Journal of Sustainable Cement-Based Materials

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Microbial induced calcite precipitation for self-

healing of concrete: a review

Rishav Garg, Rajni Garg & Nnabuk Okon Eddy

To cite this article: Rishav Garg, Rajni Garg & Nnabuk Okon Eddy (2023) Microbial induced
calcite precipitation for self-healing of concrete: a review, Journal of Sustainable Cement-Based
Materials, 12:3, 317-330, DOI: 10.1080/21650373.2022.2054477

To link to this article: https://doi.org/10.1080/21650373.2022.2054477

Published online: 03 Apr 2022.

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Journal of Sustainable Cement-Based Materials, 2023
Vol. 12, No. 3, 317–330, http://dx.doi.org/10.1080/21650373.2022.2054477

Microbial induced calcite precipitation for self-healing of concrete: a review

Rishav Garga, Rajni Gargb
and Nnabuk Okon Eddyc
*
a
Department of Civil Engineering, Galgotias College of Engineering & Technology, Greater Noida, Uttar Pradesh, India;
b
Department of Chemistry, Rayat Bahra University, Mohali, Punjab, India; cDepartment of Pure and Industrial Chemistry,
University of Nigeria, Nsukka, Enugu State, Nigeria

Microbial induced calcite precipitation (MICP) is a part and consequence of metabolic activities of many bacterial
species with involvement of various pathways in presence of specific organic and/or inorganic moieties present in the
surrounding environment. Microbial or Bioconcrete is a sustainable and promising technology based on MICP that not
only supports the self-healing of concrete structures but also mitigates the detrimental effects of CO2 emissions on
environment from the cement industry. The calcite precipitation inside concrete matrix results in crack healing with an
improvement in matrix characteristics. This paper reviews the various mechanisms involved in MICP and highlights
the role of MICP in self-healing inside cement matrix. The paper further throws light on designing of microbial
concrete and explores the beneficiary effects of MICP on various mechanical and durability properties of cementitious
materials. Further, the prospective challenges associated with microbial concrete production and the future aspects
have been discussed.
Keywords: microbial concrete; calcite precipitation; urea hydrolysis; nitrate reduction; self-healing

1. Introduction The sustainable construction practices not only involve

Concrete is the world's most commonly used construction the reduction of the usage of raw materials and energy but
material, mostly due to its excellent durability, low cost are also aimed to reduce the maintenance cost by increas-
and comparatively low energy consumption compared to ing the resilience and service life of the concrete structures
other building materials [1]. Moreover, with the rising [7]. Researchers are concerned to implement innovative
world population and the increasing urbanization of devel- strategies that can protect the concrete structures in aggres-
oping countries, concrete will be the most desirable and sive environment and minimize the repair cost. Various
consumable construction material [2]. However, a matter studies have been reported in literature on healing of
of concern is the fragility, poor resistance and regular cracks by introduction of fillers in the cracks [8]. It has
been observed that remediation of cracks at the early onset
cracking behavior of concrete [3]. Specifically, the prema-
is more beneficial in terms of better results and cost reduc-
ture degradation of concrete structures under the effect of
tion. In these situations, the concrete that self-heals, i.e.
harsh environmental conditions such as rise in temperature,
fixes cracks spontaneously and easily without external
humidity and stress leads to formation of minor and major
intervention, will most likely play a very useful role [4].
cracks. Cracks tend to weaken the structural integrity and
The phenomenon of self-healing is linked with various
performance, thereby seriously affecting the serviceability
factors including extended hydration of the embedded
and durability. The most threatening factor is the induction unhydrated cement particles, extension of the hydrated gel
of chemical attack on the matrix due to penetration of products in cracks, carbonation of the dissolved calcium
harmful agents such as carbon dioxide, sulfate ions and hydroxide, filling of cracks by shredded concrete particles
chloride ion through these cracks. Especially in case of and the blocking of cracks due to additional crystalline
reinforced structures, it may initiate corrosion of the steel salts formed by reaction with impurities present in water.
reinforcement. In particular, such vulnerabilities are a prob- Recently, the use of bacterial strains in self-healing
lem for the life cycle of intricate structures that impact the concrete has provided ‘Microbial concrete or Bioconcrete’
economy throughout the world [4]. Although the trad- as an innovative domain of research. The implementation
itional methods for maintenance and repair will effectively of a bacterial metabolic process known as Microbial
prolong the service life of the concrete structures, the exor- Induced Calcium Carbonate Precipitation of (MICP) has
bitantly high preservation and conservation costs for large provided an alternate approach for crack healing and mini-
infrastructures are the critical issue [5]. Moreover, it may mizing environmental effect in the last two decades.
be problematic to trace the crack locations, crack sizes and Bioconcrete production is based on biomineralization or
continuous service specifications for infrastructures such as inclusion of bacteria with the ability to cause mineral for-
multi-storey buildings or tunnels. Further, the social and mation in the cement matrix. The applicability of microbial
environmental impacts of structure deterioration can be concrete with potential benefits in terms of durability
detrimental as the disposal and recycling of waste construc- enhancement and ecological benefits has been well
tion materials are also associated with energy cost [6]. explored in many studies. The use of microbial concrete

Corresponding author. Email: [email protected]

*Co-Corresponding author. Email: [email protected]
ß 2022 Informa UK Limited, trading as Taylor & Francis Group
318 R. Garg et al.

Figure 1. Mechanism illustrating (a) autogenous (b) autonomous self- healing of concrete.

has been identified as sustainable and eco-friendly con- for autonomous healing of concrete as listed in Table 1
struction practices. This paper is aimed to explore the vari- [12]. The incorporation of microorganisms such as bac-
ous mechanisms used by different bacterial strains for teria or fungi has been explored to obtain bioconcrete
MICP for better understanding of the microbiological with autonomous self-healing in an eco-friendly manner.
aspects and selection of the appropriate strain. The various The specific bacterial strains have been found to be very
factors affecting the self-healing by MICP have been effective as these microorganisms can withstand harsh
reviewed and the applications in cementitious materials environmental conditions and can be introduced directly
have been explored. Further, a critical analysis of the asso- as a biological broth of nutrient medium or by encapsula-
ciated challenges and future prospective of microbial con- tion process involving use of dormant bacterial spores in
crete for sustainable construction has been presented. presence of encapsulating agents [13].

