Review Article

Harnessing microbes for self-healing concrete – A review

Farzin Asgharpour*,1,3,a, Kadir Çakiral2,3,b, Khaled Hamed Marar1,c

1Dept. of Civil Eng., Eastern Mediterranean University, Famagusta, North Cyprus Via Mersin 10, 99628
2Dept. of Chemistry, Eastern Mediterranean University, North Cyprus Via Mersin 10, 99628, Turkey
3Faculty of Medicine, Eastern Mediterranean University, North Cyprus Via Mersin 10, 99628, Turkey

Article Info Abstract

Article history: This review explores the burgeoning field of microbially enhanced construction
materials, with a special focus on self-healing concrete, through the lens of
Received 04 Mar 2024 microbial biotechnology. Central to this discourse is the innovative use of
bacteria, particularly Bacillus species, to address the pervasive issue of
Accepted 10 Apr 2024
microcracks in concrete, a fundamental material in the construction industry.
Traditional remedies, such as chemical admixtures and fiber reinforcements,
Keywords: offer partial solutions; however, self-healing concrete represents a paradigm
shift, harnessing the natural calcite-precipitating ability of bacteria to
Bacillus sp; autonomously repair cracks, thereby augmenting structural durability and
Bio-mediated repair; longevity. Delving into the mechanics, the bacteria, embedded within the
Microbial concrete; concrete matrix, remain dormant until crack formation triggers their metabolic
Self-healing concrete; pathways, leading to calcite production that effectively seals the fissures. This
Sustainable construction bio-mediated repair mechanism not only enhances the structural integrity of
concrete but also aligns with sustainable construction practices by minimizing
maintenance requirements and material wastage. The review extends beyond
self-healing phenomena, encompassing broader applications of microbial
technology in construction, including bio-concrete, bio-cement, and soil
stabilization methods. These applications underscore the versatility of microbes
in enhancing material properties such as compressive strength, tensile
resilience, and water impermeability. Empirical evidence underscores the
necessity of optimizing bacterial dosages and curing conditions to maximize the
self-healing efficiency. Future research trajectories should aim to elucidate the
complex interactions between microbial agents and concrete matrices, assess
long-term performance, and evaluate the environmental and economic
sustainability of microbial interventions in construction. The integration of
microbial technology in construction materials heralds a new epoch of
innovation, offering robust, sustainable, and resilient solutions to enduring
challenges in the industry.

© 2024 MIM Research Group. All rights reserved.

1. Introduction
In the evolving landscape of civil engineering, the quest for durable and sustainable
building materials has become paramount, driven by the urgent need to address the
environmental and structural challenges of modern construction. Concrete, the
cornerstone of global infrastructure development, is under increasing scrutiny due to its
vulnerability to degradation and the environmental impact of its production.
This paper explores the utilization of microorganisms in construction materials and the
methods employed to evaluate their effect on strength. Concrete, being the most
extensively used building material in construction projects (1) is not without its

Corresponding author: [email protected]

*

orcid.org/0000-0003-3380-8359; borcid.org/0000-0003-4639-7528; corcid.org/0000-0001-5650-2980

a

DOI: http://dx.doi.org/10.17515/resm2024.207ma0304rv
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challenges, including the presence of micro cracks (2). These micro cracks can reduce the
workability of concrete and potentially lead to structural collapse and failure.
To address these challenges, various approaches have been employed, such as the use of
chemical admixtures (3) and fiber concretes (4). Additionally, the concept of self-healing
concrete has gained attention, aiming to minimize the need for manual maintenance. One
promising self-healing approach involves the utilization of specific bacteria that operate
independently within the concrete structure (2). When tensile forces exceed the concrete's
strength, fractures occur (5). These fractures can serve as pathways for water ingress and
the infiltration of toxic substances (6). Concrete cracks can be classified into two groups:
non-hazardous cracks that are visually unappealing but do not pose a safety threat (7), and
structural cracks that require careful attention to prevent significant harm to the overall
structure. Self-healing concrete methods, such as bacterial reactions within the hardened
concrete, have shown promise in mitigating the latter type of cracks (8).
Micro cracks typically range from 0.1 to 0.05 mm in width, and their presence allows the
infiltration of water particles, acting as capillaries (9). When water droplets penetrate the
cracks, partially or completely unreacted cement hydrates, leading to expansion and
subsequent sealing of the crack (10). Recent studies, by Saravanan et al., have delved into
the compressive strength development of geopolymer concrete using manufactured sand,
highlighting the environmental benefits and performance efficiencies of alternative
materials like geopolymer concrete in reducing carbon emissions associated with
conventional cement usage (11). Similarly, the work of Chaitanya et al. on the self-healing
characteristics of GGBS admixed concrete using Artificial Neural Networks underscores
the innovative strategies being developed to enhance concrete's self-healing properties
and reduce its carbon footprint (12).
Moreover, Cappellesso et al. (2023) reviewed the efficiency of self-healing concrete
technologies, affirming the potential of microbial and chemical methods to extend the
service life of concrete structures (13). The research on Ground Granulated Blast Furnace
Slag (GGBS) in concrete by Subramanian et al. (2022) also emphasizes the improved
flexural behavior and durability of concrete beams reinforced with polymer composites
(14). In addition, the investigations into the prediction of self-healing characteristics of
concrete with GGBS by M. Chaitanya et al. (2020) and the work of Aleem et al. on the
properties of Geopolymer concrete with M-sand provide comprehensive insights into the
advancements in concrete technology that contribute to more sustainable construction
practices (15,16).
The exploration of self-healing concrete is important, particularly in the context of material
costs and the judicious selection of optimal materials tailored to project requirements.
Attaining high-quality self-healing concrete necessitates a thorough examination of prior
studies to identify effective solutions. Understanding the response and workability of
concrete, along with addressing challenges in healing or sealing cracks, remains a dilemma.
Furthermore, self-healing concrete is recognized as a viable approach to minimize
structural maintenance requirements. Following established methodologies outlined in
previous literature can lead to reduced maintenance costs and alleviate environmental
impacts associated with traditional repair methods, such as the production and
transportation of materials. The field of bacterial concrete has rapidly evolved, offering
innovative solutions for enhancing the durability and sustainability of construction
materials. This technology is believed to not only promise extended lifespans for concrete
structures but also align with environmental sustainability goals. The integration of
bacterial spores into concrete matrices for self-healing purposes represents a significant
breakthrough in construction material science (17).

