ABSTRACT
The evaluation of the physicochemical parameters, biosurfactant production and metagenomic analysis of hydrocarbon polluted soil in Opuama, Ogini, Oteghele, Transforcados pipeline and Transforcados right of way sites located at Isoko North, Warri North and South LGA of Delta State Nigeria, was carried out using standard microbiological and physicochemical methods. Hydrocarbon utilizing bacteria (HUB), total heterotrophic bacterial count (THB), physicochemical analysis including heavy metals, screening and characterization, optimization of biosurfactant using response surface methodology (RSM) and metagenomic analysis were all carried out. The DNA of the microorganisms from the soil samples were extracted using ZymoDNA extraction kit, amplified and subjected to next generational sequencing (NGS) on Pacbio SMR sequencing platform. Following NGS, gene calling was performed using freg gene scan and the resulting metagenome were then functionally annotated onto two pipelines namely: cluster of orthology group (COG) and Kyoto encyclopedia of genes and genomes (KEGG) for functional gene analysis. The bacteria screened and characterized for biosurfactant production were Escherichia coli, Bacillus subtilis, Enterobacter aerogenes, Klebsiella pneumoniae, Pseudomonas aeruginosa, Providencia stuartii and Burkholderia pseudomallei. All of the screened bacteria, Bacillus subtilis indicated high level of biosurfactant production and was subsequently subjected to optimization for biosurfactant production. The emulsification index (E1%) due to change in temperature and pH on glucose/KN03 is 77.28, sucrose/KN03 is 59.80 and spent vegetable oil/KN03 is 57.16. The mean value for hydrocarbon utilizing bacteria and total heterotrophic bacterial count were 1.5×105 cfu/g and 3.76×108 cfu/g for Opuama flow station, 5.8×104 cfu/g and 2.60×108 cfu/g for Ogini flow station, 2.80×105 cfu/g and 2.55×108 cfu/g for Oteghele river spill, 4.2×104 cfu/g and 3.04×108 cfu/g for Transforcados pipeline spill and 1.72×105 cfu/g and 2.75×108 cfu/g for Transfoecados right of way spill. Oteghele river spill had the highest concentration of crude oil utilization bacteria 2.80×105 cfu/g followed by Transforcados right of way spill 1.72×105 cfu/g while Opuama flow station had the highest concentration of heterotrophic bacteria 3.76×108 cfu/g. The performance of the pathogen for hemolytic reaction on blood agar, oil spreading on glass slide, blue plate agar reaction, emulsification index and lypolytic enzyme reaction indicated that they were significant at (p<0.05). There was significant difference in the physicochemical parameters at (p<0.05). The mean values of the pH range from 4.15d±0.1-5.75b±0.1, electrical conductivity 1179.61e±7.51- 2079.55b±1.45. Some of the heavy metals were higher in value when compared with the WHO standard. The structural metagenomics revealed eight top phyla; Proteobacteria, Actinobacteriota, Acidobacteriota, Bacteroidota, Planctomycetota, Firmicutes and Cloroflexi. The functional gene analysis revealed Carbohydrate metabolism, energy metaboilism, environmental information processing, degradation genes and various protein classes in the samples as contained in the KEGG and COG pipelines. Giving the findings of this study microorganisms found in petroleum hydrocarbon polluted soil were able to produce biosurfactants which helped in biodegradation.
