FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Issue No. 5
July 2005
Message from Malaysia
Dear Colleagues,
I am honoured to be given the opportunity to coordinate inputs on biofertilizer research andbiofertilizer industry in Malaysia in the present issue of the FNCA Biofertilizer Newsletter. There aremany institutions in Malaysia that are involved in scientific research on biofertilizer microorganisms
and biofertilizer inoculum productions, amongst them are: Universiti Pertanian Malaysia (UPM),
Malaysian Agricultural Research and Development Institute (MARDI), Malaysian Rubber Board(MRB), Universiti Sains Malaysia (USM), Universiti Malaya (UM), Universiti Kebangsaan Malaysia(UKM) and Malaysian Institute for Nuclear Technology Research (MINT).
Of these, MRB, or Rubber Research Institute Malaysia (RRIM), as it was formerly known, beganearliest in the 1970’s in mass production of Rhizobium inoculum, for use on leguminous cover crops inrubber plantations. The production of the inoculum in latter years includes sterilization of coconut coir
dust as carrier material, packed in sturdy and commercializable packets, at the MINTec-SINAGAMA
gamma irradiation plant in MINT. Use of 15N to quantify biological nitrogen fixation contribution inparticular cropping systems and to evaluate efficiency of different N2 fixing microorganisms, including
Bradyrhizobium and Azolla was also noted, as recorded in publications by scientists from universities
and research institutions in Malaysia since the 1980’s. Application of nuclear technology inbiofertilizer research and biofertilizer production is thus quite close to the hearts of many of us.
Private enterprises and companies have been
actively involved in production and marketingof biofertilizer products; Malaysian Agri HiTech Sdn. Bhd. has been producing arbuscular
mycorrhizal fungi (AMF) inocula suitable foruse in plant nurseries, oil palm plantations,fruit orchards, golf courses and landscape on alarge scale.
With agriculture (food production, foodsafety) and biotechnology being in thelimelight at present in the country, it isenvisaged that the local biofertilizer industry
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Projectwill flourish, too, capitalizing on the fusion of biotechnology and other technologies including radiation
technology.
This issue will highlight on AMF and the N2 fixers. Many thanks to all contributors; it is also mypersonal wish that the information collated here will benefit all readers.
Happy reading.
Thank you and Warm Regards.

Khairuddin Abdul Rahim
FNCA Biofertilizer Project Leader, Malaysia

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

USE OF 15N ISOTOPE DILUTION TECHNIQUE IN FIELD EVALUATIONOF N2 FIXATION IN YOUNG OIL PALM
INOCULATED WITH BACILLUS SPHAERICUS UPMB 10
Zakry, F.A.A.1, Anuar, A.R.1, Zin, Z.Z.2, #Shamsuddin, Z.H.1 and Khairuddin, A.R.3
1
Universiti Putra Malaysia,
2
Malaysian Palm Oil Board,
3
Malaysian Institute for Nuclear Technology Research# zulsham@agri.upm.edu.my
Introduction2001). N2-fixing Bacillus sphaericus UPMB 10
is a Gram-variable bacteria isolated locally fromoil palm roots (Shamsuddin and Roslina, 1995).
Results from nursery trials have shown UPMB
10 strain to be a potential biofertilizer for oilpalm seedlings with a N2 fixing capacity of up to
33% Ndfa (539 mg N plant-1) in non-sterile soil
(Amir, 2001). However, beneficial effects of theplant growth promoting rhizobacteria (PGPR)
have not been demonstrated in increasing yieldof young oil palms under field conditions. The
present paper outlines the steps undertaken to
conduct the field trial.

The 15N isotope dilution technique has been
extensively used in the quantification of
biological N2 fixation (BNF) in a cropmicroorganism symbiotic system. An advantageof this technique is its accuracy indiscriminating N2 fixed from the atmosphere andN assimilated from the soil, employing the use
an appropriate non-N2 fixing plant as thereference crop (Danso, 1986; Barrie, 1991).
Quantification of BNF by diazotrophic bacteria(e.g. Azospirillum spp. and Bacillus spp.) in
association with non-leguminous crops is lesscommon. However, BNF involving nonlegumes
andassociative
N2
fixingmicroorganisms is becoming increasingly
imperative in attempts to develop a sustainableagricultural system. The BNF process isenvisaged to reduce dependency on N fertilizer.
Since a huge portion of crop production goes to
N fertilizer, a good BNF system can help reduce
production cost, especially in the tropics where
there is an dire need to increase food crop
production (Cocking, 2000).