2. Self-healing concrete
The term auto-healing or self-healing essentially refers to a 3. Microbial induced calcium carbonate
material which has the capability to spontaneously heal precipitation (MICP)
small cracks in absence of any external treatment or human Calcium carbonate (CaCO3) is a very important biomin-
involvement [1]. Self-healing can be implemented in an eral existing in three polymorphic forms namely, calcite,
autogenous or autonomous manner. Autogenous healing
aragonite and vaterite [14]. Biomineralization or precipita-
refers to natural healing of concrete that takes place by fill-
tion of various polymorphs of calcium carbonate by sev-
ing of cracks due to deposition of carbonates or hydrates
eral biological species in soil, sea and freshwater is a
formed during the hydration of unhydrated cementitious
natural phenomenon [1]. It generally takes place either in
materials in presence of moisture as shown in Figure 1. This
an induced manner through involvement of microbial
inherent process is responsible for auto-filling of cracks and
metabolic activity or through a mediated pathway involv-
is strongly dependent upon the age, composition and envir-
ing the calcium ions binding with the negatively charged
onmental conditions of concrete [9]. On the other hand, the
functional groups in an alkaline environment [15]. During
autonomous healing is an artificial healing process taking
their metabolic pathways, the vegetative cells generate
place due to presence of a suitable chemical or biological
healing agent in the concrete matrix. Nevertheless, it is carbon dioxide that interacts with calcium ions leading to
understood that both methods of self-healing are only cap- precipitation of calcium carbonate that gets deposited out-
able of repairing cracks within several hundred micro- side the cells via an extracellular [16], intracellular [17] or
meters, which means that structural damage repair is not intercellular [18] modes as shown in Figure 2.
conceivable. The respiratory process of all aerobic bacteria, involv-
The presence of minor cracks in concrete structures ing the urea hydrolysis, oxidation of organic compounds
leads to water ingress that reinitiates the hydration pro- such as calcium lactate, and nitrate reduction, releases car-
cess in the matrix. It results in crack healing due to leach- bon dioxide which leads to the induced biomineralization
ing of calcium hydroxide and deposition of calcium of CaCO3 in the presence of sufficient amounts of water
carbonate in presence of dissolved carbon dioxide in and calcium ions [17]. The process is often known as
water [10]. This process is dependent upon the tempera- microbial induced calcium carbonate precipitation
ture as well as pH of the concrete matrix. The autogenous (MICP) and plays an vital role in building up the earth's
self-healing process has been found to accelerate by the natural segments such as soils, sediments, caves, etc. [19].
participation of the incorporated cemented materials such MICP has been used on similar tracks as promising self-
as nanofillers, fibers, mineral admixtures and other heal- healing tool for cemented structures that are highly vulner-
ing agents in the pozzolanic reaction [11]. On the other able to cracks and can be used either inside the structure
hand, autonomous healing involves the use of the or on the outer surface [12, 20]. The eco-friendly concrete
advanced technologies integrated with non-cementitious so produced is known as bioconcrete with self-healing
materials that can cure micro as well as macro-cracks at ability and leads to saving of recurring maintenance and
room temperature. Many chemical compounds such as restoration cost of the concrete based infrastructures
polymers, crystalline admixtures, fibers have been used including historical monuments [21].
 Journal of Sustainable Cement-Based Materials 319

Table 1. Comparison of autogenous and autonomous self-healing concrete.

Healing Technique Healing material Healing product Advantages Disadvantages
Autogenous self-healing
Use of Fly ash, blast furnace Hydrated CSH gel 1. Reduction in pores 1. High dosage
mineral admixtures slag, lime stone, with filler effect required except
hydrated lime, rice and pozzolanic nano-fillers
husk ash, sugarcane action except 2. Risk of decreased
bagasse ash, super- workability and
silica fume absorbent prolonged
Use of super- Polyacrylate/ Swelled products polymers. setting time
absorbent polymers polyacrylamide 2. Highly effective in 3. High heat of
copolymers very low dosage hydration
Use of nano-fillers Nano-silica, nano- Hydrated metal oxides with 4. Losses effect
alumina, nano-zinc and CSH gel microstructure beyond optimized
oxide, nano-iron development dosage and results
oxide and other nano except in decreased
metal oxides mineral strength
Use of fibers Polyethylene, Hydrated CSH gel and admixtures. 5. Risk of
polypropylene, steel calcium carbonate 3. Lead to early age agglomeration
fibers, carbon fibers strength gain. 6. Healing is
dependent upon
age of the
specimen
7. Suitable for
internal healing
with limited
crack healing
Autonomous self-healing
Use of microvascular Glass fibers, glass Hydrated products 1. Effective in 1. Expensive and
and tubular capsules tubes, polymer tubes macro-crack temperature
and clay healing with dependent
Use of coated granules Epoxy resins Hydrated products increase in 2. Involvement of
Use of shape memory NiTi, NiTiNb Hydrated CSH gel mechanical special techniques
embedded alloys and CuAlNi strength, durability and safety
Use of Bacteria and fungi Calcium carbonate and corrosion protocols in case
Micro-organisms resistance. of micro-
2. Ease of tailoring organisms
as per 3. Loss of
requirement. workability at
3. Can be used for higher dosage
external and 4. Possibility of
internal healing. unwanted
4. Initial cost can be chemical reactions
counterbalanced hindering the
by gain in healing process
service life. 5. Scaling-up
5. Micro-organisms requires
aid to carbon- extensive research.
dioxide
sequestration.

3.1. Mechanisms for MICP performance of concrete structures gets significantly

The process mainly involves the interaction of calcium improved. The assimilative analysis of the existing MICP
ions and the metabolic byproducts in the microbial cell mechanisms in the presence of nutrient medium is highly
matrix, however both intracellular and extracellular pre- desirable. Literature attributes the beneficial role of
cipitation has been reported in the literature. During the microbes in cementitious materials to the two broad path-
intracellular process, the whole bacterial cell becomes ways according to the cement matrix conditions as dis-
encapsulated due to accumulation of calcium carbonate cussed ahead and illustrated in Figure 3.
crystals as observed in case of Achromatium oxaliferum,
the large sulfur bacteria [22], while in the extracellular
course, the mineral vesicles are observed outside bacterial
membrane of species belonging to Bacillus sp.[16]. The 3.1.1. Autotrophic pathway
deposition of calcium carbonate leads to crack repairing This pathway involves either photosynthetic process, with
and provides the self-healing ability to the concrete. consumption of carbon dioxide or oxidation of methane,
Consequently, the mechanical as well as durability in both oxygenic and an-oxygenic environment.
320 R. Garg et al.

