Journal Clean WAS (JCleanWAS) 5(1) (2021) 31-34
Journal Clean WAS (JCleanWAS)
DOI: http://doi.org/10.26480/jcleanwas.01.2021.31.34
ISSN: 2521-0912 (Print)
ISSN: 2521-0513 (Online)
CODEN: JCOLBF
RESEARCH ARTICLE
ISOLATION AND MORPHOLOGICAL IDENTIFICATION OF SOIL FUNGI FROM
AGRICULTURAL SOIL IN KUANTAN
Nur Sabrina Ahmad Azmi*, Asma Adiba Hisham
Department of Plant Science, Kulliyyah of Science, International Islamic University Malaysia, Jalan Sultan Ahmad Shah, Bandar Indera
Mahkota, 25200 Kuantan, Pahang, Malaysia
*Corresponding author email: sabrinaazmi@iium.edu.my
This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction
in any medium, provided the original work is properly cited.
ARTICLE DETAILS ABSTRACT
Article History: Soil fungi possess a great number of potential benefits that could be applied in various fields. They are well-
known for acting as plant-growth promoter, biocontrol agent of plant diseases and involves in
Received 24 April 2021 bioremediation. In this study, the fungi were isolated from used agricultural soil in Glasshouse and Nursery
Accepted 28 May 2021
Available online 18 June 2021
Complex (GNC), International Islamic University Malaysia, Kuantan, Pahang using serial dilution and plating
techniques. Around 10 isolates of soil fungi were successfully isolated and the identification of all isolates
were based on their cultural and morphological characteristics. The fungi were discovered to be from genus
Cladorrhinum, Penicillium, Paecilomyces and Aspergillus.
KEYWORDS
Soil fungi, Cladorrhinum, Penicillium, Paecilomyces, Aspergillus, agricultural soil.
1. INTRODUCTION use of synthetic mineral nitrogen source as fertilizer negatively influenced
the soil microorganisms’ communities and diversity (Gryndler et al.,
Soil is the outer layer of the Earth’s crust which is loosely-arranged and 2006). Following these practices, an effective method of soil management
has many forms (Baxter and Williamson, 2001). It generally consists of is therefore vital in order to maintain crop productivity, environmental
organic and inorganic matter. The organic fraction of soil is divided into sustainability, and human health. Due to the steady increase of world
biomass that includes living organisms and necromass which is the dead population and thus escalating food production needs, proper actions
organisms and their transformation products (Nortcliff et al., 2006). A should be taken to overcome this problem in the coming years.
gram of soil is estimated to accommodates around 90–100 million bacteria
and about 200 000 fungi, with most of these organisms being located at One of the methods being taken into action as a solution is practicing
the rhizosphere (Glick, 2018). These organisms play an important role in sustainable agriculture. This includes all of the systems and practices that
maintaining soil health, quality and fertility through their natural will improve the protection of the environment and natural agricultural
interactions and processes (Lori et al., 2017). resources necessary to ensure the production of adequate and high-
quality foodstuffs at affordable costs which the rapidly growing world
The physical, chemical and biological properties of soil significantly affect population needs (Tuğrul, 2020). It mainly focuses on increasing the
the production and sustainability of agricultural practices. This is because productivity of the soil and reducing the harmful effects of agricultural
the soil acts as a medium for plant growth which stores nutrients, water practices on climate, soil, water, environment and human health.
and also provides an environment for the breakdown and immobilization
of materials such as fertilizers (Nortcliff et al., 2006). Growing media mixes The decrease of arable land and resources had become the major reasons
mainly includes topsoil, sawdust, perlite, vermiculite, peat moss, bark and for the need to have sustainable agricultural practices. However, most of
composts (Gruda, 2019). Topsoil is recognized as the outermost few the agricultural activities nowadays have been causing the quality of soil
centimetres or more of soil which is rich with organic matter and health to decrease especially by affecting the soil microbiome. The soil
microorganisms (Darmody et al., 2009). Agricultural activities cause both fungal species have both good and bad impacts on agriculture. Therefore,
good and bad effects to soil organisms. For instance, the application of raw the isolation and identification of soil fungi is important to further
and composted organics as fertilizer promote microbial proliferation in characterize their functions in soil ecosystem. Therefore, the aim of this
the soil (Bunemann et al., 2006). study are i) to isolate soil fungi communities from used agricultural soil in
Glasshouse and Nursery Complex, IIUM using serial dilution and plating
However, agricultural management such as soil tillage had been found to techniques and ii) to identify the species of isolated soil fungi based on
decrease the population of soil organisms due to mechanical destruction their cultural and morphological characteristics.
