NACE Standard TM0106-2016 Reaffirmation 2016-03-20

Item No. 21248 Revised 2015-02-01

Approved 2006-06-23

Detection, Testing, and Evaluation of

Microbiologically Influenced Corrosion
(MIC) on External Surfaces of Buried
Pipelines

This NACE International standard represents a consensus of those individual members ABSTRACT
who have reviewed this document, its scope, and provisions. Its acceptance does not in This standard describes types of microor-
any respect preclude anyone, whether he or she has adopted the standard or not, from ganisms, mechanisms by which MIC oc-
manufacturing, marketing, purchasing, or using products, processes, or procedures not curs, methods of testing for the presence of
in conformance with this standard. Nothing contained in this NACE International stan- bacteria, research results, and interpreta-
dard is to be construed as granting any right, by implication or otherwise, to manufacture, tion of testing results for external surfaces
sell, or use in connection with any method, apparatus, or product covered by Letters of buried, ferrous-based metal pipelines
Patent, or as indemnifying or protecting anyone against liability for infringement of Let- and related components. Appendixes are
ters Patent. This standard represents minimum requirements and should in no way be included for media specifications (nonman-
interpreted as a restriction on the use of better procedures or materials. Neither is this datory Appendix A), dilution procedures
standard intended to apply in all cases relating to the subject. Unpredictable circum- (nonmandatory Appendix B), and site in-
stances may negate the usefulness of this standard in specific instances. NACE Interna- spection and testing (nonmandatory Ap-
tional assumes no responsibility for the interpretation or use of this standard by other pendix C). This standard is maintained Task
parties and accepts responsibility for only those official NACE International interpreta- Group 237.
tions issued by NACE International in accordance with its governing procedures and pol-
icies which preclude the issuance of interpretations by individual volunteers.

Users of this NACE International standard are responsible for reviewing appropriate KEYWORDS
health, safety, environmental, and regulatory documents and for determining their appli- MIC, microorganisms, sampling, MMM, bio-
cability in relation to this standard prior to its use. This NACE International standard may film, bacteria, Archaea
not necessarily address all potential health and safety problems or environmental haz-
ards associated with the use of materials, equipment, and/or operations detailed or re-
ferred to within this standard. Users of this NACE International standard are also respon-
sible for establishing appropriate health, safety, and environmental protection practices,
in consultation with appropriate regulatory authorities if necessary, to achieve compli-
ance with any existing applicable regulatory requirements prior to the use of this stan-
dard.

CAUTIONARY NOTICE: NACE International standards are subject to periodic review, and
may be revised or withdrawn at any time. NACE International requires that action be tak-
en to reaffirm, revise, or withdraw this standard no later than five years from the date of
initial publication. The user is cautioned to obtain the latest edition. Purchasers of NACE
International standards may receive current information on all standards and other NACE
International publications by contacting the NACE International Membership Services
Department, 15835 Park Ten Place, Houston, Texas 77084 (telephone +1 [281] 228-
6200).

ISBN 1-57590-206-0
©2016, NACE International
 Foreword
In NACE standards, the terms shall, must, Microbiologically influenced corrosion (MIC) is corrosion caused by the presence or ac-
should, and may are used in accordance tivity (or both) of microorganisms in biofilms on the surface of the corroding material.
with the definitions of these terms in the Many materials, including most metals and some nonmetals, can be degraded in this
NACE Publications Style Manual, 4th ed., manner. Microbiologically mediated reactions can alter both rates and types of electro-
Paragraph 7.4.1.9. Shall and must are used chemical reactions in a corrosion cell. These reactions influence pitting, crevice corro-
to state mandatory requirements. The term sion, differential aeration cells, concentration cells, dealloying, and galvanic corrosion.
should is used to state something consid- Therefore, MIC investigations require microbiological, chemical, and metallurgical test-
ered good and is recommended but is not ing for proper diagnosis. The conclusion that MIC has taken place should be based on
mandatory. The term may is used to state the preponderance of circumstantial evidence. Microorganisms are often resistant to
something considered optional. many control methods and can be a serious external corrosion threat to pipelines.

This NACE standard test method applies to the external surfaces of ferrous-based metal
pipeline facilities and describes types of microorganisms, mechanisms by which MIC
occurs, methods for sampling, and testing for the presence of microorganisms, research
results, and interpretation of testing results. Sections 1 through 4 of this standard dis-
cuss the technical aspects of MIC. Sections 5 through 7 discuss field equipment and
testing procedures.

This standard is intended for use by pipeline operators, pipeline service providers, gov-
ernment agencies, and any other persons or companies involved in planning or manag-
ing pipeline integrity. Portions of Sections 3 and 4 of this standard are excerpted from
Peabody’s Control of Pipeline Corrosion, Chapter 14—“Microbiologically Influenced
Corrosion,”1 and enclosed in quotation marks.

This standard was prepared by Task Group (TG) 237, “Microbiologically Influenced Corro-
sion on External Surfaces of Buried Pipelines: Detection, Testing, and Evaluation—Stan-
dard.” It was revised in 2015 by TG 237. TG 237 is administered by Specific Technology
Group (STG) 35, “Pipelines, Tanks, and Well Casings,” and is sponsored by STG 60, “Cor-
rosion Mechanisms.” This standard is issued by NACE International under the auspices
of STG 35.

2 TM0106-2016 NACE International

 NACE International Test Method (TM0106-2016)

Detection, Testing, and Evaluation of

Microbiologically Influenced Corrosion (MIC)
on External Surfaces of Buried Pipelines
1. General.....................................................................................................................4
2. Definitions.................................................................................................................4
3. Introduction...............................................................................................................8
4. External MIC of Pipelines........................................................................................12
5. Sampling Equipment...............................................................................................14
6. Sampling and Testing Procedures..........................................................................15
7. Testing Guidelines...................................................................................................17
8. Application of Test Methods to Pipelines and Interpretation of Data.......................25
References..............................................................................................................27
Bibliography............................................................................................................29
Appendix A (Nonmandatory) Site Inspection and Testing.......................................31

Figures
Figure 1: Examples of Various Pit Morphologies as Viewed in Cross Section.... 15
Figure 2: (Live, Inactive, and Dead) Typically Present in Samples from the Oil
Industry that are Enumerated Using Various MMMs as Compared to the MPN
(Culturing) Method..................................................................................................21
Table 1: Comparisons of MMMs........................................................................21

NACE International TM0106-2016 3

 Section 1: General
1.1 While the evaluation, monitoring, and mitigation of MIC cannot be prescribed in
one particular manner for any given pipeline, this standard describes methodolo-
gies by which the appropriate tools and techniques may be selected and practi-
cally applied to external surfaces of buried, ferrous-based metal pipelines and
related components. The methods presented in this standard represent the gen-
eral consensus of industry experts in pipeline corrosion and microbiology at the
time this standard was published.

1.2 Appendix A (Nonmandatory) provides a site inspection and testing checklist.

1.3 All applicable safety and environmental codes, rules, and regulations must be
followed when using this standard.

1.4 The term “pipeline” as used in this standard generally refers to any pipe or com-
ponent of a pipeline system for which the mechanism of external MIC is of interest
to the user of this standard.

Section 2: Definitions
Abiotic: The absence of living organisms, their biological components, or the metabolic
activities of living organisms.

Acid-producing bacteria (APB): Aerobic or anaerobic bacteria that produce organic ac-
ids as an end product of their metabolism. A few organisms (e.g., Thiobacillus) are also
capable of producing mineral acids (typically under aerobic conditions).

Aeration: (1) Exposing to the action of air. (2) Causing air to bubble through. (3) Introduc-
ing air into a solution by spraying, stirring, or similar method. (4) Supplying or infusing with
air, as in sand or soil. (5) The introduction of air into the pulp in a flotation cell to form air
bubbles.

Aerobic: Containing air or free molecular oxygen.

Aerobic microorganism (aerobe): A microorganism that uses oxygen as the final elec-
tron acceptor in metabolism.

Anaerobic microorganism (anaerobe) bacteria: A microorganism that does not require

oxygen for metabolism.

Archaea: Unicellular microorganisms that are genetically distinct from bacteria and eu-
karyotes, which often inhabit extreme environmental conditions. Archaea include halo-
philes (microorganisms that may inhabit extremely salty environments), methanogens (mi-
croorganisms that produce methane), and thermophiles (microorganisms that can thrive in
extremely hot environments). Archaeoglobus is a common Archaea.

Archaeoglobus: Microorganisms that grow at high temperatures between 60 and 95 °C

(140 and 203 °F), with optimal growth at 83 °C (181°F) (ssp. A. fulgidus VC-16).2 They are
sulfate-reducing archaea, coupling the reduction of sulfate to sulfide with the oxidation of
many different organic carbon sources, including complex polymers. Archaeoglobus spe-
cies have been isolated from oil reservoirs and production systems; however, this group of
microorganisms is normally not measured with current culturing techniques.

Autoclave: A pressurized, steam-heated vessel used for sterilization.

4 TM0106-2016 NACE International

Basal: The minimal level for, or essential for maintenance of vital activities of an organism,
such as basal metabolism.

Biofilm: Microbial growth at an interface in which individual cells are bound within a matrix
of extracellular polymeric materials.

Biotic: Involving the presence or metabolic activities of living organisms.

Carbohydrate: Any of the group of organic compounds composed of carbon, hydrogen,

and oxygen, including sugars, starches, and celluloses.

Cathodic protection (CP): A technique to reduce the corrosion rate of a metal surface by
making that surface the cathode of an electrochemical cell.

Coating: (1) A liquid, liquefiable, or mastic composition that, after application to a surface,
is converted into a solid protective, decorative, or functional adherent film. (2) (In a more
general sense) a thin layer of solid material on a surface that provides improved protective,
decorative, or functional properties.

Coating system: The complete number and types of coats applied to a substrate in a
predetermined order. (When used in a broader sense, surface preparation, pretreatments,
dry film thickness, and manner of application are included.)

Culture medium: A sterile solution or other substrate that is formulated to promote the
growth of a particular type or group of microorganisms. (Also called growth medium.)

4-,6-diamidino-2-phenylindole (DAPI): A stain for optical microscopy that targets the De-
oxyribonucleic acid (DNA) in all (i.e., living and inactive) microbial cells.

Denaturing gradient gel electrophoresis (DGGE): A molecular microbiological method

used to profile the most abundant microbial groups in a sample.

Dielectric coating: A coating that does not conduct electricity. For the purposes of this
standard, it also inhibits the passage of an electric current relative to the coating’s dielectric
strength.

Disbondment: The loss of adhesion between a coating and the substrate.

Dissimilatory: Metabolic reactions in which a reductant is used as an electron acceptor

and not incorporated into the cell (e.g., dissimilatory sulfate or nitrate reduction); metabolic
changes that convert complex molecules into simple ones.

Eukaryotes: Cells having a true nucleus, bound by a double membrane. Prokaryotic cells
have no nucleus.

Facultative: Capable of growing either with or without the presence of a specific environ-
mental factor, e.g., oxygen.

