American Journal of Applied Science and Technology
12
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VOLUME
Vol.05 Issue 07 2025
PAGE NO.
12-17
10.37547/ajast/Volume05Issue07-03
The Comparative Analysis of Indigenous Soil Bacteria
and Their Oil-Degrading Potential in Certain Areas of
the “Jarkurgan
-
Neft” Oil Fields
Abdullaeva Yulduz Alisher qizi
PhD Student Institute of Microbiology, Uzbekistan Academy of Sciences, Uzbekistan
Ahmedova Zakhro Rakhmatovna
Head of Laboratory, Doctor of Biological Sciences, Professor, Institute of Microbiology, Uzbekistan Academy of Sciences, Uzbekistan
Received:
15 May 2025;
Accepted:
11 June 2025;
Published:
13 July 2025
Abstract:
This article investigates the oil-degrading potential of indigenous bacterial strains isolated from various
oil-contaminated soils and products in the Jarkurgan and Kumkurgan districts of Surkhandarya region. Based on the
analysis of the quantity and quality of microorganisms present in collected samples of soil, oil sludge, crude oil, and
water, 24 pure bacterial isolates
—
predominantly occurring species
—
were isolated. These strains were
comparatively characterized in terms of their growth and oil-degrading activity in a minimal medium supplemented
with crude oil as the sole carbon source. Additionally, nine bacterial isolates exhibiting high oil-degrading potential
were studied under conditions of Raimondo synthetic medium containing 1.0
–
2.0% crude oil, focusing on their
morphological characteristics and hydrocarbon utilization efficiency.
Keywords:
Indigenous bacteria, oil-contaminated soil, bioremediation, crude oil, Raimondo nutrient medium,
microbiota, bacteria, assimilation, growth rate, activity, oil degradation.
Introduction:
With population growth and industrial development,
the demand for fossil fuels has increased, leading to
the expansion of oil and gas industry activities. The
processes
of
oil
extraction,
refining,
and
transportation generate waste that contains large
amounts of toxic substances, which have a significant
impact on the environment, particularly the soil
ecosystem. Pollution can spread not only locally but
also over long distances. As a result of such
contamination, the stability of the soil biocenosis
—
especially the microbial communities
—
is disrupted,
leading to ecological imbalance [1]. Pollution
resulting from oil extraction can spread through soil
and groundwater at various depths, thereby
exacerbating environmental problems. Petroleum
products are among the most significant chemical
pollutants of essential natural components such as
soil, water, and air. These substances contribute to
the phytotoxicity of soil and hinder plant
development [2].
Crude oil is a liquid fossil fuel composed of various
high-molecular-weight
hydrocarbons,
and
its
presence in the soil environment leads to alterations
in the microbiota. Although high concentrations of oil
can cause the death of many microorganisms, over
time,
microbial
communities
specialized
in
hydrocarbon degradation may develop, thereby
enhancing
bioremediation
processes
[3].
Microorganisms capable of oxidatively degrading
hydrocarbons can remain active for extended periods
in oil-contaminated environments. Even when
residual oil concentrations are below 10 g/kg, these
microorganisms retain their biological activity. The
presence
of
bacteria
capable
of
utilizing
hydrocarbons as the sole source of carbon and energy
was first discovered
approximately 80 еars ago [4].
However, high concentrations of oil (1
–
30 ml/kg)
negatively affect the microbiological and ecological
stability of the soil, disrupting its water-air balance.
Microorganisms are ancient and widespread life
American Journal of Applied Science and Technology
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American Journal of Applied Science and Technology (ISSN: 2771-2745)
forms that play a crucial role in maintaining metabolic
balance in natural environments. They regulate redox
processes in the soil and, due to their sensitivity to
environmental changes, are considered reliable
bioindicators. According to research findings, when
the concentration of petroleum hydrocarbons in soil
exceeds 10%, it has a detrimental effect on the
number of microorganisms [5].
However, low concentrations of oil can stimulate
microbial growth, as oil-degrading microorganisms
become activated. Oil also affects the activity of soil
enzymes in different ways: in some cases, it enhances
enzymatic activity, while in others, it suppresses it [6].
This effect depends on the type of substance and the
properties of the soil. For instance, oil-contaminated
soil often develops anaerobic conditions, which
significantly alter the activity of soil enzymes
—
particularly oxidoreductases such as ferric reductase
and catalase [7]. Once oil enters the soil, processes of
fractionation and degradation begin. The efficiency of
these processes depends on the enzymatic activity of
the microbial communities present in the soil.
Microorganisms
capable
of
fully
oxidizing
hydrocarbons to CO₂ and water (mineralization) are
widespread in natural ecosystems, and their activity
is largely dependent on the presence of oxidase-
group enzymes [8]. The sensitivity of enzyme activity
can be ranked as follows: ferric reductase > catalase
> urease > invertase [7]. The primary microorganisms
responsible for hydrocarbon degradation in soil are
bacteria
and
certain
fungi.
