Journal of Nuclear and Radiochemical Sciences, Vol. 6, No. 1, pp. 39-42, 2005

AcceptedRole of Microbial Activity in Fe and S Cycling in Sub-Oxic to Anoxic Sulfide-Rich
Mine Tailings: A Mini-Review
Danielle Fortin* and Tanmay Praharaj
Department of Earth Sciences, University of Ottawa, Ottawa, ON, Canada, K1N 6N5
Received: November 15, 2004; In Final Form: November 15, 2004
Acid-mine drainage is an important environmental problem associated with base-metal mining and numerous
studies have looked at the chemical and microbial processes involved in the oxidation of metal sulfides under oxic
conditions. However, little is known about the activity of bacteria living in the same sulfide-rich tailings, but
under sub-oxic to anoxic conditions. Recent work on Cu-Zn mine tailings have shown that sulfate-reducing
bacteria (SRB) are present and active in these tailings (under a wide range of pH conditions) and that their activity
is season dependent. In fact, acidic conditions and low organic carbon availability in the spring tend to greatly
limit the activity of SRB in the tailings. On the other hand, IRB populations tend to increase in number in thespring, maybe as a result of the lack of activity of SRB. These findings are in agreement with studies on acidic
coal mining lakes in Germany, which showed that IRB were mainly active in acidic and oxic sediments whereas
SRB dominated in the more anoxic and pH neutral sediments. The presence and activity of SRB and IRB also
represents a potential bioremediation tool, because both microbial pathways generate alkalinity. Recent work on
acidic mining lakes indicates that IRB and SRB activity could be enhanced in a control manner and be used as anacidity neutralizing process to treat environments impacted by AMD.

Introduction
Base-metal mines generate large quantities of mining residues,
which are commonly called mine tailings. These mine tailings
often contain large concentrations iron sulfide minerals (pyrite,
pyrrhotite), along with small amounts of the ore itself and they
are generally disposed of in open-air impoundments. Once
exposed to air and humidity (from rain or snow), the sulfiderich minerals can undergo oxidation as a result of various
microbial and chemical processes.1 The oxidative dissolutionof iron sulfides generate acidity and releases large quantities of
sulfate and soluble heavy metals (Fe, Cu, Zn, Ni, Pb, Hg, As,etc.), producing acid-mine drainage (AMD). A large body ofliterature is available on the generation of AMD,2 the microbial
populations involved in the oxidation of metal sulfides3 and on
potential remediation technologies4−7 because AMD represents
a serious environmental problem facing the mining sector.
Despite all of the above research, the fact remains that little is
known about microbial processes occurring in the deeper suboxic to anoxic portion of the tailings, where metal sulfide
oxidation is no longer occurring. Until recently, it was thoughtthat anaerobic microbial processes could not be sustained in
the anoxic portion of tailings because of the lack of organic
electron donors essential to several heterotrophic bacteria.
However, studies performed on Cu-Zn and Au tailings andacidic mining lakes (from coal mining) have shown that
sulfate-reducing bacteria are indeed present in these environments and that they actively participate in Fe and S cycling.8−13
Moreover, iron-reducing bacteria are also thought to play animportant role in iron cycling and alkalinity generation.14 The
present paper is intended as a mini-review on the occurrence,role and activity of sulfate- and iron-reducing bacteria in minetailings. However, given the limited number of studies on
sulfide-rich mine tailings, this mini-review also includes recent
work on acidic mining lakes associated with coal mining andother environments impacted by mining activity. These envi*Corresponding author. E-mail: dfortin@uottawa.ca. FAX: +1613-562-5192.

ronments possess chemical and mineralogical conditions
similar to those of sulfide-rich mine tailings, i.e., presence oflarge quantities or Fe-rich minerals, acidic and oxic conditions
and low organic carbon content.
Microbial Sulfate Reduction
In aquatic systems (freshwater and marine environments),
microbial sulfate reduction represents an important carbon
mineralization process, where sulfate serves as the final electron acceptor in the oxidation of simple organic carbon
substrates (eq 1).
2CH2O + SO4−2 → 2HCO3− + H2S(1)

