence analysis 19 of the 49 differential clones could be annotated. Real-time RT-PCR of a reference culture and tolerant populations revealed significant gene responses upon Cd exposure. The reference culture showed stress induced gene expression, whereas the tolerant animals maintained normal gene expression or down regulation of cDNAs upon Cd exposure. Seven cDNAs with homology to genes involved in cell signaling, transport and programmed cell death showed an interaction between treatment and population.     

Stress responses and metal exchange kinetics following transplant of the aquatic moss Fontinalis antipyretica

1. We studied stress responses and metal exchange (uptake and loss) kinetics in the aquatic moss Fontinalis antipyretica Hedw. following transplant (in plastic mesh bags) to clean and metal-contaminated river sites and under laboratory conditions. 2. The stress response (estimated on the basis of chlorophyll:phaeophytin ratio) was more pronounced, and the moss took longer to recover, following transplant to heavily contaminated sites. 3. Metal uptake over the 28 day exposure period was predicted well by a two-compartment kinetic model: uptake velocity was initially high and gradually declined, with metal concentration in the moss showing a tendency to reach an equilibrium with metal concentration in the water. Mean uptake rate, time to reach equilibrium and metal concentration in moss at equilibrium all increased with increasing metal concentration in the water. 4. Bioconcentration constants calculated on the basis of our data rank the metals studied in the order Zn < Ni < Co < Cu < Pb < Cd. 5. Following retransplant of mosses to a clean site, metal loss was not predicted well by a passive exchange mode of the above type; instead, two phases (rapid and slow) were apparent. Loss rates during both phases, and proportion of metal load lost during the rapid phase, were proportional to the concentration of metal in the moss at the start of the recuperation trial. 6. We present nomograms to allow estimation of severity of contamination on the basis of metal concentration in moss and duration of transplant.     

Biochemical responses of the aquatic moss Fontinalis antipyretica to Cd,Cu, Pb and Zn determined by chlorophyll fluorescence and protein levels

Biochemical reactions to Cu, Cd, Zn and Pb in the aquatic moss Fontinalis antipyretica were studied in order to characterize the physiological background of the metal response. Chlorophyll fluorescence and intracellular metal localization and stress protein levels were measured. Exposure to 25 or 100 μM Cu over a 7-day period resulted in a decline of chlorophyll fluorescence to about 70% and 52%, respectively. Up to 100 μM Cd caused a decrease in chlorophyll fluorescence to 75%. With all metals used at 25–100 μM concentrations, the intracellular uptake increased. For all metals investigated at 25–100 μM, the intracellular uptake increased. Maximum values were reached at 100 μM Cu, Pb, Zn or Cd exposure. As shown by analytical electron microscopy (EDX, EELS) moss material treated with 50 μM Cu exhibited reduced sulphur levels in the cytoplasm and an increase in phosphate in vacuolar dense particles. EEL-spectra indicated that Cu is chelated in the cytoplasm by SH-groups and coprecipitated with orthophosphate in vacuoles. To monitor the stress response at the protein level, heavy metal induced heat shock protein 70 (hsp70) was measured. An antibody was raised against conserved plant metallothionein p2 motifs derived from Brassica juncea. In all metal-treated samples the antibody bound to proteins of about 8 kDa. However, sequencing failed to reveal significant homologies to known proteins. These experiments provide for the first time results on protein level after heavy metal stress in the aquatic bioindicator moss.     

Interactions between Mitochondria and Chloroplasts in Cells: I. Action of Cyanide and of 3-(3,4-Dichlorophenyl)-1,1-dimethylurea on the Spore of Funaria hygrometrica

The effects of cyanide and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) on photosynthesis and respiration of intact chlorophyllic moss (Funaria hygrometrica) spore was investigated. Thirty micromolar cyanide strongly inhibited dark respiration, was without effect on photosynthesis at high light intensities (above the saturation plateau values), and stimulated photosynthesis at low light intensities (below the saturation plateau values). Three hundred nanomolar DCMU inhibited the photosynthesis and was without effect, even under light conditions, on the dark respiration. It seems likely, therefore, that in the chlorophyllic moss spore the cytochrome oxidase pathway is not functioning under high light intensities unless the photosynthesis is inhibited by DCMU.     