and soil compaction (Garcia-Orenes et al., 2013). Besides, the extensive
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Cite the Article: Nur Sabrina Ahmad Azmi, Asma Adiba Hisham (2021). Isolation and Morphological Identification of Soil Fungi from Agricultural Soil in Kuantan.
Journal Clean WAS, 5(1): 31-34.
Journal Clean WAS (JCleanWAS) 5(1) (2021) 31-34
2. MATERIALS AND METHODS
2.1 Sample collection
Soil sample was collected from the used agricultural soil in Glasshouse and
Nursery Complex (GNC), IIUM in September 2019. The soil sample has
been used by the students for laboratory practicals or experiment
purposes, which is a mixture of top soil, peat moss and sand. Firstly, the
soil sample was air-dried for 3 days in the laminar air flow and was ground
by using mortar and pestle. Ten grams of the dried soil was weighed.
2.2 Preparation of soil dilution and microbial culture
Isolation of soil fungi by soil dilution plating was adapted from method by
Pepper and Gerba (2004). Ten grams of the soil sample was added to 95
mL of deionized water (solution A) and shaken well to disperse the
organism. This yielded a 10-fold dilution series. Next, 1 mL of the
suspension was removed from solution A using a sterile pipette and then
transferred to a new tube containing 9 mL dilution blank (solution B). The
tube was capped and vortexed. The dilution factor for solution B is 1×10ˉ².
The dilution series was continued to obtain solutions with dilution factors Figure 1: Cultural and morphological characteristics of Cladorrhinum sp.
of 1×10ˉ³ (solution C), 1×10ˉ⁴ (solution D) and 1×10ˉ⁵ (solution E). Next, 1
3.3 Paecilomyces sp.
mL of each soil dilution was transferred into petri dish containing molten
rose bengal agar supplemented with 25 μg/mL chloramphenicol. The All colonies demonstrate white color with some exhibit pale green at the
medium and inoculum were mixed before the agar solidifies. After all center on PDA. The colonies shows cottony texture and filamentous
plates have solidified, they were incubated at 28˚C for 1 week. margin with a diameter range in between 3.5 cm- 4.2 cm. The conidia are
phialospore-type, 1 celled, aggregate in a row with limoniform structure;
2.3 Isolation of fungi
and conidiophores are well developed without an inflated apical cell and
The presence of fungi isolates were observed every day for 7 to 10 days. hyaline (Figure 2).
The fungi were then separated based on color and surface morphology to
Potato Dextrose agar (PDA) plates with each type of fungi was transferred
to 3 plates. Next, the plates were incubated at room temperature for 7
days. Then, in order to obtain the pure culture, the fungi colony formed
were inoculated on water agar for 5 to 7 days to isolate single spore or
hyphal tip. The single spore or hyphal tip for each isolate was then
inoculated on new PDA as the pure culture. The pure culture fungi were
inoculated on slanted PDA for long term storage.
2.4 Cultural, morphological and microscopic identification of soil
fungi isolates
For cultural and morphological characteristics of the fungi, colony color,
texture, diameter, hyphae and conidia were observed. As for microscopic
identification, the slides were prepared and examined using compound
and light microscope (Leica ICC50 HD) equipped with LAS EZ software.
Different genera are identified using identification key from Pictorial Atlas
of Soil and Seed Fungi (Watanabe, 2010).