Soluble sulfide is produced during the reduction, along withalkalinity. Sulfide can either accumulate in the sediment porewaters or react with soluble reduced iron (Fe(II) to form ironmonosulfides, commonly known as acid volatile sulfides(AVS). These iron monosulfides can however transform overtime into pyrite, a common mineral of sedimentary environments. 15 Microbial sulfate reduction is in fact the driving
process behind the formation of metal sulfides in low temperature environments. 15 The reduction of sulfate to sulfide iscarried out by sulfate-reducing bacteria (SRB). SRB are
considered strictly anaerobic bacteria and tend to prefer neutral
pH conditions, 16 even though they have been recovered inacidic environments.17 SRB use several low molecular weightcompounds, such as acetate, formate, propionate, butyrate,
pyruvate, lactate, but also hydrogen.16
SRB Occurrence and Activity in Mine Tailings and Mining
Impacted Environments
A priori, sulfide-rich mine tailings do not represent a suitable habitat for sulfate-reducing bacteria, because they are
generally acidic and oxic and contain very little organic carbon(Figure 1). However, large SRB populations have been recovered in several acidic and oxic Cu-Zn mine tailings,9−13 as© 2005 The Japan Society of Nuclear and Radiochemical Sciences
Published on Web 07/31/2005

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Figure 1. pH, redox potential and organic carbon content of acidic
and oxic Cu-Zn tailings.

Figure 3. pH, redox potential and organic carbon content of Cu-Zn
tailings showing the presence of oxic conditions at the surface andanoxic conditions in the sub-surface.
IRB (CFU/g dry wt. sed.)

SRB (CFU/g dry wt. sed.)

1.0E+04

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summerspring-40

Figure 2. Seasonal variations in SRB and IRB populations andsulfate-reduction rates in acidic and oxic Cu-Zn tailings.

Figure 4. Seasonal variations in SRB and IRB populations andsulfate-reduction rates in Cu-Zn tailings showing the presence of oxic
conditions at the surface and anoxic conditions in the sub-surface.

shown in Figure 2. Large SRB populations have also been
recovered from the anoxic portion of some Cu-Zn tailings(Figures 3 and 4), where anoxic and slightly acidic pH condition prevail.9−12 Studies of mining impacted environments,
such as acidic mining lakes and river sediments have also beenshown to host large SRB populations. 18−19 Past research on
Cu-Zn mine tailings showed that there was a close relationshipbetween the presence of SRB and the decline of sulfate in theporewaters and pyrite enrichment zones within the tailings.9−11
However, the activity of SRB within these tailings was always
based on circumstantial evidence because their microbialactivity was never established. In addition, it was thought the
SRB were not active under oxic and acidic conditions, but
merely present as dormant cells,9−13 because Gram-positive
SRB (belonging to the Desulfotomaculum genius) are known
to sporulate under stressful conditions. 16 However, recent
work by Praharaj and Fortin,8 has shown that SRB are indeed
active in Cu-Zn tailings and their activity appears to be season
dependent (Figures 2 and 4). The study showed that SRB
populations were particularly active during the summer months
when the tailings contained more organic carbon, as comparedto the spring, when the tailings were depleted of organiccarbon and were more acidic, following snowmelt. In Cu-Zn
mine tailings sites where anoxic and pH neutral conditions
prevailed in the subsurface, sulfate reduction rates were ashigh as 1000 nmol/cm3/day during the summer.8 Interestingly,
SRB activity was not limited to anoxic tailings, high sulfate
reduction rates (ranging from 100 – 450 nmol/cm3/day) were
measured in very acidic and oxic tailings (pH 2–3) in thesummer.8 These high rates (~ 1000 nmol/cm3/day) are in factcomparable to rates measured in salt marsh environments,
where large sulfate concentrations exist.20 The rates measured

in mine tailings are also in the same order of magnitude as theones reported for natural acidic lakes,17 but slightly higher thanthe ones measured in acidic lakes impacted by coal mining.18, 21
In these acidic lakes, Blodau et al.18 reported that sulfate reduction rates were generally very low in the surficial oxic andacidic sediments (pH 3) but rapidly increased with depth andpH.
The microbial diversity of SRB living in mine tailings isstill under investigation, but preliminary results have indicatedthat they can use simple organic acids, such as lactate, acetate
and formate, which have been identified in the porewaters ofacidic Cu-Zn tailings. 12 In addition, SRB can completely
oxidize these electron donors, but results from batch experiments have also shown that some SRB are incomplete
oxidizers, i.e., they oxidize lactate to acetate (unpublisheddata).
The activity of SRB in sulfide-rich mine tailings suggeststhat new iron sulfides are being formed in situ. Sequential
chemical extractions performed on Cu-Au tailings have indeed
shown that there exists zones of pyrite enrichment (extractedwith concentrated HNO3) at various depths, which coincidewith large SRB populations.9−11 Large chromium-reducible
sulfide fractions (CRS) also appeared to coincide with zones ofhigh sulfate reduction rates in Cu-Zn tailings, as shown by
Praharaj and Fortin.8 The presence of high SRR also corresponded to slightly negative δ34S values of the CRS fractions
around the same depth, which is generally indicative of thepresence of biogenic sulfides.8 However, the δ34S values of the
CRS fractions in the sulfide-rich tailings were less negative
than the values generally reported for biogenic pyrite in cold
marine sediments.8 It is suspected that the isotopic composition of the CRS fractions in sulfide-rich tailings corresponds to-40

