-
For Review OnlyPHYSICO-CHEMICAL PROPERTIES, HEAVY METALS AND
METAL-TOLERANT BACTERIA PROFILES OF ABANDONED GOLD MINE TAILINGS IN
KRUGERSDORP, SOUTH AFRICA
Journal: Canadian Journal of Soil Science
Manuscript ID CJSS-2018-0161.R2
Manuscript Type: Article
Date Submitted by the Author: 03-Feb-2020
Complete List of Authors: Fashola, Muibat; Lagos State
University, Ojo Lagos, Department of MicrobiologyNgole-Jeme ,
Veronica ; College of Agriculture and Environmental Sciences,
UNISA, Florida, Environmental SciencesBabalola, Olubukola; North
West University, Department of Biological Sciences
Keywords: Soil properties, Metal tolerant bacteria, Mine
tailings, Tudor shaft
Is the invited manuscript for consideration in a Special
Issue?:Not applicable (regular submission)
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CJSS-2018-0161
TITLE PAGE
PHYSICO-CHEMICAL PROPERTIES, HEAVY METALS AND METAL-TOLERANT
BACTERIA PROFILES OF ABANDONED GOLD MINE TAILINGS IN
KRUGERSDORP,
SOUTH AFRICA
Muibat Omotola Fashola 1, 2, Veronica Mpode Ngole-Jeme3 and
Olubukola Oluranti Babalola 1
1. Food Security and Safety Niche Area, Faculty of Agriculture,
Science and Technology, North-West
University, Private Bag X2046, Mmabatho 2735, South Africa;
[email protected]
(O.O.B.)
2. Department of Microbiology, Faculty of Science, Lagos State
University, Ojo, Lagos, Nigeria
[email protected] (M.O.F.)
3. Department of Environmental Sciences, College of Agriculture
and Environmental Sciences,
UNISA, Florida, Private Bag X6 Florida, Roodepoort 1710, South
Africa;
[email protected]
Correspondence: [email protected]; Tel.:
+27183892568
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ABSTRACT
Mine tailings are a potential source of heavy metals that can be
toxic to microbes, plants and animals in
aquatic and terrestrial ecosystems. Bacteria have evolved
several mechanisms to tolerate the uptake of
heavy metal ions. This study aimed to assess the physicochemical
properties, concentrations of selected
heavy metals and metalloids (As, Ni, Pb, Zn, Cd, and Co) and
isolate potential metal tolerant bacteria
present at three abandoned gold mining sites with a view of
understanding how tailings characteristics
vary and the implications on microbial activities in tailings
dumps. Heavy metal tolerant bacteria were
isolated from the samples using minimum inhibitory and maximum
tolerable concentrations of the Ni,
Pb, Zn, Cd, and Co. The substrates of the studied sites were
acidic and deficient in nutrients. High
metals and metalloid concentrations in the order Zn > Ni >
Co >As > Pb > Cd were recorded in some of
the studied sites and its adjacent soil which exceeded South
African recommended values for soil and
sediments. Heavy metal tolerant bacteria that showed multiple
tolerances to Ni, Pb and Zn were
isolated and putatively identified using biochemical tests as
belonging to the phylum Proteobacteria,
Actinobacteria, and Firmicute. Gold mine tailings enriched the
soil with heavy metals and also affects
soil physicochemical properties. Proper management of mine
wastes must be ensured to prevent their
adverse effects on the diversity, composition and activity of
soil microorganisms that help in
maintenance of the ecosystem.
Keywords: Soil properties, Metal tolerant bacteria, Mine
tailings, Tudor shaft
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INTRODUCTION
Heavy metal (HM) and metalloid contamination associated with
gold mining is one of the foremost
environmental concerns in areas where such mines are located.
Heavy metals are naturally occurring
metals having an atomic number greater than 20 with a density
greater than 6 g cm-3 and a molecular
weight greater than 53 (Howlett and Avery, 1997; Ali and Khan,
2017). Metalloids on the other hand
are chemical elements which in their standard state, have (a)
the electronic band structure of a
semiconductor or a semimetal, (b) an intermediate first
ionization potential (750−1,000 kJ/mol), and (c)
an intermediate electronegativity (1.9−2.2) (Vernon, 2013). The
metalloid arsenic (As) and heavy
metals cadmium (Cd), cobalt (Co), copper (Cu), manganese (Mn),
mercury (Hg), nickel (Ni), uranium
(U), and zinc (Zn) have been reported in soils around gold mines
in different parts of the globe (Lee et
al., 2005; Jian-Min et al.; 2007; Abdul-Wahab and Marikar, 2012;
Armah et al., 2014). These elements
persist for long periods in both aquatic and terrestrial
ecosystems and adverse effects can span across
trophic webs from soil through plants to animals including
humans (Chary et al., 2008). Elevated levels
of HM in soils may lead to their uptake by plants which can
reduce crop productivity as a result of their
inhibitory effects on multiple physiological processes in plants
(Singh and Kalamdhad, 2011).
Contamination of aquatic environments with HM has also been
shown to alter food webs and reduce
diversity in aquatic ecosystems (Förstner and Wittmann, 2012).
Human health effects associated with
HM ingestion (primarily via contaminated aquatic biota) include
complications of the nervous, skeletal
endocrine and digestive systems (Fashola et al., 2016).
In addition to enriching the soil with HM, mining activities and
mine waste generation also can disrupt
nutrient dynamics in soils as a result of dynamic and
interaction alterations in physical, chemical and
microbiological processes (Adewole and Adesina, 2011). Numerous
studies have emphasized the
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significance of soil properties including organic matter (OM),
particle size distribution including clay
content, redox potential, electrical conductivity (EC), moisture
content, cation exchange capacity
(CEC), and pH on the behavior of HM in soils (Sourkova et al.,
2005; Keskin and Makineci, 2009).
Metal mobility has been shown to be lower in fine compared to
coarse textured soils especially if the
mineralogical assemblage of the clayey soil is dominated by 2:1
tetrahedral: octahedral silicate clay
minerals (e.g illite or vermiculite with high cation exchange
capacities). A high content of OM also can
enhance metal adsorption, thereby reducing mobility in the
environment whereas acidic conditions
decrease soil exchange capacities of metal cations and increases
metal solubility in the soil environment
making them more mobile (Sheoran and Choudhary, 2010 ;
Ayangbenro and Babalola, 2017).
Acid mine drainage originating from abandoned gold mines has
resulted in acidification of nearby soils
with consequences for the mobility of HM and microbial diversity
in the soil (Fashola et al., 2016).
Zinc solubility in soil has been reported to increase 100-fold
for every unit decrease in soil pH
(Mortvedt et al., 1991). Chuan et al. (1996), also reported high
solubility of Pb, Cd and Zn in
contaminated soils as the pH decreased from 5.0 to 3.3. Changes
in soil pH have been known to disrupt
certain microbial metabolic pathways via inhibiting activities
of pH-dependent enzymes or altering the
availability of key nutrients and HM, the latter which can be
toxic to soil bacteria (Kapoor et al., 2015;
Ndeddy Aka and Babalola, 2017). Changes in soil microbial
community structure and activities as a
result of mining related changes in soil physico-chemistry could
affect such key ecosystem processes as
soil organic matter turn over leading to a decline in overall
ecosystem functioning and may also lead to
indirect cascading impacts on metal mobility.
