Selenium alleviates chromium stress and promotes chromium uptake in As-hyperaccumulator Pteris vittata: Cr reduction and cellar distribution.

Arsenic-hyperaccumulator Pteris vittata exhibits remarkable absorption ability for chromium (Cr) while beneficial element selenium (Se) helps to reduce Cr-induced stress in plants. However, the effects of Se on the Cr uptake and the associated mechanisms in P. vittata are unclear, which were investigated in this study. P. vittata plants were grown for 14 days in 0.2-strength Hoagland solution containing 10 (Cr10) or 100 µM (Cr100) chromate (CrVI) and 1 µM selenate (Se1). The plant biomass, malondialdehyde contents, total Cr and Se contents, Cr speciation, expression of genes associated with Cr uptake, and Cr subcellular distribution in P. vittata were determined. P. vittata effectively accumulated Cr by concentrating 96-99% in the roots under Cr100 treatment. Further, Se substantially increased its Cr contents by 98% to 11,596 mg kg-1 in the roots, which may result from Se's role in reducing its oxidative stress as supported by 27-62% reduction in the malondialdehyde contents. Though supplied with CrVI, up to 98% of the Cr in the roots was reduced to insoluble chromite (CrIII), with 83-89% being distributed on root cell walls. Neither Cr nor Se upregulated the expression of sulfate transporters PvSultr1;1-1;2 or phosphate transporter PvPht1;4, indicating their limited role in Cr uptake. P. vittata effectively accumulates Cr in the roots mainly as CrIII on cell walls and Se effectively enhances its Cr uptake by reducing its oxidative stress. Our study suggests that Se can be used to enhance P. vittata Cr uptake and reduce its oxidative stress, which may have application in phytostabilization of Cr-contaminated soils.

Cooperation and competition between denitrification and chromate reduction in a hydrogen-based membrane biofilm reactor.

Competition and cooperation between denitrification and Cr(VI) reduction in a H2-based membrane biofilm reactor (H2-MBfR) were documented over 55 days of continuous operation. When nitrate (5 mg N/L) and chromate (0.5 mg Cr/L) were fed together, the H2-MBfR maintained approximately 100 % nitrate removal and 60 % chromate Cr(VI) removal, which means that nitrate outcompeted Cr(VI) for electrons from H2 oxidation. Removing nitrate from the influent led to an immediate increase in Cr(VI) removal (to 92 %), but Cr(VI) removal gradually deteriorated, with the removal ratio dropping to 14 % after five days. Cr(VI) removal resumed once nitrate was again added to the influent. 16S rDNA analyses showed that bacteria able to carry out H2-based denitrification and Cr(VI) reduction were in similar abundances throughout the experiment, but gene expression for Cr(VI)-reduction and export shifted. Functional genes encoding for energy-consuming chromate export (encoded by ChrA) as a means of bacterial resistance to toxicity were more abundant than genes encoding for the energy producing Cr(VI) respiration via the chromate reductase ChrR-NdFr. Thus, Cr(VI) transport and resistance to Cr(VI) toxicity depended on H2-based denitrification to supply energy. With Cr(VI) being exported from the cells, Cr(VI) reduction to Cr(III) was sustained. Thus, cooperation among H2-based denitrification, Cr(VI) export, and Cr(VI) reduction led to sustained Cr(VI) removal in the presence of nitrate, even though Cr(VI) reduction was at a competitive disadvantage for utilizing electrons from H2 oxidation.

Sphingobium sp. SJ10-10 encodes a not-yet-reported chromate reductase and the classical Rieske dioxygenases to simultaneously degrade PAH and reduce chromate.