J. Nucl. Radiochem. Sci., Vol. 6, No. 1, 2005

Role of Microbial Activity in Fe and S Cycling

a mixture of abiotic pyrite (present during the ore formation)and biogenic pyrite formed following FeS formation andmicrobial sulfate reduction.22 It is important to mention herethat the AVS fraction of sulfide-rich mine tailings, known to
include biogenic iron mono-sulfides,23 does not provide useful
information on the activity of SRB, because pyrrhotite, acommon component of sulfide-rich tailings (associated withthe ore), is also extracted, along with biogenic Fe-monosulfides, thus biasing the results.22 Sulfur isotopic composition of
sediments from acidic lakes impacted by coal mining activity
has however been used to infer microbial sulfate reduction andsulfur cycling.24−25 In summary, the best indicator of microbial
sulfate reduction in mining environments is the in situ
measurement of sulfate reduction rate,20 however when it is notpossible to do so, the combination of porewater and solid
phase geochemistry, along with sulfur isotopic analyses can beused to assess the role of SRB in Fe and S cycling in theseenvironments.
Microbial Iron-Reduction
Fe(III) reduction in sedimentary environments is driven byboth abiotic and biotic reactions.26−27 However, microbial iron
reduction represents an important carbon mineralization
process in non-sulfidogenic sediments, because abiotic iron
reduction is limited. Various Fe(III)-oxides27 can serve as the
final electron acceptor in the oxidation of organic carbon (eq 2)
Fe(OH)3 + 0.25 CH2O + 2H+→ Fe2+ + 0.25 CO2 + 2.75 H2O(2)

The microbial reduction of ferric iron oxides releases
soluble reduced iron in solution and generates alkalinity.
Various dissimilatory iron-reducing bacteria are involved in
the reduction of Fe(III) in sedimentary environments. 2 8
Several of them can gain energy from the direct oxidation of
simple electron donors, such as acetate and formate, but othermore complex electron donors can be used.26 Unlike SRB,
iron-reducing bacteria (IRB) can carry Fe(III) reduction underboth oxic and anoxic conditions and under a wide range of pH
conditions. Acidophilic iron-reducing bacteria tend to be
active under oxic conditions, whereas neutrophilic iron
reducing bacteria are generally active under anoxic
conditions.29
IRB Occurrence and Activity in Mine Tailings and Mining
Impacted Environments
The presence and activity of iron reducing bacteria havebeen investigated in acidic lakes impacted by coal and basemetal mining activity14, 18, 21, 30−32 and fluvial sediments.19 In
these environments, large populations of IRB were recoveredin Fe-rich oxic and anoxic sediments showing a wide range ofpH conditions. In acidic lake sediments, the availability ofFe(III) is not the limiting factor in the in situ activity of IRB, it
is rather the availability of suitable electron donors.21 In addition, Fe is readily recycled in these acidic lakes, especiallynear the sediment-water interface. Depending on the season
and on the concentration of dissolved oxygen, Fe(II) oxidation
rates can surpass Fe(III) reduction rates.21 The rapid cycling of
Fe in the upper portion of the sediments is partially caused by
the absence of soluble sulfide in the porewaters, which prevents
FeS precipitation and immobilization. Iron reduction rates inthese sediments can be as high as 250 nmol/cm3/day in theupper acidic and oxic sediments, but decline with depth.14 The
low Fe reduction rates in the deeper sediments is linked to theincreased in situ activity of SRB and to the potential competition for electron donors between IRB and SRB.14 Iron reducers
present in these acidic and oxic sediments are either autotrophic

