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DOI: 10.1007/s11665-013-0627-7
Microbial Corrosion in Linepipe Steel Under the Influence
of a Sulfate-Reducing Consortium Isolated
from an Oil Field
Faisal M. AlAbbas, Charles Williamson, Shaily M. Bhola, John R. Spear, David L Olson, Brajendra Mishra, and Anthony E. Kakpovbia
(Submitted January 13, 2013; in revised form June 4, 2013)
This work investigates microbiologically influenced corrosion of API 5L X52 linepipe steel by a sulfatereducing bacteria (SRB) consortium. The SRB consortium used in this study was cultivated from a sour oil
well in Louisiana, USA. 16S rRNA gene sequence analysis indicated that the mixed bacterial consortium
contained three phylotypes: members of Proteobacteria (Desulfomicrobium sp.), Firmicutes (Clostridium
sp.), and Bacteroidetes (Anaerophaga sp.). The biofilm and the pits that developed with time were characterized using field emission scanning electron microscopy (FE-SEM). In addition, electrochemical
impedance spectroscopy (EIS), linear polarization resistance (LPR) and open circuit potential (OCP) were
used to analyze the corrosion behavior. Through circuit modeling, EIS results were used to interpret the
physicoelectric interactions between the electrode, biofilm and solution interfaces. The results confirmed
that extensive localized corrosion activity of SRB is due to a formed biofilm in conjunction with a porous
iron sulfide layer on the metal surface. X-ray diffraction (XRD) revealed semiconductive corrosion products
predominantly composed of a mixture of siderite (FeCO3), iron sulfide (FexSy), and iron (III) oxidehydroxide (FeOOH) constituents in the corrosion products for the system exposed to the SRB consortium.
Keywords
biofilm, carbon steel API 5L X52, microbiologically
influenced corrosion, pipeline, sulfate-reducing
bacteria
1. Introduction
Microbiologically influenced corrosion (MIC), or biocorrosion, is of considerable concern to the oil and gas
industry. MIC is induced by indigenous microorganisms that
naturally reside in hydrocarbon and secondary water injection
systems (Ref 1-3). Microbial metabolic products, for example,
sulfide and organic acids such as acetic acid, may alter interface
chemistry, resulting in increased corrosion rates. Furthermore,
biofilms consisting of complex communities of microbes and
extracellular polymeric substances (EPS) that have developed
on the metal surface create gradients of pH and dissolved
oxygen, leading to localized forms of corrosion, such as pitting
and crevice formation (Ref 1-4).
Since the advent of modern oil and gas production, scientists
and engineers have faced problems caused by microorganisms
(Ref 5). Hydrocarbons are an excellent carbon (food) source for
a wide variety of microbes in all three domains of life—the
Faisal M. AlAbbas, Shaily M. Bhola, David L Olson, and Brajendra
Mishra, Department of Metallurgical and Materials Engineering,
Colorado School of Mines, Golden, CO 80401; Charles Williamson
and John R. Spear, Department of Civil and Environmental
Engineering, Colorado School of Mines, Golden, CO 80401;
and Anthony E. Kakpovbia, Inspection Department, Saudi Aramco,
Dhahran 31311, Saudi Arabia. Contact e-mail: falabbas@mines.edu.
Journal of Materials Engineering and Performance
Bacteria, Archaea and Eucarya, and microbial representatives of
all the three domains likely play roles in MIC (Ref 2). Bacterial
activity (e.g., sulfate reduction) is believed to be responsible for
greater than 75% of the corrosion in production oil and gas wells,
and it is likely that greater than 50% of failures of buried
pipelines and cables are due to the metabolic activities of
microbes (Ref 5). The main types of bacteria associated with
metals in pipeline systems are sulfate-reducing bacteria, iron
reducing bacteria, iron and manganese oxidizing bacteria, and
acid producing bacteria (Ref 1, 3, 5). For several reasons, SRB
have been recognized as the major contributor to MIC in pipeline
systems primarily due to their predominantly anaerobic lifestyle
and continuous production of corrosive hydrogen sulfide (Ref 3, 5).
However, in practical situations, MIC results from synergistic
interactions of different microbial consortia, which coexist in the
environment and are able to affect the electrochemical processes
through co-operative metabolisms (Ref 4).
As early as 1926, traditional microbiologists detected the
presence of SRB in oil environments (Ref 5). Studies have shown
that most kinds of cultivated SRB are responsible for the
production of hydrogen sulfide, which is a toxic and corrosive gas
in both aerial and aqueous form (Ref 4, 5). At a minimum,
hydrogen sulfide is thought to be responsible for a variety of
environmental effects which include reservoir souring, contamination of natural gas and oil with H2S, corrosion of metal
surfaces, the ‘‘plugging’’ of reservoirs due to the formation of
extensive pore-space microbial biofilms and the precipitation of
metal sulfides (Ref 5). SRB has received much attention in the oil
& gas industry and MIC investigations, as these microorganisms
have several detrimental metabolic activities including the ability
to: (1) oxidize hydrogen, (2) use O2 and Fe3+, (3) utilize aliphatic
and aromatic hydrocarbons, (4) couple sulfate reduction to the
intracellular production of magnetite and (5) compete with
nitrate-reducing/sulfur-oxidizing bacteria (NRB-SOB) (since
they may have a nitrite reducing activity) (Ref 5, 6). In total, all
of these activities can significantly degrade the quality of oil and
gas in a reservoir to be economically extractable. Moreover, SRB
adversely impacts the integrity of oil and gas installations.
