Contribution Of Microbial Fermentation To Iron(?) Reduction In Submerged Paddy Soils

Iron(?)[Fe(?)] reduction in paddy soils is closely related to the geochemical cycling of carbon,nitrogen,sulfur and phosphorus.It is of geochemical and environmental significance due to its key role in the degradation of organic pollutants,redox of heavy metals with variable valency and the suppression of methane emissions.Over the last few decades,researchers have extensively investigated obligate Fe(?)-reducing bacterium in sedimentary environments,revealing their unique metabolic patterns and considerable contribution to Fe(?)reduction.However,facultative Fe(?)reduction that accompanied with the anaerobic decomposition of organic matter is considers as a minor pathway,resulting in a lack of knowledge regarding the contribution of fermentative processes to microbial Fe(?)reduction.Unlikely naturally saturated sediments,paddy fields experience the alternations of drainage and flooding,which drives the microbial hydrogen production process coupling with organic matter fermentation as a crucial part on the electron transport chain.Therefore,further researches need to be conducted to investigate the contribution of organic matter fermentation to Fe(?)reduction in paddy soils.In the present study,paddy soils were collected from twenty drained post-harvest paddy fields representative of China's major rice production provinces.Anaerobic incubations of paddy soils were conducted with a constant temperature to determine the basic properties and Fe(?)reducing characteristics.The dominant factors influencing Fe(?)reducing capacities were discussed.To simulate the effect of mesostate and pH changes by microbial fermentation on Fe(?)reduction,paddy soils were amended with low-molecule weight organic carbon(glucose,pyruvate,lactate)or pH regulator,respectively.Dehydrogenase activity,hydrogen production,Fe(?)reducing characteristic and microbial community by high-thought-sequencing were determined to assess the relationship between Fe(?)reduction and microbial community,and the associated fermentative processes in paddy soils.The fluorescence components of dissolved organic carbon extracted from paddy soils were analyzed to reveal the contribution of the humic acid-like components to microbial Fe(?)reducing characteristics.By taking biochar as adsorptive carrier to increase the input of electron donors and shuttles,biochar and glucose-modified biochar were also amended to paddy soils to evaluate its potential to accelerate Fe(?)reduction in paddy soils.This applied the mechanism of contribution of microbial organic matter fermentation and electron transfer of dissolved organic matter to Fe(?)reduction into practice.The major results and conclusions are listed below:(1)Fe(?)accumulation exhibited an increasing trend over incubation time in submerged paddy soils.There was a significant difference in Fe(?)reducing characteristics among soils collected from different rice production provinces.Among soil basic properties,content of organic matter and amorphous iron was the dominate factor influencing microbial Fe(?)reducing characteristic,with a significant correlation between content of organic matter and amorphous iron,and Fe(?)reducing potential and the maximum Fe(?)reducing rate,respectively.Moreover,soil pH value played an key role in influencing the time to the maximum Fe(?)reducing rate.(2)An obvious difference was found in the response of microbial Fe(?)reduction to carbon sources in paddy soils from different sampling sites.The pH decrease by microbial fermentation of glucose inhibited the Fe(?)reduction in the acidic paddy soil.With different carbon sources as substrates,microbial fermentative dehydrogenation and hydrogen(H2)production mainly occurred in the initial stage of anaerobic incubation,being the maximum hydrogen production with glucose,fewer with pyruvate,and the minimum with lactate.Meanwhile,there was a consistency between the time to maximum dehydrogenase activity and the time to maximum Fe(?)reduction rate.The decrease of pH value in the culture system indicated that H+ was a major product of organic matter fermentation by microorganisms.Taking Fe(OH)3 as electron acceptor,Fe(?)was significantly reduced coupling with the consumption of H2 which produced by fermenting different carbon sources.There was a negative correlation between pH and the partial pressure of H2,as well as the concentration of Fe(?).The dehydrogenation and H2 production was regulated by the organic acid produced by microbial fermentation,which could further influence the microbial Fe(?)reducing characteristic.(3)With the depletion of bioavailable nutrition substrate and iron acceptor,the microbial community and potential Fe(?)-reducing bacteria in flooded paddy soils underwent distinct successions.At the phylum level,Firmicutes were the most dominant bacterial group and their relative abundance gradually decreased with the extension of flooding time.Besides,Acidobacteria,Bacteroidetes,Chloroflexi,Proteobacteria and Ignavibacteriae were the second most dominant bacterial group and their relative abundances showed increase tread as a function of flooding time.As the addition of different concentrations of glucose provided enough carbon sources for microorganisms,plenty of Firmicutes were enriched,particularly referring to fermentative H2-producing Clostridium and Bacillus.Meanwhile,the succession of microbial communities also lagged and the lag effect enhanced with the increased concentration of carbon sources.In paddy soil enrichment cultures by adding different concentrations of glucose,Clostridium and Bacillus were the most abundant potential Fe(?)