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Efficient Arsenic Methylation and Volatilization Mediated by a Novel Bacterium From an Arsenic-Contaminated Paddy Soil Ke Huang, Chuan Chen, Jun Zhang, Zhu Tang, Qirong Shen, Barry P. Rosen, and Fang-Jie Zhao Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.6b01974 • Publication Date (Web): 03 Jun 2016 Downloaded from http://pubs.acs.org on June 4, 2016
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Environmental Science & Technology
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Efficient Arsenic Methylation and Volatilization Mediated by a Novel Bacterium
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From an Arsenic-Contaminated Paddy Soil
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Ke Huang1, Chuan Chen1, Jun Zhang1, Zhu Tang1, Qirong Shen1, Barry P. Rosen2,
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Fang-Jie Zhao1, 3*
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Innovation Center for Solid Organic Waste Resource Utilization, College of Resources
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and Environmental Sciences, Nanjing Agricultural University, Nanjing 210095, China
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Jiangsu Key Laboratory for Organic Waste Utilization, Jiangsu Collaborative
Department of Cellular Biology and Pharmacology, Herbert Wertheim College of
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Medicine, Florida International University, Miami, Florida, USA
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Rothamsted Research, Harpenden, Hertfordshire AL5 2JQ, UK
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* Author for correspondence
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Fang-Jie Zhao, College of Resources and Environmental Sciences, Nanjing
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Agricultural University, Nanjing 210095, China
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Email:
[email protected] 17
Phone: +86 25 84396509
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Fax: +86 25 84399551
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ABSTRACT: Microbial arsenic (As) methylation and volatilization are important
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processes controlling the As biogeochemical cycle in paddy soils. To further
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understand these processes, we isolated a novel bacterial strain, SM-1, from an
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As-contaminated paddy soil. SM-1 showed strong As methylation and volatilization
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abilities, converting almost all arsenite (10 µM) to dimethylarsenate and
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trimethylarsenic oxide in the medium and trimethylarsine gas into the headspace within
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24 h, with trimethylarsine accounting for nearly half of the total As. Based on the 16S
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rRNA sequence, strain SM-1 represents a new species in a new genus within the family
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Cytophagaceae. Strain SM-1 is abundant in the paddy soil and inoculation of SM-1
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greatly enhanced As methylation and volatilization in the soil. An arsenite
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methyltransferase gene (ArarsM) was cloned from SM-1. When expressed in
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Escherichia coli, ArArsM conferred the As methylation and volatilization abilities to E.
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coli and increased its resistance to arsenite. The high As methylation and volatilization
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abilities of SM-1 are likely attributed to an efficient ArArsM enzyme coupled with low
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arsenite efflux. These results suggest that strain SM-1 plays an important role in As
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methylation and volatilization in the paddy soil and has a great potential for As
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bioremediation.
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INTRODUCTION
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Arsenic (As) is a toxic metalloid and a non-threshold class-one carcinogen to humans.1
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Arsenic is ubiquitous in the environment, derived from both geogenic and
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anthropogenic sources. Millions of people worldwide suffer from chronic As poisoning,
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especially in south and southeast Asia.2 Humans are exposed to As mainly through
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drinking water and food. Rice, the staple food for more than half of the world
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population, is a major source of dietary As for populations in south and southeast
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Asia.3-5 Large areas of paddy soils in south and southeast Asia are contaminated with
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As due to mining, smelting, irrigation with high As groundwater, and uses of
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As-containing agrochemicals.2, 6, 7 It is therefore important to understand the
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biogeochemical cycling of As in paddy systems.
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Episodic flooding and draining of paddy soil during rice cultivation have profound
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impact on the As biogeochemical cycling. Upon flooding, soil redox potential
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decreases, leading to reductive dissolution of iron oxides/hydroxides together with the
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adsorbed arsenate [As(V)], which is then reduced by microorganisms to arsenite
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[As(III)].8, 9 In addition, adsorbed As(V) can be reduced to As(III); the latter is less
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strongly adsorbed and has a greater tendency to partition into the soil solution phase.8-10
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Flooding of paddy soil thus results in increased bioavailability of As to rice plants.11, 12
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Another important change upon flooding of paddy soil is that microbial As methylation
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is enhanced.13, 14 This could be because As(III), the substrate of As methylation, is
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mobilized and/or anaerobic microorganisms capable of As methylation become more
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abundant.13, 15 Microbial As methylation is an important component of the global 3
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biogeochemical cycle of As,16 and is also a prerequisite for the production of volatile
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methylarsine gases.17, 18 Biovolatilization of As from the terrestrial environment is
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estimated to range from hundreds to tens of thousands ton per annum, but the pathway
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is poorly understood.18 Microbial As methylation in paddy soil also impacts As
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speciation in rice grain. Rice grain contains both inorganic and organic (methylated) As
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species, with methylated As species accounting for between 10% and 90% of the total
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As in rice grain depending on the geographical region and the growth conditions of
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rice.15, 19, 20 Methylated As species in rice are derived from soil microorganisms,
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because rice plants do not appear to be able to methylate As.21
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Many microorganisms are able to methylate As, some of which are also able to
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volatilize As.22 Arsenic methylation is catalyzed by As(III) S-adenosylmethionine
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(SAM) methyltransferase enzymes (ArsM), which transfer methyl group from SAM to
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As(III) to produce mono-, di- and trimethyl arsenical compounds.17, 23 Depending on
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the microorganism studied, a range of different volatile or non-volatile methylated As
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compounds are produced.17, 23-28 Genes encoding ArsM appear to be abundant and
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diverse in paddy soils,29, 30 but to date only a few studies have investigated microbial
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isolates from paddy soils for their As methylation abilities. Kuramata et al. 27 isolated
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an aerobic bacterium belonging to Streptomyces sp. from a paddy rhizosphere soil and
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showed that it can methylate As(III) to methyarsenate [MAs(V)] and dimethylarsenate
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[DMAs(V)]. Wang et al. 26 reported that an anaerobic sulfate-reducing bacterium
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belonging to Clostridium sp. isolated from a paddy soil also methylates As(III) to
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MAs(V) and DMAs(V). Both isolates appeared to produce very little volatile As 4
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species. In both laboratory and field studies, methylarsine gases, especially TMAs(III),
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have been detected from paddy soils,13, 14 but the microorganisms mediating As
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biovolatilization remain unknown.
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In the present study, we isolated a novel bacterial strain SM-1 from an
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As-contaminated paddy soil. The strain represents a new genus in the family of
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Cytophagaceae and has a strong ability to methylate and volatilize As. Here, we
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characterize the molecular mechanisms underpinning the high As methylation and
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volatilization in strain SM-1 and its role in As biogeochemical cycle in paddy soil.
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MATERIALS AND METHODS
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Soil Incubation and As Speciation in Porewater. A paddy soil was collected from
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Shimen city, Hunan Province in southern China. The soil is moderately contaminated
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with As (30.0 mg kg-1) due to mining activities nearby. The soil contains 11.1 g kg-1 of
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organic carbon and has a pH of 6.85. The soil was air-dried, sieved to