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Witwatersrand in South Africa is home to the largest known gold
reserves in the world, making the
country one of the world leaders in gold mining (Hart, 2014).
However, gold mining has also resulted
in the generation of thousands of tailings dumps along the gold
mining corridor from Nigel to
Randfontein (Bobbins, 2015). Gold mine tailings present great
risk to soils, plants, surface and
groundwater because of the dissemination of particles containing
potentially toxic metals and
metalloids through wind action and/or by runoff from the
tailings to streams that drain these tailings
(Naicker et al., 2003; Ayangbenro and Babalola, 2018). In many
residential areas near abandoned
mines in Krugersdorp, dust from tailings has been a long-term
environmental hazard (Bobbins, 2015).
Multiple studies have reported high concentrations of toxic
metals and radioactive elements in different
gold mine tailing dumps and adjacent water resources in this
area (McCarthy, 2011; Kamunda et al.,
2016; Olobatoke and Mathuthu, 2016; Ngole-Jeme and Fantke, 2017)
as well as in many other gold
mines tailings and surrounding soils in other areas in South
Africa (Mitileni et al., 2011; Matshusa et
al., 2012) and beyond (Rafiei et al., 2010; Bempah et al.,
2013).
An understanding of the soil/substrate physicochemical
characteristics and microorganisms around gold
mines may be important to inform remediation strategies aimed at
reducing HM concentrations or
bioavailability (Fashola et al., 2016). In the present study, we
investigated physicochemical properties
and six HM (Pb, Ni, Zn, Cr, Cd, Co) in abandoned gold mine
tailings dams and adjacent soils in
Krugersdorp, South Africa. Additionally, we enriched for and
isolated HM tolerance bacteria on
modified L-B medium from these substrates to help determine how
the selected physicochemical
properties may impact on the activities of microorganisms in the
tailings and the implications of these
impacts on potential bioremediation strategies.
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DESCRIPTION OF STUDY AREA
The study area was Krugersdorp (260 61S and 270 461E), a mining
town in Gauteng province, South
Africa. This province houses the Witwatersrand basin which
covers an area of 1600 km2 and has some
400 km2 of mine tailings dams and 6 billion tons of pyrite
tailings containing 430,000 tons low-grade
uranium (CSIR., 2009).The soils are dominated by plinthic,
duplex and hydromorphic soils, which all
carry land and soil limitations for agricultural crop production
(Bredenkamp, 2002). The vegetation
around the study area is made up of grassland and savanna biomes
comprising 71% and 29%
respectively of Gauteng area.
MATERIALS AND METHODS
Sample collection and preparation
Tailings and soil samples were collected from three abandoned
gold mine sites in Krugersdorp at
depths of 0-15 cm with a Dormer steel soil auger. The three
sites as indicated in Figure 1 were MA
(27.80764 E, -26.14265 S), MB (27.81576, -26.12771) and TS
(27.80362, -26.13191). Samples
collection was done once every two months between February and
September, spanning over a time
period of eight (8) months to accommodate the different seasons
in South Africa. From each of the
three sites, three tailings and three soil samples were
collected a few meters apart and combined to
obtain a composite sample representative of the site. During
each sampling period, samples were
collected at different sites of the three tailings dumps. The
samples were air-dried, crushed, and sieved
through a 2 mm sieve and the sieved soils used for the analyses
of selected physical and chemical
parameters.
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Samples characterization
Physicochemical characterization of samples.
The moisture content of samples was determined using the
gravimetric method described by Odu et al.
(1986) whereas particle size distribution of the < 2 mm
fraction of the samples was determined using
the Bouyoucos hydrometer method (Gee et al., 1986; Filgueira et
al., 2006) after dispersing the
particles with sodium hexametaphosphate, (Na6(PO3)6). A Jenway
3520 - pH meter was used to
determine sample pH, in a 1:2.5 (m: v) soil-water suspension
(Van Reewijk, 1992). Sample EC was
determined in the same suspension using a Hanna multi range
HI8733 EC meter (Thomas, 1996). The
CEC and OM content of the soil samples were determined using the
ammonium acetate extraction
technique and the modified Walkley-Black methods respectively
(Abollino et al., 2002).
Chemical characterization of samples.
Sample chemical properties determined included sulphate, carbon,
nitrogen, sulphur and HM contents.
To determine sulphate content, samples were extracted with a
solution comprising 39 g NH4OAC in 1
liter of 0.25 M acetic acid on a shaker at 200 rpm for 30 mins
after which 0.25 g of activated charcoal
was added. The mixture was again shaken for an additional 3 mins
and filtered through a sulphate free
filter paper (Whatman no 42) which had been washed with the
extracting solution. Ten milliliters of the
filtrate were then pipetted into a 50 ml Erlenmeyer flask and 1
ml of acid seed (6 M HCl + 20 mg of
K2SO4 and 50 ml of 40 mg standard solution plus 50 ml of
concentrated HCl) solution was added. The
solution was swirled, 0.5 g of Bacl2.H20 crystals added, and
allowed to rest for another minute with
frequent swirling to dissolve the crystals. Sulphate content in
the resulting solution was determined
using a UV spectrophotometer at a wavelength of 420 nm (Singh et
al., 1995). Nitrate content of the
samples was determined using the equilibrium extraction method
described by Willis and Gentry
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(1987) using a spectrophotometer automated flow analyzer. The
content of total C, N, and S in each
sample was determined using a LECO CNS Trumac Analyzer
calibrated with a LECO soil reference
standard (CNS LECO part no 502-309). The instruments settings
and operations conditions were done
in accordance with the manufacturer’s specifications.
Conventional aqua regia (3:1 HCl: HNO3)
digestion technique was performed according to the method of
Chen and Ma (2001), and Ngole and
Ekosse (2012) to extract metalloid and HM including As, Pb, Ni,
Zn, Cr, Co from the soil samples.
Heavy metal and metalloid concentrations in the sample extracts
were determined using a contraAA
300 Atomic Absorption Spectrometer.
Isolation and identification of heavy metal Tolerant bacterial
strains.
Metal tolerant bacteria were isolated from the samples using the
spread plate method as follows. One
gram of duplicate composite tailings and soil samples was
suspended in 9 mL of saline solution (8.5 g/
L of NaCl) in distilled water and vortexed for 1-2 mins at room
temperature. Each suspension was
serially diluted (10-1 to 10-7) and aliquots of 0.1 ml dilution
from 10-4, 10-5 and 10-6 spread with a glass
rod over triplicate Luria–Bertani (LB) medium supplemented with
1 mM of ZnS04, NiCl2·H20, Pb
(N03)2, CrCl2, CdCl2·2H20 and 0.5 mM CoCl2·6H20 (Van Nostrand et
al., 2007). To minimize metal
ion complexation, the medium was adjusted to pH 7 using 1.0 N
NaOH or 1.0 N HCl as was necessary.
