Construction and Building Materials 242 (2020) 118059

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Construction and Building Materials

journal homepage: www.elsevier.com/locate/conbuildmat

Use of bacteria for repairing cracks and improving properties of concrete

containing limestone powder and natural zeolite
Maedeh Sadat Jafarnia a, Mehdi Khodadad Saryazdi a,⇑, Seyed Mohammad Moshtaghioun b
a
Department of Civil Engineering, Yazd University, Yazd, Iran
b
Department of Biology, Yazd University, Yazd, Iran

h i g h l i g h t s

Cement substitution by natural zeolite increased compressive strength.

Substitution of limestone powder decreased the compressive strength unless combined with zeolite.

Properties of concrete improved by using bacteria cells in the mixtures.

Utilizing bacteria in curing environment is very effective in crack healing.

a r t i c l e i n f o a b s t r a c t

Article history: Precipitation of calcium carbonate crystals caused by the metabolic activities of certain microorganisms
Received 27 February 2019 is a relatively new method which can improve the properties of concrete and repair concrete cracks. The
Received in revised form 25 November 2019 present study investigated the effects of Sporosarcina pasteurii bacteria on healing cracks, compressive
Accepted 2 January 2020
strength, tensile strength, ultrasonic pulse velocity, electrical resistivity and microstructure of concrete
containing various percentages of limestone powder and natural zeolite. Experimental results show that
the microbial calcite precipitations enhanced compressive strength, tensile strength, ultrasonic pulse
Keywords:
velocity and electrical resistivity of all specimens at all ages. The maximum values of these parameters
Concrete
Bacteria
are related to the bacterial specimen containing 10% zeolite without limestone powder. The SEM images
Limestone powder of the specimens show that the amount of calcite crystals in the bacterial treated specimen containing
Natural zeolite limestone powder is more than the specimen without that. In addition, crack healing of the specimen
Compressive strength containing limestone powder was also slightly better than the specimen without that. The results of
Crack healing Fourier-Transform Infrared spectroscopy show that the precipitation formed at crack surfaces of speci-
mens is CaCO3.
Ó 2020 Elsevier Ltd. All rights reserved.

1. Introduction
Because of affordable prices, strength and durability properties,
nowadays concrete is one of the most consumed construction
materials in the world [1–3]. But the use of concrete also has its
own issues. Cracks in concrete are caused by various factors includ-
ing shrinkage, chemical reaction, tensile loading, differential settle-
ment and thermal gradients. Without appropriate repairs, the
cracks will grow and expanded cracks would jeopardize the dura-
bility of concrete. Furthermore, repairing expanded cracks costs
more [2,4–6]. Nowadays, there are various techniques for repairing
⇑ Corresponding author at: Yazd University, Safaeeyah, Yazd, Iran.
E-mail addresses: maedeh_jafarnia@yahoo.com (M.S. Jafarnia), mkhodadad@
yazd.ac.ir (M. Khodadad Saryazdi), moshtaghiun@yazd.ac.ir (S.M. Moshtaghioun).
cracks of concrete. But traditional repair techniques are time con-
suming and have limitations. In recent years, utilizing microbial
calcite precipitation in concrete to repair cracks and improve its
properties has attracted attention of many researchers [7–13]. By
filling the pores of the concrete matrix and the cracks with calcite
precipitations, the possibility for corrosion of the embedded steel
reduces.
Bacteria are single-celled organisms that display various
shapes and sizes. There are incredibly diverse bacterial species.
Various bacterial species that contribute to carbonate deposition
exist in different natural environments, including soils, oceans
and lakes. Ureolytic bacteria are one of the most impressive spe-
cies of microorganisms in producing large amounts of carbonate.
The optimum pH range for ureolytic activity of soil urease bac-
teria is from about 6.5 to 9 and the high alkaline condition of

