Biodegradation of tributyl phosphate using Klebsiella pneumoniae sp. S3.

Tributyl phosphate (TBP) has enormous applications in the field of extraction, fuel reprocessing, as defoamers and/or plasticizers. Excessive usage of this organophosphorus compound, poses an environmental threat. The present study deals with microbial degradation of TBP using Klebsiella pneumoniae S3 isolated from the soil. Diauxic growth curve pattern explains a preferential utilization of TBP. The strain S3 was able to biotransform TBP (1,000 mg L⁻¹) to dibutyl phosphate within 48 h and showed higher tolerance towards TBP up to 17.0 g L⁻¹. Toxicity of the parent as well as degraded product was assessed using comet assay. Generation of reactive oxygen species elaborates the oxidative stress imposed upon the bacterial strain by TBP. The antioxidant defense mechanism was studied using various biomarkers namely catalase, glutathione-S-transferase, and superoxide dismutase. The present study describes a faster and eco-friendly alternative for disposal of TBP.

Identification and profiling of arsenic stress-induced microRNAs in Brassica juncea.

MicroRNAs (miRNAs) constitute a novel mechanism of gene regulation affecting plant development, growth, and stress response. To study the role of miRNAs in arsenic (As) stress, microarray profiling of miRNAs was performed in Brassica juncea using a custom Phalanx Plant OneArray containing 381 unique miRNA probes representing 618 miRNAs from 22 plant species. miRNA microarray hybridization of roots exposed to As for 1h and 4h revealed that a total of 69 miRNAs belonging to 18 plant miRNA families had significantly altered expression. The As-responsive miRNAs also exhibited a time- and organ-dependent change in their expression. Putative target prediction for the miRNAs suggested that they regulate various developmental processes (e.g. miR156, miR169, and miR172), sulphur uptake, transport, and assimilation (miR395, miR838, and miR854), and hormonal biosynthesis and/or function (e.g. miR319, miR167, miR164, and miR159). Notable changes were observed in the level of auxins [indole-3-acetic acid (IAA), indole-3- butyric acid, and naphthalene acetic acid], jasmonates [jasmonic acid (JA) and methyl jasmonate], and abscisic acid. The exogenous supply of JA and IAA improved growth of plants under As stress and altered expression of miR167, miR319, and miR854, suggesting interplay of hormones and miRNAs in the regulation of As response. In conclusion, the present work demonstrates the role of miRNAs and associated mechanisms in the plant's response towards As stress.

Quantitative real-time expression profiling of aquaporins-isoforms and growth response of Brassica juncea under arsenite stress.

The present study analyzed the expression level of aquaporins of plasma membrane intrinsic protein (PIP) class in response to arsenite (AsIII) exposure of 100 µM from 0.5 h to 8 days in Brassica juncea. The expression levels of most of the PIPs were down-regulated during the course of AsIII exposure. This led to decrease in total water content of plants, which in turn hampered seedling growth. The level of reactive oxygen species (superoxide radicals and hydrogen peroxide), lipid peroxidation and root oxidizability increased significantly upon exposure to AsIII as compared to that of control leading to an increase in cell death. The study proposes that the down-regulation of PIPs happened presumably to regulate AsIII levels, which, however, occurred at the cost of reduced growth, disturbed water balance and induced oxidative stress.

Genome-wide analysis of thiourea-modulated salinity stress-responsive transcripts in seeds of Brassica juncea: identification of signalling and effector components of stress tolerance.

BACKGROUND AND AIMS: Abiotic stresses including salinity are the major constraints to crop production. In this regard, the use of thiourea (TU) in imparting salinity-stress tolerance to Indian mustard (Brassica juncea) has been demonstrated earlier. To gain an insight into the mechanism of TU action, various molecular and biochemical studies were conducted. METHODS: Microarray analysis was performed in seeds subjected to distilled water (control), 1 m NaCl, 1 m NaCl + 6·5 mm TU and 6·5 mm TU alone for 1 h. Real-time PCR validation of selected genes and biochemical studies were conducted under similar treatments at 1 h and 6 h. KEY RESULTS: The microarray analysis revealed a differential expression profile of 33 genes in NaCl- and NaCl + TU-treated seeds, most of which are established markers of stress tolerance. The temporal regulation of eight selected genes by real-time PCR indicated their early and co-ordinated induction at 1 h in NaCl + TU only. Besides, NaCl + TU-treated seeds also maintained a higher level of abscisic acid, reduced to oxidized glutathione (GSH : GSSG) ratio and activities of catalase, phenylalanine ammonia lyase and glutathione-S-transferases, as compared with that of NaCl treatment. The addition of LaCl(3) (a specific calcium-channel blocker) restricted the responses of TU both at molecular and biochemical level suggesting the possible involvement of a cytosolic calcium burst in the TU-mediated response. The TU-alone treatment was comparable to that of the control; however, it reduced the expression of some transcription factors and heat-shock proteins presumably due to the stabilization of the corresponding proteins. CONCLUSIONS: The TU treatment co-ordinately regulates different signalling and effector mechanisms at an early stage to alleviate stress even under a high degree of salinity. This also indicates the potential of TU to be used as an effective bioregulator to impart salinity tolerance under field conditions.