Control plates were also prepared with the same medium but with
no HM supplement added. Both the
control and experimental plates were incubated at 37°C and 25°C
for 24 - 48 h. Growth of bacteria was
observed and morphologically distinct colonies were selected and
purified in the same medium and
growth conditions (Raja and Selvam, 2009). All cultures were
stored at -80°C in LB broth with 20%
glycerol for further studies. To identify the isolates, the
following tests were performed;
Gram staining was done to classify the bacteria into Gram
positive and Gram-negative bacteria based
on their cell wall composition. Biochemical tests were performed
to determine the metabolic diversity
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of the bacteria present. Indole, Methyl red, Voges-Proskauer,
and citrate utilization test were done to
identify the coliform group that produces carbinol from glucose
fermentation. Catalase, oxidase, nitrate
reduction, hydrogen sulphide production, starch hydrolysis was
done to determine other intracellular
enzymes of the isolates while the sugar (glucose, lactose,
sucrose and maltose etc) utilization tests were
performed to show the ability of the bacteria to ferment
carbohydrates with the release of acid and gas.
The various tests were carried out as described by Cowan and
Steel (Barrow and Feltham, 2004) and
the bacterial isolates were identified accordingly using this
manual.
Determination of multiple metal tolerances levels of
isolates.
The Maximum tolerable concentration (MTC) of Pb, Cr, Zn, Ni, Cd
and Co by the identified bacteria
was determined by agar dilution method (Kannan and
Krishnamoorthy, 2006). A log-phase culture of
the isolates was spot inoculated onto LB agar plates
supplemented with increasing concentration of
metals. The concentrations in mM of the metal species were as
indicated below:
Pb2+: 1, 2, 3, 4, 5, 6, 7, 8, 9
Ni2+: 1, 2, 3, 4, 5
Zn2+ :1,2, 3, 4, 5, 6, 7, 8, 9
Co2+ : 0.5, 1, 2
Cd2+ :1, 2, 3
Cr2+: : 1, 2, 3, 4, 5, 6, 7, 8, 9, 10
All metal salts used were added to the LB agar after autoclaving
and cooling to 50°C from filter-
sterilized stock solutions. The plates were incubated at 37°C
and 25°C for 24 - 48 h and observed for
bacterial growth. The highest concentration of the metal at
which no bacteria growth was seen was
designated as the MTC.
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Quality control
All equipment used were calibrated with reference standards.
Glass wares used for HM analyses were
rinsed in dilute HNO3 before used. Heavy metals used in bacteria
isolation were filtered and sterilized
using membrane filters. All reagents and HM metal standards used
were of analytical grade. Analyses
were carried out in duplicate to ensure reproducibility of the
data obtained.
Statistical analysis
ANOVA was used to determine differences in the properties of the
tailings and adjacent soil at the
different sites whereas correlation coefficient and regression
analyses were used to understand any
relationship that existed between heavy metals and the
physicochemical properties in the tailings and
adjacent soils and to determine the influence of soil
physicochemical properties on HM behavior and
bacterial diversity. All tests were performed at a significance
level of 5% using the SPSS statistical
package programme (version 21).
RESULTS AND DISCUSSION
Physicochemical properties of tailings and adjacent soils
Tailing moisture contents ranged between 7.23% - 12.07% with a
mean of 6.61% whereas the moisture
content of adjacent soils ranged between 3.39% - 6.56% with a
mean of 4.22%. Moisture content was
therefore higher in the tailings compared to the adjacent soils
at all sites (Table 1). Moisture content in
mine soil is however not a stable parameter as it is affected by
sampling period, organic carbon content,
height of dump, stone content, soil texture and thickness of
litter layers on the dump surface (Maiti,
2006).The pattern observed may therefore change depending on the
prevailing condition. The particle
size distribution of the samples ranged between 16-28% for clay,
12-34% for silt and 28-64% for sand.
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Clay contents in the adjacent soil samples were higher than that
in the tailings (Table 1). Both the
tailings and adjacent soil samples had significant amount of
clay-sized particles.The textural
classifications of the samples were sandy loam, silt loam, and
sandy clay loam (Table 1). The host
rocks from which the Au is extracted is usually crushed to a
fine texture to increase the surface area of
the ore body exposed for chemical extraction. This could explain
the finer texture for the tailings
compared to the soil. Most of the tailings and soil samples
analyzed over the eight-month period were
acidic to moderately acidic (Table 1) with the pH values ranging
from 2.17-6.34 and mean of 4.48. All
tailings samples had lower pH values compared to the adjacent
soils. The acidic values recorded in the
tailing soils may be due to oxidation of sulphide bearing
minerals contained in the tailings, which
resulted in low pH (Dold, 2014).Generally,redox level varied
between 194 - 380 mV, with highest level
observed in tailings samples. Soils around the tailings from all
three sites had lower redox values than
the tailings (Table 1). Values for EC at the three sites varied
with values ranging from 5.33 mS/m to
11.52 mS/m in the tailing’s samples and 0.49 mS/m to 3.39 mS/m
in the adjacent soil samples (Table
1). The cation exchange capacities of the tailings varied from
one site to the other (Table 1) with the
adjacent soils having higher CEC values compared to the
tailing’s samples. The CEC values were
generally low reflecting the dominance of 1:1 clay mineralogy
and further confirming that the fineness
of the tailings was not as a result of geogenic processes which
would have resulted in the accumulation
of clay minerals that contribute significantly to soil CEC, but
the physical process of grinding. Organic
matter also affects soil CEC with CEC generally increasing
proportionately with OM content (Ngole-
Jeme, 2016), but the results in this study indicated low OM
content (Table 1) as reflected by the mean
OM content of 0.89% in the tailings and 1.13 % in the adjacent
soils also partially explaining the low
CEC. Soil organic matter in mine tailings has been reported to
be low due to low vegetation cover and
bacterial diversity which play a major role in organic matter
accumulation in soils and sediments
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(Sadhu et al., 2012). Our results are consistent with the
findings of Rösner and Van Schalkwyk (2000);
Yang et al. (2003) and Ashraf et al. (2012) who all reported
that mine associated soils generally were
low in organic matter.
Nitrate levels were mostly higher in the soils compared to the
tailings samples (Table 1). Sulphate
content in samples from the tailings and adjacent soils followed
the order TS > MA > MB >TSC >
MAC > MBC with values from site TS being above the
permissible limit of 200 mg/ml stipulated by
WHO in soils. Sulphate content was higher in the tailings than
in the adjacent soil samples. High
sulphate content in the tailings could be indicative of the
presence of sulphur containing minerals such
as pyrite, and chalcopyrite in the Au host rock (Bosman, 2009).