https://doi.org/10.1016/j.conbuildmat.2020.118059
0950-0618/Ó 2020 Elsevier Ltd. All rights reserved.
2 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059
concrete is not the best pH for urease activity of the bacteria.
Sporosarcina pasteurii is a non-pathogenic soil-inhabiting ure-
olytic bacterium that has been used throughout this study and
is not harmful to human health. Sporosarcina pasteurii is a robust
species in alkaline environments that can produce endospore in
harsh environmental conditions [14–16]. Ureolytic bacteria such
as Sporosarcina pasteurii need urea and calcium source to precip-
itate CaCO3. In this process, during the enzymatic hydrolysis of
urea, urea is converted to ammonias and carbonate. If there is
a source of calcium, bacteria act as a nucleation site to form cal-
cite precipitation [5,17,18].
Cement industry has important environmental impact on
carbon dioxide emission and global warming, since it is respon-
sible for about 5–8% of total anthropogenic carbon dioxide
emissions in the world [19–21]. Nowadays, researchers are
seeking to reduce the pollution caused by cement productions
by reducing cement consumption in concrete. In recent years,
according to environmental benefits, Portland limestone
cements (PLC) has been considered [22–24]. So that the Euro-
pean standard EN 197-1-2000 has different groups of PLC con-
taining 6–20% and 21–35% limestone powder [25]. The
Institute of Standards and Industrial Research of Iran ISIRI
4220-2005 also allows the use of 6–20% limestone powder as
cement replacement in PLC [26]. Moreover, nowadays utilizing
natural pozzolanic materials has become a common way to
improve concrete properties, enhance its strength and decrease
its permeability. One of the natural pozzolanic material is zeo-
lite. A large number of studies have been devoted to investigate
effects of natural zeolite on the mechanical and durability prop-
erties of concrete [27–30].
Fig. 1. Light micrograph of gram-positive rods of Sporosarcina pasteurii PTCC 1645
(Grams stain, 1000X).
Table 1
Physical properties of aggregates.
Material Specific gravity (SSD) Water absorption (% dry mass)
Fine aggregate 2.62 1.23
Coarse aggregate 2.69 0.96
Table 2
The chemical composition of cement, natural zeolite and limestone powder (%).
SiO2 Fe2O3 K2O
Ordinary Portland Cement 21.86 3.7 0.6
Natural Zeolite 68.95 0.97 0.95
Limestone Powder 0.32 0.1 – 0.02
So far, several studies have examined the effects of the use of
microbial calcite precipitation in various types of concrete, includ-
ing fly ash and silica fume concrete [31–33], rice husk ash concrete
[34], and lightweight aggregate concrete [35]. But no such work
has been reported on the effect of bacteria on repairing cracks
and improving properties of concrete containing limestone powder
and zeolite as partial replacement to cement. This paper investi-
gates the effect of bacteria on mechanical properties and durability
in terms of ultrasonic pulse velocity and electrical resistivity. Also,
the crack healing of specimens was evaluated by handheld micro-
scope and the microstructure of the concrete specimens and the
cracks filler was investigated by SEM analysis and Fourier-
Transform Infrared spectroscopy.
2. Materials and methods
2.1. Microorganism preparation
In this study Sporosarcina pasteurii PTCC 1645 was used in concrete. Microscopic
image of this bacterial strain is shown in Fig. 1. Bacteria were cultured in liquid
medium containing 5.0 g of peptone and 3.0 g of meat extract per liter of distilled
water (pH 7.0). Culture medium was autoclaved for 15 min at 121.5 °C. After inoc-
ulation under laminar flow hood the medium culture was incubated at 30 °C on a
shaker for 72 h at 150 rpm. After 72 h of incubation the liquid medium was cen-
trifuged at 4000 rpm for 15 min. After re-suspending bacterial cells in water, the
optical density of solution was measured by spectrophotometer at 600 nm wave-
length and the concentration of bacteria in the suspension was adjusted to about
107 cells/cm3.
2.2. Materials and mixture design
Ordinary Portland cement (CEM I-42.5), natural zeolite, limestone powder, fine
aggregate (nominal size of 4.75 mm) and coarse aggregate (nominal size of
12.5 mm) were used in this work. Physical properties of aggregates are shown in
Table 1 and the chemical properties of cement, natural zeolite and limestone pow-
der which were used in concrete mixes, are shown in Table 2. Two groups of mix-
tures were considered: one without bacteria (mix ID starts with letter C) and the
other with bacteria in mixing water (mix ID starts with letter S). Composition of
concrete mixture designs are presented in Table 3. As shown in Table 3, all mixtures
were prepared with constant amount of water, binder, fine and coarse aggregate.
Control mixtures (C-0-0 and S-0-0) containing only cement (450 kg) as the binder
and in other mixtures, various percentages by mass of cement were replaced with
limestone powder (first number followed by letter C or S in mix ID: 10, 20 and 30%)
and/or natural zeolite (second number in mix ID: 10%). Therefore, water to binder
ratio (W/B) of all mixtures is equal to 0.4, but the water to cement ratio (W/C)
increases by reducing amount of cement.
2.3. Curing environment
24 h after casting, the specimens were demolded and cured in different solution
for at least 27 days. As shown in Table 3, two group of specimens were used in this
study and specimens were kept in two different curing environment. The first group
which was cured in water, has no bacteria in mixture design. The second group has
bacteria in mixing design water and cured in aqueous molar solution of urea and
calcium chloride to investigate the effect of bacteria on properties of concrete.
The concentrations of calcium chloride and urea used in curing environment were
49 g/L and 20 g/L, respectively.
2.4. Testing procedure
2.4.1. Compressive strength and tensile strength
Compressive strength test was performed on cubic specimens with dimension
of 100 mm  100 mm  100 mm in accordance with EN 12390 [33] at ages of 28
and 60 days. Split tensile test (Brazilian test) were conducted as per ASTM C496
[34] on cylindrical specimens with dimension of 100 mm in diameter and
200 mm in height at age of 28 days.
AL2O3 CaO SO3 MgO Na2O TiO2 P2O5 Loss on ignition
4.8 64.4 2.4 1.2 0.55 – – 0.55
11.14 4.83 0.07 0.79 0.95 – – 10.64
55.3 – 0.23 – 0.01 0.02 43.43
 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059 3

