Exogenous Electron Shuttle-Mediated Extracellular Electron Transfer of <i>Shewanella putrefaciens</i> 200: Electrochemical Parameters and Thermodynamics

Despite the importance of exogenous electron shuttles (ESs) in extracellular electron transfer (EET), a lack of understanding of the key properties of ESs is a concern given their different influences on EET processes. Here, the ES-mediated EET capacity of <i>Shewanella putrefaciens</i> 200 (SP200) was evaluated by examining the electricity generated in a microbial fuel cell. The results indicated that all the ESs substantially accelerated the current generation compared to only SP200. The current and polarization parameters were linearly correlated with both the standard redox potential (<i>E</i>_ES⁰) and the electron accepting capacity (EAC) of the ESs. A thermodynamic analysis of the electron transfer from the electron donor to the electrode suggested that the EET from <i>c</i>-type cytochromes (<i>c</i>-Cyts) to ESs is a crucial step causing the differences in EET capacities among various ESs. Based on the derived equations, both <i>E</i>_ES⁰ and EAC can quantitatively determine potential losses (Δ<i>E</i>) that reflect the potential loss of the ES-mediated EET. In situ spectral kinetic analysis of ES reduction by <i>c</i>-Cyts in a living SP200 suspension was first investigated with the <i>E</i>_ES, <i>E</i>_{<i>c</i>‑Cyt}, and Δ<i>E</i> values being calculated. This study can provide a comprehensive understanding of the role of ESs in EET

Heterogeneous Photodegradation of Pentachlorophenol with Maghemite and Oxalate under UV Illumination

The degradation of pentachlorophenol (PCP) in a heterogeneous system with maghemite (γ-Fe2O3) and oxalate under UV illumination was investigated in this study. The results of adsorption experiments demonstrated competitive adsorption between PCP and oxalic acid on the surface of γ-Fe2O3. The results of photodegradation experiments showed that the rate of PCP degradation strongly relied on the oxalic acid concentration and that an optimal tested initial concentration of oxalic acid (C0ox) of 0.8 mM was obtained under our experimental conditions. It was observed that a sufficient amount of oxalic acid can be adsorbed on the γ-Fe2O3 to form various Fe(III)-oxalate complexes at C0ox = 0.8 mM. During the photoreaction, Fe(C2O4)2− and Fe(C2O4)33− were found to be the dominant Fe(III)-oxalate complexes at different C0ox, while Fe(C2O4)22− was the dominant Fe(II)-oxalate complex at C0ox ≥ 0.8 mM. The mechanism of H2O2 formation and consumption in the UV-irradiated γ-Fe2O3/oxalate system was proposed and evaluated. Furthermore, six intermediates of PCP degradation were identified by GC/MS, HPLC, and IC analyses, respectively, and a possible pathway of PCP degradation in such a system was proposed

Soil Humic Acid Stimulates Potentially Active Dissimilatory Arsenate-Reducing Bacteria in Flooded Paddy Soil as Revealed by Metagenomic Stable Isotope Probing

Dissimilatory arsenate reduction contributes a large proportion of arsenic flux from flooded paddy soil, which is closely linked to soil organic carbon input and efflux. Humic acid (HA) represents a natural ingredient in soil and is shown to enhance microbial arsenate respiration to promote arsenic mobility. However, the community and function profiles of metabolically active arsenate-respiring bacteria and their interactions with HA in paddy soil remain unclear. To probe this linkage, we performed a genome-centric comparison of potentially active arsenate-respiring bacteria in anaerobic microcosms amended with 13C-lactate and HA by combining stable-isotope probing with genome-resolved metagenomics. Indeed, HA greatly accelerated the microbial reduction of arsenate to arsenite. Enrichment of bacteria that harbor arsenate-respiring reductase genes (arrA) in HA-enriched 13C-DNA was confirmed by metagenomic binning, which are affiliated with Firmicutes (mainly Desulfitobacterium, Bacillus, Brevibacillus, and Clostridia) and Acidobacteria. Characterization of reference extracellular electron transfer (EET)-related genes in these arrA-harboring bacteria supports the presence of EET-like genes, with partial electron-transport chain genes identified. This suggests that Gram-positive Firmicutes- and Acidobacteria-related members may harbor unspecified EET-associated genes involved in metal reduction. Our findings highlight the link between soil HA and potentially active arsenate-respiring bacteria, which can be considered when using HA for arsenic removal

Incorporation of Cu into Goethite Stimulates Oxygen Activation by Surface-Bound Fe(II) for Enhanced As(III) Oxidative Transformation