According to Rosner (2007), a total
sulphur content of approximately 0.1% within the first meter
below the surface of tailings indicates that
the tailings have almost fully oxidized. The values obtained for
this study indicate that the tailings have
not been fully oxidized and may still present a risk of AMD
generation. Results in Figure 2 showed that
mean total carbon (0.48%) and nitrogen (0.13 %) content in the
tailings samples were lower than in the
adjacent soils (1.74% for C and 0.15% for N). The samples
therefore had a low C: N ratio of 1: 0.086:
1:0.027. Nitrogen is generally known to be deficient in mine
dumps (Sheoran and Choudhary, 2010),
which was observed in both the tailings and adjacent soil
samples. This is as a result of decrease in
microbial population and unfavorable conditions for maintenance
of soil vegetation cover and
formation of soil humus in the mining environment. The nitrogen
content recorded in this study also
corroborates the values reported by Saviour and Stalin (2012),
who also noted that the inadequate
mineralizable organic nitrogen and reduced mineralization rates
of mine dumps lower nitrogen
contents. The results of the physicochemical properties of the
tailings indicate the variations that may
exist in gold mine tailings and adjacent soils. These variations
may be influenced by the nature of the
host rock at the site as well as the gold mining process
utilized at the mine. It also generally shows that
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tailings have a significant influence on the physico-chemical
properties of adjacent soils as shown in
the pattern of values for the different parameters at the
different sites.
Heavy metals concentrations in tailings and adjacent soils
The concentrations of As, Cd, Co, Ni, Pb and Zn recorded in the
tailings and adjacent soils after acid
digestion are shown in Figure 3. Arsenic content in tailings MA
and MB and the adjacent soils MAC
and MBC were below the detection limit of the equipment. All
metal concentrations excluding Cd in
tailings from TS and its adjacent soil TSC exceeded the
recommended values of different standard
bodies as shown in Table 2. Cadmium concentration in site TS had
values which fell within the
permissible levels recommended by South Africa standards for
soils and sediment quality guidelines
but above Dutch pollutant standard. Highest concentration of all
the metals in tailings and adjacent soils
were recorded in site TS and TSC (Figure 3 and Table 2). The
concentration of Cd, Co and Ni found in
tailings from sites MA and MB fall within the acceptable limits
recommended by several countries
(Table 2). The maximum and mean values of Pb and Zn (Figure 3)
recorded in MAC and MBC
exceeded the recommended values stipulated by South Africa
standards for soils and sediment quality
guidelines, Dutch pollutant standard and Canadian soil quality
guidelines for the protection of human
and environmental health (Table 2).
Among all the samples analyzed, samples from sites TS and TSC
had the highest values for HM
concentrations, with Zn having the highest concentrations among
the six metals studied (Figure 3).The
concentrations of the elements are typical of what has been
reported in gold mine tailings in other
studies and also highlight the variability that exist in the
properties of mine tailings soil. The
concentration values for most of the metals were higher in the
adjacent soil compared with the tailings.
This could be attributed to the properties of the soils relative
to the tailings. Lower CEC values for the
tailings imply the cations are likely to be leached into
adjacent soil where they may accumulate because
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they have a higher OM and CEC. All the metals except Pb were
highly positively correlated (P ≤ 0.01
and P ≤ 0.05) with r values of 0.432 for Cd and As, 0.534 for Co
and As, 0.592 for Ni and As, 0.582 for
As and Zn, 0.861 for Co and Cd, 0.863 for Ni and Cd, 0.848 for
Zn and Cd, 0.964 for Co and Ni, 0.974
for Co and Zn and 0.976 for Ni and Zn indicating a common source
for these HM. Lack of correlation
with Pb could be an indication that the Pb contained in the
samples could be from other anthropogenic
inputs aside from gold mining activities. Gold mining activities
in the studied areas has resulted in
deterioration of the surrounding environment through
contamination of HM and metalloids in the soil.
This observation agrees with several studies conducted on gold
mine tailings dams as potential source
of HM contamination in adjoining soils and sediments in South
Africa (Bempah et al., 2013; Olobatoke
and Mathuthu, 2016).
Metal tolerant bacterial strains identified in the tailings and
adjacent soils
Metal tolerant bacteria are usually found in sites polluted by
high concentrations of metal species (Wei
et al., 2009 ; Xie et al., 2016). A total of 117 metal tolerant
bacteria species were isolated from both the
tailings and the adjacent soils studied using 1 mM
concentrations each of Pb, Zn, Ni, Co, Cr and Cd
(Table 3). There seem to be more metal tolerant isolates in the
tailings compared to the adjacent soils
for two of the three sites studied. After screening using
colonial and morphological characteristics;
thirty-five isolates showed distinct colony characteristics and
were selected and putatively identified
using biochemical tests (Table 4 and 5). The various colonial
and cellular morphology of the bacterial
isolates are shown in Figure 3. Out of the 35 isolates selected,
31.43% were Gram negative while the
remaining 68.57% were Gram positive. The biochemical results of
the selected isolates (Table 4)
showed the effects of the HM on the various biochemical
properties determined. Over sixty-eight
(68.57%) of the isolates were urease negative while 57.14% were
oxidase positive and 97.14% had
catalase enzyme. Most of the sugars were also utilized by the
isolates. Similar observations were made
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by Oste et al.(2001),who reported a decrease in the activities
of various soil enzymes such as urease due
to metal pollution. The ability of the bacterial strains to
utilize the sugars signifies aerobic metabolic
pathways (Samanta et al., 2012). The sugars are converted into
pyruvate that are decarboxylated and
oxidized to become acetyl Co A which condenses with oxaloacetate
to enter the TCA cycle. Also, the
utilization of the different carbon sources by the identified
bacteria indicated diversity of metabolic
patterns that could enhance their degradative ability. The
results of the biochemical tests (Table 4)
putatively revealed that the bacterial isolates belong to three
phyla: Actinobacteria, Proteobacteria and
firmicutes (Table 5).
Inability to isolate the acidophilic heterotrophic iron and
sulphur oxidizing bacteria such as
Acidiphilium, Acidithiobacillus ferrooxidans, Leptospirillum
ferrooxidans, and some other iron
oxidizing bacteria that are known to only grow and proliferate
in acidic pH 2.4 -5.0 in mine tailings
might have been due to the neutral pH of the culture medium used
in enriching and isolating the
bacteria.
Previous researches have shown the abundance of these phyla in
HM and metalloid impacted
environments (Jamaluddin et al., 2012; Bajkic et al., 2013;
Hookoom and Puchooa, 2013). Their ability
to survive in these environments has been attributed to the
composition of their cell walls that are able
to interact and bind effectively with the metals (Shin et al.,
2012) as well as various genetic
mechanisms that enable them to overcome the effects of the toxic
metals (Rensing and Grass, 2003;
Abou-Shanab et al., 2007; Dupont et al., 2011). Ability of a
native bacteria to tolerate HM stress is a
major determinant in the utilization of such bacteria as a
bioremediating agent. There are various
reports on different bioremediation strategies employed by
indigenous bacteria isolated from mine
tailings. Autochthonous Bacillus sp isolated from mine tailing
in South Korea with the ability to
effectively biomineralize active Pb ions to inactive form has
been reported by Govarthanan et al.