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This research assumes strategic significance by aiming to comprehensively assess and

analyze the impacts of self-healing on key concrete properties, including compressive
strength, tensile strength, and water absorption. The insights gleaned from this review are
pivotal for researchers, guiding the trajectory of advancements in self-healing concrete.
Ultimately, this contributes to the development of infrastructure that is both more resilient
and sustainable.

2. Application of Microbes in Construction

Microorganisms, particularly bacteria, have various applications in the construction
industry. In other words, bacteria bring a world of possibilities to the construction
industry, opening doors to innovative and sustainable practices. These tiny living
organisms play a significant role in enhancing various aspects of construction materials
and processes, paving the way for a more eco-friendly and resilient future (18).
Recent advancements in microbial applications for concrete have highlighted the efficacy
of various bacterial strains, such as Bacillus pseudofirmus and Bacillus cohnii. These
bacteria have been shown to significantly enhance concrete’s durability through the
process of calcium carbonate precipitation. This biogenic mineralization contributes to
self-healing of micro-cracks, thereby extending the lifespan of concrete structures. The
integration of these specific bacterial strains represents a promising development in
sustainable construction materials (19). In the following, the most famous applications of
microorganisms and using them in the construction is examined:
2.1. Self-Healing Concrete
In the domain of construction, self-healing concrete emerges as a captivating and
groundbreaking innovation. Imagine concrete structures imbued with the extraordinary
ability to mend themselves! This remarkable achievement is made possible through the
incorporation of specific bacterial strains from the Bacillus genus into the concrete mix.
These highly adept bacteria remain quiescent within the concrete until the emergence of
cracks, at which point they are triggered into action. In a process akin to skilled artisans,
they facilitate the production of calcium carbonate, serving as a natural adhesive that
adeptly seals the cracks and fortifies the concrete's structural integrity. The enchantment
of self-healing concrete extends beyond its capability to diminish maintenance
requirements; it also bestows an extended lifespan upon our structures, imbuing them
with heightened resilience and reliability as time unfolds (20,21).
2.2. Biodegradable Construction Materials
Microbes have emerged as a significant catalyst in the development of biodegradable
construction materials, including bioplastics and biocomposites. The incorporation of
these materials signifies a notable stride towards fostering eco-friendliness and mitigating
the environmental impact of construction activities. Leveraging the remarkable potential
of these microorganisms, we can fabricate bioplastics and biocomposites that epitomize a
more environmentally sensitive approach to construction. The intrinsic value of these
materials lies in their inherent propensity for natural degradation, leaving behind a
minimal ecological footprint while contributing to the reduction of waste in construction
projects. Such advancements underscore the crucial role of microorganisms in promoting
sustainable practices within the construction industry (22–24).
2.3. Bioconcrete and Biocement
The fascinating world of bioconcrete and biocement unfolds as it harnesses the power of
microbes in their production. Bioconcrete ingeniously incorporates bacteria capable of
limestone production, fortifying the concrete and enhancing its durability. On the other

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hand, bio cement relies on microorganisms that induce calcite precipitation within the
concrete matrix, bestowing it with heightened strength and resilience. Just imagine the
potential of these microbe-powered wonders-construction materials that not only meet
our practical needs but also demonstrate a profound commitment to environmental
stewardship. Through these advancements, we find ourselves on a path where sustainable
construction practices and ecological well-being go hand in hand (18,25,26).
2.4. Soil Stabilization
The other microbial application in construction involves soil stabilization, where certain
microbial species come to the rescue, especially in areas with weak or loose soil. These
remarkable microorganisms possess the fantastic ability to fortify the soil, rendering it
suitable and robust for construction purposes. By promoting microbial-induced calcite
precipitation, we can establish a solid foundation for our building endeavors, ensuring
stability and longevity in our construction projects. It's like nature's own construction
crew working behind the scenes to strengthen the very ground we build upon (27–29).

