Volume 8, Issue 6, June – 2023 International Journal of Innovative Science and Research Technology

ISSN No:-2456-2165

Evaluation of Growth Parameters of Tomato Seeds

Planted in Crude Oil Polluted Soil Remediated
with Strains of Bacillus subtilis and
Pseudomonas aeruginosa
1
Onuoha, U.N. ; 2Appeh, G.O.
1
Department of Microbiology, Michael Okpara University of Agriculture,
Umudike Abia State, Nigeria

Abstract:- The beneficial effects of bioremediation with I. INTRODUCTION

oil-degrading microorganism, Pseudomonas aeruginosa
and Bacillus subtilis, on tomato seedlings growth in oil- Pollution of arable land by industrial effluents, liquid
contaminated soil was investigated. The effect of wastes such as crude petroleum, including gaseous
bioremediation was examined with Pseudomonas petroleum products, are common phenomenon in many
aeruginosa and Bacillus subtilis on phytotoxicity, as countries of the world. This is especially true of oil
measured by tomato seed growth. Tomato growth producing countries (Basumatory et al., 2012; Nwaugo et
parameters (leaf length, height, seed germination, al., 2007; Brooijmans et al., 2009).
chlorosis, stunted growth) were evaluated daily for
15days. In the experimental samples that was polluted Crude oil contamination is a serious environmental
with crude petroleum and inoculated with Pseudomonas threat for soil and plants growing in it. Contamination of soil
aeruginosa, the percentage seed germination was 99% with oil results in serious depression of growth of most
while the seedlings reached a mean height of plants, primarily due to its effects on the physical and
66.9±0.20mm with mean leaf length of 8.1±0.10mm, chemical properties of soil and soil water properties. Crude
while in the control set-up without the isolate, percentage oil affects the environment by changing the essential
seed germination, height and leaf length were 39%, elements of habitat. To overcome this problem, different soil
38.6% and 4.4mm respectively. Records from remediation technologies have been developed such as soil
experimental sample inoculated with Bacillus subtilis, washing with surfactants, biological treatment, thermal
percentage seed germination at the end of the 15days treatment, etc. Bioremediation has the potential to change
monitoring period was 98%, while mean height of the the soil physicochemical properties (Brennan et al., 2014),
tomato seedlings was 66.0±0.20mm and mean length was improve the water holding capacities of soil (Evangelou et
7.9±0.20mm, at the end of the monitoring period was al., 2014) eventually resulting in increased soil fertility (Mia
3.2±0.20mm. Some qualitative morphological et al., 2017); with potential beneficial effects on crop
observations made on the seedlings were also observed productivity, plant establishment, and growth (Brennan et
for seedlings. In the untreated control as well as the al., 2014).
treated experimental samples (Pseudomonas aeruginosa
and Bacillus subtilis), sprouting of the seeds were rapid Bioassays such as measurements of seed germination
and completed in a few days, the seedlings showed steady and early seedling growth have been used to monitor
growth and morphologically normal. In the control soils treatment effects and restoration of oil-contaminated sites
however, sprouting was sluggish, and growth was (Sverdrup et al., 2003). Dorn and Salanitro (2000) found
stunted, yellowing of the leaves was observed (chlorosis); that seed germination and plant growth using corn, wheat,
a number of the seedlings were folded at the tips. These and oats differed from different soils and oil combinations
were indications of phytotoxicity resulting from oil before, during, and after bioremediation. Sayles et al. (1999)
pollution. The results of the present research have showed that oil-contaminated soil treated with aerobic
proven the bioremediation potential of bacterial isolates, biodegradation was less toxic to lettuce and oat root
for crude oil contamination which also showed elongation. Hanson et al. (2007) reported that Acinetobacter
significant growth potential for the tomato seeds. sp. A3-treated soil permitted better germination and growth
of mungbeans, as evidenced by better plant length, weight,
Keywords:- Phytotoxicity, Pollution, Crude Oil, and leaf chlorophyll content. This indicated that
Pseudomonas aeruginosa, Tomatoes. Acinetobacter sp. A3 was capable of reducing crude oil
phytotoxicity through biodegradation.