Both polycyclic aromatic hydrocarbons (PAHs) and heavy metals persist in the environment and are toxic to organisms. Their co-occurrence makes any of them difficult to remove during bioremediation and poses challenges to environmental management and public health. Microorganisms capable of effectively degrading PAHs and detoxifying heavy metals concurrently are required to improve the bioremediation process. In this study, we isolated a new strain, Sphingobium sp. SJ10-10, from an abandoned coking plant and demonstrated its capability to simultaneously degrade 92.6 % of 75 mg/L phenanthrene and reduce 90 % of 3.5 mg/L hexavalent chromium [Cr(VI)] within 1.5 days. Strain SJ10-10 encodes Rieske non-heme iron ring-hydroxylating oxygenases (RHOs) to initiate PAH degradation. Additionally, a not-yet-reported protein referred to as Sphingobium chromate reductase (SchR), with low sequence identity to known chromate reductases, was identified to reduce Cr(VI). SchR is distributed across different genera and can be classified into two classes: one from Sphingobium members and the other from non-Sphingobium species. The widespread presence of SchR in those RHO-containing Sphingobium members suggests that they are excellent candidates for bioremediation. In summary, our study demonstrates the simultaneous removal of PAHs and Cr(VI) by strain SJ10-10 and provides valuable insights into microbial strategies for managing complex pollutant mixtures.

A PadR family transcriptional repressor regulates the transcription of chromate efflux transporter in Enterobacter sp. Z1.

Chromium is a prevalent toxic heavy metal, and chromate [Cr(VI)] exhibits high mutagenicity and carcinogenicity. The presence of the Cr(VI) efflux protein ChrA has been identified in strains exhibiting resistance to Cr(VI). Nevertheless, certain strains of bacteria that are resistant to Cr(VI) lack the presence of ChrB, a known regulatory factor. Here, a PadR family transcriptional repressor, ChrN, has been identified as a regulator in the response of Enterobacter sp. Z1(CCTCC NO: M 2019147) to Cr(VI). The chrN gene is cotranscribed with the chrA gene, and the transcriptional expression of this operon is induced by Cr(VI). The binding capacity of the ChrN protein to Cr(VI) was demonstrated by both the tryptophan fluorescence assay and Ni-NTA purification assay. The interaction between ChrN and the chrAN operon promoter was validated by reporter gene assay and electrophoretic mobility shift assay. Mutation of the conserved histidine residues His14 and His50 resulted in loss of ChrN binding with the promoter of the chrAN operon. This observation implies that these residues are crucial for establishing a DNA-binding site. These findings demonstrate that ChrN functions as a transcriptional repressor, modulating the cellular response of strain Z1 to Cr(VI) exposure.

Bioreduction of chromate in a syngas-based membrane biofilm reactor.

This study leveraged synthesis gas (syngas), a renewable resource attainable through the gasification of biowaste, to achieve efficient chromate removal from water. To enhance syngas transfer efficiency, a membrane biofilm reactor (MBfR) was employed. Long-term reactor operation showed a stable and high-level chromate removal efficiency > 95%, yielding harmless Cr(III) precipitates, as visualised by scanning electron microscopy and energy dispersive X-ray analysis. Corresponding to the short hydraulic retention time of 0.25 days, a high chromate removal rate of 80 µmol/L/d was attained. In addition to chromate reduction, in situ production of volatile fatty acids (VFAs) by gas fermentation was observed. Three sets of in situ batch tests and two groups of ex situ batch tests jointly unravelled the mechanisms, showing that biological chromate reduction was primarily driven by VFAs produced from in situ syngas fermentation, whereas hydrogen originally present in the syngas played a minor role. 16 S rRNA gene amplicon sequencing has confirmed the enrichment of syngas-fermenting bacteria (such as Sporomusa), who performed in situ gas fermentation leading to the synthesis of VFAs, and organics-utilising bacteria (such as Aquitalea), who utilised VFAs to drive chromate reduction. These findings, combined with batch assays, elucidate the pathways orchestrating synergistic interactions between fermentative microbial cohorts and chromate-reducing microorganisms. The findings facilitate the development of cost-effective strategies for groundwater and drinking water remediation and present an alternative application scenario for syngas.

