Effects of Cadmium on Root Growth, Cell Division and Nucleoli in Root Tip Cells of Garlic

The effects of different concentrations (10⁻⁷ to 10⁻² M) of cadmium chloride on root growth, cell division and nucleoli in root tip cells of Allium sativum L. were investigated. At lower concentrations of Cd²⁺ (10⁻⁷ to 10⁻⁶ M), Cd²⁺ did not influence the root growth, even had a stimulation effects during a short treatment. The results showed that the rate of root growth per day at the treatment groups (10⁻⁴ to 10⁻² M Cd²⁺) decreased with increasing duration of the treatment and increasing Cd²⁺ concentration. Cd²⁺ induced c-mitosis, anaphase bridges, chromosome stickiness and on nucleoli, causing some particles of similar silver-stained material scattered in the nuclei and making the silver staining reaction at the periphery of the nucleolus weaker. This revised version was published online in July 2006 with corrections to the Cover Date.     

Contribution of <Emphasis Type="Italic">Glomus intraradices</Emphasis> inoculation to nutrient acquisition and mitigation of ionic imbalance in NaCl-stressed <Emphasis Type="Italic">Trigonella foenum-graecum</Emphasis>

The study aimed to investigate the effects of an AM fungus (Glomus intraradices Schenck and Smith) on mineral acquisition in fenugreek (Trigonella foenum-graecum) plants under different levels of salinity. Mycorrhizal (M) and non-mycorrhizal (NM) fenugreek plants were subjected to four levels of NaCl salinity (0, 50, 100, and 200 mM NaCl). Plant tissues were analyzed for different mineral nutrients. Leaf senescence (chlorophyll concentration and membrane permeability) and lipid peroxidation were also assessed. Under salt stress, M plants showed better growth, lower leaf senescence, and decreased lipid peroxidation as compared to NM plants. Salt stress adversely affected root nodulation and uptake of NPK. This effect was attenuated in mycorrhizal plants. Presence of the AM fungus prevented excess uptake of Na⁺ with increase in NaCl in the soil. It also imparted a regulatory effect on the translocation of Na⁺ ions to shoots thereby maintaining lower Na⁺ shoot:root ratios as compared to NM plants. Mycorrhizal colonization helped the host plant to overcome Na⁺-induced Ca²⁺ and K⁺ deficiencies. M plants maintained favorable K⁺:Na⁺, Ca²⁺:Na⁺, and Ca²⁺:Mg²⁺ ratios in their tissues. Concentrations of Cu, Fe, and Zn²⁺ decreased with increase in intensity of salinity stress. However, at each NaCl level, M plants had higher concentration of Cu, Fe, Mn²⁺, and Zn²⁺ as compared to NM plants. M plants showed reduced electrolyte leakage in leaves as compared to NM plants. The study suggests that AM fungi contribute to alleviation of salt stress by mitigation of NaCl-induced ionic imbalance thus maintaining a favorable nutrient profile and integrity of the plasma membrane.     

Role of VA-mycorrhiza in growth and mineral nutrition of apple (Malus pumila var.domestica) rootstock cuttings