Figure 2. Schematic representation of modes for MICP.

Figure 3. Pathways for MICP.

3.1.1.1. Photosynthetic pathway (Equation (1)) that diffuse across the cellular membrane
It involves the assimilation of atmospheric carbon dioxide [25].
by autotrophic microbes including algae and cyanobac- CO2 þ H2 O $ H2 CO3 $ H þ þ HCO3  (1)
teria, through common photosynthetic pathways involving
the active involvement of ATPase leading to an exchange The enzyme, carbonic anhydrase (CA) catalyzes the
of Ca2þ and Hþ ions [23]. Both intracellular as well as dissociation of bicarbonate ions into hydroxide ions and
extracellular precipitation has been reported, but extracel- carbon dioxide (Equation (2)) leading to generation of an
lular process has been considered the prime process [24]. alkaline environment [26].
Fresh water Cyanobacteria such as Synechococcus leopo- HCO3  $ CO2 þ OH  (2)
liensis has been found to result in increased microbial pre-
The interaction of the bicarbonate ions on the extracel-
cipitation of calcite at extracellular level while the alkaline
lular surface with excess of calcium ions contributes to
lake cyanobacteria, such as Candidatus Gloeomargarita
the precipitation of calcium carbonate (Equation (3)) [23].
lithophora and Thermosynechococcus elongates, can live
in poor nutritional media by using carbon dioxide as a Ca2þ þ HCO3  ! H þ þ CaCO3 (3)
source of carbon and can induce intracellular precipitation
of calcium carbonate [17].
Bacteria convert carbon dioxide into glucose via the 3.1.1.2. Methane oxidation
photosynthetic cycle that is again converted to carbon Aerobic and anaerobic oxidation of methane by methane
dioxide in a number of steps. This carbon dioxide dis- oxidizing bacteria such as Methylocystis parvus in fresh-
solves in presence of water molecules in the cement water as well as marine ecosystems, also results in the pre-
matrix and produces bicarbonate ion along with Hþ ions cipitation of calcium carbonate [27].
 Journal of Sustainable Cement-Based Materials 321

The aerobic oxidation of methane proceeds through pasteurii [29], Bacillus megaterium [30], Bacillus firmus
the formation of methanol (Equation (4)) catalyzed by and Bacillus sphaericus [16] etc. in addition to
methane mono-oxygenase in the cellular periplasm [19]. Escherichia coli [31], Protius mirabilis[1] and ACRN
Methane monooxygenase [32]. Hydrolysis of urea by bacterial urease and conver-
CH4 þ O2 ! CH3 OH þ H2 O (4) sion of carbon dioxide to carbonate ions by carbonic anhy-
NADH
drase are the two synergistic processes that are mainly
The subsequent enzymatic processes convert methanol
to formaldehyde, formate and ultimately to carbon dioxide dependent upon the pH, concentration of carbonate and
in presence of methanol dehydrogenase and formate calcium ions in the cellular environment [25, 30].
dehydrogenase (Equations (5)–(7)). Literature reports decrease in substrate affinity and activ-
ity of urease with decrease in pH [33] while immobiliza-
Methanol dehydrogenase
CH3 OH 
! HCHO (5) tion of enzyme has also been found to decrease the
PQQ
reaction rate [31].
Formate dehydrogenase
! HCOO þ H þ
HCHO þ H2 Oþ (6) The mechanism can be represented in a series of steps.
NAD
 The hydrolysis of urea first creates carbamate and ammo-
HCOO þ H2 O ! CO2 þ OH (7)
nia by the enzyme urease (Equation (12)).
Carbon dioxide further gets converted to bicarbonate
NH2 CONH2 þ H2 O ! NH2 COOH þ NH3 (12)
ions leading to calcite precipitation in presence of the
alkaline calcium rich environment (Equation (8)). The subsequent hydrolysis of carbamate (Equation
Ca 2þ 
þ CO2 þ OH ! CaCO3 þ H2 O (8) (13)) produces additional ammonia along with carbonic
acid [34].
In the presence of sulfate ions, oxidation of methane
by anaerobic methanotrophic bacteria produces bicarbon- NH2 COOH þ H2 O ! H2 CO3 þ NH3 (13)
ate ions and bisulfite ions (Equation (9)) [28]. In another carbonic anhydrase driven hydrolysis
  (Equations (14) and (15)), ammonia reacts with water to
CH4 þ SO4 2
! HCO3 þ HS þ H2 O (9)
produce hydroxide ions which convert carbonic acid to
The subsequent reaction of the bicarbonate ions with
carbonate ions [35].
sufficient calcium ions produces calcium carbonate.
NH3 þ H2 O ! NH4 þ þ OH  (14)
3.1.2. Heterotrophic pathway H2 CO3 þ 2OH  ! 2H2 O þ CO3 2 (15)
This pathway involves production of bicarbonate ions
These reactions result in increased pH around the bac-
through metabolic assimilation of sulfur or nitro-
terial cell and the bacterial surface gets negatively
gen compounds.
charged, thereby attracting the positively charged calcium
ions acting as nucleation site for calcium carbonate pre-
3.1.2.1. Sulfate reduction cipitation [1]. The active transport and accumulation of
Calcium carbonate may be precipitated by sulfate reduc- calcium ions in the cell takes place along with expulsion
ing bacteria such as Desulfovibrio vulgaris in the presence of protons from the cell. The resulting high concentration
of an oxygen deficient environment rich in organic matter of calcium ions with low concentration of protons induces
such as hypersaline lagoons, geological sediments and supersaturation of carbonate ions in the cellular environ-
hypersaline lakes [23]. The process is initiated by gypsum ment and facilitates the precipitation of calcium carbonate
dissolution (Equation (10)) releasing the calcium ions and [26]. Various polymorphs such as calcite, vaterite and ara-
creating a sulfate-rich atmosphere in the microenviron- gonite have been observed depending upon the amino
ment [15]. acid composition in the microenvironment [36].
ƒ!H2 OCa2þ þ SO4 2
CaSO4 ƒƒ (10)
Bacterial reduction of sulfate produces hydrogen sul- 3.1.2.3. Nitrate reduction
fide and bicarbonate ions in the presence of organic matter Anaerobic denitrifying bacteria such as Pseudomonas aer-
(Equation (11)). uginosa and Diaphorobacter nitroreducens convert nitrate
2CH2 O þ SO4 2 ! 2HCO3  þ H2 S (11) to nitrogen gas and produce carbon dioxide during meta-
bolic oxidation of organic matter (Equation (16)) resulting
The degasification of hydrogen sulfide and substantial in increased pH of the system due to H þ ion uptake from
increase in pH facilitates the precipitation of calcium car- the system [37].
bonate as dolomite, aragonite or calcite depending upon
the bacterial strains [23]. 5HCOO þ 2NO3  þ 7H þ ! 5CO2 þ 8H2 O þ N2 (16)
Owing to the insufficient oxygen supply, nitrate reduc-
3.1.2.2. Degradation of urea tion not only results in CaCO3 precipitation but also pro-
The enzymatic hydrolysis of urea aids in the precipitation duces carbon dioxide. The carbon dioxide that is produced
of calcium carbonate in the microbial environment of also interacts with calcium hydroxide and increases the
many bacterial species of Bacillus including Bacillus amount of calcium carbonate.
322 R. Garg et al.