-reducing bacteria followed by Desulfitobacterium,Paenibacillus,Anaeromyxobacter and Solibacillus,Combing with the characteristics of microbial dehydrogenation,H2 production,and Fe(?)reduction,it was obvious that the shifts in the community structure of Fe(?)-reducing bacterium,as well as dehydrogenation and H2 production responded to the shifts in the community structure of Clostridium and Bacillus,further influenced microbial Fe(?)reduction.(4)During anaerobic incubation,as the initial soil pH of acidic and alkaline paddy soils shifts to acidic or lower levels,dehydrogenase activity were greatly inhibited.Similarly,H2 production in alkaline paddy soils was also inhibited,whereas this process in acidic paddy soils was promoted.Although acid-soluble Fe(?)level significantly increased compared with that in the control treatment,Fe(?)reduction was still inhibited due to the decrease of initial pH.These results indicated that under low pH conditions,the dissolution of Fe(?)oxides was not the main factor affecting the Fe(?)reduction process.As the initial soil pH shifts to neutral and higher levels,pH had no significant effect on microbial dehydrogenation process and acid-soluble Fe(?).However,partial pressure of H2 significantly declined accompanied by earlier time to maximum Fe(?)reduction rate in the neutral treatment of alkaline paddy soil and in the neutral,alkaline,and strongly alkaline treatments of acidic paddy soil,implying the supporting role of H2 consumption for microbial Fe(?)reduction.During the rapid Fe(?)reduction stage,dehydrogenase activity representing the activity of microorganisms for metabolizing organic matters was correlated with Fe(?)accumulation in paddy soils.The influences of initial soil pH on microbial Fe(?)reduction could be attributed to the changes of enzymatic dehydrogenation and H2 production by pH shifts.(5)The initial soil pH shift from acidic to alkaline was less influential on the microbial community structure,with a slight increase in the relative abundance of Firmicutes.As the initial soil pH shift from alkaline to acidic,microbial community structure was changed intensively,with a remarkable increase in the relative abundance of Proteobacteria.The stimulative growth of potential Fe(?)reducing bacterium with pH shift from acidic to alkaline was accompanied by the enhanced consumption of Fe(?)reduction to H2 and the shorter time to reach the maximum Fe(?)reducing rate.When regulating the soil initial pH from acidic to alkaline,the relative abundance of potential Fe(?)reducing bacterium was depressed during the early stage of anaerobic incubation,while enriched during the middle and later stage of incubation.Coincidentally,as the initial soil pH shift from alkaline to acidic,there was an inhibition effect of pH regulation on Fe(?)accumulation during the early stage of anaerobic incubation and a remarkable Fe(?)accumulation was detected during the middle and later stage as the initial soil pH shift from alkaline to acidic.The above results revealed that the Fe(?)reduction process was affected by the variation in potential Fe(?)reducing bacterium community structure with pH regulation directly and the fermentative dehydrogenation and hydrogenation indirectly.(6)All the fluorescence EEM spectra of solutions extracted from paddy soils were decomposed into a four-component model based on the residual analysis and split half analysis.The fluorescence index of paddy soils revealed that dissolved organic matter was terrestrially derived rather than microbial derived.The humification index was positively related to the Fe(?)reducing capacity of paddy soils.The maximum contribution of highmolecule-weight UVC terrestrial humic acid-like component to Fe(?)reducing capacity was detected,while the content of dissolved organic matter showed the minimum contribution.There were significant correlation between the Logistic kinetics parameters of microbial Fe(?)reduction and the fluorescence of UVC+UVA terrestrial humic acid-like component and the UVA marine humic acid-like component.It is speculated that the humification degree of dissolved organic matter and the content of each humic acid-like component play a positive role in the Fe(?)reducing characteristics of paddy soils(7)Biochar addition at different particle sizes led to a promotion to Fe(?)reduction in submerged paddy soils.A more remarkable increase was detected as particle size decreased.This is partly attributed to the dissolved organic matter of biochar which was considered as electron shuttle and responsible for the majority of the increase in Fe(?)accumulation.Another reason could be that biochar addition increased the contribution of free Fe and nitrate nitrogen to Fe(?)reduction.Glucose-modified biochar can effectively stimulate Fe(?)reduction in paddy fields while simultaneously alleviating the pH increase usually caused by pristine biochar application,with a more significant effect as the decreased particle size.This study extended and deepened our knowledge about the influence factors and microbial community related to Fe(?)reduction.The contribution of microbial fermentation to Fe(?)reduction revealed in this study turned the research focus of Fe(?)reduction from “obligate” to “facultative”.Therefore,this thesis provides scientific support for revealing the mechanism of Fe(?)reduction in paddy soil and theoretical support for degradation of organic pollutant via accelerating Fe(?)reduction in paddy soils.