Fluorescence in situ hybridization (FISH): A molecular microbiological method used for

enumeration of microorganisms. The method is based on gene probes targeting ribosomal
Ribonucleic acid (RNA) (16S or 23S rRNA) in microbial cells. Only living and active cells
contain sufficient ribosomes that can be detected by FISH. Gene probes consist of two
parts: (1) an artificial DNA strand complementary to the ribosomal RNA in the target cell;
and (2) a fluorescing molecule covalently attached to the probe that enables observation
of the target microorganism in the microscope.

Fungi: Nucleated, usually filamentous, spore-bearing parasitic organisms devoid of chlo-

rophyll, including molds, mildews, smuts, mushrooms, yeast, and others.

NACE International TM0106-2016 5

 Growth: Increase in the quantity of metabolically active protoplasm, accompanied by an
increase in cell number, cell size, or both.

Growth medium: See Culture medium.

Heterotrophic: An organism that obtains nourishment from the ingestion and breakdown
of organic matter.

Holiday: A discontinuity in a protective coating that exposes unprotected surface to the

environment.

Inoculum: A small quantity of microorganisms used to start a new culture.

Inorganic acid: A compound composed of hydrogen and a nonmetal element or radical.

They can range from acids of great strength (e.g., hydrochloric acid [HCl] and sulfuric acid
[H2SO4] to those that are very weak (e.g., boric acid). A substance that yields hydrogen
ions (which can act as a proton donor) and the conjugate base ions when dissolved in
water. (Also called a mineral acid.)

Isotonic: Having uniform tension of a solution; having the same osmotic pressure as the
fluid phase of a cell or tissue.

Metal-oxidizing bacteria: Bacteria (most notably iron and manganese oxidizing bacteria)
that derive their energy from oxidizing one oxidation state of a metal to another.

Metal-reducing bacteria (MRB): Bacteria that when in direct contact with solid iron (Fe+3)
and manganese (Mn+4) oxides them and produces soluble ions (Fe+2 and Mn+2), resulting
in dissolution of surface oxides and localized corrosion.1

Methanogens: Microorganisms that produce methane as a metabolic byproduct in anoxic

(i.e., oxygen-free) conditions. They are classified as Archaea, a group quite distinct from
bacteria. Some are extremophiles and found in environments such as oil field systems, hot
springs, and submarine hydrothermal vents, as well as in the “solid” rock of the Earth’s
crust, kilometers below the surface. Methanogens are common Archaea in oil production
systems; however, they are normally not measured with current culturing techniques.
Methanogens are involved in MIC by consuming hydrogen at the metal surface and there-
by creating a depolarization.

Microaerophilic: Pertaining to those microorganisms requiring free oxygen, but in very

low concentration for optimum growth.

Microbiologically influenced corrosion (MIC): Corrosion affected by the presence or

activity, or both, of microorganisms. (Note: The microorganisms that are responsible for
MIC are typically found in biofilms on the surface of the corroding material. Many materials,
including most metals and some nonmetals, can be degraded in this manner.)

Microbe: See microorganism.

Microorganism: An organism of microscopic or ultramicroscopic size. Bacteria, Archaea,

and fungi are microorganisms. Bacteria and Archaea are combined and called prokary-
otes. Fungi belong to eukaryotes (Eukarya).

Mineral acid: See inorganic acid.

Monosaccharide: A carbohydrate that cannot be hydrolyzed to a simpler carbohydrate.

Morphology: A branch of biology that deals with structure and form of an organism at any
stage of its life history.

6 TM0106-2016 NACE International

Most probable number method (MPN): A technique that does not rely on quantitative
assessment of individual cells; instead, it relies on specific qualitative attributes of the mi-
croorganism being counted. The important aspect of MPN methodology is the ability to
estimate a microbial population size based on a process-related attribute. The MPN tech-
nique estimates microbial population sizes in a liquid substrate. The methodology for the
MPN technique is dilution and incubation of replicated cultures across several serial dilu-
tion steps.

Motile: Exhibiting or capable of movement.

Organic acid: Weak acid that contains carbon (correctly classified as carboxylic acids
because they contain a carboxyl group, -COOH). Organic acids (e.g., formic, acetic, lactic)
are the end product of metabolism by a variety of microorganisms. (Also called short-chain
fatty acids.)

Organism: A complex structure of interdependent and subordinate elements whose rela-

tions and properties are largely determined by their function as a whole. (Also see micro-
organism.)

Oxidation-reduction potential: The potential of a reversible oxidation-reduction reaction

in a given electrolyte reported on the standard hydrogen electrode scale. (also called redox
potential)

Permeation: The migration of water from the soil through the coating to the pipe surface
by diffusion.

Phosphate buffer: Solution made of dibasic potassium phosphate (K2HPO4) and sodium
phosphate (Na2HPO4).

Pipe-to-soil potential: See structure-to-electrolyte potential.

Polymerase chain reaction (PCR): A molecular technique which allows the production of
large quantities of a specific DNA from a DNA template using a simple enzymatic reaction
without a living organism. A quantitative version of PCR is called quantitative polymerase
chain reaction (qPCR).

Polysaccharide: A carbohydrate composed of many monosaccharides.

Prokaryotes: The prokaryotes are divided into two domains: the bacteria and the Archaea.
Archaea were originally thought to live only in inhospitable conditions such as extremes of
temperature, pH, and radiation, but have been found in all types of habitats. Sulfate-reduc-
ing prokaryotes (SRP) consist of both sulfate-reducing bacteria and sulfate-reducing Ar-
chaea.

Quantitative polymerase chain reaction (qPCR): A molecular microbiological method used

to quantify the total number of microorganisms or a specific genus/species of microorganisms
in nearly any type of sample. qPCR can be used for both fluid and solid samples, as well as
microorganisms collected via membrane filtration. This method does not underestimate micro-
organisms that do not grow in culture. This method uses synthetic DNA (called primers) tagged
with a fluorescent molecule or synthetic DNA mixed with a DNA intercalating agent (dye) to
quantify microorganisms using a modified version of polymerase chain reaction (PCR).

Redox potential (Eh): See oxidation-reduction potential.

Ringers solution: An aqueous solution of chlorides that is isotonic to animal tissues.

Sterile: (1) Free of any living microorganisms. (2) Not introducing microorganisms that are
foreign to the host body or subject under study.

NACE International TM0106-2016 7

 Structure-to-electrolyte potential: The potential difference between the surface of a bur-
ied or submerged metallic structure and the electrolyte that is measured with reference to
an electrode in contact with the electrolyte.

Substratum: A solid surface; often refers to a surface colonized by microorganisms.

Sulfate-reducing Archaea (SRA): A group of anaerobic Archaea that perform dissimilato-

ry reduction of sulfate, resulting in sulfide formation. They are most likely to grow at reser-
voir conditions (60 to 95 °C [140 to 203 °F]).

Sulfate-reducing bacteria (SRB): A group of anaerobic bacteria that perform dissimilato-

ry reduction of sulfate, resulting in sulfide formation. They grow at a broad range of tem-
peratures.

Sulfate-reducing prokaryotes (SRP): A group of microorganisms that consists of both

sulfate-reducing bacteria (SRB) and sulfate-reducing Archaea (SRA).

Tenting: A tent-shaped void formed along the girth weld or longitudinal seam-weld rein-
forcement in a pipe when the external coating is not in continuous intimate contact with the
pipe and weld surfaces.

Water leaching: The removal of soluble constituents from a coating by water.

Section 3: Introduction
3.1 MIC is corrosion affected by the presence or activity (or both) of microorganisms in
biofilms on the surface of the corroding material. Many materials, including most
metals and some nonmetals, can be degraded in this manner. MIC can result from
the activities of microorganisms, including bacteria, Archaea, and fungi1 in biofilms or
in the local environment directly in contact with the corroding material.2 This standard
is primarily focused on the effects of bacteria and Archaea. In one survey, Jack et al.3
reported that MIC was responsible for 27% of the corrosion deposits on the exterior
of line pipe. Pope and Morris4 reported that almost all cases of MIC on external sur-
faces of coated pipes were associated with disbonded or damaged coatings. The
following general statements are commonly accepted regarding microorganisms.1

3.1.1 Individual microorganisms are usually small in size (they typically are
from 0.2 to 10 μm (8 to 400 µin) in length by up to 2 or 3 µm [80 to 120 µin] in
width)—a quality that allows them to penetrate crevices and other areas. Colo-
nies of microorganisms can grow to macroscopic proportions.1

3.1.2 Microorganisms may be motile, capable of migrating to more favorable

conditions or away from less favorable conditions (e.g., toward food sources or
away from toxic materials).

3.1.4 Certain microorganisms can withstand a wide range of temperatures (at

least -10 to 99 °C [14 to 210 °F]), pH levels, and oxygen concentrations.1

3.1.5 Microorganisms grow in colonies and form biofilms, making survival

more likely under adverse conditions.

3.1.6 Under favorable conditions, microorganisms can reproduce very quickly

(generation times of 18 min have been reported).1

3.1.7 Individual microbial cells can be widely and quickly dispersed by water
or other modes, thus the potential for some cells in the population to reach more
favorable environments is good.

8 TM0106-2016 NACE International

 3.1.8 Many microorganisms can quickly adapt to use a wide variety of different
nutrient sources. For example, Pseudomonas fluorescens can use more than 100
different compounds as sole sources of carbon and energy, including sugars,
lipids, alcohols, phenols, and organic acids.1

3.1.9 Many microorganisms form extracellular polysaccharide materials (cap-

sules or slime layers). The resulting slimes are sticky and trap organisms and
debris (food), resist penetration of toxicants, (e.g., biocides or corrosion inhibi-
tors), and hold the cells between the source of the nutrients (the bulk fluid) and
the surface.1

3.1.10 Many bacteria and fungi produce spores that are resistant to tempera-
ture, acids, alcohols, disinfectants, drying, freezing, and other adverse conditions.
Spores may remain viable for hundreds of years and germinate on finding favor-
able conditions. In the natural environment, there is a difference between survival
and growth. Microorganisms can withstand long periods of starvation and desic-
cation. If conditions are alternating wet and dry, microbes may survive dry periods
and grow only during the wet periods.

3.1.11 Microorganisms may be resistant to many chemicals (antibiotics, disin-

fectants, and others) by virtue of their ability to degrade these chemicals or by
being impenetrable (because of slime, cell wall, or cell membrane characteris-
tics). Resistance may be easily acquired by mutation or acquisition of a plasmid
(essentially by naturally occurring genetic exchange between cells, i.e., genetic
engineering in the wild).

3.2 Mechanisms for MIC

MIC typically takes place in the presence of microbial consortia that are com-
prised of more than one physiological type of microorganism. Depending on the
environment, these microbes may include metal-oxidizing bacteria, sulfate-reduc-
ing prokaryotes (SRP), acid-producing bacteria (APB), metal-reducing bacteria
(MRB), and methanogens that interact in complex ways within the structure of
biofilms.5,6 MIC does not produce a unique form or morphology of corrosion. In-
stead, MIC can result in pitting, crevice corrosion, underdeposit corrosion (UDC),
and dealloying, in addition to galvanic corrosion.