3.1.2.4. Metabolic degradation of decreases the workability and rheological performance of

organic compounds concrete [51]. Previous studies indicate a significant
The self- healing properties of concrete can also be impact of the ingredients, media, method, speed and dur-
strengthened by using heterotrophic aerobic bacteria such ation of mixing upon the self-healing by MICP. Mainly,
as Bacillus cohnii and Bacillus pseudofirmus. This process use of protecting agent and nutrient medium in addition to
involves the oxidation of organic compounds by aerobic specific bacteria has been identified in the literature as dis-
bacteria leading to calcite precipitation and production of cussed ahead.
carbon dioxide (Equation (17)).
CaC6 H10 O6 þ 6O2 ! CaCO3 þ 5CO2 þ 5H2 O (17) 4.1. Use of specific bacteria
Selection of suitable bacteria with efficient survival rate
This pathway can be used to solve problems such as and suitable metabolic activity is highly critical while
excess ammonia production during the ureolytic process, designing bio-concrete. Due to short lifetime and patho-
because the bacteria directly use the organic materials and genicity of many bacteria, their suitability as self-healing
CaCO3 is produced in their metabolic activity. This meta- agents is limited [20]. Some non-spore forming bacteria
bolic process is more important as the precipitation of belonging to Shewanella species have been reported to
CaCO3 increases when the CO2 generated during the bac- survive only for 6–7 days in the cement matrix as the
terial respiration interacts with calcium hydroxide gener- growing calcium carbonate precipitation in pores checks
ated by hydration of the cement (Equation (18)). the nutrient flow [52]. Some ureolytic bacteria, such as
CaðOH Þ2 þ CO2 ! CaCO3 þ H2 O (18) Bacillus licheniformis, Bacillus flexus, and Bacillus
megoaterium can resist alkaline medium and can live
without a nutrient for hundreds of years due to the exten-
4. Designing of bioconcrete sive dormant period of their spores. These bacteria have
The concept of exploring MICP for the surface treatment been reported to almost completely heal the cracks leading
of natural stone was first introduced in 1990 [38] that was to significant improvement in strength of the microbial
explored later on as bioconcrete or microbial concrete. concrete [53]. In addition, ureolytic bacteria isolated from
Microbial concrete is typically obtained by adding the marble sludge has been used to study the properties of
viable yet dormant bacteria to the usual concrete compos- concrete made from rice husk ash [54].
ition in the presence of a suitable nutrient medium such as A number of urea hydrolyzing bacteria, such as
calcium lactate [39], yeast [40], urea [41, 42], proteins Sporosarcina ureae, Proteus vulgaris, Proteus mirabilis,
[43] etc. The metabolism of dormant bacteria gets acti- Bacillus sphaericus, Bacillus megaterium, and Bacillus
vated by the water, entering freshly formed cracks, which Subtilis, have been explored to exhibit a strong positive
results in CaCO3 precipitation in addition to other impact on crack filling and strength of self-healing con-
hydrated products that fills the pores as well as the micro- crete. While ureolytic bacteria may dramatically improve
cracks. This improves the microstructure of the cement the crack healing and strength of various types of con-
matrix and improves the macro properties of the cementi- crete, urea production produces a large quantity of carbon
tious materials [44].The microbial precipitation inside the dioxide in the ureolytic phase followed by production of
concrete is generally carried out by introducing specified ammonia with a pungent odor. These issues can be over-
doze of suitable calcite precipitating bacteria in the mix come by using Bacillus pseudofirmus, and Bacillus cohnii,
design containing nutrient medium [45]. Usually, the mix the spore-forming and alkaliphilic bacteria that undergo
proportions of the bioconcrete are determined using the metabolic conversion of organic compounds such as cal-
conventional mix design technique. Literature reports cium lactate for precipitation of calcium carbonate [13].
optimal bacterial concentration varying from 105–109 These bacteria also show improved response in presence
cells/mL. The biological mixture of bacteria and nutrient of various calcium sources, such as calcium chloride, cal-
medium can also be applied directly on the cracked sur- cium nitrate and calcium acetate and oxide. Bacillus subti-
face so as to induce autonomous healing by calcite pre- lis can consume free calcium oxide in the concrete to
cipitation in the cracks [46]. Furthermore, reinforcement ensure precipitation of calcium carbonate in absence of
in the form of fibers, metal wires, or steel bars can be any outer calcium sources at an optimal bacterial concen-
used in bioconcrete to reduce the width of cracks and pre- tration [20]. Researchers have also used Diaphorobacter
nitroreducens as an active and compact denitrifying core
vent premature failure due to crack formation [20].
to have marginal effects on setting time opposed to
Mineral admixtures are also used as partial substituents of
Bacillus sphaericus that results in significant delayed set-
cement in order to reduce the use of cement in bioconcrete
ting time [48].
[47]. Depending on the situation, the fresh properties as
well as the mechanical properties of concrete get modified
significantly [48]. The incorporation of bacterial strains in 4.2. Protecting agent
concrete has been reported to have negligible impact on Self-healing tendency of bacterial concrete is also depend-
workability [49, 50]. However, the use of nanomaterials ent upon the high metabolic activity of bacteria. Hence,
or materials with swelling behavior in combination with suitable encapsulation treatment is essential for protection
bacterial strain contributes to water adsorption and of the bacteria from the detrimental environmental factors
 Journal of Sustainable Cement-Based Materials 323