3. History and Importance of Use of Microbes

In the history of concrete, the introduction of calcium-rich bacteria during the mixing stage
has emerged as a significant development. These bacteria play a crucial role in the self-
healing process of concrete. When cracks form within the concrete, the introduced bacteria
initiate the precipitation of calcium carbonate. This natural process effectively seals the
cracks and reinforces the overall structure. Consequently, bacterial concrete exhibits
higher strength compared to conventional concrete. By leveraging a biotechnological
approach centered around calcite precipitation, it becomes possible to enhance both the
strength and durability of structural concrete (30,31).
The use of bacteria in concrete is an innovative method that harnesses the power of
microbial activity to improve the performance and longevity of concrete structures. The
formation of calcium carbonate through the action of bacteria leads to the creation of a
more robust and resilient construction material (32–34). Self-healing concrete can be
achieved through three main methods: natural, chemical, and biological. The biological
method can be further categorized into three subcategories: bacteria, fungi, and viruses.
Among these methods, the utilization of bacteria, particularly specific strains within the
concrete, has garnered significant interest (35). Self-healing concrete can be achieved
through three main methods: natural, chemical, and biological. The biological method can
be further categorized into three subcategories: bacteria, fungi, and viruses. Among these
methods, the utilization of bacteria, particularly specific strains within the concrete, has
garnered significant interest (35,36). The historical development and increasing interest
in using microbes, especially bacteria, in the self-healing process of concrete highlight the
potential of biotechnological advancements in construction materials. By incorporating
microbial activity, concrete can exhibit improved resilience and longevity, addressing the
challenges associated with cracks and structural deterioration (37). The historical
development of bio-concrete represents a significant milestone in construction material
technology. This evolution from traditional concrete to bio-concrete highlights a shift
towards sustainable, self-healing materials, emphasizing the crucial role of microbes in
modern construction practices (38).
OPC accounted for approximately 12% of global CO2 emissions in 2020. Alkali-activated
slag (AAS) has emerged as a potentially sustainable alternative to ordinary OPC. Research
suggests that producing AAS composites with intelligent properties, enabling maintenance
and repair with minimal external assistance, may be a sustainable solution. Additionally,
bacteria-based self-repairing represents a promising and sustainable alternative method
for repairing and conducting regular maintenance (39).

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3.1 Various Types of Bacteria Used in Concrete

Bacterial strains used in concrete are carefully selected for their ability to thrive in high-
pH environments. Typically, microorganisms cannot survive in alkaline conditions with a
pH value of 10 or higher (40,41). Table 1 provides a list of bacteria capable of withstanding
pH levels equal to or greater than 10.
Table 1. Bacteria that can be usable in concrete in the alkaline environment (41)

Type of the Compressive

No. Application References
Bacteria Strength (+ or -)
Mortar
Sporosarcina
1 Concrete (42–46)
Pasteurii +
Fly Ash
Mortar
2 Bacillus cereus NA (47–49)
Concrete
3 Bacillus flexus NA Mortar (50–52)
Bacillus Concrete
4 + (51,53–55)
megaterium Bricks

Bacillus Mortar
5 (56–61)
sphaericus + Concrete

Bacillus Concrete with 10%

6 NA (62,63)
halodurans Cement kiln dust
7 Bacillus cohnii + Mortar (64,65)
Bacillus
8 - Cement stone (66,67)
pseudofirmus
Mortar
9 Bacillus subtilis + (68–71)
Concrete
Diaphorobacter
10 - Mortar (61,72)
nitroreducens

11 Shewanella sp. + Mortar (73)

Mortar
12 Escherichia coli NA (74,75)
Concrete

13 Bacillus aureus + Rice Husk Ash (39)

Among these bacteria, those belonging to the Bacillus sp., such as Bacillus subtilis and
Bacillus megaterium, are well-known for their ability to thrive in highly alkaline
environments. In such conditions, these bacteria produce spores that resemble plant
seeds. These spores have robust walls and remain dormant until cracks develop in the
concrete, allowing water to penetrate the structure. When exposed to the pH range of 10
to 11.5, typical of highly alkaline concrete, these bacterial spores become active. Apart from
Bacillus sp., other bacterial species have also been found to survive in alkaline
environments (41,76).
In addition to pH, other factors play a role in the biochemical processes involving bacteria,
including the concentration of Ca+2 ions, the presence of nucleation sites, and the