Oil palms at nursery stage

Exploitation of the BNF process and plant
growth stimulation involving associative N2
fixing bacteria on oil palm (Elaeis guineensis)
can potentially make Malaysian palm oil
industry more sustainable and environmental
friendly (Dobereiner and Baldani, 1998;
Shamsuddin et al., 1999). The palm oil industry
plays a significant role in Malaysian economy
through its contribution of about 4.3% (RM 14.5
billion) of the total GDP in 2000 (RM 341
billion) (Department of Statistics Malaysia,

Typical oil palm plantation
3

Bunches of oil palm
fruits after harvest

Oil palm fruits on trees

FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Production of Bacillus sphaericus UPMB 10 inoculum

PRODUCTION LINE

GOPF
JITU®

Carrier Production
Mixing50kGyGamma-irradiation
Mintec-Sinagama,
MINT

Packing

dH2O

Ready
to
Use

Quality
Assurance

Gamma-irradiatedbags

Sterile conditions
Mother culture
Purity test

Incubation
28°C ± 2

Batch Culture

Inoculation

Process flow for the production of the PGPR

Ground oil palm frond

MixingPacking in polyethylene bag

Ready to gamma-irradiation
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
Field application of 15N dilution to young
oil palm trees

fixed by B. sphaericus UPMB 10 in association
with young oil palms under field situations.

The 15N labelled ammonium sulphate enrichedwith 10% atom excess (a.e.) equivalent to 10 kg
N ha-1 will be dissolved in distilled water. The
15
N solution (1000 mL) will be applied to eachisotopic-plot (one palm m-2) inoculated with
UPMB 10 and non-inoculated (reference plants)
which is equivalent to 1 g N plant-1 at planting.
a.

b.

＜References＞
Amir, H.G. 2001. Nitrogen fixation and plant
growth enhancement by beneficial rhizobacteria
in association with oil palm seedlings. Ph.D.
thesis. Universiti Putra Malaysia. 155 pp.

Labelling technique
Soil labelling will be carried byincorporating the 15N labelled (NH4)2SO4
into the soil at planting time by uniformly
spraying the fertilizer solution within 1m2
of the isotopic-plot. 15N is applied only
once during the study period of 1 year.

Barrie, A. 1991. New methodologies in stable
isotope analysis. In: Anon., Stable Isotope inPlant Nutrition, Soil Fertility and Environmental
Studies. IAEA Publication, Vienna, pp. 3-25.
Cocking, E.C. (2000). Helping plants get more
nitrogen from the air. European Review 8 (2):
193-200.

Plant sampling
Samples will be obtained by destructive
harvest of the oil palm trees one-year after
planting. Each palm will be sectioned into
leaves, stem and roots. A few oil palm
trees will be destructively sampled at startof experiment to determine initial N
content.

Danso, S.K.A. 1986. Review: Estimation of N2fixation by isotope dilution: an appraisal oftechnique involving 15N enrichment and theirapplication – comments. Soil. Biol. Biochem. 18:
234-244.
Department of Statistics Malaysia (2001).
Dobereiner, J. and Baldani, V.L.D. (1998).
Biological nitrogen fixation by endophytic
diazotrophs in non-legume crops in the tropics.
In Nitrogen fixation with Non-legumes, ed. K.A.
Malik, M.S. Mirza and J.K. Ladha, pp 3-7.
Kluwer Academic publisher, Great Britain.

Inoculation of UPMB 10 on the oil palms

Each N2-fixing isotopic-plot of young oil palm
will receive 3 kg of the PGPR inoculum. The
inoculum is thoroughly mixed with planting soil
and applied under and around the oil palm
seedling at planting. Subsequent application willbe spread uniformly within the weeded circle.
Each isotopic-plot will be inoculated withrespective inoculum treatments at planting andat four monthly intervals(3 kg
inoculum/isotopic-plot/application).
The
uninoculated control treatment plots will receive
inoculum carrier and sterilized (γ-irradiated, 50kGy) inoculum.