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(2013). This bacterium also increases the urease enzyme activity
of the tailings as well as significantly
reduced the bioavailability of Pb in the environment.
Furthermore, Ndeddy Aka and Babalola (2016),
described the enhancement of phytoextraction capacity of
B.juncea by three indigenous HM resistant
bacteria; Pseudomonas aeruginosa, Alcaligenes faecalis and
Bacillus subtilis isolated from mine
tailings in South Africa. It was observed that the inoculation
of the bacteria with B. Juncea, an
hyperaccumulator plant grown in soil spiked with different
concentrations of Ni, Cd and Cr resulted in
higher accumulation of the three metals compared with the
uninoculated plant. This shows that
indigenous HM tolerant bacteria could bring about revegetation
of mine tailings and surrounding
polluted sites.
Determination of multiple metal tolerances
The isolates were screened against six metals (Co2+, Ni2+ Pb2+
Cd2+ Zn2+ Cr2+) to examine the extent of
multiple tolerances. All the isolates showed the ability to
tolerate multiple metals found in the tailings
and surrounding soils. All the isolates could not grow beyond 1
mM concentration of Co. The pattern of
toxicity of the metals to the bacterial isolates was Co > Cd
> Ni > Pb > Zn > Cr, which shows that Co is
the most toxic to the bacterial isolates out of the six metal
species as shown in Table 6. The isolates
showed a high level of tolerance for Cr with 51.42% of the
isolates tolerating 9 mM and 17.14%
tolerating 10 mM of Cr. 25.71% of the bacterial isolates
tolerated up to 7 mM of Zn, 11.42% up to 7
mM of Pb and 17.14% tolerated 5 mM of Ni (Table 6).
The high level and widespread tolerance shown by the bacterial
isolates against multiple metals (Table
6) could be attributed to the elevated concentrations of the
metals recorded in the different samples
where the bacteria were isolated from. Bacteria when exposed to
high concentrations of HM devise
various physiological and genetic mechanisms needed for their
adaptation and survival under such
conditions (Fashola et al., 2016). Fashola et al. (2019), have
also highlighted that exposure of bacteria
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to high concentrations of heavy metals extends the lag phase of
their life cycle resulting in increased
survival rate. In addition, the efflux system which enables
bacteria to pump out metals from the
cytoplasm to the periplasmic space with the help of ATPase found
in the internal membrane of the
bacteria has been reported in Gram-negative bacteria (Blair et
al., 2014). Other mechanisms such as
complexation, precipitation, biotransformation, bioaccumulation,
oxidation-reduction reactions and
biosorption have also been reported to be responsible for
bacterial tolerance to metals (Wei et al., 2009;
Sahmoune and Louhab, 2010 ; Govarthanan et al., 2013).
The MTC values recorded for Ni, Pb and Zn by the bacterial
isolates obtained in this study are however
much higher than those reported by Choudhary and Sar (2009) and
Govarthanan et al. (2013) in mine
polluted tailings and soil. The Ni tolerance level recorded is
lower than the values reported by Bajkic et
al. (2013), where Ni tolerance as high as 11 mM was attained by
the bacteria strain MS108 identified as
Staphylococcus sp. from a copper mining and smelting complex in
Serbia. The differences in metal
tolerance by the bacterial isolates could be attributed to many
factors such as the strength of the
medium used in isolation, presence of negatively charged ions
like chloride, organic constituents, and
nature of the medium which determines availability of the metals
to the bacteria (Kannan and
Krishnamoorthy, 2006). The multiple metals tolerant patterns
observed by the bacterial isolates could
be attributed to the fact that, mine tailings are usually
contaminated with a cocktail of metals, and
hence, bacteria surviving in mine tailings are likely to possess
the ability to withstand high
concentrations of different metals. For example, Staphylococcus
sp isolated from copper mining soil
was able to tolerate Cr, Ni and Cd (Bajkic et al., 2013).
Similarly, the bacterial strain CCNWRS33-2
isolated from Lespedeza cuneate in gold mine tailings also
showed high tolerance to Cu, Cd, Pb and Zn
(Wei et al., 2009). The present study also shows high tolerance,
as well as tolerance to multiple metals
by bacterial isolates.The multiple heavy metal tolerance
exhibited by the isolates could be used to bio
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augment environments contaminated with a cocktail of metals to
improve the rate of natural
attenuation.
Implications of the physicochemical properties of soils and
tailings on HM behavior and effects on
the adjacent environment
Some very strong negative and positive correlations were
observed between HM and analyzed
parameters including pH, EC, and nitrates in the adjacent soils
(Table 7). These correlations were none
existent in the tailings samples (Table 8). In both the soil and
tailings samples, there were significant
correlations among the heavy metals (Table 7 and 8). Soil
properties and heavy metals concentrations
in uncontaminated soils display natural associations because of
the influence of these properties on
heavy metal behavior in the soils. Lack of correlation between
soil properties and the heavy metals in
the tailings is an indication that these have been affected by
anthropogenic activities, which in this case
may be the process of mining. There is strong correlation
between the heavy metals in the tailings
compared to the adjacent soil indicating that these are possibly
from the same source. Worthy of note is
the lack of correlation between the heavy metals studied in the
adjacent soils and Pb. This may indicate
that the origin of Pb in the soils is not the tailings but some
other activity.
The fate and activities of HM in soils and sediments are
influenced by the physicochemical properties
of the soils which dictate their mobility and bioavailability
(Osakwe, 2010; Ngole-Jeme, 2016). Particle
size distribution, pH, CEC, redox conditions, Sulphur (sulphide
and sulphate content) and organic
matter contents are known to be the major soil properties
affecting HM behavior in mining
environment. The acidic pH of the tailings and soil samples
indicates that the metals and metalloids in
the samples around the different sites will be labile as HM
mobility increases under acidic conditions.
The mobility of these contaminants would also be enhanced by the
low organic matter content recorded
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in both the tailings and adjacent soils, as organic matter
contains various functional groups which
improve the capacity of soil to bind metal ions
(Hernandez-Soriano and Jimenez-Lopez, 2012). The
fine texture of the exposed tailings will increase their
erodibility by wind which would result in their
dispersal to surrounding environments. The fine texture of the
tailings is therefore a contributory factor
to the spatial extent of HM contamination around mining
environments. The presence of the sulphide of
most metals are precipitated over a wide pH range whereas
sulphates increases acidity in soil which
mobilizes HM. The effect of sulphur on the behavior of HM may
therefore depend on whether the
reduced form of sulphur (sulphide) or the oxidized form
(sulphate) is present.
