Microbial Chromate Reduction Coupled to Anaerobic Oxidation of

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Remediation and Control Technologies

Microbial Chromate Reduction Coupled to Anaerobic Oxidation of Elemental Sulfur or Zerovalent Iron Jiaxin Shi, Baogang Zhang, Rui Qiu, Chunyu Lai, Yufeng Jiang, Chao He, and Jianhua Guo Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.8b05053 • Publication Date (Web): 18 Feb 2019 Downloaded from http://pubs.acs.org on February 18, 2019

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Environmental Science & Technology

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Microbial Chromate Reduction Coupled to Anaerobic Oxidation of

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Elemental Sulfur or Zerovalent Iron

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Jiaxin Shia, Baogang Zhanga,*, Rui Qiua, Chunyu Laib, Yufeng Jianga, Chao Hea,

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Jianhua Guob,*

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a School

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Circulation and Environmental Evolution, China University of Geosciences (Beijing),

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Beijing 100083, P. R. China

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of Water Resources and Environment, MOE Key Laboratory of Groundwater

b Advanced

Water Management Centre, The University of Queensland, St Lucia, Queensland 4072, Australia

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*Corresponding

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Baogang Zhang: Tel.: +86 10 8232 2281; Fax: +86 10 8232 1081. E-mail:

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[email protected].

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Jianhua Guo: Tel.: + 61 7 3346 3222; Fax: + 61 7 3365 4726; E-mail:

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[email protected].

authors.

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ACS Paragon Plus Environment


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ABSTRACT: Chromate (Cr(VI)), as one of ubiquitous contaminants in groundwater,

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has posed a major threat to public health and ecological environment. Although

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various electron donors (e.g., organic carbon, hydrogen and methane) have been

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proposed to drive chromate removal from contaminated water, little is known for

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microbial chromate reduction coupled to elemental sulfur (S(0)) or zerovalent iron

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(Fe(0)) oxidation. This study demonstrated chromate could be biologically reduced by

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using S(0) or Fe(0) as inorganic electron donor. After 60-day cultivation, the sludge

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achieved a high Cr(VI) removal efficiency of 92.9 ± 1.1% and 98.1 ± 1.2% in two

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independent systems with S(0) or Fe(0) as the sole electron donor, respectively. The

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deposited Cr(III) was identified as the main reduction product based on X-ray

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photoelectron spectroscopy. High-throughput 16S rRNA gene sequencing indicated

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that Cr(VI) reduction coupled to S(0) or Fe(0) oxidation was mediated synergically by

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a microbial consortia. In such the consortia, S(0)- or Fe(0)-oxidizing bacteria (e.g.,

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Thiobacillus or Ferrovibrio) could generate volatile fatty acids as metabolites, which

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were further utilized by chromate-reducing bacteria (e.g., Geobacter or Desulfovibrio)

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to reduce chromate. Our findings advance our understanding on microbial chromate

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reduction supported by solid electron donors and also offer a promising process for

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groundwater remediation.

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TOC Art

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INTRODUCTION

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Although chromium (Cr) is required by human and other organisms as an essential

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micronutrient element, it becomes toxic and carcinogenic in the form of chromate

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(Cr(VI)).1 Cr(VI) is one of widely detected contaminants in groundwater.2 The natural

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concentration of Cr in groundwater is usually low (5 μg/L).3 However, serious Cr(VI)

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pollutions occurred in groundwater at numerous places over the past few decades,

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mainly due to Cr-containing wastewater released from tannery, metallurgical and

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nuclear industrial sectors.4,5 According to a survey conducted by Environmental

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Protection Agency (EPA) in the US, the concentration of Cr(VI) is above 10 μg/L in

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4% of groundwater systems, with the maximum concentration up to 97 μg/L.6 The

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exposure of Cr(VI) can cause severe adverse effects on human health, e.g., respiratory

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irritation, nasal or lung cancer, and damage to the gastrointestinal tract.5 In terms of

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its toxicity, the Cr(VI) concentration in drinking water is limited to 10 μg/L in

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California US.7

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Removing excessive Cr(VI) from groundwater has become an urgent

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environmental problem to be solved.5 Conventional remediation methods include

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physical adsorption, chemical reduction, ion exchange, and phytoremediation.8,9

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However, both physical and chemical processes are usually costly and may cause a

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secondary pollution (e.g., residual metal sludge), while the cycle time for

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phytoremediation is generally long (more than 4 months).5,10,11 Biological chromate

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removal methods provide a promising pathway for Cr(VI)-contaminated groundwater

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remediation.12 Recently, some strains such as Pseudomonas dechromaticans, 4


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Desulfovibrio vulgaris and Leucobacter sp. have been reported to possess potential

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capacities for microbial Cr(VI) reduction.13-15 Cr(VI) can be microbially reduced to

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Cr(III) when it is utilized as an electron acceptor by these microorganisms with

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organic carbons as electron donors.16 Compared to Cr(VI), Cr(III) is less toxic and

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shows limited mobility due to its low solubility in groundwater.17

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Due to the low bioavailability of organic carbons in groundwater, additional

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supplies of electron donors are usually required to support heterotrophic Cr(V)

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reduction. However, the extra addition of organic carbons may cause a secondary

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pollution.18 In addition, the formation of organic-Cr(III) complexes increases the

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solubility of Cr(III).19 Recently, microbial reduction of Cr(VI) has also been proposed

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by using gaseous substrates as electron donors, such as hydrogen or methane in terms

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of low operation cost and substantial availability.20 Although hydrogen- or methane-

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based methods are efficient to reduce Cr(VI) to Cr(III) and satisfactory results have

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been achieved,21,22 the safety issues resulting from the usage and storage of these

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explosive gases require precautionary measures before their wide applications in

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practice. Very recently, batch and continuous flow bioreactors utilizing S(0) or Fe(0)

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as electron donor have been evaluated to achieve removals of nutrients and toxic

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metals in groundwater.23-25 Since particulate S(0) is non-toxic and insoluble in water,

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it can be persistently fixed in biosystems, then release electrons slowly for microbes

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to remove contaminants. The application of S(0) as electron donor have advantages,

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including less biomass production and enhanced biological stability. Meanwhile, S(0)

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can be obtained from the waste byproduct of oil refining.26 Similarly, Fe(0) enables to

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react with water through iron corrosion process under anaerobic conditions, thus

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releasing hydrogen to drive microbial reactions.25 For example, Geets et al.27 added

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Fe(0) to permeable reactive barriers to support sulfate-reducing bacteria (SRB) to

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improve removal efficiencies of heavy metals (e.g., Cu and Zn). For in situ

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application of Fe(0) for groundwater remediation, it can reduce biomass clogging due

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to a lower biomass yield compared to using organic carbons as electron donors.28

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Nevertheless, the feasibility of S(0) or Fe(0) as the sole electron donor for microbial

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Cr(VI) reduction has not been well explored so far.