2.4.2. Electrical resistivity

Curing condition Chloride ions can penetrate through the network of concrete pores and cause
corrosion of reinforcement in concrete. Studies show that there is a strong correla-
tion between electrical resistivity and chloride ions penetration in different con-

Urea-CaCl2
Urea-CaCl2
Urea-CaCl2
Urea-CaCl2
Urea-CaCl2
Urea-CaCl2
Urea-CaCl2
Urea-CaCl2
crete mixtures [37–39]. Electrical resistivity of concrete is a characteristic of

Water
Water
Water
Water
Water
Water
Water
Water
concrete, indicating the resistance against the flow of the electrical current which
is the result of ions movement in concrete. Electrical resistivity of concrete is the
most important parameter to estimate the corrosion of the reinforcements in con-
crete. In general, these two parameters are inversely related. So that by increasing
the electrical resistivity of concrete, the corrosion rate of the reinforcements in con-
crete decreases [36,38]. So far, several studies have been conducted on the relation-
Bacteria content

ship between electrical resistivity and other parameters of concretes [35–37].

In this study electrical resistivity was measured on cylindrical cores, taken from
(cell/ml)

100 mm cubic specimen, with diameters of 54 mm and heights of 100 mm. To mea-
sure the electrical resistivity, the specimens were placed between two copper

107
107
107
107
107
107
107
107
plates. The test set-up is shown in Fig. 2. The electrical resistivity of the specimens
0
0
0
0
0
0
0
0
was determined according to Eq. (1).

VA
q¼ ð1Þ
IL
Coarse aggregate

where q is electrical resistivity, V is potential difference, I is current, A is surface area

of specimen and L is height of the concrete specimen.
(kg/m3)

712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3
712.3 2.4.3. Ultrasonic pulse velocity
Ultrasonic pulse velocity measurement is a non-destructive test that are
influenced by microstructure, mechanical properties and concrete quality. As
the quality of concrete improves, the speed of ultrasonic waves through
concrete specimen increases [27,40]. Cubic specimens with dimension of
100 mm  100 mm  100 mm were used for measuring ultrasonic pulse veloc-
aggregate
(kg/m3)

ity as per ASTM C597 [41] at ages of 28 and 60 days. Two probes were posi-
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
1040.6
Fine

tioned on two parallel and smooth surfaces of the test specimens. These two
probes emit and receive the ultrasound waves and the device records the time
required for the ultrasound pulse to pass through the width of specimen. The
pulse velocity is computed by dividing the distance to the time measured.
(kg/m3)