including high pH of cement matrix. The protection of reported as an excellent nutrient medium for growth of
bacterial cells in the aggressive cement matrix is ensured Bacillus Subtilis resulting in complete self-healing of con-
through incorporation of suitable encapsulating agents crete at 44 days [3]. Improved cement mortar and sand-
such as silica, polyurethane [55] and calcium sulphoalu- stone strength has been achieved through MICP by
minates [56]. In presence of protective materials such as Bacillus cereus using tofu wastewater as a nutrient
diatomaceous earth, extended clay, granules that encapsu- medium [64].
late bacterial spores, the bacteria can survive in concrete
even under the harsh conditions [57]. The most common 4.4. Mixing techniques
protecting agents include activated carbon, hydrogel, silica
The bacterial strains can be introduced via direct method
gel, polyurethane, diatomaceous earth, expanded particles
or indirect method. In the direct method, the bacterial
such as clay, shale aggregates, and perlite [55]. Better
strains in the active or dormant stage are added along with
results have been obtained for the performance of bacterial
concrete in presence of protecting agents [58]. the nutrient medium while in the indirect method, encap-
Microcapsules of melamine have also been used for sulated bacterial strains are used [3]. Depending upon the
encapsulation of Bacillus sphaericus spores [41]. type of the bacterial strain, it can be added either during
Immobilization of Bacillus sphaericus by silica gel and dry mixing [20], wet mixing [29] or at the final stage of
polyurethane was also found to extend the ureolytic activ- mixing [65]. However, the medium deposition offers bet-
ity and precipitation of calcium carbonate. Polyurethane ter results in terms of crack repair and water absorption in
has been found to impart better strength and decreased the case of microbial concrete [46]. An uniform distribu-
water permeability as compared to silica gel [55]. tion of healing constituents is required so as to achieve
Encapsulation of Bacillus pseudofirmus by hydrogel has sufficient self-healing. Hence it is very important to follow
been found to almost completely heal the cracks lesser correct methods of mixing and dispersion of the constitu-
than width of 1 mm [59]. Use of expanded perlite or ents. The most common method is adding raw materials
expanded clay has been reported to heal the cracks up to in several batches and mixing with a mechanical mixer
0.79 mm depth at 28 days of curing and reduced the water until a homogenized mixture is obtained [55]. Additional
permeability by 26% [2]. physical procedures such as ultrasonic treatment and
chemical treatments such as the incorporation of super-
plasticizers or water-reducing admixtures can also be used
4.3. Nutrient medium
in addition to machine-driven mixing [51]. The moulding
Selecting appropriate calcium/nitrogen sources for bacter- of self-healing concrete is the same as for regular con-
ial metabolism in the dry and basic concrete is of consid- crete, as the mix is transferred to moulds in layers using
erable importance [60]. Diaphorobacter nitroreducens is mechanical vibrations in order to compress the compo-
the typical denitrifying bacteria used in self- healing con- nents [47].
crete due to reduction of nitrates resulting in the precipita-
tion of CaCO3. These nitrate reducing bacteria have also
been reported to exhibit better results in terms of survival 4.5. Type of curing
and strength of mortars in presence of protecting agents In both autonomous and autogenous self-healing cases,
such as granular activated carbon, expanded clay and dia- cure conditions are preserved during the initial stages and
tomaceous earth. The use of encapsulation effect in com- the self-healing stages. Besides cracking width and age of
bination of calcium acetate and calcium nitrate was found curing, the curing method also has a major effect on
to provide better results with no effect on setting time. microbial concrete's self-healing effect with best results in
The use of organic medium such as yeast was reported to presence of water. Curing in still water is advantageous
retard the setting in case of ureolytic bacteria [48]. The compared to running water, as the flowing water drains
effect of calcium sources such as calcium formate, cal- out the hydroxide and calcium ions, reducing the pH val-
cium nitrate and calcium lactate has also been reported on ues and calcium ion levels that are essential for the devel-
the compressive strength and hydration kinetics of bacter- opment of healing products [66]. In case of mortars
ial concrete using spore-forming and alkali-resistant bac- containing mineral admixtures, still water healing results
teria [61]. Calcium lactate results in delayed hydration in a faster reduction of the permeability coefficient and
while the other two sources accelerate the process. the crack width [67]. Properly ordered and sequential
However, these calcium sources have been found to water curing followed by air exposure has been found to
improve the rheology of bacterial cement matrix. Calcium result in improved self-healing efficiency possibly due to
lactate and calcium formate result in increased compres- the fact that excess water evaporation during air exposure
sive strength but calcium nitrate decreases the compres- increases the ionic concentration in the minor cracks
sive strength and is hence, not suggested as a suitable which facilitates the formation of varied gel and expansion
calcium source for bacterial growth [61]. Formate has also products [66].
been reported to accelerate the setting and considered
essential in MICP [62]. In presence of calcium acetate and
calcium chloride as a calcium source, an improvements in 5. Applications of MICP in cementitious materials
the efficiency of Bacillus sphaericus in crack healing has The basic application of these MICP processes to micro-
been reported [63]. Peptone and yeast extract has been bial concrete often addresses some critical issues
Table 2. Effect of MICP in presence of various nutrient media on the cementitious materials.
324

Material Bacteria Nutrient medium Effect on material References

Photosynthetic pathway
Mortar Synechococcus sp. PCC 7002 Gelatin-low salt media Increased compressive and flexural strength with high [70]
regenerative ability
Sulfate reduction
Stone Desulfovibrio vulgaris subsp. Vulgaris Desulfovibrio DSMZ 63 medium Black crust removal [83]
ATCC 29579
Concrete SRB Nutrient broth and Increased compressive strength (13%) and decreased [76]
magnesium sulfate water absorption (8.5%)
Concrete SRB Nutrient broth and Increased compressive strength (60.87%), tensile [76]
sodium chloride strength (48.34%) and flexural strength (52.30%)
Degradation of urea
Sand Column Bacillus sphaericus MCP-11 Yeast extract, ammonia sulfate Uniform cementation and increased strength [89]
and nickel (II) chloride
Concrete Sporosarcina pasteurii (NCIM 2477) Yeast extract, urea, and lactose Increased Compressive strength (17%) and crack [90]
mother liquor healing (24.0-28.4% of the total sand weight)
Concrete Sporosarcina pasteurii (NCIM 2477) Nutrient broth, urea, and corn Increased Compressive strength (35%) and crack [91]
steep liquor healing (26.33-30.12% of the total sand weight)
Concrete ACRN Yeast extract, urea Increased Compressive strength (21.92%) [92]
Concrete Immobilized microcapsules of Bacillus Minimal basal salt medium Crack width healing (970 mm) with 40% reduction in [41]
sphaericus LMG 22557 water permeability with reduction in
compressive strength
Concrete Encapsulated Sporosarcina pasteurii Peptone, Beef extract, urea, and Increased compressive strength (84%), decreased water [56]
ATCC 11859 manganese sulfate of absorption (50%) and crack healing upto 417 mm
R. Garg et al.