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The objective of this study is to investigate the potential of microbial Cr(VI)

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reduction coupled with S(0) or Fe(0) as the sole electron donor. Six lab-scale

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bioreactors were set up to test the feasibility of chromate removal after seeding

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anaerobic digestion sludge as inoculums. Firstly, we monitored chromate removal

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efficiency and identified the intermediates and final products of different systems with

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S(0) or Fe(0) alone as electron donors. Then, potential influencing factors (including

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initial Cr(VI) and bicarbonate concentrations) during the Cr(VI) reduction were

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assessed. Finally, the potential players for microbial reduction of Cr(VI) were

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identified based on 16S rRNA gene sequencing. In this study, microbial chromate

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reduction coupled to S(0) or Fe(0) oxidation was stably achieved after 60-day

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cultivation. The findings of this study shed a light on microbial reduction of Cr(VI)

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and could guide a process design to achieve groundwater remediation with Cr(VI)

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contamination.

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MATERIALS AND METHODS

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Bioreactor setting-up and the inoculum

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Six identical serum bottles with a working volume of 250 mL were set up as

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bioreactors to test chromate removal in this study. Each of bottle was sealed with

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butyl rubber stoppers and aluminum crimp caps to maintain a strict anaerobic

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condition. Initially, 50 mL anaerobic sludge obtained from an up-flow anaerobic

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sludge blanket reactor (Beijing YanJing Brewery Co., Ltd., China) were inoculated

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into each bioreactor. These six bioreactors were divided into three groups equally

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(each group with duplicate reactors). The first group (B-S) was operated with 5 g S(0)

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supplement as electron donors, while the second group (B-Fe) with 5 g Fe(0) addition.

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Neither S(0) nor Fe(0) was added into the third group (B-I) in order to confirm

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whether chromate reduction was available under the absence of electron donor. In

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addition, extra two reactors filled with the same amount of S(0) and Fe(0) but without

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inoculum were named as “C-S” and “C-Fe” as abiotic controls, respectively.

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The original cultivation was done in six bioreactors with 200 mL synthetic

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groundwater containing the following components per liter: 0.504 g NaHCO3, 0.2464

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g CaCl2, 0.035 g NH4Cl, 1.0572 g MaCl2·6H2O, 0.4459 g NaCl, 0.0283 g KCl, 0.029

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g KH2PO4. In the medium, bicarbonate was supplemented as inorganic carbon sources

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and K2CrO4 with a given concentration was added to the bioreactors as the source of

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Cr(VI). Both S(0) and Fe(0) (Analytical reagent) were purchased from Sinopharm

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Chemical Reagent Co., Ltd. (Beijing, China) and used directly without further

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processing. The whole experiments were performed in an anaerobic chamber, which 7



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avoided S(0) and Fe(0) oxidation. According to the manufacture information, both

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S(0) and Fe(0) have a similar range of particle size (i.e., 0.8-4.0 mm).

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Experimental procedure

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Long-term incubation was conducted in all six bioreactors by feeding 200 mL

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synthetic groundwater with 50 mg/L Cr(VI) at the beginning of each cycle (120 h),

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refreshing the aqueous solution every 120 h. In each bioreactor, 5 g S(0) or 5 g Fe(0)

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was supplemented initially at the incubation stage. After 60-d incubation, each

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bioreactor achieved a steady state at room temperature (22 ± 2 ºC). Soluble organics

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existing in the original inoculum were exhausted after 60-d cultivation in terms of

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total organic of less than 5 mg/L. Then three consecutive cycle studies were

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conducted to evaluate the performance of microbial reduction of Cr(VI) with S(0) and

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Fe(0) as the sole electron donor, respectively. For B-S and B-Fe systems, 5 g S(0) and

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5 g Fe(0) were freshly supplied to B-S and B-Fe at the beginning of these three

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consecutive cycles, respectively. In addition, both Cr(VI) and bicarbonate were

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provided to keep initial Cr(VI) concentration of 50 mg/L and initial bicarbonate

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concentration of 360 mg/L. During the cycle study, 5 mL supernatants in bioreactors

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were collected every 12 h and filtered through 0.22 μm membrane filter to determine

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the Cr(VI) concentrations and identify reaction metabolites. Furthermore, in order to

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investigate the effect of solution conditions on the microbial Cr(VI) reduction, batch

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experiments with different initial Cr(VI) concentrations (25, 50, 75, and 100 mg/L)

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and various initial bicarbonate concentrations (0, 180, 360, and 540 mg/L) were

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carried out in triplicates. 8


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Analytical methods

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The concentration of Cr(VI) was measured by the colorimetric method by using an

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UV-visible spectrophotometer (DR 6000, HACH, USA) at 540 nm.29 The

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determination of dissolved total Cr was performed by an Inductively Coupled Plasma

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Mass Spectrometry (ICP-MS, X Series II, Thermo Fisher Scientific, USA). The pH

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level was determined using a pH-201 meter (Hanna, Italy). The sulfate, sulfite and

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thiosulfate concentrations in B-S were measured using an ion chromatography (ICS-

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1100, Thermo Fisher Scientific, USA ). Gas chromatograph (Agilent 4890, J&W

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Scientific, USA) equipped with a flame ionization detector was used to identify the

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potential reaction intermediates (mainly volatile fatty acids, VFAs) produced in

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bioreactors. The morphology of solid S(0) and Fe(0) were observed through scanning

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electron microscope (SEM) with energy dispersive X-ray (EDS) operated at 20 kV

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(JEOL JAX-840, Hitachi Limited, Japan). The precipitates adhered to the bioreactors

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were collected and characterized by X-ray photoelectron spectroscopy (XPS)

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(XSAM-800, Kratos, UK).

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Microbial analysis

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In order to identify potential players for microbial chromate reduction, sludge samples

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(0.5 g) were collected from B-S and B-Fe systems after the three consecutive cycles

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for high-throughput 16S rRNA gene sequencing. Microbial DNA was extracted by

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using FastDNA® SPIN Kit for Soil (Qiagen, CA, the USA) according to

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manufacturer’s protocols. We used primers 338F (ACTCCTACGGGAGGC

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AGCAG) and 806R (GGACTACHVGGGTWTCTAAT) to amplify the V4-V5

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regions of the bacteria 16S ribosomal RNA gene (GeneAmp® 9700, ABI, the

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USA).30 The primers 524F10ext (TGYCAGCCGCCGCGGTAA) and Arch958R

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(YCCGGCGTTGAVTCCAATT) were used to amplify archaea. The PCR

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amplification and high-throughput 16S rRNA gene sequencing were performed by

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Majorbio Technology (Shanghai, China) on an Illumina HiSeq4000 platform

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(Illumina, Inc., San Diego, CA, USA). Based on 16S rRNA gene amplicon

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sequencing data, the microbial community structure was analyzed by following the

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pipeline described previously by Zhang et al.23

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RESULTS AND DISCUSSION

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Microbial Cr(VI) reduction performance

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With an initial Cr(VI) concentration of 50 mg/L, Figure 1 shows the variations of

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Cr(VI) and total Cr concentrations over time in three consecutive cycles for both of B-

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S and B-Fe systems after 60-d cultivation. When S(0) or Fe(0) was supplied as the