3. RESULTS
3.1 Cultural, morphological and microscopic characterization of
soil fungi isolates
A total of 10 fungi isolates were obtained from the used agricultural soil in
GNC IIUM and were identified based on their cultural and morphological
characteristics such as colony color, reverse color, texture, margin,
presence of sulcation and colony diameter. The fungi isolates obtained can
be grouped into 4 genus based on their morphological characteristics
which are Cladorrhinum sp., Paecilomyces sp., Aspergillus sp. and
Penicillium sp. suggesting that the diversity of fungal population in the
used agricultural soil.
3.2 Cladorrhinum sp. Figure 2: Cultural and morphological characteristics of Paecilomyces sp.
Three isolates of fungi (IIUM C1, IIUM C2 and IIUM E13) exhibit white to 3.4 Aspergillus sp.
pale cream colony on PDA with a cottony texture except for IIUM E13
The colony is white yellowish on PDA with a granular texture and
which demonstrate velvety texture. The colony margin is smooth (IIUM C1
filamentous margin. It is a fast growing fungi with the colony diameter
and IIUM C2) and filamentous (IIUM E13) with the diameter range
range between 6.5 cm to 7.0 cm at 7 days. The hyphae is septate and
between 3.0 cm- 3.6 cm. The colonies exhibit septate hyphae and the
conidiophores are biseriate with apically inflated globose, and bearing
conidia are phialospore-type, 1 celled, hyaline, globose with
numerous phialides (Figure 3).
conidiophores are without an inflated apical cell (Figure 1).
Cite the Article: Nur Sabrina Ahmad Azmi, Asma Adiba Hisham (2021). Isolation and Morphological Identification of Soil Fungi from Agricultural Soil in Kuantan.
Journal Clean WAS, 5(1): 31-34.
Journal Clean WAS (JCleanWAS) 5(1) (2021) 31-34
cadmium, Cd through bioaccumulation of the heavy metal
(Woldeamanuale, 2017), thus is beneficial in water treatment systems.
5. CONCLUSION
In conclusion, 10 fungi isolate from 4 different genera were successfully
isolated from the used agricultural soil in Glasshouse and Nursery
Complex, IIUM. The results showed significant variants in morphological
characteristics thus it is speculated that the used agricultural soil harbors
Figure 3: Cultural and morphological characteristics of Aspergillus sp.
diverse fungal populations. Besides, in reference to previous studies, the
3.5 Penicillium sp. identified genus of Cladorrhinum, Penicillium, Paecilomyces and Aspergillus
sp. exhibit many potential applications that can be explored in various
Colony is green with white margin on PDA with a velvety texture and industries, not only limited to agriculture. However, further studies such
smooth margin. The colony is 3.0 cm to 3.2 cm in diameter. The colony as molecular identification is important in identifying and classifying soil
shows the presence of septate hyphae and the conidia are phialospore- fungi.
type, 1 celled, dry, and globose, with a well-developed penicillus and
conidiophores are well developed without an inflated apical cell, hyaline REFERENCES
and the spores are aggregated in a row (Figure 4).
Abdallah, R.A.B., Khiareddine, H.J., Mejdoub-Trabelsi, B., Daami-Remadi,
M., 2015. Soil-borne and compost-borne Aspergillus species for
biologically controlling post-harvest diseases of potatoes incited by
Fusariumsambucinum and Phytophthora erythroseptica. Journal of
Plant Pathology & Microbiology, 6 (10), Pp. 1–9.
https://doi.org/10.4172/2157-7471.1000313
Baxter, N.M., Williamson, J., 2001. Introduction to soils. In L. Macartney
(Ed.), Know Your Soils. Victoria: Department of Natural Resources and
Figure 4: Cultural and morphological characteristics of Penicillium sp. Environment, Pp. 3–24.
Bilal, S., Khan, A.L., Shahzad, R., Asaf, S., Kang, S.M., Lee, I.J., 2017.
4. DISCUSSION Endophytic Paecilomyces formosus LHL10 augments Glycine max L.
The used agricultural soils in GNC, IIUM harbors many potential microbes adaptation to Ni-contamination through affecting endogenous
that can be further explored for their effectiveness in agriculture and many phytohormones and oxidative stress. Frontiers in Plant Science, 8, Pp. 1–
other industries. One of the significant ability of fungi belonging to the 17. https://doi.org/10.3389/fpls.2017.00870
genus Cladorrhinum is their biocontrol potential against plant diseases.