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Volume 8, Issue 6, June – 2023 International Journal of Innovative Science and Research Technology
ISSN No:-2456-2165
Considering the detrimental effects of crude oil III. RESULTS AND DISCUSSION
pollution on plants and its attendant implications for food
security and environmental integrity in oil rich regions, it The percentage seed germination of the tomato seeds
has become necessary to preserve the available arable land (Lycopesicon esculetum) and the mean seedling height; The
through bioremediation approaches mean leaf length and the mean largest leaf breath of the
experimental set ups (ESU) as well as Control 1 (C1),
In this study, we investigated the remediation of crude Control 2 (C2) and Control 3 (C3), are presented in Tables 1
oil contaminated soil using isolates of Bacillus subtilis and to 4 for polluted soils bioremediated with polluted with
Pseudomonas aeruginosa. Treated soils were used to study Pseudomonas aeruginosa and Bacillus subtilis respectively;
the germination of tomato seedlings and its morphological, for 15days, which marked the end of the monitoring period
and physiological responses were used as indicators for soil for planting. 100% percentage seed germination was
toxicity. observed in the untreated control samples (C3), while the
mean height of the seedlings was 71.0±0.20mm. Mean leaf
II. METHODS length recorded was 8.9±0.00mm while the largest leaf
breath was 3.8±0.20mm. In the experimental samples that
 Study Design was polluted with crude petroleum and inoculated with
Experiments to achieve ex-situ bioremediation were Pseudomonas aeruginosa, the percentage seed germination
set up in sizable plastic containers of equal dimensions. 5% was 99% while the seedlings reached a mean height of
pollution of the freshly obtained garden soil sample was 66.9±0.20mm. The mean leaf length observed for this set up
prepared in the laboratory set ups designated as follow: was 8.1±0.10mm, while the mean largest leaf breath was
3.3±0.00mm. For the control set ups of sample A; the
 Experimental set up (ESU) which contained polluted records were as follows: percentage seed germination for
agricultural soil (5% v/w) and 250ml of the controls 1and 2 are 39.0% and 35.0% respectively, while
Pseudomonas aeruginosa broth at its exponential the mean height of seedlings for same controls were
growth phase evenly dispersed all over the set up. 38.6±0.00mm for control 1 and 36.8±0.00 for control 2
 A second but similarly designed representing control 1 respectively. The mean leaf length of sample A controls
(CSU 1) which contained the polluted agricultural soil were: 4.4±0.00mm for control 1 and 4.1±0.10 for control 2
(5%v/w) and 250ml sterile prepared microbiological respectively, while the largest leaf breath recorded were,
media (UDBB), but devoid of any microorganism and 2.5±0.10mm for control 1 and 2.2±0.20mm for control 2
evenly dispersed in the soil. respectively(Tables 1 and 2)
 A third also similarly designed representing control 2
(CSU 2) which contained the polluted agricultural soil Records from experimental sample B and controls
(5%v/w) and 250ml sterile distilled water (SDW) which was crude petroleum polluted agricultural soil, seeded
evenly dispersed all over the soil. with Bacillus subtilis are presented in Table 3 and 4. The
record shows the following values: The percentage seed
The above designs were set up for both Pseudomonas germination at the end of the 15days monitoring period is
aeruginosa denoted sample A and for Bacillus subtilis 98%, while mean height of the tomato seedlings was
denoted sample B. 66.0±0.20mm ; mean length length recorded was
7.9±0.20mm, while the largest leaf breath at the end of the
 Testing for Decontamination of the Polluted Soil monitoring period was 3.2±0.20mm.
The experimental and control soils were examined
nine (9) weeks after bioremediation of the oil pollution From the control soils, at the end of the 15day
treatment process. This was to check the recovery of the monitoring period, percentage seed germination recorded
agricultural soil from the oil impaction. To achieve this test, was 38.0% and 34.0% for B controls 1 and 2 respectively.
100 Tomato seeds (Lycopersicon esculantum) were planted The mean height of seedlings was 38.0±0.10mm for controls
per replicate set up and the results obtained compared to 1 and 36.1±0.20mm for control 2.The mean leaf length for
those of a third control (contr 3) which was not treated controls 1 and 2 were 3.8±0.40mm and 3.6±0.10mm
(untreated and unpolluted agricultural soil). The seeds were respectively, while the largest leaf breath were 2.4± 0.00mm
watered at weekly intervals and the daily records of: for control 1 and 1.9 ±0.00mm for control 2.
Number of germination, their morphology, mean seedling
height, mean length of leaves and mean leaf breadth were Some qualitative morphological observations made on
taken. Observations e.g leaf folds/curls, leaf droppings, the seedlings were also recorded in Table 5 for sample A
chlorosis, stunted growth etc were also made (Akujobi et al., soils treated with Pseudomonas aeruginosa, and Table 6 for
2011). sample B soils treated with Bacillus subtilis. In the untreated
control (C3) as well as the treated experimental samples A
and B, sprouting of the seeds were rapid and completed in a
few days, the seedlings showed steady growth and
morphologically normal. In the control soils however,
sprouting was sluggish, and when they did sprout, growth
was stunted, yellowing of the leaves was observed
(chlorosis); A number of the seedlings were folded at the

