Silver nanoparticles inhibit nitrogen fixation in soybean (Glycine max) root nodules.

Antimicrobial silver nanoparticles (AgNPs) are popular in consumer and industrial products, leading to increasing concentrations in the environment. We tested whether exposure to AgNPs could be detrimental to a microbe, its host plant, and their symbiotic relationship. When subjected to 10 µg/mL AgNPs, growth of Bradyrhizobium japonicum USDA 110 was halted. Axenic nitrogen-fertilized Glycine max seedlings were unaffected by 2.5 µg/mL of 30 nm AgNPs, but growth was inhibited with the same dose of 16 nm AgNPs. With 2.5 µg/mL AgNPs, biomass of inoculated plants was 50% of the control. Bacteroids were not found in nodules on plants treated with 2.5 µg/mL AgNPs and plants given 0.5-2.5 µg/mL AgNPs had 40-65% decreased nitrogen fixation. In conclusion, AgNPs not only interfere with general plant and bacterial growth but also inhibit nodule development and bacterial nitrogen fixation. We should be mindful of not releasing AgNPs to the environment or to agricultural land.

Uptake and phytotoxic effect of benzalkonium chlorides in Lepidium sativum and Lactuca sativa.

Cationic surfactants such as benzalkonium chlorides (BACs) are used extensively as biocides in hospitals, food processing industries, and personal care products. BACs have the potential to reach the rooting zone of crop plants and BACs might thereby enter the food chain. The two most commonly used BACs, benzyl dimethyl dodecyl ammonium chloride (BDDA) and benzyl dimethyl tetradecyl ammonium chloride (BDTA), were tested in a hydroponic system to assess the uptake by and phytotoxicity to lettuce (Lactuca sativa L.) and garden cress (Lepidium sativum L.). Individually and in mixture, BACs at concentrations up to 100 mg L-1 did not affect germination; however, emergent seedlings were sensitive at 1 mg L-1 for lettuce and 5 mg L-1 for garden cress. After 12 d exposure to 0.25 mg L-1 BACs, plant dry weight was reduced by 68% for lettuce and 75% for garden cress, and symptoms of toxicity (necrosis, chlorosis, wilting, etc.) were visible. High performance liquid chromatography-mass spectroscopy analysis showed the presence of BACs in the roots and shoots of both plant species. Although no conclusive relationship was established between the concentrations of six macro- or six micro-nutrients, growth inhibition or BAC uptake, N and Mg concentrations in BAC-treated lettuce were 50% lower than that of control, indicating that BACs might induce nutrient deficiency. Although bioavailability of a compound in hydroponics is significantly higher than that in soil, these results confirm the potential of BACs to harm vascular plants.

Sorption and leaching of benzalkonium chlorides in agricultural soils.

The adsorption and leaching characteristics of two commonly used benzalkonium chlorides (BACs), benzyl dimethyl dodecyl ammonium chloride (BDDA) and benzyl dimethyl tetradecyl ammonium chloride (BDTA) using three agricultural soils with varied proportions of silt, sand, clay, and organic matter were determined. BACs are cationic surfactants used in large quantities for sanitary and personal care products and are abundant in environmental samples. Adsorption isotherm data (aqueous concentration in the range of 25-150 mg L-1) fitted the Langmuir model better than the Freundlich model. BDTA with a longer alkyl chain adsorbed more to soil compared to BDDA, and the soil with the highest percentage of clay adsorbed the most. Column tests conducted using soils amended with lime stabilised biosolids and artificial rain water at a flow rate of 0.2 mL min-1 indicate very low leaching of BACs. Less than 1% of the available BDDA leached through sandy loam soil column with a depth of 9 cm. Therefore, the possibility of BACs to become bioavailable through leaching is very low at environmentally relevant concentrations.

Differential accumulation of cadmium in near-isogenic lines of durum wheat: no role for phytochelatins.

Certain cultivars of some crops, including durum wheat (Triticum durum Desf.), have a propensity to accumulate cadmium in the grain. In the 1980s, a Canadian wheat breeding program generated five pairs of near-isogenic lines of durum wheat that vary in cadmium-accumulation. Within each pair, one member accumulates twofold to threefold higher concentrations of cadmium in the shoot and grain. However, the physiological explanation for the high-low phenotype is unknown. We studied correlations between concentrations of cadmium and non-protein thiols, including phytochelatins, in these five pairs of near-isogenic lines to test the hypothesis that differential retention of cadmium-binding complexes in the root would explain the phenotype. The expected high-low pattern of cadmium accumulation was found in three of the pairs. In one pair, cadmium was positively correlated with cysteine and glutathione in the roots and with phytochelatins 2 and 4 in the shoots but in another pair cadmium was strongly negatively correlated with phytochelatins 2 and 4 in the shoots and unrelated to cysteine or glutathione. No correlations between concentrations of cadmium and the non-protein thiols were found in the third pair or in the remaining two pairs. The production of phytochelatins is a well-described response to cadmium but the lack of consistent correlation between cadmium and non-protein thiols in these five near-isogenic lines indicates that complexation with non-protein thiols does not explain differential translocation of cadmium in durum wheat.