This study investigates the impact of environmental SRB
(cultivated from oil field samples rather than obtained from a
culture collection) on the corrosion behavior of API 5L X52
linepipe steel. The SRB consortium used in this study was
cultivated from an oil well in Louisiana, USA (Ref 7). 16S
rRNA gene sequence analysis was performed to identify the
SRB species. The nature and kinetics of chemical and
electrochemical reactions introduced by SRB activities on
API 5L X52 carbon steel coupons were characterized using
electrochemical impedance spectroscopy, linear polarization
resistance (LPR) and open circuit potential (OCP). The biofilm
and corrosion morphology were investigated using field
emission scanning electron microscopy coupled with energy
dispersive spectroscopy (EDS). The composition of corrosion
products was evaluated using x-ray diffraction analysis.
2. Materials and Methods
2.1 Microorganisms and Testing Medium
The SRB consortium used in this study was cultivated from
water samples obtained from an oil well located in Louisiana,
USA. The water samples were collected and bottled at the well
head from an approximate depth of 2200 ft. as described under
the NACE Standard TM0194 (2004) (Ref 8). The SRB were
cultivated in a modified Baar!s medium (ATCC medium 1250).
Baar!s medium is reported to be suitable when studying the
influence of mixed bacterial communities on steel corrosion,
though many different media recipes exist (e.g., the Postgate
series of media) and any number of them likely work (Ref 9).
This growth medium was composed of magnesium sulfate (2.0 g),
sodium citrate (5.0 g), calcium sulfate di-hydrate (1.0 g),
ammonium chloride (1.0 g), sodium chloride (25.0 g),
di-potassium hydrogen orthophosphate (0.5 g), sodium lactate
60% syrup (3.5 g), and yeast extract (1.0 g). All components
were per liter of distilled water. The pH of the medium was
adjusted to 7.5 using 5 M sodium hydroxide and sterilized in an
autoclave at 121 "C for 20 min. The SRB species were cultured
in the growth medium with filter-sterilized 5% w/w ferrous
ammonium sulfate added to the medium at a ratio of 0.1-5.0
mL respectively. The bacteria were incubated for 72 h at 37 "C
under an oxygen-free nitrogen headspace.
2.2 Sulfate-Reducing Consortium Identifications
Genomic DNA was extracted from the bacterial consortium
using the MoBio Powersoil DNA extraction kit (MoBio,
Carlsbad, CA) with the 10-min vortexing step replaced by
1 min of bead beating. Primers 515F (5¢-GTGCCAGCMG
CCGCGGTAA-3¢) and 1391R (5¢-GACGGGCGGTGWGT
RCA-3¢) (Ref 7, 10) were used to amplify 16S rRNA genes.
Polymerase chain reaction (PCR), cloning and transformation
were conducted as described by Sahl et al. (Ref 11). Unique
restriction fragment length polymorphisms (RFLP) were
sequenced on an ABI 3730 DNA sequencer at Davis Sequencing, Inc. (Davis, CA), and Sanger sequence reads were called
with PHRED (Ref 12, 13) via Xplorseq (Ref 14). Sequences
were aligned with the SINA aligner (Ref 15) and added
(parsimony insertion) to the Silva SSURef111_NR guide tree
(Ref 16) with the ARB software package (Ref 17). Sequences
were also compared to the Genbank database (Ref 18) via the
Basic Local Alignment Search Tool (BLAST) (Ref 19).
Phylogenetic trees were created with RAxML (Ref 20).
Relevant sequences were obtained from the Genbank database
and aligned and masked with ssu-align (Ref 21). Phylogenetic
trees were created using the GTR substitution model and the
gamma distribution of rate heterogeneity. The number of
bootstrap replicates was determined using the RAxML frequency-based criterion (Ref 22). 16S rRNA gene sequences
produced during this research have been deposited in the
Genbank with accession numbers KC756849-KC756851.
2.3 Sample Preparation
Pipeline steel (API 5L X52) coupons, provided by a local
energy company (Saudi ARAMCO), were used for this study,
and their chemical compositions are shown in Table 1.
Metallographic specimens from the received materials were
prepared with standard methods for optical microscopy (1 lm
final polish and 2% nital etch). Representative microstructure
contains a mixture of polygonal ferrite and pearlite structures.
For corrosion evaluations, the coupons were machined to a
size of 10 mm 9 10 mm 9 5 mm and embedded in a mold of
non-conducting epoxy resin, leaving an exposed surface area of
100 mm2. For electrical connection, a copper wire was soldered
at the rear of the coupons. The coupons were polished with a
progressively finer sand grinding paper until a final grit size of
600 lm was obtained. After polishing, the coupons were rinsed
with distilled water, ultrasonically degreased in acetone and
sterilized by exposing to pure ethanol for 24 h.
2.4 Electrochemical Tests
The electrochemical measurements were made in a conventional three electrode ASTM cell coupled with a potentiostat and a
high frequency impedance analyser (Gamry-600). The electrochemical cells were composed of a test coupon as a working
electrode (WE), a graphite electrode as an auxiliary electrode and a
saturated calomel electrode (SCE) as a reference electrode. The
glasswares were autoclaved at 121 "C for 20 min at 20 psi pressure
and then air dried. Graphite electrodes, purging tubes, rubber
stoppers and needles were sterilized by immersing in 70 vol.%
ethanol for 24 h followed by exposure to a UV lamp for 20 min.