Efficient nitrate and Cr(VI) removal by denitrifier: The mechanism of S. oneidensis MR-1 promoting electron production, transportation and consumption.

When Cr(VI) and nitrate coexist, the efficiency of both bio-denitrification and Cr(VI) bio-reduction is poor because chromate hinders bacterial normal functions (i.e., electron production, transportation and consumption). Moreover, under anaerobic condition, the method about efficient nitrate and Cr(VI) removal remained unclear. In this paper, the addition of Shewanella oneidensis MR-1 to promote the electron production, transportation and consumption of denitrifier and cause an increase in the removal of nitrate and Cr(VI). The efficiency of nitrate and Cr(VI) removal accomplished by P. denitrificans as a used model denitrifier increased respectively from 51.3% to 96.1% and 34.3% to 99.8% after S. oneidensis MR-1 addition. The mechanism investigations revealed that P. denitrificans provided S. oneidensis MR-1 with lactate, which was utilized to secreted riboflavin and phenazine by S. oneidensis MR-1. The riboflavin served as coenzymes of cellular reductants (i.e., thioredoxin and glutathione) in P. denitrificans, which created favorable intracellular microenvironment conditions for electron generation. Meanwhile, phenazine promoted biofilm formation, which increased the adsorption of Cr(VI) on the cell surface and accelerated the Cr(VI) reduction by membrane bound chromate reductases thereby reducing damage to other enzymes respectively. Overall, this strategy reduced the negative effect of chromate, thus improved the generation, transportation, and consumption of electrons. SYNOPSIS: The presence of S. oneidensis MR-1 facilitated nitrate and Cr(VI) removal by P. denitrificans through decreasing the negative effect of chromate due to the metabolites' secretion.

Chromate-induced methylglyoxal detoxification system drives cadmium and chromate immobilization by Cupriavidus sp. MP-37.

The detoxification of cadmium (Cd) or chromium (Cr) by microorganisms plays a vital role in bacterial survival and restoration of the polluted environment, but how microorganisms detoxify Cd and Cr simultaneously is largely unknown. Here, we isolated a bacterium, Cupriavidus sp. MP-37, which immobilized Cd(II) and reduced Cr(VI) simultaneously. Notably, strain MP-37 exhibited variable Cd(II) immobilization phenotypes, namely, cell adsorption and extracellular immobilization in the co-presence of Cd(II) and Cr(VI), while cell adsorption in the presence of Cd(II) alone. To unravel Cr(VI)-induced extracellular Cd(II) immobilization, proteomic analysis was performed, and methylglyoxal-scavenging protein (glyoxalase I, GlyI) and a regulator (YafY) showed the highest upregulation in the co-presence of Cd(II) and Cr(VI). GlyI overexpression reduced the intracellular methylglyoxal content and increased the immobilized Cd(II) content in extracellular secreta. The addition of lactate produced by GlyI protein with methylglyoxal as substrate increased the Cd(II) content in extracellular secreta. Reporter gene assay, electrophoretic mobility shift assay, and fluorescence quenching assay demonstrated that glyI expression was induced by Cr(VI) but not by Cd(II), and that YafY positively regulated glyI expression by binding Cr(VI). In the pot experiment, inoculation with the MP-37 strain reduced the Cd content of Oryza sativa L., and their secreted lactate reduced the Cr accumulation in Oryza sativa L. This study reveals that Cr(VI)-induced detoxification system drives methylglyoxal scavenging and Cd(II) extracellular detoxification in Cd(II) and Cr(VI) co-existence environment.

Reduction of Chromate via Biotic and Abiotic Pathways in the Presence of Three Co-contaminating Electron Acceptors.