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availability of dissolved inorganic carbon (77,78). Bacteria play a crucial role in creating
an alkaline environment through various pathways, including autotrophic and
heterotrophic processes. Among these pathways, enzymatic hydrolysis of urea, aerobic
oxidation of organic carbon, and anoxic oxidation of organic carbon have been extensively
studied and recognized as significant contributors to alkalinity generation.
These processes are essential for establishing favorable conditions for bacteria to thrive
and actively participate in the self-healing process of concrete. Extensive research has
demonstrated the pivotal role of bacteria in creating an alkaline environment and
promoting the healing capabilities of concrete structures (66,79–89).
The specific strains Bacillus subtilis and Bacillus megaterium, both gram-positive bacteria,
are commonly employed in concrete applications. Bacillus subtilis possesses a remarkable
ability to form highly resistant dormant endospores as a response to nutrient deprivation
and environmental stresses. It is commonly found as a gut commensal in humans and can
also be present in the upper layer of soil. On the other hand, Bacillus megaterium, besides
being prevalent in soil, can be found in various environments, including certain food items
like honey, as well as on surfaces such as clinical specimens, paper, and stone (90).
In the context of evaluating bacterial influence on concrete properties, it is imperative to
discuss the concept of optical density (OD). Optical density is a quantitative measure of the
attenuation of light as it passes through a sample containing particles or solutes. In
microbiological assays, this measure is often used to estimate the concentration of bacteria
within a culture by assessing the light absorption at a specific wavelength, typically 600
nm (OD600). The attenuation is due to both the scattering and absorption of light by the
bacterial cells, which corresponds to their concentration in the culture medium. The
relevance of OD measurements in microbial concrete research lies in its ability to correlate
bacterial concentrations with the observed effects on concrete's mechanical properties. In
self-healing concrete, where bacterial activity is pivotal, an optimal OD value indicates the
effective concentration of bacteria required to precipitate calcium carbonate to heal cracks
and enhance the structural integrity of the concrete. It is this precise and careful calibration
of bacterial density, ascertained through OD measurements, that allows for the targeted
improvement of compressive strength and other key concrete properties.
By carefully selecting bacterial strains that can survive and thrive in alkaline
environments, concrete applications can harness the self-healing properties of these
microorganisms, contributing to the durability and resilience of concrete structures.

4. Results and discussions:

4.1 Compressive Strength
After using Bacillus cereus and Bacillus subtilis, it is observed that compressive strength of
the concrete upsurged in both 7 and 28 days. However, the optimum amount of using these
kinds of bacteria (optimum amount of Bacillus cereus and Bacillus subtilis) is 103 CFU/mL
and 105 CFU/mL respectively (48).
The research utilized Ordinary Portland cement (OPC) with 25 grades along with two
different bacterial strains, namely Bacillus cereus and Bacillus subtilis, in the study.
Concrete cubes with dimensions of 150×150×150 mm was used for testing. The data
presented in Fig. 1 demonstrates the impact of these bacteria on concrete strength.

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Fig. 1. A comparison about compressive strength of using different types of Bacillus

cereus and Bacillus subtilis with different dosages (48)

It can be observed that the optimal concentration of Bacillus cereus resulted in a significant
enhancement in concrete strength, with a notable 20% increase at both the 7-day and 28-
day curing periods. Similarly, the application of Bacillus subtilis showcased even more
promising results, exhibiting a remarkable 32% increase in concrete strength at the 7-day
mark and a commendable 25% increase at the 28-day stage.

Fig. 2. comparison compressive strength of concrete with different types of bacteria

(Bacillus subtilis and Bacillus megaterium) (91)

Based on Figure 2, compressive strength values for four different concrete batches tested
at three different curing times: 7 days, 14 days, and 28 days. The concrete utilized is OPC
53 grade, molded into cubes with dimensions of 150mm per side. The chart demonstrates
a distinct increase in compressive strength for each batch as the curing time extends from
7 days to 28 days. This suggests that the incorporation of Bacillus sp. Bacteria (specifically
Bacillus subtilis and Bacillus megaterium) contributes positively to the strength
development of concrete. For instance, Batch 1 shows an appreciable increase in strength
at each testing interval, which indicates that the bacterial treatment could be influencing
the curing process and improving the concrete's mechanical properties. This pattern is
consistent across all batches, confirming the benefit of bacterial additives in concrete