Comparison of different microbial biomass and activity measurement methods in metal-contaminated soils

The aims of this study were: (1) to compare different microbial methods of detecting the effects of heavy metals on the functioning of the soil ecosystem; and (2) to evaluate the effect of incubation on microbial biomass and microbial activity in soils that were not pre-incubated after sampling in order to determine their suitability for measuring the effects of heavy metals on the soil microbial ecosystem. The microbial biomass methods (included: biomass C, N and ninhydrin-N by fumigation-extraction (FE); substrate-induced respiration (SIR); soil ATP content and microbial activity as evolved CO2-C and arginine ammonification. All were tested in soils from the Woburn Market Garden Experiment. Due to past sludge application the soils contained, Zn, Cu or Ni at around current European Union upper limits and Cd at up to three times the limit. The amount of microbial biomass in metal-contaminated soils was about half of that found in soils from the experiment that received uncontaminated organic manure or inorganic fertilizer. The amount of biomass measured by FE and soil ATP content in incubated soils showed little change over 20 days incubation. However, SIR measurements were statistically affected over the first few days of incubation. The rates of arginine ammonification were higher in this order: farmyard manure (FYM)>inorganic fertilizer>sewage-sludge throughout the incubation. However, the evolved CO2-C rates were not significantly different among the treatments. Discriminant analysis confirmed smaller amounts of biomass in the metal-contaminated soils than in the other treatments. Linked properties, such as relationships between biomass and soil organic matter, or biomass-specific respiration rates, may provide "internal control" which may help overcome problems of establishing suitable control, or comparative measurements, when moving from experimental to natural environments.     

Adaptation to metal-contaminated soils in populations of the moss,Ceratodon purpureus: vegetative growth and reproductive expression

Many observations suggest that morphological evolution occurs slowly in bryophytes, and this has been suggested to reflect low genetic diversity within species. Isozyme studies, however, stand in apparent contrast and have shown that bryophytes can contain high levels of genetic variability within and among populations. In light of this conflict, we tested the potential of the moss, Ceratodon purpureus, to undergo adaptive change (i.e., ecotypic differentiation) in response to soils that have been contaminated with high levels of metals for 90 years by measuring gametophytic growth and reproductive expression under experimental conditions. Variation in protonemal growth in sterile culture indicates that plants from one population growing on contaminated soil near a smelter are significantly more tolerant of zinc, cadmium, and lead than plants from uncontaminated sites. Results from a common garden experiment, in which plants were grown on soil from the smelter site, indicate that plants from near the smelter are significantly more tolerant of contaminated soils than plants from uncontaminated sites for vegetative growth. The same experiment suggests that plants from the smelter site are also more tolerant in terms of gametangial production (although we could not test this statistically). Our results demonstrate that C. purpureus has been able to undergo relatively rapid evolution in response to strong selective pressures.     

Heavy Metal Resistance and Accumulation Characteristics in Willows

The resistance of Salix to Cu, Cd, Ni, and Zn was investigated in hydroponic culture, with phytoextraction potential evaluated for Cu. Root elongation (indicative of resistance level) was significantly affected, with considerable variation between and within individual clones. Resistance appeared to be clone- or hybrid-specific, rather than species-specific. S. caprea clones (and hybrids) were among the most resistant, but a secondary S. caprea clone from a different provenance was much less tolerant. S. viminalis and S. triandra clones were the most sensitive. Highest resistance was found in response to Cd, while Cu and Ni were extremely toxic. A resistant S. caprea ecotype originating from a metalliferous mine spoil was identified using this technique. Copper concentration reached a maximum of 2000, 400, and 82 μg g^-1 (d.wt) in roots, wood, and foliage, respectively, after 1 month in hydroponic culture. The level of variation in the response of Salix to metals may cause difficulties in phytoremediation screening programs, but may be essential in providing genetic variation for selection of metal resistance traits, where the contaminant profile is heterogeneous, mixed, or subject to change. Clone selection for metal phytoextraction is feasible, but a longer field-scale study on metal-contaminated soils is needed before their role in phytoremediation can be confirmed.     

