Production of indole-3-acetic acid via the indole-3-acetamide pathway in the plant-beneficial bacterium Pseudomonas chlororaphis O6 is inhibited by ZnO nanoparticles but enhanced by CuO nanoparticles.
ABSTRACT: The beneficial bacterium Pseudomonas chlororaphis O6 produces indole-3-acetic acid (IAA), a plant growth regulator. However, the pathway involved in IAA production in this bacterium has not been reported. In this paper we describe the involvement of the indole-3-acetamide (IAM) pathway in IAA production in P. chlororaphis O6 and the effects of CuO and ZnO nanoparticles (NPs). Sublethal levels of CuO and ZnO NPs differentially affected the levels of IAA secreted in medium containing tryptophan as the precursor. After 15 h of growth, CuO NP-exposed cells had metabolized more tryptophan than the control and ZnO NP-challenged cells. The CuO NP-treated cells produced higher IAA levels than control cultures lacking NPs. In contrast, ZnO NPs inhibited IAA production. Mixing of CuO and ZnO NPs resulted in an intermediate level of IAA production relative to the levels in the separate CuO and ZnO NP treatments. The effect of CuO NPs on IAA levels could be duplicated by ions at the concentrations released from the NPs. However, ion release did not account for the inhibition caused by the ZnO NPs. The mechanism underlying changes in IAA levels cannot be accounted for by effects on transcript accumulation from genes encoding a tryptophan permease or the IAM hydrolase in 15-h cultures. These findings raise the issue of whether sublethal doses of NPs would modify the beneficial effects of association between plants and bacteria.
Project description:Nanoparticle (NPs) containing essential metals are being considered in formulations of fertilizers to boost plant nutrition in soils with low metal bioavailability. This paper addresses whether colonization of wheat roots by the bacterium, Pseudomonas chlororaphis O6 (PcO6), protected roots from the reduced elongation caused by CuO NPs. There was a trend for slightly elongated roots when seedlings with roots colonized by PcO6 were grown with CuO NPs; the density of bacterial cells on the root surface was not altered by the NPs. Accumulations of reactive oxygen species in the plant root cells caused by CuO NPs were little affected by root colonization. However, bacterial colonization did reduce the extent of expression of an array of genes associated with plant responses to stress induced by root exposure to CuO NPs. PcO6 colonization also reduced the levels of two important chelators of Cu ions, citric and malic acids, in the rhizosphere solution; presumably because these acids were used as nutrients for bacterial growth. There was a trend for lower levels of soluble Cu in the rhizosphere solution and reduced Cu loads in the true leaves with PcO6 colonization. These studies indicate that root colonization by bacterial cells modulates plant responses to contact with CuO NPs.
Project description:The unregulated discharge of nanoparticles (NPs) from various nanotechnology industries into the environment is expected to alter the composition and physiological functions of soil microbiota. Considering this knowledge gap, the impact of five NPs (Ag, ZnO, CuO, Al2O3, and TiO2) differing in size and morphology on growth behavior and physiological activity of Azotobacter chroococcum, Bacillus thuringiensis, Pseudomonas mosselii, and Sinorhizobium meliloti were investigated. Various biochemical and microscopic approaches were adopted. Interestingly, all bacterial strains were found sensitive to Ag-NPs and ZnO-NPs but showed tolerance toward CuO, Al2O3, and TiO2-NPs. The loss of cellular respiration due to NPs was coupled with a reduction in population size. ZnO-NPs at 387.5 ?g mL-1 had a maximum inhibitory impact on A. chroococcum and reduced its population by 72%. Under Ag-NP stress, the reduction in IAA secretion by bacterial strains followed the order S. meliloti (74%) > P. mosselii (63%) > A. chroococcum (49%). The surface of bacterial cells had small- or large-sized aggregates of NPs. Also, numerous gaps, pits, fragmented, and disorganized cell envelopes were visible. Additionally, a treated cell surface appeared corrugated with depressions and alteration in cell length and a strong heterogeneity was noticed under atomic force microscopy (AFM). For instance, NPs induced cell roughness for P. mosselii followed the order 12.6 nm (control) > 58 nm (Ag-NPs) > 41 nm (ZnO-NPs). TEM analysis showed aberrant morphology, cracking, and disruption of the cell envelope with extracellular electron-dense materials. Increased permeability of the inner cell membrane caused cell death and lowered EPS production. Ag-NPs and ZnO-NPs also disrupted the surface adhering ability of bacteria, which varied with time and concentration of NPs. Conclusively, a plausible mechanism of NP toxicity to bacteria has been proposed to understand the mechanistic basis of ecological interaction between NPs and resourceful bacteria. These results also emphasize to develop strategies for the safe disposal of NPs.