TABLE OF CONTENTS
TITLE
PAGE i
DECLARATION ii
CERTIFICATION iii
DEDICATION iv
ACKNOWLEDGEMENTS v
TABLE
OF CONTENTS vi
LIST
OF TABLES xii
LIST
OF FIGURES xv
ABSTRACT xix
CHAPTER 1: INTRODUCTION
1.1
Background of the Study 1
1.2
Justification 6
1.3
Aim and Objectives of the
Study 6
1.4
Significance of the Study 7
CHAPTER 2: LITERATURE REVIEW
2.1
Exploration of Petroleum Hydrocarbon in
Nigeria 8
2.2
Origin of Petroleum Hydrocarbon 8
2.3
Petroleum
Hydrocarbon Formation in Various Depth Zones 9
2.4
Formation and Types of Kerogen 10
2.5
Chemical
Composition of Petroleum Hydrocarbon (crude oil) 11
2.6
Crude
Oil Classification 12
2.7
Petroleum
Biotechnology 13
2.8
Factors Affecting Hydrocarbon Degradation 13
2.9
Degradation Mechanism of Petroleum
Hydrocarbon 15
2.10
Biosurfactants
Biotechnology
18
2.11
Biosurfactants
Classification 19
2.12
Properties of Biosurfactant 27
2.13
Mechanisms
of Interaction 27
2.14
Applications
of Biosurfactants 30
2.14.1
Microbial Enhanced Oil Recovery 30
2.14.2
Cleanup of Spilled Oil 30
2.14.3 Antimicrobial Activity 30
2.14.4 Medical and Therapeutic Applications 30
2.14.5 Anti-adhesive Agents 31
2.14.6 Control of Disease and Plant Pathogen 31
2.14.7 Applications in Food Processing 31
2.15 Advantages of Biosurfactants 32
2.16 Disadvantages of Biosurfactants 33
CHAPTER 3: MATERIALS AND
METHODS
3.1 Study
Site 35
3.2 Sample
Collection 35
3.2.1 Preparation
of Soil Samples for Serial Dilution 38
3.3 Microbiological Analysis 38
3.3.1 Estimation of Hydrocarbon Utilizing Bacteria
Count (HUB) of 38
the Soil Samples
A-E
3.3.2 Estimation of Total Heterotrophic Bacterial
Count (THB) of 39
the Soil Samples
A-E
3.3.3 Characterization
and Identification of Bacterial Isolates for 39
the Soil
Samples A-E
3.4 Physicochemical
Analysis 39
3.4.1 Determination
of pH 40
3.4.2 Determination
of Conductivity and Redox Potential 40
3.4.3 Determination
of Soil Total Nitrogen 40
3.4.4 Determination
of Total Phosphorus Content 41
3.4.5 Determination of Percentage Total Organic Carbon 41
3.4.6 Determination
of Total Hydrocarbon Content 42
3.4.7 Determination
of Total Petroleum Hydrocarbon 42
3.4.7.1 Preparation of Soil Sample Extracts 43
3.4.8 Digestion
of Soil Samples for Metal Analysis 43
3.4.9 Extraction of Cations in the Soil Samples 43
3.4.9.1 Determination of Exchangeable Cations 44
3.4.10 Determination of Potassium and Sodium 44
3.4.11 Determination of Calcium and Magnesium 44
3.4.12 Determination of Exchangeable Acidity 45
3.4.13 Titration for Hydrogen ion (H+) 45
3.4.14 Determination
of Effective Cation Exchange Capacity (ECEC) 46
3.4.15 Determination of Soil Base Saturation 46
3.4.16 Determination of Heavy Metals 46
3.5 Screening of Biosurfactant Producing
Bacteria for the Soil Samples A- E 47
3.5.1 Blood Agar Reaction (Haemolysis) 47
3.5.2 Oil Spreading on Glass Plate 47
3.5.3 Blue Agar Plate Reaction 47
3.5.4 Measurement of Emulsification Index (E24) 48
3.5.5 Lipolytic Enzyme Production 48
3.6 Optimization
of Biosurfactant Producing Microorganisms Using 48
Response Surface Methodology (RSM) for the
Isolate
3.6.1 Inoculum
Development of the Isolate Used for Optimization 49
3.7 Metagenomic Analysis of the Soil Samples
A-E 49
3.7.1 Bacterial
DNA Isolation from Samples 50
3.7.2 Lysis of the Samples 50
3.7.3 Binding to Column 50
3.7.4 Column
Wash 51
3.7.5 DNA
Elution 51
3.7.6 DNA Amplification 51
3.7.7 Metagenomic
Sequencing of DNA 51
3.7.8 Functional
Analysis of Sequence Reads 52
3.8 Molecular
Analysis 52
3.8.1 Extraction
of DNA 53
3.8.2 PCR
Reaction 54
3.9 Heat
Map 54
3.10 Experimental
Framework and Data Analysis 55
CHAPTER
4: RESULTS AND DISCUSSION
4.1 Results 56
4.1.1 Microbiological Characteristics of the Soil Samples 56
4.1.2 Physicochemical
Analysis of the Soil Samples (Sample A-E) 60