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Volume 8, Issue 6, June – 2023 International Journal of Innovative Science and Research Technology
ISSN No:-2456-2165
tips while their stems were pale and etiolated. These were the control stems with leaves felled and dropped on their
indications of phytotoxicity resulting from oil. A number of sides.

Table 1 Mean percentage seed germination (%) and height of seedlings (mm) of tomato seedlings (Lycopersicon esculetum)
planted in crude petroleum polluted agricultural soil after bioremediation using Pseudomonas aeruginosa [SAMPLES A]
Mean Percentage Seed Germination (%) Mean Height Of Seedling ( mm)
Time in Days Exptl Contr 1 Contr 2 Contr 3 Exptl Contr 1 Contr 2 Contr 3
0 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
1 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
2 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
3 4.0±0.50 0.0±0.00 0.0±0.00 5.0±0.50 5.0±0.50 0.0±0.00 0.0±0.00 3.8±0.10
4 20.0±0.30 2.0±0.50 1.0±0.50 25.0±0.50 4.6±0.00 1.2±0.10 1.0±0.10 6.2±0.10
5 25.0±0.20 8.0±0.50 3.0±0.00 30.0±0.00 10.2±0.00 3.4±0.10 2.1±0.00 14.8±0.00
6 32.0±0.10 8.0±0.50 9.0±0.00 42.0±0.50 20.2±0.00 6.1±0.10 5.9±0.10 22.6±0.10
7 45.0±0.50 23.0±0.00 22.0±0.50 68.0±0.00 28.0±0.10 8.6±0.10 6.1±0.10 30.0±0.10
8 58.0±0.30 26.0±0.50 24.0±0.00 82.0±0.50 34.2±0.10 14.6±0.10 12.5±0.00 38.8±0.30
9 62.0±0.20 32.0±0.00 26.0±0.00 96.0±0.00 40.4±0.10 20.1±0.00 18.8±0.10 46.9±0.40
10 71.0±0.20 34.0±0.50 29.0±0.00 100.0±0.00 46.5±0.00 34.9±0.00 21.5±0.10 50.4±0.10
11 86.0±0.10 36.0±0.50 30.0±0.50 100.0±0.00 52.8±0.00 28.0±0.00 22.4±0.00 58.8±0.20
12 92.0±0.00 38.0±0.50 32.0±0.50 100.0±0.00 59.4±0.10 30.1±0.00 28.2±0.10 62.5±0.00
13 97.0±0.00 38.0±0.50 32.0±0.50 100.0±0.00 62.0±0.20 32.2±0.10 31.1±0.10 62.6±0.00
14 99.0±0.10 38.0±0.00 33.0±0.00 100.0±0.00 48.5±0.10 36.8±0.20 33.4±0.20 68.1±0.10
15 99.0±0.00 38.0±0.00 33.0±0.00 100.0±0.00 66.9±0.20 38.0±0.00 36.9±0.00 69.0±0.00