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sole electron donor, both B-S and B-Fe systems exhibited a high Cr(VI) removal

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efficiency, reaching 92.9 ± 1.1% and 98.1 ± 1.2%, respectively. The Cr(VI) removal

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efficiencies in both B-S and B-Fe were comparable with the previous results by using

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organic electron donors (e.g. acetate).31 The profile of total Cr concentration in the

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supernatant of B-S and B-Fe followed the similar trend with the profile of Cr(VI)

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(Figure 1). The obtained removal efficiency of total Cr in B-S and B-Fe was 88.9 ±

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1.2% and 97.4 ± 0.9%, respectively. Compared to Cr(VI) reduction rate ( 0.39 ±

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0.04 mg/L·h in B-S and 0.42 ± 0.02 mg/L·h in B-Fe), the obtained total Cr reduction

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rate was slower (0.38 ± 0.07 mg/L·h in B-Fe and 0.41 ± 0.03 mg/L·h). In contrast,

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only 3.9 ± 0.9% of Cr(VI) was removed in B-I without any supply of S(0) or Fe(0)

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(Figure S1), confirming that Cr(VI) reduction is associated with the presence of S(0)

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or Fe(0) in the proposed bioprocess. In addition, abiotic experiments without biomass

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(C-S and C-Fe) were conducted to verify whether chemical reduction also made

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contributions for chromate removal. Results showed that the C-S system without

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biomass did not exhibit any Cr(VI) removal capacity (Figure S1), ruling out the

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contribution of chemical reaction in the B-S system. Differently, Cr(VI) removal

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efficiency reached 14.8 ± 0.7% in the first cycle of C-Fe, while this percentage

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decreased to 7.9 ± 0.5% after 3-cycle operation (Figure S1a). Eventually, only 1.9 ±

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0.3% of Cr(VI) was removed after 60-day operation (Figure S1b). Therefore, a higher

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removal efficiency of Cr(VI) in B-Fe (98.1 ± 1.2%) might be partially associated with

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the combined microbial reduction and chemical reaction. Chemical reduction of

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chromate might be driven by electrons released from Fe(0) (Reaction 1).32 Iron

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corrosion could also occur and the corresponding products (e.g., H2 and Fe2+) would

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further contribute to Cr(VI) reduction (Reactions 2-4).33

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CrO42- + Fe(0) + 4H2O → Cr3+ + Fe3+ + 8OH-

(1)

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Fe(0) + 2H2O → H2 ↑ + Fe2+ + 2OH-

(2)

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CrO42- + 1.5H2 + H2O → Cr3+ + 5OH-

(3)

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CrO42- + 3Fe2+ + 4H2O → Cr3+ + 3Fe3+ + 8OH-

(4)

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It should be noted that a distinct decrease of chemical chromate reduction from

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Day 1 (14.8 ± 0.7%) to Day 60 (1.9 ± 0.3%) was due to the Fe(0) passivation, which

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was evidenced by iron oxides coating on the surface according to EDS spectrums and

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SEM images. The EDS results showed that the initial S(0) and Fe(0) are pure (Figure

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S2a), while iron oxides were observed in C-Fe after the three operating cycles (Figure

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S2b) and 60-day operation (Figure S2c). The SEM images showed that the surface of

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S(0) in C-S appeared to be uniform and smooth (Figure S3a). In C-Fe, botryoidal

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clusters depositions on the surface of Fe(0) were found (Figure S3b), which might be

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precipitates of iron oxides derived from chemical oxidation of Fe(0) by Cr(VI).34 This

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Fe(0) passivation could inhibit chemical reactions in which Fe(0) acted as electron

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donor for Cr(VI) reduction. Obvious morphology variations of S(0) and Fe(0) in

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bioreactors were also observed. Surfaces of S(0) and Fe(0) in B-S and B-Fe were

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relatively rough, with microbes attaching on the surface (Figure S3c and d). Microbes

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used S(0) and Fe(0) as electron donors to accelerate the Cr(VI) reduction.26, 35 The

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EDS spectrums also indicated solid Cr component was formed on the surface of S(0)

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and Fe(0) in both B-S and B-Fe systems (Figure S2d).

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Identification of final products

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It was observed that gray-blue precipitates appeared in the solution of B-S and B-Fe

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(Figure S4). The produced precipitates were identified by XPS. As shown in Figure

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2a, two distinct peaks appeared between 576.0-578.0 eV and 586.0-588.0 eV,

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corresponding to Cr 2p3/2 and Cr 2p1/2, respectively. Such the spectrum indicated

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that the precipitates were Cr(III) species.36 The XPS results further indicated that Cr3+

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from biotransformation of Cr(VI) formed Cr(III) precipitates immediately after its 12


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production in bioreactors, which was consistent with the main components of

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Cr(OH)3 and Cr2O3 reported previously.36 In addition, the produced Cr(III) could also

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form complexes with extracellular polymeric substances (EPS) secreted by

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microorganisms.37 Considering Cr(III) is less toxic and insoluble under near neutral

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pH condition,17 the remediation process based on S(0) or Fe(0) could be practically

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useful to treat Cr(VI) contaminated groundwater.

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In order to investigate whether electron donors, including both Fe(0) and S(0),

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were oxidized or not, both sulfur and iron species were monitored during microbial

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chromate reduction. The sulfate concentration gradually increased in a cycle in B-S

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(Figure 2b), indicating the transformation of S(0) to sulfate. This also confirmed that

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microbial Cr(VI) reduction was coupled with S(0) oxidation, similar to a previous

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study.24 Other sulfur-related intermediates such as sulfite and thiosulfate were

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undetected in B-S. Regarding the B-Fe system, soluble iron species in the supernatant

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were hardly found, while brown precipitates adhering to the bottle were observed

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(Figure S4). The XPS analysis showed these precipitates had two typical peaks

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assigned to Fe 2p1/2 (725.0-727 eV) and Fe 2p3/2 (710.0-712.5 eV) (Figure 2c),

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which are similar to the peaks of Fe3O4 oxides obtained in a previous study.38 This

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result suggested that Fe(0) might have been oxidized into Fe3O4, which could release

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energy to synthesize intermediates for microbial Cr(VI) reduction.

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Accumulations of VFAs were observed within a cycle for 120 h (Figure 2d), in

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which 8.63 ± 0.15 mg/L and 18.09 ± 0.44 mg/L VFAs were produced in B-S and B-

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Fe, respectively. Isobutyrate was found as the main form of residual VFAs in both B-

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S and B-Fe, which is consistent with our previous study in S(0)- or Fe(0)-based

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biological vanadate reduction.23 Differently, acetate was observed as the dominant

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component of VFAs in methane-based bromate or nitrate removals.39,40 The total

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concentration of residual VFAs in B-Fe was higher than that in B-S, which might lead

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to a faster Cr(VI) removal rate. In autotrophic biosystems, VFAs could be derived

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from bicarbonate reduction or carbon dioxide fixation.41,42 In this bioprocess,

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autotrophs employed bicarbonate or carbon dioxide to synthesize VFAs via the Calvin

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cycle.43,44 These synthesized VFAs could be utilized as electron donors to achieve

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Cr(VI) reduction.