4.1.3 Screening of Biosurfactant Producing Bacteria
from the Soil Samples 64
4.1.4 Optimization
Using Response Surface Methodology 66
4.1.5 Structural
Metagenomics Analaysis of the Soil Samples 79
4.1.6 Funtional
Gene Analysis 123
4.1.7 PCR
Reaction and Gel Electrophoresis 123
4.2 Discussion
of Findings 162
4.2.1 Microbiological
Analysis 162
4.2.2 Biosurfactant
Producing Bacteria Analysis 164
4.2.3 Optimization
of Biosurfactant Producing Bacteria by (RSM) 165
4.2.4 Physicochemical
Analysis 166
4.2.5 Metals Analysis 170
4.2.6 Structural
Metagenomics Analysis 171
4.2.6 Functional
Gene Analysis 181
CHAPTER 5: CONCLUSION AND RECOMMENDATIONS
5.1 Conclusion 184
5.2 Recommendation 185
5.3 Contributions to Knowledge 186
REFERENCES
APPENDICES
LISTS OF TABLES
Table Title page
2.1 Classification of
biosurfactant and important microorganisms 26
4.1 Cell morphology and biochemical
identification of isolates 59
4.2 Physicochemical
properties of the different soil samples 62
(sample A-E)
4.3 Metal
properties of the different soil samples (sample A-E) 63
4.4 Performance
of the bacterial isolate based on different screening 65
Methods
4.5 Carbohydrate metabolism found in Opuama flow
station sample 124
4.6 Energy metabolism found in Opuama flow station
sample 125
4.7 Environmental
information processing found in Opuama flow 126
station
4.8 Degradation genes found in Opuama flow station
sample 127
4.9 Carbohydrate metabolism found in Ogini flow station
sample 128
4.10 Energy metabolism found in Ogini flow station sample 129
4.11 Environmental
information processing found in Ogini flow 130
station sample
4.12
Degradation genes found in
Ogini flow station sample 131
4.13 Carbohydrate metabolism found in Oteghele spill
sample 132
4.14 Energy metabolism found in Oteghele spill sample 133
4.15 Environmental
information processing found in Oteghele 134
spill
4.16 Degradation genes found in Oteghele spill sample 135
4.17 Carbohydrate
Metabolism found Transforcados pipeline 136
spill sample
4.18 Energy metabolism found in Transforcados pipeline spill sample 137
4.19 Environmental information processing found in Transforcados 138
pipeline spill sample
4.20 Degradation genes found in Transforcados pipeline spill sample 139
4.21 Carbohydrate
metabolism found in Transforcados right of 140
way spill
4.22 Energy
metabolism found in Transforcados right of way 141
spill sample
4.23 Environmental
information processing found in Transforcados 142
right of way spill sample
4.24 Degradation
genes found in Transforcados right of way spill sample143
4.25
Cluster of orthology group
(COG) class of Opuama flow 144
station
sample
4.26 Cluster of orthology group (COG)
families of Opuama flow 145
station sample
4.27 Phosphoribulokinase (PRK) of
Opuama flow station sample 146
4.28
Cluster of orthology group
(COG) class of Ogini flow 147
station
sample
4.29
Cluster of orthology group
(COG) families of Ogini flow 148
station
sample
4.30 Phosphoribulokinase
(PRK) of Ogini flow station sample 149
4.31
Cluster of orthology group
(COG) class of Oteghele 150
spill
sample
4.32
Cluster of orthology group
(COG) family of Oteghele 151
spill
sample
4.33 Phosphoribulokinase
(PRK) of Oteghele spill sample 152
4.34 Cluster of orthology group (COG)
class of Transforcados pipeline 153 spill sample
4.35 Cluster of orthology group (COG)
family of Transforcados 154
Pipieline spill
sample
4.36 Phosphoribulokinase
(PRK) of Transforcados pipeline spill sample 155
4.37 Cluster of orthology group (COG)
class of Transforcados right of 156
way spill sample
4.38 Cluster of orthology group
(COG) families of Transforcados right 157
of way spill
sample
4.39
Phosphoribulokinase
(PRK) of Transforcados right of way 158
spill