(kg/m3)
Water

2.4.4. Crack healing

180
180
180
180
180
180
180
180
180
180
180
180
180
180
180
180

For measuring crack healing capacities, cylindrical specimens with heights of

200 mm and diameters of 100 mm were used. To induce crack in the specimen, it
was loaded similar to splitting tensile test but only half of the height of the cylin-
drical specimen was placed under loading plates, in this way cracks with different
widths were created without splitting the specimen in half. Points were marked
45
45
45
45

45
45
45
45
0
0
0
0

0
0
0
0
substitution
(% by mass)

along the induced cracks and the crack width at each point was measured by a
Zeolite As

hand-held microscope with a measurement accuracy of 0.02 mm. After measuring

Natural

cement

the initial crack width, the specimens were immersed in the curing environment, an
aqueous molar solution of urea and calcium chloride. They were taken out of the
10
10
10
10

10
10
10
10
0
0
0
0

0
0
0
0

curing media at different times and the crack width at the marked points were mea-
sured. Then they were immersed back into the curing environment. Crack widths of
healing specimens were measured after 5, 15, and 30 days of curing. To evaluate the
crack-healing capacity of different specimens, the percentage of crack healing was
(kg/m3)

calculated as follows (Eq. (2)).

As cement substitution
Limestone Powder

135

135

135

135
45

45

45

45
90

90

90

90
0

0
mass)
(% by

10
20
30

10
20
30

10
20
30

10
20
30
0

0
Cement
(kg/m3)

315

315

315

315
450
405
360

405
360

270
450
405
360

405
360

270
Mixture I.D.

C-10-10
C-20-10
C-30-10

S-10-10
S-20-10
S-30-10
C-10-0
C-20-0
C-30-0
C-0-10

S-10-0
S-20-0
S-30-0
S-0-10
C -0-0

S-0-0
Concrete mix designs.

Second
Group
Table 3

First

Fig. 2. Electrical resistivity test set-up.

