FORMULATION AND TESTING OF LOCALLY ISOLATED EFFECTIVE
MICROBES FOR THE DEVELOPMENT OF BIOFERTILIZERS
Shruti Prashant Talwar
Bachelor of Science with Honours
(Biotechnology Resource)
2013
Faculty of Resource Science and Technology
FORMULATION AND TESTING OF LOCALLY ISOLATED EFFECTIVE
MICROBES FOR THE DEVELOPMENT OF BIOFERTILIZERS
SHRUTI PRASHANT TALWAR
This project report is submitted in partial fulfilment of the requirement for the degree
of Bachelor of Science with Honours (Resource Biotechnology)
Resource Biotechnology Programme
Department of Molecular Biology
Faculty of Resource Science and Technology
Universiti Malaysia Sarawak
2013
ACKNOWLEDGEMENT
Above all, I am grateful to God for all his blessings.
I owe sincere gratitude, to my supervisor, Dr Awang Ahmad Sallehin Awang Husaini for
giving me this invaluable opportunity to work under him and also for his patience,
motivation, enthusiasm, and immense knowledge and most important of all, for believing in
me. His guidance has helped me through this research and writing of this thesis. I could not
have imagined having a better advisor and mentor for my final year project.
Also, I would like to express my deepest gratitude to my amazing parents for their
never – ending love and support through good times and through challenging times.
I would also like to acknowledge 1IPTA 1Menteri (Kementerian Tenaga, Teknologi Hijau
dan Air), for providing a financial grant for my research.
I would also like to acknowledge and express my thanks to the postgraduate students from
the Molecular Genetics lab for all their help and guidance.
And last but not the least I would like to thank my dearest friends who have come into my
life and inspired, touched, and illuminated me with their presence.
Table of Contents
Title Page
Acknowledgement………………………………………………………... I
Table of Contents…………………………………………………………. II
List of Abbreviations…………………………………………………….. V
List of Tables and Figures………………………………………………… VI
Abstract.....………………………………………………………………... 1
1.0 Introduction ......................................................................................... 2
2.0 Literature Review………………………………................................. 6
2.1 Sustainable Agriculture………………………………………….. 6
2.2 Biofertilizer ……………………………………………………... 7
2.3 Effective Microbes………………………………………………. 8
2.3.1 Plant Growth Promoting Rhizobacteria (PGPR)………...... 10
2.3.2 Nitrogen Fixing Bacteria (NFB)………………………….. 11
2.3.3 Phosphate Solubilizing Bacteria (PSB)…………………… 12
2.4 Compost as the Natural Carrier…………………………………... 12
2.5 Biofertilizer in Malaysia………………………………………… 13
3.0 Materials and Method………………………………………………... 16
3.1 Soil Sampling…………………………………………………… 16
3.2 Media Preparation ……………………………………………… 16
3.3 Sampling………………………………………………………... 19
3.4 Isolation………………………………………………………… 19
3.5 Characterization and Identification……………………………... 19
3.6 Genomic DNA isolation………………………………………… 20
3.7 16S rDNA Analysis……………………………………………... 21
3.8 Polymerase Chain Reaction…………………………………….. 21
3.9 Agarose Gel Electrophoresis……………………………………. 22
3.10 Maintenance of Isolates………………………………………... 22
3.11 Developing Biofertilizer using OPEFB As Base Medium…….. 23
3.12 Compost Analysis……………………………………………... 23
3.12.1 Bacterial Count………………………………………….. 23
3.12.2 Moisture Content………………………………………… 24
3.12.3 pH ……………………………………… ……………….. 24
3.12.4 Phytotoxicity Test……………………………………… 25
3.12.5 Germination Index………………………………………. 25
3.13 Pot Trial Testing……….……………………………………… 26
4.0 Results and Discussion…………………………………………. 27
4.1 Characterization and Identification…………………………. 27
4.2 Genomic DNA Isolation…………………………………….. 28
4.3 Polymerase Chain Reaction………………………………… 29
4.4 DNA Purification……………………………………………. 30
4.5 Sequencing Results…………………………………………. 31
4.6 Compost Analysis ………………………………………….. 33
4.6.1 Bacterial Count……………………………………….. 33
4.6.2 Moisture Content………………………………………. 34
4.6.3 pH……………………………………………………... 35
4.6.4 Phytotoxicity Test……………………………………. 36
4.7 Pot Trials…………………………………………………… 37
5.0 Conclusion ………………………………………………........... 38
6.0 References……………………………………………………... 40
Appendix A: Sequencing results of unknown isolates using 16S r DNA 45
List of Abbreviations
AMF Arbuscular mycorrhizial fungi
BNF Biological nitrogen fixation
BLAST Basic local alignment tool
CTAB Cetyl trimethylammonium bromide
DNA Deoxyribonucleic acid
EM Effective microbes
LB Luria Bertani
NFB Nitrogen fixing bacteria
NA Nutrient agar
PGPM Plant growth promoting microbes
PGPR Plant growth promoting rhizobacteria
PSB Phosphate solubilizing bacteria
List of Tables and Figures
Tables
Table 1: Optimized PCR reaction mixture……………………………………………………… 21
Table 2: Optimized thermocycling profile………………………………… 21
Table 3: Morphological characteristics of the unknown isolate…………... 27
Table 4: Profiles of moisture content of inoculated compost and control
compost……………………………………………………………….