Shamsuddin, Z.H. and Roslina, A.W. (1995).
Isolation of Azospirillum from oil palm roots.
18th Microbiology Symposium, Kuching,
Sarawak.
Shamsuddin, Z.H., Amir, H.G., Roslina, A.W.,
Halimi, M.S., Nazalan, N., Ramlan, M.F. andMarziah (1999). Growth of oil palm seedlings
inoculated with N2 fixing Azospirillum andrhizobacteria. In Colloqium on advances in oilpalm research under IRPA-funded programmes
in 7th Malaysia plan. PORIM Kuala Lumpur.

Conclusion

It is envisaged that this isotopic dilution
technique could elucidate quantification of N2
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
COMMERCIAL POTENTIAL OF AMF APPLICATION
IN THE AGRICULTURE AND SOIL REHABILITATIONAND CONSERVATION SECTOR
P. Raja* and C.K. Tang
Malaysian Agri Hi-Tech Sdn. Bhd.
* raja2020@yahoo.com
What is AMF?

mycorrhizas, which included at the time only
ericaceous and orchid mycorrhiza. The namefor the arbuscular mycorrhizal symbiosis haschanged through the years.

Arbuscular mycorrhizal Fungi (AMF) is adistinct group of fungi that form obligate
symbiotic relationships with the roots offlowering plants. Today, mycologists and plantresearchers are typically well versed in thefunction of arbuscular mycorrhizas and theirconsequences for nutrient cycling and plant
productivity. AMF may have been described as
early as 1842 (Nageli 1842), but most of
Nageli’s drawings only remotely resemble thearbuscular mycorrhiza. Rayner (1926-1927) andTrappe and Berch (1985) cited other earlyobservations of the symbiosis during the period
1875-1895. Most surveys of host plants andsophisticated anatomical descriptions of what
are most certainly arbuscular mycorrhizas are
given by Schlicht (1889), Dangeard (1896), Petri(1903), Jones (1924) and Lohman (1927).
Schlicht (1889) had already observed the basic
anatomical relationships between host andfungal tissues.
Janse (1897) called theinteramatrical spores “vesicles” and determinedthat other structures, named “arbuscules” by
Gallaud (1905), were located in the inner cortex.
Gallaud (1905) made very accurate observationsof the arbuscule is enirely surrounded by a hostmembrane, which was later confirmed by Cox
and Sanders (1974) using Transmission
Electrom Microscopy (TEM).
Frank (1885) gave the name “mycorrhiza” tothe peculiar association between plant roots andectomycorrhizal fungi. A thorough discussionof the derivation of the word “mycorrhiza”,
including the incorporation of the secondr isgiven by Kelly (1931, 1950). In anotherpublication, Frank (1887) recognized adistinction between ectotrophic and endotrophic

AMF benefits
The mycorrhiza colonized plant is better
nourished and better adapted to its environment.
It obtains increased protection against
environmental stresses (Sylvia and Williams,
1992), including drought (Subramanian et al1995) cold (Charest et al 1993) Paradis et al1995), Salinity (Davis and Young, 1985) andpollution (Leyval et al 1994; Shetty et al 1995).
In addition, symbiosis tends to reduce theincidence of root diseases and minimizes theharmful effect of certain pathogenic agents(Dehne, 1982). By and large, the growth andhealth of colonized plants is improved. At thesame time they obtain increased protection
against environmental conditions detrimental totheir survival.
AMF biotechnology in agriculture can be
gradually
integrated
with
conventional
agricultural practices. Over the medium term, it
is quite possible to restore soil quality andimprove soil sustainability (Betlenfalvay and
Linderman. 1992). It has been shown thatappropriate management of mycorrhizae in
agriculture allows a substantial reduction in theuse of chemicals, thus lessening the level ofpollution of surface waters, reducing farm laborand costs of production, while maintaining
yields at their highest levels. The advantages andbenefits of adopting mycorrhizae in agriculture,
allows us to better visualize the scope of thisphenomenon at the crop level and in turn, theimpact of its long term adoption on the qualityof life.
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

General advantages and benefits of adopting AMF in agriculture system
Field
Agriculture

Advantages
Agrosystem stability
Plant production

Plant physiology

Plant morphology

Benefits
Improved plant roots, soil
microflora and the abiotic
geochemical soil matrix.
Increase shoot and root
biomass

Stress resistanceDrought, cold andpollution

Resistance to pathogens

Crop protection, reduced
use of pesticides,
fungicides, improvement
in animal, plant, humanhealth and product quality

Value added to productIncreased synthesis ofprimary and secondary
metabolites
Reduced chemicalfertilizers (10-30%)