Simply by their presence on a metal surface, microorganisms may set up the proper
conditions for pitting or crevice corrosion. Once localized corrosion has been initiated,
microbial reactions can maintain suitable conditions (e.g., low oxygen concentration)
for continued pit/crevice growth. Under anaerobic reducing conditions, MIC may be
observed when there is some mechanism for the removal or transformation of corro-
sion products (e.g., a transition from stagnation to flow) or the introduction of oxygen
to a previously anaerobic environment. The following discussion about individual MIC
mechanisms is directly related to carbon steel.1

3.2.1 Sulfate Reduction

Sulfate-reducing bacteria (SRB) are a diverse group of anaerobic microorgan-

isms that can be isolated from a variety of subsurface environments. If the aerobic
respiration rate within a biofilm is greater than the oxygen diffusion rate during
biofilm formation, the metal/biofilm interface can become anaerobic and provide
a niche for sulfide production by SRB. The critical thickness of the biofilm required
to produce anaerobic conditions depends on the availability of oxygen and the
rate of respiration. SRB concentrations may be proportional to sulfate concentra-
tions. The distribution of favorable pH ranges from 6 to 12, although SRB can
adapt to other less optimum conditions. SRB grow in soil, fresh water, or salt
water under anaerobic conditions.

NACE International TM0106-2016 9

 Many species of SRB have been identified, differing in morphology and in the
organic substances that they can metabolize. They have in common the ability to
oxidize certain organic substances to organic acids or carbon dioxide by reduc-
tion of inorganic sulfate to sulfide. In the absence of oxygen, the metabolic activ-
ity of SRB causes accumulation of hydrogen sulfide near metal surfaces. This is
particularly evident when metal surfaces are covered with biofilms. The concen-
tration of sulfide is highest near the metal surface.

Iron sulfide forms quickly on carbon steels and covers the surface if both ferrous
and sulfide ions are available. Formation of iron sulfide minerals stimulates the
cathodic reaction. Once electrical contact is established, a galvanic couple devel-
ops with the mild steel surface as an anode, and electron transfer occurs through
the iron sulfide. Under conditions such as low ferrous ion concentrations, adher-
ent and temporarily protective films of iron sulfides are formed on the steel sur-
face, with a consequent reduction in corrosion rate.7,8

Although SRB are anaerobic in their metabolism, studies by Hardy and Brown
demonstrated that the availability of oxygen can increase corrosion in the pres-
ence of SRB.9 They found that the corrosion rates of mild steel in anaerobic cul-
tures of SRB were low (6.74 μm/y [0.265 mpy]), while subsequent exposure to air
caused higher corrosion rates (610 μm/y [24 mpy]).

The most prevalent mechanism for the observed corrosion in a study reported by
Jack et al.10 was the formation of a galvanic couple between steel and microbio-
logically produced iron sulfides. The couple is normally short-lived because the
iron sulfide matrix becomes saturated with electrons derived from the corrosion
process. In the presence of SRB, however, the corrosion process is perpetuated
because SRB removes electrons (in the corrosion process) from the iron sulfide
surface. This process may involve the formation of cathodic hydrogen on the iron
sulfide or the direct transfer of electrons from the iron sulfide matrix to redox pro-
teins in the bacterial cell wall. Corrosion rates associated with this mechanism
were proportional to the amount of iron sulfide in the corrosion cell.

Sulfate-reducing Archaea (SRA) are like SRB, obtaining their energy by oxidizing
organic compounds or molecular hydrogen (H2) while reducing sulfates to sul-
fides, especially to hydrogen sulfide.11 SRA consist of the genera Archaeoglobus.
Archaeoglobus grow at temperatures in the range of 60 to 95 °C (140 to 203 °F),
with optimal growth at 83 °C (181 °F) (ssp. A. fulgidus VC-16).2 Previously, Ar-
chaeoglobus species have been isolated from oil reservoirs and oil production
systems; however, this group of microorganisms is normally not measured with
current culturing techniques. SRA are known to cause the corrosion of iron and
steel in oil and gas processing systems by producing iron sulfide.

The formation of H2S by SRA activity can have profound effects in terms of reser-
voir souring, health/safety/environmental threats, and materials degradation.

3.2.2 Acid Production

Organic acids are produced by both bacteria and fungi. This process is anaerobic for
some microorganisms or aerobic for other microorganisms and fungi. Most final prod-
ucts of APB are short-chained fatty acids (e.g., acetic, formic, and lactic acids). The
role of APB in MIC is controversial. Pope et al.12 proposed that APB produce biogen-
ic organic acids that are directly responsible for corrosion in the absence of SRB.
Jack et al.13 reported that the main role of APB is to provide the environment and
nutrients for SRB growth. Other bacterial species can produce aggressive inorganic
acids, such as sulfuric acid (H2SO4), in aerobic environments. Microorganisms in the
soil as well as other environments can locally generate high concentrations of carbon
dioxide. The carbon dioxide dissolves in the water, producing carbonic acid. Carbon-

10 TM0106-2016 NACE International

 ic acid solution is corrosive to pipeline steels and can lead to general attack, pitting
attack, and stress corrosion cracking.1

3.2.3 Metal Deposition

3.2.3.1 Microorganisms can also affect corrosion by creating differen-

tial aeration cells on the surface of the metal and fixing the location of
anodic sites beneath colonies of microorganisms. The organisms most
often cited as causing differential aeration cells are those organisms
capable of depositing iron and manganese oxides.

3.2.3.2 Iron-oxidizing bacteria produce orange-red tubercles of iron ox-

ides and hydroxides by oxidizing ferrous ions from the bulk medium or
the substratum. Iron-depositing bacteria are microaerophilic and may
require synergistic associations with other bacteria to maintain low oxy-
gen conditions in their immediate environment. Deposits of cells and
metal ions create oxygen concentration cells that effectively exclude ox-
ygen from the area immediately under the deposit and initiate a series of
events that individually or collectively are very corrosive.

In an oxygenated environment, the area immediately under individual

deposits becomes deprived of oxygen. That area becomes a relatively
small anode compared to the large surrounding oxygenated cathode.
Cathodic reduction of oxygen can result in an increase in pH of the solu-
tion in the vicinity of the metal. The metal forms metal cations at anodic
sites. If the metal hydroxide is the thermodynamically stable phase in
the solution, the metal ions are hydrolyzed by water, forming hydrogen
(H+) ions. If cathodic and anodic sites are separated from one another,
the pH at the anode decreases and that at the cathode increases.

The pH at the anode depends on specific hydrolysis reactions. In addi-

tion, chloride (Cl-) ions from the electrolyte migrate to the anode to neu-
tralize any buildup of charge, forming heavy metal chlorides that are
extremely corrosive. Under these circumstances, pitting involves the
conventional features of differential aeration, a large cathode-to-anode
surface area, and the development of acidity and metallic chlorides. Pit
initiation depends on mineral deposition by microorganisms.

3.2.3.3 Manganese oxidation and deposition is coupled to cell growth

and metabolism of organic carbon. The reduced form of manganese
(Mn+2) is soluble and the oxidized forms (Mn2O3, MnOOH, Mn3O4, and
MnO2) are insoluble. As a result of microbial action, manganese oxide
deposits are formed on buried or submerged materials including metal,
stone, glass, and plastic, and can occur in natural waters that have man-
ganese concentrations as low as 10 to 20 ppb. For mild steel corrosion
under anodic control, manganese oxides can elevate corrosion current.
The current can be significant for biomineralized oxides that provide
large mineral surface areas. Given sufficient conductivity, manganese
oxide can sometimes serve as a cathode to support corrosion at an ox-
ygen-depleted anode within the deposit.

3.2.4 Metal Reduction

Dissimilatory iron and/or manganese reduction occurs in several microorganisms,

including anaerobic and facultative aerobic bacteria. Inhibitor and competition ex-
periments suggest that iron (Fe+3) and manganese (Mn+4) are efficient electron
acceptors that are similar to nitrate in redox ability and are capable of out-compet-
ing electron acceptors of lower potential, such as sulfate or carbon dioxide.14

NACE International TM0106-2016 11

 MRB in direct contact with solid Fe+3 and Mn+4 oxides produce soluble ions (Fe+2
and Mn+2). The result is dissolution of surface oxides and localized corrosion.1

3.2.5 Methanogens

Methanogens produce methane as a metabolic by-product in anoxic conditions.

They are classified as Archaea, a group quite distinct from bacteria.

Methanogens typically thrive in environments in which all electron acceptors oth-

er than CO2 (such as oxygen, nitrate, sulfate, and trivalent iron) have been deplet-
ed. They are common in wetlands, where they are responsible for marsh gas, and
in the guts of animals such as ruminants and humans. Others are extremophiles,
found in environments such as oilfield systems, hot springs, and submarine hy-
drothermal vents, as well as in the “solid” rock kilometers below the surface of the
Earth’s crust. Methanogens are common Archaea in oil production systems; how-
ever, they are normally not measured with current culturing techniques.15,16

Methanogens are known to promote MIC in steel and other metal structures by
consuming hydrogen formed at the corrosion cathode.17

Section 4: External MIC of Pipelines

4.1 Environment

The potential for MIC of buried pipelines is controlled by the availability of nutri-
ents, water, and electron acceptors. Peabody reported data from Harris18 that in-
dicated soil moisture content and bacterial cell counts were greater in backfill
material than in undisturbed earth adjacent to a pipeline. Trench backfill is not as
consolidated and allows greater penetration of moisture and increased oxygen
diffusion. Anaerobic bacteria thrive in waterlogged, dense soil. Alternating mois-
ture and oxygen concentrations influence the growth of bacterial populations. De-
spite the numerous mechanisms that one would predict for MIC of buried pipe-
lines, most failures have been attributed to the presence and activities of SRB
and APB. In general, sandy soils favor APB; high-clay soils support populations of
both kinds of organisms. To protect against all forms of external corrosion and
cracking, several coating materials have been used, including coal tar deriva-
tives, asphalts, polyolefin tapes, and fusion-bonded epoxies (FBE). Line pipe has
been further protected by CP. Although these are sound measures, MIC can oc-
cur in the presence of these preventative measures, if one or both break down or
are not properly maintained.1

4.2 Coatings

4.2.1 Differing environmental conditions (e.g., soil moisture, microflora, nutri-

ents) in both field surveys and laboratory experiments make it difficult to interpret
coatings performance and draw comparisons. Comparisons between field sur-
veys and laboratory experiments must not exceed stated condition limitations.
Tenting of coatings along irregularities on the pipe surface, especially at long
seam or girth welds, can create gaps between the coating and the pipe surface
that fill with groundwater and introduce microorganisms that may create corrosion
cells under the disbonded coating. Tape coatings are particularly susceptible to
tenting while liquid- or powder-applied coatings are more resistant to this type of
failure mechanism. Tenting has been most prevalent in wet high-clay soils on
unstable, geologically active slopes that are subject to high service temperatures.
The high service temperatures promote coating disbondment. Not all coating ma-
terials are affected by soil bacteria under all conditions. Coatings derived from
both coal (tars) and petroleum (asphalts) pass some exposure tests and fail oth-

12 TM0106-2016 NACE International

 ers. Materials that, by themselves, show resistance to attack by microorganisms
fail when combined or reinforced with other materials.1

4.2.2 Jack et al.10 demonstrated that certain coatings disbond more readily
after being exposed to soils containing SRB and APB. Polyethylene (PE) coating
damage proceeded linearly with time. PE tape coatings supported higher bacteri-
al counts than extruded PE or FBE, presumably because of the presence of bio-
degradable adhesive/primer components in the coating system. Susceptibility to
disbonding was high with FBE, higher with extruded PE, and highest with PE
tape. Two types of coating damage were reported: 1) damage caused by water
leaching and 2) damage caused by permeation. Both mechanisms affect intact
coatings and coatings around holidays. At existing holidays, damaged FBE coat-
ings experience an increased susceptibility to coating disbondment.