sample
LISTS
OF FIGURES
Figure Title page
2.1 Hydrocarbons
degradation by microorganisms 17
2.2 Examples of glycolipids and surfactin 22
2.3
Reaction of surface
tension and biosurfactant concentration 28
2.4 Biosurfactants
at water and air Interface 29
3.1 Map
of the study area 37
4.1 Hydrocarbon
utilizing bacterial count (HUB) of the 57
sampling sites (sample A-E)
4.2 Total heterotrophic bacteria
count (THB) of the sampling sites 58
(sample A-E)
4.3 The
distribution of Emulsification index (EI %) due to variation in 67
Temp and pH using glucose/KN03 as supplements
4.4 The distribution of
Emulsification index (EI %) due to variation 68
in Temp and pH using glucose/yeast extracts as
supplements
4.5 The
distribution of Emulsification index (EI %) due to variation in 69
Temp and pH using glucose/soy bean source as
supplements
4.6 The
distribution of Emulsification index (EI %) due to variation in 70
Temp and pH using glucose/locally prepared meat
extract as
supplements
4.7 The distribution of Emulsification
index (EI %) due to variation in 71
Temp and pH using sucrose/KN03 as
supplements
4.8 The
distribution of Emulsification index (EI %) due to variation in 72
Temp and pH using sucrose/yeast extract as
supplements
4.9 The
distribution of Emulsification index (EI %) due to variation in 73
Temp and pH using sucrose/soy bean extract as
supplements
4.10 The distribution of
Emulsification index (EI %) due to variation 74
in Temp and pH using
sucrose/locally produced meat extract as supplements
4.11 The
distribution of Emulsification index (EI %) due to 75
variation in Temp and pH using spent vegetable oil/KN03
as supplements
4.12 The
distribution of Emulsification index (EI %) due to variation 76
inTemp and pH using spent vegetable oil/yeast
extract as
supplements
4.13 The distribution of
Emulsification index (EI %) due to variation 77
inTemp and
pH using spent vegetable oil/soy beans source
as
supplements
4.14 The
distribution of Emulsification index (EI %) due to variation 78
inTemp and pH
using spent vegetable oil/ locally prepared
meat extract as supplements
4.15 Kingdom classification of Opuama
flow station 81
4.16 Top
phylum classification of Opuama flow station 82
4.17 Top
class classification of Opuama flow station 83
4.18
Top order classification
of Opuama flow station 84
4.19
Top family classification
of Opuama flow station 85
4.20
Top genus classification
of Opuama flow station 86
4.21
Top specie classification
of Opuama flow station 87
4.22 Top
kingdom classification of Ogini flow station 90
4.23 Top
phylum classification of Ogini flow station 91
4.24 Top
class classification of Ogini flow station 92
4.25 Top
order classification of Ogini flow station 93
4.26 Top
family classification of Ogini flow station 94
4.27 Top
genus classification of Ogini flow station 95
4.28 Top
species classification of Ogini flow station 96
4.29 Top
kingdom classification of Oteghele spill 99
4.30 Top
phylum classification of Oteghele spill 100
4.31 Top
class classification of Oteghele spill 101
4.32 Top order classification of Oteghele
spill 102
4.33 Top
family classification of Oteghele spill 103
4.34 Top
genus classification of Oteghele spill 104
4.35 Top
specie classification of Oteghele spill 105
4.36 Kingdom
classification of Transforcados pipeline spill 107
4.37 Phylum
classification of Transforcados pipeline spill 108
4.38 Class
classification of Transforcados pipeline spill 109
4.39 Order
classification of Transforcados pipeline spill 110
4.40 Family
classification of Transforcados pipeline spill 111
4.41 Genus
classification of Transforcados pipeline spill 112
4.42 Specie classification of
Transforcados pipeline spill 113
4.43 kingdom classification of transforcados right