Improvement in nutrition
Carbohydrate level

Improved photosynthetic
activity

Tolerance to water stress

Cultivation of arid soils orsoils unfit for agriculture
and altered hormonalbalance

Resistance to lowtemperatures
Transformation of rootsystem and rootarchitecture

Diversity of crops ininhospitable areas
Adaptation to stress,
increased resistance toerosion, fixation of soils,
increased numerous andbranched roots, root apices
and improved production
with certain root crops

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Plant community

Microorganism diversity
in the subsoil

Reestablishment of soil
microflora andimprovement of soil
quality

Habitat restoration

Stabilize thebiogeochemical cycling

Improvement in yields,
better acclimatization attransplanting and diversityof plant cover
Micropropagation plants
High value plants
Uniform size, better
growth, shorten nursery
periodWhen these filaments come in contact with ayoung root, they thread their way between the
The benefits of AM fungi are not only limited tocortical cells and quickly propagate, forming
improved uptake of nutrients and water from soil.
intracellular arbuscules and in some cases,
In recent years, it has been shown that theseintercellular vesicles.
These arbuscular fungi
fungi interact with other soil microorganisms
are so called because of the tree-like structureswith same special positive attitude. For example,
that are differentiated in the roots. Spores are
it has been shown that the conversions ofalso differentiated in the soil and roots. They act
atmospheric nitrogen from a gas into a form ofas reserve, propagation organs and as a reference
fertilizer nitrogen (biological nitrogen fixation)
structure for species identification. Thus far weby soil bacteria can be greatly enhanced by thehave no serious clues to the sexuality of AMF.
presence of AMF (Camel et al 1991; Finlay and
Their association with the Zygomycetes is based
Sonderstrom, 1992; Jakobsen and Rosendahl,
on the similarity of their spores to the spores of1990). The myriad of interactions that can occur
other known representatives of this class.
in the soil around roots with AMF is unlimited,
The close association created between the plantbut recent results indicate that the microbial
and fungus through the hyphal network allows
associations with AMF play significant roles inthe exchange of nutrients for the survival andthe benefits observed by inoculation with AMF
growth of the two partners. First, the wide
fungi. Establishing the mycorrhiza “team” ondispersal of the fungus in the soil through its
plant roots early in early stages of growth cycle
large filament network gives the fungus access tois clearly the right approach for exploiting thesea much larger volume of soil and higher area forremarkable fungi in agriculture. Inoculation withobsorption than the root system itself. The
mycorrhizal fungi and their microbial associates
fungal hyphal act more or less as a pump,
can maximize crop yields and quality while
supplying the root with a supplement of water
minimizing applications of chemical fertilizers
and mineral salts to which it normally would not
and pesticides that can be harmful to people andhave access.
In return, the fungus receives
the environment.
metabolized nutrients from the plant that is
unable to synthesize by itself, such as sugars,
Biological and biochemicalamino acids and secondary metabolites.
mechanism of AMF
The following evidence obtained from research
to date as biological control by AMF
Arbuscular mycorrhizal fungi establish a networkof microscopic hyphal filaments in the soil.
Survival of partners

8

FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer ProjectAs a matter of fact colonized plants are better
able to obtain their nourishment in the soil andresist environmental stresses gives fungal
symbionts a biofertilizing and crop protection
role. In agriculture, the increased uptake of soil
minerals by colonized plants means that it ispossible to consider substantially reducing
applications of fertilizers and pesticides and at
the same time obtain equivalent or even higher
crop yields (Abbott and Robson, 1991). Throughappropriate management of mycorrhizae in
agriculture, it is also possible to maintain soil
quality and sustainability while protecting theenvironment over the long term and reducing
costs of production.