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mixtures. The data signifies that both early age and longer-term strength properties are
enhanced, highlighting the potential for these biological agents to improve construction
materials' performance.
Also, it is observed that concrete batches treated with Bacillus megaterium initially
demonstrate lower compressive strength at the 7-day mark compared to those treated
with Bacillus subtilis and the combined consortia. This initial lag suggests that Bacillus
megaterium may have a slower start in the bio-mineralization process which contributes
to the concrete's strength. However, as the curing period extends to 28 days, a remarkable
phenomenon occurs: the Bacillus megaterium-treated batches exhibit superior
compressive strength compared to their Bacillus subtilis counterparts and the consortia.
The reasons behind this trend could be multifold. It is possible that Bacillus megaterium
engages in a more gradual but ultimately more effective calcium carbonate precipitation
process, which is a critical factor in concrete strengthening. The delay in strength gain
could also be attributed to the specific metabolic pathways of Bacillus megaterium that
may take longer to kickstart but result in more robust crystal formation over time. Another
aspect to consider is the possibility that Bacillus megaterium could be more effective at
pore-filling within the concrete matrix, which becomes evident only in the later stages of
curing.

Fig. 3. Compressive strength of using Enterococcus faecalis, Bacillus sp. and mix of
Enterococcus faecalis and Bacillus sp. per MPa (92)

In this experiment, the compressive strength of concrete was evaluated over an extended
curing period by examining four distinct batches: Batch 1 (control), Batch 2 (Enterococcus
faecalis), Batch 3 (Bacillus sp.), and Batch 4 (combination of Bacillus sp. and Enterococcus
faecalis). The primary objective was to assess the impact of these bacterial compositions
on the concrete's strength development. The cube specimens used in the study were
150mm ×150mm ×150mm, and Portland composite cement (PCC) was utilized. The
presented graphical representation (Fig. 3) displayed the results obtained from the
experiment, with Batch 3 (Bacillus sp.) demonstrating the most favorable and optimal
outcomes in terms of compressive strength. Notably, the concrete's strength noticeably
increased after 7 and 14 days of curing, with Batch 3 exhibiting the highest strength values
among all the batches. The inclusion of Bacillus sp. at a 5% concentration proved to be
effective in enhancing the concrete's compressive strength within the specified time frame.
Through this analysis, valuable insights were gained regarding the influence of different

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bacterial additives on the concrete's performance, contributing to a deeper understanding

of their role in enhancing the material's mechanical properties.

Fig. 4. Compressive strength of concretes with different concentrations of Bacillus

subtilis (93)

Fig. 4 presents the results of the compressive strength test, revealing a consistent
enhancement in the strength of the various concrete samples over the curing duration.
After 28 days of curing, the compressive strengths of the concrete specimens with different
concentrations of Bacillus subtilis (0, 105, 107, and 109 cell/ml) were measured at 26.2 MPa,
29.9 MPa, 27.4 MPa, and 26.8 MPa, respectively. Notably, all the tested concretes exhibited
higher compressive strengths than the target strength, with all samples incorporating
different concentrations of B. subtilis outperforming the control sample. The percentage
increases in strength relative to the control sample were 14%, 5%, and 2% for B. subtilis
concentrations of 105, 107, and 109, respectively, after 28 days of curing. Similarly, after 56
days of curing, the percentage increases in strength compared to the control sample were
13%, 4%, and 1% for B. subtilis concentrations of 105, 107, and 109, respectively, consistent
with the 28-day curing results. The obtained results underwent statistical analysis using
Analysis of Variance (ANOVA), confirming that both bacterial concentrations and curing
duration significantly influenced the compressive strengths of the concrete samples (p <
0.05). This analysis highlights the critical role of these factors in determining the
performance and quality of the concretes. It is worth noting that for this research, Portland
cement (32.5 N) was used as the binder material.
Fig. 5 presents a comparison of different mixes, denoted as Mix 1 to Mix 5, with varying
culture densities (OD600) of 0, 0.107, 0.2, 0.637, and 1.221 respectively. The graphical
representation highlights the relationship between compressive strengths and optical
densities across various curing periods. The control specimen or Mix 1 exhibited
compressive strengths of 20.1, 31.8, and 38.9 MPa at the age of 28 days, as per the designed
strengths. Notably, all bacterial groups displayed an increase in compressive strength upon
the addition of B. subtilis, with the highest values recorded in Mix 4 at 23.7, 35.6, and 42.5
MPa respectively.