4.2.3 Peabody reported that coal tars, coal-tar epoxies, and coal-tar enamels
were immune to disbonding because of activities of microorganisms, which would
likely lessen the chances of corrosion, including MIC. Early coatings based on
asphalt were subject to oxidation and loss of low-molecular-weight components
through biodegradation and biodeterioration, resulting in a permeable, embrittled
coating.3 Pendrys19 demonstrated that, with time, asphalt could be degraded by
microorganisms selected from soil. Harris18 demonstrated that bacteria common-
ly found in pipeline soils can degrade asphalt, tape adhesives, kraft paper (ex-
pendable once the pipeline is in place), as well as the binders and fillers used in
felt pipeline wrappers. The next-generation coatings were based on polyvinyl
chloride (PVC) or PE. The PVC tape was unstable in service. Plasticizers consti-
tute up to 50 wt% of a PVC product and can be effectively lost through biodeteri-
oration and water dissolution. PE coatings rely on adhesives to attach the poly-
olefin layer to the primed steel surface.1

4.3 Cathodic Protection

For CP levels more negative than –850 mV (–880 mV to –1,000 mV) polarized vs.
a saturated copper/copper sulfate (Cu/CuSO4) reference electrode, it has been
demonstrated that bacteria levels, including SRB, can increase in saturated soils,
seawater, and marine sediments1,20-22 so that if the CP is intermittent, MIC can
occur at a higher rate than if CP were not previously present.23 MIC has at least
three effects on CP of pipelines.

4.3.1 When microbial activity is present and the coating is compromised, the
potential level required to mitigate corrosion is a more negative value. Pope and
Morris4 found that pipeline failures were often in contact with wet clays with little
scaling potential, creating a situation in which the demand for CP continues at a
high level over long periods of time and in which CP may not be distributed equal-
ly over the surface of coating holidays and surrounding disbondments. Microor-
ganisms colonize and initiate corrosion at such sites. Research by Barlo and
Berry24 determined that the current criteria specified in NACE SP016925 for CP of
buried pipelines (0.85 V versus Cu/CuSO4) were generally valid in concept. How-
ever, the critical values for the criteria varied with the environment. Elevated tem-
perature (60 °C [140 °F]), mill scale, and anaerobic bacteria affect CP require-
ments—100 to 200 mV more cathodic (negative) change in protection potential is
required compared to the condition absent those factors.1

4.3.2 MIC can increase the kinetics of corrosion reactions, increasing the CP
current necessary to achieve a given level of polarization.

4.3.3 Microorganisms can attack certain pipeline coatings, increasing exposed

metal surface area and further increasing the CP current used to achieve a given
level of polarization. Water intrusion at breaks in the coating may cause corrosion

NACE International TM0106-2016 13

 where the remaining dielectric coating may block CP. It should be noted, however,
that when sufficient CP levels exist, corrosion is mitigated even in the presence of
increased levels of bacteria. The difficulty is determining what is adequate for provid-
ing protection in each situation without adversely affecting the coating.1

Section 5: Sampling Equipment

5.1 Certain procedures may be used to collect samples.

5.1.1 Typical equipment and supplies should include some or all of the following:

(a) Sterile plastic or glass containers (10 to 125 mL [0.33 to 4.20 oz]).
(b) Sterile plastic sample collection bags.
(c) Sterile metal scalpels.
(d) Sterile cotton or polyester-fiberfill swabs.
(e) Sterile wooden spatulas (tongue depressors).
(f) Sterile 1 to 5 mL (0.03 to 0.1 oz) syringes.
(g) Sterile disposable plastic pipettes.
(h) Sterile latex gloves.
(i) Ice chest with refrigerant.
(j) Digital or film camera.
(k) Magnifying lens (5 to 60x).
(l) Marking pens (for wet surfaces).
(m) Nylon bristle brushes.
(n) Mechanical pit depth gauge.
(o) Labels.
(p) pH paper or meter.
(q) Culture media.
(r) Ultrasonic thickness (UT) meter to measure metal thickness.
(s) Sterile vials of phosphate-buffer solution.
(t) Formaldehyde (36.5%) for fixation on site (for use with DAPI method) or
2% glutaraldehyde as a general fixative.

5.2 Sterile containers must not contain any chemical that inhibits microbial survival. Col-
lection containers may contain sterile phosphate buffer, Ringers solution, or other
holding medium for suspension of solid samples. If samples are to be analyzed for
chemical composition, they must be maintained separately from samples transferred
to a holding medium. Most sampling containers are glass or plastic.

5.2.1 Glass containers should have screw-down caps and must have been
sterilized using a combination of pressure and temperature over time (100 kPa
[14.5 psig] steam at approximately 121 °C [250 °F] for a minimum of 20 min).
Glass containers may be reused many times after cleaning and sterilization.

5.2.2 Plastic containers are usually disposable; however, some can be

cleaned and autoclaved. In most cases, plastic sampling containers have rigid
walls, but polyethylene flexible-walled containers with closure ties can be used.

5.2.3 Sterile plastic bags provide a lightweight sample container. Plastic bags de-
signed for domestic use are not sterile and must not be used for sample collection.

5.2.4 Sterile, individually wrapped supplies can be purchased from most phar-
macies and all scientific supply stores. Sterile bags and tubes are usually ordered
in bulk, and the interiors remain sterile until opened.

14 TM0106-2016 NACE International

Section 6: Sampling and Testing Procedures
6.1 Site Inspection

6.1.1 Before excavation of the pipeline, the topography and soil type of the
area surrounding the dig should be noted. The site should be photographed both
close up and from a distance. The photos should complement one another so that
they can be aligned and compared. A ruler should be included in photographs to
indicate relative dimensions.

6.1.2 The following measurements provide useful information:

6.1.2.1 Soil resistivity using ASTM(1) G5726 Wenner 4 point or similar

method.

6.1.2.2 Pipe-to-soil potential.

6.1.2.3 Redox potential (Eh) as the potential of a platinum electrode

using a saturated copper copper-sulfate reference electrode.

6.1.2.4 Soil pH to compare with under-coating pH measurements to

determine effectiveness of CP.

6.1.3 Exposure of the pipe should be slow and careful to avoid damage to the
pipe and areas of sampling. Soil should be removed from the pipe surface care-
fully to expose the coating surface. In most cases, products adhering to the coat- Elliptical
ing surface shall not be removed until they are at a laboratory or other suitable
location where analyses can be performed. Color photographs of corroded areas
and their relationship to the pipe surface should be taken.

Shallow, Parabolic
6.1.4 The following should be noted and recorded (see checklist in Appendix A):

6.1.4.1 Coating type (e.g., coal tar, asphalt, bitumen, tape, FBE), type
of damage (e.g., disbonding, holidays, blistering, seam tenting, crack-
ing, wrinkling, or none), extent of damage (percentage of exposed area), Deep, Narrow
and location (circumferential and longitudinal position on the pipe in re-
lation to weld seams and coating seam, if present).

6.1.4.2 Characteristics of soil: moist, dry, or wet; silt, sand, gravel, rock, or
clay; odor; discoloration associated with a buried object; local chemical Grain Attack, Vertical
composition, e.g., presence of salts, cations, anions, such as chlorides,
sulfates, carbonates, etc.

6.1.4.3 Relationship of corroded area within overall system: depth,

Undercut
light, engineering design (welds, seams), etc.

6.2 Description of Corrosion Products

6.2.1 Corrosion products: color (brown, black, white, or gray), shape (deposit, Sub-Surface
nodule, or films), texture, and odor.

6.2.2 The form of any visible corrosion: shapes, sizes, and depths of pits;
crevice corrosion; UDC should be noted (See Figure 1).
Grain Attack, Horizontal
6.2.3 Any visible biological accumulations in corroded areas: form, color, tex-
ture, or odor (e.g., none, earth, rotten eggs, etc.). FIGURE 1: Examples of Various Pit Mor-
phologies as Viewed in Cross Section.27
(1)
ASTM International (ASTM), 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428.
(2)
European Federation of Corrosion (EFC), 1 Carlton House Terrace, London, SW 1Y 5DB, UK.

NACE International TM0106-2016 15

 6.2.4 Chemical testing for sulfides and carbonates.

6.3 Coating Inspection

6.3.1 Once the pipe is exposed, the soil, water, and deposits must be sampled
immediately and stored or tested. The coating surrounding the suspected area of
corrosion should be carefully removed using a knife or similar instrument. Sample
contamination of corrosion with soil or water must be kept to a minimum. Contact
with the soil, corrosion product, or film with hands or tools other than sterile imple-
ments must be avoided.

6.3.2 Coating inspection for MIC testing purposes must precede any other
coating evaluation. Likely areas of active corrosion or MIC should be identified
using ILI tools or visual inspection. Damaged coatings should be carefully re-
moved using a sterile scalpel to expose the steel.

6.4 Sample Collection

6.4.1 Whenever possible, a clean working surface should be used, because if

proper precautions are not taken, dust or dirt on the working surface or general
area can potentially contaminate the inside of sample containers when a sterile
sample bottle is uncapped. Ideally, the containers should be handled by gripping
the lower part of the bottle, and skin contact with the upper part of the container
must be avoided.

6.4.2 If a small volume of liquid is present under the coating, a sample should
be taken using a sterile syringe or polyester-fiberfill swab. Both the liquid in the
syringe and on the swab may be used for the enumeration of microorganisms as
described later. The swab must be stored in a sterile plastic tube until tested.

6.4.3 The pH of any liquid found under the coating should be tested using pH
(1 to 14) paper or a meter with a microsensor electrode. The coating should be
carefully sliced to a length to allow the test paper to be slipped behind the coating.
The coating should be pressed against the pH paper for a few seconds and then
lifted. The pH paper may then be removed. The color of the paper in relation to
the chart provided with the paper should be noted and recorded. A syringe or pi-
pette should be used to extract a small amount of liquid to measure pH by putting
a drop or two of liquid on the microsensor. If possible, the pH of any available
groundwater (typically at pipe level) away from the pipe should be determined for
reference.