4 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059

W0  Wt compressive strength of all the mixtures. So that the addition of

Healing percentageð%Þ ¼  100 ð2Þ
W0
bacteria to the control specimen enhances the compressive
where W 0 is the initial width of the cracks created and W 1 is the width of the cracks strength about 3.9% at 28 days. The compressive strengths of
at the time of the measuring. Three mixtures were selected to be investigated. The nonbacterial specimens containing 10%, 20% and 30% limestone
first one is the bacterial specimen without limestone powder and zeolite (S-0-0).
powder (C-10-0, C-20-0, C-30-0) are 8.6, 11.8, 17%, lower than
The second one is also a specimen with bacterial in its mixture but containing 20%
limestone powder and 10% zeolite (S-20-10). This mixture was selected because it the compressive strength of the control specimen (C-0-0) at age
contains less cement but has good compressive strength. Third specimen (S-CE) of 28 days. While the compressive strengths of bacterial specimens
was similar to first one (0% lime stone powder and 0% zeolite) but instead of using S-10-0, S-20-0 and S-30-0 are 6.4%, 8.3%, 12.2% lower than the
bacterial in the mixture, rather this specimen was cured in the suspension of bacte- compressive strength of control specimen (C-0-0) at the age of
ria and a reactive solution. The concentration of bacteria in curing environment was
107 cell/ml.
28 days. As stated, using 10% of zeolite as cement replacement
enhances the compressive strength of the specimens. So that the
2.4.5. Microstructure of specimens compressive strengths of the specimens C-0-10, C-10-10 and
To analyze the microstructure of the bacterial and nonbacterial specimens, C-20-10, are about 20.6% and 7.5% and 0.6% more than the com-
Scanning electron microscopic (SEM; Vega3 Tescan) analysis at the accelerating pressive strength of the control specimen at 28 days. These
voltage of 26 KV was performed on small broken pieces of concrete specimens. enhancements in compressive strength for the bacterial specimens
Hence, six different specimens (bacterial and nonbacterial specimens without lime-
(S-0-10, S-10-10 and S-20-10) compared to the control specimen
stone powder and zeolite, and mixtures containing 30% limestone powder and 10%
zeolite) were examined with scanning electron microscope. Calcite precipitation of are respectively about 23%, 13.2% and 9%. Although the compres-
crack filler was also visualized by a field emission Scanning electron microscope sive strength of the specimen containing 30% limestone powder
(FESEM; Mira3 Tescan, Czech republic) analysis at the accelerating voltage of 15 and 10% zeolite is about 8.2% less than the compressive strength
KV. Moreover, these precipitates were examined by Fourier-transform infrared
of the control specimen at 28 days, the use of bacteria in this con-
spectroscopy (FT-IR). Infrared spectroscopy (IR) is used to determine the molecular
structure and identification of chemical species. FTIR measurement was performed
crete mixture has reduced this reduction in compressive strength
using an Avatar spectrometer (Thermo, USA). to 4%. The compressive strength of the bacterial specimen contain-
ing 20% limestone powder, without zeolite is about 4% more than
3. Results and discussion the compressive strength of non-bacterial specimen containing
the same percentage of limestone powder and zeolite at the age
3.1. Compressive strength of 28 days; and use of bacteria in specimen containing 30% lime-
stone powder enhances the compressive strength about 5.7% at
The results of the compressive strength are provided in Fig. 3. 28 days. While these values are 3.9% for the control specimen.
The compressive strength of nonbacterial concrete containing
varying percentages of limestone powder and zeolite was between
44.3 and 64.4 MPa at age of 28 days and between 50 and 65.8 MPa
at 60 days. According to the results, compressive strength of con-
crete mixtures containing limestone powder decreased with
increase in limestone powder content up to 30% as cement replace-
ment and concrete mixtures containing 10% zeolite as cement
replacement displayed higher compressive strength than speci-
mens containing the same percentage of limestone powder with-
out zeolite. The compressive strength of specimens containing
zeolite with increase in limestone powder up to 20% is more than
the compressive strength of the control specimen (C-0-0), but the
specimen containing 30% limestone powder and 10% zeolite
(C-30-10) displayed lower compressive strength than the control
specimen (C-0-0). According to the results, the compressive
strength of specimens containing varying percentages of limestone
powder and zeolite with bacteria cells was between 46.9 MPa and
65.7 MPa at age of 28 days and between 51.5 MPa and 69.8 MPa at
60 days. Use of bacteria cells in the mixtures resulted in increase in Fig. 4. Tensile strength of specimens at 28 days.

Fig. 3. Compressive strength of 100 mm cubic specimens; (a) at 28 days, (b) at 60 days.
 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059 5

Therefore, the effect of using bacteria on increasing the compres-
sive strength of specimens containing 20% and 30% limestone pow-
der is slightly higher than the control specimen. The variations in
compressive strength of specimens at 60 days are similar to the
observed change at the age of 28 days.
3.2. Split tensile test
The test results are illustrated in Fig. 4. According to the results,
the trend of tensile strength variations of the specimens is similar
to the compressive strength changes. The results show that the
presence of bacteria in mixture design has increased the tensile
strength between 0.37 MPa and 0.77 MPa. The tensile strength of
the bacterial specimen containing 10% limestone powder and
10% zeolite (S-10-10) is similar to the tensile strength of the con-
trol specimen (C-0-0). It means that by using bacteria in concrete
mixture and reducing 20% cement weight, the tensile strength
can be similar to that of the control specimen. The tensile strength
of the bacterial specimen without limestone powder and zeolite is
about 10% higher than the tensile strength of the control specimen.

Fig. 5. Electrical resistivity of specimens; (a) at 28 days, (b) at 60 days.

Fig. 6. Relationship between electrical resistivity and compressive strength of specimens; (a) at 28 days, (b) at 60 days.

Fig. 7. Ultrasonic pulse velocity of specimens; (a) at 28 days, (b) at 60 days.

6 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059

Fig. 8. Relationship between ultrasonic pulse velocity and compressive strength of specimens; (a) at 28 days, (b) at 60 days.