Since the awareness of the society on environmental conservation is on Bunemann, E.K., Schwenke, G.D., Zwieten, L.V., 2006. Impact of
the rise, the use of microbial-based fungicide showed an increase in Agricultural Inputs on Soil Organisms - A Review. Australian Journal of
significance as an alternative to chemical products which will inevitably Soil Research, 44, Pp. 379–406.
cause negative impacts on the environment. The study of fungi from genus
Cladorrhinum by found that it has the ability to reduce disease incidence Coelho, E., Reis, T.A., Cotrim, M., Rizzutto, M., Corrêa, B., 2020.
in cotton crops infested by Rhizoctonia solani (Gasoni and Gurfinkel, Bioremediation of water contaminated with uranium using Penicillium
2009). In addition, cotton plants growing in phosphorus deficient piscarium. Biotechnology Progress, Pp. 1–30.
substrate showed a significant increase in plant height and phosphorus https://doi.org/10.1002/btpr.3032
content at blossom stage when the root were colonized with C.
feocundissimum (Gasoni and Gurfinkel, 1997). Darmody, R.G., Daniels, W.L., Marlin, J.C., Cremeens, D.L., 2009. Topsoil:
what is it and who cares? Journal American Society of Mining and
This proved that the infestation by C. foecundissimum could increase the Reclamation, (1), Pp. 237–269.
availability of phosphorus for root uptake. However, the mechanisms on
how the fungi enhance nutrient uptake are yet to be known. Aside of García-Orenes, F., Morugán-Coronado, A., Zornoza, R., Scow, K., 2013.
Cladorrhinum sp., the biocontrol ability of some Aspergillus sp is also Changes in soil microbial community structure influenced by
notable. For instance, 3 Aspergillus isolates inhibited the growth of agricultural management practices in a Mediterranean agro-
Fusarium oxysporum, a causal agent of Fusarium wilt in banana (Hidayat et ecosystem. PLOS ONE, 8 (11), Pp. 1–9.
al., 2019). Aside of that, Aspergillus strains were found to have reduce
mycelial growth of F. sambucinum and Phytophthora erythroseptica by Gasoni, L., Gurfinkel, B.S., 1997. The endophyte
27% to 68% and 16% to 25% respectively, therefore having potential to Cladorrhinumfoecundissimum in cotton roots: phosphorus uptake and
be used as natural fungicide (Abdallah et al., 2015). In addition, the host growth. Mycological Research, 101 (7), Pp. 867–870.
inoculation of A. niger in garlic portrayed positive impact in overcoming
https://doi.org/10.1017/s0953756296003462
wilt caused by Fusarium while at the same time increased the plant growth
by 30% to 40% (Sugiharto, 2019). Gasoni, L., Gurfinkel, B.S., 2009. Biocontrol of Rhizoctonia solaniby the
endophytic fungus Cladorrhinum foecundissimumin cotton
Furthermore, some species of Penicillium and Paecilomyces had been
plants. Australasian Plant Pathology, 38 (4), Pp. 389–391.
explored as potential bioremediator. For instance, Penicillium citrinum
isolated from mangrove soil could absorb heavy metals notably lead, Pb https://doi.org/10.1071/ap09013
(II) in an optimum condition where pH is 7, temperature of 30˚C and 60
Glick, B.R., 2018. Soil microbes and sustainable
minutes contact time (Wahab et al., 2017). A study also supported the
same result but using different species; Penicillium janthinellum and agriculture. Pedosphere, 28 (2), Pp. 167–169.