The dark production of reactive oxygen species (ROS) coupled to biogeochemical cycling of iron (Fe) plays a pivotal role in controlling arsenic transformation and detoxification. However, the effect of secondary atom incorporation into Fe(III) oxyhydroxides on this process is poorly understood. Here, we show that the presence of oxygen vacancy (OV) as a result of Cu incorporation in goethite substantially enhances the As(III) oxidation by Fe(II) under oxic conditions. Electrochemical and density functional theory (DFT) evidence reveals that the electron transfer (ET) rate constant is enhanced from 0.023 to 0.197 s–1, improving the electron efficiency of the surface-bound Fe(II) on OV defective surfaces. The cascade charge transfer from the surface-bound Fe(II) to O2 mediated by Fe(III) oxyhydroxides leads to the O–O bond of O2 stretching to 1.46–1.48 Å equivalent to that of superoxide (•O2–), and •O2– is the predominant ROS responsible for As(III) oxidation. Our findings highlight the significant role of atom incorporation in changing the ET process on Fe(III) oxyhydroxides for ROS production. Thus, such an effect must be considered when evaluating Fe mineral reactivity toward changing their surface chemistry, such as those noted here for Cu incorporation, which likely determines the fates of arsenic and other redox sensitive pollutants in the environments with oscillating redox conditions

Iron Vacancy Accelerates Fe(II)-Induced Anoxic As(III) Oxidation Coupled to Iron Reduction

Chemical oxidation of As(III) by iron (Fe) oxyhydroxides has been proposed to occur under anoxic conditions and may play an important role in stabilization and detoxification of As in subsurface environments. However, this reaction remains controversial due to lack of direct evidence and poorly understood mechanisms. In this study, we show that As(III) oxidation can be facilitated by Fe oxyhydroxides (i.e., goethite) under anoxic conditions coupled with the reduction of structural Fe(III). An excellent electron balance between As(V) production and Fe(III) reduction is obtained. The formation of an active metastable Fe(III) phase at the defective surface of goethite due to atom exchange is responsible for the oxidation of As(III). Furthermore, the presence of defects (i.e., Fe vacancies) in goethite can noticeably enhance the electron transfer (ET) and atom exchange between the surface-bound Fe(II) and the structural Fe(III) resulting in a two time increase in As(III) oxidation. Atom exchange-induced regeneration of active goethite sites is likely to facilitate As(III) coordination and ET with structural Fe(III) based on electrochemical analysis and theoretical calculations showing that this reaction pathway is thermodynamically and kinetically favorable. Our findings highlight the synergetic effects of defects in the Fe crystal structure and Fe(II)-induced catalytic processes on anoxic As(III) oxidation, shedding a new light on As risk management in soils and subsurface environments

Table_1_New Arsenite Oxidase Gene (aioA) PCR Primers for Assessing Arsenite-Oxidizer Diversity in the Environment Using High-Throughput Sequencing.DOCX

Gene encoding the large subunit of As(III) oxidase (AioA), an important component of the microbial As(III) oxidation system, is a widely used biomarker to characterize As(III)-oxidizing communities in the environment. However, many studies were restricted to a few sequences generated by clone libraries and Sanger sequencing, which may have underestimated the diversity of As(III)-oxidizers in natural environments. In this study, we designed a primer pair, 1109F (5′-ATC TGG GGB AAY RAC AAY TA−3′) and 1548R (5′-TTC ATB GAS GTS AGR TTC AT−3′), targeting gene sequence encoding for the conserved molybdopterin center of the AioA protein, yielding amplicons approximately 450 bp in size that are feasible for highly parallel amplicon sequencing. By utilizing in silico analyses and the experimental construction of clone libraries using Sanger sequencing, the specificity and resolution of 1109F/1548R are approximated with two other previously published and commonly used primers, i.e., M1-2F/M3-2R and deg1F/deg1R. With the use of the 1109F/1548R primer pair, the taxonomic composition of the aioA genes was similar both according to the Sanger and next-generation sequencing (NGS) platforms. Furthermore, high-throughput amplicon sequencing using the primer pair, 1109F/1548R, successfully identified the well-known As(III)-oxidizers in paddy soils and sediments, and they also revealed the differences in the community structure and composition of As(III)-oxidizers in above two biotopes. The random forest analysis showed that the dissolved As(III) had the highest relative influence on the Chao1 index of the aioA genes. These observations demonstrate that the newly designed PCR primers enhanced the ability to detect the diversity of aioA-encoding microorganisms in environments using highly parallel short amplicon sequencing.</p

Soil Humic Acid Stimulates Potentially Active Dissimilatory Arsenate-Reducing Bacteria in Flooded Paddy Soil as Revealed by Metagenomic Stable Isotope Probing