Project description:Black mustard (Brassica nigra) was grown in pots amended with 41 nm ZnO (200-600 mg/kg soil) and 47 nm CuO (12.5-50 mg/kg soil) nanoparticles (NPs) to analyze growth response and yield characteristics. B. nigra seed germination was not affected by CuO NPs, but significant toxicity was observed by ZnO NP treatment. Both NPs significantly increased the growth profile of B. nigra, i.e., the stem height, number of leaves, average leaf area, number of branches, and number of nodes per plant. Application of ZnO and CuO NPs brought a significant dose-dependent decrease in primary root length; however, the number of secondary roots increased in the presence of CuO NPs. The average number of flowers and pods per plant significantly increased in the presence of CuO NPs. The seed yield, average seed weight per plant, and seed diameter parameters were observed to be better in the presence of CuO NPs as compared with ZnO NPs. Total protein contents and glucosinolates increased in the seeds grown in the NP-amended soil, while total oil contents decreased. Oil analysis depicted that oleic acid and linolenic acid percentage decreased while erucic acid percentage increased in seeds in the presence of both NPs in the soil. An atomic absorption spectrophotometer showed accumulation of Cu and Zn in B. nigra in the following order: root > stem > leaves > seeds. The study concludes that CuO and ZnO NPs have detrimental effect on the B. nigra plant and yield. The release of NPs and type of metal in NPs might also have a positive effect on the plant; however, their concentration in the soil also matters.
Project description:Zinc Oxide nanoparticles (ZnO NPs) are being rapidly developed for use in consumer products, wastewater treatment and chemotherapy providing several possible routes for ZnO NP exposure to humans and aquatic organisms. Recent studies have shown that ZnO NPs undergo rapid dissolution to Zn+2, but the relative contribution of Zn+2 to ZnO NP bioavailability and toxicity is not clear. Gene expression profiling of D. magna exposed to ZnO NPs or ZnSO4 at equitoxic concentrations demonstrated that the particles cause toxicity through a distinct mechanism compared with Zn+2. D. magna were also exposed to a SiO NPs as a particle control at equimolar concentrations. The SiO NPs resulted in few differentially expressed genes and there was very little overlap between the genes affected by the ZnO NPs and the SiO NPs, suggesting that ZnO NPs cause a distinct pattern of differentially expressed genes. In the ZnO NP exposures, effects were observed to genes involved in cytoskeletal transport, cellular respiration and reproduction. Three biomarker genes including a multi-cystatin, ferritin and a C1q containing gene were confirmed as differentially expressed in a specific pattern by ZnO NP and provide a suite of biomarkers for identifying environmental exposure to ZnO NP and differentiating between NP and ionic exposure. Overall design: We exposed Daphnia magna to the 1/10 LC50 and LC25 of ZnO nanoparticles and Zn++ as ZnSO4 for 24-h. For each exposure condition, we performed 3 exposures and 2 technical replicates (as dye swap) for each exposure (6 microarrays total). All exposures were compared to a unexposed laboratory control
Project description:The effects of engineered nanoparticles (ENPs) on heavy metal fate and biotoxicity in farmland soil are mostly unknown. A flooding-drying simulation experiment was conducted to study the effects of three typical metal oxide nanoparticles (TiO2-NPs, ZnO-NPs and CuO-NPs) on the chemical speciation of heavy metals and micronutrient bioavailability in paddy soil. The results showed that the addition of ZnO-NPs and CuO-NPs caused significant increases in soil pH, Eh and EC after a 90-d flooding-drying process. ZnO-NPs and CuO-NPs addition caused clearly increase in the Zn and Cu concentrations in the acid-soluble fraction, Fe/Mn oxides-bound fraction and organic-bound fraction, leading to higher bioavailability in the soil. DTPA-extractable Zn and Cu increased to 184.6 mg kg-1 and 145.3 mg kg-1 in the maximum ZnO-NPs and CuO-NPs concentration treatments (500 mg kg-1). TiO2-NPs promoted the transformation of Mn from a Fe/Mn oxides-bound fraction to an acid-soluble fraction. Soil Cd bioavailability obviously decreased in the TiO2-NPs treatment but increased in the ZnO-NPs and CuO-NPs treatments.