A multielement analysis of Cu induced changes in the mineral profiles of Cu sensitive and tolerant populations of Silene paradoxa L.

This work investigates the Cu induced changes in element profiles in contrasting ecotypes of Silene paradoxa L. A metallicolous copper tolerant population and a non-metallicolous sensitive population were grown in hydroponics and exposed to different CuSO₄ treatments. Shoot and root concentrations of Ca, Cu, Fe, K, Mg, Mn, Mo, Na, P, S and Zn were evaluated through ICP-OES.Results indicated that increasing the environmental Cu concentration had a population dependent effect on element profiles, shoot-to-root ratios and correlations among the elements. Generally, in the tolerant population Cu treatment induced a higher element accumulation in roots and had minimal effects on the shoot element profile, thus resulting in a progressively decreasing shoot-to-root ratio for each element. In the sensitive population element concentrations in root and shoot were much more affected and without a consistent trend. Copper treatment also affected the correlations between the elements, both in roots and shoots of the two populations, but more so in the sensitive population than in the tolerant one. Thus, Cu exposure strongly disturbed element homeostasis in the sensitive population, but barely or not in the tolerant one, probably mainly due to a higher capacity to maintain proper root functioning under Cu exposure in the latter. Differences in element profiles were also observed in the absence of toxic Cu exposure. These differences may reflect divergent population-specific adaptations to differential nutrient availability levels prevailing in the populations’ natural environments. There is no evidence of inherent side-effects of the Cu tolerance mechanism operating in the tolerant population.     

Accumulation, subcellular localization and ecophysiological responses to copper stress in two Daucus carota L. populations

Copper accumulation, subcellular localization and ecophysiological responses to excess copper were investigated using pot culture experiments with two Daucus carota L. populations, from a copper mine and an uncontaminated field site, respectively. Significant differences of malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) concentrations and antioxidant enzyme [superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX)] activities of leaves under Cu treatment were observed between the two populations. At high Cu concentrations (400 and 800 mg kg⁻¹), a significant increase in contents of MDA and H₂O₂ but a significant decrease in activities of SOD, CAT and APX were observed in uncontaminated population. Contrarily, the population from copper mine maintained a lower level of MDA and H₂O₂ but higher activities of SOD, CAT and APX. Copper accumulation in roots and shoots increased significantly with the increase of copper concentrations in soils in the two populations. No significant difference of the total Cu in roots and shoots was found between the two populations at same copper treatment. There were also no striking differences of cell wall-bound Cu and protoplasts Cu of leaves between the two populations. The difference was that Cu concentration in vacuoles of leaves was 1.5-fold higher in contaminated site (CS) population than in uncontaminated site population. Hence, more efficient vacuolar sequestration for Cu and maintaining high activities of SOD, CAT and APX in the CS population played an important role in maintaining high Cu tolerance.     

Methodological aspects of moss sample preparation. Effects of freezing and duration of washing on the cellular distribution of elements in Fontinalis squamosa Hedw.

Aquatic bryophytes are widely used in biomonitoring studies, mainly because of their capacity to act as bioaccumulators. As different methods can be used to preserve and process moss samples, the results of elemental analyses may also vary, particularly if the contents of different cellular compartments are analyzed separately. In the present study, we compared the total concentrations of some elements and the concentrations of these in different cellular locations in frozen-thawed and fresh samples of the aquatic moss Fontinalis squamosa that were also washed for different lengths of time before analysis. We used the sequential elution technique (SET) to extract the different fractions, and we determined the concentrations of K, Mg, Ca, Cd, Co, Cu, Al and Zn in the extracts. The results varied depending on the element, cellular location and moss sampling site. In the moss samples processed after freezing, the greatest differences were in the intracellular concentrations of K, Mg and Cd, which were lower than in the fresh samples. Minor differences were found in the concentrations of elements in other cellular locations and in the total concentrations of elements. The increase in the duration of the initial washing step, carried out to remove soluble and particulate intercellular elements, may also cause the release of elements (e.g. K and Mg) bound to extracellular exchange sites. The concentrations in the other cellular fractions and the total concentrations were less affected by the washing duration. Neither freezing nor the use of long washing times are recommended for processing moss samples prior to extraction of elements by the SET. Further studies are needed to confirm and clarify the observations.     