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Fig. 5. Comparison of Compressive Strength and Optical Density of Bacillus subtilis

(94)

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The compressive strengths of Mix 1 at a curing period of 120 days were determined to be
30.6, 35.8, and 47.1 MPa, while Mix 4 exhibited superior performance compared to the
other mixes with recorded values of 35.2, 43.2, and 57.2 MPa for the specified design
strengths. Moreover, it illustrates the increase in strengths on the 28th day for different
culture densities. It is evident that Mix 4, with a culture density of OD600=0.637, displays a
notable improvement across all design strengths. This can be attributed to the enhanced
formation of mineral deposits within the internal structures of the concrete. Same results
reported from Nivedhitha et. Al while they used the Bacillus subtilis for self-healing
concrete (95).
In the domain of microbial-enhanced concrete, the introduction of Bacillus subtilis is of
paramount significance, which could be elaborated in the context of the experimental
observations discussed in Figure 2. Notably, this strain has demonstrated a consistent
ability to increase the compressive strength across all bacterial groups. This enhancement
is attributable to the unique properties of Bacillus subtilis that include promoting better
cohesion in the cement matrix, instigating biogenic mineral precipitation, and fostering
synergistic interactions within the microbial community embedded in the concrete. Such
cohesive interactions are believed to reduce the porosity of concrete, thus densifying the
matrix and enhancing the mechanical properties. Furthermore, Bacillus subtilis facilitates
calcium carbonate precipitation, not merely as a crack-filling agent but as a means to bind
concrete constituents more effectively, thereby improving compressive strength. Its ability
to stimulate other beneficial bacteria within the concrete also contributes to a more robust
and uniformly healed material. The intricacies of these interactions and their influence on
the mechanical properties of concrete provide valuable insights into the biological
mechanisms underpinning the improved durability and structural integrity of the
construction material.
4.2 Tensile Strength
The inclusion of bacteria in concrete demonstrates a parallel effect on the tensile strength,
mirroring the notable improvements observed in compressive strength. Tensile strength
is an important measure of a material's ability to resist breaking under tension and is
crucial for the overall structural integrity of concrete. Fig. 6 displays the outcomes of the
tensile strength tests conducted on the concrete samples.

Fig. 6. Comparison tensile strength of concrete with different types of bacteria

(Bacillus subtilis and Bacillus megaterium) (91)

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Fig. 7. Comparison of Tensile Strength and Optical Density of Bacillus subtilis (94)

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The results indicate that the presence of bacteria has a substantial and successful impact
on the tensile strength of the concrete, particularly after a curing period of 7 and 28 days.
It is worth noting that the tensile strength values are measured in megapascals (MPa).
These findings further support the efficacy of bacteria in enhancing the tensile properties
of the concrete over time. Fig. 7 presents a comparative analysis of several concrete mixes,
labeled as Mix 1 to Mix 5, with each mix associated with different culture densities
represented by the optical density at 600nm (OD600) values. The evaluated mixes
correspond to OD600 values of 0, 0.107, 0.2, 0.637, and 1.221, respectively, enabling a
thorough examination of how varying culture densities influence the performance and
properties of the concrete. At a curing period of 28 days, mix 1 exhibited split tensile
strengths of 3.2 MPa, 3.5 MPa, and 3.8 MPa for the specified design strengths.
Notably, mix 4 demonstrated the highest increase in split tensile strength, reaching values
of 3.5 MPa, 3.9 MPa, and 4.2 MPa, respectively, indicating a significant enhancement in its
tensile properties compared to the other mixes. Moreover, after an extended curing period
of 120 days, Mix 4 exhibited substantial improvements ranging from 13% to 18% in split
tensile strength across all grades. This extended curing duration facilitated further
development and strengthening of the concrete's tensile capabilities. The results
underscore the effectiveness of Mix 4 in enhancing split tensile strength, suggesting its
potential for applications requiring improved tensile performance in various construction
projects. For this research, specimens were prepared using ordinary Portland cement
(OPC) with strength class 42.5 N.

Fig. 8. splitting tensile strength of standard and Bacillus megaterium bacteria concrete.
(96)

According to Fig 8, the addition of Bacillus megaterium into the concrete mixture results in
a noticeable improvement in splitting tensile strength. The concrete samples with Bacillus
megaterium showed an increase of 9.88% at 7 days and 10.28% at 28 days, compared to
the standard concrete samples.
4.3 Water Absorption
Water absorption is a significant and widely recognized test for concrete. In this study, we
will delve into the water absorption test while incorporating bacteria and examine their
impact on the concrete's strength and the rehabilitation of micro cracks. This investigation
aims to shed light on the potential benefits of utilizing bacteria in enhancing concrete
properties and durability.

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Fig. 9. Analysis of water absorption on calcined clay concrete and OPC in different
dosages of bacteria. (97)

Fig. 9 illustrates the distinctions between Ordinary Portland Cement (OPC) and the
addition of a pozzolan replacement, specifically calcined clay concrete (CC), with varying
dosages of Bacillus subtilis. The incorporation of bacteria in calcined clay concrete led to a
notable reduction in water absorption capacity compared to regular concrete and calcined
clay concrete. Over a 28-day curing period, the inclusion of B. subtilis in calcined clay
concrete resulted in water absorption reductions of 18.30%, 17.38%, and 13.59% for 10%,
15%, and 20% calcined clay replacements, respectively. These findings indicate that the
water absorption of the specimens decreased in bacterial-infused CC when compared with
specimens without bacteria. The study utilized OPC grade 43 and casted M25 grade of
concrete cubes with dimensions of 150×150×150 mm.