Buckling-driven morphological transformation of droplets of a mixed colloidal suspension during evaporation-induced self-assembly by spray drying.

Morphological transformation during evaporation-induced self-assembly of a mixed colloidal suspension in micrometric droplets has been investigated. It has been demonstrated that a buckling-driven shape transition of drying droplets of mixed colloidal suspension takes place during evaporation-induced self-assembly. Further, it is also shown that the distortion modulations get significantly amplified with enhancement in volume fraction of anisotropic soft colloidal component of the mixed colloids. It has been argued that the reduction in elastic modulus of formed shell, at the boundary of a drying droplet, and the anisotropic nature of one of the colloidal components facilitate the deformation process. Hierarchical structures of these assembled colloidal grains have been probed using electron microscopy and scattering techniques.

Survival of phosphate-solubilizing bacteria against DNA damaging agents.

Phosphate-solubilizing bacteria (PSBs) were isolated from different plant rhizosphere soils of various agroecological regions of India. These isolates showed synthesis of pyrroloquinoline quinone (PQQ), production of gluconic acid, and release of phosphorus from insoluble tricalcium phosphate. The bacterial isolates synthesizing PQQ also showed higher tolerance to ultraviolet C radiation and mitomycin C as compared to Escherichia coli but were less tolerant than Deinococcus radiodurans. Unlike E. coli, PSB isolates showed higher tolerance to DNA damage when grown in the absence of inorganic phosphate. Higher tolerance to ultraviolet C radiation and oxidative stress in these PSBs grown under PQQ synthesis inducible conditions, namely phosphate starvation, might suggest the possible additional role of this redox cofactor in the survival of these isolates under extreme abiotic stress conditions.

Effect of variable sulfur supply on arsenic tolerance and antioxidant responses in Hydrilla verticillata (L.f.) Royle.

In the present study, Hydrilla verticillata plants were exposed to arsenate (AsV; 50 microM) and arsenite (AsIII; 5 microM) under variable S supply: deficient (2 microM S, -S), normal (1 mM S, +S) and excess (2 mM S, +HS). Arsenic accumulation (microg g(-1) dw) in +HS plants was about 2-fold higher upon exposure to both AsV (30) and AsIII (50) than that observed in +S (12 & 24) and -S (14 & 26) plants. Despite lower As accumulation, -S plants experienced the maximum oxidative stress owing to an inadequate response of enzymatic and molecular antioxidants and significant decline in total thiols and the ratio of reduced glutathione (GSH) to oxidized glutathione (GSSG). By contrast +HS plants had significant increase in total thiols and an improved redox status, did not demonstrate any negative impact to antioxidants except catalase and hence experienced the least increase in oxidative stress parameters. In conclusion, an increase in S supply to plants may improve their accumulation capacity for As through enhanced tolerance caused by a positive effect on thiol metabolism and antioxidant status of the plants.

Comparative biochemical and transcriptional profiling of two contrasting varieties of Brassica juncea L. in response to arsenic exposure reveals mechanisms of stress perception and tolerance.

The mechanisms of perception of arsenic (As)-induced stress and ensuing tolerance in plants remain unresolved. To obtain an insight into these mechanisms, biochemical and transcriptional profiling of two contrasting genotypes of Brassica juncea was performed. After screening 14 varieties for As tolerance, one tolerant (TPM-1) and one sensitive (TM-4) variety were selected and exposed to arsenate [As(V)] and arsenite [As(III)] for 7 d and 15 d for biochemical analyses. The tolerant variety (TPM-1) demonstrated higher accumulation of As upon exposure to both 500 microM As(V) and 250 microM As(III) [49 microg g(-1) and 37 microg g(-1) dry weight (dw) after 15 d] as well as a better response of thiol metabolism as compared with the responses observed in the sensitive variety (TM-4). Transcriptional profiling of selected genes that are known to be responsive to sulphur depletion and/or metal(loid) stress was conducted in 15-d-old seedlings after 3 h and 6 h exposure to 250 microM As(III). The results showed an up-regulation of sulphate transporters and auxin and jasmonate biosynthesis pathway genes, whereas there was a down-regulation of ethylene biosynthesis and cytokinin-responsive genes in TPM-1 within 6 h of exposure to As(III). This suggested that perception of As-induced stress was presumably mediated through an integrated modulation in hormonal functioning that led to both short- and long-term adaptations to combat the stress. Such a coordinated response of hormones was not seen in the sensitive variety. In conclusion, an early perception of As-induced stress followed by coordinated responses of various pathways was responsible for As tolerance in TPM-1.