of way spill 116
4.44 Top
phylum classification of transforcados right of way spill 117
4.45 Top
class classification of transforcados right of way spill 118
4.46 Top
order classification of transforcados right of way spill 119
4.47 Top
family classification of transforcados right of way spill 120
4.48 Top
genus classification of Transforcados right of way spill 121 4.49 Top
specie classification of Transforcados right of way spill 122
4.50 Gel
electrophoresis showing amplification bands of isolates 159
4.51
Heatmap of the top ten species from each sample (A to E) 160
4.52 Heatmap
of the top ten genus from each sample (A to E) 161
CHAPTER 1
INTRODUCTION
1.1
BACKGROUND
OF THE STUDY
World over, Overcoming the negative
impacts on soil, air and water contamination is a problem for scientists and
environmentalists (Imeh and Sunday, 2012). According to Mehdi (2014) and Patowary et al. (2017), hydrocarbons
are one of the environmental contaminants and the important source of energy in
industry and day to day living. Their release in the environment, whether
unintentionally or as a result of human activities has greatly destroyed the
flora and fauna, polluted the water supply and destroyed human lives and
properties. The discovery, extraction, refinement, transportation, and storage
of petroleum and its byproducts also contribute to a serious environmental
problem (Eze, 2010). According to Das and Chandran (2011), the
major causes of oil spills in Nigeria are corrosion of oil pipelines and
storage tanks, pipeline sabotage and carelessness during oil production
activities.
Also about 40,000 barrels of oil from Mobil platform in
Akwa Ibom State in 1998 were spilled into the environment causing serious harm
to the coastal environment (Eze, 2010). Hence, the oil-rich Niger Delta region
of Nigeria is currently dealing with a significant ecological issue brought on
by widespread crude oil contamination (Chikere et al., 2011).
All
petroleum hydrocarbons originate from crude oil, which is made up of the
fossilized remains of zooplankton and algae from ancient times that underwent
catagenesis to form hydrocarbons. As the name implies, it is a heterogeneous
and complex mixture of hydrocarbons made primarily of hydrocarbons like
asphaltenes, resins, mono and polyaromatics, cycloalkanes, and aliphatic
compounds (linear, branched, saturated, and unsaturated compounds). According
to Joanna and Pawet (2018) and Arjoon and Speight (2020), crude oil is
typically made up of 80% aromatic and saturated hydrocarbons. Petroleum
hydrocarbon is primarily made up substituted hydrocarbons, with carbon making
up 85%-90% and hydrogen accounting for 10%–14% of the mixture. The remaining
components are non-hydrocarbon elements such as sulfur, nitrogen, vanadium, nickel,
arsenic, lead and other metals in small quantities. Also Sodium chlorides, magnesium
chloride and other metallic compounds like vanadium, phosphorus, lead, and
nickel are part of it (Chaudhuri, 2011; Abdulatif, 2015).
Despite the economic advantages of the crude oil
business, oil spillage affects and changes the composition of the soil's
microbial community and biogeochemical cycles and has a significant detrimental
impact on environmental quality and sustainable soil fertility (Milena et al., 2019). Cleanup methods are
required in contaminated areas due to the harmful impacts of petroleum
hydrocarbons on numerous ecosystem components. Burning, sinking, mechanical
removal and the application of detergents are examples of physical and chemical
cleanup techniques and these techniques do however, have drawbacks. The
majority of them are pricey, while some merely move the oil to another site or
possibly worsen environmental pollution (Eze, 2010).