One-year-old apple cuttings (Malus pumila var.domestica cv. M26) were grown for 6 months in pot culture with and without inoculum of the VA-mycorrhizal fungus (VAMF)Glomus macrocarpum in soil from a long-term fertilizer field experiment with different P availability (20, 210, and 280 mg CAL-extractable P kg⁻¹). The indigenous VAMF propagule density was reduced by 0.5 Mrad X-irradiation. At harvest, non-inoculated and inoculated plants had similar proportions of root length bearing vesicles. Net dry weight of tree cuttings was significantly increased by inoculation only at 20 mg P kg⁻¹ (+62%). Increasing P availability from 210 to 280 mg P kg⁻¹ led to a 4-week depression of shoot elongation rate only in the inoculated plants. Uptake of P was significantly enhanced by inoculation at 20 and 210 mg P kg⁻¹ (+64 and +12%, respectively). On average, inoculated plants had significantly higher concentrations of Zn in leaves and in roots (+16 and +14%, respectively) and of copper in stems and in roots (+13 and +126%, respectively). Proportion of vesicle bearing root length was significantly correlated with root caloric content. A lipid content of 0.9–4.5% in the root dry matter was attributed to the presence of vesicles corresponding to 1.6–8.2% of total root caloric content. As the control plants were also infected, the beneficial effect of VA-mycorrhiza on nutrient uptake and growth of apple cuttings was underestimated at all P levels. Furthermore, VAM-potential at the lowest P level was not fully exploited as onset of infection was most certainly delayed because of a decreased photosynthetic rate due to P deficiency. Energy drain by VAMF-infection was most probably underestimated considerably, due to, among others, loss of infected root cortex during root growth, sampling and staining. It is concluded that apple cuttings rely on VA-mycorrhizal P-uptake at least in low P soils. In high P soils, apple cuttings may profit predominantly from the uptake of Zn and Cu by the fungal symbionts.     

Effects of Zn<Superscript>2+</Superscript> on nodulation and growth of a South American actinorhizal plant, <Emphasis Type="Italic">Discaria americana</Emphasis> (Rhamnaceae)

Discaria americana is a xerophytic shrub which lives in symbiosis with an actinomycete of the genus Frankia. The objective of this paper was to investigate the effects of high soil Zn²⁺ concentrations on growth and nodulation on the association Discaria americana–Frankia with the aim of determining if this association is suitable for improving contaminated soils. Two experiments were performed in 1 dm³ pots containing soil and different Zn additions, from 0 to 2,000 mg Zn²⁺ kg⁻¹ dry soil, with or without N fertilization. Zn additions strongly delayed shoot and root growth, but once growth was initiated, the biomass production of the plants supplied with moderate Zn amounts did not differ from the control plants. Zn reduced the final nodule number, but not the total nodule biomass. At the end of the experiment only the highest Zn treatments showed a lower nodule weight than the control plants, while N addition completely inhibited nodulation. It is concluded than Zn reduces the number of Frankia infections, but once the actinomycete is inside the roots, nodules can continue growing according to plant demand for N, compensating the reduced nodule number with more biomass. On the other hand, there is a toxic effect of Zn itself on plants when present in very high concentrations.     

Cd-induced growth reduction in the halophyte <Emphasis Type="Italic">Sesuvium portulacastrum</Emphasis> is significantly improved by NaCl

The effects of Cd²⁺ and NaCl, applied together or separately, on growth and uptake of Cd²⁺ were determined for the halophyte Sesuvium portulacastrum L. Seedlings were cultivated in the presence of 50 or 100 μmol L⁻¹ Cd²⁺ alone or combined with 100 or 400 mmol L⁻¹ NaCl. Data showed that alone, Cd²⁺ induced chlorosis, necrosis, and inhibited growth. Addition of NaCl to Cd²⁺-containing medium restored growth and alleviated the toxicity, however. NaCl also enhanced the amounts of Cd²⁺ accumulated in the shoots. All Cd²⁺ treatment reduced K⁺ and Ca²⁺ uptake and transport to the shoots. Accumulation of Na⁺ in the shoots was not affected by Cd²⁺, however. Thus S. portulacastrum maintained its halophytic characteristics in the presence of Cd²⁺. We suggest this halophyte could be used for phytoextraction of Cd²⁺ from salt-contaminated sites.     