In fact, by filling the pores of the concrete matrix with calcite pre- specimens made with different percentages of limestone powder
cipitations, the tensile strength of the specimens has increased to and zeolite. The high value of coefficient R2 indicates a good rela-
some extent compared to similar specimens without any bacteria. tionship between these two parameters.

3.4. Ultrasonic pulse velocity

3.3. Electrical resistivity
Ultrasonic pulse velocity of specimens is shown in Fig. 7.
Fig. 5 illustrates the electrical resistivity of all specimens at the
According to the results, the ultrasonic pulse velocity of the bacte-
ages of 28 and 60 days. According to the results electrical resistiv-
rial specimen without zeolite and limestone powder (S-0-0) is
ity of specimens containing limestone powder decreased with
about 0.98% and 0.93% more than the ultrasonic pulse velocity of
increase in limestone powder as cement replacement. As the elec-
the control specimen (C-0-0) respectively, at the ages of 28 and
trical resistivity of specimen containing 30% limestone powder is
60 days. The use of zeolite also increases the ultrasonic pulse veloc-
about 39.7% and 38.4% less than the electrical resistivity of the con-
ity of the specimens. So that the ultrasonic pulse velocity of the
trol specimen at ages of 28 and 60 days, respectively. While replac-
nonbacterial specimen containing 10% zeolite (C-0-10) is about
ing 10% of cement weight with zeolite, has led to a significant
2.1% and 0.37% more than the ultrasonic pulse velocity of the con-
increase in the electrical resistivity of the specimens. So that the
trol specimen (C-0-0) respectively, at 28 and 60 days. The use of
electrical resistivity of the non-bacterial specimen containing 10%
limestone powder as cement replacement has reduced the ultra-
zeolite (C-0-10) is 53% higher than the electrical resistivity of the
sonic pulse velocity of specimens. The lowest value of ultrasonic
control specimen (C-0-0) at age of 28 days. According to the
pulse velocity is related to the specimen containing 30% limestone
results, the electrical resistivity of the mixtures containing 20%
powder (C-30-0). So that the ultrasonic pulse velocity of this spec-
and 30% limestone powder with 10% zeolite (C-20-10 and
imen is about 7% and 10.2% less than the ultrasonic pules velocity
C-30-10) is approximately equal to the electrical resistivity of the
of the control specimen at 28 and 60 days. The highest rate of the
control specimen (C-0-0). Thus the negative effect of cement sub-
ultrasonic pulse velocity is related to the bacterial specimen con-
stitution with lime stone powder (i.e. increasing W/C) can be com-
taining 10% zeolite (S-0-10). The ultrasonic pulse velocity of this
pensated by adding 10% zeolite to the mix as well. According to the
specimen is about 3.7% and 2% more than the ultrasonic pules
results, filling the pores by microbial calcite precipitation increases
velocity of the control specimen respectively at 28 and 60 days.
the electrical resistance of all the specimens. The bacterial speci-
men containing 10% zeolite (S-0-10), displayed highest electrical
resistance (180.5 X.m and 208.7 X.m respectively at 28 and
60 days) among all specimens. Use of bacteria has increased the
electrical resistivity of the control specimen by 6.7% at 28 days.
Also, the electrical resistivity of bacterial specimens containing
10%, 20% and 30% are about 11.4%, 11.6% and 18%, respectively,
more than nonbacterial specimens of similar mixtures at 28 days.
This increment for the specimen containing 10% zeolite is about
7.5% and for the specimens containing 10% zeolite with 10%, 20%
and 30% limestone powder are about 13%, 21% and 17.5% respec-
tively at 28 days. This increment for both of the specimens without
limestone powder and zeolite and without limestone powder, con-
taining 10% zeolite is 6.3% at 60 days. While this increment is
higher for the specimens containing limestone powder. So that
the bacterial specimen containing 30% limestone powder has the
highest increase and this amount is 14% for this specimen. This
indicates that the use of bacteria in the specimens containing lime-
stone powder has been more effective in increasing the electrical
resistivity. Fig. 6 demonstrates the relationship between electrical Fig. 9. Relationship between ultrasonic pulse velocity and tensile strength of
resistivity and compressive strength of bacterial and non-bacterial specimens at 28 days.
 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059 7

Fig. 10. SEM images of (a) C-0-0; (b) S-0-0; (c) C-0-10; (d) S-0-10; (e) C-30-0, (f) S-30-0.