Investigating the ability of Pseudomonas fluorescens UW4 to reduce cadmium stress in Lactuca sativa via an intervention in the ethylene biosynthetic pathway.

A typical plant response to any biotic or abiotic stress, including cadmium (Cd), involves increased ethylene synthesis, which causes senescence of the affected plant part. Stressed plants can experience reduced ethylene and improved growth if they are inoculated with bacteria that have the enzyme ACC deaminase, which metabolizes the ethylene precursor ACC (1-aminocyclopropane-1-carboxylate). We investigated whether one such bacterium, Pseudomonas fluorescens UW4, reduces the production of ethylene and improves the growth of lettuce (Lactuca sativa) sown in Cd-contaminated potting material (PRO-MIX® BX). Plants were inoculated with the wild-type P. fluorescens UW4 or a mutant strain that cannot produce ACC deaminase. Cadmium-treated plants contained up to 50 times more Cd than did control plants. In noninoculated plants, Cd induced a 5-fold increase in ethylene concentration. The wild-type bacterium prevented Cd-induced reductions in root biomass but there was no relationship between Cd treatment and ethylene production in inoculated plants. In contrast, when the concentration of ethylene was plotted against the extent of bacterial colonization of the roots, increased colonization with wild-type P. fluorescens UW4 was associated with 20% less ethylene production. Ours is the first study to show that the protective effect of this bacterium is proportional to the quantity of bacteria on the root surface.

Biodegradation of benzalkonium chlorides singly and in mixtures by a Pseudomonas sp. isolated from returned activated sludge.

Bactericidal cationic surfactants such as quaternary ammonium compounds (QACs) are widely detected in the environment, and found at mg kg(-1) concentrations in biosolids. Although individual QACs are amenable to biodegradation, it is possible that persistence is increased for mixtures of QACs with varying structure. The present study evaluated the biodegradation of benzyl dimethyl dodecyl ammonium chloride (BDDA) singly and in the presence of benzyl dimethyl tetradecyl ammonium chloride (BDTA) using Pseudomonas sp., isolated from returned activated sludge. Growth was evaluated, as was biodegradation using (14)C and HPLC-MS methods. BDTA was more toxic to growth of Pseudomonas sp. compared to BDDA, and BDTA inhibited BDDA biodegradation. The benzyl ring of [U-(14)C-benzyl] BDDA was readily and completely mineralized. The detection of the transformation products benzyl methyl amine and dodecyl dimethyl amine in spent culture liquid was consistent with literature. Overall, this study demonstrates the antagonistic effect of interactions on biodegradation of two widely used QACs suggesting further investigation on the degradation of mixture of QACs in wastewater effluents and biosolids.

Does the response of insect herbivores to cadmium depend on their feeding strategy?

Phytoremediation has been proposed for the elimination of toxic metals in soil, yet little attention has been given to the performance of insects that feed on contaminant-tolerant plants. We tested the performance of two herbivores with different feeding behaviors, the cabbage looper, Trichoplusia ni, and the green peach aphid, Myzus persicae, reared on cadmium-tolerant Brassica juncea plants that contained different concentrations of cadmium. We also tested the performance of the aphid parasitoid Aphidius colemani developing in aphids reared on plants with different levels of cadmium. The hypothesis tested was that the chewing insect would be more negatively affected than the sucking insect, because of the localization of cadmium within the host plant, and that the aphid parasitoid would not be affected. We also compared the performance of T. ni on artificial diet with different levels of cadmium. Neither the phloem-feeding aphid nor its parasitoid was affected by cadmium in the host plant. The effects of cadmium on the foliage-feeding cabbage looper varied, with negative effects on development observed in experiments with artificial diet but not in those using natural host plants. These data, together with information available in the literature, support the idea that the effects of toxic metals present in a host plant may be influenced by a herbivore's feeding strategy. However, a wide range of chewing and sucking species needs to be tested to confirm this hypothesis.

Reduced translocation of cadmium from roots is associated with increased production of phytochelatins and their precursors.

Cadmium (Cd) is a non-essential trace element and its environmental concentrations are approaching toxic levels, especially in some agricultural soils. Understanding how and where Cd is stored in plants is important for ensuring food safety. In this study, we examined two plant species that differ in the distribution of Cd among roots and leaves. Lettuce and barley were grown in nutrient solution under two conditions: chronic (4 weeks) exposure to a low, environmentally relevant concentration (1.0 µM) of Cd and acute (1 h) exposure to a high concentration (5.0 mM) of Cd. Seedlings grown in solution containing 1.0 µM CdCl2 did not show symptoms of toxicity and, at this concentration, 77% of the total Cd was translocated to leaves of lettuce, whereas only 24% of the total Cd was translocated to barley leaves. We tested the hypothesis that differential accumulation of Cd in roots and leaves is related to differential concentrations of phytochelatins (PCs), and its precursor peptides. The amounts of PCs and their precursor peptides in the roots and shoots were measured using HPLC. Each of PC2₋4 was synthesized in the barley root upon chronic exposure to Cd and did not increase further upon acute exposure. In the case of lettuce, no PCs were detected in the root given either Cd treatment. The high amounts of PCs produced in barley root could have contributed to preferential retention of Cd in barley roots.