6.4.4 If surface deposits or corrosion products are fragile, they should be

scraped from the pipe. Corrosion products must be collected by scraping the area
with a sterile scalpel, or swabbing with a sterile polyester-fiberfill swab.

6.4.5 Multiple samples, when present, are typically taken from one or more of
the following locations:

6.4.5.1 Undisturbed soil immediately next to the exposed pipe-steel

surface or at an area of coating damage.

6.4.5.2 Deposits associated with visual evidence of pipe corrosion, es-

pecially those pit contents underneath nodules or tubercules that have
been removed and are more likely to be uncontaminated with soil or
groundwater. (Note: it should be identified when corrosion samples are
known or suspected to be contaminated from outside sources).

6.4.5.3 Scale or biofilm on the steel surface or the backside of the coating.

16 TM0106-2016 NACE International

 6.4.5.4 Liquid trapped behind the coating.

6.4.5.5 Fresh, undisturbed soil at pipe depth at least 1 m (3 ft) trans-

verse to the pipe. This location, such as the ditch wall, acts as a refer-
ence from which to determine whether the microbial population near the
pipe is elevated.

6.4.5.6 Additional samples should be taken from other locations where

bacterial activity is suspected and tested.

6.4.5.7 Solid and liquid samples must be placed in clean, sterile,

sealed, and carefully labeled containers. The containers should be filled
to no more than 60% maximum volume. The bag or tube must be sealed
tightly, and the whole bagged sample sealed again within a second plas-
tic bag to reduce the risk of leakage.

6.4.5.8 Sample containers should be labeled with the location of the

dig site, sample origin, date, sampling time, and the tests that have been
performed on the sample. Extraneous dust, dirt, and debris shall not be
introduced into the sample at the time of collection. The interior surfaces
of sample containers or any other parts likely to be in direct contact with
the sample must not have been directly handled.

6.5 Sample Transport

6.5.1 Changes in detectable numbers and types of microorganisms can occur

rapidly after removal of samples from the environment. One should arrive at the
corrosion site with all solutions, media, and other necessary materials. If that is
not possible, liquids and solids can also be transported to a laboratory or other
facility for processing and testing. The major concern in transporting samples for
microbiological evaluation is to ensure that microorganisms remain alive and ac-
tive without multiplication. Sample collection may expose microorganisms to
abrupt changes in pressure, temperature, atmosphere, and light, causing redistri-
bution in numbers and types of microorganisms in the original sample.

6.5.2 After collection, samples should be stored in the dark away from tem-
perature extremes. When transit times are less than 6 h, liquid samples can be
maintained at the original collection temperature by storing in an insulated con-
tainer. If the sample has been collected from sources above 30 °C (86 °F), intrin-
sic heat can maintain the microbial population without significant changes. If the
transit time is longer than 6 h, the sample temperature should be lowered to less
than 10 °C (50 °F) to restrict growth and competition. A standard method used to
control post-sampling shifts in microflora has been to provide a cold-temperature
shock by packing ice packs around sample containers to bring the temperature
down to within the range of 1 to 4 °C (34 to 39 °F), which reduces microbial activ-
ity to a basic survival metabolic mode. Prolonged storage for periods of longer
than a few days can cause changes in the microflora and should be avoided.

Section 7: Testing Guidelines

7.1 Field vs. Laboratory Testing

7.1.1 Bacteria, being living organisms, are highly sensitive to changes in their
environment (e.g., temperature, salinity, and dissolved gases). Additionally, many
chemical species associated with microbial metabolism, such as organic acids or
sulfide compounds, can be rapidly oxidized or degraded. Thus, to obtain results
that accurately represent pipeline conditions, certain tests relevant to MIC inves-

NACE International TM0106-2016 17

 tigation or monitoring must be performed within minutes or hours of sample col-
lection. Historically, this has been one of the factors that resulted in the lack of
meaningful data for use in MIC assessment of pipelines because the analytical
results may be misleading. Therefore, collecting samples for microbiological anal-
ysis should be avoided when pipe and coating have been extensively handled,
exposed to atmosphere or sunlight for extended periods of time, or otherwise
exposed to conditions where dehydration, extreme temperature changes, or con-
tamination could occur. If such samples are analyzed, the compromising condi-
tions should be documented so they may be taken into consideration when a final
evaluation of the data at that site is performed.

7.1.2 As technology has improved, more types of tests and more sophisticat-
ed analyses have been made available for use in the field (i.e., near the point
where samples are collected). Because indicators of the environmental condi-
tions under which MIC may occur are readily degraded or lost after removal of a
sample from the pipeline, a general rule for improving the quality of data is to
perform testing on location whenever practical. Proper sample preservation and
handling procedures must be followed diligently and consistently in cases in
which testing cannot be performed in the field.

7.1.3 Results from both corrosion and microbiological tests should be integrat-
ed when evaluating the threat, likelihood, or presence of MIC. Information on both
types of tests is provided in Section 7. Many of the test procedures described
here are equally useful for evaluating external MIC.

7.2 Microbiological Culture Testing

7.2.1 Although MIC is often attributed to a single type of microorganism, more

often the corrosion is caused by the activities of several different organisms that
form a community. Microbiological testing of buried pipelines has generally includ-
ed testing for SRB, APB, general aerobic organisms, general anaerobic organ-
isms, and in some cases, iron-depositing and iron-reducing bacteria. It should be
noted that iron-depositing and iron-reducing bacteria are very difficult to grow in
culture, and microscopic analysis is more commonly used for detection of these
bacteria.

7.2.2 The objective of the microbiological culture techniques described in this

standard is to approximate the size of the viable bacterial population in a solid or liquid
sample using semi-quantitative estimates, or preferably, the most probable number
(MPN) method when sample replication is employed.28,29 Such estimates are based
on the assumption that bacteria are normally distributed in liquids or solids. Microor-
ganisms in solid and liquid environmental samples are usually enmeshed in particu-
lates. Typically, a suspension of solid samples is made in a Ringers solution or phos-
phate buffer. Particles should be dispersed so that colony forming units are separated
to maximize the accuracy of the estimate. Water samples should be mixed by shak-
ing or stirring for 10 to 60 sec just prior to dilution.

7.2.3 To culture microorganisms, a small amount of liquid or a suspension of

a solid is added to a solution or solid that contains nutrients. The small sample is
called an inoculum, and the nutrient is called the culture or growth medium. There
are normally three considerations when growing microorganisms: type of culture
medium, incubation temperature, and length of incubation.

7.2.4 The type of medium used to culture microorganisms determines to a

large extent the numbers and types of microorganisms that grow. No growth me-
dium can approximate the complexity of a natural environment. Under ideal cir-
cumstances, liquid culture provides favorable growth conditions for 1 to 10% of
the natural population.30 Typically, the presence of specific types of organisms is

18 TM0106-2016 NACE International

 established, and a standard methodology is used so that comparisons can be
made. There are no “correct” culture media. A convenient method is to purchase
prepared, pre-measured, pre-sterilized media. Test kits also generally include sy-
ringes, swabs, and marking pens. Several culture media formulations for various
groups of microorganisms are included in NACE Standard TM0194.28

7.2.5 Highly colored samples may sometimes interfere with culture interpretations.

7.3 Microscopy Methods—General

7.3.1 Microscopy is most commonly used to examine liquid or solid samples

directly to determine the overall numbers of microorganisms present without re-
gard to their viability or species. The procedure involves placing a few µL of sam-
ple on a glass slide, preparing the slide for examination using various staining
techniques, and examining the slide with a light microscope at magnifications
from approximately 500x to 1,500x. Depending on the method of sample prepa-
ration, the detection limit for all microorganisms present in a liquid sample, includ-
ing viable, nonviable, and dead cells, is approximately 102 to 103 cells per mL.
Levels of microorganisms in coastal seawaters are typically > 106 cells per mL.
This procedure is typically performed in the laboratory.

7.3.2 Microorganisms structurally consist of lipids and proteins that degrade

and break down once the cell dies if the sample is not preserved after collection.
Samples collected for microscopy are often preserved (fixed) using a formalde-
hyde or glutaraldehyde phosphate buffer solution. This type of preservation kills
the microorganisms but preserves or fixes the structure of the cell. Fixative solu-
tion vials are commercially available. Typically only a few mL of the liquid or solid
sample are preserved in the fixative.

7.3.3 One advantage of microscopy is that only a minute amount of sample is

required for examination. A surface swab can provide adequate sample material
(e.g., when no bulk liquid or solid is present).

7.4 Epifluorescent Microscopy

7.4.1 Epifluorescent microscopy involves treating the sample with a stain that
fluoresces when viewed under a specific wavelength of ultraviolet light. This tech-
nique helps distinguish microorganisms from debris, or may be used to examine
specific cellular structures of microorganisms. Hydrocarbons and some organic
materials may interfere with epifluorescent microscopy as they give auto-fluores-
cence and obscure the signal from the biological material. This procedure is typi-
cally performed in the laboratory.

7.4.2 Biological stains such as acridine orange (N,N,N’,N’-tetramethylacri-

dine-3,6-diamine), fluorescein isothiocyanate (FITC), and DAPI are used for epi-
fluorescent microscopy. A variety of microbiological test kits are commercially
available. Acridine orange is a common nucleic acid stain that permeates cells to
interact with DNA and RNA.

7.4.3 Fluorescent probes have been developed to “label” specific groups of

microorganisms, or to distinguish live vs. dead cells in a sample. Fluorescent in
situ hybridization (FISH) probes are used to identify and quantify certain species
and groups of microorganisms. Because FISH labels only microorganisms with a
certain content of ribosomal RNA, it is only active cells or cells that have recently
been active that are enumerated. Quantitative use of FISH probes is discussed in
Paragraph 7.8.2.

7.4.4 The DAPI method quantifies all intact microorganisms containing DNA

NACE International TM0106-2016 19

 (both living and inactive cells) in almost any type of liquid sample.31,32 DAPI is often
used in combination with FISH analysis to help distinguish the total cell count from the
number of cells that are labeled using the FISH probes. The full DAPI method may be
completed in less than half a day in a laboratory. The liquid sample is filtered whereby
microorganisms (cells of bacteria, Archaea, and fungi) are collected on a filter. These
are subsequently stained with a fluorescent dye that binds to the DNA in the cells and
then washed to remove excess dye. Cells are manually counted using an epifluores-
cence microscope. Fluid samples should be fixed in the field with formaldehyde
(36%) to a final concentration of 2% in the sample. Adequate sample amounts for this
type of testing are 50 to 100 mL (1.6 to 3.40 oz) of fluid depending on the cell density.