Table 2 Mean leaf length (mm) and largest leaf breath (mm) of tomato seedlings (Lycopersicon esculetum) planted in crude
petroleum polluted agricultural soil after bioremediation using Pseudomonas aeruginosa [Sample A]
Mean Leaf Length (mm) Mean Largest Leaf Breath (mm)
Time in days Exptl Contr 1 Contr 2 Contr 3 Exptl Contr 1 Contr 2 Contr 3
0 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
2 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
3 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
4 0.0±0.00 0.0±0.00 0.0±0.00 2.6±0.10 1.2±0.10 0.0±0.00 0.0±0.00 1.4±0.20
5 2.0±0.10 1.0±0.10 0.0±0.0 3.0±0.10 1.2±0.10 0.1±0.10 0.2±0.10 1.6±0.10
6 2.9±0.30 1.3±0.00 1.0±0.10 3.6±0.10 1.5±0.30 0.6±0.40 0.4±0.20 1.0±0.10
7 0.40 1.6±0.20 1.2±0.20 4.0±0.10 1.6±0.10 1.2±0.10 0.6±0.10 1.9±0.20
8 4.6±0.10 2.0±0.20 1.7±0.30 4.0±0.30 1.8±0.20 1.3±0.00 1.2±0.20 2.0±0.10
9 5.2±0.20 2.2±0.20 2.0±0.00 4.5±0.20 2.1±0.20 1.3±0.00 1.2±0.20 2.4±0.30
10 5.9±0.30 2.6±0.10 2.1±0.10 5.0±0.40 2.2±0.30 1.5±0.10 1.3±0.10 2.7±0.20
11 6.5±0.10 3.2±0.20 2.3±0.30 5.8±0.10 2.4±0.40 1.7±0.30 1.4±0.10 2.9±0.10
12 7.1±0.10 3.6±0.40 2.5±0.30 6.8±0.20 2.6±0.20 1.9±0.10 1.5±0.20 3.0±0.00
13 7.2±0.20 3.6±0.10 2.8±0.30 7.9±0.00 2.8±0.00 2.2±0.10 1.6±0.10 3.3±0.20
14 7.4±0.20 3.6±0.10 2.8±0.20 8.0±0.10 3.0±0.10 2.3±0.20 1.8±0.10 3.5±0.40
15 7.8±0.10 3.8±0.10 2.9±0.10 8.0±0.10 3.1±0.10 2.4±0.10 1.9±0.00 3.8±0.20
16 8.0±0.10 3.8±0.10 2.9±0.10 8.1±0.00 3.2±0.00 2.5±0.10 2.2±0.20 3.8±0.20

Table 3 Mean percentage seed germination (%) and height of seedlings (mm) of tomato seedlings (Lycopersicon esculetum)
planted in crude petroleum polluted agricultural soil after bioremediation using Bacillus subtilus [SAMPLES B]
Mean Percentage Seed Germination (%) Mean Height of Seedling ( mm)
Time in days Exptl Contr 1 Contr 2 Contr 3 Exptl Contr 1 Contr 2 Contr 3
0 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
1 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
2 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
3 1.0±0.20 0.0±0.00 0.0±0.00 5.0±0.10 2.0±0.00 1.0±0.20 0.0±0.00 3.8±0.50
4 3.0±0.30 0.0±0.00 1.0±0.20 25. 0±0.40 3.2±0.50 1.1±0.10 1.1±0.40 6.2±0.20
5 26.0±0.20 6.0±0.10 1.0±0.10 30.0±0.10 6.6±0.30 2.4±0.30 1.9±0.40 14.8±0.40
6 37.0±0.10 20.0±0.10 8.0±0.30 42.0±0.30 16.9±0.20 5.5±0.20 4.0±0.20 22.6±0.00
7 40.0±0.00 23.0±0.30 20.0±0.30 68.0±0.20 26.4±0.10 8.6±0.40 6.2±0.30 30.0±0.10
8 50.0±0.50 27.0±0.40 22.0±0.20 82.0±0.10 30.5±0.00 14.0±0.30 12.9±0.10 38.8±0.20
9 68.0±0.10 30.0±0.20 28.0±0.10 96.0±0.30 36.2±0.40 16.4±0.20 15.0±0.40 46.9±0.40

