Phytoextraction of Heavy Metals Induced by Bioaugmentation of a Phosphate Solubilizing Bacterium

Arunakumara, K.K.I.U.,Walpola, Buddhi Charana,Song, Jun-Seob,Shin, Min-Jung,Lee, Chan-Jung,Yoon, Min-Ho The Korean Society of Environmental Agriculture 2014 한국환경농학회지 Vol.33 No.3

BACKGROUND: Excessive metals in the soil have become one of the most significant environmental problems. Phytoremediation has received considerable attention as a method for restoring the contaminated soils. The microbes having remarkable metal tolerance and plant growth-promoting abilities could also play a significant role in remediation of metal-contaminated soils, because bioaugmentation with such microbes could promote phytoextraction of metals. Therefore, the present study was focused on evaluating the phytoextraction of heavy metals (Co, Pb and Zn) in Helianthus annuus (sunflower) induced by bioaugmentation of a phosphate solubilizing bacterium. METHODS AND RESULTS: A phosphate solubilizing bacterium was isolated from metal-contaminated soils based on the greater halo size (>3 mm) with solid NBRIP agar medium containing 10 g glucose, 5 g $Ca_3(PO_4)_2$, 5 g $MgCl_2{\cdot}6H_2O$, 0.25 g $MgSO_4.7H_2O$, 0.2 g KCl, 0.1 g $(NH_4)_2SO_4$ in 1 L distilled water. Isolated bacterial strain was assessed for their resistance to heavy metals; $CoCl_2.6H_2O$, $2PbCO_3.Pb(OH)_2$, and $ZnCl_2$ at various concentrations ranging from $100-400{\mu}g/mL$ (Co, Pb and Zn) using the agar dilution method. A pot experiment was conducted with aqueous solutions of different heavy metals (Co, Pb and Zn) to assess the effect of bacterial strain on growth and metal uptake by Helianthus annuus (sunflower). The impact of bacterial inoculation on the mobility of metals in soil was investigated under laboratory conditions with 50 mL scaled polypropylene centrifuge tubes. The metal contents in the filtrate of plant extracts were determined using an atomic absorption spectrophotometer (Perkinelmer, Aanalyst 800, USA). CONCLUSION: Inoculation with Enterobacter ludwigii PSB 28 resulted in increased shoot and root biomass and enhanced accumulation of Co, Pb and Zn in Helianthus annuus plants. The strain was found to be capable of promoting metal translocation from the roots to the shoots of H. annuus. Therefore, Enterobacter ludwigii PSB 28 could be identified as an effective promoter of phytoextraction of Co, Pb and Zn from metal-contaminated soils.

Alleviation of Phyto-toxicity of Copper on Agricultural Plants

K.K.I.U. Arunakumara,윤민호,Buddhi Charana Walpola 한국응용생명화학회 2013 Applied Biological Chemistry (Appl Biol Chem) Vol.56 No.5

Copper (Cu), a redox-active transition metal, is known to be involved in protein metabolism, photosynthetic and respiratory electron transport, cell wall metabolism, antioxidant activity,nitrogen fixation, ion metabolization, and hormone perception,among others in plants. Though Cu has been listed among the essential elements, it could potentially result in complete inhibition of plant growth and development at excess concentrations. Measures available for alleviating Cu toxicity in plants are discussed in the present paper. Exogenous application of nitric oxide through up-regulating the components of antioxidant defense system [catalase (CAT-EC 1.11.1.6), peroxidase (POD-EC 1.11.1.7), superoxide dismutase (SOD-EC 1.15.1.1), and ascorbate peroxidase (APX-EC 1.11.1.11) activities] and stimulating the enzyme P5CS (D1-pyrroline-5-carboxylate synthetase), which catalyzes proline biosynthesis, has been proved to stand against the adverse impacts of Cu toxicity. Addition of cations (such as Ca2+ and Mg2+) through stimulating site-specific competition for metal ions could also prevent excess accumulation of Cu in cell interior. Silicon application, through nutrient balancing and physically blocking the apoplastic bypass flow has also been recognized to be effective in alleviating Cu toxicity. Addition of organic amendments and use of arbuscular mycorrhizal fungi as soil inoculants have also proved successful in amelioration of Cucontaminated soils. Though molecular and physiological mechanisms associated with Cu toxicity have been substantially investigated, information on the regulation of the expression of stress-related genes in key agricultural plant species is still lacking. Additional research efforts focusing at field validation of the toxicity alleviation methods are also equally important.

How do Citrus Crops Cope with Aluminum Toxicity?