Cadmium-induced stress on the seed germination and seedling growth of <Emphasis Type="Italic">Brassica napus</Emphasis> L., and its alleviation through exogenous plant growth regulators

Because of its prolific growth, oilseed rape (Brassica napus L.) can be grown advantageously for phytoremediation of the lands contaminated by industrial wastes. Therefore, toxic effect of cadmium on the germination of oilseed rape, the capability of plants for cadmium phytoextraction, and the effect of exogenous application of plant growth regulators to mitigate phytotoxicity of cadmium were investigated. For the lab study of seedlings at early stage, seeds were grown on filter papers soaked in different solutions of Cd²⁺ (0, 10, 50, 100, 200 and 400 μM). In greenhouse study, seedlings were grown in soil for 8 weeks, transferred to hydroponic pots for another 6 weeks growth, and then treated with plant growth regulators and cadmium. Four plant growth regulators viz. jasmonic acid (12.5 μM), abscisic acid (10 μM), gibberellin (50 μM) and salicylic acid (50 μM); and three levels of Cd²⁺ (0, 50 and 100 μM) were applied. Data indicated that lower concentration of Cd²⁺ (10 μM) promoted the root growth, whereas the severe stresses (200 or 400 μM) had negative effect on the establishment of germinating seedlings. Plants treated with any of the tested plant growth regulators alleviated cadmium toxicity symptoms, which were reflected by more fresh weight, less malondialdehyde concentration in leaves and lower antioxidant enzyme activities. The application of abscisic acid to the plants cultivated in the medium containing 100 μM Cd²⁺ resulted in significantly lower plant internal cadmium accumulation. Huabing Meng and Shujin Hua contributed equally to this paper.     

镉胁迫下硒对罗汉果组培苗光合特性的影响

实验以罗汉果组培苗为材料,室内栽培在内装市售营养土的塑料盆中,以0、10、50、100、200mg·kg-1浓度镉离子和1mg·kg-1浓度硒处理,培养20d后分析罗汉果幼苗的相关光合生理指标。结果表明:低浓度Cd2+对叶片叶绿素含量、光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)影响不大或稍有上升,但高浓度镉离子处理植株叶片的叶绿素含量、光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)明显下降;随Cd2+处理浓度的增加,叶片胞间CO2浓度(Ci)呈现上升趋势;加硒则延缓叶绿素下降,促进光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)上升,降低叶片胞间CO2浓度(Ci)。表明高浓度镉离子的毒害导致罗汉果组培苗叶片光合性能受到伤害,从而影响罗汉果幼苗生长。镉硒混合处理反映出硒对镉的毒害有缓解作用。     

Effects of pre-inoculation with a vesicular-arbuscular mycorrhizal fungus on growth of onions transplanted to the field as multi-seeded peat modules

Summary A 1984 field experiment tested the effect of inoculation with a vesicular-arbuscular mycorrhizal fungus on yield of onions (Allium cepa L. cv. Balstora) grown under commercial conditions from seedlings raised in peat modules. Roots in commercial blocking compost (M 64) could not be infected, so a modified peat, containing 50% of sterilized clay soil, was used to produce mycorrhizal seedlings. Treatments to seedlings were: uninoculated in M64 compost (K), uninoculated in modified medium (NM) and inoculated withGlomus mosseae in modified medium (M). There were two blocks of plots, one irrigated, one not. At harvest the yields of marketable (>20 mm bulb diameter) onions from M seedlings were generally about twice those from NM seedlings. On non-irrigated plots M seedlings yielded 30.3 tha⁻¹, slightly less than did K seedlings (36.6 t ha⁻¹). On irrigated plots M seedlings yielded 35.3 t ha⁻¹ and K seedlings 34.9 t ha⁻¹, but this difference was not significant. Differences in size of bulbs at harvest were small even though rates of vegetative growth differed markedly between treatments during crop development. Variations in final yield arose largely from differences in numbers of onions that failed to bulb (thicknecks). Irrigation increased mean bulb weight in all treatments but also markedly increased the number of thicknecks. Unexpectedly, the increase in thicknecks was much less in inoculated plants. This effect of mycorrhizal infection did not seem to be related to improved phosphorus nutrition.     