Cement paste Bacillus mucilaginous Brewers yeast, nutrient, glucose, Increased flexural strength (56-72%) with crack healing [71]
and calcium nitrate area of 87.5%
Cement paste Soil Bacteria Lentil seed powder, sugar, Increased compressive strength (23.49%), reduced water [35]
gypsum, beef extract, and urea absorption (15.40%)
Cement mortar Bacillus cereus Tofu water and Nutrient Broth Increased compressive strength (27.8%) [64]
Cement Mortar Encapsulated Bacillus sphaericus in Urea, yeast extract, calcium Increased setting time and decreased compressive [48]
presence of granular activated carbon, nitrate and calcium acetate strength (68%)
zeolite, diatomaceous earth and
Activated Compact Denitrifying Core
Nitrate reduction
Cement Mortar Encapsulated Diaphorobacter Urea, yeast extract, calcium Negligible effect on setting time and [48]
nitroreducens in presence of granular nitrate and calcium acetate compressive strength
activated carbon, zeolite, diatomaceous
earth, and Activated Compact
Denitrifying Core
Concrete Encapsulated Pseudomonas aeruginosa in Urea, yeast extract, calcium Decreased water absorption (42%) with crack healing [74]
presence of expanded clay particles formate and calcium nitrate upto 200 mm
Concrete Encapsulated Diaphorobacter Urea, yeast extract, calcium Decreased water absorption (47%) with crack healing [74]
nitroreducens in presence of expanded formate and calcium nitrate upto 250 mm
clay particles
 Journal of Sustainable Cement-Based Materials 325

providing potential advantages. It helps in matrix develop- the crack healing upto a width of 0.46 mm at 100 days of
ment with formation of calcite as the healing product that curing indicating the viability and functionality of the
results in filling of pores and cracks thereby significantly embedded soil-isolated bacteria due to protection by clay
affecting the structural, strength and durability characteris- particles [75]. Researchers have also used bacterial spores
tics of the concrete. The oxidation process absorbs the dis- embedded in calcium alginate beads in the presence of an
solved oxygen in the embedded water so that the concrete adequate nutrient medium for the self-healing of concrete
structure can sustain corrosion [68], and increases the structures in marine environment, since standard methods
resistance of steel reinforcement [60]. The effect of MICP are not successful in the icy climatic areas [68]. Recently,
on crack healing, strength, durability and corrosion resist- Sporosarcina pasteurii and Bacillus sphaericus have been
ance of cemenetitious materials has been shown in Table used to heal thermal cracks with clearly visible surface as
2 and discussed ahead. evident in [42].

5. 1.Repairing of cracks 5.2. Strength enhancement

Autotrophic bacteria can be used very effectively in off- The calcium carbonate formed by the MICP in the cemen-
shore environments with exposure to carbon dioxide and titious samples increases the strength by improving the
sunlight for repairing fine pores and restoring concrete inter-particle bonding and decreasing the porosity by
structures [69]. Recently, MICP tendency of effectively filling the matrix pores and cracks. The impact
Synechococcus sp. PCC 7002 have been explored to of varying concentration of bacterial strain, Sporoscarcina
obtain living building materials with self-sustaining func- Pasteurii has been investigated on the water absorption,
tions. These materials were found to possess significantly compressive strength, and rapid chloride permeability of
higher fracture energy as compared to conventional mate- concrete in absence and presence of fly ash and silica
rials with good viability and regenerative potential of the fume. Best performance was observed at an optimum con-
bacterial species [70]. Bacillus mucilaginous is another centration of bacterial strain (105 cell/mL) [47]. The com-
aerobic heterotrophic and denitrifying bacterium that have pressive strength of the bacterial (Sporosarcina pasteurii)
a strong ability to heal cracks at early-age of concrete. It incorporated shotcrete specimens with coarse aggregates
produces carbonic anhydrase enzyme that assimilates presoaked in the bacterial solution (urea-CaCl2 as nutrient
atmospheric carbon dioxide into carbonate ions. The bac- medium) was 26% higher compared to control specimens
teria have been reported to help in healing of cracks by at 150 days [72]. On the other hand, an optimized dosage
almost 87.5% after 28 days [71]. Sporosarcina pasteurii, of Bacillus pasteurii (107 cell/mL) in presence of rice
the gram-positive bacterium with a high spore- forming husk ash has been reported to result in compressive
potential, has been used to repair the cracks and enhance strength enhancement of self-compacting concrete by
properties of light weight aggregate concrete under curing 21% [49]. In comparison to control specimen, 30%
by urea-CaCl2 solution [72]. improved strength of the bacterial cement mortar has been
By using Bacillus cohnii in the presence of suitable recorded by using effluent from chicken manure bio-gas
nutrient medium, calcite gains of size varying from plant as a cost-effective nutrient medium for Bacillus pas-
20–80 lm were found to be precipitated on broken con- teuri [43]. A significant increase in compressive strength
crete surfaces [13]. Immobilization of Bacillus sphaericus of material containing Bacillus mucilaginous has been
by diatomaceous earth resulted in substantially higher ure- observed with appearance of clearly visible white powder
olytic activity leading to filling of cracks in mortar up to a in the healed cracks that was identified as calcite in crys-
width of 0.17 mm at bacterial concentration of 109 cells/ tallographic analysis [71]. Bacillus pseudofirmus, in pres-
mL in presence of a deposition medium. The quantity of ence of calcium alginate and chitosan as the nutrient
decomposed urea was found to increase with increasing medium, have been reported to significantly impact the
concentration of diatomaceous earth with an optimal con- compressive and flexural tensile strength of bacterial con-
centration as 60% (weight/volume) of the bacterial sus- crete with an increase of 10.28% and 13.79%, respectively
pension. Further, calcite grains of size in the range of [59]. Domestic acidic water has been used as a source to
5–25 lm were obtained during the use of Bacillus sphaeri-
extract sulfate reducing bacteria to significantly enhance,
cus for crack healing [73]. In addition, Crack healing by
the compressive, tensile and flexural strength of the bio-
calcite grains of size up to 100 mm depth has been reported
concrete with significant decline in water permeability
in presence of silica gel as a bacteria (Bacillus sphaericus)
[76, 77].
protective material [55]. On the other hand, a crack heal-
ing up to the width of 970 lm has been observed in mor-
tars containing microencapsulated spores of Bacillus 5.3. Durability enhancement
sphaericus. Further, use of wet-dry cycles was found to Concrete degradation and deterioration leading to decline
promote the self-healing of specimens indicating the in its service life is a great matter of concern. The princi-
requirement of water during the cycle [41]. pal degradation factors impacting the durability of cemen-
Effective crack closure has also been achieved by titious materials include seasonal fluctuations, freeze-thaw
using Diaphorobacter nitroreducens for healing of cracks cycles, relative humidity, and chemicals in the environ-
up to 350 mm in 28 days [74]. On the other hand, use of ment. Researchers have obtained a significant enhance-
expanded clay in presence of calcium lactate resulted in ment in durability of cementitious materials with self-
326 R. Garg et al.