Fig. 10. Water absorption of standard and Bacillus megaterium concrete. (96)

In Fig. 10, the study utilized Ordinary Portland Cement (OPC) of grade 43, and Bacillus
megaterium at a concentration of 108 CFU/mL was employed. The specimens were of size
150mm x 150mm x 150mm. The results demonstrate that when B. megaterium is
incorporated into the concrete mix, it leads to a reduction in water absorption both at 7
and 28 days, by 5.25% and 7.35%, respectively, as compared to the control samples.

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Table 2. Water absorption of different bacteria after 28 days of curing. (98)

Type of the Results of water
Researchers
Bacteria absorption after 28 days
The concrete sample
De Muynck et al. (99), exhibited a reduction of
B. sphaericus
Achal et al. (100) 45–50% compared to the
controlled sample.
The concrete sample
showed a reduction of
Reddy et al. (101), Muhammad et al.
B. subtilis nearly 50% compared to
(102),
the controlled concrete
Pei et al. (103)
sample.
The concrete sample
exhibited a decrease of
B. megaterium Dhamia et al. (53) 46% compared to the
controlled concrete
sample.
The concrete sample
showed a reduction of
Sporosarcina Achal et al. (104),
50-70% compared to the
pasteurii Pei et al. (105)
controlled concrete
sample.

The concrete sample

De Muynck et al. (99), exhibited a 50%
B. pseudofirmus
Maheshwaran et al. reduction compared to
the controlled concrete
sample.
Based on the data presented in Table 2, a comparison was made regarding the impact of
various bacterial types on water absorption. The results indicate that Sporosarcina
pasteurii exhibited the highest effectiveness in reducing water absorption, achieving a
remarkable reduction of up to 70%. On the other hand, other Bacillus sp. showed a water
absorption reduction ranging from 45% to 50%. Consequently, it can be inferred that the
selection of bacterial species does not significantly influence the water porosity variation.
Recent studies, as explored in 'Calcium Carbonate Precipitation by Different Bacterial
Strains for Concrete Repair', have demonstrated the significant role of bacterial strains like
Bacillus sphaericus and Bacillus pasteurii in enhancing concrete's compressive strength
and reducing its water absorption. These strains facilitate the biomineralization process,
leading to calcium carbonate formation that not only aids in sealing micro-cracks but also
improves the overall structural integrity of the concrete. This innovative approach
promises a new dimension in prolonging the durability of concrete structures, particularly
in environments subjected to extensive wear and tear (106).

5. Discussion and Recommendation for Future Studies

The exploration into microbial concrete has laid a foundation for innovation within the
construction industry, yet it is clear that we have only begun to unearth the full spectrum
of its capabilities. Future research directives must be comprehensive and multifaceted to
span the depths of this burgeoning field.
Firstly, the scope of microbial agents employed in concrete must be broadened. Diverse
microorganisms, including fungi and algae, should be investigated for their unique

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biogenic processes that may yield more robust and resilient self-healing concrete
formulations. Uncovering alternative biological pathways could pave the way for concrete
that is not only self-repairing but also possesses enhanced mechanical properties.
Longitudinal studies are imperative to ascertain the long-term viability and efficacy of
microbial concrete. Such studies should rigorously evaluate the performance of these
innovative materials against the rigors of environmental stresses encountered in situ, such
as freeze-thaw cycles, corrosive chemical exposures, and abrasion. Understanding how
microbial concrete withstands such conditions will inform the engineering of materials
that can endure through decades rather than just years.
Further, there is an essential need to refine the delivery and maintenance of microbial
concentrations within the concrete matrix. Innovative techniques, such as advanced
encapsulation or the development of biofilms, must be optimized to balance the microbial
activity with structural integrity. These methods should ensure the longevity of microbial
viability and consistent healing action throughout the concrete’s lifecycle.
Additionally, a thorough ecological and economic analysis is essential to evaluate the
sustainability of microbial concrete. This encompasses a complete lifecycle assessment
that addresses the environmental impacts from production to end-of-life recycling or
disposal. The integration of such assessments will ensure that the advancement in
construction materials technology is in concert with our sustainability objectives,
minimizing the carbon footprint while maintaining cost-efficiency.
In the vein of scientific inquiry, there is an urgency to decode the biochemical intricacies
of microbial calcite precipitation. Understanding the molecular mechanisms that govern
this process could lead to the optimization of microbial consortia, tailored for enhanced
self-healing efficiency. Investigating how Bacillus subtilis and other bacteria interact with
varying concrete compositions will allow for broader application and more customized
material solutions.
The path to harnessing the full potential of microbial technologies within construction
materials is complex, demanding a confluence of expertise from disciplines such as
microbiology, material science, and civil engineering. Collaborative research efforts must
converge to innovate resilient, self-sustaining, and environmentally benign construction
materials. As we stride toward this goal, we must also ensure that our scientific and
engineering pursuits are scalable and transferable to practical applications, cementing the
legacy of microbial concrete as a pillar of sustainable development in the construction
industry.