34
Table 5: Germination Index of water spinach seeds in inoculated compost and
control compost on day 30 of composting period……………………..
36
Table 6: Statistical analysis of the pot trials using t – test……………………... 37
Figures
Figure 1: Results of Gram staining visualized with 1000 X modification…….. 27
Figure 2: Visualised genomic DNA extracted from unknown bacteria………. 28
Figure 3: Visualized PCR product showing dominant band of 1500bp……….. 29
Figure 4: Results of purification of the PCR product………………………….. 30
Figure 5: Total bacterial count on day 10 in the compost inoculated with
bacteria and control compost………………………………………….
34
Figure 6: Total bacterial count on day 30 in the compost inoculated with
bacteria and control compost………………………………………….
34
Figure 7: pH profiles of inoculated compost and control compost…………….. 35
1
Formulation and testing of locally isolated effective microbes for the development of
biofertilizers
Shruti Prashant Talwar
Resource Biotechnology Faculty of Resource Science and Technology
Universiti Malaysia Sarawak
ABSTRACT
The soil is a complex and heterogeneous environment constituting a large diversity of effective microbes
which can be utilized as biofertilizers. In the present study, local soil samples were collected for isolation of
beneficial bacteria. Genomic DNA was extracted from the unknown isolate and subjected to the
amplification of 16S rDNA gene for identification. The similarity searching of the sequence obtained after
sequencing showed 99% similarity with Enterobacter Cloacae, a gram negative rod shaped bacteria
belonging to the community of PGPR. This makes it a promising strain to be developed as a biofertilizer.
Additionally, biofertilizers were developed using four bacterial isolates namely, Bacillus cereus, Bacillus
amyloliquefaciens, Bacilus licheniformis and Pseudomonas aeruginosa This consortium was then inoculated
onto OPEFB compost and compared with uninoculated compost as control. Over a 30 day period, various
types of compost analysis were also conducted. On day 30, the moisture content of inoculated compost was
83.58%. The pH was slightly acidic at 6.42 with bacterial count higher than uninoculated compost. The
Germination Index (GI) was at 95% indicating that the compost was mature and free from phytotoxins. After
curing, pot trials evaluating the effects of this formulation on the growth factors of Capsicum annum L,
showed a significant increase in root and shoot length. Hence, combination of these bacterial strains could be
a good biofertilizer for sustainable agriculture.
Key Words: Biofertilizer, Enterobacter Cloacae, consortium, OPEFB compost.
