Impact of Crude Oil Polluted Soil on Seedling Morphological Characteristics and Biomass Accumulation of Monodora myristica (African Nutmeg)

: This study was designed to investigate the effect of crude oil polluted soil on the morphological characteristics and biomass accumulation of Monodora myristica seedling. The study was carried out in the forest nursery of the Department of Forestry and Wildlife Management, University of Port Harcourt, Nigeria using standard field experimental methods. Significant variations (P ≤ 0.05) were observed in seedling height and biomass while number of leaves and collar diameter displayed no significant differences (P > 0.05) between treatments. The result shows that M. myristica seedlings are not tolerant to crude oil pollution. The trend in growth parameters measured showed that increased level of crude oil reduced growth. It is recommended that M. myristica seedlings should not be raised in crude oil polluted soils or be used in crude oil pollution remediation programmes due to its poor growth performance.

Monodora myristica, commonly called African nutmeg or calabash nutmeg is a perennial edible plant of the family Annonaceae (Onyenibe et al., 2015).It is primarily found in the evergreen forests of West Africa in nations like Liberia, Nigeria, Cameroon, Angola, Uganda and west Kenya (Onyenibe et al., 2015) and its common names in Nigeria are ehuru, ariwo, ehiri and airama (Burabai et al., 2008).Its seed is a common spice in West African cooking and has a taste and aroma comparable to nutmeg.This tropical shrub is of the family of flowering plants (Okafor, 1976).The fruit of M. myristica is a 20 cm diameter berry that is smooth, green, spherical, and when it ages, it turns woody.It is joined to an elongated stalk that can reach a length of 60 cm.The many long, pale brown seeds, which are about 1.5 cm long and encircled by a white, aromatic pulp, are found inside the fruit.Studies have shown that almost every part of M. myristica tree is important economically.The timber is hard, easy to work with and is used for carpentry, house fittings and joinery while the seeds are also made into necklaces (Nguefack et al., 2004).Reports abound in the literature as to the medicinal use of M. myristica, the stem bark is used in the treatments of haemorrhoids, stomach ache, fever pains and eye diseases (Nwaoguikpe and Uwakwe, 2005), while the seeds are used in treating headache and hypertension in Central African Republic (Koudou et al., 2007).In Eastern Nigeria, the seeds are used as condiment and is one of the major spices used as postpartum tonic.M. myristica has been proven to have anti-sickling properties (Nwaoguikpe and Uwakwe, 2005).When grounded to powder, the kernel is used to prepare soup as stimulant to relieve constipation and control passive uterine haemorrhage in women immediately after child birth (Udeala, 1980;Iwu et al., 1987) is a natural occurring complex mixture of hydrocarbon and non-hydrocarbon compounds and at high concentrations can be toxic to living organisms (Nelson-Smith, 1973).The discovery of crude oil in Nigeria has significantly improved her economy due to its importance as a significant source of foreign exchange and in boosting the Gross Domestic Product (GDP) of the country.However, its exploration and exploitation has posed immense danger and harm to the oil-producing communities due to the resultant effect it has on the aquatic life and soil topography of the oil-rich region.The Niger Delta region of Nigeria is well-known for the production of oil that commenced as far back as 1956 and the discovery of oil in the Niger Delta region improved the financial state of the economy but was immediately accompanied by a series of crude oil spillage on land and water giving rise to soil and water pollution (Adekunle and Adeniyi, 2015).The major cause of oil spills has been identified to be from vandalization and leakages of pipelines.The effects of environmental pollution resulting from the production of oil became a topical issue at both national and international levels (Adekunle and Adeniyi, 2015).Crude oil contamination in and around the exploration and spillage sites of oil industries is a burning problem throughout the world.Crude oil releases toxic hydrocarbons in nature which causes pollution.Soil contamination has become a global phenomenon.Chemicals and heavy metals have contaminated soil in many regions of the world as a result of industrial, agricultural, and uncontrolled waste disposal practices.(Tang and Angela, 2019).Contamination of soil with oil results in serious retardation of growth of most plants, primarily due to its effects on the physical and chemical properties of soil and soil water relations (Saini and Arya, 2016).There are several reports of the effect of crude oil pollution on plant growth (Osuji and Nwoye, 2007).Oil pollution causes loss of plant covering and plants diversity (Maranho et al., 2006).As important as M. myristica is, its natural population is depleting very fast in the wild as a result of overexploitation, deforestation and forest degradation consequent upon many underlying causes many of which are no more obscure.Excessive pressure on tropical forests by people and their animals have posed a very serious challenge to natural regeneration of many tropical tree species because the mother trees that should serve as seed trees are being removed for one reason or the other (Longman, 1993).Ngwuta et al. (2016) reported that though Monodora myristica is one of the valued forest tree species, it is facing the threat of extinction caused by large scale exploitation and destruction of the natural forests.The silvicultural requirements of most forest tree species including M. myristica are inconsistent and results vary from location to location.Information on the silvicultural requirements of M. myristica have not been well documented.There is also no adequate information on the effect of crude oil pollution on M. myristica seedlings thereby creating need for this study.
There are compelling reasons why the research on the effect of crude oil pollution on seedlings morphological characteristics and biomass accumulation of Monodora myristica is necessary.Obtaining adequate knowledge on the impact of crude oil polluted soil on the seedlings of Monodora myristica is very essential to determining the level of pollution this species can tolerate.Findings from the study can inform the development of regulations and policies related to the protection of indigenous plants and ecosystems in regions where Monodora myristica is found.It can also contribute to guidelines for responsible resource extraction and environmental protection.Hence, the objective of this paper was to investigate the effect of crude oil polluted soil on the morphological characteristics and biomass accumulation of Monodora myristica seedling.Materials: Materials used for the study include: Viable seeds of Monodora myristica; germination basket and medium sized polybags for transplanting; a field notebook and pen for data collection; sterilized forest topsoil used as growth medium for transplanting seedlings; sharp sand used as medium for seed germination; and crude oil for polluting the soil.