Thiourea modulates the expression and activity profile of mtATPase under salinity stress in seeds of Brassica juncea.

BACKGROUND AND AIMS: Large areas of the globe are becoming saline due to evapotranspiration and poor irrigation practices, and sustainability of agriculture is being seriously affected. Thiourea (TU) has been identified as an effective bioregulator imparting stress tolerance to crops. The molecular mechanisms involved in the TU-mediated response are considered in this study. METHODS: Differential display was performed in order to identify TU-modulated transcripts in Brassica juncea seeds exposed to various treatments (distilled water; 1 m NaCl; 1 m NaCl + 500 p.p.m. TU). The differential regulation of these transcripts was validated by quantitative real-time PCR. KEY RESULTS: Thiourea treatment maintained the viability of seeds exposed to NaCl for 6 h. Expression analysis showed that the transcript level of alpha, beta, gamma, delta and epsilon subunits of mitochondrial ATPase (mtATPase) varied in seeds subjected to the different treatments for 1 h: expression level was significantly altered by 1 m NaCl relative to controls; however, in the NaCl + TU treatment it reverted back in an integrated manner. Similar results were obtained from time-kinetics studies of beta and delta subunits in roots of 8-d-old seedlings. These observations were also confirmed by the mtATPase activity from isolated mitochondria. The reversal in the expression and activity profile of mtATPase through the application of a bioregulator such as TU is a novel finding for any plant system. CONCLUSIONS: The results suggest that TU treatment maintains the integrity and functioning of mitochondria in seeds as well as seedlings exposed to salinity. Thus, TU has the potential to be used as an effective bioregulator to impart salinity tolerance under field conditions, and might prove to be of high economic importance by opening new avenues for both basic and applied research.

Uranium and thorium sequestration by a Pseudomonas sp.: mechanism and chemical characterization.

The mechanism and chemical nature of uranium and thorium sequestration by a Pseudomonas strain was investigated by transmission electron microscopy, energy dispersive X-ray (EDX) analysis, FTIR spectroscopy and X-ray diffractometry. Atomic force microscopy (AFM) used in the tapping mode elucidated the morphological changes in bacterial cells following uranium and thorium binding. Transmission electron microscopy revealed intracellular sequestration of uranium and thorium throughout the cell cytoplasm with electron dense microprecipitations of accumulated metals. Energy dispersive X-ray analysis confirmed the cellular deposition of uranium and thorium. EDX and elemental analysis of sorption solution indicated the binding of uranium and thorium by the bacterial biomass via displacement of cellular potassium and calcium. The strong involvement of cellular phosphate, carboxyl and amide groups in radionuclide binding was ascertained by FTIR spectroscopy. X-ray powder diffraction (XRD) analyses confirmed cellular sequestration of crystalline uranium and thorium phosphates. Overall results indicate that a combined ion-exchange-complexation-microprecipitation mechanism could be involved in uranium and thorium sequestration by this bacterium. Atomic force microscopy and topography analysis revealed an undamaged cell surface with an increase in cell length, width and height following radionuclide accumulation. The arithmetic average roughness (R(a)) and root mean square (RMS) roughness (R(q)) values indicated an increase in surface roughness following uranium and thorium sequestration.

Potential of Chromolaena odorata for phytoremediation of (137)Cs from solution and low level nuclear waste.

Potential of Chromolaena odorata plants for remediation of (137)Cs from solutions and low level nuclear waste was evaluated. When plants were exposed to solutions spiked with three different levels of (137)Cs, namely 1 x 10(3) kBqL(-1), 5 x 10(3) kBqL(-1) and 10 x 10(3) kBqL(-1), 89%, 81% and 51% of (137)Cs was found to be remediated in 15 d, respectively. At the lowest Cs activity (1 x 10(3) kBqL(-1)), accumulation of Cs was found to be higher in roots compared to shoots, while at higher Cs activities (5 x 10(3) kBqL(-1) and 10 x 10(3) kBqL(-1)), Cs accumulation was more in shoots than roots. When plants were incubated in low level nuclear waste, 79% of the activity was removed by plants at the end of 15 d. The present study suggests that C. odorata could be used as a potential candidate plant for phytoremediation of (137)Cs.