Evaluation of methods for celery (<Emphasis Type="Italic">Apium Graveolens</Emphasis> L.) transformation using <Emphasis Type="Italic">Agrobacterium tumefaciens</Emphasis> and the <Emphasis Type="Italic">bar</Emphasis> gene as selectable marker

Callus selection (CS) and the flamingo-bill explant (FB) methods were evaluated for efficacy in transformation for celery. Agrobacterium tumefaciens strains EHA105 and GV3101, each with the bar gene under the promoters NOS (pGPTV-BAR) or 35S (pDHB321.1), were used. Leaf explants were inoculated and co-cultivated for 2 d in the dark. Calluses emerged on the explants on callus medium (C), Murashige and Skoog (MS) medium + 2,4-Dichlorophenoxyacetic acid (2,4-D) (2.3 μM) + kinetin (2.8 μM) + timentin (300 mg·l⁻¹). Calluses 4- to 6-wk-old were selected for glufosinate (GS) resistance by a two step method. First, calluses were transferred to C medium + GS 0.35, 0.5, 1, 2, 5, or 10 mg·l⁻¹; calluses formed only with 0, 0.35 and 0.5 mg·l⁻¹ GS. All growing calluses from 0 and 0.35 mg·l⁻¹ and a few from 0.5 mg·l⁻¹, were divided and placed back on C + GS 0.35–0.5 mg·l⁻¹ for another 5–6 wk. Second, tolerant clones were again divided and placed on C + GS 1–50 mg·l⁻¹. When cultivar XP85 was inoculated with both strains, using pGPTVBAR, 19 glufosinate resistant (GR) callus clones were selected, but shoots regenerated only for strain EHA105 inoculations. When both of the strains (each with pDHB321.1) were inoculated on cv. XP166, 3 and 12 GR calluses occurred for EHA105 and GV3101, respectively. Using CS, a total of 34 GR callus clones were selected, and shoots were regenerated from over 50% of them on Gamborg B5 medium + 6-(γ, γ-dimethylallylamino) purine 2ip (4.9 μM) + naphthaleneacetic acid (NAA; 1.6 μM) and rooted on MS in 5–6 mo total time. Conversely, using FB with inoculation by GV3101/pDHB321.1 on cv. XP166 yielded putative transgenic celery plants confirmed by polymerase chain reaction (PCR) in just 6 wk. Transformation of the bar gene into celery was confirmed by PCR for 5 and 6 CS and FB lines, respectively. Southern blot analyses indicated 1–2 copies in CS lines and 1 copy in FB lines. Herbicide assays on whole plants with 100 and 300 mg·l⁻¹ glufosinate indicated a range of low to high tolerance for lines derived by both methods. The bar gene was found to be Mendelian inherited in one self-fertile CS derived line.     

Zinc-tolerant Suillus bovinus improves growth of Zn-exposed Pinus sylvestris seedlings

Scots pine (Pinus sylvestris L.) seedlings inoculated or not (NM) by a Zn-sensitive or a Zn-tolerant isolate of the ectomycorrhizal fungus Suillus bovinus (L. Fr.) Roussel were exposed to 0.1 or 150 μM Zn²⁺ for 9 months. We hypothesized that inoculation with a Zn-tolerant S. bovinus isolate should result in added Zn resistance of the host plant. Plant and fungal growth as well as nutrient profiles and photosynthetic pigments in pine needles were quantified. In NM plants and in plants colonized by the Zn-sensitive isolate, plant growth, N, P, Mg and Fe assimilation were strongly inhibited under Zn stress and concurred with significantly reduced chlorophyll concentrations. In contrast, plants colonized by the Zn-tolerant isolate grew much better and remained physiologically healthier when exposed to elevated Zn. These results provide further evidence for the important role metal-adapted mycorrhizal fungi play as an effective biological barrier against metal toxicity in trees.     