Bioreduction of Cr(VI) to Cr(III) is a promising technology for removing Cr(VI), but Cr(VI) reduction alone cannot support microbial growth. This study investigated the reduction of Cr(VI) in the presence of three electron acceptors that typically coexist with Cr(VI): NO3-, SO42-, and Fe(III). All three systems could reduce Cr(VI) to Cr(III), but the fate of Cr, its impacts on reduction of the other acceptors, and its impact on the microbial community differed. Although Cr(VI) was continuously removed in the NO3--reduction systems, batch tests showed that denitrification was inhibited primarily through impeding nitrite reduction. The SO42- and Fe(III) reduction systems reduced Cr(VI) using a combination of biotic and abiotic processes. Across all three systems, the abundance of genera capable of reducing Cr(VI) increased following the introduction of Cr(VI). Conversely, the abundance of genera that cannot reduce or resist Cr(VI) decreased, leading to restructuring of the microbial community. Furthermore, the abundance of sulfide oxidizers and Fe(II) oxidizers substantially increased after the introduction of chromate. This study provides fundamental knowledge about how Cr(VI) bioreduction interacts with bioreductions of three other co-contaminating electron acceptors.

Hexavalent chromate bioreduction by a magnetotactic bacterium Magnetospirillum gryphiswaldense MSR-1 and the effect of magnetosome synthesis.

Magnetotactic bacteria (MTB) are receiving attention for heavy metal biotreatment due to their potential for biosorption with heavy metals and the capability of the magnetic recovery. In this study, we investigated the characteristics of Cr(VI) bioreduction and biosorption by an MTB isolate, Magnetospirillum gryphiswaldense MSR-1, which has a higher growth rate and wider reflexivity in culture conditions. Our results demonstrated that the MSR-1 strain could remove Cr(VI) up to the concentration of 40 mg L-1 and with an optimal activity at neutral pH conditions. The magnetosome synthesis existed regulatory mechanisms between Cr(VI) reduction and cell division. The addition of 10 mg L-1 Cr(VI) significantly inhibited cell growth, but the magnetosome-deficient strain, B17316, showed an average specific growth rate of 0.062 h-1 at the same dosage. Cr(VI) reduction examined by the heat-inactivated and resting cells demonstrated that the main mechanism for MSR-1 strain to reduce Cr(VI) was chromate reductase and adsorption, and magnetosome synthesis would enhance the chromate reductase activity. Finally, our results elucidated that the chromate reductase distributes diversely in multiple subcellular components of the MSR-1 cells, including extracellular, membrane-associated, and intracellular cytoplasmic activity; and expression of the membrane-associated chromate reductase was increased after the cells were pre-exposed by Cr(VI).

Chromium in plant growth and development: Toxicity, tolerance and hormesis.

Research over the last three decades showed that chromium, particularly the oxyanion chromate Cr(VI) behaves as a toxic environmental pollutant that strongly damages plants due to oxidative stress, disruption of nutrient uptake, photosynthesis and metabolism, and ultimately, represses growth and development. However, mild Cr(VI) concentrations promote growth, induce adventitious root formation, reinforce the root cap, and produce twin roots from single root meristems under conditions that compromise cell viability, indicating its important role as a driver for root organogenesis. In recent years, considerable advance has been made towards deciphering the molecular mechanisms for root sensing of chromate, including the identification of regulatory proteins such as SOLITARY ROOT and MEDIATOR 18 that orchestrate the multilevel dynamics of the oxyanion. Cr(VI) decreases the expression of several glutamate receptors, whereas amino acids such as glutamate, cysteine and proline confer protection to plants from hexavalent chromium stress. The crosstalk between plant hormones, including auxin, ethylene, and jasmonic acid enables tissues to balance growth and defense under Cr(VI)-induced oxidative damage, which may be useful to better adapt crops to biotic and abiotic challenges. The highly contrasting responses of plants manifested at the transcriptional and translational levels depend on the concentration of chromate in the media, and fit well with the concept of hormesis, an adaptive mechanism that primes plants for resistance to environmental challenges, toxins or pollutants. Here, we review the contrasting facets of Cr(VI) in plants including the cellular, hormonal and molecular aspects that mechanistically separate its toxic effects from biostimulant outputs.