ABSTRAK
Tanah adalah satu persekitaran yang kompleks dan berbeza-beza yang membentuk satu kepelbagaian
mikrob efektif yang mungkin boleh digunakan sebagai baja bio. Dalam kajian ini, sampel tanah tempatan
telah dikumpulkan untuk pengasingan bakteria berfaedah. DNA genomik telah diambil dari pengasingan
yang tidak diketahui dan merujuk kepada amplifikasi gen 16S rDNA untuk pengenalan. Persamaan mencari
jujukan diperolehi setelah jujuk menunjukkan 99% persamaan dengan Enterobakter cloacae, satu rod gram
negatif membentuk bakteria kepunyaan komuniti PGPR. Ini menjadikan ia satu terikan yang baik untuk
dikembangkan sebagai satu baja bio. Sebagai tambahan, baja bio dikembangkan menggunakan empat
bakteria terasing iaitu, Bacillus cereus, Bacillus amyloliquefaciens, Bacilus licheniformis dan Pseudomonas
aeruginosa. Konsortium ini kemudiannya diinokulat ke kompos OPEFB dan dibanding dengan kompos
tanpa inokulasi sebagai kawalan. Selepas tempoh 30 hari, pelbagai jenis analisis baja telah dijalankan.
Pada hari ke 30, kandungan kelembapan kompos disuntik adalah 83.58%. pH adalah sedikit berasid pada
tahap 6.42 dengan kiraan bakteria yang tinggi daripada baja pokok tanpa inokulasi. Indeks Percambahan
(GI) adalah pada 95% menunjukkan bahawa kompos itu matang dan bebas daripada phytotoxins. Selepas
pengawetan, ujian periuk menilai kesan penggubalan ini kepada faktor-faktor pertumbuhan Capsicum
annum L, menunjukkan peningkatan yang ketara dalam akar dan panjang menembak. Oleh itu, gabungan
jenis bakteria ini boleh menjadi biobaja baik untuk pertanian lestari.
Kata Kunci: Biofertilizer, Enterobakter cloacae, konsortium , kompos OPEFB.
2
1.0 INTRODUCTION
Agriculture has sustained human lives since ancient times. Today, with global population
exceeding 7 billion, agriculture inexorably continues to play a very important role in the
survival of mankind.
Since many years, farmers have been depending on chemical fertilizers for enhancing
the growth of plants. But the continuous use of these chemical fertilizers and pesticides has
not only made it difficult to sustain the soil fertility but also contaminated and damaged
soil health. Hence, it has become necessary to reduce the use of chemical fertilizers in
order to lessen the pressure on the environment due to irresponsible agricultural practises.
Nowadays, factors such as soil degradation, chemical pollution, the demand for safe
food and more importantly, the rising cost of petroleum have forced farmers to seek other
alternatives (Zakaria, 2006). This has also led researchers to devise methods to increase
soil fertility and to develop sustainable agricultural cultivation techniques, more than ever
before.
In this context, biofertilizers can be considered as key components of integrated
nutrient management (Mohammadi & Sohrabi, 2012). The term “Biofertilizer” or more
appropriately “Microbial inoculants” can generally be defined as preparation containing
live or latent cells of efficient strains of Nitrogen fixing, Phosphate solubilising or
cellulolytic microorganisms used for application to seeds, soil with the objective of
increasing the number of such microorganisms and accelerate those microbial process
which augment the availability of nutrients that can be easily assimilated by plants (Pandit
et al., 2011). Biofertilizers enhance the productivity and sustainability of soil as they are
3
low cost, eco-friendly and renewable source of plant nutrients. Thereby, they play an
important role in sustainable agricultural system.
In recent years there has been an upsurge into the research related to biofertilizers
since they act as natural stimulators of plant growth and development. Consequently, there
is considerable interest in the possible use of inoculants of effective microbes for the
development of biofertilizer. This mainly involves the selection and multiplication of
plant-beneficial microorganisms such as bacteria, algae and fungi, either alone or in
combination.
Since, bacteria are extremely perishable and sensitive to environmental factors, there is
always a need for developing efficient and hardy strains of these microbes which can
withstand local ecological conditions, replenish soil fertility and improve nutrient uptake
for the plant. These improved strains can also be designed so that they are suitable for
different conditions and thereby for greater crop diversification.
The formulation of biofertilizer typically consists of establishing viable bacteria in a
suitable carrier together with additives that aid in stabilization and protection of microbial
cell during storage, transport and at the target (Brahmaprakash & Sahu, 2012). A good
quality carrier material for the microbial inoculants should generally consist of carbon,
nitrogen and vitamin sources, which can promote growth and survival of bacteria.
Compost is one such carrier material which has these characteristics.
