Study On Dissimilatory Reduction Of Iron Oxides In Paddies And Its Influential Factors

Dissimilatory Iron Reduction (DIR), a kind of metabolism of microorganism, and Fe(III) as the terminal electron acceptor was reduced to Fe(II) in this process, coupled with the oxidation of organo/abio electron donors. DIR occurs generally in the natural anaerobic environment, where Dissimilatory Iron Reduction Bacteria (DIRB) exists universally. But, it was not focused on till last decades. Hypothetically, it is the first form of microbial respiration. Since it could be inherited in some degree in the evolution process, there must be much more microbial are capable to dissimilatory reduce Fe(III). Therefore, investigations focused on DIR process could help better understanding the evolution process and the energy metabolism in early stages, theoretically. Redox of iron oxides is prevalent in paddies, the model system in studying the redox process in natural eco-systems. It is influenced by microbial under waterlogging soil system. Study on characteristics of dissimilatory reduction of iron oxides in paddies and its influence factors, might help better understanding the paddies formation, its microbial ecology, also could help to elucidate the remediation and transformation of contamination in paddy soils.DIR is closely related to the electron donors, acceptors and shuttles, affected by environmental conditions such as temperature, light. In the dissertation, effects of electron donors such as organic acids, saccharides, acceptors such as Cr(VI), Cu(II), SO42-, As(V) , and electron shuttles such as AQDS, FA, on the DIR process were investigated, and environmental conditions such as temperature, light were also discussed. 16S rDNA sequencing analyses was employed to genealogical classified the Fe(III)-reducers isolated from paddies using purified amplification culture method. The main objectives and results obtained are listed as follows:1. Effects of electron donors such as organic acids, saccharides on the dissimilatory reduction of amorphous Fe(OH)3 synthesized in the laboratory was studied by Co-culture incubation method. Control experiments showed that there is no obviously accumulation of Fe(II) in the system in the absence of electron donors, which suggested that the DIR is a microbiological process, typically. Addition of electron donors such as organic acids could enhance the accumulation of Fe(II) in the system and the rate constant of the DIR reaction. The dissimatory reduction ratio of Fe(OH)3 is 56.70%, 45.47%, 35.10%, and the rate constant is 0.816, 0.318, 0.520 d-1, in the presence of lactate, pyruvate, and acetate, respectively. The enhancement of organic acids is related to its molecule structure such as carbon chain length and function group. In addition, electron donors such as saccharides could also enhance the accumulation of Fe(II) in the system and the rate constant of the DIR reaction, and the accumulation of Fe(II) and the rate constant could be enhanced by the addition of saccharides concentration increased. The dissimilatory reduction ratio of Fe(OH)3 is 94.40%, 93.23%, 97.38%, 87.68% and the rate constant is 0.937, 1.035, 0.218, 0.183 d-1, in the presence of glucose, fructose, starch, and cellulose, respectively. Effects of amylose such as starch and cellulose on the dissimilatory reduction of iron oxides in the paddy soils were also investigated by soil slurry anaerobic incubation method. The acumulation of ferrous and the rate constant could be enhanced by the addition of starch and cellulose with its concentration increased from 0 to 20 g L-1. The enhancement is correlated with soil characters such as pH, organic matter, and amorphous iron content. Compare with the kinetics parameters of Fe(III) reduction in anaerobic incubation, in the range microbe normal growing pH, the lower pH is more appropriate.2. Cr(VI), Cu(II), As(V) and SO42- could be used as alternative electron acceptor by Dissimilatory Iron Reduction Bacteria. The investigation showed that Fe(III) reduction started only when the reduction of Cr(VI) was accomplished, and the generation of ferrous was delayed by the addition of alternative electron acceptors as Cr(VI), and it was accelerated by the increasing concentration of Cr(VI), suggested that the effect of Cr(VI) on the DIR might be an combined bio-abio transformation, in which Cr(VI) could be reduced by Fe(II) generated from DIR. When Cu(II) was used as alternative electron acceptor, both the accumulation of Fe(II), and the rate constant of DIR were decreased, and accelerated by the increasing concentration. The DIR was inhibited obviously when the concentration of Cu(II) added increased to 800 mg L-1. As(V) could served as an alternative electron acceptor, thus inhibit the DIR with the increasing concentration. Compared with Cr(VI), As(V) could not be reduced entirely, which suggested Fe(III) could regulate dissimilatory reduction of As(V) under