healing and reduction of pores and/or cracks as a result of [79]. Water absorption in the crashed mortar containing
MICP. Precipitation of calcite in pores and cracks signifi- Diaphorobacter nitroreducens has been observed to
cantly reduces the permeability of the cementitious struc- decrease by 50% compared to the reference sample [74].
tures as evident from the comparative results reported for A highly alkaliphilic and novel facultative anaerobic
conventional and bacterial concrete. A significant strain of BKH4 bacteria isolated from hot springs has
decrease in the gas permeability and water absorption of been reported to increase the durability of mortars with
mortar and concrete has been reported after the surface significant increase in chloride and sulfate resistance at an
treatment by Bacillus sphaericus cultures due to calcium optimized concentration of 104 cell/mL [80].
carbonate precipitation [46]. Precipitation of flower
shaped grains of calcite with size of 15–20 mm in cracks 5.4. Corrosion resistance
increased the resistance to water penetration of mortar Methanotrophs use methane and sulfate ions in their meta-
samples containing diatomaceous earth immobilized bolic pathways and hence, can be used to induce a protect-
Bacillus sphaericus by 30% as compared to conventional ive coating against corrosion of metallic sheets. A recent
specimens [73]. Likewise, incorporation of Bacillus aerius research on Methylocystis parvus, a methanotroph has
(105 cells/mL) in RHA concrete resulted in decreased por- shown the possibility to precipitate calcite in the presence
osity, permeability, and water absorption and enhanced of calcium formate [19]. Hydrogen sulfide, is a corrosive
density of the matrix at all the examined ages, owing to gas which can cause biological corrosion on the internal
filling of pores and cracks by calcite precipitation [54]. and external surfaces of concrete structures in presence of
The ureolytic Bacillus megaterium and Bacillus pas- oxygen leading to decreased efficiency of the process with
teurii along with non-ureolytic Bacillu. Cohnii incorpo- undesirable outcomes [81]. The anoxygenogenic and
rated concrete have been observed to show reduction in phototrophic Desulfovibrio vulgaris have been reported to
concentration of sulfate ions by 21%, 26% and 32%, convert hydrogen sulfide to elemental sulfur through con-
respectively due to filling of pores due to MICP [44]. sumption of gypsum [82]. The bacterial species have been
Introduction of Bacillus pasteurii at a maximal concentra- used to remove black crusts deposited upon historical
tion of 107 cell/mL in rice husk ash (RHA) based self- stones along with calcite precipitation [83]. The process
compacting concrete has been reported to decline the can be represented as shown in Equation (19) [15]:
water permeability by 80% owing to the pore reduction
with pozzolanic action of RHA and calcite production by 6CaSO4 þ 6CO2 þ 4H2 O ! 6CaCO3
(19)
the bacteria [49]. On the other hand, use of calcium acet- þ 4H2 S þ 2S þ 11O2
ate as a calcium source has been found to exhibit better Furthermore, this type of healing agent can protect
results in terms of more pronounced reduction in pore size steel reinforcement against corrosion, acting as an oxygen
and permeability of microbial mortars incorporated with diffusion barrier, because aerobic bacteria consume oxygen
wild strain of Sporosarcina pasteurii [60]. The durability in the metabolic conversion pathway [75]. The nitrate
of light weight aggregate has been also found to increase reducing bacteria namely Diaphorobacter nitroreducens,
with a reduction in the water absorption, porosity, and per- and Pseudomonas aeruginosa encapsulated with granular
meability due to use of bacterial (Sporosarcina pasteurii) activated carbon, diatomaceous earth, and expanded clay
curing solution [72]. The optimal dosage of Sporosarcina have been found to exhibit better survival and inhibit the
pasteurii (105 cells/mL) in the silica-fume based concrete corrosion of steel at controlled pH conditions [62].
demonstrated enhanced durability with excellent resist- Reinforced concrete specimens containing Sporosarcina
ance to chloride penetration after 91 days of curing [47]. pasteurii and cured in the urea–calcium lactate medium
Zeolite-immobilized bacteria, Sposarcina ureae and showed improved corrosion resistance with increased elec-
Sporosarcina pasteurii introduced mortar composites trical resistance than compared to concrete specimens with-
have exhibited decreased sorptivity and rapid chloride per- out steel reinforcement as well those containing Bacillus
meability with pronounced self-healing efficiency in four subtilis. An increase of calcite sediments inside concrete
months with better performance of the specimens with pores was credited for both a reduction in porosity and an
Sporosarcina pasteurii due to more calcite production and increase in conductivity [84]. Researchers found that a new
better crack healing [78]. bio-concrete made from anaerobic granular sludge showed
Introduction of Bacillus subtilis at a concentration of great promise in reducing sewer corrosion. The sulfate
107 cell/mL has been found to result in a decreases of reducing bacteria in the sludge promoted the sulfur cycle
chloride penetration and carbonation depth of light weight between aerobic and anaerobic sub-layers of the corrosion
aggregate concrete by about 20.5% and 27.2%, respect- leading to a net reduction in biogenic sulfuric acid gener-
ively. Water absorption of light weight concrete reinforced ation. Consequently, corrosion rate was almost decreased
with steel fibers was reduced by 13.1% and the results to half as compared in conventional samples [85].
were attributed to the sealing of fine cracks by the calcite
precipitation by the bacteria [20]. Samples cultured in
media containing an alkali tolerant bacteria, Bacillus cer- 6. Drawbacks of microbial concrete
eus were reported to have decreased chloride permeability Implementation of biotechnology in construction sector is
and water absorption by 10.9% and 12.0%, respectively associated with various risks and safety as well as eco-
with healing of cracks having a width of 100–800 mm nomic concerns with requirement of specialized
 Journal of Sustainable Cement-Based Materials 327