These
include
representatives of the genera Pseudomonas,
Arthrobacter,
Mycobacterium,
Rhodococcus,
Bacillus, Micrococcus, Klebsiella, among others [9].
This research is dedicated to studying the
composition of the microflora present in the soils and
certain products of the "Jarkurgan-Neft" area,
isolating
and
purifying
active
strains,
and
investigating representatives with high oil-degrading
potential.
METHODOLOGY
Soil samples were collected for microbiological
analysis from oil-contaminated areas of the
“Petromaruz
-
Uzbekistan” LLC site in Jarkurgan district
and the “Lalmikor Neft” oil field in Kumkurgan district
of Surkhandarya region. Sampling was carried out
using a grid method from a depth of 0
–
20 cm in
accordance with GOST 17.4.4.02
–
84 and R 54039
–
2010 standards. Additionally, samples of oil-
separated water, oil sludge, and crude oil were
selected as research objects.
Under laboratory conditions, the soil samples were
cleaned and passed through a 2 mm mesh sieve. From
each sample, 10 g of soil was taken and serially
diluted in 90 ml of sterile water, then inoculated onto
sterilized selective nutrient media. The oil sludge
samples were processed in the same way, while the
water samples were inoculated directly. As nutrient
media, meat-peptone broth was used for bacteria,
and Czapek medium was used for microscopic fungi.
The diluted samples were inoculated using the deep
plating method and incubated in a thermostat at 25
–
30 °C for 3–
8 days [10]. Based on growth rate,
morphological characteristics, quantity, and quality
indicators, a total of 24 pure bacterial isolates were
obtained from the studied samples. To assess the
viability,
growth
activity,
and
hydrocarbon
assimilation and degradation abilities of the isolated
strains in an oil-containing environment
—
and to
select promising strains for bioremediation
—
a solid
artificial Raimondo nutrient medium was prepared.
The medium consisted of the following components
(g/L): Na₂CO₃ –
0.1; CaCl₂ –
0.1; MnSO₄ –
0.02; FeSO₄
–
0.01; MgSO
₄ –
0.2; NH₄Cl –
0.2; NaCl
–
3.0; Na₂HPO₄
–
1.5; KH₂PO₄ –
1.0. To this, 1% crude oil and 2% agar
were added to prepare a solid medium. The medium
was sterilized in an autoclave at 120 °C under 1
atmosphere of pressure for 30 minutes. After
sterilization, the medium was poured into Petri dishes
and allowed to solidify. Four bacterial strains were
inoculated into each dish, which were then placed in
a thermostat at 37 °C to begin the incubation process.
The growth rate of the strains and their colony-
forming activity in the presence of crude oil were
observed.
A qualitative analysis was then conducted on the
strains, and based on growth indicators, 14 out of the
24 isolated strains were selected. Subsequently, a
new solid Raimondo nutrient medium was prepared
by adding 2% crude oil and 2% agar. The medium was
sterilized in an autoclave at 120 °C under 1
atmosphere of pressure for 30 minutes. After
sterilization, it was poured into Petri dishes. Four
bacterial strains were inoculated into each dish,
which were t
hen placed in a thermostat at 37 °C, and
monitoring was carried out.
RESULTS AND DISCUSSION
According to the results of the study, 24 bacterial
strains were incubated on solid nutrient medium
containing 1.0% crude oil over varying time intervals
(24
–
360 hours), and their growth was evaluated.
Fourteen strains were individually inoculated into
sectors of Petri dishes. After 5 days of incubation at
37 °C, the growth intensity and colony
-forming
characteristics of the strains were visually assessed.
Based on the size of the growth and assimilation
zones in the 1.0% oil-containing medium, the activity
American Journal of Applied Science and Technology
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American Journal of Applied Science and Technology (ISSN: 2771-2745)
levels of the isolates obtained from soils in different
zones of the “Jarkurgan
-
Neft” site varied.
01, 02, 03, 04
05, 06, 07, 08
09, 10, 11, 12
13, 14, 15, 16
17, 18, 19, 20
21, 22, 23, 24
Figure 1.
Oil assimilation levels of selected bacterial isolates obtained from the
Jarkurgan oil fields on Raimondo nutrient medium supplemented with 1.0% crude
oil. *Note: 01–24 refer to isolate numbers.
