Mechanism Of Microbial Arsenic Reduction In Flooded Paddy Soil In The Presence Of Humic-like Substances

Arsenic contamination in paddy fields is a widespread environmental issue,and the dietary intake of rice has become a major source of arsenic in the human body;this is especially acute in South and Southeast Asian countries where rice is commonly consuming as a staple food.Arsenic biogeochemical behavior and toxicity vary greatly among different chemical species.The development of anoxic conditions in paddy field leads to an increase in arsenic reduction,which can facilitate arsenic mobility.Arsenic biogeochemical cycling is often coupled to the cycling of iron,carbon,nitrogen and sulfur.Biochar,riboflavin and humic substances are all electroactive humic-like organic matter,which can potentially participate in the reduction of arsenic and affect its behavior in the environment.So far,the potential role of the above-described humiclike substances in arsenic reduction and the related mechanism of microbial arsenic reduction in flooded paddy soil are still poorly characterized.In this study,anaerobic microcosms were set up with an arsenic-contaminated paddy soil and amendments of biochar,riboflavin and three extracted humic fractions(fulvic acid,FA;humic acid,HA;and humin),in combination with RNA-based high-throughput sequencing and quantitive real-time PCR to investigate the potential mechanism of microbial arsenic reduction in flooded paddy soil.The main findings were as follows:(1)Firstly,the role of artificial solid biochar on arsenic reduction in flooded paddy soil was evaluated.Chemical analyses revealed that biochar amendment can significantly enhance microbially mediated reduction of As(V),releasing amounts of As(III)into soil solution in the presence of lactate,mainly due to biochar's electron shuttle function.Microbial community profiling was also performed targeting As(V)-respiring gene arrA and arsenic resistance gene arsC.Results indicated that both arrA closely associated with Geobacter and arsC related to Enterobacteriaceae were selected by lactate,and lactate and biochar.Compared with the lactate alone microcosms,transcriptions of the bacterial 16 S rRNA gene,Geobacter spp.and arrA gene were increased in the lactate and biochar microcosms,where transcript abundance of Geobacter and arrA closely tracked with dissolved As(V)concentrations.Thus,lactate and biochar in flooded paddy soils can stimulate the active As(V)-respiring bacteria Geobacter spp.and arrA gene transcripts for arsenic reduction and release.(2)To further investigate the influence of soluble electroactive organic matter secreted by microorganisms on arsenic reduction in flooded paddy soil,riboflavin was thus selected as a model substance to conduct this experiment.Similar to biochar,addition of 5-50 ?M riboflavin can dramatically increase microbial arsenic reduction in the presence of lactate in flooded paddy soil.Analysis of arrA gene based transcript clone libraries showed that the community composition of active dissimilatory arsenate-reducing bacteria was different between 5 ?M and 50 ?M riboflavin amended microcosms,with the dominant bacteria detected as Geobacter spp.Transcriptions of bacterial 16 S rRNA gene,Geobacter spp.and arrA gene were higher in lactate and riboflavin microcosms than that in lactate alone microcosms.Furthermore,the abundance of arrA gene transcripts closely tracked with dissolved As(V)concentrations in lactate and 50 ?M riboflavin microcosms.Thus,similar to that of biochar,5-50 ?M riboflavin can also stimulate the active As(V)-respiring bacteria Geobacter spp.and arrA gene transcripts for arsenic reduction and release in the presence of lactate in flooded paddy soils.(3)On basis of knowledge on arsenic reduction in the presence of artificial biochar and soluble electroactive riboflavin,then anaerobic microcosms were set up amended with three extracted soil humic fractions(fulvic acid,FA;humic acid,HA and humin,HM)to comprehensively investigate its influence on arsenic reduction in flooded paddy soil.Different from biochar and riboflavin,the rate and extent of microbial As(V)reduction was significantly enhanced by humic substances,following the order FA > HA > HM.In addition,the abiotic process,that is microbially reduced FA,produced approximately 50-70% of arsenic equivalent to that of microbial As(V)reduction.arrA gene-based microbial community profiling indicated that FA addition selected for ArrA related to Geobacter sp.OR-1 while HA and HM selected for ArrA that were related to uncultured organisms.Quantitative reverse transcription-PCR analysis revealed that FA significantly stimulated the transcriptions of Geobacter spp.and arr A gene.Comprehensive analysis of physical and chemical properties of humic substances,arsenic reduction transcription,functional bacterial community and quantification of transcripts,we may conclude that the electron transfer capacities of humic substances(FA > HA > Humin)highly affected arsenic reduction.