1. Colonization by AMF may offer protection
against soil-borne pathogens (Davis andMenge, 1980; Dehne, 1982; Krishna andBagyaraj, 1983).
2. Infection of roots by nematodes may be
reduced when they are colonised by AMF( Sitaramaiah and Sikora 1982; Cooper andGrandisum 1987)
3. Increased nurient concentrations in the leavesof AMF colonised plants may lead toenhanced development but not increased
incidence of foliar pathogens (Graham andMenge 1982).
Agricultural and land reclamation
potential
Research work on the potential value of
Arbusculr mycorrhizal fungi in agriculture andland reclamation followed from the findings in
the 1950s,1960s and 1970s that they could
substantially increase P uptake and plant growthunder certain circumstances.
However, theincreasing number of observations that suchfungi already exist in most agricultural soils led
some to conclude that there would be little value
in inoculation (Menge 1985). Khan (1972) may
have been among the first to demonstrate thatsuch a practice could be beneficial in somecircumstances, but it would frequently prove to
be uneconomic because of the large cost of
inoculum production relative to the cost ofphosphate fertilizer (Menge 1985) However, thepracticality of inoculation soils that are
inherently low in inoculumpotential such as
sterile citrus nursery beds, sterile potting media
or soils that are highly disturbed may be greater.
For example, the revegetation of disturbed lands,
and the course of plant succession in suchenvironments may be strongly influenced byinoculation with mycorrhizal fungi Much of thepertient literature on use of arbuscular
mycorrhizal fungi in land reclamation was
summarized in a publication edited by Williams
and Allen (1984).

Future perspectives
Future needs are of two major kinds. The
mechanism by which AMF influence root systemneeds to be understood at a basic level so thatroot system design and modification can be used
as a management tool, especially within intensive
horticultural systems. In addition the use of
AMF within sustainable systems needs to bebetter understood in relation to nutrient cycling.
It is important that AMF should not be seen as
the single and final solution for the control ofplant pathogens, chemical fertilizers, pesticides
and insecticides but as part of an integrated
approach. In ‘ open systems’ management islikely to be the way forward, with practices
shaped by research aimed at a greater
understanding of the role of AMF in plant-soil
systems.
However, in ‘closed systems’
inoculation is likely to be the way forward inmany cases.
For many years difficulties
associated with inoculum production have
prevented large-scale use. However, inoculum is
now available commercially throughout Western
Europe, Japan, USA, Malaysia and SouthAmerica. It is certain that only by adopting theseapproaches is it likely that the benefits of AMF
to agricultural production will be optimized,
leading to a reduction in chemical inputs andincreased sustainability.
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
microorganisms may also be involved in such
positive interactions. Mycorrhizas are commonly
defined as plant-fungus associations; infunctional terms however they may better be
regarded as a value regulating plant-soil
interactions, possibly increasing carbon sink toroots and carbon efflux to soil (Garbaye, 1991).
Their management in consideration of other soil
microorganisms is a challenge and an
opportunity for sustainable plant-soil systems(Staley et al 1992).
Further research is necessary, both at a basic andapplied level, i.e., regarding the maintenance of
inoculated organisms in the field. It is worth
noting, however, that the number of field
experiments is increasing every year. It istherefore to be hoped that, not withstanding thedifficulties, and even some failures andcontradictions, this body of experience will soon
allow a wider utilization of mycorrhizas andassociated organisms in plant generations.

Malaysian government has promoted Malaysia to
be a garden country.
The importance oflandscape is apparent in the recent and fastdevelopment of Malaysian highway projects.
However, the development of these areas haveoften been characterised by poor establishment ofplants due to the disturbed and compacted soils
found along highways.
In this study theapplication of commercially available AMF
inoculum (MycoGoldTM) and organic matter(MycoOrganicTM) in Malaysian high way
landscape plants are UPM Interchange (PLUS)
and Sepang Toll Plaza (ELITE) to improve root
growth.
In UPM Interchange the soil pH
ranged from 4.6-6.5 and bulk density is 5.2g/ cm3. Total of 7 trees, one shrub and 7 cover crops
were treated with MycoGoldTM
andMycoOrganicTM. All plant roots had colonizedwith AM fungi.
The highest per cent infection(70 %) and spore count (227/10g soil) wererecorded in Acacia mangium roots andrhizosphere soils. In Sepang Toll Plaza soil pH
ranged between 5.2-6.8 and bulk density was
about 4.8 g/ cm-3. Mycorrhizal infection wasnoted on all treated plants. Out of 6 plants thehighest percentage of infection (35%) and spore
count (20/10g soil) were recorded in Duranta
repen ‘gold’. Twelve AM fungal spores wereisolated from both rhizosphere soil samples. Outof 12, nine species from Commercial product and3 from native spores. After application ofcommercial AMF inoculum and organic matterin landscape plants, we observed the fast
recovery of cover crops and trees are as
evidenced by the high production of new leaflets.
These preliminary results are indicates that AMF
inoculation can be profitably exploited for
Malaysianhighwaylandscapeplantdevelopment. (Raja et al 2001).