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The pH decreased gradually in B-S within a typical operating cycle (120 h),

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which is due to the generation of hydrogen ion during the S(0) bio-oxidation

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process24. In contrast, the pH profile showed an ascending trend in B-Fe (Figure S5),

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as hydroxyl ion can be produced from both chemical corrosion and biological

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oxidation of Fe(0).25 The decreasing bicarbonate concentration in both biosystems

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indicated that bicarbonate acted as an inorganic carbon source for autotrophic

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microbes to synthesize VFAs, which supported microbial Cr(VI) reduction with S(0)

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and Fe(0) as electron donors, respectively.

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Process elucidation and evaluation

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The processes of reduction of Cr(VI) coupled with S(0) and Fe(0) oxidations

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(Reactions 5 and 6, respectively) were proposed in terms of determined reaction

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products, and stoichiometric relationship:

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CrO42- + 10.5S0 + 15HCO3- + 3NH4+ → 3C5H7NO2 + Cr3+ + 10.5SO42- + H2O + 4H+ 14


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(5)

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CrO42- + 1.5Fe0 +0.375HCO3- + 0.075NH4+ + 4.825H2O → 0.075C5H7NO2 + Cr3+ +

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1.5Fe(OH)3↓ + 5.3OH(6)

284 285

Both mass and electron balances were conducted to elucidate these two

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processes. In B-S, the measured sulfate concentration (93.7 ± 1.6 mg/L) within in a

287

typical operating cycle (120 h), indicating that solid phase S(0) could be oxidized to

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SO42-.45,46 The released electrons resulting from S(0) oxidation to sulfate was 1.46 ±

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0.03 mmol, which is higher than 0.69 ± 0.06 mmol electrons required for transfering

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Cr(VI) to Cr(III). It is assumed that approximately 47.3% of released electrons were

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transferred from S(0) to Cr(VI) in one cycle, while the remaining electrons might

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support microbial growth and intermediates synthesis.23 According to Reaction 5, the

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theoritical required bicarbonate were 839.5 ± 73.2 mg/L in B-S based on the

294

measured reduction of Cr(VI), while the measured bicarbonate consumption was

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353.2 ± 4.1 mg/L. The insufficient bicarbonate consumption in B-S might be

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compensated by carbon dioxide production from anaerobic respiration of accumulated

297

organic intermediates.47

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It is too difficult to perform the electron balance in B-Fe, because no soluble iron

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species was availably detected. The consumption of bicarbonate was monitored in B-

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Fe. It is found that a larger amount of bicarbonate (177.1 ± 0.9 mg/L) was reduced

301

than the theoretical value (21.9 ± 1.8 mg/L) in B-Fe, likely due to the growth of

302

autotrophic bacteria and the transformation of bicarbonate to carbonate along with

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increasing of pH. Furthermore, the biomass production was stoichiometrically 0.16 g

304

to remove 1 g Cr(VI) supported by Fe(0) (Reaction 6), which was much lower than

305

that in bioreactors using lactose and cheese whey as electron donors for Cr(VI)

306

reduction.48 The low biomass production of this reaction was also comparable to the

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cell yield of the autotrophic microorganisms (ranging 0.073 to 0.280 g cell dry weight

308

per gram chemical oxygen demand equivalent of hydrogen).23 Such a low biomass

309

yield might be a significant advantage of our proposed biosystems using Fe(0) as an

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electron donor, since aquifer clogging in organic carbon-based systems could be

311

alleviated.49

312

Influencing factors examination

313

The effects of initial Cr(VI) and bicarbonate concentration on microbial Cr(VI)

314

reduction performance were further investigated. The initial Cr(VI) concentrations

315

would determine the reaction duration to achieve a complete Cr(VI) reduction to the

316

final Cr(VI) concentration below the detection limit. It was observed that the

317

complete reduction of Cr(VI) with concentration of 25 mg/L required approximately

318

72 h for both B-S and B-Fe, whereas the time needed for a complete removal of 100

319

mg/L Cr(VI) took more than 120 h in both systems (Figure 3a and 3b). However, the

320

maximum reduction rate increased from 0.70 ± 0.11 mg/L·h to 1.59 ± 0.17 mg/L·h for

321

B-S, and the rate increased from 0.69 ± 0.02 mg/L·h to 2.73 ± 0.07 mg/L·h for B-Fe.

322

These results demonstrated that microbial chromate reduction processes with S(0) and

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Fe(0) as electron donors were very robust to treat a wide range of Cr(VI)

324

concentrations (25 to 100 mg/L). In addition, B-Fe seems to be more robust towards 16


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higher Cr(VI) concentration than B-S. Increasing bicarbonate concentration could provide sufficient carbon source for

327

microbes to promote Cr(VI) reduction in both B-S and B-Fe.50 The removal efficiency

328

of Cr(VI) was 70.1 ± 1.5% and 80.4 ± 1.2% in B-S and B-Fe, respectively, without

329

the addition of bicarbonate. With the increase of initial bicarbonate concentration, the

330

reduction rate and the removal efficiency of Cr(VI) in B-S and B-Fe were enhanced.

331

When the bicarbonate concentration increased from 0 mg/L to 540 mg/L, the

332

percentage of Cr(VI) removal in B-S and B-Fe increased by 26.4 ± 1.1% and 19.0 ±

333

1.3% respectively (Figure 3c and 3d). Meanwhile, the removal rates increased from

334

0.29 ± 0.01 to 0.40 ± 0.02 mg/L·h and from 0.34 ± 0.01 to 0.42 ± 0.01 mg/L·h with

335

the same bicarbonate concentration change, respectively (Figure 3c and 3d).

336

Microbial community structure

337

In order to compare microbial community structure at different conditions, 16S rRNA

338

gene sequencing was performed for both the inoculum and two biomass samples (one

339

from B-S and another from B-Fe) after 75-day cultivation. Compared to the inoculum,

340

microbial richness was decreased in both B-S and B-Fe systems in terms of

341

rarefaction curves (Figure S6). In particular, the B-S system harbored less diverse

342

microbial communities in terms of OTU numbers, Simpson and Shannon indexes,

343

suggesting microbial communities tended to be highly selected (Table S1). However,

344

microbial diversity increased in B-Fe, implying more species existed due to Fe(0) as

345

well as its corrosion products.51

346

Compared to the inoculum, microbial communities changed significantly at the 17



347

class level after feeding solid electron donors (Figure 4a). The predominant classes in

348

B-S were Betaproteobacteria, Gammaproteobacteria and Bacteroidia, with a relative

349

abundance of 31.1%, 22.8% and 18.3%, respectively. These three representative

350

classes accounted for around 72.1% of the total bacterial population in B-S, while

351

accounted for only about 14.1% in the inoculum. Meanwhile, a new class Bacilli with

352

a relative abundance of 4.17% appeared in B-S, which was not detected in either

353

inoculum or B-Fe. In B-Fe, Alphaproteobacteria, Spirochaetes and

354

Betaproteobacteria were enriched by 2.5%, 5.3% and 10.9% compared to the

355

inoculum, with the proportions of Anaeroline and Gammaproteobacteria remained

356

basically constant.