Two solutions were used in this experiment. Under a sterilized
condition (in a sterilized laminar flow hood), the first cell was
prepared with 600 mL of sterilized modified Baar!s growth media
(described above) and the second cell was prepared with 600 mL
sterilized Baar!s media inoculated with 5 mL of the SRB
consortium at 106 cell/mL. The electrochemical cells were purged
for 1 h with pure nitrogen gas to establish the anaerobic
environment. Electrochemical tests were performed at a flow rate
of 0.9 m/s (100 rpm) under atmospheric pressure conditions at
30 "C, to simulate the flow conditions of Saudi Aramco oil
production pipeline.
Open circuit potential values of the systems were monitored
with time during the immersion period followed by periodic
readings up to 336 h. Impedance measurements were performed
on the system at the OCP for various time intervals from
immersion up to 288 h. The frequency sweep was applied from
105 to 10!2 Hz with an AC amplitude of 10 mV.
Journal of Materials Engineering and Performance
Table 1 The chemical composition of API-5L X52 carbon steel coupons, wt.%
Fe
C
Si
Cr
Ni
Mn
Cu
Mo
Nb
Ti
Al
V
S
P
Bal.
0.070
0.195
0.03
0.02
1.05
0.05
0.004
0.021
0.001
0.029
0.003
0.008
0.008
During the LPR technique, the polarization resistance (Rp)
was measured on the system at a scanning amplitude of
±10 mV with reference to the OCP for various time intervals
from immersion up to 336 h. The LPR plots were then fitted
using the Gamry-600 electrochemical software to obtain the
goodness of fit for corrosion rate values. The values were then
compared using the mathematical calculation as determined by
the equation given below (Ref 23);
!
"
1
ba bc
ðEq 1Þ
icor ¼
Rp 2:3ðba þ bc Þ
where ba and bc is the anodic and cathodic slope, respectively.
2.5 Surface Analysis and Corrosion Product Compositions
of the Coupons Exposed to SRB
At the conclusion of each test, the WEs were carefully
removed from the system for fixation. To fix the grown biofilm
to the steel surface, the coupons were immersed for 1 h in a 2%
glutaraldehyde solution, dehydrated with 4 ethanol solutions
(15 min each): 25, 50, 75, and 100% successively, air dried
overnight and then gold sputtered (Ref 24). After fixation, the
coupons were examined using field emission scanning electron
microscopy coupled with EDS to evaluate the morphology and
chemical composition of the biofilm. Corrosion product
composition was obtained using the x-ray diffraction method
with a Philips PW 3040/60 spectrometer using Cu Ka radiation
source. The coupons were then cleaned using a standard
protocol described under the ASTM-GI-03 (Ref 25), and the pit
morphology and density on the exposed coupons were
examined using FE-SEM.
3. Results and Discussion
3.1 Identification of the Sulfate-Reducing Consortium
16S rRNA gene sequence analysis indicated that the mixed
bacterial culture consortium contained three phylotypes that are
close to members of the Proteobacteria (Desulfomicrobium
sp.), Firmicutes (Clostridium sp.), and Bacteroidetes (Anaerophaga sp.) (Ref 7). The phylogenetic tree representative of the
bacterial consortium has been shown in Fig. 1. Desulfomicrobium, including mesophiles and thermophiles, are known to
be commonly associated with oil reservoirs. They have been
isolated previously from different oil fields in the North Sea
(Ref 26) and in 5 of 6 different Alberta oil fields in Canada as
observed by Voordouw and colleagues (Ref 27). Leu et al.
(1999) reported the presence of different strains of Desulfomicrobium obtained from various samples in distant oil fields
such as formation water and drilling mud (Ref 26). Desulfomicrobium sp. are anaerobic, Gram-negative, rod-shaped,
sulfate-reducing bacteria that grow on different carbon source
substrates that include lactate, pyruvate, glycerol, and ethanol
with optimal growth temperatures between 25 and 35 "C
Journal of Materials Engineering and Performance
(Ref 27). They are capable of using sulfate, thiosulfate or sulfite
as a terminal electron acceptor (Ref 6, 28). Several researchers
have further reported that in the presence of 0.5% NaCl,
Desulfomicrobium metabolize lactate and sulfate to produce
acetate, CO2 and H2S as major end products (Ref 26-28). These
microorganisms can play a significant role in oil field reservoir
souring by the reductive generation of hydrogen sulfide from
these sulfur containing electron acceptors (Ref 28). Clostridium
sp. are anaerobic microbes that are Gram-positive, sporeforming bacteria. This type of bacteria is capable of surviving
high temperatures due to the formation of heat-resistant
endospores (Ref 1, 6, 28). Anaerophaga sp. have also been
previously identified in samples from a produced water
obtained from the high-temperature Troll oil formation in the
North Sea (Ref 28). The cultivation of this consortium was
previously reported (Ref 7).
3.2 Surface Morphology and Element Analysis
At the conclusion of the tests, the visual inspection of the
coupon exposed to the biotic system revealed dense, thick and
black products covered on the surface. There is a significant
difference in the appearance, structure, and morphology of the
corrosion products developed on the steel coupons exposed to
the biotic system compared to that exposed to the abiotic
system as shown in Fig. 2 and 3. Both the morphological
observations and EDS elemental analysis of corrosion products
of API X52 steel immersed in the biotic system are shown in
Fig. 2(a)-(c). As shown in Fig. 2(a), there are two distinctive
areas: A1 and A2. Quantitative EDS analysis shows A2 is
composed of a higher amounts of sulfide, sodium salt,
phosphate and carbon (Fig. 2(b) and Table 2). The light region
(A2) is considered the outer layer where the sulfur is the
predominant constituent, Fig. 2(b). On the other hand, the dark
region, A1, is considered to be the inner layer in which the iron,
oxygen, and carbon, in addition to sulfur and phosphorous, are
predominant, as shown in Fig. 2(c) and Table 2. The outer and
inner areas were chosen based on our previous research
experience, and no cross section view was performed on the
sample (Ref 7). The spherical characteristic structures shown in
Fig. 2(a) might represent siderite (FeCO3) surrounded by a
mixture of iron sulfide (FeS) and iron oxides.