A novel gene from the acidophilic bacterium Leptospirillum sp. CF-1 and its role in oxidative stress and chromate tolerance.

BACKGROUND: Acidophilic microorganisms like Leptospirillum sp. CF-1 thrive in environments with extremely low pH and high concentrations of dissolved heavy metals that can induce the generation of reactive oxygen species (ROS). Several hypothetical genes and proteins from Leptospirillum sp. CF-1 are known to be up-regulated under oxidative stress conditions. RESULTS: In the present work, the function of hypothetical gene ABH19_09590 from Leptospirillum sp. CF-1 was studied. Heterologous expression of this gene in Escherichia coli led to an increase in the ability to grow under oxidant conditions with 5 mM K2CrO4 or 5 mM H2O2. Similarly, a significant reduction in ROS production in E. coli transformed with a plasmid carrying ABH19_09590 was observed after exposure to these oxidative stress elicitors for 30 min, compared to a strain complemented with the empty vector. A co-transcriptional study using RT-PCR showed that ABH19_09590 is contained in an operon, here named the "och" operon, that also contains ABH19_09585, ABH19_09595 and ABH19_09600 genes. The expression of the och operon was significantly up-regulated in Leptospirillum sp. CF-1 exposed to 5 mM K2CrO4 for 15 and 30 min. Genes of this operon potentially encode a NADH:ubiquinone oxidoreductase, a CXXC motif-containing protein likely involved in thiol/disulfide exchange, a hypothetical protein, and a di-hydroxy-acid dehydratase. A comparative genomic analysis revealed that the och operon is a characteristic genetic determinant of the Leptospirillum genus that is not present in other acidophiles. CONCLUSIONS: Altogether, these results suggest that the och operon plays a protective role against chromate and hydrogen peroxide and is an important mechanism required to face polyextremophilic conditions in acid environments.

A novel gene from the acidophilic bacterium Leptospirillum sp. CF-1 and its role in oxidative stress and chromate tolerance

BACKGROUND: Acidophilic microorganisms like Leptospirillum sp. CF 1 thrive in environments with extremely low pH and high concentrations of dissolved heavy metals that can induce the generation of reactive oxygen species (ROS). Several hypothetical genes and proteins from Leptospirillum sp. CF 1 are known to be up regulated under oxidative stress conditions. RESULTS: In the present work, the function of hypothetical gene ABH19_09590 from Leptospirillum sp. CF 1 was studied. Heterologous expression of this gene in Escherichia coli led to an increase in the ability to grow under oxidant conditions with 5 mM K2CrO4 or 5 mM H2O2. Similarly, a significant reduction in ROS production in E. coli transformed with a plasmid carrying ABH19_09590 was observed after exposure to these oxidative stress elicitors for 30 min, compared to a strain complemented with the empty vector. A co transcriptional study using RT PCR showed that ABH19_09590 is contained in an operon, here named the "och" operon, that also contains ABH19_09585, ABH19_09595 and ABH19_09600 genes. The expression of the och operon was significantly up regulated in Leptospirillum sp. CF 1 exposed to 5 mM K2CrO4 for 15 and 30 min. Genes of this operon potentially encode a NADH:ubiquinone oxidoreductase, a CXXC motif containing protein likely involved in thiol/disulfide exchange, a hypothetical protein, and a di hydroxy acid dehydratase. A comparative genomic analysis revealed that the och operon is a characteristic genetic determinant of the Leptospirillum genus that is not present in other acidophiles. CONCLUSIONS: Altogether, these results suggest that the och operon plays a protective role against chromate and hydrogen peroxide and is an important mechanism required to face polyextremophilic conditions in acid environments.

Sulfur transfer from the endophytic fungus Serendipita indica improves maize growth and requires the sulfate transporter SiSulT.