Acknowledgement
The Authors would like to express ourappreciated to the Staffs, Management andShareholders of Malaysian Agri Hi-Tech Sdn.
Bhd. for their continuous supports in the
promotion of AMF towards Sustainable
Agriculture.

＜References＞
Abbott, L.K. and A.D. Robson. (1991). The
effect of root density, inoculum
placement and infectivity of inoculum on
the development of vesicular-arbuscular
mycorrhizas. New Phytologist. 97: 285299.
Betlenfalvay, G.J. and R.G. Linderman. 1992.
Mycorrhizae in Sustainable Agricultrue,
American Society of Agronomy Specail
publication 54, American Society ofAgronomy, Madison.
Camel, S.B. M.G.Reyes-Solis., R. Ferrera-Cerrat,
R.L. Franson, M.S. Brown, and G.J.
Bethlenfalvay. (1991).
Growth ofConclusions
Beneficial effects of mycorrhizas including plant
growth are well established in controlled
conditions and field trials.
Nevertheless,although frequently reported as mycorrhizal
effects, it should not be excluded that other10

FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
Graham, J.H. and J.A. Menge. 1982. Influenceof vesicular-arbuscular mycorrhizal andsoil phosphorous on tale-all disease ofwheat. Phytopathology. 72: 95-98.
Jackobsen, I. and L. Rosendahl. (1990). Carbon
flow into soil and external hyphae fromroots of mycorrhizal cucumber plants.
New Phytologists 115: 77-83.
Janse J.M. 1897. Les endophytes radicaux dequelques plantes Javanaises. Ann Jardin
Bot Buitenzorg. 14: 53-201.
Jones, F.R. 1924. A mycorrhizal fungus in theroots of legumes and some other plants.
Journal of Agriculture Research 29: 5460.
Kelley, A.P. 1931. The concept of mycorrhiza.
Mycorlogia. 23: 147-151.
Kelley, A.P. 1950.
Mycotrophy in plants.
Chronica Botanica, Waltham, Mass.
Khan, A.G. 1972. The effect of vesiculararbuscular mycorrhizal associations on
growth of cereals. I. Effects on maize
growth. New Phytologist. 71: 613-619.
Krishna, K.R. and D.J. Bagyaraj, 1983.
Interaction between Glomus fasciculatumand Sclerutium rolfissi in peanut.
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Leyval, C., I.I. Weissenhorn, A. Glashoff and J.
Berthelin. (1994). Influence of heavy
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Lohman, M.L. 1927. Occurrence of mycorrhiza
in Iowa forest plants. University of Iowa
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Menge, J.A. 1985. Developing widescale VA
mycorrhizal inoculations: is it practical ornecessary?
In:
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ForestResearch Laboratory, Oregon State
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Nageli C., 1842. Pilze im Innern von Zellen.
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Petri, L. 1903.
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vesicular-arbuscular
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Charest, C. Y. Dalpe and A. Brown. 1993. The
effectofvesicular-arbuscular
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Cooper, K.M. and G.S. Grandison. 1987. Effects
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Cox, G.G. and F.E.T. Sandrs. 1974.
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Davis, E.A. and J. L. Young.
(1985).
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Canadian Journal of Botany. 63: 11991203.
Davis, R.M. and J.A. Menge. 1980. Influence ofGlomusfasciculatus
andsoil
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Dehne, H.W. (1982).
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Garbaye J. (1991). Biological interaction in the
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and birdsfoot trefoil growth andnodulation. Biology Fertility Soils. 14:
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di Janse) nelle micorrize endotrrofiche.
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Raja, P., J. Herdhawati, C.A. Nazam1 and I.
Zubaidah. 2001. Rehabilitation of
Malaysian highway landscape plants
using commercial arbuscular mycorrhizal
fungi. ICOM3. Diversity and Integration
in Mycorrhizas. Adelaide, Australia
Rayner, M.C. 1926-1927. Mycorrhiza. New
Phytologist. 25: 1-50.
Schlichet A. 1889. Beitrag zur Kenntniss der
Verbreitung
undBedeutungderMycorhizen.
Landwirtschaftiche
Jahrbucher. 18: 478-506.
Shetty, K.G., B.A.D. Herick and A.P. Schwab.
(1995). Effects of mycorrhizae fertilizer
amendments on zinc tolerance of plants.
Environmental Pollution. 88: 307-314.
Sitaramaiah, K. and R.A. Sikora. 1982. Effectof the mycorrhizal fungus Glomus
fasciculatus on the host parasite
relationship of Rotylenchus reniformis intomato. Nematology. 28: 412-419.
Staley, T.E., E.G. Lawrence and E.L. Nance.
(1992).
Influence of a plant growthpromoting pseudomonad and vesiculararbuscular mycorrhizal fungus on alfalfa

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

A Few Photographs

Conducting 15N-aided biofertilizer field trials on rice at MADA, Kedah.