Compost is a natural agro-management method, utilizing agricultural waste and
indigenous soil microorganisms (Phua et al., 2012). Composting through the modified
‘Natural Farming’ method, has been gaining acceptance in several countries and in
Malaysia, by the Department of Agriculture (Phua et al., 2012). It is a simple and cheap
4
method to turn wastage like empty fruit bunches (EFB) of oil palm industries into
compost. Composts which are inoculated with biofertilizer containing appropriate
functional microbes increase the decomposition rate, shorten the maturity period and
thereby improve the compost quality (Wei et al., 2007).
At present, Malaysia is striving to adopt sustainable and zero waste agricultural
practises in response to growing environmental concerns. And, in keeping with the spirit of
times, there is an ongoing attempt to promote the use of biofertilizers in the Malaysian
farming systems so that they can be used as eco-friendly and cost effective inputs by the
farmers.
The ultimate goal of sustainable agriculture is to develop farming systems that are
productive, profitable, energy conserving, environmentally sound, conserving of natural
resources and that ensures food safety and quality (Namasivayam & Kirithiga, 2010). This
study endeavours to give a new input into the research of using biofertilizers containing
effective microbes so as to move away from conventional and chemical based agriculture.
This basically involves reducing the loss to the environment by increasing organic
farming systems and decreasing the input of inorganic chemical fertilizers. Hence, the
need for the amalgamation of microbial waste management into agro-industries, and their
roles in creating a better environment and sustain agriculture, cannot be over emphasized.
In view of these facts, the main objectives of this research study were:
• Isolation and characterization of unknown bacteria from local soil samples.
• Formulation of Bacillus cereus, Bacillus amyloliquefaciens, Bacilus licheniformis and
Pseudomonas aeuriginosa for the development of biofertilizer, so as to enhance plant
yield and growth.
5
• To characterize the biofertilizer produced in terms of their physiochemical and
microbiological properties.
• Pot trial testing using the formulated biofertilizer, on selected plants so as to enhance
plant yield and growth.
6
2.0 LITERATURE REVIEW
2.1 Sustainable Agriculture
Over the past decades, world population has increased dramatically. According to United
Nations (UN, 2009) reports, the global population, which was approximately 6 billion in
2000 is likely to increase to 9 billion by the year 2050. With focus on feeding a rapidly
growing human population, world agriculture too has shown phenomenal growth. This
increase in crop productivity has mainly been due to intensive off- farm inputs such as
synthetic chemical fertilizers and pesticides.
Unfortunately, the excessive and indiscriminate use of these chemical inputs for
enhancing agricultural production has led to a number of environmental problems. Amidst
the current situation, there is a growing awareness and concern about the harmful effects of
these agrochemicals which has led to increased demand for sustainable food production
and agriculture. Therefore, improving agricultural sustainability has become an important
goal (Food and Agriculture Organization of the United Nations [FAO], 2002).
The term ‘sustainable agriculture’ implies regenerative practices which optimally use
locally available resources and natural processes, such as nutrient recycling; build on
biodiversity; regenerate and develop natural resources; and limit the use of external inputs
of agro-chemicals, minerals and non-renewable energy (Roling & Wagemakers, 1998).
The challenge for agriculture over the coming decades, therefore, will be to meet the
worlds increasing demand for food in a sustainable way (Gruhn et al., 2000)
In order to reach such a situation, research and development plays a key role in
providing the technologies and products to enable all this to happen. In this present
7
context, the role of biofertilizers in sustainable agriculture assumes special significance
(Kannaiyan & Kumar, 2006).
2.2 Biofertilizer
According to Mosttafiz et al. (2012), the most promising strategy to reach the goal of
sustainable agriculture, is to substitute hazardous agrochemicals with environment-friendly
preparations for symbiotic microbes which could improve the nutrition of crops and
livestock, as well as their protection from biotic (pathogens, pests) and abiotic (including
pollution and climate change) stresses. Consequently, biofertilizers have emerged as one of
the alternatives for transition towards more sustainable development pathways through
biological nitrogen fixation (BNF) (Sangar, 2010) and have become important components
of integrated nutrient management (Mohammadi & Sohrabi , 2012).