The implication of the observed associations could be felt in
various environmental receptors. Metals
associated with mines located in this area have been reported to
travel hundreds of kilometres and
impact downstream ecosystems (Olobatoke and Mathuthu, 2016).
Considering that these particles could
be laden with HM, they could be a source of human exposure to
HM. The health risk associated with
incidental inhalation and ingestion of HM contaminated tailings
particles have been highlighted by
Ngole-Jeme and Fantke (2017). High incidence of dust related
diseases has been reported among the
residents living around the mine dumps and this problem continue
to increase as a result of increased
re-mining of the tailing dams due to recent high price of gold
and development of new extraction
technology (Cairncross,2013). The particle size of the tailings
as determined in this study will pose a
significant health problem. A number of potentially hazardous
substances are usually found in the dust
particles, some of which are more soluble in human physiological
fluids and available for absorption by
the body. These particles when inhaled are usually deposited in
the respiratory system which can results
in a series of respiratory disorders.
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High soil organic matter and clay content increase cation
adsorption in the soil and consequently the
CEC of the soil. The tailings and the surrounding soils samples
however had very low CEC which
would further enhance cation mobility in the soil environment.
The various values recorded for redox in
the tailings and surrounding soils showed that all the samples
examined had moderately reduced redox
conditions (Seo and DeLaune, 2010). This is as a result of
oxygen consumption by lithotrophic iron
reducers present in the samples to enable them use the metals
for their growth and metabolic activities.
This would have resulted in the lowering of the redox potential
of the samples (Chuan et al., 1996;
Bohrerova, 2004).
Implications of physicochemical properties of the tailings,
soils and HM contents on bacterial
activities and diversities
As a result of the low pH and high metal concentrations recorded
in the tailings and adjacent soil
samples, the metallophilic and acidophilic sulphur and iron
oxidizing bacteria such as the
Acidithiobacillus spp, Acidiphillum spp are the predominant
bacteria that can survive in these tailings
and the adjacent soil samples. This has also been explained by
Natarajan, (2008), and Fashola et al.,
(2015). Several other bacteria such as Bacillus spp,
Arthrobacter spp, Pseudomonas spp,
Achromobacter spp, Streptomyces spp and many others with ability
to thrive in the acidic and high
metals conditions have also been identified (Jamaluddin et al.,
2012; El Baz et al., 2015; Ndeddy Aka
and Babalola, 2017). The low content of organic matter in both
the tailings and adjacent soils will result
in reduction of bacterial biomass and extractable carbon as well
as bacterial community structure and
biodiversity. Similar observations have been made by Šourková et
al., (2005), and Laudicina et al.,
(2015). But the low C:N ratio will help the bacteria present to
synthesize their proteins.Insufficient
moisture content will lower microbial activity as microbes
require adequate moisture for their growth
and metabolic activities. Redox potential is also an important
soil property that affects all biochemical
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reactions and enzymatic activity of the bacteria cells (Husson,
2013). It greatly affects availability of
substrate and energy transformation which play a vital role in
regulating soil microbial abundance,
diversity and community structure (Song et al., 2008). The
moderately reducing conditions observed in
the tailings and surrounding soils will limit the bacterial
community to those that can survive the
moderately reduced redox conditions.
The characteristics of the tailings and adjacent soil present
potential challenges for the establishment of
a healthy soil bacterial community. There are likely important
dynamic feedbacks between the tailings
and soil bacteria and plant establishment and broader ecosystem
restoration. Understanding what
stimulates soil-like microbial communities could help spur this
restoration.
CONCLUSION
Gold mining represent one of the major anthropogenic activities
that leads to the release of HM into the
broader environment. Physicochemical properties of the mine
waste sites and the adjacent soils pose
substantial challenges to reclamation. However, HM resistant
bacteria as we have isolated from the
mine wastes could be explored as a potential future in-situ
inoculant to reduce the bioavailability and
mobility of HM and spur plant growth and shifts towards more
healthy soil-like communities
dominated by heterotrophs. Next-steps in this regionally and
globally important African mining region
could build on the current work using molecular methods to
better characterize community structure
and network interactions by determining mechanisms of bacterial
resistant to the HM, by determining
optimal environmental conditions needed for inoculate
utilization in removing HM from these and
similar polluted environments.
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Acknowledgments: Fashola Muibat Omotola acknowledges the
sponsorship of Tertiary Education
Trust fund (TETFUND) and the scholarship of North West
University, Mafikeng. OOB also gratefully
acknowledges NRF for grant (UID81192) that supports work in her
laboratory.
Author Contributions: Muibat Omotola Fashola, Veronica Mpode
Ngole-Jeme and Olubukola
Oluranti were involved in samples collection, wet laboratory
analyses, and drafting and finalization of
the manuscript for publication.
Conflict of interest: There is no competing interest in relation
to this manuscript.
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Figure 1: Map showing the sampling locations
Figure was created using Arc Map version 10.2 and assembled from
the following data sources: Data
points (from field trip excel sheet), Contour lines (WR2000),
Shape files (WR2000). Base map from
WR2000, courtesy of WRC South Africa’’. The study area was
Krugersdorp (26061S and 270 461E), a
mining town in Gauteng province, South Africa. The three sites
as indicated in Figure 1 were MA
(27.80764 E, -26.14265 S), MB (27.81576, -26.12771) and TS
(27.80362, -26.13191).
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MA MAC MB MBC TS TSC0
0.5
1
1.5
2
2.5
3
N
C
S
key
Sampling sites
% o
f nut
rient
s
Figure 2: Average concentration of total nitrogen (N), carbon
(C) and Sulphur (S) across mine sites.
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Conc
entr
atio
n (m
g/kg
)Co
ncen
trat
ion
(mg/
kg)
Conc
entr
atio
n (m
g/kg
)Co
ncen
trat
ion
(mg/
kg)
Conc
entr
atio
n (m
g/kg
)Co
ncen
trat
ion
(mg/
kg)
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Figure 3: Total concentrations of As, Cd, Co, Ni, Pb, and Zn
recorded during the 8 months sampling periods.
C L Y O B L S T C F I I S E F R R C P N0
10
20
30
40
50
60
70
80
90
100
Frequency %
Figure 4: Colonial and cellular morphology of the bacterial
isolates
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Keys: R, L, Y, O and B = cream, light brown, yellow, orange and
brown coloration, L, S and T = large, small and tiny sizes, C, F
and I = circular,
filamentous and irregular shape, I, S and E = irregular,
serrated and entire margin, F, R = flat and raised elevation, R and
C = rod and cocci cell
shape, P and N = Gram positive and Gram negative Gram
reaction.