iron-reducing conditions. Though the rate constant was decreased with the increasing concentration of sulphate, the cumulation of Fe(II) during the reduction process was increased. And the further investigation showed that the decrease of the rate constant was accelerated with the increasing electron donors could be used by DIRB.3. Co-culture incubation and soil slurry anaerobic incubation method were employed to study the effect of electron shuttles on the dissimilatory reduction of Fe(OH)3 and iron oxides in different paddy soils such as ZJ, JL, SC, and JX. The obtained results showed that the accumulation of Fe(II) caused by dissimilatory reduction of Fe(OH)3 was not affected by the addition of AQDS or FA, but the rate constant was increased when the electron shuttle such as AQDS or FA was added. As contrast, the accumulation of Fe(II) caused by dissimilatory reduction of iron oxides in paddy soils and the rate constant was increased by the addition of AQDS.4. Soil slurry incubated under anaerobic constant and variable temperature to investigate the effect of temperature on DIR process. The result showed that, both Fe(II) accumulation amount and rate constant were increased at higher temperature. The Fe(III)/Fe(II) ratio could has a corelationship with Eh. Decomposition of soil organic matter was improved at higher temperature, thus the reductive capacity of iron oxides in the soils was increased, and the Fe(III)/Fe(II) ratio decreased dramatically with the increase of temperature at the meantime. Therefore, DIR process was improved, and the Eh was decreased at the same time. And the consumption of free iron oxides by DIRB was increased with the increase of temperature, which could also improve the DIR process. The results from soil slurry incubation under variable temperature also showed that, the effect of temperature might closely related to the mineralization of soil organic matter, which could provide a low molecular carbon source-facilitated the consumption by DIRB. Thus, the maximum Fe(II) accumulation amount might decreased under the higher temperature only when soil slurry incubation under constant temperature first. This might attributed to the depletion of soil organic matter which was facilitated to mineralization.5. Soil slurry was incubated under anaerobic constant temperature conditions, treatments including illuminous incubation, dark incubation, illuminous to dark incubation, and dark to illuminous incubation were setup to simulate natural conditions to investigate the effect of illumination on the dissimilatory reduction of iron oxides in paddy soils. Under illuminous conditions, the trend of Fe(II) vs incubation time is different in anaerobic incubated calcareous paddy soils, it increased at the initial stages and then decreased to a very low concentration. To SC and TJ, the accumulation of Fe(II) was decreased in by 54.75% and 78.13%. When illumination was ceased, Fe(II) content increase dramatically. Phototrophic bacteria, identified to be cyanobacteria by Optics Image microscope, can oxidizing Fe(II) to Fe(III) through photosynthesis respiration by photosynthetic system II which producing O2. Added sulphate can inhibit the process of Fe(II) oxidation, and the phototrophs bacteria were poisoned by excess sulphate.6. An anaerobic, gram-negative, rod-shaped organism P4 was isolated from Sichuan paddy soil using purified amplification culture method. Adopting the PCR acquired parts of 16S rDNA sequences of Fe(III)-reducting strain P4 with 1325 bp. Through NCBI MegaBlast homology analysis and 16S rDNA phylogenetic tree construction, strain P4 belonged to Clostridium butyricum. With different carbon sources, the maximal reaction rate of iron reduction was in proper order for: glucose>pyruvate>lactate>>succinate>propionat >acetate. With glucose as carbon source, The dissimatory reduction ratio of Fe(OH)3 is 63.79%, pyruvate is 22.19%, the others are all below 10%, showing glucose and pyruvate be the dominant carbon source for strain P4.Aimed at the dissimilatory reduction of iron oxides in paddy soils, taken the environmental conditions into account, the DIR process was investigated by Co-culture and soil slurry incubation method under anoxic conditions. Effect of intrinsic factor such as electron donors, acceptors, shuttles, and environmental factors such as temperature, light were investigated to disclose mechanism of dissimilatory iron reduction. This work indicated that the DIR process could be regulation and controlled to decontaminate reducible pollutants in natural environments, and will be helpful to understand the DIR process and would also provide basic understanding to the natural attenuation of reducible organic pollutants in the contaminated sediments, subsurface environments, and soil profiles. This investigation provides a feasible remediation technology and might potentially be used in conjunction with the existing remediation strategies.