According to the definition proposed by Vessey (2003), Biofertilizer is a substance
which contains living microorganisms which, when applied to seed, plant surfaces, or soil,
colonizes the rhizosphere or the interior of the plant and promotes growth by increasing
the supply or availability of primary nutrients to the host plant.
Research in the field of biofertilzer has resulted in the development of different kinds
of microbial inoculant or biofertilizers such as nitrogen fixing bacteria, phosphate
solubilizing microorganism, vesicular-arbuscular mycorrhizae (VAM) and PGPR
(Dhamangaonkar & Misra, 2009). Moreover, studies on the interaction between plant, soil
and the different microorganisms have shed light on their inter-relationships thus providing
new possible ways to exploit them for agricultural purposes (Malusa et al., 2011).
As reported by Wani et al, (1995), biofertilizers are now being increasingly used as a
part of Integrated Plant Nutrient System (IPNS) that advocate involving a combination of
8
fertilizers, organic manures and microbial inoculants which are imperative to sustain crop
production and maintain soil health and soil diversity in the long run. The use of BNF
technology for maintenance of soil health and sustainable agriculture can be an alternate to
chemical fertilizer.
According to Chien et al. (2007), the main and direct purposes of applying
biofertilizers to the soil are: to provide nutrient sources and good soil conditions for the
growths of crops when used as a live body; to partially substitute and enhance the function
of chemical fertilizer and then subdue the application quantities of fertilizers and still
maintain the same crop yields and the capital used for making bio-fertilizers is cheaper
than that of chemical fertilizers and to lessen the negative effect aroused from applying
chemical fertilizers to soil. Chien et al. (2007) also stated that, the indirect purposes of
using bio-fertilizers to soil are: to enhance the growth of root system to increase the water
and nutrient absorption abilities of crops, extend the life of root, neutralize and degrade
harmful materials accumulated in soil, promote survival efficiency of seedling after
transplanting and get shorter time for the flower to come out.
Thus the manifold advantages of biofertilizer leads to its wide applicability in
sustainable agriculture (Bashan, 1998). The success of biofertilzer depends on several
factors, such as selecting the most effective microbial strain, seeking optimum conditions
for its growth, formulation of the inocula and the method of its application (Bashan, 1998).
2.3 Effective Microbes
Microbes are an important component of world biodiversity (Phua et al., 2012). Effective
Microorganisms (EM), is a concept suggested by Professor Teruo Higa. (Higa & Parr
,1994) from the University of Ryukyus, Japan, and it consists of beneficial and naturally
9
occurring microorganisms that can be applied as inoculants so as to shift the soil
microbiological equilibrium in ways that can improve soil quality, enhance crop
production and protection, conserve natural resources, and ultimately create a more
sustainable agriculture and environment. According to Abdullah et al. (2011), EM
solutions which were used for the preparation of biofertilizer helped to increase the
number of beneficial microbes in the soil; which in turn improved the soils microbial
health and thereby promoted a healthy environment for plants.
According to Parr et al. (2010), the exact mechanisms of how EM interacts and
functions in the soil-plant ecosystem is not known. However, there is evidence that
supports several theories concerning its action including a) suppression of plant pathogens
and diseases, b) enhanced nutrient availability, c) stimulated plant growth (i.e., auxin-
mediated effects), and d) improved root surface-rhizosphere relationships (Higa &
Wididana, 1991 as cited in Parr et al., 2010).
Numerous field and greenhouse trials indicate the benefits of EM as a biofertilizer in
crop production, as a probiotic in poultry and livestock rations, and as a starter to improve
composting and recycling of municipal/industrial wastes and effluents (Hussain et al.,
1999 as cited in Javaid, 2010).
Biofertilisers based on beneficial and effective microorganisms belong to a wide array
of genera, classes and phyla; ranging from bacteria to yeasts and fungi (Malusa et al.,
2012). A specific group of this kind of biofertiliser includes products based on plant
growth-promoting microorganisms (PGPM), which include nitrogen fixing
microorganism, mycorrhizal fungi and plant growth-promoting rhizobacteria (Sahai,
1999).
10
The prospects for improved agriculture by the use of effective microbial inoculant as
biofertiliser are particularly good, especially in developing countries, because they give
better yields, have lower costs and thereby lead to reduced dependence on chemicals
(Sahai, 1999).