Effect of bioaccumulation of cadmium on biomass productivity,essential trace elements,chlorophyll biosynthesis,and macromolecules of wheat seedlings

Soil contamination with heavy metals has become a worldwide problem, leading to losses in agricultural yield and hazardous human health effects as they enter the food chain. The present investigation was undertaken to examine the influence of cadmium (Cd²⁺) on the wheat (Triticum aestivum L.) plant. Cd²⁺ accumulation and distribution in 3-wk-old seedlings grown in nutrient medium containing varying concentrations of Cd²⁺ (control, 0.25, 0.50, 1.0, 2.5, and 5.0 mg/L) was monitored. The effect of varying Cd²⁺ concentrations up to 21 d on biomass productivity, plant growth, photosynthetic pigments, protein, amino acids, starch, soluble sugars, and essential nutrients uptake was studied in detail to explore the level up to which the plant can withstand the stress of heavy metal. Plants treated with 0.5, 1.0, 2.5, and 5.0 mg/L Cd²⁺ showed symptoms of heavy-metal toxicity as observed by various morphological parameters which were recorded with the growth of plants. The root, shoot-leaf length and the root, shoot-leaf biomass progressively decreased with increasing Cd²⁺ concentration in the nutrient medium. Cd²⁺ uptake and accumulation was found to be maximum during the initial growth period. Cd²⁺ also interfered with the nutrients uptake, especially calcium (Ca²⁺), magnesium (Mg²⁺), potassium (K⁺), iron (Fe²⁺), zinc (Zn²⁺), and manganese (Mn²⁺) from the growth medium. Growth reduction and altered levels of major biochemical constituents such as chlorophyll, protein, free amino acids, starch, and soluble sugars that play a major role in plant metabolism were observed in response to varying concentrations of Cd²⁺ in the nutrient medium. In the present study, the effects of Cd²⁺ on growth, biomass productivity, mineral nutrients, chlorophyll biosynthesis, protein, free amino acid, starch, and soluble sugars in wheat plants was estimated to establish an overall picture of the Cd²⁺ toxicity at structural and functional levels.     

Dynamics of zinc uptake and accumulation in the hyperaccumulating and non-hyperaccumulating ecotypes of Sedum alfredii Hance

Sedum alfredii Hance has been identified as a Zn-hyperaccumulating plant species native to China. The characteristics of Zn uptake and accumulation in the hyperaccumulating ecotype (HE) and non-hyperaccumulating ecotype (NHE) of S. alfredii were investigated under nutrient solution and soil culture conditions. The growth of HE was normal up to 1000 μM Zn in nutrient solution, and 1600 mg Zn kg⁻¹ soil in a Zn-amended soil. Growth of the NHE was inhibited at Zn levels ≥250 μM in nutrient solution. Zinc concentrations in the leaves and stems increased with increasing Zn supply levels, peaking at 500 and 250 μM Zn in nutrient solution for the HE and the NHE, respectively, and then gradually decreased or leveled off with further increase in solution Zn. Minimal increases in root Zn were noted at Zn levels up to 50 μM; root Zn sharply increased at higher Zn supply. The maximum Zn concentration in the shoots of the HE reached 20,000 and 29,000 mg kg⁻¹ in the nutrient solution and soil experiments, respectively, approximately 20 times greater than those of the NHE. Root Zn concentrations were higher in the NHE than in the HE when plants were grown at Zn levels ≥50 μM. The time-course of Zn uptake and accumulation exhibited a hyperbolic saturation curve: a rapid linear increase during the first 6 days in the long-term and 60 min in the short-term studies; followed by a slower increase or leveling off with time. More than 80% of Zn accumulated in the shoots of the HE at half time (day 16) of the long-term uptake in 500 μM Zn, and also at half time (120 min) of the short-term uptake in 10 μM ⁶⁵Zn²⁺. These results indicate that Zn uptake and accumulation in the shoots of S. alfredii exhibited a down-regulation by internal Zn accumulated in roots or leaves under both nutrient solution and soil conditions. An altered Zn transport system and increased metal sequestration capacity in the shoot tissues, especially in the stems, may be the factors that allow increased Zn accumulation in the hyperaccumulating ecotype of S. alfredii. Section Editor: F. J. Zhao     