The ultrasonic pulse velocity of the nonbacterial specimen contain- has caused the ultrasonic pulse velocity to be 0.98% higher than
ing 10% zeolite and 10% limestone powder (C-10-10) is 0.98% less the control specimen.
than the ultrasonic pulse velocity of the control specimen (C-0-0) The relationships between compressive strength and ultrasonic
at 28 days. While, adding the bacteria to this concrete mixture pulse velocity of bacterial specimens and non-bacterial specimens
8 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059

Fig. 11. Images of crack healing processes of specimens after different repair time, (a) bacterial specimen without limestone powder and zeolite (S-0-0); (b) bacterial
specimen contains 20% limestone powder and 10% zeolite (S-20-10); (c) crack healing by using bacteria in curing environment (S-CE).

at the age of 28 and 60 days are presented in Fig. 8. The relation- (S-0-10) provides dense structure than the nonbacterial specimen
ship between tensile strength and ultrasonic pulse velocity of with the same mixture design (C-0-10). The SEM image of the
specimens is illustrated Fig. 9. specimen containing 30% limestone powder (Fig. 10e, f) revealed
that the amount of calcite crystals in the bacterial treated speci-
men containing limestone powder (S-30-0) is more than the
3.5. Microstructure of concrete specimens
amount of crystals formed in the bacterial specimen without lime-
stone powder (S-0-0). When the bacteria were exposed to lime-
Fig. 10a, c, e refer to the non-bacterial mixtures (without lime-
stone powder, they had better activities in order to produce
stone powder and zeolite (C-0-0), containing 30% limestone pow-
calcite precipitations.
der (C-30-0) and containing 10% zeolite (C-0-10)) and the images
Fig. 11 illustrates direct observation of cracks healing of speci-
of bacterial specimens (without limestone powder and zeolite
mens. As can be seen in this figure, compared with the specimens
(S-0-0), containing 30% limestone powder (S-30-0) and containing
that bacteria were used in their mixture design, the amount of sed-
10% zeolite (S-0-10)) are presented in Fig. 10b, d, f. The images of
iments in the cracks of the specimen cured in the suspension of
the specimens indicate the presence of calcite precipitation in
bacteria and the reactive solution are much higher. Fig. 12 shows
bacterial specimens. It is obvious that the voids of the bacterial
the microscopic images of crack healing processes of three differ-
specimens are filled by calcite precipitations. Fig. 10a relates to
ent specimens. Fig. 12a refers to the crack healing of the bacterial
the control specimen without limestone powder and zeolite
specimen without limestone powder and zeolite (S-0-0) with a
(C-0-0). The existence of voids in the structure of this specimen
width of 0.28 mm that was nearly fully healed after 30 days and
is clearly evident. As shown in Fig. 10b, calcite precipitations cre-
Fig. 12b shows the crack healing of the bacterial specimen contain-
ated by bacteria have filled up the voids. Fig. 10c, d shows the bac-
ing zeolite and lime stone powder (S-20-10) with a width of
terial and nonbacterial specimen containing 10% zeolite. The
0.34 mm. According to the images, after 28 days of healing, this
presence of a bacteria in the mixture design containing 10% zeolite
 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059 9

Fig. 12. Microscopic image of healing the cracks of different specimens with different widths during the time, (a) bacterial specimen without limestone powder and zeolite
(S-0-0); (b) bacterial specimen contains 20% limestone powder and 10% zeolite (S-20-10); (c, d) crack healing by using bacteria in curing environment (S-CE).

crack was completely healed. In Fig. 12c and d a crack treated with
using bacteria in curing environment is shown (S-CE). As it can be
seen in Fig. 12c, completely healing of the crack with the width of
0.4 mm was at earlier healing time (5 days), while the crack with a
width of 1.1 mm was only partially healed after 28 days (Fig. 12d).
Fig. 13 illustrates the crack healing percentage of specimens at
different healing times. As seen from this figure, the presence of
bacteria in curing environment (S-CE) has the best healing results
among the other specimens. It can be seen that the trend line of the
crack healing of the specimen containing limestone powder and
zeolite is slightly higher than the specimen without limestone
powder and zeolite. So, apparently the cracks of the specimen con-
tains 20% limestone powder and 10% zeolite (S-20-10) seem to be
healed better than the specimen without limestone and zeolite
(S-0-0). As shown in Figs. 14, 15 all the cracks up to 0.58 mm were
completely healed at 30 days, when the bacteria have been used in
10 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059

Fig. 13. Crack healing percentage of different specimens as a function of the initial crack width after curing time of (a) 5 days (b) 15 days and (c) 30 days.