Similar features have been reported by Hendrik Venzlaff et al.
for steel surfaces exposed to a SRB strain, Desulfopila
corrodens. The fact that Desulfomicrobium have the capacity
to convert the carbon source (lactate) through pyruvate to acetate
along with the production of carbonate supports the formation of
FeCO3 (Ref 6, 29). Furthermore, the presence of di-potassium
hydrogen orthophosphate and sodium chloride in the growth
media might lead to the precipitation of phosphorous-based
compounds and sodium chloride on the surface as suggested by
the EDS spectra (Ref 30).
The morphological observations and elemental analysis of
surface deposits and corrosion products on API X52 carbon
steel coupons exposed to the abiotic system are shown in
Fig. 3(a) and (b). There is one coherent homogenous layer of
corrosion product with salt crystal deposits on the surface. As
Fig. 1
The phylogenetic characterization of SRB consortium
Fig. 2 FESEM and EDS analysis for the system under biotic conditions. (a) FESEM Image of carbon steel exposed to Barr!s medium inoculated with SRB, at 1009. Arrow points to a magnified image (2009) of iron carbonate layer. EDS analysis corresponding to the FESEM outer
and inner region shown in Fig. 2(b) and (c), respectively
Journal of Materials Engineering and Performance
Fig. 3
FESEM and EDS analysis on the API X52 exposed to sterilized Baar!s medium
Table 2 Comparative of EDS analysis corresponding to the abiotic and the biotic systems, respectively
Element, wt.%
Abiotic system
Whole region
Biotic system
Outer region (A2)
Inner region (A1)
Fig. 4
C
O
Na
Si
Fe
S
Cl
Mn
Ca
P
Total
0.01
18.73
4.54
…
68.38
1.54
4.61
2.11
0.08
0.00
100
14.99
10.35
4.20
23.58
8.66
6.96
1.69
0.89
10.06
26.98
55.09
22.43
0.00
0.00
0.00
0.00
0.00
0.00
6.84
8.72
100
100
FESEM image for the biofilm developed on the API X52 exposed to the SRB-containing medium 10009 and 20009
shown in Fig. 3(b) and Table 2, quantitative EDS analysis
shows that this layer might be composed of iron oxides mixed
with sodium chlorides, calcium and carbon-based compounds
that accumulated from the growth medium (Ref 30).
As shown in Table 2 and Fig. 2 and 3, the biotic and abiotic
systems displayed significant differences in distribution and
composition of corrosion products. In the presence of the SRB
consortium, there is significant accumulation of sulfur-based
compounds and FeCO3, and a substantial amount of corrosion
Journal of Materials Engineering and Performance
products growing upward is observed, and the interface does
not have a rigid appearance, most likely due to the biofilm
matrix, nature of which is polysaccharidic and viscoelastic
(Fig. 2 and 4), while the corrosion products for the abiotic
system exhibits a completely different thin-flat layer with a hard
texture (Fig. 3) (Ref 1-4).
The biofilm developed in the presence of the SRB
consortium together with the produced corrosion products have
a heterogeneous morphology and thickness (Fig. 4). The
ο
14000
∇
12000
ο
•
⊗
Intensity (CPS)
10000
8000
6000
4000
2000
⊗
∇
•
ο
∇
⊗
+
ο
ο
0
20
40
60
80
100
2θ (degree)
Fig. 5 XRD spectra for the system under biotic conditions for 14
days exposure with SEM image of the examined surface
FESEM micrograph (Fig. 4) reveals the presence of the
corrosion products, cells, spores, and EPS fibers distributed
over the coupon. At the conclusion of the experiment, the steel
substrate was hardly visible. It was covered with a porous black
layer. A jelly-like substance could be observed among the
corrosion products, which was speculated to be biofilm
produced EPS (Ref 30, 31). Rod-shaped and round shaped
bacteria occupied a significant volume fraction qualitatively,
(no quantitative measurements were performed); however, EPS
and corrosion products have been reported to occupy 75-95%
of biofilm volume, while 5-25% is occupied by the metabolizing cells (Ref 1, 30). Higher concentrations of hydrogen
sulfide, phosphate-based compounds, and other potentially
biotically-generated, corrosion-influencing compounds are
likely to be promoted by SRB metabolism and biofilm
formation (Ref 1, 3, 30, 31). Conversely, the nature of
corrosion film for the abiotic system exhibits a completely
different thin-flat layer with a hard texture (Fig. 3).
3.3 XRD Results
The XRD spectra of API X52 steel coupons exposed to the
biotic system are displayed in Fig. 5. The XRD pattern
confirmed the formation of a significant amount of iron sulfide
compounds that include mackinawite (Fe1+xS), other biogenetic
iron sulfide (FeS), siderite (FeCO3), and iron (III) oxidehydroxide (FeOOH) (Ref 29, 32, 33). The formation of pyrite
and mackinawite films in the presence of SRB is expected as a
consequence of their production of sulfide. Due to the very high
reactivity of H2S with iron, the mackinawite layer has been
shown to form, which is much faster than what would have
been expected from the typical kinetics for a precipitation
process. The formed mackinawite is not stable and may
dissolve depending on the solution saturation level. For the pH
ranging between 4 to 7, which is a typical SRB-containing
environment, the solution is often supersaturated with respect to
iron sulfide, and the layer does not dissolve (Ref 34). The iron
sulfide films could protect or deteriorate the surface. Thin
protective films are associated with low ferrous ion concentration rates while active films are believed to form in the presence
of high ferrous ion concentrations. These protective films have
been proposed to fail due to disruption by microbial action,
bulky growth, and oxidation (Ref 34). It has been proposed that
the corrosion rate for steel can be defined by the rates of iron
sulfide layer formation and breakdown (Ref 7).