Population density-dependent metal tolerance: One possible basis and its ecological implications

A population density-dependent copper (Cu) resistance mechanism in a gram-negative soil bacterium, strain TDCd1, was shown to be inducible and was accompanied by changes in the protein composition of the outer membrane of the cell envelope. Characteristically, following inoculation of TDCd1 into Cu-supplemented growth media, there was a period of growth inhibition during which the number of individuals tolerant to Cu gradually increased, even though microcultural experiments indicated that some cells died during this period. We concluded that the population density dependency of the resistance mechanism resulted from the interactions between the rate of cell death, the time taken for Cu resistance to develop, and the size of the initial population. Therefore, the ability of relatively large populations of microorganisms to grow in metal-supplemented media under laboratory conditions may have little ecological significance for sparse populations in natural environments.     

Patterns of rapd markers and heavy metal concentrations in Perna viridis (L.), collected from metal-contaminated and uncontaminated coastal waters: are they correlated with each other?

Genetic variation due to heavy metal contamination has always been an interesting topic of study. Because of the numerous contaminants being found in coastal and intertidal waters, there is always much discussion and argument as to which contaminant(s) caused the variations in the genetic structures of biomonitors. This study used a Single Primer Amplification Reaction (SPAR) technique namely Random Amplified Polymorphic DNA (RAPD) to determine the genetic diversity of the populations of the green-lipped mussel Perna viridis collected from a metal-contaminated site at Kg. Pasir Puteh and those from four relatively' uncontaminated sites (reference sites). Heavy metal levels (Cd, Cu, Pb and Zn) were also measured in the soft tissues and byssus of the mussels from all the sites. Cluster analyses employing UPGMA done based on the RAPD makers grouped the populations into two major clusters; the Bagan Tiang, Pantai Lido, Pontian and Kg. Pasir Puteh populations were in one cluster, while the Sg. Belungkor population clustered by itself. This indicated that the genetic diversity based on bands resulting from the use of all four RAPD primers on P. viridis did not indicate its potential use as a biomarker of heavy metal pollution in coastal waters. However, based on a correlation analysis between a particular metal and a band resulting from a specific RAPD primer revealed some significant (P < 0.01) correlations between the primers and the heavy metal concentrations in the byssus and soft tissues. Thus, the correlation between a particular metal and the bands resulting from the use of a specific RAPD primer on P. viridis could be used as biomonitoring tool of heavy metal pollution.     

Differential Sensitivity of Lichens to Heavy Metals

Zinc, Cd and Cu inhibited photosynthesis in lichens containingcyanobacterial phycobionts at substantially lower concentrationsthan those causing decreased photosynthesis in lichens containingchlorophycean phycobionts. This distinction was not relatedto differences in total thallus concentrations of Zn, Mg, Caor K or to the quantity of Zn taken up to intracellular sites.When incubated with concentrated Zn solutions the chlorophyceanlichen Cladonia rangiformis accumulated more Zn on extracellularexchangeable sites than did the cyanobacterial lichen Peltigerahorizontalis. algae, cyanobacteria, chlorophyceae, lichens, heavy metal, photosynthetic inhibition, zinc, cadmium, copper, cation uptake     

Carbon dioxide exchange of two ecodemes of Schistidium antarctici in Continental Antarctica