Fig. 15. Maximum width of healed crack.

Fig. 14. Maximum width of 100% healed crack.

curing environment and complete crack healing for the widths up
to 1.1 mm were achieved for this specimen at 30 days. This values
are respectively 0.16 and 0.42 for both of the specimens that bac-
teria have been used in their mixture design (S-0-0 and S-20-10).
3.6. Microstructure of cracks filler
The carbonates have three to four intense bands in Infrared
spectroscopy (IR) region. The 1420 cm1 and 876 cm1 region
bands are characteristic for identification of carbonates and the
711 cm1 region band is characteristic of calcite among various
carbonate rocks [36]. The results of Fourier-Transform Infrared
spectroscopy of the precipitation at cracks surface show in
Fig. 16. Existence of peaks at 1423 and 874 induced by the pres-
ence of carbonate and the 710 cm1 region band shows that the
precipitation at cracks surface is calcite which matched with the
mineralization by bacteria. The SEM image of the precipitation cre-
ated on the surface of the cracks is presented in Fig. 17.
 M.S. Jafarnia et al. / Construction and Building Materials 242 (2020) 118059
Fig. 16. Details of the Fourier-Transform Infrared spectroscopy of the precipitation
formed at cracks surface.
Fig. 17. Details of the SEM image of the precipitation formed at cracks surface.
4. Conclusions

Calcite precipitations in bacterial specimens increased com-
pressive strength, tensile strength, ultrasonic pulse velocity
and electrical resistivity of all specimens.

Compressive strength, tensile strength, ultrasonic pulse velocity
and electrical resistivity of all specimens decreased by increas-
ing the percentage of limestone powder replacement of cement.
But this decrease for specimens containing bacteria is lower
than nonbacterial specimens.

Partial replacement of cement with zeolite significantly
increased compressive strength and electrical resistivity of the
specimens. For example, when 10% zeolite is used, the 28-
days compressive strength of the non-bacterial specimen
increased by 20% and that of bacterial specimen increased by
23%. These values for 60-days specimen without bacteria was
11% and with bacteria was 18%. Furthermore, the electrical
resistivity of the specimen containing 10% zeolite is about by
11
53% and 31% higher than the electrical resistivity of the control
specimen at 28 and 60 days, respectively. This values for the
bacterial specimen containing 10% zeolite improves to values
of 64% and 39%.

According to the SEM images, the use of limestone powder has a
positive effect on the formation of calcite crystals in the
microstructure of the bacterial specimens. This effect is also
reflected in the average percentage of crack healing. As the
average percentage of crack healing of the specimen containing
limestone powder is slightly higher than the specimen without
limestone powder.

The percentage of crack healing by using the bacteria in curing
environment is significantly higher than the percentage of crack
healing by using the bacteria in concrete mixture design. All the
cracks up to 0.58 mm wide were completely healed at 30 days
by using bacteria in curing environment, while if bacteria are
used in concrete mixture only cracks with up to 0.16 mm width
were completely healed.

From the findings, considering a curing aqua containing bacte-
ria for applying to the cracked surfaces of a floor slab is sug-
gested. Using bacteria in the wet curing environment is very
promising method for healing cracks.
CRediT authorship contribution statement
Maedeh Sadat Jafarnia: Investigation, Writing - original draft.
Mehdi Khodadad Saryazdi: Project administration, Validation,
Writing - review & editing. Seyed Mohammad Moshtaghioun:
Writing - review & editing.
Declaration of Competing Interest
The authors declare that they have no known competing finan-
cial interests or personal relationships that could have appeared
to influence the work reported in this paper.
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