Project description:The plant pathogen Agrobacterium tumefaciens infects plants and introduces the transferred-DNA (T-DNA) region of the Ti-plasmid into nuclear DNA of host plants to induce the formation of tumors (crown galls). The T-DNA region carries iaaM and iaaH genes for synthesis of the plant hormone auxin, indole-3-acetic acid (IAA). It has been demonstrated that the iaaM gene encodes a tryptophan 2-monooxygenase which catalyzes the conversion of tryptophan to indole-3-acetamide (IAM), and the iaaH gene encodes an amidase for subsequent conversion of IAM to IAA. In this article, we demonstrate that A. tumefaciens enhances the production of both IAA and phenylacetic acid (PAA), another auxin which does not show polar transport characteristics, in the formation of crown galls. Using liquid chromatography-tandem mass spectroscopy, we found that the endogenous levels of phenylacetamide (PAM) and PAA metabolites, as well as IAM and IAA metabolites, are remarkably increased in crown galls formed on the stem of tomato plants, implying that two distinct auxins are simultaneously synthesized via the IaaM-IaaH pathway. Moreover, we found that the induction of the iaaM gene dramatically elevated the levels of PAM, PAA and its metabolites, along with IAM, IAA and its metabolites, in Arabidopsis and barley. From these results, we conclude that A. tumefaciens enhances biosynthesis of two distinct auxins in the formation of crown galls.
Project description:Nanoparticles (NPs) are widely used in various domestic products and their usage is constantly increasing which in turn can raise several environmental health issues. Like other abiotic stresses, nanomaterials also affect the growth of crop plants. Solanum melongena is a common vegetable crop grown in the tropics and subtropics regions with medicinal properties. In this study, S. melongena was analyzed for its response to three commercially important metallic nanoparticles, namely NiO, CuO, and ZnO, at four different concentrations (100, 250, 500 and 1000 mg/L). The growth of the eggplant seedlings was suppressed by all the NPs in a concentration-dependent manner and among them, NiO was shown to be more toxic as it suppressed the root and shoot growth effectively. Total chlorophyll contents were decreased in the NP-treated plants compared to control plants. Significant changes were found in the secondary metabolites such as anthocyanins, total phenolic and total flavonoid contents in the NP-treated plants. A dose-dependent increase in the reactive oxygen species (ROS) generation was noticed in the NP-treated plants which are evidenced by the 4-nitro blue tetrazolium chloride (NBT) and 3,3'-diamiobenzidine (DAB) histochemical staining. The DNA damage imposed by the NP in the seedlings of eggplants may be due to the elevated ROS and MDA (malondialdehyde) production. NiO NP was found to be more toxic comparable to CuO and ZnO NPs in the present study. Apart from the toxic effects, nanoparticles also showed profound effects on the production of important secondary metabolites such as phenolics and flavonoid compounds.