Effects of cadmium on photosynthetic oxygen evolution from single stomata in Brassica juncea (L.) Czern.

Scanning electrochemical microscopy (SECM) was utilized to investigate photosynthetic oxygen evolution from single stomata in leaves of live Brassica juncea (L.) Czern cultured in nutrient solution to which 0.2 or 0.01 mM CdC12 had been added. The bulk leaf surface serves as an insulator normally; therefore, a typical negative feedback was observed on the probe approach curves (PACs) when the probe approached epidermal cells. When the probe tip approached an open stoma, a higher tip current was detected due to the O2 release from this stoma. Thus, SECM can be used to map the O2 concentration profile near the leaf surface and study stomatal complex structure size and density. The oxygen release from single stomata was also analyzed by comparison of experimental PACs with those simulated by COMSOL multiphysics software (version 3.4). In addition to an increase in the stomatal complex size and a decrease in the complex density, the Cd accumulation caused up to a 26% decrease in photosynthetic rate determined at the level of a single stoma. The O2 evolution was also monitored by recording the tip current vs time when a tip sat above the center of a stoma. Periodic peaks in O2 release-time curves were observed, varying from 400 to 1600 s. The opening and closing activities of single stomata were also imaged by SECM.

Cadmium and zinc accumulation in soybean: A threat to food safety?

A greenhouse study was conducted to quantify cadmium and zinc accumulated by soybean (Glycine max (L.) Merr.) when the metals were supplied separately and together. The highest dose of cadmium (100 mg/kg) reduced plant height and dry weight (down to 40% and 34% of control, respectively); the highest dose of zinc (2000 mg/kg) reduced plant height to 55% of control and dry weight to 70% of control. With both metals present, the plants were approximately the same size as those treated with cadmium only. The concentration of cadmium in the roots was unaffected by zinc. In other tissues, the effect of zinc on the accumulation of cadmium depended on the doses provided. At low doses, the addition of zinc reduced the concentration of cadmium in aboveground tissues to 40-50% of that found in plants exposed to cadmium only. However, when applied in high doses, the presence of zinc in cadmium-contaminated soils increased the uptake and accumulation of cadmium in aboveground tissues by up to 42%. In contrast, at high doses, the presence of cadmium in zinc-contaminated soil resulted in approximately 35% lower concentrations of zinc in all tissues. At a lower dose, cadmium had no effect on concentration of zinc in the plant tissues. The effects of high doses of one metal on the uptake of the other metal can be partially explained by the effects of one metal on the bioavailability of the other metal. In soils to which only one metal was added, bioavailable cadmium was 70-80% of the total cadmium, and bioavailable zinc was 50-70% of the total zinc. When both metals were added to the soil, 80-100% of the cadmium and 46-60% of the zinc were bioavailable. Concentrations of both metals were highest in root tissues (10-fold higher for cadmium, and up to 2-fold higher for zinc). Although relatively little cadmium was translocated to pods and seeds, the seeds of all plants (including those from control and zinc-treated plants) had concentrations of cadmium 3-4 times above the limit of 0.2 mg/kg set by the Codex Alimentarius Commission. This was surprising given that cadmium in the soil was only 1 mg/kg well below the maximum allowable amount for agricultural soil. While it is possible that more cadmium was accumulated by plants in this study than that which might occur under agricultural field conditions, these results reinforce the need to monitor concentrations of toxic metals in food crops.

Cadmium-induced plant stress investigated by scanning electrochemical microscopy.

In vivo oxygen evolution above single stomata in Brassica juncea has been used to investigate, for the first time, the effect of Cd-induced stress as imaged by scanning electrochemical microscopy (SECM). SECM images showed a clear stomatal structure-a pore, whose aperture is modulated by two guard cells, serving as the conduit for the oxygen produced. Lower stomatal density and larger stoma size were found in plants treated with 0.2 mM CdCl2 compared with control plants. Either the introduction of Cd caused a slower cell replication in the plane of the epidermis, hence fewer stomata, and/or the number of open stomata was reduced when plants were under Cd-stress. Oxygen evolution above individual stomatal complexes in Cd-treated plants was lower than that from control plants, as determined from the electrochemical current above the middle of each stoma. All guard cells under illumination were swollen, indicating that the stomata were open in both control and treated plants. Thus, decreased oxygen evolution in response to Cd cannot be attributed to simple closing of the stomata, but to a lower photosynthetic yield. SECM provides an excellent tool for monitoring the effects of Cd on photosynthetic activity at the scale of individual stomata.