7.4.5 Microscopy and biochemical methods have been used for many years, and
genetic methods are now becoming available for the detection, quantification, and in
some cases identification of microorganisms present at corrosion sites.33-35

7.5 Adenosine Triphosphate Photometry (ATP)

7.5.1 ATP, related to energy production and consumption, is present in all liv-
ing cells. When cells die, however, ATP rapidly degrades. Consequently, the
quantity of ATP in field samples is approximately proportional to the number of
living microorganisms in that sample. ATP may give an indication of the viable
biomass present in living organisms, and may be measured using an enzymatic
reaction that generates light when ATP is present. The intensity of the light is
measured in a photomultiplier, the output being proportional to the amount of ATP.

7.5.2 Several commercial field test kits are available for ATP quantification.
Quantification of ATP typically relies on photometers that measure the amount of
light emitted when the ATP within the sample is allowed to react with a particular
enzyme. Before the reaction, the sample to be quantified is filtered and treated
with gold buffers. These buffers assist in releasing the ATP from the organism so
they can react with the enzyme. Finally, the sample and enzyme are combined
and analyzed. Advantages of ATP measurement include speed to results (less
than 10 min per sample), no underestimation of unculturable organisms, and use
in any sample type including produced fluids, oil/emulsions, and solids. ATP mea-
sures should be backed up with at least one additional method for more specific
quantification of the microorganism (e.g., SRB).

7.6 Hydrogenase Measurements

7.6.1 Hydrogenase is an enzyme produced by bacteria that use hydrogen as an

energy source. Testing for the presence of the hydrogenase is one method used to
enumerate bacteria populations in corrosion deposits and water samples in the field.

7.6.2 Quantification of bacteria populations using this method first involves

extraction of the hydrogenase enzyme from the sample. The extracted enzyme is
preserved in a solution that maintains enzyme activity and then placed within a
reaction chamber where hydrogen is introduced. The hydrogen is oxidized and a
redox indicator color change reveals the presence of the hydrogenase enzyme
(refer to NACE Standard TM0194). The reaction typically is not rapid; it can take
anywhere from 30 min to 4 h. The reaction time and developed color intensity
together are used to measure the relative activity of the enzyme.

7.7 Adenosine Phosphosulfate Reductase (APS)

7.7.1 APS reductase is an enzyme specifically associated with SRB. Mea-

surement of the APS reductase present in a bacterial sample provides an indica-
tion of the active SRB concentration. Detection and measurement are based on
immunological methods and may be performed using a field kit.

20 TM0106-2016 NACE International

 7.7.2 The test involves exposure of the sample to small particles containing
antibodies. These particles specifically capture the APS reductase enzyme. The
particles, now mixed with APS reductase, are subsequently isolated on a porous
membrane and exposed to specific indicator chemicals. Reaction between the
particles and chemicals results in a color change that is proportional to the con-
centration of the APS reductase in the sample.

7.8 Molecular Microbiological Methods (MMM)

7.8.1 MMM, also referred to as genetic methods, are culture-independent ap-

proaches that provide direct analysis of samples without the bias introduced by
the growth process used during culturing. Because no prior growth of microorgan-
isms is required, MMMs accept very small amounts of any type of sample (liquid,
biofilm, solid) with or without live bacteria. After genetic materials are extracted
from the sample, assays that are very specific and render a more precise quanti-
fication of various types of bacteria than culture tests are performed in the labora-
tory.36,37 A comparison of MMMs is shown in Table 1 and Figure 2.

Table 1
Comparison of MMMs31
Living
Method Dead Cells Quantitative
MMM Cells Information Yielded
Based On Counted? Method?
Counted?
DAPI Microscopy Yes Yes Yes Total cell counts (live and dead)
Total numbers of live bacteria
Total numbers of live Archaea
FISH Microscopy Yes No Yes
Total numbers of live SRB
Total numbers of live SRA
Comparison of populations
DGGE PCR Yes Yes No
Identification of abundant microorganisms
Numbers of total bacteria
Numbers of total Archaea
qPCR PCR Yes Yes Yes Numbers of SRB
Numbers of SRA
Numbers of three groups of methanogens2

7.8.2 Quantitative FISH—quantitative FISH is a MMM in which only living and

active cells are stained with a fluorescent dye visible during epifluorescence mi-
croscopy. Unlike the DAPI method, FISH probes may be designed to attach only
to selected groups of microorganisms (e.g., specific types of SRB or SRA). There-
fore, only the specific target microorganisms are visible and may be enumerated
during subsequent microscopy. The quantitative FISH method does not underes-
timate organisms that do not grow in culture. FISH is a microscopy method that
uses synthetic oligonucleotides (synthetic DNA) tagged with a fluorescent mole-
cule (dye). Together, the synthetic DNA and the fluorescent molecule are referred
to as a probe. The probe is mixed with the bacteria in a fluid sample and the
numbers of bacteria (or Archaea) that take up the probe and have it hybridize to
their rRNA are counted. The quantitative FISH method differentiates active/alive
microorganisms from dead microorganisms. A probe may be designed to detect
a general population (e.g., total bacteria or total Archaea) or a specific genus or
species (e.g., Desulfovibrio desulfuricans). A given sample is analyzed using sev- FIGURE 2: An Illustration of the Portions
eral different probes to understand the prokaryotic diversity in a sample, and of the Different Pools of Microorganisms
which types/species of microorganisms are most abundant. The preparation (Live, Inactive, and Dead) Typically Pres-
steps for the quantitative FISH method are different from the DAPI method. How- ent in Samples from the Oil Industry that
ever, cells stained for the quantitative FISH method are counted in the same are Enumerated Using Various MMMs as
manner as in direct bacterial counts (for the DAPI method) in the laboratory. Ade- Compared to the MPN (Culturing) Meth-
quate sample amounts for this type of testing are in the range of 50 to 100 mL (1.6 od. Each of the Methods Indicated is
to 3.40 oz) of fluid depending on the cell density. Samples are filtered onto 0.2 μm Discussed Further in the Text.37

NACE International TM0106-2016 21

 filters. FISH probes are selected to target total bacteria and Archaea, as well as
specific groups of relevance (e.g., SRB). The samples are inspected by epifluo-
rescence microscopy at 1,000x magnification. The cells stained with each probe
are counted and related to the overall number of cells obtained by the DAPI meth-
od (see Paragraph 7.4.4).

7.8.3 qPCR—PCR and qPCR is a MMM to amplify a single or few copies of a

piece of DNA across several orders of magnitude, generating millions of copies of a
particular DNA sequence. The qPCR is an emerging method for enumerating micro-
organisms in complex environmental samples, particularly in solid samples where
epifluorescence microscopy (used for the DAPI method and FISH method) may be
difficult to perform because of background interference. The qPCR method enumer-
ates genes rather than individual cells by applying a modified polymerase chain reac-
tion (PCR) method. Like the quantitative FISH method, the qPCR method may be
applied to count total cells or specific groups of microorganisms (e.g., SRP or metha-
nogenic Archaea).9,15,38,39 Because qPCR targets the DNA in all prokaryotes, the
qPCR method measures living, inactive, and dead microorganisms. qPCR may be
used to quantify the total number of microorganisms or a specific genus/species of
microorganisms in nearly any type of sample, including produced fluids, oil/emulsion,
and solids. The qPCR method does not underestimate organisms that do not grow in
culture. qPCR may be done on both fluid and solid samples as well as microorgan-
isms collected via membrane filtration. qPCR uses synthetic DNA (called primers)
tagged with a fluorescent molecule or synthetic DNA mixed with a DNA intercalating
agent (dye) to quantify organisms using a modified version of the PCR method. Brief-
ly, total prokaryotic DNA is extracted and amplified using primers that target to a
conserved region of the bacterial DNA. The region used may make the assay very
general (meaning counting total bacteria or Archaea), or very specific (meaning
counting a single genus or species). Similar to quantitative FISH, the qPCR method
may be used to enumerate a very general group of bacteria (i.e., total bacteria or
Archaea) or very specific organism (e.g., Desulfovibrio desulfuricans). Adequate
sample amounts for this type of testing are 50 mL (1.7 oz) of fluid, 2.0 to 5 g (0.07 to
0.1 oz) of solid, or a 0.2 or 0.45 μm filter with a minimum of 10 mL (0.30 oz) of fluid
passed through it.

7.8.4 DGGE—MMM method based on the PCR method that is used for com-
paring microbial communities across a number of different samples. During
DGGE, genetic material in individual samples is amplified by PCR and subse-
quently compared by electrophoresis. DGGE is used for identifying dominant
groups of microorganisms in individual samples and for evaluating how the micro-
organisms are distributed between samples.41-43 DGGE may be performed on
any fluid or solid sample, as well as bacteria collected via membrane filtration.
Adequate sample amounts for this type of testing are 50 mL (1.7 oz) of fluid, 2 to
5 g (0.07 to 0.1 oz) of solid, or a 0.2 or 0.45 μm filter with a minimum of 10 mL
(0.30 oz) of fluid passed through it.

7.9 Organic Acids

Organic acids such as butyric, pyruvic, propionic, and acetic acid are by-products or
intermediary species of microbial metabolism. Gas chromatography-mass spectrom-
etry (GC-MS) or high-pressure liquid chromatography (HPLC) are used in the labora-
tory to identify and quantify organic acid species in properly preserved samples. Or-
ganic acids quickly degrade; therefore, samples are preserved by filtration into
nitrogen-purged vials and maintained at 4 °C (40 °F) until analysis is performed.

7.10 Chemical Analysis

7.10.1 Corrosion products should be taken from the steel surface, coating, or opti-
mally, from the pit contents underneath a deposit that has been removed. The color

22 TM0106-2016 NACE International

 and type of sample collected should be noted in each case. Although liquid or corro-
sion product samples are often limited in volume, when sampled from externally cor-
roded pipe, on-site tests should be performed because significant chemical changes
can occur over a short period of time. Additionally, compositional analyses, i.e., an-
ions, cations/metals, and organic acids should be performed in the laboratory.

7.10.2 Field tests on liquids should include pH, total alkalinity, and dissolved hydro-
gen sulfide. Field tests on solids and corrosion products should include pH and a
qualitative analysis for the presence of sulfides and carbonates. Carbonates are pres-
ent if noticeable bubbling occurs when a drop of dilute hydrochloric acid is placed on
a small portion of the corrosion product. Sulfides can be detected by the characteris-
tic odor of rotten eggs or by exposing the acid-treated corrosion product to lead-ace-
tate test paper. A white-to-brown color change occurs in the presence of sulfides.
Follow-up testing in the laboratory with more sophisticated analytical equipment (e.g.,
energy dispersive spectroscopy [EDS], x-ray diffraction [XRD], etc.) to determine the
elemental and mineral phases present should be performed to verify field tests. When
collecting corrosion product samples from steel surfaces, coating, or pit contents for
laboratory analysis, a large enough sample should be taken to enable testing by var-
ious methods. Avoid reusing the material from one test method for another test meth-
od if multiple tests are performed.