Removal of copper ions by the filamentous fungus, Rhizopus oryzae from aqueous solution.

Removal of heavy metals present in wastewaters has been a major concern due to their non-biodegradability and toxicity. Removal of copper ion using NaOH treated Rhizopus oryzae biomass was investigated in a batch reactor. The copper uptake exhibited substantial enhancement both in terms of kinetics of uptake as well as the loading capacity. The copper biosorption by viable and pretreated fungal biomass fit well to a Lagergren's pseudo second order reaction in comparison to pseudo first order kinetics. Investigation on effect of pH indicated improved performance in the range of pH 4-6 in alkali treated biomass. Copper uptake exhibited by viable biomass was highest at 21 degrees C, unlike pretreated biomass that showed maximum uptake across the range of temperature 21-55 degrees C. The maximum copper loading capacity of the viable and pretreated biomass according to Langmuir isotherm was 19.4 and 43.7 mg/g, respectively. Distribution coefficient of pretreated biomass showed improvement at lower residual concentration, indicating a change in the nature of binding by the treated biomass. Copper uptake decreased with an increasing dose of biosorbent, although enhancement in the total metal ion removal was observed at higher dose.

Potential of vetiver (Vetiveria zizanoides L. Nash) for phytoremediation of phenol.

Aseptically grown Vetiveria zizanoides were evaluated for their potential for phytoremediation of phenol from Murashige and Skoog's liquid medium. Phenol was found to be completely removed from incubation medium at the end of 4 days by V. zizanoides plantlets, when medium was supplemented with 50 and 100 mg L(-1) phenol, while with 200, 500, and 1000 mg L(-1) of phenol, 89%, 76% and 70%, respectively, were removed. Phenol removal was found to be associated with inherent production of peroxidase and hydrogen peroxide. Coupled with H(2)O(2) formation, the levels of antioxidant enzymes like superoxide dismutase and peroxidase showed an enhancement when plants were exposed to phenol, whereas catalase levels initially showed a decline due to the utilization of H(2)O(2) by peroxidase for phenol oxidation. However, when peroxidase levels declined, there was an enhancement in catalase levels to minimize the presence of H(2)O(2) in the medium. Having confirmed that the removal of phenol was by V. zizanoides plantlets, in the next phase, micropropagated plantlets and well-developed plants grown in hydroponics were used under in vivo conditions to study the effect of phenol (200 mg L(-1)) on plant growth and reuse. Although plant growth was reduced in presence of phenol, the results of the reuse study indicated the possibility of plants getting adapted to phenol without any decline in potential for phenol remediation.

Biosorption characteristics of uranium(VI) from aqueous medium onto Catenella repens, a red alga.

The biosorption characteristics of uranium(VI) onto Catenella repens (a red alga), were evaluated as a function of pH, biosorbent size, time, biomass dosage, initial uranium concentration and temperature. Within the pH range studied (1.5-7.5), 4.5 was the optimum pH for the uptake of uranium(VI) by C. repens. Reduction in particle size did not increase the biosorption capacity. The metal removal was rapid, with more than 90% of total biosorption taking place in 30 min, and equilibrium was attained in 45 min. The maximum metal loading capacity of the alga was 303 mg/g. Within the temperature range studied (15-55 degrees C), there was no significant change in biosorption, under optimal conditions. Adsorption process could be well defined by both the Langmuir and Freundlich isotherms with r(2) of 0.94 and 0.96, respectively. The kinetic data fitted the pseudo-second-order kinetic model with the r(2) value of 0.99. At a low pH of 2.5, where most of the biomasses show either no or less metal uptake, a good (>15%) metal loading capacity of 25% was achieved. Therefore biosorption characteristics were also evaluated at pH 2.5.

Advances in development of transgenic plants for remediation of xenobiotic pollutants.

Phytoremediation-the use of plants for cleaning up of xenobiotic compounds-has received much attention in the last few years and development of transgenic plants tailored for remediation will further enhance their potential. Although plants have the inherent ability to detoxify some xenobiotic pollutants, they generally lack the catabolic pathway for complete degradation/mineralization of these compounds compared to microorganisms. Hence, transfer of genes involved in xenobiotic degradation from microbes/other eukaryotes to plants will further enhance their potential for remediation of these dangerous groups of compounds. Transgenic plants with enhanced potential for detoxification of xenobiotics such as trichloro ethylene, pentachlorophenol, trinitro toluene, glycerol trinitrate, atrazine, ethylene dibromide, metolachlor and hexahydro-1,3,5-trinitro-1,3,5-triazine are a few successful examples of utilization of transgenic technology. As more genes involved in xenobiotic metabolism in microorganisms/eukaryotes are discovered, it will lead to development of novel transgenic plants with improved potential for degradation of recalcitrant contaminants. Selection of suitable candidate plants, field testing and risk assessment are important considerations to be taken into account while developing transgenic plants for phytoremediation of this group of pollutants. Taking advantage of the advances in biotechnology and 'omic' technologies, development of novel transgenic plants for efficient phytoremediation of xenobiotic pollutants, field testing and commercialization will soon become a reality.