MATERIALS AND METHOD
Fruit collection and processing: The seeds of Monodora myristica were gotten from Okomu National Park in Edo State, Nigeria.The floatation method of seed viability test was employed to detect the viable seeds.Seeds that floated after putting them in a container filled with water were regarded as nonviable and discarded while the seeds that sank were regarded as viable and used for the experiment.
Experimental Design: The experiment was laid out in a completely randomized design (CRD).One hundred and fifty (150) seeds were directly sown in a germination tray filled with sharp sand.Watering was done as necessary.After one month of establishment, 10 seedlings were pricked into poly-bags filled with topsoil for each the nine treatments.The treatments involved polluting 10 kg of topsoil with different levels of crude oil (0 control), 25, 50, 75, 100, 125, 150, 175 and 200 ml).The topsoil was mixed thoroughly with different volumes of crude oil and left for 2 days to allow for proper mixing.Prior to the transplanting operation, the poly-bags were fully watered.Watering and weeding were carried out regularly and when required throughout the period of the experiment.
Seedling Growth Data Collection: Seedling growth data were collected on seedling height (cm), diameter (mm) and leaf number.The data collection began one month after planting and monthly thereafter for six months.The seedling height was measured from the substrate level to the tip of the youngest leaf using a meter rule; stem collar diameter was measured at the collar position using a digital vernier caliper; the numbers of fully expanded leaves were determined `by directly counting.
Seedling Biomass and Moisture Content: Five seedlings were chosen from each treatment at the end of the experiment and carefully removed from the polybags and the root system exposed by carefully washing off the growth media from the roots.Absorbent paper was used for blotting excess moisture from the plants.Seedlings were then separated into shoot and root components by cutting at the collar.The root length, fresh weight of shoot (including the leaves) and root were taken and then placed in a paper bag for drying.The shoot and the root samples were dried in the oven at 70°C for three days (72 hours).The biomass (g) was determined by weighing both wet and dry weight with a weighing scale while the moisture content was determined by subtracting the dry weight from the wet weight.
Data Analysis: Data collected were analyzed using SPSS statistical software (SPSS version 18, SPSS Inc.).One-way analysis of variance was used to test for significant differences (p ≤ 0.05) in the evaluated seedling attributes among the treatments.Duncan multiple range test (DMRT) was used to determine the variation in means.All graphs were plotted using Microsoft excel 2010.