Possibly, the capacity of the bacterial consortium (Desulfomicrobium & Clostridium) to convert the carbon source
(lactate) through pyruvate to acetate and produce carbonate
resulted in the formation of FeCO3 (Ref 29). Some bacteria
of the Clostridium genus (present in the consortium) are
capable of fermentative utilization of lactate depending on
the pH medium and produced acetate, bicarbonate or
propionic acid that may be relevant to our system; however,
due to our limitations, such measurements have not been
made (Ref 6). It might be possible that the bacterial
consortium work in a synergistic way and affect the
corrosion of the alloy steel.
EDS and XRD results suggested that the following reactions
occurred as a result of metabolic activities of the SRB
consortium and corrosion behavior of the steel surface. The
general chemical and electrochemical reactions can vary
according to the surface content and EDS analyses, but
generally include a cathodic reaction under natural deaerated
conditions (Ref 1, 3, 30, 31, 35):
2H2 O þ 2e! ! H2 þ 2OH!
ðEq 2Þ
The SRB Desulfomicrobium sp. may use cathodic hydrogen
to reduce sulfate to sulfide as follows (Ref 1-4, 31)
þ
!
!
SO2!
4 þ 9H þ 8e ! HS þ 4H2 O
ðEq 3Þ
Reaction [3] normally happens at a slow rate without biocatalysis from bacteria, however; in systems that contain
microbes, the reaction can be rapid, being enzymatically
catalyzed by the hydrogenase enzyme system of Desulfomicrobium sp. Some hydrogen sulfide ions will convert to hydrogen
sulfide especially at acidic pH as described (Ref 3, 30, 33)
HS! þ Hþ ! H2 S
ðEq 4Þ
Reaction [4] is rapidly facilitated by the presence of
Desulfomicrobium sp. at a pH between 1 and 7. The production
of hydrogen sulfide and the oxidation of iron (anodic reaction) lead
to the formation of different types of iron sulfide as follows
(Ref 30, 31):
Fe0 ! Fe2þ þ 2e!
ðEq 5Þ
Fe2þ þ H2 S ! Fey Sx þ 2Hþ
ðEq 6Þ
According to Liu et al. (Ref 36), the ferrous ions (Fe2+)
produced by the dissolution process react with sulfide
metabolized by the bacteria with the subsequent production
of different forms of iron sulfide (FeSx). Therefore, it is
expected that the structure of the biofilm changes according
to the different growth phases of the SRB (Ref 30, 31, 35,
36).
The rate determining step (RDS) for the SRB metabolic
activities, Reaction [3], is the proton reduction to form
hydrogen. The kinetics of this hydrogen reduction are expected
to be slow due to the fact that the concentration of protons is
extremely low under natural de-aerated conditions. It has been
reported that in a de-aerated water environment, the formation
Journal of Materials Engineering and Performance
Fig. 6
FESEM analysis for the API X52 coupon surface after cleaning for the system under biotic conditions at 2009, 20009 and 40009
of hydrogen on the surface is extremely low because of limited
protons (Ref 37).
Hþ þ 1e! ! Hads
ðEq 7Þ
Therefore, when there is not enough hydrogen to drive the
metabolic reaction (Reaction [3]) Desulfomicrobium sp. may
switch to convert the carbon source (lactate) through pyruvate
to acetate with the production of carbonate (Ref 6, 29, 30).
When SRB utilize lactate as an electron donor, they produce
extra ATPs by the proton motive force and the oxidation of
lactate to acetate plus carbon dioxide (Ref 6). The produced
hydrogen crosses the cytoplasmic membrane and is oxidized by
periplasmic hydrogenase to initiate a proton motive force,
which in turn bio-catalyzes Reaction [3] (Ref 4, 6). The prime
reactions of this process are as follows:
2CH3 CHOHCOO! þ 2SO2!
4
!
þ
! 4CH3 COO! þ HCO!
3 þ H2 S þ HS þ H þ CO2
ðEq 8Þ
Carbonate ions will react with ferrous ions and form
insoluble siderite (FeCO3) in addition to iron sulfide (FeS). The
net reaction will be as follows (Ref 29):
Journal of Materials Engineering and Performance
!
4Fe þ SO2!
4 þ 3HCO3 þ H2 O
! FeS ðs) þ 3FeCO3 ðs) þ 5OH! ;
DG& ¼ !86:1 kJ/mol Fe
ðEq 9Þ
Closer inspection of the API X52 carbon steel electrode in
Fig. 6 shows variations in surface roughness. Generalized
corrosion was observed in an irregular pattern on the coupon as
in Fig. 6(a). The images at higher magnifications in Fig. 6(b) also
show preferential etching of selected pearlite grains in a regular,
rectangular, and repetitive manner. The attack appears to be in an
un-preferred orientation. An inter-granular attack that produces
microstructural relief and etching of pearlite microstructures can
be observed in Fig. 6(c) and (d).