357

Potential bacteria associated with Cr(VI) reduction and oxidation of S(0) and

358

Fe(0) were further analyzed at the genus level (Figure 4b). The relative abundance of

359

Geobacter was increased both in B-S and B-Fe, which had been reported performed

360

well in situ bioremediation performance under high concentration of Cr(VI)

361

contamination.52 Desulfovibrio was enriched in B-Fe and its ability of reducing

362

Cr(VI) had been previously confirmed.53 In B-Fe, an iron-oxidizing bacteria

363

Ferrovibrio was accumulated, which used Fe(0) as direct electron donor.54 The

364

increased Syntrophus (0.86%) might be involved in the production and consumption

365

of VFAs.55 In B-S, chemoautotrophic Thiobacillus increased significantly from 0.25%

366

in the inoculum to 17.1% after 75-day cultivation. It has been reported that

367

Thiobacillus could oxidize S(0) to release electrons and gain energy to synthesize

368

metabolites to reduce Cr(VI).56,57 The accumulated Blvii28 (17.9%) might play an

18


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Page 19 of 37


369

important role in autotrophic S(0) oxidation in B-S, which has been reported to be

370

able to generate hydrogen for S(0) autotrophic microbes.58

371

In both B-S and B-Fe, archaeal communities were also observed to shift

372

significantly. Methanobacteria became the only dominating archaea at the class level

373

in B-S and B-Fe, compared to the diverse archaeal species in the inoculum (Figure

374

S7a). The genus Methanobacterium was less in the inoculum (7.6%) but

375

predominated in B-S (91.0%) and B-Fe (81.0%) (Figure S7b). It was reported that

376

Methanobacterium have the potential of detoxifying Cr(VI) through bio-reduction.59

377

Potential microbial C(VI) reduction mechanism and environmental implications

378

In the light of the performances of batch tests, intermediate identification and microbial

379

community structure, a synergistic mechanism for the reduction of Cr(VI) coupled with

380

oxidation of S(0) and Fe(0) was proposed (Figure 5). Firstly, autotrophic microbes such

381

as Thiobacillus and Ferrovibrio might have synthesized VFAs as byproducts through

382

bicarbonate reduction with energy from oxidation of S(0) to sulfate or Fe(0) to

383

Fe(II)/Fe(III), respectively.54,56,57 Secondly, heterotrophic Cr(VI)-reducing microbes

384

such as Geobacter in B-S and Desulfovibrio in B-Fe would reduce Cr(VI) by using

385

VFAs as electron donors and carbon sources. Both extracellular and intracellular

386

reduction of Cr(VI) could occur, since mixed anaerobic culture was employed.11 Fox

387

example, Geobacter could reduce Cr(VI) extracellularly with the aid of c type

388

cytochromes on out membrane.60 Finally, the reduction products, Cr(III) would be

389

precipitated naturally in a near-neutral pH environment. Similar synergistic

390

mechanisms had also been reported in the bio-reduction of bromate driven by 19



391

methane.39 However, it should be noted that more studies are needed to elucidate the

392

pathways involved and reveal the functions of the key members of a complex microbial

393

community in our S(0)- and Fe(0)-based systems through meta-omics (e.g.,

394

metagenomics and metatranscriptomics).

395

Considering Cr(VI) contamination in aquifer becomes serious, bioremediation

396

has been proposed as a promising pathway to alleviate its negative environmental

397

impacts.61 This study demonstrates for the first time that Cr(VI) could be biologically

398

reduced to Cr(III) coupled with oxidation of S(0) or Fe(0) with an efficient removal

399

rate. We further compared chromate removal performance with different electron

400

donors in literature. Cr(VI) reduction rate in both B-S (0.39 ± 0.04 mg/L·h) and B-Fe

401

(0.42 ± 0.02 mg/L·h) was much higher than that obtained with gaseous electron

402

donors (0.003 mg/L·h for hydrogen and 0.01~0.06 mg/L·h for methane),19, 21 and

403

comparable to the rate obtained using organics, such as acetate (0.62 mg/L·h) and

404

glucose (0.60 mg/L·h) (Table S2).31, 62 Moreover, employing solid inorganic electron

405

donors such as S(0) and Fe(0) can minimize possible secondary contamination and

406

aquifer clogging induced by organic substrates injection. The proposed bioprocesses

407

can be implemented as bio-permeable reactive barriers (bio-PRB) for efficient

408

bioremediation of Cr(V) contaminated groundwater. Bio-PRB is an in-situ

409

remediation project based on a wall of porous material cross the path of contaminated

410

groundwater flow which mainly removes contaminants by biodegradation combined

411

with adsorption, oxidation/reduction and other processes. 63 In such the bio-PRB

412

system, indigenous microorganisms can be accumulated and solid inorganic electron

20


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Page 21 of 37


413

donors can be periodically replaced to maintain a high-efficiency. In addition, nitrate,

414

as a frequent co-contaminant in groundwater, might compete with chromate to utilize

415

electron donor, thus decreasing Cr(VI) removal efficiency,64,65 which should be

416

further investigated in the future.

417

Supporting Information

418

The bacterial richness and diversity of inoculated sludge and bioreactors;

419

comparison of common bioprocesses for Cr(VI) removal with different electron

420

donors; changes of Cr(VI) in C-S, C-Fe and B-I at different stages; EDS patterns

421

of S(0) and Fe(0) at initial states and after use at different states in employed

422

reactors; corresponding SEM images of solid electron donor in reactors; images

423

of the precipitates generated in employed bioreactors during operation; changes

424

of pH and bicarbonate concentration in employed bioreactors within a typical

425

operating cycle; rarefaction curves for species abundance in the inoculum,

426

biomass in B-S and B-Fe; archaeal community compositions.

427

ACKNOWLEDGEMENTS

428

This research work was supported by the National Natural Science Foundation of

429

China (NSFC) (No. 41672237, No. 91647115), Beijing Nova Program (No.

430

Z171100001117082) and the Fundamental Research Funds for the Central

431

Universities (No. 2652018193).

432

NOTES

433

The authors declare no competing financial interest.

21



434

REFERENCES

435

(1) Li, J. X.; Zhang, X. Y.; Liu, M. C.; Pan, B. C.; Zhang, W. M.; Shi, Z.; Guan, X.

436

H. Enhanced reactivity and electron selectivity of sulfidated zerovalent iron

437

toward chromate under aerobic conditions. Environ. Sci. Technol. 2018, 52,

438

2988-2997.

439

(2) Jamieson-Hanes, J. H.; Gibson, B. D.; Lindsay, M. B. J.; Kim, Y.; Ptacek, C. J.;

440

Blowes, D. W. Chromium isotope fractionation during reduction of Cr(VI) under

441

saturated flow conditions. Environ. Sci. Technol. 2012, 46, 6783-6789.