A deficiency of the essential macronutrient sulfur leads to stunted plant growth and yield loss; however, an association with a symbiotic fungus can greatly improve nutrient uptake by the host plant. Here, we identified and functionally characterized a high-affinity sulfate transporter from the endophytic fungus Serendipita indica. SiSulT fulfills all the criteria expected of a functional sulfate transporter responding to sulfur limitation: SiSulT expression was induced when S. indica was grown under low-sulfate conditions, and heterologous expression of SiSulT complemented a yeast mutant lacking sulfate transport. We generated a knockdown strain of SiSulT by RNA interference to investigate the consequences of the partial loss of this transporter for the fungus and the host plant (maize, Zea mays) during colonization. Wild-type (WT) S. indica, but not the knockdown strain (kd-SiSulT), largely compensated for low-sulfate availability and supported plant growth. Colonization by WT S. indica also allowed maize roots to allocate precious resources away from sulfate assimilation under low-sulfur conditions, as evidenced by the reduction in expression of most sulfate assimilation genes. Our study illustrates the utility of the endophyte S. indica in sulfur nutrition research and offers potential avenues for agronomically sound amelioration of plant growth in low-sulfate environments.

Chromate fate and effect in bioelectrochemical systems for remediation of chlorinated solvents.

A continuous-flow bioelectrochemical reactor was developed in a previous study to address the bioremediation of groundwater contaminated by trichloroethene (TCE). The present report investigated the applicability of the same system in the presence of Cr(VI) and its possible inhibitory effect on dehalorespiring bacterial populations. Preliminary batch tests were performed at the optimal cathodic reducing potential for the reductive dechlorination (RD) of TCE (-0.65 V vs. the standard hydrogen electrode) with two different dechlorinating microorganism consortia. The results demonstrated that Cr(VI) removal efficacy was increased by microorganisms that had been previously acclimatised to Cr(VI). Specifically, Cr(VI) was completely reduced only in the presence of acclimated microorganisms. The presence of chromate negatively affected RD performance, by either (i) limiting the TCE transformation to cis-dichloroethene at lower concentrations, or (ii) completely inhibiting RD at higher concentrations. In contrast, after the acclimation period, RD was extended down to vinyl chloride, which is the main TCE daughter product. Finally, the continuous flow reactor was fed by synthetic groundwater contaminated with TCE (50 µM) and Cr(VI) (45 µM), and the experimental results showed that Cr(VI) was completely reduced under RD conditions. Moreover, TCE removal was complete, with vinyl chloride and ethene as the main intermediates, thus indicating that chromate inhibition was decreased by Cr(VI) removal.

Phytohormones producing rhizobacterium alleviates chromium toxicity in Helianthus annuus L. by reducing chromate uptake and strengthening antioxidant system.

Contamination of agricultural land with heavy metal is a serious biological and environmental issue. Such threat can be challenged by exploring the plant symbiotic microbes that can improve plant growth through phyto-hormones secretion and chromate chelation. In the current study, chromate resistant rhizospheric Staphylococcus arlettae strain MT4 was isolated from the rhizosphere of Malvestrum tricuspadatum L. The strain showed potential to secrete phytohormones and plant growth promoting secondary metabolites under induced chromate stress, making it a best suitable candidate in chromate stress alleviation. Moreover, the rhizobacterium MT4 significantly promoted the net assimilation and relative growth rate of sunflower grown in the presence of chromate (100 ppm). Chromate stress alleviation strategy of MT4 strain was three-fold. MT4 alleviated chromate stress and promoted the sunflower growth by suppressing the chromate intake by the host, modulating phytohormones and strengthening of the host's antioxidant system. The improved antioxidant system was confirmed by noticing lower ROS accumulation and improved ROS scavenging, lower peroxidase activity and higher accumulation of phenols and flavonoids.

Shewanella decolorationis LDS1 Chromate Resistance.

