Concentration Dependent Effect of Heavy Metals on Soil Carbon Mineralization

Walpola, Buddhi Charana,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2012 한국토양비료학회지 Vol.45 No.4

The present laboratory investigation was conducted to assess the effect of heavy metals on carbon mineralization. Soil was treated with three concentrations (50, 100 and $150{\mu}mol\;g^{-1}$ soil) of two heavy metals (Cd and Zn) in a factorial combination of treatments replicated four times. Determination of carbon mineralization was carried out at 3, 7, 14, 21, 28, 42 and 56 days after metal treatments.. The amount of $CO_2$-C released from heavy metal treated soils was found to be decreased at an increasing rate during the first 28 days, followed by slow release as incubation progressed. The total amounts of $CO_2$-C released were 448, 382 and $348mg\;kg^{-1}$ soil respectively for soils treated with 50, 100 and $150{\mu}mol\;g^{-1}$ soil of Zn. The corresponding figures for Cd treated soils were 406, 354 and $282mg\;kg^{-1}$ soil implying that dose-dependent reduction in cumulative $CO_2$-C released from soils. The inhibition of carbon mineralization was found to be high in Cd treated soils than that of Zn treated. Therefore, tolerance and adaptation of the microbial community is likely to be related to the concentration and the type of metal. According to the results, carbon mineralization can be considered as possible indicator of soil pollution by means of heavy metals.

Stress Induced Phosphate Solubilization by Aspergillus awamori bxq33110 Isolated from Waste Mushroom Bed of Agaricus bisporus

Walpola, Buddhi Charana,Song, June-Seob,Jang, Kab-Yeul,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2012 한국토양비료학회지 Vol.45 No.3

A fungal strain, capable of solubilizing insoluble phosphate under diverse temperature, pH and salt conditions was isolated from Waste Mushroom bed of Agaricus bisporus in South Korea. Based on 18S rRNA analysis, the strain was identified as Aspergillus awamori bxq33110. The strain showed maximum phosphate solubilization in AYG medium (525 ${\mu}g\;mL^{-1}$) followed by NBRIP medium (515 ${\mu}g\;mL^{-1}$). The strain solubilized $Ca_3(PO_4)_2$ to a greater extent and rock phosphate and $FePO_4$ to a certain extent. However $AlPO_4$ solubilizing ability of the strain was found to be very low. Glucose at the rate of 2% ($561{\mu}g\;mL^{-1}$) was found be the best carbon source for Aspergillus awamori bxq33110 to solubilize maximum amount of phosphate. However, no significant difference ($P{\leq}0.05$) in phosphorus solubilization was found between 1% and 2% glucose concentrations. $(NH_4)_2SO_4$ was the best nitrogen source for Aspergillus awamori bxq33110 followed by $NH_4Cl$ and $NH_4NO_3$. At pH 7, temperature $30^{\circ}C$ and 5% salt concentration (674 ${\mu}g\;mL^{-1}$) were found to be the optimal conditions for insoluble phosphate solubilization. However, strain Aspergillus awamori bxq33110 was shown to have the ability to solublize phosphate under different stress conditions at $30-40^{\circ}C$ temperature, pH 7-10 and 0-10% salt concentrations indicating it's potential to be used as bio-inoculants in different environmental conditions.

Assessment of Plant Growth Promoting Activities of Phosphorus Solubilizing Bacteria

Walpola, Buddhi Charana,Song, June-Seob,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2012 한국토양비료학회지 Vol.45 No.1