Hydrocarbonoclastic bacteria are found in various habitats,
including soils and water sediments and can degrade or alter petroleum
hydrocarbons without causing any negative impacts. The bulk of natural
hydrocarbon compounds can be broken down by microorganisms. Resins,
asphaltenes, and polycyclic hydrocarbons with high molecular weights are less
biodegradable. The nature of the majority of insoluble petroleum hydrocarbons
inhibits this interaction, which is required for hydrocarbon uptake to occur by
hydrocarbon degrading microorganisms using their substrate (Sampson, 2016).
Globally bioremediation is the use of the metabolic
capacity of biological systems i.e. plants and microbes, to breakdown hazardous
compounds into non harmful or harmless ones in the environment and so far have
received much attention. This is accepted by the public for the cleanup of
contaminated environment due to the fact that it is a natural method using the
natures recycling and self-decontamination capabilities (Chikere et al., 2011). It is a favored treatment
method for cleaning up hydrocarbon-contaminated soils and several studies have
confirmed its efficacy and environmental friendliness in various geographical
and ecological settings. It is also widely known that bacteria
that use hydrocarbons are common in both polluted and unpolluted soil and that
the availability of microorganisms with the required catabolic reaction is
crucial for effective bioremediation. The three primary natural
processes that remove hydrocarbons from the environment involve evaporation, photo-oxidation
and microbial degradation. The reaction might take some time to stabilize
contaminated areas. However, the development of low-cost, high-efficiency
remediation techniques is therefore essential (Eze, 2010).
Chemically made surfactants
are made to increase contaminant solubility; often they are toxic causing additional
contamination to the environment. Similar qualities are
shared by surface-active chemicals produced by microbes, although they are
non-hazardous, biodegradable and produced at
the polluted site. Surfactants and emulsifiers are released by bacteria that
break down hydrocarbons to aid in the assimilation of these insoluble
substrates. In contaminated
areas, microorganisms that can emulsify and solubilize hydrophobic contaminants
at site may have a distinct competitive advantage over rivals. As a result, it
is often beneficial to isolate from such areas and the sites are usually
abundant with the desired microorganisms with the required traits for both on
site and off site bioremediation methods (Nwaguma et al., 2016). There are two primary categories of substances made
by microorganisms: (1) biosurfactants that reduce surface tension at the
air-water interface (2) bioemulsifiers, which lower interfacial tension between
immiscible liquids or at the solid-liquid interface. Although biosurfactants
typically have emulsifying abilities while bioemulsifiers do not always lower
surface tension. According to Batista et
al. (2006), biosurfactant is applied to improve oil recovery from wells,
lessen viscosity of heavy oil, clean oil storage tanks, increase flow through
pipelines, and stabilize fuel water-oil emulsions.
There is a high need for surfactants and was
anticipated to have a global market between 30 to 64 billion US dollars in 2016
and 39 to 86 billion US dollars in 2021. The market for biosurfactants was
344068.40 tons in 2013 and is expected at yearly growth rate of 3–4% from 2014
to 2020, reaching 461991.67 tons. Revenue from biosurfactant market exceeded
$1–8 billion in 2016 and is anticipated to increase to $2–6 billion by 2023,
with the rhamnolipid market likely to grow by over 8% (Singh et al., 2018).
Surfactants contain amphiphilic molecules having polar
and nonpolar moieties in one single molecule. They have a tendency to disperse liquids,
lower surface tensions and cause solubility of non-polar compounds in polar
solvents (Fenibo et al., 2019). Using
biosurfactant producing microorganisms is a good strategy that is employed to
accelerate the degradation of sites polluted with hydrocarbon. Varieties of microorganisms produce
biosurfactants with many chemical compositions, including glycolipids, fatty
acids, lipopeptides and lipoproteins, phospholipids and neutral lipids. Some
microbes produce biosurfactants into the growth medium during the hydrocarbon
breakdown process, changing the cell surface's hydrophobicity in the process
(Thavasi et al., 2011; Patowary et al., 2017; Al-Tamimi et al., 2019).