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10 78.0±0.10 34.0±0.30 29.0±0.10 100.0±0.00 42.1±0.30 20.5±0.30 18.9±0.20 50.4±0.30
11 82.0±0.10 36.0±0.10 30.0±0.10 100.0±0.00 50.5±0.10 26.2±0.10 26.4±0.00 58.8±0.10
12 88.0±0.40 38.0±0.30 33.0±0.40 100.0±0.00 55.9±0.50 30.1±0.00 27.0±0.50 62.5±0.30
13 96.0±0.30 38.0±0.00 33.0±0.20 100.0±0.00 60.1±0.30 33.0±0.20 30.1±0.40 65.6±0.20
14 98.0±0.10 38.0±0.00 34.0±0.50 100.0±0.00 62.0±0.00 36.6±0.30 34.2±0.10 68.1±0.10
15 98.0±0.10 39.0±0.20 34.0±0.00 100.0±0.00 66.2±0.20 39.8±0.10 36.3±0.20 71.0±0.20

Table 4 Mean leaf length (mm) and largest leaf breath (mm) of tomato seedlings (Lycopersicon esculetum) planted in crude
petroleum polluted agricultural soil after bioremediation using Bacillus subtilus [SAMPLE B]
Mean leaf length (mm) Mean largest leaf breath (mm)
Time in days Exptl Contr 1 Contr 2 Contr 3 Exptl Contr 1 Contr 2 Contr 3
0 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
1 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
2 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00
3 0.0±0.00 1.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 0.0±0.00 1.4±0.20
4 1.6±0.10 1.0±0.10 0.0±0.00 3.0±0.10 1.1±0.50 0.1±0.10 0.15±0.10 1.6±0.50
5 2.5±0.40 1.1±0.20 1.0±0.20 3.6±0.30 1.3±0.30 0.4±0.30 0.4±0.10 1.6±0.40
6 3.1±0.20 1.3±0.00 1.2±0.10 4.0±0.40 1.5±0.10 1.0±0.30 0.4±0.10 1.9±0.20
7 4.4±0.00 1.8±0.10 1.5±0.00 4.0±0.40 1.7±0.20 1.2±0.40 0.8±0.10 2.0±0.30
8 4.9±0.00 1.9±0.20 1.9±.040 4.5±0.30 2.1±10 1.2±0.40 1.1±0.20 2.4±0.40
9 5.8±0.10 2.4±0.10 1.9±0.40 5.0±0.10 2.3±0.30 1.4±0.20 1.2±0.10 2.7±0.30
10 6.4±0.20 3.3±0.40 2.4±0.30 5.8±0.30 2.3±0.30 1.6±0.30 1.4±0.30 2.9±0.20
11 6.4±0.10 3.6±0.10 2.6±0.10 6.8±0.10 2.5±0.40 1.85±0.20 1.6±0.40 3.0±0.20
12 7.3±0.20 37.1±0.10 2.8±0.00 7.9±0.00 2.8±0.50 1.3±0.40 1.7±0.10 3.3±0.30
13 7.3±0.00 4.4±0.40 2.9±0.10 8.0±0.10 2.7±0.20 2.3±0.10 1.8±040 3.5±0.10
14 7.9±0.20 4.4±0.40 2.9±0.10 8.0±0.10 3.2±0.30 2.4±0.10 1.9±0.10 3.8±0.00
15 7.9±0.20 4.4±0.40 3.0±0.10 8.1±0.10 3.2±0.20 2.4±0.00 1.9±0.00 3.8±0.10

Table 5 Some morphological observation made on the tomatoes seedlings (Lycopersicon esculentum) on polluted agricultural soil
after bioremediation using Pseudomonas aeruginosa

 Legend

 NG = No germination seen yet

 ― = Absent
 + = Present
 ++ = Higher percentage present
 +++ = Highest percentage present

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Table 6 Some Morphological Observations Made on the Tomato Seedlings (Lycopersicon Esculetum) Planted on Crude Petroleum
Polluted Soil After Biomediation Using Bacillus Subtilus [SAMPLE B]