The genus Shewanella is well known for its genetic diversity, its outstanding respiratory capacity, and its high potential for bioremediation. Here, a novel strain isolated from sediments of the Indian Ocean was characterized. A 16S rRNA analysis indicated that it belongs to the species Shewanella decolorationis It was named Shewanella decolorationis LDS1. This strain presented an unusual ability to grow efficiently at temperatures from 24°C to 40°C without apparent modifications of its metabolism, as shown by testing respiratory activities or carbon assimilation, and in a wide range of salt concentrations. Moreover, S. decolorationis LDS1 tolerates high chromate concentrations. Indeed, it was able to grow in the presence of 4 mM chromate at 28°C and 3 mM chromate at 40°C. Interestingly, whatever the temperature, when the culture reached the stationary phase, the strain reduced the chromate present in the growth medium. In addition, S. decolorationis LDS1 degrades different toxic dyes, including anthraquinone, triarylmethane, and azo dyes. Thus, compared to Shewanella oneidensis, this strain presented better capacity to cope with various abiotic stresses, particularly at high temperatures. The analysis of genome sequence preliminary data indicated that, in contrast to S. oneidensis and S. decolorationis S12, S. decolorationis LDS1 possesses the phosphorothioate modification machinery that has been described as participating in survival against various abiotic stresses by protecting DNA. We demonstrate that its heterologous production in S. oneidensis allows it to resist higher concentrations of chromate.IMPORTANCEShewanella species have long been described as interesting microorganisms in regard to their ability to reduce many organic and inorganic compounds, including metals. However, members of the Shewanella genus are often depicted as cold-water microorganisms, although their optimal growth temperature usually ranges from 25 to 28°C under laboratory growth conditions. Shewanella decolorationis LDS1 is highly attractive, since its metabolism allows it to develop efficiently at temperatures from 24 to 40°C, conserving its ability to respire alternative substrates and to reduce toxic compounds such as chromate or toxic dyes. Our results clearly indicate that this novel strain has the potential to be a powerful tool for bioremediation and unveil one of the mechanisms involved in its chromate resistance.

Anaerobic methane oxidation coupled to chromate reduction in a methane-based membrane biofilm batch reactor.

Chromate can be reduced by methanotrophs in a membrane biofilm reactor (MBfR). In this study, we cultivated a Cr(VI)-reducing biofilm in a methane (CH4)-based membrane biofilm batch reactor (MBBR) under anaerobic conditions. The Cr(VI) reduction rate increased to 0.28 mg/L day when the chromate concentration was ≤ 2.2 mg/L but declined sharply to 0.01 mg/L day when the Cr(VI) concentration increased to 6 mg/L. Isotope tracing experiments showed that part of the 13C-labeled CH4 was transformed to 13CO2, suggesting that the biofilm may reduce Cr(VI) by anaerobic methane oxidation (AnMO). Microbial community analysis showed that a methanogen, i.e., Methanobacterium, dominated in the biofilm, suggesting that this genus is probably capable of carrying out AnMO. The abundance of Methylomonas, an aerobic methanotroph, decreased significantly, while Meiothermus, a potential chromate-reducing bacterium, was enriched in the biofilm. Overall, the results showed that the anaerobic environment inhibited the activity of aerobic methanotrophs while promoting AnMO bacterial enrichment, and high Cr(VI) loading reduced Cr(VI) flux by inhibiting the methane oxidation process.

Efficient bioaccumulation of tungsten by Escherichia coli cells expressing the Sulfitobacter dubius TupBCA system.