2.3.1 Plant Growth Promoting Rhizobacteria (PGPR)
Most of the bacteria included in biofertilizer have close relationship with plant roots.
Rhizobium has symbiotic interaction with legume roots and rhizobacteria inhabit on root
surface or rhizosphere of soil (Forum for Nuclear Cooperation in Asia [FNCA], 2006).
These species of soil bacteria flourish in the rhizosphere of plants, may grow in, on, or
around plant tissues and stimulate plant growth (Muraleedhara et al., 2010). They are
collectively known as PGPR and among them are strains from genera such as Alcaligenes,
Acinetobacter, Arthrobacter, Azospirillum, Bacillus, Burkholderia, Enterobacter, Erwinia,
Flavobacterium, Paenibacillus, Pseudomonas, Rhizobium, and Serratia (Sharma et al.,
2011).
PGPR have been reported to directly enhance plant growth by a variety of
mechanisms: fixation of atmospheric nitrogen, solubilisation of minerals such as
phosphorus, production of siderophores, and synthesis of plant growth hormones i.e.
Indole-3- acetic acid (IAA), gibberellic acid, cytokinins, and ethylene (Nelson, 2004 as
cited by Kumar et al., 2012) Indirect mechanisms involves the biological control of plant
pathogens and deleterious microbes, through the production of antibiotics, lytic enzymes,
hydrogen cyanide, catalase and siderophore or through competition for nutrients and space
11
can improve significantly plant health and promote growth, as evidenced by increases in
seedling emergence, vigour, and yield (Khan, 2006 as cited by Kumar et al., 2012).
A range of PGPR have shown their ability to significantly increase the vegetative
growth and grain yield of crop plants like rice, wheat, maize, sugarcane and cotton
(Figueiredo et al., n.d).
2.3.2 Nitrogen Fixing Bacteria (NFB)
Nitrogen which is one of the major nutrients required for the growth of crops is often
limited. Bacteria mediate fixation of nitrogen at temperature and pressure enzymatically,
by a process known as biological nitrogen fixation (BNF) (Brahmaprakash & Sahu, 2012)
According to Mohammadi and Sohrabi (2012), nitrogen-fixing bacteria (NFB)
transforms inert atmospheric N2 to organic compounds; and are grouped into free-living
bacteria (Azotobacter and Azospirillium) and the blue green algae and symbionts such as
Rhizobium, Frankia and Azolla. According to Brahmaprakash and Sahu, (2012) biological
nitrogen fixation by prokaryotes is a beneficial process in returning nitrogen to the soil
towards crop production and leading to sustainable nutrient management.
Nitrogen-fixing bacteria of Azotobacter and Azospirillum genera have been widely
tested to increase yield of cereals and legumes under field conditions. Azolla biofertilizer
were used for rice cultivation in different countries such as Vietnam, China, Thailand and
Philippines; and the field trials indicated that rice yields increased by 0.5-2 t/ha (Gupta,
2004 as cited by Mohammadi & Sohrabi, 2012). According to Mohammadi and Sohrabi
(2012), co-inoculation of some Pseudomonas and Bacillus strains along with effective
Rhizobium spp. has shown to stimulate chickpea growth, nodulation and nitrogen fixation.
12
2.3.3 Phosphate Solubilizing Bacteria (PSB)
After nitrogen fixation, phosphate solubilisation is a very important plant growth
promoting activity. Several soil bacteria particularly belonging to genera Bacillus and
Pseudomonas, possess the ability to change insoluble forms into soluble form by secreting
organic acids as formic acid, acidic, propionic, lactic, glycolic, fumaric and succinic acid
(Vazquez et al., 2000). According to Sharma et al. (2011) phosphorus solubilizing bacteria
play an important role in phosphorus nutrition by enhancing its availability to plants
through the release from inorganic and organic soil phosphorous pools, by solubilisation
and mineralization.
Crop plants such as peanut, various horticultural plants and vegetables were
successfully inoculated with PSBs to obtain higher yields. PSB such as Pseudomonas spp.
enhanced the number of nodules, dry weight of nodules, yield components, grain yield,
nutrient availability and uptake in soybean crop (Sharma et al., 2011). Several field
experiments concluded that PSBs not only improved the growth and quality of crops but
also drastically reduced the usage (by 1/3-1/2) of chemical or organic fertilizers (Chien et
al., 2007). As reported by Sharma et al. (2011) Phosphate solubilizing bacteria enhanced
the seedling length of Cicer arietinum while co-inoculation of PSM and PGPR reduced
phosphorous application by 50 % without affecting corn yield.