The rectangular attack, found predominantly in the more
corroded areas on the surface, may be the result of the galvanic
coupling between the semiconductive iron sulfide, which acts
as a cathode, and the steel surface, or anode. Galvanic corrosion
is either a chemical or an electrochemical corrosion phenomenon. The latter is due to a potential difference between two
different surfaces connected through a circuit that allows for
current flow to occur from more active metal (anode), to less
active surface (cathode). In our case, the iron sulfide is
considered the cathode where the calculated equilibrium
FESEM analysis for the API X52 coupon surface after cleaning for the system under abiotic conditions exposure at 1009 and 5009
potential of sulfate reduction (SO42!/FeS) is !0.25 V, and the
steel surface is the anode with calculated equilibrium potential
of !0.44 V (Ref 29). Anodic corrosion of iron by galvanic
coupling with iron sulfide has been proposed previously
(Ref 37, 38). Galvanic corrosion attacks the iron matrix
adjacent to iron sulfide deposits.
Furthermore, the conductivity of the iron sulfide augments
this coupling between the solution and underlying surface that,
in turn, catalyzes the corrosion process. The conductivity of
heterogeneous corrosion products composed of FeS and
FeCO3 in SRB cultures has been reported to be around 50
Sm!1, which is higher than that of many typical semiconductors such as silicon. This conductivity is mainly due to the
presence of FeS as FeCO3, which is considered an insulating
mineral (Ref 29).
The preferred attacks shown in the pearlite microstructures
(Fig. 6c) are attributed to the nature of heterogeneous structures
of pearlite. Pearlite consists of plates of cementite (Fe3C) in a
matrix of ferrite (Ref 33). The structural and compositional
inhomogeneity within the pearlite structure invites the possibility of this preferred attack and enhances the localized
corrosion (Ref 39). When pearlite is exposed to localized
corrosion, as in our case, galvanic microcells between ferrite
(cathode) and cementite (anode) are generated. Consequently,
the surface will exhibit deeper and larger pits. However, in a
real system where oxygen might exist, the corrosion mechanism becomes quite complicated, and not only the sulfate and
sulfide but also several other ionic species (e.g. chlorides) may
be involved. On the other hand, minimal corrosion damage was
observed in the abiotic system where the polishing marks are
still visible (Fig. 7).
3.4 Open Circuit Potential/Polarization Resistance
The OCP variations for the biotic and abiotic systems are
shown in Fig. 8. The Ecorr as a function of time data revealed
that in the biotic system, a substantial shift of Ecorr towards
noble values (-600 mV/SCE) occurred for the first 100 h and
then remained more or less stable throughout the period of
exposure. This potential shift was attributed to the growth of
the SRB species, their metabolic activities, and subsequent
accumulation of iron sulfide on the surface. SRB attached to the
Biotic System
Abiotic System
-600
OCP (mV) /SCE
Fig. 7
-650
-700
-750
0
50
100
150
200
250
300
350
Time (hrs)
Fig. 8 OCP variations under biotic and abiotic conditions
coupon surface, colonized and reproduced to form a biofilm,
and the activity of microbes in this biofilm subsequently altered
the electrochemical processes taking place at the steel surface.
These alterations include pH changes, H2S production, iron
sulfide formation and even EPS production. These factors
collectively enhance the reduction capacity of the system and
accelerate anodic dissolution (Ref 30, 31, 35, 36).
In stark contrast, the abiotic system had a notable increase of
the Ecorr, which then remained more or less steady at
approximately !690 mV/SCE. This potential shift might be
attributed to the accumulation of the growth medium constituents such as organic compounds, potassium, sodium chloride,
and phosphorous on the coupon surface (Ref 31). There is a
difference in noble direction of approximately 90 mV/SCE
between the biotic and abiotic systems. This positive shift in
Ecorr is known as ennoblement. Ennoblement has been reported
for different alloys exposed to microbes. It is probably the most
Journal of Materials Engineering and Performance
3000
70
Biotic System
Abiotic System
60
Corrosion Rate (MPY)
Rp (Ω.cm2)
2500
2000
1500
1000
500
50
40
30
20
10
0
0
0
(a)
Fig. 9
Biotic System
Abiotic System
50
100
150
200
250
300
350
Time (hrs)
(b)
0
50
100
150
200
250
300
350
Time (hrs)
(a) Polarization resistance (Rp) and (b) corrosion rate variations under biotic and abiotic conditions
notable phenomenon in many MIC investigations (Ref 1-3).
The ennoblement has been attributed to microbial colonization,
biofilm formation and the deposition of sulfide, which collectively result in organometallic catalysis and acidification of the
electrode surface (Ref 1). The accumulation of corrosion
products such as iron sulfide will result in galvanic coupling
with the steel surface and may contribute to this ennoblement
phenomenon. Ennoblement promotes pitting corrosion, which
is more critical for passive alloys (Ref 1-3).
The polarization resistance variations for biotic and abiotic
systems are shown in Fig. 9(a). The Rp as a function of time
data revealed that in the biotic system, a substantial decrease of
Rp to 500 X cm2 was followed by another decrease to
approximately 250 X cm2 at 350 h. There was a noticeable
increase in the Rp to '3000 X cm2 at 100 h. This increase is
attributed to the formation of a film of siderite (FeCO3) and
mackinawite that provide short-term protection of the steel
surface. It has been reported that the mackinawite layer partially
protects the steel from corrosion (Ref 7). The substantial drop
in the Rp is attributed to the formation of a stable biofilm and
conductive iron sulfide mixed layers on the surface. SRB
metabolic activities produce hydrogen sulfide and form organic
compounds such as an EPS with acids at the metal/biofilm
interface (Ref 1, 3, 30). These factors, along with galvanic
coupling between the iron sulfide and underlying surface, create
an aggressive environment that leads to this substantial
decrease of polarization resistance.