41

21

or hetorotrophic acidophilic bacteria. Autotrophic IRB
include Acidithiobacillus ferrooxidans, whereas acidophilic
heterotrophic bacteria belonging to the genus Acidiphilium
have been identified as key players in the microbial reductionof Fe(III) in acidic mining lakes.33 In particular, A. cryptumJF-5 has been shown to oxidize several sugars, alcohols, H2,
and most organic acids of the tricarboxylic acid cycle.21, 33
In mine tailings, little is known about the in situ activity of
IRB, even though they can be present in large numbers. They
have been recovered under acidic and oxic tailings (Figure 2),
but also in anoxic and slightly acidic tailings (Figure 4).
Interestingly, populations numbers do not appear to be affectedby seasonal variations, in fact, results indicate that they might
be more abundant in the spring shortly after snowmelt, than in
the summer (Figures 2 and 4). Their in situ activity has notbeen measured as a function of sampling season, but the results
tend to indicate that IRB populations might be sustained andeven thrive under more acidic conditions in the spring, when thein situ pH drops.8 In addition, in the spring, SRB activity has
been shown to decline, as a result of various physico-chemicalfactors, including low pH, thus lowering any potential competition for suitable electron donors between IRB and SRB. If it
is the case, the behaviour of IRB in mine tailings would be inagreement with the findings about acidic lakes impacted bymining activity, where IRB are more active in the upper acidic
sediments, due to low pH conditions and less active SRB populations. Preliminary batch experiment results have shown that
IRB isolated from mine tailings can completely oxidize formate,lactate and acetate, but cannot compete with SRB under anoxic
and more neutral pH conditions (unpublished data). It is clearthat more work is needed on the ecology of IRB in sulfide-richmine tailings since they can affect Fe cycling and the generationof alkalinity, as discussed in the last section of this paper. Suchwork in currently underway in the author’s laboratory.
Stimulation of Microbial Iron and Sulfate Reduction as a
Potential Bioremediation Technology
Equations 1 and 2 indicate that alkalinity is being generated
during the microbial reduction of sulfate and iron. As a result,these two anaerobic processes could be used to increase alkalinity in sulfide-rich tailings and sediments of mining lakes andcombat acidity production or existing acidity originating fromacid-mine drainage. Some studies have been performed onacidic coal mining lakes in Germany,34−38 where the sediments
are characterized by low pH (pH ~2) and low organic carbon
content.21 Given the fact that microbial sulfate and iron reduction are carried out by heterotrophic bacteria (with the exception of A. ferrooxidans), it is essential to have a large supply ofsuitable organic carbon in these sediments in order to stimulate
microbial growth and alkalinity generation. Laboratory experiments performed with sediments from acidic mining lakes
have shown that the addition of simple organic substrates(lactate, pyruvate, etc.) and cheap industrial wastes can stimulate microbial sulfate reduction, which had for effect toincrease the pH of the sediment slurry by several pH units. 36
Small microcosms filled up with lake sediments and waterfrom acidic mining lakes have also been tested to determine
the effect of organic carbon substrate addition on alkalinity
production.34 Results showed that the addition of cheap industrial wastes (a combination of Carbokalk (a mixture of organiccarbon and lime from the sugar industry), ethanol and whey)
stimulated microbial sulfate reduction, neutralized the acidity
and increased the pH of the microcosm to near neutral-pH
conditions. 34 Large enclosures (amended with Karbokalk)were also setup in these acidic mining lakes in order to assess
the importance of Fe(III) reduction in alkalinity generation.37
Results showed that microbial iron reduction was the initial
microbial process to take place, but the presence of suitable

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J. Nucl. Radiochem. Sci., Vol. 6, No. 1, 2005

anions for the precipitation of Fe(II) minerals (either carbonate
or sulfides) is critical in maintaining the gain of alkalinity
because Fe(II) has the potential to be re-oxidized and generate
acidity.37 Overall, these studies indicate that controlled microbial iron and sulfate reduction processes might represent an
important remediation tool to maintain alkalinity generation inenvironments impacted by acid-mine drainage. In sulfide-minetailings, several studies have looked at chemical and physicalprocesses designed to eliminate acidity generation within themining waste or prevent it, prior to open-air disposal.39−40
Controlled in situ iron and sulfate reduction experiments have
however never been investigated in such environments, but
based on the information that in situ SRB and IRB populations
are present and active in Cu-Zn tailings, these alkalinity generation processes should be further investigated.
Conclusion
Base-metal mines and their associated tailings can generate
acidity and release large concentrations of toxic heavy metals
in the surrounding ecosystems. If left unattended and untreated,these sulfide-rich tailings can oxidize for hundreds of years. It
is then important to find practical and cost-effective treatment
processes to reduce or stop acidity generation. Based on the
research that has been carried out in Cu-Zn mine tailings andacidic coal mining lakes in Germany, it appears that in situ
microbial processes, namely iron and sulfate reduction, might
represent an interesting approach in controlling and neutralizing
acidity within these mining-impacted sites. However, several
factors must be kept in account, such as the type of organic
substrates used to stimulate microbial activity, the in situ conditions of the sites (temperature, pH, Eh, concentration of electron
acceptors, etc.) and the potential competition between variablemicrobial communities for the available electron donors. All of
the above have the potential to severely impact the activity of
SRB and IRB and the long term effectiveness of the bioremediation processes.
Acknowledgements. Part of the research presented in thispaper was funded by an NSERC Discovery Grant to D. Fortin.
The authors also like to thank all the mining companies thathave granted us access to their mine tailings sites over theyears and all the graduate and under-graduate students who
have participated in the NSERC-funded project.
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