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In situ stimulation of thiocyanate biodegradation through phosphate amendment in gold mine tailings water Mathew P. Watts, Han M. Gan, Lee Y. Peng, Kim-Anh Lê Cao, and John W. Moreau Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.7b04152 • Publication Date (Web): 24 Oct 2017 Downloaded from http://pubs.acs.org on October 26, 2017
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Environmental Science & Technology
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In situ stimulation of thiocyanate biodegradation through phosphate amendment in gold
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mine tailings water
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Mathew P. Watts1, Han M. Gan2,3,4, Lee Y. Peng3, 4, Kim-Anh Lê Cao5 and John W. Moreau1, *
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1
School of Earth Sciences, The University of Melbourne, Parkville, Vic., Australia
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Centre for Integrative Ecology, School of Life and Environmental Sciences, Deakin University,
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Geelong, Vic, Australia
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3
School of Science, Monash University Malaysia, Bandar Sunway, Petaling Jaya, Selangor,
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Malaysia
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4
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Bandar Sunway, Petaling Jaya, Selangor, Malaysia
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5
14
Melbourne, Parkville, Vic., Australia
Genomics Facility, Tropical Medicine and Biology Platform, Monash University Malaysia,
Centre for Systems Genomics, School of Mathematics and Statistics, The University of
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*Corresponding Author :
[email protected] 17
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ABSTRACT
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Thiocyanate (SCN-) is a contaminant requiring remediation in gold mine tailings and
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wastewaters globally. Seepage of SCN--contaminated waters into aquifers can occur from
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unlined or structurally-compromised mine tailings storage facilities (TSFs). A wide variety of
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microorganisms are known to be capable of biodegrading SCN-; however, little is known
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regarding the potential of native microbes for in situ SCN- biodegradation, a less costly
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remediation option to engineered approaches. Here we experimentally characterize the principal
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biogeochemical barrier to SCN--biodegradation for an autotrophic microbial consortium enriched
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from mine tailings, to arrive at an environmentally realistic assessment of in situ SCN-
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biodegradation potential. When amended with phosphate, the consortium completely degraded
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up to ~10 mM SCN- to ammonium and sulfate, with some evidence for nitrification of the
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ammonium to nitrate. Although similarly enriched in known SCN--degrading strains of
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thiobacilli, this consortium differed in its source (mine tailings) and metabolism (autotrophy)
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from those of previous studies. Our results provide proof-of-concept that phosphate limitation
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may be the principal barrier to in situ SCN- biodegradation in mine tailing waters, and also yield
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new insights into the microbial ecology of in situ SCN- bioremediation involving autotrophic
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sulfur-oxidising bacteria.
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1. INTRODUCTION
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Thiocyanate (SCN-) commonly forms in waste streams of the gold mining and coal gasification
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industries1. During gold processing, cyanide (CN-) is applied as a lixivant for gold; however,
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excess CN- readily reacts with reduced sulfur (in sulfide ore minerals) to form SCN- 2. The
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chemical stability of SCN-, and re-use of mine wastewater for gold extraction, can result in high
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accumulation (>1 g L-1) of SCN- in mine waste streams3. The presence of this SCN- decreases
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the efficiency of gold extraction4, and more importantly presents a toxicity problem for aquatic
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organisms5, with LC50 values of 0.55 to 33.47 mg L-1 reported for Daphnia magna6.
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Conventionally, the waste products of the gold extraction process are stored in large, often
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unlined, open-air tailings storage facilities (TSFs)7-9. These structures allow sedimentation of
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solid-phase waste and retention of SCN--contaminated water, therefore posing a potential
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environmental risk from accidental water release due to dam failure or seepage into
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groundwater10. The remediation of SCN- in tailing waters therefore mitigates a major potential
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environmental impact of gold mining.
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A number of chemical methods have been proposed for the removal of SCN- from contaminated
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waters, typically utilizing costly strong oxidizing agents3 that can produce hazardous waste3, 5.
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However, the ability of microbes to degrade SCN- offers an effective and potentially economical
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alternative11,
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degradation have been isolated from a range of environments13. These microbes grow either
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autotrophically via sulfur oxidation, utilizing SCN- as an energy source and/or as a sulfur or
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nitrogen source, or heterotrophically via an organic carbon substrate, utilizing SCN- as a nitrogen
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. Taxonomically and metabolically diverse microorganisms capable of SCN-
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source5. Two biodegradation pathways have been proposed: one mediated by SCN--hydrolase
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through the intermediate carbonyl sulfide (COS)14, 15, and the other via the intermediate cyanate
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(CNO-) mediated by a thiocyanate:cytochrome c oxidoreductase identified in Thioalkalivibrio
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species16. Both pathways result in release of carbon dioxide (CO2), sulfide (S2-) or elemental
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sulfur (S0), and ammonium (NH4+)14.
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Harnessing the metabolic activity of SCN--degrading microorganisms, several bioreactor systems
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have been designed and tested12,
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throughput sequencing to characterise SCN--degrading bioreactor communities13, and
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Thiobacillus species tend to be highly represented19, 20, 23-25. These bioreactors can also contain
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microorganisms capable of cycling the nitrogen and sulfur released from SCN- degradation23.
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The removal of by-product NH4+ is key to many bioreactor designs, with promotion of the
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growth of nitrifying bacteria19, 25 and nitrogen assimilation by algae20. To our knowledge, no
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studies have examined the potential for in situ biostimulation of SCN- degradation in
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contaminated mine tailing waters.
17-22
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Furthermore, some studies have employed high-
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The aim of this study was to assess the potential for biostimulation of an extant microbial
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community from gold mine tailing waters to biodegrade SCN-. Through the sole addition of
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phosphate (PO43-), complete degradation to NH4+ and sulfate (SO42-) was achieved in replicate
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batch incubations, with some evidence of limited nitrification to NO3-. High-throughput Illumina
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sequencing of 16S and 18S rRNA genes was performed to characterise the microbial community
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composition of tailings surface water, and to evaluate any changes observed during
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biostimulation. Through monitoring of geochemical and microbial community dynamics, we
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inferred that metabolic responses to biostimulation resulted in rapid, complete SCN-
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biodegradation. This study therefore represents proof-of-concept for in situ biodegradation of
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SCN- in gold mine tailings, and yields important insights into the roles of microbial community
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members in this process.
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2. MATERIAL AND METHODS
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2.1. Field site description. Samples were collected from a TSF located at the Stawell Gold Mine
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in Victoria, Australia, that historically has received large quantities of slurried waste from both
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milled sulfidic and oxide-rich ores26, 27. Some of the SCN--contaminated water from this slurry
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has seeped through the unlined TSF into the underlying aquifer, while the residual surface water
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has been re-used in the gold extraction process. Frequent historical monitoring (>20 years) of
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the surface water geochemistry of the TSF records consistently saline and slightly alkaline (pH
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~8) conditions, typically with 500-1000 mg L-1 of SCN- and