In contrast, in the abiotic system, Rp decreased to about
1000 X cm2 at the first 50 h followed by an increase to about
1200 X cm2, which then remained more or less steady
throughout the experiment. The initial drop in the Rp was due
to the corrosion effects of the deposited nutrients (i.e., sulfide
and sodium chloride) on the surface. However, when stable
deposits and a corrosion film were formed on the surface, it
provided protection as indicated by a steady resistance
afterwards (Ref 40).
The polarization resistance is inversely proportional to the
corrosion rate, implying a higher corrosion rate at lower
resistance. The corrosion rate plots over time for the biotic and
abiotic systems are shown in Fig. 9(b).
Journal of Materials Engineering and Performance
The corrosion rate for the biotic system increased significantly after 100 h, which is in agreement with the shift of OCP.
The corrosion rate reached a maximum value of 45 mpy after
250 h, whereas the corrosion rate for the abiotic system for the
same interval is approximately 15 mpy. It should be noted that
the utilization of the LPR method in this study should not
replace or underestimate the weight loss (WL) method in
determining the corrosion rate. The WL is absolutely more
reliable as it provides direct proof of corrosion rather than
indirect electrochemical measurements (e.g., LPR).
Several general statements could be made to explain the
high corrosion rates observed under the biotic conditions. The
microenvironment at the metal-liquid layer could have been
altered by biofilms via attachment of bacteria and deposition of
organic by-products such as: EPS, hydrogen sulfide, and iron
sulfide (Reaction [3] and [8] and Fig. 4). These changes
enhance the kinetics of the corrosion processes at the metal
surface, which was reflected in an increase in Ecorr accompanied by a decrease in Rp. Moreover, the accumulation of iron
sulfide (Reaction [5] and [8]) on the carbon steel coupons forms
a galvanic cell with the adjacent steel surface, resulting in
further enhancement of the corrosion process (Ref 1, 3). The
observed rectangular pits and preferred pearlite microstructures
attacks (Fig. 6) are due to these galvanic couplings. However, it
was unclear what portion of an increase of corrosion resulted
from the changes induced by the bacterial metabolic activities
and what portion was due to other redox couples promoted by
the iron sulfide galvanic coupling.
3.5 Electrical Impedance Spectroscopy Results
Figure 10(a) displays the Nyquist plots for a carbon steel
coupon exposed to the abiotic system. At low frequencies (LF)
(Fig. 10a), the magnitude of the capacitive loop represented by
the semicircle diameter increased with time. These LF magnitudes represent the change in the charge transfer resistance (Rct)
that describes the evolution of the anodic reaction controlled by
charge transfer processes (Ref 30, 31, 35, 41). The resistance
increased to values around 4000 X cm2. The diagram obtained
at 192 and 288 h exhibited a bigger loop diameter, which
6000
Phase Angle (degree)
96 h
4000
-Z" (Ω.cm2)
0h
-80
0 hrs
96 hrs
192 hrs
288 hrs
2000
192 h
-60
288 h
-40
-20
0
0
(a)
Fig. 10
0
1500
3000
4500
6000
(b)
2
Z' (Ω.cm )
10-1
100
101
102
103
104
105
Frequency (Hz)
EIS date abiotic system; (a) Nyquist plots and (b) phase angle plots
Fig. 11 Circuits model used to fit the EIS data for the abiotic system
indicates an increase in charge transfer resistance. Under these
conditions, it was found that the corrosion process occurred in
the first 50 h, and a steady corrosion rate was observed as the
exposure time increased (Fig. 9b). In the abiotic system, the
anodic reaction is represented by reaction (5), and the cathodic
reaction is shown by reaction (2). The steady corrosion rate is
possibly due to the protective effect of a mixed layer of deposits
and corrosion products that are possibly composed of sodium
chloride, sulfide, potassium, and carbon-based compounds on
the electrode surface (Ref 40). The formation of a capacitive
layer on the steel surface is confirmed by phase angle spectra
(Fig. 10b) that displayed one time constant or one peak at 10
Hz. The equivalent electrical circuit based on the minimum
deviation between the measured and fitted data for the abiotic
condition is shown in Fig. 11. The circuit includes:
•
•
10-2
A resistance Rs considered as the solution resistance.
Parallel connection of a charge transfer resistance (Rct) for
the steel surface and constant phase element (CPE) associated with a double layer capacitance due to the formation
of a heterogeneous layer composed of corrosion products
along with other compounds deposited from the growth
media.