Dissimilatory arsenate reduction contributes a large proportion of arsenic flux from flooded paddy soil, which is closely linked to soil organic carbon input and efflux. Humic acid (HA) represents a natural ingredient in soil and is shown to enhance microbial arsenate respiration to promote arsenic mobility. However, the community and function profiles of metabolically active arsenate-respiring bacteria and their interactions with HA in paddy soil remain unclear. To probe this linkage, we performed a genome-centric comparison of potentially active arsenate-respiring bacteria in anaerobic microcosms amended with 13C-lactate and HA by combining stable-isotope probing with genome-resolved metagenomics. Indeed, HA greatly accelerated the microbial reduction of arsenate to arsenite. Enrichment of bacteria that harbor arsenate-respiring reductase genes (arrA) in HA-enriched 13C-DNA was confirmed by metagenomic binning, which are affiliated with Firmicutes (mainly Desulfitobacterium, Bacillus, Brevibacillus, and Clostridia) and Acidobacteria. Characterization of reference extracellular electron transfer (EET)-related genes in these arrA-harboring bacteria supports the presence of EET-like genes, with partial electron-transport chain genes identified. This suggests that Gram-positive Firmicutes- and Acidobacteria-related members may harbor unspecified EET-associated genes involved in metal reduction. Our findings highlight the link between soil HA and potentially active arsenate-respiring bacteria, which can be considered when using HA for arsenic removal

Image_1_New Arsenite Oxidase Gene (aioA) PCR Primers for Assessing Arsenite-Oxidizer Diversity in the Environment Using High-Throughput Sequencing.TIF

Gene encoding the large subunit of As(III) oxidase (AioA), an important component of the microbial As(III) oxidation system, is a widely used biomarker to characterize As(III)-oxidizing communities in the environment. However, many studies were restricted to a few sequences generated by clone libraries and Sanger sequencing, which may have underestimated the diversity of As(III)-oxidizers in natural environments. In this study, we designed a primer pair, 1109F (5′-ATC TGG GGB AAY RAC AAY TA−3′) and 1548R (5′-TTC ATB GAS GTS AGR TTC AT−3′), targeting gene sequence encoding for the conserved molybdopterin center of the AioA protein, yielding amplicons approximately 450 bp in size that are feasible for highly parallel amplicon sequencing. By utilizing in silico analyses and the experimental construction of clone libraries using Sanger sequencing, the specificity and resolution of 1109F/1548R are approximated with two other previously published and commonly used primers, i.e., M1-2F/M3-2R and deg1F/deg1R. With the use of the 1109F/1548R primer pair, the taxonomic composition of the aioA genes was similar both according to the Sanger and next-generation sequencing (NGS) platforms. Furthermore, high-throughput amplicon sequencing using the primer pair, 1109F/1548R, successfully identified the well-known As(III)-oxidizers in paddy soils and sediments, and they also revealed the differences in the community structure and composition of As(III)-oxidizers in above two biotopes. The random forest analysis showed that the dissolved As(III) had the highest relative influence on the Chao1 index of the aioA genes. These observations demonstrate that the newly designed PCR primers enhanced the ability to detect the diversity of aioA-encoding microorganisms in environments using highly parallel short amplicon sequencing.</p

Dependence of Secondary Mineral Formation on Fe(II) Production from Ferrihydrite Reduction by <i>Shewanella oneidensis</i> MR‑1

Although dissimilatory iron reduction and secondary mineral formation by <i>Shewanella oneidensis</i> MR-1 have been widely recognized, questions remain about the effects of biogenic Fe­(II) on the rate and extent of secondary mineral formation and the importance of biogenic Fe­(II)-induced crystallization processes. In this study, we investigated the effects of different mutants of MR-1 on the bioreduction and mineralization of ferrihydrite. The results indicate that while the reduction rates of ferrihydrite by Δ<i>mtrD</i>, Δ<i>mtrF</i>, and Δ<i>omcA</i> are similar to that of the wild type (WT), the capacity to reduce ferrihydrite decreased dramatically in the mutants Δ<i>cymA</i> and Δ<i>mtrA</i>. The order for Fe­(III) reduction by MR-1 WT and mutants was ranked as follows: WT ≈ Δ<i>mtrD</i> ≈ Δ<i>mtrF</i> > Δ<i>omcA</i> > Δ<i>mtrC</i> > Δ<i>cymA</i> > Δ<i>mtrA</i>. Secondary minerals of ferrihydrite were characterized using X-ray diffraction, Fourier transform infrared spectra, and scanning electron microscopy. The results show that goethite and hematite were the main secondary minerals formed during the first 2 days in all treatments, and then magnetite appeared in the WT, Δ<i>mtrD</i>, Δ<i>mtrF</i> and Δ<i>omcA</i> treatments, whereas magnetite began to appear from the sixth day onward in the Δ<i>mtrC</i> treatment. However, no magnetite was observed during the 6 days in the Δ<i>mtrA</i> and Δ<i>cymA</i> incubation treatments. The plausible electron transfer pathways of bioreduction and phase transformation were also verified using thermodynamic calculations of elementary reactions. This study clarified the importance of Fe­(II) production in secondary mineral formation processes and highlighted the significance of biogenic Fe­(II)-catalyzed crystallization. This information may, in turn, help us to better understand natural microbe–mineral interaction processes