Plant growth promoting traits like production of indoleacetic acid (IAA), ammonia, hydrogen cyanide (HCN), siderophore, and like the enzyme activities of catalase, ACC deaminase, cellulase, chitinase and protease were assayed in vitro for twenty one phosphorus solubilizing bacteria isolated from soil isolates. Except SPP-5 and SPP-15 strains, all the other isolated strains produced IAA in various amounts of 10 to $23{\mu}g\;ml^{-1}$. All strains showed positive response for ammonia production and ACC deaminase activity implying that they are capable of growing in a N-free basal medium. Catalase activity was found to be superior in SPP-2, SPP-7, SPP-12 and SPP-17 compared to the other strains tested. HCN production was detected by 15 strains and among them SPP-9, SPP-15, SAph-11, and SAph-24 were found to be strong HCN producers. Except the isolates SPP-10, SPP-12, SPP-13 and SPP-14, all the other isolates produced more than 80% siderophore units. None of the strains showed cellulose and chitinase activity. SAph-8, SAPh-11, SAPh-24 and SPP-15 strains showed 35.84, 50.33, 56.64 and 34.78 U/ml protease activities, respectively. SPP-1, SPP-2, SPP-3, SPP-11, SPP-17, SPP-18, SAph-11 and SAph-24 strains showed positive response for all the tested plant growth promotion traits except cell wall degrading enzyme activities. According to the results, all the tested phosphorus solubilizing isolates could exhibit more than three or four plant growth promoting traits, which may promote plant growth directly or indirectly or synergistically. Therefore, these phosphorus solubilizing strains could be employed as bio-inoculants for agriculture soils.

Application of Immobilization Technology in Solubilization of Rock Phosphate

Walpola, Buddhi Charana,Kim, Ah Young,Jeon, Ju Hyeon,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2014 한국토양비료학회지 Vol.47 No.4

Phosphates solubilizing bacterial strains belong to Pantoea, Burkholderia and Enterobacter were isolated and employed in assessing their solubilization ability of Ca phosphate and ER phosphate (Eppawala Rock Phosphate). Among the bacterial strains used, PSB-13 (Pantoea rodasii) showed higher Ca-phosphate solubilization ($1100{\mu}g\;ml^{-1}$) as well as rock phosphate solubilization ($168{\mu}g\;ml^{-1}$). The strain was then immobilized in agar to further assess its phosphate solubilization ability. According to the results, agar encapsulated strain solubilized 0.3%, 7.31%, 20.24%, and 20.62% more Ca-phosphate and 11.53%, 15.29%, 28.48%, 36.55% (respectively in 4 cycles) more ER-phosphate than free cells. The reuse efficiency of agar entrapped bacterial cells for Ca-phosphate and ER-phosphate solubilization was greater than that by freely suspended bacterial cells. In conclusion, immobilization could enhance the phosphate solubilization capacity of the strains and thus could be used effectively in enhancing solubilization of ER phosphate.

Influence of Different pH Conditions and Phosphate Sources on Phosphate Solubilization by Pantoea agglomerans DSM3493

Walpola, Buddhi Charana,Keum, Mi-Jung,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2012 한국토양비료학회지 Vol.45 No.6

Pantoea agglomerans DSM3493 was isolated from green house soils collected from Chungchugnam-do province, Gongju-Gun area in South Korea and phosphate solubilization and organic acid production of the strain were assessed using three types of insoluble phosphate sources (Ca phosphate, Fe phosphate and Al phosphate) under three different pH conditions (7, 8 and 9). The highest Ca phosphate solubilization ($651{\mu}g\;mL^{-1}$) was recorded at pH 7 followed by pH 8 and 9 (428 and $424{\mu}g\;mL^{-1}$ respectively). The solubilization rate was found to be 80.4, 98.1 and $88.7{\mu}g\;mL^{-1}$ (for Fe phosphate containing medium) and 9.3, 12.1 and $29.8{\mu}g\;mL^{-1}$ (for the Al phosphate containing medium) respectively at pH 7, 8 and 9. Though increasing pH of the medium caused reduction in the rate of solubilization of Ca phosphate, solubilization of Fe and Al phosphates enhanced with increasing pH. By contrast, the highest amount of organic acid was produced with Ca phosphate while the lowest was recorded with the presence of Al phosphate. Among the organic acids, gluconic acid production was found to be the highest, followed by oxalic acid and citric acid regardless the source of phosphate. Results can thus be concluded that the production of organic acids appears to play a significant role in the inorganic phosphate solubilization.

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.