Penicillium brasilianum which removed 91.2% and 74.4% lead
Gruda, N.S., 2019. Increasing Sustainability of Growing Media Constituents
respectively (Martins et al., 2016). In addition, a most recent research
revealed the potential of Penicillium piscarium an important alternative for and Stand-alone Substrates in Soilless Culture Systems. Agronomy, 9 (6),
treating water contaminated with uranium. This fungus was able to Pp. 298. https://doi.org/10.3390/agronomy9060298
remove between 93.2% to 97.5% uranium from solution contaminated
with of 1 to 100 mg Lˉ¹ of uranium nitrate at pH 3.5, increasing the Gryndler, M., Larsen, J., Hršelová, H., Řezáčová, V., Gryndlerová, H., Kubát,
solution’s pH to levels above 5.6 (Coelho et al., 2020). As for Paecilomyces J., 2006. Organic and mineral fertilization, respectively, increase and
sp., P. formosus could accumulate nickel, Ni in the mycelia without affecting decrease the development of external mycelium of arbuscular
its growth rate. This resulted in lower intake of Ni by soybean that was mycorrhizal fungi in a long-term field experiment. Mycorrhiza, 16, Pp.
inoculated with the fungi and therefore alleviates the stress induced by Ni 159–166.
toxicity (Bilal et al., 2017). In addition, two Paecilomyces strain isolated
from industrial dumping area could tolerate up to 6000 mg Lˉ¹ of Hidayat, I., Dewi, L.C., Sukmawati, D., 2019. Antagonistic activity of three
Cite the Article: Nur Sabrina Ahmad Azmi, Asma Adiba Hisham (2021). Isolation and Morphological Identification of Soil Fungi from Agricultural Soil in Kuantan.
Journal Clean WAS, 5(1): 31-34.
Journal Clean WAS (JCleanWAS) 5(1) (2021) 31-34
Aspergillus isolates against Fusarium wilt of banana. Journal of and Fusarium sp. inoculant. IOP Conference Series: Earth and
Microbial Systematics and Biotechnology, 1 (1), Pp. 1–10. Environmental Science, 308, Pp. 1–11. https://doi.org/10.1088/1755-
https://doi.org/10.37604/jmsb.v1i1.16 1315/308/1/012058
Lori, M., Symnaczik, S., Mader, P., Deyn, G.D., Gattinger, A., 2017. Organic Tuğrul, K.M., 2020. Soil Management in Sustainable Agriculture.
farming enhances soil microbial abundance and activity meta-analysis Sustainable Crop Production, Pp. 1–16.
and meta-regression. PLOS ONE, 12 (7), Pp. 1–25. https://doi.org/10.5772/intechopen.88319
Martins, L.R., Lyra, F.H., Rugani, M.M.H., Takahashi, J.A., 2016. Wahab, A.A., Awang, A.S.A.H., Azham, Z., Tay, M.G., Adeyemi, F.M., 2017.
Bioremediation of metallic ions by eight Penicillium species. Journal of Biosorption of lead (II) ion using Penicillium citrinum KR706304
Environmental Engineering, 142 (9), Pp. 1–8. isolated from the mangrove soil environment of southeast Borneo. Ife
https://doi.org/10.1061/(asce)ee.1943-7870.0000998 Journal of Science, 19 (2), Pp. 341–349.
https://doi.org/10.4314/ijs.v19i2.14
Nortcliff, S., Hulpke, H., Bannick, C.G., Terytze, K., Knoop, G., Bredemeier,
M., Dworshak, U., 2006. Soil, definition, function and utilization of Watanabe, T., 2010. Pictorial atlas of soil and seed fungi: morphologies of
soil. Ullmanns Encyclopedia of Industrial Chemistry, 33, Pp. 399–419. cultured fungi and key to species, Third Edition (Mycology) (3rd ed.).
CRC Press.
Pepper, I.L., Gerba, C.P., 2004. Filamentous fungi. In E. R. Loya
(Ed.), Environmental Microbiology: A Laboratory Manual, Pp. 27–36. Woldeamanuale, T.B., 2017. Isolation, screening and identification of
UK: Elsevier Academic Press. cadmium tolerant fungi and their removal potential. Journal of Forensic
Sciences & CriminalInvestigation, 5 (2), Pp. 1–
Sugiharto, A., 2019. Response of growth of garlic towards Aspergillus niger 7.https://doi.org/10.19080/jfsci.2017.05.555656.
Cite the Article: Nur Sabrina Ahmad Azmi, Asma Adiba Hisham (2021). Isolation and Morphological Identification of Soil Fungi from Agricultural Soil in Kuantan.
Journal Clean WAS, 5(1): 31-34.