Tungsten (W) is a valuable element with considerable industrial and economic importance that belongs to the European Union list of critical metals with a high supply risk. Therefore, the development of effective and new methods for W recovery is essential to ensure a sustainable supply. In the present study, the Sulfitobacter dubius W transport system TupABC was explored in order to demonstrate both its functionality in Escherichia coli cells and to construct a bioaccumulator (EcotupW). The complete gene cluster tupBCA or partial gene cluster tupBC were cloned in an expression vector and transformed into E. coli. Metal accumulation was evaluated when each construct strain was grown with three separate metal oxyanions (tungstate, molybdate or chromate). The specificity of the bioaccumulator was determined by competition assays using cells grown with mixed solutions of metal oxyanions (W/Mo and W/Cr). The results showed the relevance of the TupA protein in the TupABC transporter system to W-uptake and also allowed Mo and Cr accumulations, although with amounts 1.7 and 2.9-fold lower than W, respectively. To identify the importance of the valine residue in the accumulation efficiency of the VTTS motif, site-directed mutagenesis of tupA was performed. A mutant with a threonine residue, instead of the respective valine, confirmed that W was internalized by nearly double the amount compared to the native form. The findings indicated that cells carrying the native S. dubius TupABC system were great W-bioaccumulators and could be promising tools for W recovery.

Microbial chromate reduction coupled with anaerobic oxidation of methane in a membrane biofilm reactor.

It has been reported that microbial reduction of sulfate, nitrite/nitrate and iron/manganese could be coupled with anaerobic oxidation of methane (AOM), which plays a significant role in controlling methane emission from anoxic niches. However, little is known about microbial chromate (Cr(VI)) reduction coupling with AOM. In this study, a microbial consortium was enriched via switching nitrate dosing to chromate feeding as the sole electron acceptor under anaerobic condition in a membrane biofilm reactor (MBfR), in which methane was continuously provided as the electron donor through bubble-less hollow fiber membranes. According to long-term reactor operation and chromium speciation analysis, soluble chromate could be reduced into Cr(III) compounds by using methane as electron donor. Fluorescence in situ hybridization and high-throughput 16S rRNA gene amplicon profiling further indicated that after feeding chromate Candidatus 'Methanoperedens' (a known nitrate-dependent anaerobic methane oxidation archaeon) became sole anaerobic methanotroph in the biofilm, potentially responsible for the chromate bio-reduction driven by methane. Two potential pathways of the microbial AOM-coupled chromate reduction were proposed: (i) Candidatus 'Methanoperedens' independently utilizes chromate as electron acceptor to form Cr(III) compounds, or (ii) Candidatus 'Methanoperedens' oxidizes methane to generate intermediates or electrons, which will be utilized to reduce chromate to Cr(III) compounds by unknown chromate reducers synergistically. Our findings suggest a possible link between the biogeochemical chromium and methane cycles.

Insights into the mode of flavin mononucleotide binding and catalytic mechanism of bacterial chromate reductases: A molecular dynamics simulation study.

Enzymes from natural sources protect the environment via complex biological mechanisms, which aid in reductive immobilization of toxic metals including chromium. Nevertheless, progress was being made in elucidating high-resolution crystal structures of reductases and their binding with flavin mononucleotide (FMN) to understand the underlying mechanism of chromate reduction. Therefore, herein, we employed molecular dynamics (MD) simulations, principal component analysis (PCA), and binding free energy calculations to understand the dynamics behavior of these enzymes with FMN. Six representative chromate reductases in monomeric and dimeric forms were selected to study the mode, dynamics, and energetic component that drive the FMN binding process. As evidenced by MD simulation, FMN prefers to bind the cervix formed between the catalytic domain surrounded by strong conserved hydrogen bonding, electrostatic, and hydrophobic contacts. The slight movement and reorientation of FMN resulted in breakage of some crucial H-bonds and other nonbonded contacts, which were well compensated with newly formed H-bonds, electrostatic, and hydrophobic interactions. The critical residues aiding in tight anchoring of FMN within dimer were found to be strongly conserved in the bacterial system. The molecular mechanics combined with the Poisson-Boltzmann surface area binding free energy of the monomer portrayed that the van der Waals and electrostatic energy contribute significantly to the total free energy, where, the polar solvation energy opposes the binding of FMN. The proposed proximity relationships between enzyme and FMN binding site presented in this study will open up better avenues to engineer enzymes with optimized chromate reductase activity for sustainable bioremediation of heavy metals.