Early growth and oil assimilation were observed at
48
–
72 hours post-inoculation in the following
isolates: JN-01, JN-04, JN-05, JN-06, JN-07, JN-13, JN-
15, JN-18, JN-19, and JN-20. These isolates
demonstrated good adaptability to the hydrocarbon-
enriched medium, along with high activity and
relatively strong oil-degrading capacity. Moderate-
phase growth (96
–
168 hours) was observed in strains
JN-08, JN-09, and JN-10. Although these strains
became active more slowly, they still exhibited stable
growth. Strains showing high growth activity
(indicated in blue and dark blue): JN-04, JN-06, JN-07,
JN-13, JN-18, and JN-20 demonstrated the highest
growth intensity (0.7
–
0.8) and were identified as
having significant potential for bioremediation
applications.
American Journal of Applied Science and Technology
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American Journal of Applied Science and Technology (ISSN: 2771-2745)
Figure 2.
Growth and comparative oil assimilation indicators of 20 bacterial isolates
obtained from soils of various zones of the “Jarkurgan-Neft” (JN) oil fields on oil-
containing nutrient medium.
For each strain, the onset time of growth and growth
intensity were monitored, and a total of 14 active
strains were selected. These strains were re-
incubated
on
Raimondo
nutrient
medium
supplemented with 2% crude oil, and growth was
assessed in a second phase. After 48 hours of
incubation, clear signs of growth were observed in
strains JN
–
01, JN
–
04, JN
–
06, JN
–
10, JN
–
13, JN
–
15,
JN
–
18, and JN
–
20. Additionally, growth was recorded
in strain JN
–
05 after 144 hours of incubation. Based
on growth intensity and stability at the end of the
incubation period, a total of 9 bacterial strains were
identified as highly active. The morphological
characteristics of these strains were then examined
using microscopy.
JN - 1
JN - 4
JN - 5
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American Journal of Applied Science and Technology (ISSN: 2771-2745)
JN - 6
JN - 10
JN - 13
JN - 15
JN - 18
JN - 20
Figure 3.
Microscopic images of the most active bacterial isolates obtained
from oil-contaminated soils. JN – Jarkurgan-Neft.
Microscopic examination of the isolates was
performed using Gram staining (1×1000). The strains
exhibited morphological diversity, with the majority
identified as Gram-negative rods and coccus-shaped
forms. Some strains formed densely clustered
colonies in the oil-containing medium, indicating their
strong growth and assimilation activity in the
presence of crude oil.
Table 1. – Protein synthesis dynamics by bacterial isolates in oil-containing liquid
nutrient medium (mg/ml).
Isolated strains
Protein production dynamics (mg/ml) during growth over time
(hours)
48
72
96
120
JN - 01
4,0
3,0
3.5
2.5
JN - 04
2.3
4,0
4.3
4,0
JN - 05
2,0
3.9
4.3
2.8
JN - 06
1.8
4,0
4,0
3.5
JN - 10
1.3
3.6
2.8
2.3
JN - 13
5.5
4.5
4,0
2.8
JN - 15
2.3
3.8
3.3
3.5
JN - 18
3.3
3.9
4
4.3
JN - 20
2.8
3,0
2.8
2.8
The growth activity of bacterial strains in liquid
nutrient medium supplemented with 1% crude oil
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American Journal of Applied Science and Technology (ISSN: 2771-2745)
was evaluated at different incubation times.
According to the results, some strains began active
growth as early as 48 hours (e.g., JN
–
13
—
5.5
mg/ml), while others reached their maximum growth
levels at 72
–
96 hours (e.g., JN
–
04, J.N
–
18). Notably,
strain JN
–
18 exhibited a growth level of 4.3 mg/ml at
120 hours. These findings indicate differences in the
strains’ adaptability to crude oil and their
hydrocarbon assimilation activity. During incubation,
most strains showed an initial increase in growth,
followed by stabilization or a slight decline. This trend
indicates that the substrate in the nutrient medium
gradually approached saturation, leading to
limitations in microbial activity.
CONCLUSION
The conducted research confirmed the presence of
indigenous bacteria in oil-contaminated soils
surrounding the “Jarkurgan
-Ne
ft” oil field and
revealed their high adaptability to petroleum-
polluted environments. A total of 24 bacterial isolates
were obtained, of which 14 demonstrated active
growth in media containing 1
–
2% crude oil. Among
them, 9 strains stood out with high growth intensity
and strong oil-degrading capabilities. Their growth
dynamics on Raimondo nutrient medium and visible
colony formation indicated not only efficient
hydrocarbon assimilation but also stable survival in
oil-rich environments.
Furthermore, experiments conducted in oil-
containing liquid media recorded protein synthesis
dynamics over various incubation periods. Notably,
strains such as JN
–
13 and JN
–
18 exhibited the highest
protein production levels, reflecting their elevated
metabolic activity and relevance for bioremediation
processes.
These results confirm the potential to isolate effective
oil-degrading strains from local microflora and
develop environmentally friendly biotechnological
methods and biopreparations based on them. In the
future, it is advisable to investigate the genomic
structure, enzymatic activity, and interactions of
these strains with plants.
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