In general, a CPE is used instead of capacitor to compensate for
the deviation from ideal behavior. The impedance of CPE is
defined by the following equation (Ref 29, 41-43):
ZCPE ¼ ðCPEÞ!1 ðjxÞ!a
ðEq 10Þ
When the carbon steel was exposed to the biotic system, the
EIS spectra varied significantly with exposure time as shown in
Fig. 12(a). The LF magnitude, represented by the semicircle
diameter, significantly decreased with time, indicating a
decrease in charge transfer resistance (Rct) and a subsequent
increase in the corrosion rate as supported by Fig. 9(b). The SRB
consortium impacts the corrosion rate by at least three mechanisms, via biofilm formation, reduction of sulfates and
production of hydrogen sulfide and subsequent formation of
iron sulfide (Ref 30, 31, 35). For the first 48 h, the medium
frequency (MF) response presented in the phase diagram in
Fig. 12(c) shows one time constant that indicates an activation
control process, which is represented by the circuit diagram for
the abiotic system shown in Fig. 11. This behavior is attributed
to the formation of an unstable conditioning layer based on a
mixture of inorganic / organic compounds; it is essentially the
time of biofilm formation (Ref 30, 31, 35). However, after
biofilm formation in conjunction with the development of iron
sulfide and siderite film and when a steady state is reached at 96
h, another time constant shows up in the phase angle spectra in
Fig. 12(c). Figure 13(a) shows the circuit model used to fit the
EIS data at 96 and 192 h, where the compact biofilm (bf) formed
constitutes an additional parallel combination of resistance and
capacitance. At 288 h, the compact biofilm layer and iron sulfide
layers start developing pores and the corresponding EIS data fit
the circuit model shown in Fig. 13(b). The enhancement of the
dissolution kinetics of the metallic surface is evidenced by the
decrease of the magnitude of charge transfer resistance (Rct) with
time as shown in Fig. 12(a) and 13(b). EIS spectra suggest that
the formation of an adherent biofilm along with a mixed layer of
iron sulfide and siderite established electrochemical cells on the
steel surface and subsequently enhanced the corrosion process.
EIS results draw general statements about how the corrosion
proceeds in the biotic system:
•
Before 96 h, the MF response showed one time constant
(Fig. 12c) that was attributed to the formation of an unstable layer of a mixture of corrosion products, mainly ferric
hydroxide and organic compounds. At this stage, the SRB
bacteria attached to the surface, assimilated lactate and
reduced sulfate to sulfide ions. Biogenic hydrogen sulfide
and a subsequent mixed layer of iron sulfide and siderite
along with EPS are formed by the precipitation of ferrous
ions with sulfide and carbonate ions.
Journal of Materials Engineering and Performance
3000
200
0h
96 h
288 hrs
366 hrs
150
288 h
-Z'' (Ω*cm2)
-Z" (Ω*cm2)
192 h
2000
1000
100
50
0
(a)
0
1000
2000
0
3000
0
Z' (Ω*cm )
100
150
200
2
Z'(Ω*cm )
-80
Phase Angle (degree)
50
(b)
2
0h
96 h
192 h
288 h
-60
-40
-20
0
(c)
Fig. 12
10-2
10-1
100
102
103
104
105
Frequency (Hz)
EIS date for biotic system; (a), (b) Nyquist plots, and (c) phase angle plots
Fig. 13 Circuit models used to fit the EIS data for the biotic system (a) at 96 and 192 h (b) at 288 and 366 h
•
101
At 96 h, the mixed layers of EPS and semiconductive corrosion products were stable as evidenced by the phase
angle spectra that reveal two time constants (Fig. 12c). At
this stage, the microenvironment changes induced by the
bacterial metabolic activities in the biofilm and the galvanic coupling between the iron sulfide with the underlying surface increased the corrosion rate significantly, as
shown in Fig. 9(b). The galvanic coupling attacks were
Journal of Materials Engineering and Performance
more pronounced with pearlite microstructures, as represented in Fig. 6, due to their structure and compositional
heterogeneity that induced electrochemical potential gradients.
• At the final stage, 288 h, with the proliferation of the
SRB, production of excess hydrogen sulfide and accumulation of excess corrosion products, the biofilm and iron
sulfide film decomposed, cracked and became loose and
porous due to the production of polysulfide products and
the induced intrinsic physical growth stresses (Ref 7, 32,
35, 44). Subsequently, the steel surface was exposed to
the aggressive medium again, which accelerated the corrosion rate significantly ('45 mpy), as shown in Fig. 9(b).
4. Conclusions
In this study, MIC of API 5L X52 carbon steel coupons was
investigated by exposure of the coupons to a SRB consortium
cultivated from produced water from a production sour oil well.
16S rRNA gene sequence analysis indicated that the mixed
bacterial culture consortium contained three phylotypes that are
close to members of the Proteobacteria (Desulfomicrobium
sp.), Firmicutes (Clostridium sp.) and Bacteroidetes (Anaerophaga sp.); all described previously as species associated with
MIC. In the presence of a SRB-biofilm, substantial levels of
sulfide were detected. XRD revealed the presence of different
phases, siderite (FeCO3), iron sulfide (FexSy), and iron oxide
(FeOOH) constituents in the corrosion products for the system
exposed to the SRB consortium. The microenvironment at the
metal-liquid layer became highly altered by the bacterial
biofilms via attachment of bacteria, deposition of organic byproducts, (e.g., EPS and the production of hydrogen sulfide)
and subsequent development of semiconductive iron sulfide.
The metabolic activities resulted in accumulation of a
substantial amount of iron sulfide that promoted galvanic
coupling with the underlying surface and resulted in general
corrosion. Thereafter, the corrosion rate increased dramatically.
The corrosion of the steel coupons was significantly more
severe in the biotic conditions compared to the abiotic control.
The corrosion rate was ' 45 mpy in the biotic system, while it
was '15 mpy for the abiotic system. The nature of the
corrosion was generalized with preferential etching of select
pearlite microstructures in a regular, rectangular, and repeating
manner. The galvanic coupling attacks were more marked with
pearlite microstructures due to high electrochemical potential
gradients. Moreover, the biofilm, as well as iron sulfide altered
the kinetic behavior of the system by inducing an extra time
constant in the circuit model.
Acknowledgments
The authors acknowledge and appreciate the Saudi Aramco and
Inspection Department Management for their continual support for
this project.
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