Arunakumara, K.K.I.U.,Walpola, Buddhi Charana,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2012 한국토양비료학회지 Vol.45 No.6

World Agriculture faces daunting challenges in feeding the growing population today. Reduction in arable land extent due to numerous reasons threatens achievement of food and nutritional security. Under this back ground, agricultural use of acidic soils, which account for approximately 40 % of the world arable lands is of utmost important. However, due to aluminum (Al) toxicity and low available phosphorous (P) content, crop production in acidic soils is restricted. Citrus, in this context, gains worldwide recognition as a crop adapted to harsh environments. The present paper reviewed Al toxicity and possible toxicity alleviation tactics in citrus. As reported for many other crops, inhibition of root elongation, photosynthesis and growth is experienced in citrus also due to Al toxicity. Focusing at toxicity alleviation, interaction between boron (B) and Al as well as phosphorus and Al has been discussed intensively. Al toxicity in citrus could be alleviated by P through increasing immobilization of Al in roots and P level in shoots rather than through increasing organic acid secretion, which has been widely reported in other crops. Boron-induced changes in Al speciation and/or sub-cellular compartmentation has also been suggested in amelioration of root inhibition in citrus. Despite the species-dependent manner of response to Al toxicity, many commercially important citrus species can be grown successfully in acidic soils, provided toxicity alleviation Agro-biological tactics such as addition of phosphorous fertilizers are used properly.

Pterocarpus santalinus Linn. f. (Rath handun): A Review of Its Botany, Uses, Phytochemistry and Pharmacology

Kodithuwakku K. I. U.i Arunakumara,Siripala Subasinghe,윤민호 한국응용생명화학회 2011 Applied Biological Chemistry (Appl Biol Chem) Vol.54 No.4

Pterocarpus santalinus Linn.f., commonly known as Red sanders, belongs to the family Fabaceae. It is endemic to India and considered globally endangered, with illegal harvest being a key threat. The plant is renowned for its characteristic timber of exquisite color, beauty, and superlative technical qualities. The red wood yields a natural dye santalin, which is used in coloring pharmaceutical preparations and foodstuffs. In the traditional system of medicine, the decoction prepared from the heartwood is attributed various medicinal properties. It has been used in inducing vomiting and treating eye diseases, mental aberrations, and ulcers. The heartwood of Red sanders is known to have antipyretic, anti-inflammatory, anthelmintic, tonic, hemorrhage, dysentery, aphrodisiac, and diaphoretic activities. It has also been used as a cooling agent. Ethanol extract of stem bark was reported to possess anti-hyperglycaemic activity. The wood in combination with other drugs is also prescribed for snake bites and scorpion stings. Phytochemical investigations of aqueous and ethanol extracts of stem bark revealed the presence of alkaloids, phenols, saponins, glycosides, flavonoides, triterpenoides, sterols, and tannins. The heart wood contains isoflavone glucosides and two anti-tumour lignans, viz., savinin and calocedrin. However, the species has remained unexplored for many pharmacological activities claimed. The present paper reviewed botany, uses, phytochemistry, and pharmacology of P. santalinus.

Evaluation of Nitrogen Mineralization in Soil Polluted by Zinc and Cadmium

Buddhi Charana Walpola,K.K.I.U. Arunakumara,윤민호 한국토양비료학회 2011 한국토양비료학회지 Vol.44 No.4

Soil microbial functions are considered to be effective in assessing the severity of heavy metal pollution. Therefore, this study was carried out to examine the effect of heavy metals on nitrogen mineralization by measuring the releasing pattern of inorganic nitrogen (NH_4^+-N and NO_3^--N) in a soil treated with heavy metals. A factorial combination of two heavy metals (Zn and Cd) treated with three concentrations (50, 100and 150 μmol g^-1 soils) was used in a laboratory incubation. Nitrogen mineralization was determined at 3, 7,14, 21, 28, 42 and 56 days after the treatments replicated four times. Soil sample free from heavy metals was served as the control. The amount of nitrogen mineralization from heavy metal treated soils was found to be decreased at an increasing rate during the first 21 days of incubation. However, as the incubation progressed,nitrogen mineralization was found to be decreased at decreasing rates. Furthermore, during this period,nitrogen mineralization in Cd treated soils was significantly lower (P≤0.05) than that of the control. Soils treated with Cd at the concentration of 150 μmol g^-1 showed the lowest N mineralization throughout the incubation. Nitrogen mineralization in Zn treated soils (50 μmol g^-1) was found to be higher than the other heavy metal treated soils. On the base of present findings, nitrogen mineralization of soil could be considered as a viable assessment of the degree of heavy metal pollution.

Mobilization of Heavy Metals in Contaminated Soils induced by Bioaugmentation of Shewanella xiamenensis HM14

Walpola, Buddhi Charana,Arunakumara, K.K.I.U.,Song, Jun-Seob,Lee, Chan-Jung,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2014 한국토양비료학회지 Vol.47 No.4

A bacterial strain with the potential ability to solubilize heavy metals was isolated from heavy metal contaminated soils collected from abandoned mines of Boryeong area in South Korea. The bacterial strain with the highest degree of metal resistance was shown to have close proximity with Shewanella xiamenensis FJ589031, according to 16S rRNA sequence analysis, and selected for investigating the mobilization of metals in soil or plant by the strain. The strain was found to be capable of solubilizing metals both in the absence and in the presence of metals (Co, Pb and Cd). Metal mobilization potential of the strain was assessed in a batch experiment and the results showed that inoculation could increase the concentrations of water soluble Co, Pb and Cd by 48, 34 and 20% respectively, compared with those of non-inoculated soils. Bacterial-assisted growth promotion and metal uptake in sunflower (Helianthus annuus) was evaluated in a pot experiment. In comparison with non-inoculated seedlings, the inoculation led to increase the growth of H. annuus by 24, 18 and 16% respectively in Co, Pb and Cd contaminated soils. Moreover, enhanced accumulation of Co, Pb and Cd in the shoot and root systems was observed in inoculated plants, where metal translocation from root to the above-ground tissues was also found to be enhanced by the strain. Plant growth promotion and metal mobilizing potential of the strain suggest that the strain could effectively be employed in enhancing phytoextraction of Co, Pb and Cd from contaminated soils.