Summary Schistidium antarctici is the commonest of five bryophytes known in the Windmill Islands area of Wilkes Land, Greater Antarctica. In moist habitats it forms closed carpets, but in dry sites it develops a short cushion growth form. Carbon dioxide exchange of both a mesic (Sm) and a xeric growth form (Sx) was investigated by means of an IRGA system in the field near Casey Station under natural light and simulated ambient or controlled temperature conditions in the plant chamber. The chlorophyll content in Sm was three times higher than in Sx. The light compensation point of Sm was lower than in Sx. The data for photosynthesis and dark respiration were computed by means of non-linear and linear regression analysis. Sm was more productive and had a wider temperature range of positive net photosynthesis than Sx under similar conditions. Dark respiration per gram of the whole moss sample was identical in both ecodemes. A decline of the photosynthesis curves at quantum flux densities above 500 mol m^-2 s^-1 PAR indicated a photoinhibitory effect in Sm. Sx was even more sensitive to high irradiance levels. Photoinhibition was not apparent in laboratory measurements under artificial light. According to our field measurements the photoinhibitory effect increases with increasing temperature. Moisture loss was avoided during the experiments by water supply from the bottom and frequently spraying the moss samples with water. In the natural habitat the desiccating effect of solar radiation is important, as it quickly causes photosynthesis to cease. The moss will dry out sooner in a xeric habitat than in one which is continuously moist. Consequently, the mesic Schistidium might particularly be subjected to photoinhibition by bright sunshine.Cordially dedicated to Professor Dr. O.H. Volk, Würzburg, on the event of his 85th birthday     

Phytoextraction of cadmium with Thlaspi caerulescens

The in situ phytoextraction of cadmium from soils can only be achieved using plants that are both tolerant to high Cd concentrations and able to extract sufficient amounts of the metal. However, very few plant species are capable of remediating Cd polluted soils in a reasonable time frame. This paper aims to show that the population of the hyperaccumulator Thlaspi caerulescens J. & C. Presl. from Viviez (south of France), which has a high Cd-accumulating capability, is an efficient tool to remove Cd from contaminated soils. Roots of T. caerulescensViviez proliferate in hot spots of metals in soils which is particularly advantageous because of heterogeneity of the distribution of metal in polluted soils. Isotopic techniques showed that plants from this population acquire Cd from the same pools as non-accumulating species, but that it was much more efficient than non-hyperaccumulators at removing the metal from the soil labile pool. This is due: to (i) a specific rooting strategy, and (ii) a high uptake rate resulting from the existence in this population of Cd-specific transport channels or carriers in the root membrane. Growth and overall extraction can be improved with appropriate N fertilisation, supplied either as mineral fertilisers or uncontaminated sewage sludge. Selecting bigger plants is possible from within a suitable Cd-accumulating population to improve the phytoextraction process. Growing the Cd-accumulating populations results in a reduction in the availability of Cd and Zn as shown with field and lysimeter experiments conducted for several years. As a result, on a practical aspect, Cd hyperaccumulating populations of T. caerulescens may be used as a tool to efficiently reduce the availability of Cd in soils, providing appropriate populations are used.     

Cadmium and copper uptake and accumulation by Sesbania rostrata seedling, a N-fixing annual plant: implications for the mechanism of heavy metal tolerance

Sesbania rostrata,an annual tropical legume,has been found to be tolerant to heavy metals,with an unknown mechanism.It is a promising candidate species for revegetation at mine tailings.In this study,sequential extractions with five buffers and strong acids were used to extract various chemical forms of cadmium and copper in S.rostrata,with or without Cd or Cu treatments,so that the mechanisms of tolerance and detoxification could be inferred.Both metals had low transition rates from roots to the aboveground of S.rostrata.The transition ratio of Cd (4.00%) was higher than that of Cu (1.46%).The proportion of NaCl extracted Cd (mostly in proteinbinding forms) increased drastically in Cd treated plants from being undetectable in untreated plants.This suggests that Cd induced biochemical processes producing proteinlike phytochelatins that served as a major mechanism for the high Cd tolerance of S.rostrata.The case for Cu was quite different,indicating that the mechanism for metal tolerance in S.rostrata is metal-specific.The proportion of water-insoluble Cu (e.g.oxalate and phosphate) in roots increased significantly with Cu treatment,which partially explains the tolerance of S.rostrata to Cu.However,how S.rostrata copes with the high biotic activity of inorganic salts of Cu,which increased in all parts of the plant under Cu stress,is a question for future studies.Sesbania rostrata is among the very few N-fixing plants tolerant to heavy metals.This study provides evidence for the detoxification mechanism of metals in Sesbania rostrata.