6. Conclusion
The study meticulously explores the innovative integration of microbial technologies in
the construction sector, specifically through the development of self-healing concrete
utilizing bacterial agents. The investigation has illuminated the significant role of bacteria,
notably the Bacillus species, in autonomously repairing microcracks that compromise the
structural integrity of concrete. This self-healing mechanism, driven by the microbial-
induced calcite precipitation process, not only seals cracks but also contributes to the long-
term durability and strength of the concrete infrastructure.
Our comprehensive analysis has revealed that the incorporation of bacteria like Bacillus
cereus and Bacillus subtilis leads to marked improvements in the compressive strength of
concrete within the initial seven days of curing. This enhancement persists and becomes
more pronounced with prolonged curing periods, underlining the enduring benefits of
microbial action on the concrete’s mechanical properties. The optimal bacterial dosages
and specific curing conditions emerge as critical factors that significantly influence the

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efficacy of the self-healing process, necessitating further research to establish

standardized protocols for practical applications.
Furthermore, the study extends beyond the immediate realm of self-healing concrete to
explore broader applications of microbial interventions in construction materials. This
includes the development of bioconcrete and biocement, which leverage microbial
activities for material fortification and sustainability. The soil stabilization techniques
enhanced by microbial action also present a promising avenue for improving the
foundational stability of construction sites, particularly in geotechnically challenging
environments.
The empirical evidence gathered points to a substantial reduction in water absorption
rates in microbially treated concrete, a factor that contributes to the material’s resilience
against environmental degradation and extends its service life. The nuanced interplay
between bacterial concentration, culture density, and environmental conditions in
dictating the performance of microbial concrete underscores the complexity of this
innovative technology.
The integration of microbial technologies in the construction industry heralds a
transformative era in material science, where sustainability, durability, and self-repairing
capabilities become intrinsic characteristics of construction materials. The potential of
bacteria-infused concrete to revolutionize building practices is immense, offering a
sustainable solution to the perennial challenges of material degradation and
environmental impact. However, the path to widespread adoption and optimization of
these technologies are paved with challenges that necessitate meticulous research,
standardized testing protocols, and a deeper understanding of microbial ecology within
construction materials.
The dynamic interplay of bacterial agents within the concrete matrix heralds a complex
ecosystem, whose understanding is pivotal for the optimization of material properties and
engineering applications. Future investigations must delve into the nuanced mechanisms
of bacterial interactions, with an emphasis on real-time dynamics and long-term impacts
on structural resilience and durability. Moreover, the potential for an engineered synergy
between diverse microbial species and concrete formulations beckons a new frontier in
construction material science. To translate this pioneering research into tangible
engineering practices, it is imperative to devise strategies for the scaled-up application that
address economic feasibility, environmental considerations, and compatibility with
existing construction methodologies. The pathway forward is collaborative and
interdisciplinary, requiring concerted efforts from microbiologists, material scientists, and
civil engineers, to harness the full spectrum of benefits offered by microbial technologies
in construction.
While this study has provided valuable insights into the potential of microbial concrete,
particularly focusing on self-healing properties and the role of Bacillus species, it
acknowledges certain limitations. The experimental conditions under which the microbial
concrete's performance was evaluated might not fully replicate the complex
environmental conditions encountered in real-world applications. The longevity and
sustainability of the microbial-induced healing process under varied climatic and loading
conditions remain to be comprehensively understood. Additionally, while the study
explored a range of bacterial species, the ecological impact of introducing these
microorganisms into construction materials warrants further investigation to ensure that
no adverse environmental consequences arise. Economic feasibility, scalability of the
microbial application process, and the potential for microbial resistance development in
the concrete matrix are aspects that need to be addressed in future research. These
limitations underline the necessity for continued and extensive investigation into the

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dynamics of microbial concrete to optimize its application in the construction industry and
to fully understand its environmental implications.

Acknowledgement
I would like to express my sincere gratitude to Associate Professor Dr. Mümtaz Güran for
his exemplary guidance, invaluable teaching methods, and insightful contributions to both
this paper and my thesis.

Author Contribution Statement

Farzin Asgharpour led the project with roles in conceptualization, methodology, data
curation, and writing the original draft, ensuring the study's integrity. Kadir Çakıral
enhanced the manuscript with visualization, investigation, and editing. Khaled Hamed
Marar provided supervision and guidance, shaping the research direction and quality.

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