Evaluation of Nitrogen Mineralization in Soil Polluted by Zinc and Cadmium

Walpola, Buddhi Charana,Arunakumara, K.K.I.U.,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2011 한국토양비료학회지 Vol.44 No.4

Soil microbial functions are considered to be effective in assessing the severity of heavy metal pollution. Therefore, this study was carried out to examine the effect of heavy metals on nitrogen mineralization by measuring the releasing pattern of inorganic nitrogen ($NH_4^+$-N and $NO_3^-$-N) in a soil treated with heavy metals. A factorial combination of two heavy metals (Zn and Cd) treated with three concentrations (50, 100 and $150{\mu}mol\;g^{-1}$ soils) was used in a laboratory incubation. Nitrogen mineralization was determined at 3, 7, 14, 21, 28, 42 and 56 days after the treatments replicated four times. Soil sample free from heavy metals was served as the control. The amount of nitrogen mineralization from heavy metal treated soils was found to be decreased at an increasing rate during the first 21 days of incubation. However, as the incubation progressed, nitrogen mineralization was found to be decreased at decreasing rates. Furthermore, during this period, nitrogen mineralization in Cd treated soils was significantly lower ($P{\leq}0.05$) than that of the control. Soils treated with Cd at the concentration of $150{\mu}mol\;g^{-1}$ showed the lowest N mineralization throughout the incubation. Nitrogen mineralization in Zn treated soils ($50{\mu}mol\;g^{-1}$) was found to be higher than the other heavy metal treated soils. On the base of present findings, nitrogen mineralization of soil could be considered as a viable assessment of the degree of heavy metal pollution.

Optimum Conditions for Transformation of Synechocystis sp. PCC 6803

Xiaonan Zang,Bin Liu,Shunmei Liu,K.K.I.U. Arunakumara,Xuecheng Zhang 한국미생물학회 2007 The journal of microbiology Vol.45 No.3

This study was conducted to determine the optimal conditions for introduction of exogenous DNA into Synechocystis sp. PCC 6803. Of the three transformation techniques studied, electroporation, ultrasonic transformation and natural transformation, natural transformation showed the highest efficiency. Additionally, this study demonstrated that the higher plasmid concentration and longer homologous recombining fragments resulted in a greater number of transformants. For successful transformation, the lowest concentration of plasmid was 0.02 μg/ml, and the shortest homologous recombining fragment was 0.2 kb. Use of Synechocystis sp. PCC 6803 in the logarithmic growth phase resulted in two-fold higher transformation rate than that of the same organism when cells in the latent phase or the plateau phase were used for transformation. Pretreatment of the host strain, Synechocystis sp. PCC 6803, with EDTA (2 mM) for two days prior to transformation increased the transformation efficiency by 23%. Additionally, incubation of the cells and DNA for 5 h under light conditions increased the transformation efficiency by two orders of magnitude. Moreover, recovery treatment of the cells before they were plated onto antibiotic medium also increased the transformation efficiency.

Optimum Conditions for Transformation of Synechocystis sp. PCC 6803

Zang, Xiaonan,Liu, Bin,Liu, Shunmei,Arunakumara, K.K.I.U.,Zhang, Xuecheng The Microbiological Society of Korea 2007 The journal of microbiology Vol.45 No.3

This study was conducted to determine the optimal conditions for introduction of exogenous DNA into Synechocystis sp. PCC 6803. Of the three transformation techniques studied, electroporation, ultrasonic transformation and natural transformation, natural transformation showed the highest efficiency. Additionally, this study demonstrated that the higher plasmid concentration and longer homologous recombining fragments resulted in a greater number of transformants. For successful transformation, the lowest concentration of plasmid was $0.02\;{\mu}g/ml$, and the shortest homologous recombining fragment was 0.2 kb. Use of Synechocystis sp. PCC 6803 in the logarithmic growth phase resulted in two-fold higher transformation rate than that of the same organism when cells in the latent phase or the plateau phase were used for transformation. Pretreatment of the host strain, Synechocystis sp. PCC 6803, with EDTA (2 mM) for two days prior to transformation increased the transformation efficiency by 23%. Additionally, incubation of the cells and DNA for 5 h under light conditions increased the transformation efficiency by two orders of magnitude. Moreover, recovery treatment of the cells before they were plated onto antibiotic medium also increased the transformation efficiency.