Project description:Arthrobacter pascens ZZ21 is a plant-beneficial, fluoranthene-degrading bacterial strain found in the rhizosphere. The production of the phytohormone indole-3-aectic acid (IAA) by ZZ21 is thought to contribute to its ability to promote plant growth and remediate fluoranthene-contaminated soil. Using genome-wide analysis combined with metabolomic and high-performance liquid chromatography-mass spectrometry (HPLC-MS) analyses, we characterized the potential IAA biosynthesis pathways in A. pascens ZZ21. IAA production increased 4.5-fold in the presence of 200 mg·L-1 tryptophan in the culture medium. The transcript levels of prr and aldH, genes which were predicted to encode aldehyde dehydrogenases, were significantly upregulated in response to exogenous tryptophan. Additionally, metabolomic analysis identified the intermediates indole-3-acetamide (IAM), indole-3-pyruvic acid (IPyA), and the enzymatic reduction product of the latter, indole-3-lactic acid (ILA), among the metabolites of ZZ21, and subsequently also IAM, ILA, and indole-3-ethanol (TOL), which is the enzymatic reduction product of indole-3-acetaldehyde, by HPLC-MS. These results suggest that the tryptophan-dependent IAM and IPyA pathways function in ZZ21.
Project description:Zinc Oxide nanoparticles (ZnO NPs) are being rapidly developed for use in consumer products, wastewater treatment and chemotherapy providing several possible routes for ZnO NP exposure to humans and aquatic organisms. Recent studies have shown that ZnO NPs undergo rapid dissolution to Zn+2, but the relative contribution of Zn+2 to ZnO NP bioavailability and toxicity is not clear. Gene expression profiling of D. magna exposed to ZnO NPs or ZnSO4 at equitoxic concentrations demonstrated that the particles cause toxicity through a distinct mechanism compared with Zn+2. D. magna were also exposed to a SiO NPs as a particle control at equimolar concentrations. The SiO NPs resulted in few differentially expressed genes and there was very little overlap between the genes affected by the ZnO NPs and the SiO NPs, suggesting that ZnO NPs cause a distinct pattern of differentially expressed genes. In the ZnO NP exposures, effects were observed to genes involved in cytoskeletal transport, cellular respiration and reproduction. Three biomarker genes including a multi-cystatin, ferritin and a C1q containing gene were confirmed as differentially expressed in a specific pattern by ZnO NP and provide a suite of biomarkers for identifying environmental exposure to ZnO NP and differentiating between NP and ionic exposure. We exposed Daphnia magna to the 1/10 LC50 and LC25 of ZnO nanoparticles and Zn++ as ZnSO4 for 24-h. For each exposure condition, we performed 3 exposures and 2 technical replicates (as dye swap) for each exposure (6 microarrays total). All exposures were compared to a unexposed laboratory control
Project description:IAA biosynthetic pathways in a basidiomycetous yeast, Rhodosporidiobolus fluvialis DMKU-CP293, were investigated. The yeast strain showed tryptophan (Trp)-dependent IAA biosynthesis when grown in tryptophan supplemented mineral salt medium. Gas chromatography-mass spectrometry was used to further identify the pathway intermediates of Trp-dependent IAA biosynthesis. The results indicated that the main intermediates produced by R. fluvialis DMKU-CP293 were tryptamine (TAM), indole-3-acetic acid (IAA), and tryptophol (TOL), whereas indole-3-pyruvic acid (IPA) was not found. However, supplementation of IPA to the culture medium resulted in IAA peak detection by high-performance liquid chromatography analysis of the culture supernatant. Key enzymes of three IAA biosynthetic routes, i.e., IPA, IAM and TAM were investigated to clarify the IAA biosynthetic pathways of R. fluvialis DMKU-CP293. Results indicated that the activities of tryptophan aminotransferase, tryptophan 2-monooxygenase, and tryptophan decarboxylase were observed in cell crude extract. Overall results suggested that IAA biosynthetic in this yeast strain mainly occurred via the IPA route. Nevertheless, IAM and TAM pathway might be involved in R. fluvialis DMKU-CP293.