RESULTS AND DISCUSSION
Growth Response of M. myristica Seedlings to Crude Oil Pollution: Seedling Height: Seedlings of M. myristica showed considerable variations (p ≤ 0.05) in height at all stages of growth.M. myristica seedlings under different levels of crude oil pollution showed a decrease in height as oil pollution increased with the Control having highest height from months 1 to 6 (21.74, 25.00, 25.77, 26.06, 26.43 and 30.58 cm, respectively), followed by 25ml (21.29, 24.10, 24.81, 25.18, 25.67 and 27.95 cm, respectively) while lowest height was observed at 200 ml (15.44, 17.69, 18.52, 18.98, 19.67 and 21.35cm) at all stages of growth (Table 1).Collar Diameter: M. myristica seedlings under different levels of crude oil pollution showed a decrease in collar diameter as oil pollution increased with the Control (0 ml) having the highest collar diameter from months 1 to 6 (2.38, 2.63, 2.73, 2.86, 3.02 and 3.31 mm, respectively), followed by seedlings grown in 25 ml level of crude oil.The lowest collar diameter was observed in seedlings grown in 200 ml (2.09, 2.30, 2.41, 2.55, 2.64 and 2.84 respectively).However, the variations in collar diameter were not significant (p > 0.05) at all stages of growth (Table 2).
Leaf Production: Seedlings of M. myristica showed no significant variations (p ≤ 0.05) in leaf number at all stages of growth.At month 6, the highest leaf production was observed in the Control (0 ml) while the lowest was observed in 200 ml level of crude oil.At month 2, the highest leaf production was observed in the Control (0ml) followed by 25ml but the lowest was observed in 200ml.This is shown in Figure 1.(Baker, 1970).Effect of crude oil levels on seedling height and biomass were observed to be significantly different while collar diameter, number of leaves and root moisture content were not significantly different although it was observed that increase in crude oil levels led to decrease in growth for all parameters measured which shows that crude oil had adverse effect on plants.This may be attributed to disruption in water and nutrient uptake owing to the effects of oil in soil, and the depletion of soil nitrogen and phosphorus content (Baran et al., 2002).The reduction in the evaluated plant growth parameters with the increased crude oil levels could be related to increased effect of the crude oil which could have inhibited cell division, enlargement and expansion and nutrient availability.These could have negatively affected the anatomical and physiological structures of the plant and hence growth reduction (Agbogidi and Eshegbeyi, 2006).Growth reduction in the presence of crude oil has also been reported by other researchers including Bamidele et al. (2007), Chima et al. (2018) and Oyedeji et al. (2012).The trend in seedling height increment of M. myristica showed that the addition of various levels of crude oil retarded growth in total height.The observed reduction in the height of M. myristica agrees with the findings of Rowell (1977) that growth retardation is possible with oil pollution of soil due to insufficient aeration caused by displacement of air from pore spaces.This may have been caused by the higher quantity of oil present in the soil which may have inhibited the flow of nutrients.
Growth reduction with increasing pollution level is suggestive of the fact that oil in the soil affects nutrient level probably as a result of immobilization (Agbogidi et al., 2009).Also, highest biomass observed in control which decreased as concentration increased agrees with the report of Baruah et al. (2016) that after harvest, plant biomass of Crotalaria pallida planted in soil contaminated with crude oil was found to be the maximum in 0 ppm (i.e. the Control) and gradually decreased according to their concentration gradient.The non-significant difference in collar diameter and leaf production among treatments suggests that crude oil-pollution of soil up to 200 ml/10kg of soil had no negative impact on the growth parameters for the sixmonth period of the experiment.

Conclusion:
Crude oil pollution significantly affected the early growth of M. myristica in height, collar diameter, leaf production and biomass.The result shows that M. myristica seedlings are not tolerant to crude oil pollution.It is recommended that M. myristica seedlings should not be raised in crude oil polluted soils or be used in crude oil pollution remediation programmes or projects due to its poor growth performance in crude oil-polluted soil.

Fig 1 :
Fig 1: Effects of crude oil pollution on leaf production of M. myristica at months 2 and 6.Bars with the same letter (s) are not significantly different at the 0.05 level.
. Crude oil Impact of Crude Oil Polluted Soil on Seedling Morphological…..
Study Area: The study was carried out at the forest nursery of the Department of Forestry and Wildlife Management., Faculty of Agriculture, University of Port Harcourt, Rivers State, Nigeria.The nursery lies at Latitude 04°53 > 38.3'N and Longitude 00.6° 54'38'E between October and April 2021.

Table 1 .
Height performance of M. myristica seedling at different levels of crude oil pollution (±SE).

Table 2 .
Collar diameter of M. myristica seedling at different levels of crude oil pollution (±SE).Means with the same alphabet on the same column do not differ significantly (p > 0.05).