Kinetic properties of a micronutrient transporter from<Emphasis Type="Italic"> Pisum sativum</Emphasis> indicate a primary function in Fe uptake from the soil

Fe uptake in dicotyledonous plants is mediated by a root plasma membrane-bound ferric reductase that reduces extracellular Fe(III)-chelates, releasing Fe²⁺ ions, which are then absorbed via a metal ion transporter. We previously showed that Fe deficiency induces an increased capacity to absorb Fe and other micronutrient and heavy metals such as Zn²⁺ and Cd²⁺ into pea (Pisum sativum L.) roots [Cohen et al. (1998) Plant Physiol 116:1063–1072). To investigate the molecular basis for this phenomenon, an Fe-regulated transporter that is a homologue of the Arabidopsis IRT1 micronutrient transporter was isolated from pea seedlings. This cDNA clone, designated RIT1 for root iron transporter, encodes a 348 amino acid polypeptide with eight putative membrane-spanning domains that is induced under Fe deficiency and can functionally complement yeast mutants defective in high- and low-affinity Fe transport. Chelate buffer techniques were used to control Fe²⁺ in the uptake solution at nanomolar activities representative of those found in the rhizosphere, and radiotracer methodologies were employed to show that RIT1 is a very high-affinity ⁵⁹Fe²⁺ uptake system (K _m =54–93 nM). Additionally, radiotracer (⁶⁵Zn, ¹⁰⁹Cd) flux techniques were used to show that RIT can also mediate a lower affinity Zn and Cd influx (K _m of 4 and 100 M, for Zn²⁺ and Cd²⁺, respectively). These findings suggest that, in typical agricultural soils, RIT1 functions primarily as a high-affinity Fe²⁺ transporter that mediates root Fe acquisition. This is consistent with recent findings with Arabidopsis IRT1 knockout mutants that strongly suggest that this transporter plays a key role in root Fe uptake and nutrition. However, the ability of RIT1 to facilitate Zn and Cd uptake when these metals are present at elevated concentrations suggests that RIT1 may be one pathway for the entry of toxic metals into the food chain. Furthermore, the finding that plant Fe deficiency status may promote heavy metal uptake via increased expression of this transporter could have implications both for human nutrition and also for phytoremediation, the use of terrestrial plants to sequester toxic metals from contaminated soil.     

Cadmium and zinc influx characteristics by intact corn (Zea mays L.) seedlings

Summary Cadmium and zinc uptake parameters were determined for intact corn (Zea mays L.) seedlings grown for 15 and 22 in nutrient solutions containing levels of Cd and Zn that were similar to those found in soil solutions. Uptake of both elements was assumed to follow Michaelis-Menten kinetics. Calculations were based on the concentrations of free ionic Cd (Cd²⁺) and Zn (Zn²⁺) rather than the total solution concentration. Rates of Zn uptake were measured by determining depletion of Zn for periods of up to 30 h from solutions containing initial concentrations of 1.5 and 10μmol Zn 1⁻¹. Depletion curves suggested that Zn uptake characteristics were similar at both levels of Zn in solution. The Imax for Zn uptake decreased from 550 to 400 pmol m⁻² root surface s⁻¹ between 16 and 22 d of growth while Km decreased from 2.2 to 1.5 μmol Zn²⁺ 1⁻¹. Cadmium uptake parameters were measured by controlling Cd²⁺ activities in nutrient solution betwen 6.3 to 164 nmol l⁻¹ by continuous circulation of nutrient solution through a mixed-resin system. Imax for Cd uptake was 400 pmol m⁻² root surface s⁻¹ at 15 and 22 d of growth. The magnitude of Km increased from 30 to 100 nmol Cd²⁺ 1⁻¹ during this time period. The Km value suggests that corn is efficient for Cd uptake. The results of these uptake studies are consistent with the observed uptake of Zn and Cd by corn seedlings in soils.     