442

(3) Lou, S.; Liu, S. G.; Dai, C. M.; Tao, A.; Tan, B.; Ma, G. F.; Chalov, R. S.;

443

Chalov, S. R. Heavy metal distribution and groundwater quality assessment for a

444

coastal area on a Chinese. Island. Pol. J. Environ. Stud. 2017, 26, 733-745.

445

(4) Zhang, B. G.; Feng, C. P.; Ni, J. R.; Zhang, J.; Huang, W. L. Simultaneous

446

reduction of vanadium (V) and chromium(VI) with enhanced energy recovery

447

based on microbial fuel cell technology. J. Power Sources 2012, 204, 34-39.

448

(5) Singh, R.; Dong, H. L.; Liu, D.; Zhao, L. D.; Marts, A. R.; Farquhar, E.; Tierney,

449

D. L.; Almquist, C. B.; Briggs, B. R. Reduction of hexavalent chromium by the

450

thermophilic methanogen Methanothermobacter thermautotrophicus. Geochim.

451

Cosmochim. Acta 2015, 148, 442-456.

452 453

(6) Gifford, M.; Chester, M.; Hristovski, K.; Westerhoff, P. Human health tradeoffs in wellhead drinking water treatment: Comparing exposure reduction to 22


Page 22 of 37

Page 23 of 37

454


embedded life cycle risks. Water Res. 2017, 128, 246-254.

455

(7) Pan, C.; Troyer, L. D.; Liao, P.; Catalano, J. G.; Li, W.L.; Giammar, D. E. Effect

456

of humic acid on the removal of chromium(VI) and the reduction of solids in iron

457

electrocoagulation. Environ. Sci. Technol. 2017, 51, 6308-6318.

458

(8) Kathiravan, M. N.; Karthick, R.; Muthukumar, K. Ex situ bioremediation of

459

Cr(VI) contaminated soil by Bacillus sp.: Batch and continuous studies. Chem.

460

Eng. J. 2011, 169, 107-115.

461

(9) Huang, H. Y.; Wu, K. J.; Khan, A.; Jiang, Y. M.; Ling, Z. M.; Liu, P.; Chen, Y.;

462

Tao, X. Y.; Li, X. K. A novel Pseudomonas gessardii strain LZ-E simultaneously

463

degrades naphthalene and reduces hexavalent chromium. Bioresour. Technol.

464

2016, 207, 370-378.

465

(10)Adiloglu, S.; Sume, M.T.S.A. Chrome (Cr) pollution in agricultural areas

466

improvement by phytoremediation method with canola (Brassica napus L.) plant

467

growing. J. Essent. Oil-Bear. Plants 2015, 18, 1180-1186.

468

(11)Wang, S.; Zhang, B. G.; Diao, M. H.; Shi, J. X.; Jiang, Y. F.; Cheng, Y. T.; Liu,

469

H. Enhancement of synchronous bio-reductions of vanadium (V) and chromium

470

(VI) by mixed anaerobic culture. Environ. Pollut. 2018, 242, 249-256.

471 472

(12)Fruchter, J. In situ treatment of chromium-contaminated groundwater. Environ. Sci. Technol. 2002, 36, 464A-472A.

23



473

(13)Romanenko, V.; Korenkov, V. N. A pure culture of bacterial cells assimilating

474

chromates and bichromates as hydrogen acceptors when grown under anaerobic

475

conditions. Mikrobiologiia 1977, 46, 414-417.

476

(14)Slobodkina, G. B.; Bonch-Osmolovskaia, E. A.; Slobodkin, A. I. Reduction of

477

chromate, selenite, tellurite, and iron(III) by the moderately thermophilic

478

bacterium Bacillus thermoamylovorans SKC1. Microbiology. 2007, 76, 530-534.

479

(15)Butler, E. C.; Chen, L. X.; Hansel, C. M.; Krumholz, L. R.; Madden, A. S. E.;

480

Lan, Y. Biological versus mineralogical chromium reduction: potential for

481

reoxidation by manganese oxide. Environ. Sci.: Processes Impacts 2015, 17,

482

1930-1940.

483

(16)Prevot, A. B.; Ginepro, M.; Peracaciolo, E.; Zelano, V.; DeLuca, D. A. Chemical

484

vs bio-mediated reduction of hexavalent chromium. An in-vitro study for soil and

485

deep waters remediation. Geoderma 2018, 312, 17-23.

486

(17)Wang, W.; Zhang, B. G.; Liu, Q. S.; Du, P. H.; Liu, W.; He, Z. Biosynthesis of

487

palladium nanoparticles using Shewanella loihica PV-4 for excellent catalytic

488

reduction of chromium (VI). Environ. Sci. Nano 2018, 5, 730-739.

489

(18)Mamais, D.; Noutsopoulos, C.; Kavallari, I.; Nyktari, E.; Kaldis, A.; Panousi, E.;

490

Nikitopoulos, G.; Antoniou, K.; Nasioka, M. Biological groundwater treatment

491

for chromium removal at low hexavalent chromium concentrations. Chemosphere

492

2017, 152, 238-244.

24


Page 24 of 37

Page 25 of 37

493


(19)Lai, C. Y.; Zhong, L.; Zhang, Y.; Chen, J. X.; Wen, L. L.; Shi, L. D.; Sun, Y. P.;

494

Ma, F.; Rittmann, B. E.; Zhou, C. Bioreduction of chromate in a methane-based

495

membrane biofilm reactor. Environ. Sci. Technol. 2016, 50, 5832-5839.

496

(20)Jiang, Y. F.; Zhang, B. G.; He, C.; Shi, J. X.; Borthwick, A. G. L.; Huang, X. Y.

497

Synchronous microbial vanadium (V) reduction and denitrification in

498

groundwater using hydrogen as the sole electron donor. Water Res. 2018, 141,

499

289-296.

500

(21)Chung, J.; Nerenberg, R.; Rittmann, B. E. Bio-reduction of soluble chromate

501

using a hydrogen-based membrane biofilm reactor. Water Res. 2006, 40, 1634-

502

1642.

503

(22)Lu, Y. Z.; Chen, G. J.; Bai, Y. N.; Fu, L.; Qin, L. P.; Zeng, R. J. X. Chromium

504

isotope fractionation during Cr(VI) reduction in a methane-based hollow-fiber

505

membrane biofilm reactor. Water Res. 2018, 130, 263-270.

506

(23)Zhang, B. G.; Qiu, R.; Lu, L.; Chen, X.; He, C.; Lu, J. P.; Ren, Z. J. Autotrophic

507

vanadium(V) bioreduction in groundwater by elemental sulfur and zerovalent

508

iron. Environ. Sci. Technol. 2018, 52, 7434-7442.

509

(24)Sahinkaya, E.; Kilic, A. Heterotrophic and elemental-sulfur-based autotrophic

510

denitrification processes for simultaneous nitrate and Cr(VI) reduction. Water

511

Res. 2014, 50, 278-286.

512

(25)Karri, S.; Sierra-Alvarez, R.; Field, J.A. Zero valent iron as an electron-donor for 25



513

methanogenesis and sulfate reduction in anaerobic sludge. Appl. Biochem.