 Legend

 NG = No germination seen yet

 - = Absent
 + = Present
 ++ = Higher percentage present
 +++ = Highest percentage present

IV. DISCUSSIONS the experimental soil samples. From the results obtained, the
recovery effects were better in the experimental and
Crude oil contamination affects the agriculture sector unpolluted soil samples in contrast to the controls. The
particularly due to soil toxicity and reduced plant growth. result on plant height agrees with previous findings of
Crude oil causes environmental risks in the soil ecological Ikhajiagbe and Anoliefo (2011) on the significant reduction
system, by inhibition of plant growth, damage to soil of plant growth occasioned by oil pollution. Ojimba and
structure, disturbance of soil water quality (Han et al., Iyagba (2012) reported the decreased output of horticultural
2016). crops in crude oil polluted farms as compared with the
unpolluted farms.
The present research was designed to observe the
growth response of tomato seedlings in crude oil polluted Various studies have reported the role of these
soil samples remediated with isolates of Pseudomonas remediation techniques in the improvement of plant growth
aeruginosa and Bacillus subtilis. It was found that these (Shahid et al., 2017; Laird et al., 2010). Our results also
methods are not only beneficial to remediate the soil but relate with Laird et al. (2010), where fresh weight and dry
also capable to overcome the toxic nature of crude oil as weight increased because of remediation of the soil. Hence
evidenced by the improved growth performance of the it could be inferred that the isolates used in this study may
seedlings in comparison with the unpolluted soil. reduce the leaching losses of nutrients by causing retention
Phytotoxicity study showed that leaf breath, heights, seed of nutrients in the soil.
germination and leaves lengths of the plants were
significantly reduced in the samples planted in control soil Disturbances of major physiological processes within
relative to the experimental and unpolluted soils. The plants systems due to hydrocarbon contamination are
percentage increase recorded for seed germination and plant common; and manifest in the forms of morphological,
height in both experimental soils (P. aeruginosa and B. pigment, chlorotic and necrotic foliar patterns alterations. In
subtilis remediated) might be due to an increase in nutrient this study, only tomato seeds grown in experimental soil and
uptake by the plant and a decrease in crude oil uptake (Mosa unpolluted soil didn’t exhibit stunted growth, chlorosis and
et al., 2016). Increased microbial activities resulting in leaf folding/dropping. For plants grown in contaminated
reduced accumulation of crude oil residues made these soil, the parameters were not observed completely. This
nutrients available in the state that they were easily finding is not without precedence as some couples of
assimilated by the test plant. This resulted to the better plant investigations claimed similar results (Chuku et al., 2018;
yield indices observed, in terms of percentage seed Okonokhua et al., 2007). Studies by Kekere et al. (2011)
germination; plant height and plant leaf breath parameters in reported that there was no crop yield recorded in cowpea

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Volume 8, Issue 6, June – 2023 International Journal of Innovative Science and Research Technology
ISSN No:-2456-2165
grown in contaminated soil due to plant mortality. They also [7]. Hanson, K.G., Nigam, A.N., Kapadia, M. and Desai,
observed that crude oil contamination at all intensities A.J. (2007). Bioremediation of crude oil
resulted in a significant reduction in leaf number when contamination with Acinetobacter sp. A3. Curr.
compared with the control treatment in a concentration- Microbiol., 35:191–193.
dependent manner. Our results correlate with Ali et al. [8]. Ikhajiagbe, B. & Anoliefo, G. O. (2011). Natural
(2017), who reported that crude oil adversely affects the attenuation of a 14- month-old waste engine oil
plant-water-soil relationship, resulting in chlorosis. The polluted soil. Journal of Soil Science and
relative plants' heights and the number of leaves were worst Environmental Management, 2(7), 184-192.
affected especially in plants grown in crude oil tainted soils [9]. Kekere, O., Ikhajiagbe, B. and Apela, B. R. (2011).
where absence of leaves was observed at the end of the Effects of crude petroleum oil on the germination,
experiments. growth, and yield of Vigna unguiculata Walp L.
Journal of Agriculture and Biological Sciences,
V. CONCLUSION 2(6).158 – 165.
[10]. Laird, D. A. et al. (2010). Impact of biochar
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