7.11 Pipeline Examination

7.11.1 When examining external surfaces of the exposed pipeline, disbonded

coating, corrosion products, soil, and other materials should be removed from the
pipe wall using a clean spatula or knife, with care taken not to scratch the metal. Any
remaining material should be removed with a clean, dry, stiff brush, e.g., nylon bristle
brush. A brush with metal bristles obscures the pit features. In cases when all of the
product cannot be removed with this method, a brass bristle brush may be used in the
longitudinal direction. The area may subsequently be cleaned with an air blast or an
alcohol swab. A shiny metallic surface in the pit suggests the possibility of active cor-
rosion. However, judgment should be used to differentiate this condition from one
created by scraping the steel surface with a metallic object, such as the knife or spat-
ula used to clean the surface or to obtain the sample product.

7.11.2 The steel surface shall be inspected for corrosion, and any damage shall
be carefully documented. When possible, gauges should be used to measure the
pit depths. Also, the length of the corroded area in relation to the circumferential
and longitudinal position should be determined. The newly cleaned corroded area
should first be examined without magnification. Then, a low-power magnifying
lens at 5 to 50x power should be used to examine the detail of the corrosion pits.
An example checklist is in Appendix A.

7.12 Analysis of Pipeline Samples

7.12.1 Careful analysis of pipeline samples (pipe sections or components that

have been removed from service) may provide useful information regarding inter-
nal corrosion mechanisms.

7.12.2 Precautions should be taken to avoid contamination of the external sur-

faces of the pipeline before, during, and after the pipe section or component is
removed from service. If inadvertent alteration of the external surface of the pipe-
line sample occurs, the nature of the alteration should be noted to aid in correct
interpretation of subsequent testing results.

7.12.3 Alteration of pipeline samples can occur as a result of removal efforts,

exposing the surface deposits to oxygen, soil, foreign matter, temperature chang-
es, and contamination from handling.

NACE International TM0106-2016 23

 7.12.4 Samples should be collected immediately after the pipe section or compo-
nent is removed from service (i.e., within minutes when possible). Corrosion products
and biofilms can change profoundly on exposure to air, affecting test results.

7.12.5 When samples cannot be collected immediately after a pipe section or

component is removed, the external surfaces and deposits may be covered tem-
porarily with new, clean, plastic sheeting to minimize exposure to air until samples
are collected. This practice may compromise the condition of the surface samples
to an unknown extent, and should be used only when samples cannot be collect-
ed immediately.

7.12.6 Culture tests of samples from pipe that has been exposed to air, dehy-
dration, and potential contamination for extended periods of time (i.e., days)
should not be relied on for providing useful data.

7.12.7 The internal condition of pipeline samples at the time of removal should be
carefully and thoroughly documented; these data are important in the interpretation of
both field and laboratory tests. An example checklist is provided in Appendix A.

7.12.8 Field and laboratory tests of pipeline samples for external corrosion
analysis should be directed toward characterization of the biological, chemical,
and metallurgical conditions present in the pipeline. In particular, distinction
should be made between samples collected in corroded areas vs. areas where no
corrosion is present.

7.12.9 Corrosion features on the pipeline sample should be protected from fur-
ther corrosion after removal of the pipeline sample. Microscopic features of corro-
sion damage are easily lost because of oxidation or improper handling.

7.12.10 Optical microscopy and scanning electron microscopy (SEM) of internal

corrosion features may provide useful information regarding the origin and growth
of localized corrosion.

7.12.11 Metallographic examination of corroded areas removed from pipeline

samples may provide information about the nature of the corrosion relative to the
microstructure of the pipeline. Particularly, selective or preferential corrosion of
microstructural features may be determined from metallographic examination.

7.12.12 Chemical analysis of microscopic corrosion features using EDS or other

microanalytical techniques may provide useful data regarding localized corrosion
initiation mechanisms.

7.12.13 Preservation methods, such as in situ histological embedment of bio-

films and corrosion products, may yield samples suitable for microscopic exam-
ination using fluorescent staining techniques, phase contrast examination of thin
sections, and transmission electron microscopy (TEM).

7.12.14 Interpretation of data collected from pipeline samples should be per-

formed as described in Section 8.

7.13 Data and Records Management

All data and information, e.g., sample collection method used, should be docu-
mented on field data sheets or in logbooks with permanent ink.

7.14 Inspection Techniques

7.14.1 Inspection data are used to detect and monitor corrosion-related dam-

24 TM0106-2016 NACE International

 age. Techniques include visual inspection, ultrasonic testing (UT), radiographic
testing (RT), and magnetic flux methods. Inspection may be used for establishing
the orientation, distribution, density, size, shape, and extent of external corrosion
damage; however, inspection results alone do not establish the presence of MIC.
Inspection data should be integrated with other information about the internal
environment of the pipeline.

7.14.2 The results of ILI may provide information about the location and sever-
ity of external corrosion relative to operating parameters, design, elevation, and
other considerations.

7.14.3 An external corrosion direct assessment (ECDA) standard has been

published for pipelines in ANSI/NACE SP0502.43 As for ILI data, ECDA methods
provide information about the location and severity of external corrosion relative
to pipeline design and operating conditions.

Section 8: Application of Test Methods to Pipelines

and Interpretation of Data
8.1 Data Interpretation

8.1.1 Because microorganisms are ubiquitous, the presence of bacteria or

other microorganisms does not necessarily indicate a causal relationship with
external corrosion observed on a pipeline. In fact, microorganisms can nearly al-
ways be cultured from natural environments. Therefore, merely detecting viable
bacteria in liquid or solid samples associated with external corrosion does not
necessarily prove that MIC has occurred.

8.1.2 To determine the cause of external corrosion, all chemical, microbiolog-

ical, metallurgical, and operational data about the pipeline site must be examined,
integrated, and analyzed. Analytical results from samples obtained in the corrod-
ed area should be compared with the results from reference samples taken out-
side the corroded area.

8.1.3 To determine the presence of external MIC on a pipeline, microbiologi-

cal, operational, and chemical data must be integrated. Analysis of the data
should demonstrate that microorganisms and their activities have provided the
predominant influence over the corrosion mechanism present on the pipeline, as
opposed to abiotic mechanisms.

8.1.4 Pipeline operators may collect data in support of internal MIC analysis in
conjunction with other routine sampling, maintenance, integrity assessment, in-
spection, and environmental and regulatory compliance activities.

8.1.5 Interpretation of data relative to the assessment or determination of MIC on

a pipeline should consider a number of factors that can influence both corrosion and
microorganism growth, including: soil type, soil composition, groundwater composi-
tion (if present), temperature, coating condition, and cathodic protection.

8.1.6 Interpretation of data related to bulk phase (macro scale) conditions

must be done in consideration of the fact that microorganisms can exist and flour-
ish in microniches. For example, pipelines may experience little or no corrosion
damage as a result of a wide range of conditions throughout the majority of the
pipeline, yet be affected by MIC in a small area of a specific section of the pipeline
because of the unique environment present only at that location.

NACE International TM0106-2016 25

 8.1.7 Because MIC is a complex mechanism that involves electrochemistry,
microbiology, corrosion control, pipeline operation and design, as well as engi-
neering and integrity assessment, plans to evaluate MIC of pipelines and analyze
data should include input from those with expertise in the respective fields. Pipe-
line operators should seek input from a multidisciplinary team whenever possible,
so as not to emphasize one aspect of science or technology over another.

8.2 Corrosion Damage Investigation

8.2.1 MIC is suggested by increased levels of viable microorganisms associ-

ated with pit areas that were uncontaminated by adjacent groundwater or soil, or
by microscopic determination of iron and manganese bacteria in corrosion depos-
its. These are often good indicators of microbial involvement.

8.2.2 To validate MIC as the cause of internal corrosion, the following three
conditions must be met:

8.2.2.1 Condition 1—Assuming there is no known or suspected con-

tamination from outside sources, demonstration of increased levels of
specific types of viable microorganisms (bacteria or fungi) associated
with the corrosion, relative to samples taken outside the corroded area.

8.2.2.2 Condition 2—Chemical indicators that support the microbio-

logical evidence (e.g., elevated levels of sulfide or sulfur in pit deposits
from SRB and SRA, or organic acids from APB) are identified in the
corroded area.

8.2.2.3 Condition 3—Biotic factors (the presence or activities of living

organisms) are identified as the primary contributor to the corrosion
damage. The objective of this verification step is to establish that the
presence of specific biotic conditions was the predominant contributor to
the corrosion observed. The influence of abiotic factors (chemical or
physical conditions unrelated to living organisms) on the corrosion
mechanism also must be considered in all cases. The nature of the cor-
rosion damage to the pipeline system should be consistent with the na-
ture of the identified microorganism(s) and their by-products, or their
physical influence on the formation of corrosion cells. For example, if
viable APB or methanogens are concentrated at the corrosion damage
relative to the environment, and evidence of their metabolic activity (or-
ganic acids) is determined to be associated with the corrosion, the na-
ture of the corrosion damage should be consistent with these observa-
tions (e.g., accelerated corrosion damage or pitting beneath biofilms or
deposits). This is an important step in the final diagnosis because it is
often difficult to discern between the relative contributions of various
factors (biotic and abiotic) affecting localized corrosion.

8.3 Corrosion Mitigation

8.3.1 The methods described in this standard may be used to determine the need
for, and effectiveness of, mitigation measures for controlling MIC. Specific procedures
for mitigating external MIC of pipelines are beyond the scope of this standard.

8.3.2 NACE SP0169 provides general information about methods for con-
trolling external MIC by design, operation, and specific measures, such as coat-
ings and cathodic protection.

8.3.3 The mitigation measures used to control external corrosion of pipelines

may be effective for both biotic and abiotic corrosion mechanisms.

26 TM0106-2016 NACE International

References
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10. T.R. Jack, G. Van Boven, M. Wilmott, R. Worthingham, “Evaluating Performance of Coatings Exposed to Biologically Active Soils,”
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11. J. Larsen, K.B. Sørensen, B. Højris, T.L. Skovhus, “Significance of Troublesome Sulfate-Reducing Prokaryotes (SRP) in Oilfield
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16. M. Davies, P.J.B. Scott, Oilfield Water Technology (Houston, TX: NACE, 2006), p. 213.

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18. J.O. Harris, “Bacterial Activity at the Bottom of Back-Filled Pipeline Ditches,” CORROSION 16, 3 (1960): p. 441.

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21. J. Guezennec, “Influence of Cathodic Protection of Mild Steel on the Growth of Sulfate-Reducing Bacteria at 35 °C in Marine Sediments,”
Biofouling 3, 4 (1991): p. 339.

NACE International TM0106-2016 27

22. J. Guezennec, M. Therene, “A Study of the Influence of Cathodic Protection on the Growth of SRB and Corrosion in Marine
Sediments by Electrochemical Techniques,” 1st European Federation of Corrosion Workshop on Microbiological Corrosion, held
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24. T.J. Barlo, W.E. Berry, “An Assessment of the Current Criteria for Cathodic Protection of Buried Steel Pipelines,” MP 23, 9 (1984):
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25. NACE SP0169 (latest revision), “Control of External Corrosion on Underground or Submerged Metallic Piping Systems” (Houston,
TX: NACE).