514

Biotechnol. 2005, 92, 810-819.

515

(26)Sahu, A. K.; Conneely, T.; Nusslein, K. R.; Ergas, S. J. Biological perchlorate

516

reduction in packed bed reactors using elemental sulfur. Environ. Sci. Technol.

517

2009, 43, 4466-4471.

518

(27)Geets, J.; Borremans, B.; Vangronsveld, J.; Diels, L.; Van der Lelie, D.

519

Molecular monitoring of SRB community structure and dynamics in batch

520

experiments to examine the applicability of in situ precipitation of heavy metals

521

for groundwater remediation. J. Soils Sediments 2005, 5, 149-163.

522

(28)Lai, K. C. K.; Lo, I. M. C. Removal of chromium (VI) by acid-washed zero-

523

valent iron under various groundwater geochemistry conditions. Environ. Sci.

524

Technol. 2008, 42, 1238-1244.

525

(29)Shao, Q. Q.; Xu, C. H.; Wang, Y. H.; Huang, S. S.; Zhang, B. L.; Huang, L. H.;

526

Fan, D. M.; Tratnyek, P. G. Dynamic interactions between sulfidated zerovalent

527

iron and dissolved oxygen: Mechanistic insights for enhanced chromate removal.

528

Water Res. 2018, 135, 322-330.

529

(30)Cao, X. L.; Diao, M. H.; Zhang, B. G.; Liu, H.; Wang, S.; Yang, M. Spatial

530

distribution of vanadium and microbial community responses in surface soil of

531

Panzhihua mining and smelting area, China. Chemosphere 2017, 183, 9-17.

532

(31)Reddy, G. K. K.; Nancharaiah, Y. V. Sustainable bioreduction of toxic levels of 26


Page 26 of 37

Page 27 of 37


533

chromate in a denitrifying granular sludge reactor. Environ. Sci. Pollut. Res.

534

2017, 25, 1969-1979.

535

(32)Lo, I. M. C.; Lam, C. S. C.; Lai, K. C. K. Hardness and carbonate effects on the

536

reactivity of zero-valent iron for Cr(VI) removal. Water Res. 2006, 40, 595-605.

537

(33)Zhong, J. W.; Yin, W. Z.; Li, Y. T.; Li, P.; Wu, J. H.; Jiang, G. B.; Gu, J. J.;

538

Liang, H. Column study of enhanced Cr(VI) removal and longevity by coupled

539

abiotic and biotic processes using Fe-0 and mixed anaerobic culture. Water Res.

540

2017, 122, 536-544.

541

(34)Mitra, P.; Sarkar, D.; Chakrabarti, S.; Dutta, B. K. Reduction of hexavalent

542

chromium with zero-valent iron: Batch kinetic studies and rate model. Chem.

543

Eng. J. 2011, 171, 54-60.

544

(35)Yu, X. Y.; Amrhein, C.; Deshusses, M. A.; Matsumoto, M. R. Perchlorate

545

reduction by autotrophic bacteria attached to zerovalent iron in a flow-through

546

reactor. Environ. Sci. Technol. 2007, 41, 990-7.

547

(36)Han, R.; Li, F. B.; Liu, T. X.; Li, X. M.; Wu, Y. D.; Wang, Y.; Chen, D. D.

548

Effects of incubation conditions on Cr(VI) reduction by c-type cytochromes in

549

intact Shewanella oneidensis MR-1 cells. Front. Microbiol. 2016, 7, 746.

550

(37)Kantar, C.; Demiray, H.; Dogan, N. M. Role of microbial exopolymeric

551

substances (EPS) on chromium sorption and transport in heterogeneous

552

subsurface soils: I. Cr(III) complexation with EPS in aqueous solution. 27



553

554

Chemosphere 2011, 82, 1496-1505. (38)Tian, X. K.; Wang, W. W.; Tian, N.; Zhou, C. X.; Yang, C.; Komarneni, S.

555

Cr(VI) reduction and immobilization by novel carbonaceous modified magnetic

556

Fe3O4 /halloysite nanohybrid. J. Hazard. Mater. 2016, 309, 151-156.

557

(39)Luo, J. H.; Wu, M. X.; Yuan, Z. G.; Guo, J. H. Biological bromate reduction

558

driven by methane in a membrane biofilm reactor. Environ. Sci. Technol. Lett.

559

2017, 4, 562-566.

560 561

562 563

564

(40)Luo, J. H.; Chen, H.; Yuan; Z. G.; Guo, J. H. Methane-supported nitrate removal from groundwater in a membrane biofilm reactor. Water Res. 2018, 132, 71-78. (41)Tabita, F. R. The hydroxypropionate pathway of CO2 fixation: Fait accompli. Proc. Natl. Acad. Sci. U. S. A. 2009, 106, 21015-21016. (42)Estelmann, S.; Hugler, M.; Eisenreich, W.; Werner, K.; Berg, I. A.; Ramos-Vera;

565

W. H.; Say, R. F.; Kockelkorn, D.; Gadon, N.; Fuchs, G. Labeling and enzyme

566

studies of the central carbon metabolism in Metallosphaera sedula. J. Bacteriol.

567

2011, 193, 1191-1200.

568

(43)Herrmann, M.; Rusznyák, A.; Akob, D. M.; Schulze, I.; Opitz, S.; Totsche, K. U.;

569

Kusel, K. Large fractions of CO2-fixing microorganisms in pristine limestone

570

aquifers appear to be involved in the oxidation of reduced sulfur and nitrogen

571

compounds. Appl. Environ. Microbiol. 2015, 81, 2384-2394.

28


Page 28 of 37

Page 29 of 37

572


(44)Dobrinski, K. P.; Boller, A. J.; Scott, K. M. Expression and function of four

573

carbonic anhydrase homologs in the deep-sea chemolithoautotroph

574

Thiomicrospira crunogena. Appl. Environ. Microbiol. 2010, 76, 3561-3567.

575

(45)Zhang, B. G.; Zhao, H. Z; Zhou, S. G.; Shi, C. H.; Wang, C.; Ni, J. R. A novel

576

UASB-MFC-BAF integrated system for high strength molasses wastewater

577

treatment and bioelectricity generation. Bioresour. Technol. 2009, 100, 5687-

578

5693.

579

(46)Boles, A. R.; Conneely, T.; Mckeever, R.; Nixon, P.; Ergas, S. J. Performance of

580

a pilot-scale packed bed reactor for perchlorate reduction using a sulfur oxidizing

581

bacterial consortium. Biotechnol. Bioeng. 2012, 109, 637-646.

582

(47)Li, C. H.; Wong, Y. S.; Wang, H. Y.; Tam, N. F. Y. Anaerobic biodegradation of

583

PAHs in mangrove sediment with amendment of NaHCO3. J. Environ. Sci. 2012,

584

30, 148-156.

585

(48)Contreras, E. M.; Orozco, A. M.; Zaritzky, N. E. Biological Cr(VI) removal

586

coupled with biomass growth; biomass decay; and multiple substrate limitation.