Effects of heavy metals on seed germination and early seedling growth of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Seed is a developmental stage that is highly protective against external stresses in the plant life cycle. In this study, we analyzed toxicity of essential (Cu²⁺ and Zn²⁺) and non-essential heavy metals (Hg²⁺, Pb²⁺ and Cd²⁺) on seed germination and seedling growth in the model species Arabidopsis. Our results show that seedling growth is more sensitive to heavy metals (Hg²⁺, Pb²⁺, Cu²⁺ and Zn²⁺) in comparison to seed germination, while Cd²⁺ is the exception that inhibited both of these processes at similar concentrations. To examine if toxicity of heavy metals is altered developmentally during germination, we incubated seeds with Hg²⁺ or Cd²⁺ only for a restricted period during germination. Hg²⁺ displayed relatively strong toxicity at period II (12–24 h after imbibition), while Cd²⁺ was more effective to inhibit germination at period I (0–12 h after imbibition) rather than at period II. The observed differences are likely to be due in part to selective uptake of different ions by the intact seed, because isolated embryos (without seed coat and endosperm) are more sensitive to both Hg²⁺ and Cd²⁺ at period I. We assessed interactive toxicity between heavy metals and non-toxic cations, and found that Ca²⁺ was able to partially restore the inhibition of seedling growth by Pb²⁺ and Zn²⁺.     

Response of citronella Java (Cymbopogon winterianus Jowitt) to VA mycorrhizal fungi and soil compaction in relation to P supply

Nutrient acquisition and growth of citronella Java (Cymbopogon winterianus Jowitt) was studied in a P-deficient sandy soil to determine the effects of mycorrhizal symbiosis and soil compaction. A pasteurized sandy loam soil was inoculated either with rhizosphere microorganisms excluding VAM fungi (non-mycorrhizal) or with the VAM fungus, Glomus intraradices Schenck and Smith (mycorrhizal) and supplied with 0, 50 or 100 mg P kg^-1 soil. The soil was compacted to a bulk density of 1.2 and 1.4 Mg m^-3 (dry soil basis). G. intraradices substantially increased root and shoot biomass, root length, nutrient (P, Zn and Cu) uptake per unit root length and nutrient concentrations in the plant, compared to inoculation with rhizosphere microorganisms when the soil was at the low bulk density and not amended with P. Little or no plant response to the VAM fungus was observed when the soil was supplied with 50 or 100 mg P kg^-1 soil and/or compacted to the highest bulk density. At higher soil compaction and P supply the VAM fungus significantly reduced root length. Non-mycorrhizal plants at higher soil compaction produced relatively thinner roots and had higher concentrations and uptake of P, Zn and Cu than at lower soil compaction, particularly under conditions of P deficiency. The quality of citronella Java oil measured in terms citronellal and d-citronellol concentration did not vary appreciably due to various soil treatments.     

Calcium protects <Emphasis Type="Italic">Trifolium repens</Emphasis> L. seedlings against cadmium stress

The effect of calcium (Ca²⁺) on Trifolium repens L. seedlings subjected to cadmium (Cd²⁺) stress was studied by investigating plant growth and changes in activity of antioxidative enzymes. Physiological analysis was carried out on seedlings cultured for 2 weeks on half-strength Hoagland medium with Cd²⁺ concentrations of 0, 400 and 600 μM, and on corresponding medium supplied with CaCl₂ (5 mM). Exposure to increasing Cd²⁺ reduced the fresh weight of the upper part (stems + leaves) of the seedlings more strongly than that of the root system. In both parts of T. repens seedlings H₂O₂ level and lipid peroxidation increased. In the upper part, Cd²⁺ exposure led to a significant decrease in the activity of superoxide dismutase, catalase and glutathione peroxidase and an increase in ascorbate peroxidase activity. In contrast, the roots showed an increase in the activity of antioxidative enzymes under Cd²⁺ stress. Ca²⁺ addition to medium reduced the Cd²⁺ accumulation, and considerably reversed the Cd²⁺-induced decrease in fresh mass as well as the changes in lipid peroxidation in the both parts of T. repens seedlings. Ca²⁺ application diminished the Cd²⁺ effect on the activity of antioxidative enzymes in the upper part, even though it did not significantly affect these enzymes in the roots. So the possible mechanisms for the action of Ca²⁺ in Cd²⁺ stress were considered to reduce Cd²⁺ accumulation, alleviate lipid peroxidation and promote activity of antioxidative enzymes.     