Electrochemical biosensor for catechol using agarose-guar gum entrapped tyrosinase.

An electrochemical biosensor using tyrosinase was constructed for the determination of catechol. The enzyme was extracted from a plant source Amorphophallus companulatus and entrapped in agarose-guar gum composite biopolymer matrix. Catechol was determined by direct reduction of biocatalytically liberated quinone species at -0.1 V versus Ag/AgCl (3M KCl). The response was found to be linear and concentration dependent in the range of 6 x 10(-5) to 8 x 10(-4)M with a lower detection limit of 6 microM. It has reusability up to 20 cycles and a shelf life of more than 2 months when stored at 4 degrees C.

Isolation of a starch utilizing, phosphate solubilizing fungus on buffered medium and its characterization.

A phosphate solubilizing fungus, Paecilomyces marquandii AA1 was isolated from phosphate deficient soil on Pikovskaya's medium buffered with Tris-HCl pH 8. The organism was identified on the basis of morphological characterization and by sequencing of 18S rRNA gene. The organism could release phosphate from both buffered and unbuffered medium and solubilized rock phosphates from various places. The effect of concentration of ore, temperature, carbon and nitrogen sources on solubilization of rock phosphate was studied. Ammonium salts were the best nitrogen source, followed by asparagine, sodium nitrate, potassium nitrate, urea and calcium nitrate in that order.

Optical microbial biosensor for detection of methyl parathion pesticide using Flavobacterium sp. whole cells adsorbed on glass fiber filters as disposable biocomponent.

An optical microbial biosensor was described for the detection of methyl parathion pesticide. Whole cells of Flavobacterium sp. were immobilized by trapping in glass fiber filter and were used as biocomponent along with optic fiber system. Flavobacterium sp. has the organophosphorus hydrolase enzyme, which hydrolyzes the methyl parathion into detectable product p-nitrophenol. The immobilized microbial biocomponent was disposable, cost-effective and showed high reproducibility and uniformity. The detection of methyl parathion by the use of disposable microbial biocomponent with optical biosensor was simple, single step and direct measurement of very low quantity of the sample. The home made reaction vessel was small and needed only 75 microl of sample. A lower detection limit 0.3 microM methyl parathion was estimated from the linear range (4-80 microM) of calibration plot of organophosphorus hydrolase enzymatic assay. The applicability to synthetic methyl parathion spiked samples was demonstrated.

Phenol removal using Brassica juncea hairy roots: role of inherent peroxidase and H(2)O(2).

Removal of phenol, a major pollutant in aqueous effluents was studied using plant hairy root cultures. Among four different species of hairy roots tested, Brassica juncea showed the highest potential for phenol removal. The effect of phenol concentration and reuse in a batch system was studied using B. juncea hairy root cultures. Unlike most of the studies reported earlier, phenol removal by the hairy roots was seen to take place without the need for addition of external hydrogen peroxide (H(2)O(2)). To understand the mechanism of phenol removal, levels of peroxidase and phenol oxidase were monitored in the hairy roots. Peroxidase activity in the roots was enhanced when exposed to phenol, while phenol oxidase remained constant. Since peroxidase has a pre-requisite for H(2)O(2), the levels of H(2)O(2) were monitored for its in situ synthesis. H(2)O(2) levels were seen to increase in the presence of phenol. Thus, a mechanism wherein hairy roots also produce H(2)O(2) besides peroxidase, as a protection strategy of plant against xenobiotic stress is plausible.

Heavy metal pollution abatement using rock phosphate mineral.

The low-grade rock phosphate of Jhabua, Madhya Pradesh (India), was investigated for its possible application in the removal of lead, copper, zinc and cobalt ions from aqueous solutions. Effects of contact time, amount of adsorbent and initial concentration of metal ions were studied. Adsorption of heavy metal ions was found to follow the order: Pb2+ > Cu2+ > Zn2+ > Co2+. The probable mechanism of metal ions removal by rock phosphate was found to be by its dissolutions followed by subsequent precipitation.