They are heterogeneous and produced by
microorganisms. It is amphiphilic in nature and made up of a hydrophilic moiety
and hydrophobic part causing their aggregation at surface between fluids with
different polarities like water and hydrocarbons. The hydrophilic part contains
monosaccharides or polysaccharides and proteins. The hydrophobic part comprises
saturated, unsaturated, hydroxylated fatty acids and alcohols. It is made up two
primary categories: biosurfactants and synthetic surfactants. Biosurfactants
are formed by biological processes, as opposed to chemical reactions, which
produced industrially (Nwaguma et al.,
2016; Beulah et al., 2018). With
their distinct characteristics like low toxicity, high biodegradability, good
biocompatibility with eukaryotic organisms, effectiveness at a wider range of
temperatures, pH values, and salinities, low irritancy, ability to be produced
from renewable and inexpensive substrates and production in a friendly conditions like low temperatures,
biosurfactants have recently attracted much more interest than their chemical
counterparts (Odalys et al., 2017).
Biosurfactants producing
microorganisms are widespread in the environment and can be found in both soil
and water. Additionally, they can survive in a varying temperatures, pH and
salinities and can be found in harsh situations (such as oil reserves).
However, the best habitat for extensive biosurfactant production remains
microbial communities that degrade hydrocarbons. Pseudomonas, Bacillus, Sphingomonas, Klebsiella, Actinobacteria,
Halomonas, Alcanivorax and Acinetobacter
are the key genera that typically dominate populations of hydrocarbon-degrading
bacteria (Zhang et al., 2012; Nwaguma
et al., 2016; Xu et al., 2018).
1.2 JUSTIFICATION
One of the challenges in the pollution of the
environment is the release of petroleum hydrocarbons and their byproducts. Both
industrialized and developing nations are focusing their attention on the
cleanup of hydrocarbon pollution (Yanan et
al., 2019). Petroleum compounds take much time to decompose because of
their ability to bind to soil constituents. Since hydrocarbon is mostly a
mixture of carbon and hydrogen, their spillage on soil brings about an imbalance
in the carbon-nitrogen composition at the spill site. This results in a
nitrogen shortage in polluted soil and inhibits bacterial growth and the uptake
of carbon sources (Olajire and Essien, 2014). Through the use of various
microorganisms that may use hydrocarbons as food, bioremediation has the
potential to degrade, modify, remove, immobilize or detoxify harmful compounds
existing in the earth's biospheres (Xu et
al., 2018).
Total degradation, mineralization, sequestration of
toxic contaminants and microbial activity result in the cleaning of the environment
(Seema and Ajinath, 2018). Traditional physical and/or chemical treatments use
a lot of energy, cost a lot of money, and leave behind residues that are
harmful to the biota. According to Vijavakumar and Saravanan (2015), the properties
of microbial surfactants include surface activity, tolerance to pH,
temperature, and ionic strength, biodegradability, low toxicity, emulsifying
and demulsifying ability, and antimicrobial activity.
1.3 AIM AND OBJECTIVES
This research is aimed at evaluating the
physicochemical parameters, biosurfactant production and metagenomic analysis
of hydrocarbon polluted soil in parts of Delta State Nigeria.
1.4 SPECIFIC OBJECTIVES
These include to:
1. Estimate
heterotrophic bacteria and hydrocarbon utilizing bacteria
2. Screen
and characterize bio-surfactant producing bacteria from hydrocarbon
contaminated soil
3. Optimize
bio-surfactants production from the isolated bacterial species
4. Determine
the physicochemical characteristics of
the various soil samples
5. Determine the total petroleum hydrocarbon and heavy
metals of the soil samples
6. Apply
metagenomic tools to examine the various microbial communities in the soil
samples
7. Determine
the functional gene analysis of the microbial communities, including the
reaction of the microorganisms in degradation of petroleum hydrocarbon
1.5 SIGNIFICANCE
OF THE STUDY
The Significance of the study was
to isolate microorganisms capable of degrading hydrocarbons contaminating the
environment. These organisms were observed to produce biosurfactant enhancing
the degradation of petroleum htdrocarbons in the studied sites.
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