Solubilization of Inorganic Phosphates and Plant Growth Promotion by Pantoea Strains

Walpola, Buddhi Charana,Kong, Won-Sik,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2013 한국토양비료학회지 Vol.46 No.6

Two phosphate solubilizing Pantoea strains (P. agglomerans and P. rodasii) were employed in elucidating their phosphate solubilizing potential under different carbon and nitrogen sources, pH, temperature and salt conditions. Plant growth promoting characteristics such as ACC deaminase activity, indole acetic acid (IAA), HCN, ammonia, and siderophore production of the two strains were assessed in vitro. Potential applicability of the strains as bio-inoculants was also evaluated in pot experiments conducted under green house conditions. Phosphate solubilization measured as the amount of phosphorous released into the medium was recorded as 810 and $788{\mu}g\;ml^{-1}$ respectively by P. agglomerans and P. rodasii. Glucose at the rate of 2% was found be the best carbon source, while $(NH_4)_2SO_4$ was the best nitrogen source for both strains. Despite a slight decrease in phosphate solubilization observed at higher temperature, pH and salt concentrations, both strains could withstand against a range of temperature ($30-35^{\circ}C$), pH (7-9) and the presence of NaCl (up to 5%) without much compromising the phosphate solubilization. Different plant growth promoting traits (ACC deaminase activity, IAA, HCN, ammonia, and siderophore production) of the strains and their ability to promote the growth of green gram seedlings indicate that both strains possess high potential to be used as bio-inoculants.

Effect of Fungicides on Phosphate Solubilization by Klebsiella oxytoca and Enterobacter ludwigii

Walpola, Buddhi Charana,Keum, Mi-Jung,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2013 한국토양비료학회지 Vol.46 No.2

The aim of the present study was to isolate phosphate solubilizing bacteria (PSB) and to assess their potential tolerance to fungicides. Out of thirty PSB, two strains Klebsiella oxytoca and Enterobacter ludwigii were selected on the basis of their tolerance to fungicides. Both strains were assessed for their phosphate solubilizing ability using three different fungicides (difenoconazole, fluazinam and streptomycin) each with the concentrations of 0, 1, 2 or 3 times of the recommended rate. Both strains showed increased phosphate solubilization with difenoconazole at 1, 2 and 3 times of the recommended rate as compared to the phosphate solubilization of the control. The phosphate solubilization in Klebsiella oxytoca was recorded as 326, 538, 518 and 481 ${\mu}g\;mL^{-1}$ at 0, 1, 2 and 3 times of the recommended rate respectively, whereas in Enterobacter ludwigii it was recorded as 395, 499, 529 and 533 ${\mu}g\;mL^{-1}$ respectively at various doses. Based on the present findings, it may be concluded that both strains have the potential to be used as bio-inoculants which can solubilize phosphate even at the higher doses as compared to the recommended rate of fungicides.

Synergistic Phosphate Solubilization by Burkholderia anthina and Aspergillus awamori

Walpola, Buddhi Charana,Jang, Hyo-Ju,Yoon, Min-Ho Korean Society of Soil Science and Fertilizer 2013 한국토양비료학회지 Vol.46 No.2

Single or co-inoculation of phosphate solubilizing bacterial and fungal strains (Burkholderia anthina and Aspergillus awamori respectively) was performed separately to assess their synergistic and antagonistic interactions and the potential to be used as bio-inoculants. Co-inoculation was found to release the highest content of soluble phosphorus (1253 ${\mu}g\;ml^{-1}$) into the medium, followed by single inoculation of fungal strain (1214 ${\mu}g\;ml^{-1}$) and bacterial strain (997 ${\mu}g\;ml^{-1}$). However, there was no significant difference between single inoculation of fungal strain and co-inoculation of fungal and bacterial strain in terms of the phosphorous release. The highest pH reduction, organic acid production and glucose consumption were observed in the sole A. awamori inoculated culture medium. According to the plant growth promotion bioassays, co-inoculation of the microbial strains resulted in 21% and 43% higher shoot and root growth of the mung bean seedlings respectively as compared to the respective controls. Therefore, co-inoculation of B. anthina and A. awamori showed better performance in stimulating plant growth than that in inoculation of each strain alone. However, assessment period of the present study being short, we recommend in engaging further experimentation under field conditions in order to test the suitability of the strains to be used as bio-inoculants.