techniques. The pH of the cement matrix, the quantifica- concrete can reduce the maintenance requirement by
tion of dissolved inorganic carbon and the accessibility of increasing the efficiency of cement structures resulting in
nucleation sites are the three major factors regulating cal- decreased cement consumption, thereby reducing carbon
cium bio-mineralization [59]. In addition, environmental dioxide emission rates [60]. In addition, its role in seques-
parameters such as temperature and atmospheric pressure tration of carbon dioxide through the enzyme catalyzed
of carbon dioxide influence the amount of the dissolved biochemical process adds to its eco-suitability. However,
inorganic carbon [86]. Selection of microbial concrete implementation of these dynamic microbial concrete tech-
bacteria based on MICP may require additional protection niques in a variety of harsh environments involves strat-
for a specific environment, because bacteria may not be egies and additional realistic application costs. Hence,
adapted in that area [59]. Further the information regard- numerical simulation and analysis for reduction of experi-
ing optimized content of bacterial cells and auxiliary mental cost is highly desirable [88]. Mechanisms for con-
materials is required so as to obtain concrete with best per- crete self-healing can be used in a particular environment
formance. However, literature reports varying information to ensure that these basic requirements are preserved and
regarding the optimization of microbial concrete. Bacillus a more sophisticated bacterial selection is carried out. A
pseudofirmus have a survival time of up to 4 months in description of the bacterial knowledge according to the
cement matrix, subjected to the pore diameter. As the various MICP mechanisms and their correct environmen-
pores get filled and the diameter shortens than 1 lm, the tal conditions is therefore required for this understanding.
approximate size of bacterial spores, the survival is limited Further, enhancement of bacterial activity may increase
[59]. A certain adverse impact on compressive strength
the service length of the microbial concrete structures. A
has also been found in some cases [87]. Owing to
natural MICP system by selecting the appropriate bacterial
increased pores in the matrix by a higher blending speed
strain in simulation with a given environment of various
and longer blending time, the fluidity and strength of
harsh situations can be considered. Use of MICP in a
cement composites are normally decreased [86].
more eco-friendly way through specific enzymes such as
Urea is the most common healing medium used in
urease instead of complete bacteria will render out the
microbial concrete in the presence of different nutrient
prospective drawbacks [57]. The incorporation of support-
sources. However, the release of two moles of ammonium
ions per one mole of carbamate molecule may lead to an ive cementitious materials can further prove a boon for
increased nitrogen load resulting in major environmental bacterial concrete and compensate the overall cost by the
issues [13]. Implementation of specific techniques for incurring benefits. Additionally, standard tests and proto-
treatment of the released ammonia and ammonium ions to cols are required for scaling up the use of microbial con-
avoid contact with the surrounding environment aids to crete in infrastructural applications.
surplus cost. Even then, these healing agents have been
extensively used due to better results.
The direct and indirect exposure to bacterial species 8. Conclusion
during fabrication of the bioconcrete may negatively This paper reviews the mechanisms and applications of
impact the health of the working staff as well as the sur- microbial induced calcite precipitation for self-healing of
rounding flora and fauna [57]. Proteus mirabilis and concrete. The cementitious structures when exposed to
Proteus vulgaris contain ammonia-binding urease enzyme surrounding aggressive environments develop cracks that
yet their suitability is limited as firstly the concrete pH is can severely impact their performance and service life.
higher than the optimal bacterial pH and secondly, these The use of MICP is a very effective tool that can be tail-
bacteria can cause the infection of the urinary tract and ored to promote self-healing of concrete by incorporation
wounds, hence a careful handling is required [1].
of suitable encapsulating and protective agents. Moreover,
Although, the efficient crack healing and corrosion pre-
it is shown that, together with self-healing, a number of
vention may also play a crucial role in the increased ser-
additional advantages can also be achieved by different
vice life of concrete structures, yet the accumulation of
MICP mechanisms including the enhancement in mechan-
toxic nitrogen oxides decreases the efficiency of the pro-
ical and durability properties of concrete. The use of suit-
cess. Selection of non-pathogenic and indigenous microor-
able and economical techniques may offset the
ganisms with due consideration of biosafety protocols
unavoidable cost related to nutrients, bacterial culture and
may complicate the procedures. Hence, extensive research
is required to confirm the fortunate gains of microbial handling. Appropriate bacteria must be selected with a
concrete in an eco-friendly and safer way. longer activity period so as to compensate the cost of
microbial concrete development with the incurred bene-
fits. In addition, the use of supportive cementitious materi-
7. Future aspects als in microbial concrete can give better results from an
The high-temperature calcination process during cement environmentally sustainable and economic point of view.
production is associated with accumulated emissions of Thus, further research in this direction can promote the
carbon dioxide that accounts for nearly 10% of global car- microbial concrete as a viable alternative to deal with the
bon dioxide emissions, and the ever-increasing global current high maintenance cost of various cementi-
trend poses a major environmental risk [13]. Microbial tious structures.
328 R. Garg et al.

Competing interests exopolysaccharides and capsular polysaccharides.

Microsc Microanal. 2007;13(1):42–50.
There are no relevant financial or non-financial competing [17] Benzerara K, Skouri-Panet F, Li J, et al. Intracellular
interests to report. Ca-carbonate biomineralization is widespread in cyano-
bacteria. Proc Natl Acad Sci USA. 2014;111(30):
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No potential conflict of interest was reported by and the problem of the vital effect. Rev Mineral
Geochem. 2003;54(1):1–29.
the authors.
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oxidation-driven calcium carbonate precipitation by
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Funding 2014;80(15):4659–4667.
The authors declare no specific funding for this work. [20] Salmasi F, Mostofinejad D. Investigating the effects of
bacterial activity on compressive strength and durability
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