Table 1: Physicochemical and chemical properties of soils and
tailings samples
Properties MA MAC MB MBC TS TSC
Location Kagiso Kagiso Chamdor Chamdor Tudor shaft Tudor
shaft
Clay (%) 16 24 24 28 26 28
Silt (%) 28 12 12 44 18 34
Sand (%) 56 64 64 28 56 38
Texture Sandy loam Sandy clay loam Sandy loam Silt loam Sandy
clay loam Sandy clay loam
Moisture content (%) 3.88 1.53 2.46 1.91 6.61 4.22
Ph 2.69 5.09 3.37 5.34 4.05 6.34
Redox (mV) 301.00 162.50 209.75 126.50 167.75 104.25
EC( mS/m) 4.59 0.09 2.49 0.20 4.07 6.34
CEC (meq/100) 15.84 21.10 20.31 13.93 10.07 13.65
Organic matter (%) 0.37 1.13 0.89 0.59 0.15 0.45
Sulphate (mg/kg) 184.25 9.19 152.64 3.72 296.62 17.77
Nitrates (mg/kg) 0.91 2.95 0.82 3.58 34.87 33.16
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Table 2: Recommended standards of heavy metals in soils and
sediments
Element MA MB TS
MA MAC MB MBC TS TSC
South
AfricaaCanada b Netherlandsc
As < 1 < 4
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Table 3: Total number of metal species tolerant bacteria
isolated from the three sites
Sampling
Site
Total number
of isolates
Number of
selected isolates
Designates
MA 19 8 OM6 142, OMF 002, OMF 811, OMF 812, OMF 813, OMF
815,
OMF 816, OMF 810
MAC 23 4 OMF 817, OMF 818, OMF 814, OMF 809
MB 22 8 OMF 001, OM4 274, OMF 808, OMF 807, 0M9 107, OMF
806,
OMF 804, OMF 805
MBC 16 3 OMF 819, OMF 820, OMF 821
TS 24 7 OMF 532, OMF 008, OMF 132, OMF 003, OMF 802, OMF
801,
OMF 321
TSC 13 5 OMF 832, OMF 835, OMF 800, OMF 803, OMF 005
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Total number of isolates 117 35 35
Key: TS (Tudor shaft), TSC (Tudor shaft surrounding soil), MA
(Mine tailings A), MAC (MA surrounding soil), MB (Mine tailings
B), MBC (MB surrounding soil).
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Table 4: Biochemical characteristics of the bacterial
isolatesTESTS
ISOLATECODES
MO
T
CA
T
OX
I
NIT
IN VP
MR
CIT
UR
E
SH GL
H2S
MA
N
XY
L
GLU
AR
A
SUC
GA
L
FRU
MA
L
LAC
OMF 012 + + + + + ─ ─ + ─ + + ─ ─ + + + + + + + +OMF 811 ─ + ─ +
─ + ─ + + + ─ ─ ─ + + + + + + + +OMF 812 ─ + ─ + ─ + + + ─ + ─ + +
+ + + + + + + +OMF 813 + + + + + + + ─ ─ ─ ─ ─ ─ ─ + + + + ─ ─ +OMF
814 + + + + + ─ ─ + ─ + + ─ ─ + + + + + + + +OMF 815 ─ + + + ─ ─ ─
─ ─ ─ ─ ─ ─ ─ + ─ ─ ─ ─ ─ ─OMF 816 + + + ─ ─ + + ─ ─ ─ + + + + + +
+ + + + +OM6 142 ─ + + + ─ + + ─ ─ + ─ ─ + + + + + + + + +OMF 810 +
+ ─ + ─ + + + ─ + + ─ + + + ─ + + + + +OM8 321 + + + ─ ─ + ─ ─ ─ ─
─ ─ + + + + + + + + +OMF 001 ─ + + + ─ + + + ─ + + ─ + ─ + ─ + + ─
+ ─OMF 809 + + ─ + ─ + ─ + ─ + ─ ─ + + + + + + + + +OMF 274 + + + ─
─ + ─ + ─ + ─ + ─ + ─ ─ ─ + ─ + +OMF 808 ─ + ─ + + ─ ─ ─ ─ + ─ ─ +
+ + + + + + + +OMF 807 + + ─ + + + + ─ ─ + + ─ + ─ ─ + ─ ─ + + ─OMF
107 + + + ─ ─ + + ─ ─ ─ ─ ─ ─ ─ + ─ + + + + +OMF 806 ─ + ─ + + ─ ─
─ ─ + ─ ─ + ─ + + + + + + +OMF 805 + + ─ + ─ + + + ─ + ─ + + + + +
+ ─ + + +OMF 804 ─ ─ + ─ ─ + ─ + + ─ + ─ + ─ ─ ─ ─ ─ ─ + ─OMF 532 ─
+ ─ + ─ ─ + + ─ + ─ ─ + + + + + ─ + + +OMF 008 ─ + + + ─ + ─ ─ ─ +
─ + + + + ─ ─ + + + +OM7 132 ─ + ─ + ─ + ─ + + + ─ ─ ─ + + + + + +
+ +OMF 003 + + + ─ ─ ─ ─ + + ─ + ─ ─ ─ + + + ─ + + +OMF 803 ─ + + ─
─ ─ ─ ─ + + + + + + + + + + + + +OMF 802 ─ + + ─ ─ ─ ─ + ─ + + ─ +
─ + + + + ─ + +OMF 801 ─ + ─ + + ─ ─ + + + + ─ + ─ + + + ─ + + +OMF
800 + + ─ + ─ + ─ + + + ─ ─ + + + + + + + + +OMF 005 ─ + + + + + +
─ ─ + ─ ─ + + + + + + + + +OMF 817 + + ─ + ─ + ─ + ─ + + ─ + ─ + ─
+ + + + +OMF 818 + + ─ + + ─ + + + + ─ ─ ─ + + + + + + + +OMF 819 ─
+ + + ─ ─ + ─ + + ─ + + + + + ─ ─ + + ─OMF 820 ─ + + + ─ ─ ─ + + +
+ ─ + + + ─ + + ─ + ─OMF 821 ─ + ─ ─ ─ ─ + + ─ ─ ─ ─ + ─ ─ + + + +
+ +OMF 832 + + + + + + ─ + ─ + + + + + + + + + + + ─OMF 835 + + + +
─ ─ ─ + + + ─ ─ + ─ + ─ ─ + + + ─Key : MOT (Motility), CAT
(Catalase), OXI (Oxidase), NIT(Nitrate), IN (Indole), VP (Voges
Proskaeur), MR (Methyl red), CIT (Citrate), URE (Urease), SH (
Starch
hydrolysis), GL (Gelatin liquefaction), MAN (Mannose), XYL
(Xylose), GLU (Glucose), ARA (Arabinose), SUC (Sucrose), GAL
(Galactose), FRU (Fructose), MAL (Maltose), LAC (Lactose).