587

Water Res. 2011, 45, 3034-3046.

588

(49)Braun, B.; Schroder, J.; Knecht, H.; Szewzyk, U. Unraveling the microbial

589

community of a cold groundwater catchment system. Water Res. 2016, 107, 113-

590

126.

591

(50)Tandukar, M.; Huber, S. J.; Onodera, T.; Pavlostathis, S. G. Biological 29



592

chromium(VI) reduction in the cathode of a microbial fuel cell. Environ. Sci.

593

Technol. 2009, 43, 8159-8165.

594

(51)Kirschling, T. L.; Gregory, K. B.; Minkley, E. G.; Lowry, G. V.; Tilton, R. D.

595

Impact of nanoscale zero valent iron on geochemistry and microbial populations

596

in trichloroethylene contaminated aquifer materials. Environ. Sci. Technol. 2010,

597

44, 3474-3480.

598

(52)Pei, Y. X.; Yu, Z. S.; Ji, J.; Khan, A.; Li, X. K. Microbial community structure

599

and function indicate the severity of chromium contamination of the Yellow

600

River. Front. Microbiol. 2018, 9, 38-50.

601

(53)Franco, L. C.; Steinbeisser, S.; Zane, G. M.; Wall, J. D.; Fields, M. W. Cr(VI)

602

reduction and physiological toxicity are impacted by resource ratio in

603

Desulfovibrio vulgaris. Appl. Microbiol. Biotechnol. 2018, 102, 2839-2850.

604

(54)Sorokina, A. Y.; Chernousova, E. Y.; Dubinina, G. A. Ferrovibrio denitrificans

605

gen. nov. sp. nov. a novel neutrophilic facultative anaerobic Fe(II)-oxidizing

606

bacterium. FEMS Microbiol. Lett. 2012, 335, 19-25.

607

(55)Deng, Y.; Huang, Z. X.; Ruan, W. Q.; Miao, H. F.; Shi, W. S.; Zhao, M. X.

608

Enriching ruminal polysaccharide-degrading consortia via co-inoculation with

609

methanogenic sludge and microbial mechanisms of acidification across

610

lignocellulose loading gradients. Appl. Microbiol. Biotechnol. 2018, 102, 1-12.

611

(56)Yang, Z. H.; StoVen, K.; Haneklaus, S.; Singh, B. R.; Schnug, E. Elemental 30


Page 30 of 37

Page 31 of 37


612

sulfur oxidation by Thiobacillus spp. and aerobic heterotrophic sulfur-oxidizing

613

bacteria. Pedosphere 2010, 20, 7-79.

614

(57)Ishii, T.; Nakagawa, H.; Hashimoto, K.; Nakamuraa, R. Acidithiobacillus

615

ferrooxidans as a bioelectrocatalyst for conversion of atmospheric CO2 into

616

extracellular pyruvic acid. Electrochemistry 2012, 80, 327-329.

617

(58)Su, X. L.; Tian, Q.; Zhang, J.; Yuan, X. Z.; Shi, X. S.; Guo, R. B.; Qiu, Y. L.

618

Acetobacteroides hydrogenigenes gen. nov. sp. nov. an anaerobic hydrogen-

619

producing bacterium in the family Rikenellaceae isolated from a reed swamp. Int.

620

J. Syst. Evol. Microbiol. 2014, 64, 2986-2991.

621

(59)Mosher, J. J.; Phelps, T. J.; Podar, M.; Hurt, R. A.; Campbell, J. H.; Drake, M.

622

M.; Moberly, J. G.; Schadt, C. W.; Brown, S. D.; Hazen, T. C.; Arkin, A. P.;

623

Palumbo, A. V.; Faybishenko, B. A.; Elias, D. A. Microbial community

624

succession during lactate amendment and electron acceptor limitation reveals a

625

predominance of metal-reducing Pelosinus spp. Appl. Environ. Microbiol. 2012,

626

78, 2082-2091.

627

(60)Gong, Y. F.; Werth, C. J.; He, Y. X.; Su, Y. M.; Zhang, Y. L.; Zhou, X. F.

628

Intracellular versus extracellular accumulation of hexavalent chromium reduction

629

products by Geobacter sulfurreducens PCA. Environ. Pollut. 2018, 240, 485-492.

630

(61)Fernandez, P. M.; Vinarta, S. C.; Bernal, A. R.; Cruz, E. L.; Figureueroa, L. I. C.

631

Bioremediation strategies for chromium removal: Current research, scale-up

31



632

approach and future perspectives. Chemosphere 2018, 208, 139-148.

633

(62)Qian, J.; Zhou, J.; Wang, L.; Wei, L.; Li, Q.; Wang, D.; Wang, Q. Direct Cr (VI)

634

bio-reduction with organics as electron donor by anaerobic sludge. Chem. Eng. J.

635

2017, 309, 330-338.

636

(63)Pagnanelli, F.; Cruz, V. C.; Mainelli, S.; Toro, L. Assessment of solid reactive

637

mixtures for the development of biological permeable reactive barriers. J.

638

Hazard. Mater. 2009, 170, 998-1005.

639

(64)Sahinkaya, E.; Kilic, A.; Calimlioglu, B.; Toker, Y. Simultaneous bioreduction of

640

nitrate and chromate using sulfur-based mixotrophic denitrification process. J.

641

Hazard. Mater. 2013, 26, 234-239.

642

(65)Zhong, L.; Lai, C. Y.; Shi, L. D.; Wang, K. D.; Dai, Y. J.; Liu, Y. W.; Ma, F.;

643

Rittmann, B. E.; Zheng, P.; Zhao, H. P. Nitrate effects on chromate reduction in a

644

methane-based biofilm. Water Res. 2017, 115, 130-137.

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Figure 1. Variations of Cr(VI) and total Cr in B-S and B-Fe during three consecutive operating cycles. Red arrows represent the replacement of synthetic groundwater.

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Figure 2. Final products during Cr(VI) reduction in B-S and B-Fe. (a) XPS spectra of Cr 2p for the generated precipitates; (b) Variations of products from S(0) oxidation with time; (c) Fe 2p XPS spectra for precipitates from Fe(0) oxidation; (d) Average concentration of residual VFAs in a typical operating cycle.

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Figure 3. Effects of different operating factors on microbial reduction of Cr(VI) in proposed bioreactors. Initial Cr(VI) concentration in (a) B-S and (b) B-Fe; initial bicarbonate concentration in (c) B-S and (d) B-Fe.

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Figure 4. Relative abundances of microbial communities revealed by high-throughput sequences for the inoculum, biomass in the B-S and B-Fe after 75 d cultivation at levels of (a) class and (b) genus.

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Figure 5. Mechanistic pathways in microbial Cr(VI) reduction processes coupled to S(0) oxidation (a) and Fe(0) oxidation (b) under anaerobic condition. In B-Fe, chemical reduction also made approximately 1.9% of chromate removal efficiency, as indicated by a dash line.

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Microbial Chromate Reduction Coupled to Anaerobic Oxidation of

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