26. ASTM G57 (latest revision), “Standard Test Method for Field Measurement of Soil Resistivity Using the Wenner Four-Electrode
Method” (West Conshohocken, PA: ASTM).

27. R.B. Eckert, Field Guide for Investigating Internal Corrosion of Pipelines (Houston, TX: NACE, 2003).

28. NACE Standard TM0194 (latest revision), “Field Monitoring of Bacterial Growth in Oilfield Systems” (Houston, TX: NACE).

29. Standard Methods for the Examination of Water and Wastewater (Washington, DC: APHA,(4) Denver, CO: AWWA,(5) and Alexan-
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31. J. Larsen, T.L. Skovhus, M. Agerbæk, T.R. Thomsen, P.H. Nielsen, “Bacterial Diversity Study Applying Novel Molecular Methods on
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32. J. Larsen, S. Zwolle, B.V. Kjellerup, B. Frølund, J.L. Nielsen, P.H. Nielsen, “Identification of Bacteria Causing Souring and Biocorro-
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paper no. 88249 (Houston, TX: NACE, 1988).

34. X. Zhu, J. Lubeck, K. Lowe, A. Daram, J.J. Kilbane II, “Improved Method for Monitoring Microbial Communities in Gas Pipelines,”
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35. T.L. Skovhus, B. Højris, A.M. Saunders, T.R. Thomsen, M. Agerbæk, J. Larsen, “Practical Use of New Microbiology Tools in Oil
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36. X.Y. Zhu, A. Ayala, H. Modi, J.J. Kilbane II, “Applications of Quantitative, Real-Time PCR in Monitoring Microbiologically Influenced
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37. C. Whitby, T.L. Skovhus, eds., Applied Microbiology and Molecular Biology in Oilfield Systems (New York, NY: Springer, 2011).

38. K. Takai, K. Horikoshi, “Rapid Detection and Quantification of Members of the Archaeal Community by Quantitative PCR Using
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(5)
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28 TM0106-2016 NACE International

40. R. Sooknah, S. Papavinasam, R. W. Revie, “Validation of a Predictive Model for Microbiologically Influenced Corrosion,” CORRO-
SION 2008, paper no. 08503 (Houston, TX: NACE).

41. J. Larsen, T.L. Skovhus, A.M. Saunders, B. Højris, M. Agerbæk, “Molecular Identification of MIC Bacteria from Scale and Produced
Water: Similarities and Differences,” CORROSION 2008, paper no. 08652 (Houston, TX: NACE, 2008).

42. V.V. Keasler, et al, “Identification and Analysis of Biocides Effective Against Sessile Organisms,” SPE International Symposium on
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Bibliography
T.J. Barlo, W.E. Berry, “An Assessment of the Current Criteria for Cathodic Protection of Buried Steel Pipelines,” MP 23, 9 (1984): p. 9.

NACE SP0169 (latest revision), “Control of External Corrosion on Underground or Submerged Metallic Piping Systems” (Houston, TX:
NACE).

ASTM G57 (latest revision), “Standard Test Method for Field Measurement of Soil Resistivity Using the Wenner Four-Electrode Method”
(West Conshohocken, PA: ASTM).

R.B. Eckert, Field Guide for Investigating Internal Corrosion of Pipelines (Houston, TX: NACE, 2003).

NACE Standard TM0194 (latest revision), “Field Monitoring of Bacterial Growth in Oilfield Systems” (Houston, TX: NACE).

Standard Methods for the Examination of Water and Wastewater (Washington, DC: APHA,(4) Denver, CO: AWWA,(5) and Alexandria,
VA: WEF,(6) 2006).

P.J.B. Scott, “Expert Consensus on MIC: Prevention and Monitoring—Part 1,” MP 43, 3 (2004): p. 50.

J. Larsen, T.L. Skovhus, M. Agerbæk, T.R. Thomsen, P.H. Nielsen, “Bacterial Diversity Study Applying Novel Molecular Methods on
Halfdan Produced Waters,” CORROSION 2006, paper no. 06668 (Houston, TX: NACE, 2006).

J. Larsen, S. Zwolle, B.V. Kjellerup, B. Frølund, J.L. Nielsen, P.H. Nielsen, “Identification of Bacteria Causing Souring and Biocorrosion
in the Halfdan Field by Application of New Molecular Techniques,” CORROSION 2005, paper no. 05629 (Houston, TX: NACE,
2005).

D.H. Pope, T.P. Zintel, “Methods for the Investigation of Under-Deposit Microbiologically Influenced Corrosion.” CORROSION/88, paper no.
88249 (Houston, TX: NACE, 1988).

X. Zhu, J. Lubeck, K. Lowe, A. Daram, J.J. Kilbane II, “Improved Method for Monitoring Microbial Communities in Gas Pipelines,”
CORROSION 2004, paper no. 04592 (Houston, TX: NACE, 2004).

T.L. Skovhus, B. Højris, A.M. Saunders, T.R. Thomsen, M. Agerbæk, J. Larsen, “Practical Use of New Microbiology Tools in Oil Produc-
tion,” SPE Offshore Europe Conference 2007, paper no. SPE 109104 (Richardson, TX: SPE, 2007).

X.Y. Zhu, A. Ayala, H. Modi, J.J. Kilbane II, “Applications of Quantitative, Real-Time PCR in Monitoring Microbiologically Influenced
Corrosion (MIC) in Gas Pipelines,” CORROSION 2005, paper no. 05493 (Houston, TX: NACE, 2005).

C. Whitby, T.L. Skovhus, eds., Applied Microbiology and Molecular Biology in Oilfield Systems (New York, NY: Springer, 2011).

K. Takai, K. Horikoshi, “Rapid Detection and Quantification of Members of the Archaeal Community by Quantitative PCR Using Fluoro-
genic Probes,” Applied and Environmental Microbiology 66, 11 (2000): p. 5066.

NACE International TM0106-2016 29

T.L. Skovhus, N.B. Ramsing, C. Holmstrom, S. Kjelleberg, I. Dahllof, “Real-Time Quantitative PCR for Assessment of Abundance of
Pseudoalteromonas Species in Marine Samples,” AEM 70, 4 (2004): p. 2373.

R. Sooknah, S. Papavinasam, R. W. Revie, “Validation of a Predictive Model for Microbiologically Influenced Corrosion,” CORROSION
2008, paper no. 08503 (Houston, TX: NACE).

J. Larsen, T.L. Skovhus, A.M. Saunders, B. Højris, M. Agerbæk, “Molecular Identification of MIC Bacteria from Scale and Produced
Water: Similarities and Differences,” CORROSION 2008, paper no. 08652 (Houston, TX: NACE, 2008).

V.V. Keasler, et al, “Identification and Analysis of Biocides Effective Against Sessile Organisms,” SPE International Symposium on
Oilfield Chemistry 2009, paper no. SPE 121082 (Richardson, TX: SPE, 2009).

ANSI/NACE Standard SP0502 (latest revision), “Pipeline External Corrosion Direct Assessment Methodology” (Houston, TX: NACE).

(7)
Gas Technology Institute (GTI), formerly Gas Research Institute (GRI), 1700 S. Mount Prospect Road, Des Plaines, IL 60018-1804.

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 Appendix A
(Nonmandatory)
Site Inspection and Testing

A1.1 A sample checklist used for site inspection and testing is provided in Table A1:

Table A1
Site Inspection and Testing Checklist

Paragraph No. Inspection Point Observations

6.1.1 Observations prior to excavation
General topography of site
Soil type
6.1.2 Measurements prior to excavation
Soil resistivity
Pipe-to-soil potential
Close interval survey
Current mapping
Redox potential
Other
6.1.3 Observations during excavation
Method of excavation
Coating damaged while digging?
Soil depth to top of pipe
Special features (bend, fitting, etc.)
Previous repair area?
Observations after excavation
6.1.4.1 Coating type (coal tar, asphalt, bitumen, tape, wax, epoxy)
Nature of coating damage (disbonding, blistering, tenting,
cracking, wrinkling)
Extent of coating damage (length, circumference, area of
pipe exposed)
Location of coating damage relative to pipe girth and seam
welds, and coating seam if applicable
Typical location of coating damage (top, bottom, sides,
random)
Coating repairs
Age of coating
Plant or field application
6.1.4.2 Soil moisture level
Soil type (silt, sand, gravel, rock, clay, peat, other)
Discoloration near pipeline
Soil strata depths
Running water (spring)
Seasonal effects at location
Soil or groundwater pH
Soil pH around pipeline
Note soil samples collected for analysis

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 6.1.4.3 Relationship of corrosion to:
Distance from nearest compressor station
Inlets, outlets, taps, fittings
Heat sources or temperature change
Construction/material changes
Recent repairs or damage
Power lines
Other pipelines or underground facilities
6.2.1 Corrosion products/deposits color
Nature of deposits (scale, nodule, film)
Deposit texture (hard, soft, friable)
Deposit odor
Deposit strata (note changes in layers if they exist)
Note visual differences between general deposits and
localized deposits associated with corrosion
Relationship between coating and deposits (e.g., beneath
coating, on top of coating, etc.)
Calcareous deposits present?
Chemical spot testing results
Note deposit samples collected for analysis
Visible biological accumulations in deposits or on pipe
Liquid present beneath coating?
6.2.2 Observations of corrosion damage
Nature of corrosion damage
Isolated pitting
Isolated pitting within areas of general corrosion
Linked pitting within areas of general corrosion
General metal loss with few deeper pits
Etching or general metal loss with no pitting
Selective attack at welds
Crevice corrosion (at flange joints, mechanical joints)
Pit morphology (elliptical, parabolic, narrow, grain attack,
subsurface). See Figure 1.
Pit features (striations, tunnels, cup shape, pits within pits,
undercutting, strata levels, grooves, shiny, dull)
Other observations
Severity of corrosion:
Longitudinal extent
Circumferential extent
Maximum wall loss
Profile of wall loss
Maximum/average pit depth
Maximum/average pit diameter
Pit length vs. pit width
Depth/diameter ratio
Where is corrosion the most severe?

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6.4 Samples collected for microbial and chemical analysis Note whether sample was collected and exact
location of sample.
Soil samples
a. Undisturbed soil next to pipe
b. Undisturbed soil, bottom of ditch
c. Undisturbed soil, ditch wall, pipe elevation
d. Soil in contact with coating damage or corrosion
Coating/deposit samples
a. Disbonded or damaged coating
b. Deposits at corrosion sites
c. Deposits where no corrosion occurred
d. Scale, biofilm, liquids from under coating
Corrosion samples
a. Corrosion products or nodules
b. Material from beneath nodules
c. Surface swab of pit contents
d. Pipe sample cut-out
General information and history
Year of installation
Pipe diameter and wall thickness
Pipe grade and manufacturer
Year of CP installation
Type of CP system

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34 TM0106-2016 NACE International
NACE International TM0106-2016 35
NACE Standard TM0106-2016 ISBN 1-57590-000-0
Item #00000 NACE International