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Table 5: Probable identity of the bacterial isolates
Isolates code Putative identity
OMF 012 B. amyloliquefaciensOMF 811 Bacillus sp.OMF 812
Enterobacter spOMF 813 B. methylotrophicusOMF 814 Bacillus spOMF
815 Micrococcus yunnanensisOMF 816 B. pumilusOM6 142 E. faecalisOMF
810 B. subtilisOM8 321 Bacillus spOMF 001 B. pumilusOMF 809 B.
anthracisOMF 274 E. aerogenesOMF 808 B. cereusOMF 807 Arthrobacter
oxydansOMF 107 B. thuringiensisOMF 806 Arthrobacter spOMF 805
Enterobacter spOMF 804 Rhodococcus spOMF 532 E. asburiaeOMF 008
Alcaligenes spOM7 132 Enterobacter spOMF 003 B. cereusOMF 803
Arthrobacter spOMF 802 A. nigatensisOMF 801 B. cereusOMF 800 B.
psychoduransOMF 005 Enterococcus spOMF 817 Serratia marcescensOMF
818 Citrobacter spOMF 819 M. variansOMF 820 M. luteusOMF 821
Delftia acidovoransOMF 832 Aeromonas hydrophiliaOMF 835 Burkhoderia
cepacia
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Table 6: Maximum tolerable concentration of the tested metals
against the bacterial isolatesIsolate codes MTC (mM)
Co2+ Ni2+ Pb2+ Cd2+ Zn2+ Cr2+OMF-012 1 2 4 1 3 8OMF-811 1 2 5 1
3 9OMF-812 1 2 4 1 4 9OMF-813 1 3 5 2 5 7OMF-814 1 3 6 2 5 9OMF-815
1 2 6 2 6 8OMF-816 1 3 4 1 4 8OM6-142 1 5 6 2 7 9OMF-810 1 5 6 2 7
9OM8-321 1 4 6 2 7 9OMF-001 1 4 5 2 7 9OMF-809 1 4 6 3 7 9OM4-274 1
4 6 3 6 7OMF-808 1 2 3 1 3 9OMF-807 1 2 4 2 4 10OMF-107 1 4 7 2 9
9OMF-806 1 4 6 2 5 9OMF-805 1 4 6 2 4 8OMF-804 1 2 5 2 4 8OMF-532 1
5 7 3 9 10OMF-008 1 4 6 3 6 9OM7-132 1 4 6 2 7 9OMF-003 1 5 7 3 9
10OMF-803 1 4 5 2 5 9OMF-802 1 4 5 2 5 9OMF-801 1 4 5 1 5 9OMF-800
1 3 6 2 5 10OMF-005 1 5 7 2 7 9OMF 817 1 2 4 2 4 7OMF 818 1 3 6 1 3
8OMF 819 1 2 5 3 3 7OMF 820 1 3 3 2 5 9OMF 821 1 2 4 2 7 6OMF 832 1
2 4 1 7 7OMF 835 1 4 4 2 5 6
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Table 7 : Correlations of the Physico chemical parameters with
heavy metals in adjacent soils
pH EC (m
S/cm
)
Redo
x
% M
oist
ure
Org
anic
m
atte
r
CEC
Nitr
ate
Sulp
hate
mg
/ L Tota
l N
itrog
en
Carb
on
Sulp
hur
As Cd Co Ni
Pb Zn
pH 1.00 EC (mS/cm) 0.71 1.00 Redox -0.39 -0.06 1.00 % Moisture
0.43 0.59 0.08 1.00 Organic matter 0.05 -0.33 0.25 -0.33 1.00 CEC
-0.62 -0.65 -0.12 -0.68 0.02 1.00 Nitrate 0.20 0.66 0.62 0.58 -0.21
-0.48 1.00 Sulphate 0.39 0.33 -0.55 0.31 -0.24 0.18 -0.03 1.00
Tot-Nitrogen 0.29 -0.13 -0.11 -0.43 0.29 0.04 -0.32 -0.06 1.00
Carbon 0.00 0.08 -0.39 0.00 -0.35 0.35 -0.17 0.66 0.16 1.00 Sulphur
0.20 0.22 -0.46 0.07 -0.38 0.16 0.04 0.35 -0.49 0.01 1.00 As 0.70
0.85 -0.27 0.68 -0.41 -0.39 0.54 0.68 -0.22 0.26 0.44 1.00 Cd 0.49
0.87 0.26 0.68 -0.30 -0.54 0.88 0.25 -0.32 -0.04 0.23 0.83 1.00 Co
0.66 0.92 -0.06 0.72 -0.37 -0.53 0.70 0.45 -0.30 0.07 0.35 0.95
0.94 1.00 Ni 0.68 0.90 -0.14 0.72 -0.40 -0.47 0.66 0.57 -0.27 0.17
0.40 0.98 0.89 0.98 1.00 Pb 0.20 0.11 -0.32 0.17 -0.30 0.17 0.03
0.60 0.05 0.47 0.31 0.32 0.00 0.12 0.33 1.00 Zn 0.68 0.86 -0.21
0.69 -0.42 -0.41 0.59 0.63 -0.23 0.23 0.43 1.00 0.86 0.96 0.98 0.29
1.00
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Table 8 : Correlations of the Physico chemical parameters with
heavy metals in the tailings samples
pH EC (m
S/cm
)
Redo
x
% M
oist
ure
OM
CEC
Nitr
ate
Sulp
hate
m
g / L
Tota
l N
itrog
en
Carb
on
Sulp
hur
As Cd Co Ni
Pb Zn
pH 1.00
EC (mS/cm) -0.35 1.00
Redox -0.65 -0.07 1.00
% Moisture 0.22 -0.15 -0.16 1.00
Organic matter 0.16 -0.09 -0.41 -0.40 1.00
CEC -0.26 0.12 0.18 -0.49 0.29 1.00
Nitrate 0.27 -0.17 -0.29 -0.01 -0.22 -0.39 1.00
Sulphate mg / L -0.23 0.38 -0.06 -0.10 -0.24 0.22 0.06 1.00
Total Nitrogen 0.41 -0.24 -0.06 0.44 -0.58 -0.21 0.40 0.37
1.00
Carbon -0.15 0.44 -0.21 -0.01 -0.24 -0.06 0.34 0.54 0.28
1.00
Sulphur 0.40 0.23 -0.51 0.43 -0.48 -0.42 0.41 0.38 0.66 0.38
1.00
As 0.28 -0.20 -0.26 -0.08 -0.23 -0.48 0.97 0.06 0.34 0.31 0.38
1.00
Cd 0.23 -0.17 -0.27 -0.06 -0.14 -0.37 0.99 -0.03 0.28 0.32 0.30
0.96 1.00
Co 0.26 -0.10 -0.32 0.06 -0.24 -0.38 0.99 0.05 0.42 0.40 0.47
0.94 0.98 1.00
Ni 0.36 -0.04 -0.43 0.18 -0.34 -0.49 0.93 0.11 0.53 0.44 0.69
0.90 0.89 0.96 1.00
Pb 0.33 -0.02 -0.50 0.43 -0.40 -0.26 0.69 0.39 0.69 0.46 0.81
0.60 0.60 0.75 0.84 1.00
Zn 0.31 -0.07 -0.39 0.16 -0.30 -0.40 0.96 0.09 0.51 0.43 0.60
0.90 0.93 0.99 0.99 0.83 1.00
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