Effect of heavy metals on the glutathione status in different ectomycorrhizal <Emphasis Type="Italic">Paxillus involutus</Emphasis> strains

The glutathione (GSH) status and heavy metal tolerance were investigated in four Paxillus involutus strains isolated from different heavy-metal-polluted and non-polluted regions of Europe. The heavy metal burden in the habitats did not affect significantly either the heavy metal (Cr₂O₇²⁻, Cd²⁺, Hg²⁺, Pb²⁺, Zn²⁺, Cu²⁺) tolerance and accumulation or the GSH production of the strains tested. Exposures to heavy metals increased the intracellular GSH concentrations in 12 from 24 experimental arrangements (four strains exposed to six heavy metals) independently of the habitats of the strains. The importance of GSH in heavy metal tolerance (high MIC values, ability to accumulate heavy metals and to grow in the presence of heavy metals) was thus demonstrated in this ectomycorrhizal fungus.     

Heavy metal resistant <Emphasis Type="Italic">Distigma proteus</Emphasis> (Euglenophyta) isolated from industrial effluents and its possible role in bioremediation of contaminated wastewaters

Summary The alga, Distigma proteus, isolated from industrial wastewater showed tolerance against Cd²⁺ (8.0 μg/ml), Cr⁶⁺ (12 μg/ml), Pb²⁺ (15 μg/ml) and Cu²⁺ (10 μg/ml). The metal ions slowed down the growth of the organism after 4–5 days of exposure. The reduction in cell population was 90% for Cu²⁺, 84% for Cd²⁺, 71% for Cr⁶⁺, and 63% for Pb²⁺ after 8 days of metal stress. The order of resistance to heavy metal, in terms of reduction in the cellular population, was Cu²⁺ > Cd²⁺ > Cr⁶⁺ > Pb²⁺. Chromium- and cadmium-processing capabilities of the alga were worked out for its potential use as a bioremediator of wastewater. The reduction in the amount of Cr⁶⁺ after 2, 4, 6 and 8 days of algal culture containing 5.0 μg Cr⁶⁺ ml⁻¹ of culture medium was 77, 85, 92 and 97%, respectively. Distigma could also remove 48% Cd²⁺after 2 days, 68% after 4 days, 80% after 6 days and 90% after 8 days from the medium. The heavy metal uptake ability of Distigma can be exploited for metal detoxification and environmental clean-up operations.     

Amelioration of phytotoxic effects of Cd on mung bean seedlings by gluconic acid secreting rhizobacterium <Emphasis Type="Italic">Enterobacter asburiae</Emphasis> PSI3 and implication of role of organic acid

Mung bean seedlings inoculated with Enterobacter asburiae PSI3, a gluconic acid-producing rhizosphere isolate, enhanced plant growth in the presence of phytotoxic levels of Cd²⁺ in gnotobiotic pot experiments as compared to the uninoculated Cd-treated plants. Addition of organic acids to Cd-stressed seedlings promoted root elongation. Hematoxylin competition assays showed that organic acids could displace Cd²⁺ from the Cd²⁺: hematoxylin complex in the same order of effectiveness as was found for restoration of root net elongation viz. oxalate > malate > succinate while gluconate was effective at higher concentrations. Root associated Cd²⁺, assessed by hematoxylin staining of roots was found to be reduced when roots were treated with organic acid. Cd stress increased antioxidant enzymes such as peroxidase and superoxide dismutase in mung bean roots while organic acid treatment suppressed the up-regulation of these enzymes by Cd.