2.4 Compost as the natural carrier
According to Bhramaprakash and Sahu (2012), carrier is a delivery vehicle which is used
to transfer live microorganism from an agar slant of laboratory to a rhizosphere and
therefore plays a major role in formulating microbial inoculant.
13
Compost can be a good nutrient carrier material for bacterial biofertilizer. It can
support the growth and survival of bacteria. It is a biodegradable and a non-polluting
material, since it is generally prepared from naturally abundant waste materials.
According to Mwegoha (2012), composting has been established as one of the low
cost alternatives for minimizing the volume of solid waste disposed off to the environment
with the potential of economic gain from resource recovery. This viable means of
transforming various organic wastes into products can be used safely and beneficially as
biofertilizers and soil conditioners (Parr et al., 2010). According to Yeoh et al. (2011), by
converting the biowastes into composts, the nutrients in the waste can be harnessed and
potentially utilized as a valuable soil amendment, hence creating a zero waste process.
2.5 Biofertilizer In Malaysia
In the early years of developing the agricultural sector, Malaysia has relied heavily on
conventional methods to produce, increase and sustain food production (Faridah, 2001).
However, in recent years responding to environmental issues, the nation is steadily
adopting sustainable agricultural practises. According to Phua et al. (2012), enhancement
of biodiversity and agrowaste management are the approaches towards sustainability. This
can be implemented by introducing integrated agriculture, with main emphasis on organic
farming and use of organic matter, composting, conservation measure and production of
organic fertilizers using the available agricultural waste (Faridah, 2001)
According to Zakaria (2006), soil enhancers in the form of compost, indigenous
microbes and enzymes from natural farming technology; effective microbes and
Arbuscular-mycorrhizal fungi are all the alternatives that are presently being used. Several
14
states in Malaysia have reported increased plant growth, weight and sizes of plants, using
these soil-enhancing technologies (Zakaria, 2006).
The effective microbe (EM) technology brought from Japan, which makes use of
isolated groups of specific microbes such as photosynthetic bacteria, lactic acid bacteria
and yeasts, is currently being practised in many states of Malaysia, including Sarawak
(Zakaria, 2006). These effective microbes have been used to accelerate decomposition of
organic residues and agricultural byproducts through various stages, with a concomitant
release of plant nutrients through mineralization process, as evident from the good, healthy
harvest of crops (Rahim, 2002).
Malaysia has an abundance of agricultural waste that can be turned into compost
(Zakaria, 2006). At present empty fruit bunches (EFB) of oil palm are one of the
agricultural waste that are building up at alarming rates at palm oil factories (Phua et al.,
2012). This waste product which is hazardous to the environment if disposed off
indiscriminately, can be treated, recycled and turned into valuable product like compost
(Faridah, 2001).
According to Rahim (2002) there is a great potential for the biofertilizer industry in
Malaysia, producing products from local sources and natural resources. However most of
the microbial inoculants available in the market are imported (Zakaria, 2006). Therefore
for quarantine purpose the Ministry of Agriculture advocates the production of EM for
biofertilizer, using selected local microorganisms (Zakaria, 2006). In Malaysia,
mycorrhizal products are perceived to be more versatile than others and therefore it has
greatly appealed to the agricultural industry. Nevertheless, according to Rahim (2002)
there is also a good potential for biofertilizer products based on Azorhizobium and
Azospirillum.
15
Effective microorganisms that have the ability to degrade fat, lignin, cellulose and
hemicelluloses are given priority while preparing the inoculum specifically for the EFB
substrates (Yeoh et al., 2011). But there is always a challenge with microbial products and
hence research in the field of effective microbes will enhance biofertilizer use in the
country. Therefore, in the current attempt to make the agriculture industry in Malaysia a
viable component of a healthy and pleasant ecosystem, the use of biofertilizer and other
microbial products is very crucial (Rahim, 2002).