Preparation of biofertilizer products for pot trials, MINT.

Analysis of N-15 using the emission spectrometer at MINT

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
N-FIXING MICROBES
Pauline Liew Woan Ying
Malaysian Institute for Nuclear Technology Research (MINT)
pauline_liew@mint.gov.my

In order to achieve the goal of using indigenous microbes for local applications, research works to look
into the availability of N-fixing microbes in local soil and water resources has been carried out. The
samples include soil samples obtained from the paddy planting plots, compost samples, water samples
etc. Potential N-fixers were isolated based on their abilities to survive and multiply under nitrogendeficient condition.
The microbes were isolated and characterized based on microbiological and molecular approaches.
Among the positive isolates were shown in figure below. Most of the isolates were identified to bemembers of the genus Bacillus. It was therefore hypothesized that Bacillus could be one of the major
contributing groups forming the N-fixing reservoir in the local soil and water resources.

Diagram shows phylogeny of representative nitrogen-fixing isolates. MBCC represents MINT Bacteria Culture
Collection. Isolates were identified based on 16S rRNA gene sequences. Data were presented at the 14th International
Congress on Nitrogen Fixation, Beijing, China (Liew et al., 2004).

Reference
Liew, Woan Ying Pauline, Jong, Bor Chyan and Abdul Rahim, Khairuddin (2004) A Study on
Microbial Nitrogen Fixation in Malaysia. In Proc. 14th International Congress on Nitrogen Fixation,
Beijing, China, 27 Oct – 1 Nov 2004.
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Project Leader of Malaysia

Dr. Khairuddin Bin Abdul Rahim
Senior Research Officer
Manager, Bioindustry Group
Division of Agrotechnology & Biosciences
Malaysian Institute for Nuclear Technology
Research (MINT)BANGI, 43000 KAJANG, SELANGOR, MALAYSIA
Tel. + 60-3-89250510, Fax. + 60-3-89282956
E.Mail: khairuddin@mint.gov.my

Research Interest••••

Biofertilizer: arbuscular mycorrhizal fungi; symbiotic N2-fixing microorganisms; plant growthpromoting rhizomorphs.
Bioprocessing: microbiology and microbial processing of Aquilaria oleoresin.
Soil bioremediation: oil sludge; ex-mining land.
Use of isotopic tracers (e.g. 15N, 32P) and other nuclear technology in soil fertility and plant
nutrition studies (sago, oil palm, Arachis hypogaea, Acacia, Hevea, Coffea, Theobroma cacao,
Lansium, leguminous cover crops).

Present Responsibilities
1. Project Leader, Malaysia, for Forum for Nuclear Application in Asia (FNCA), Biofertilizer
Research.
2. Project Leader, Microbiology and Microbial Processing of Aquilaria Oleoresin.
3. Project Leader, Bioremediation of soils contaminated with potentially toxic elements using
effective microorganisms and plants (IRPA EAR 08-01-03-0005-EA0001)
4. Project Leader, Production of Biofertilizer Microorganisms and Organic Fertilizer of Agro ByProducts.
5. Project Leader, Nitrogen Management in Rice (MoU MINT-MADA).
6. Team Member, Groundwater Microbiology of Malaysian Municipal and Industrial Landfill Sites,
under Project on Landfill and Modelling of Pollutant Transport in Groundwater from Municipal
and Industrial Disposal Sites (IRPA 08-01-03-0007 EA 0001).
7. Team member:, Studies on Ethnobotany of the Flora of Hutan Belum..

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project~Information~

FNCA Web Site: http://www.fnca.jp/english/＊＊

FNCA Biofertilizer News Letter No. 5
Editor:Khairuddin Bin Abdul Rahim
Project Leader of FNCA Biofertilizer of Malaysia

Secretariat:

Yuko Wada
Asia Cooperation Center (ACC)Japan Atomic Industrial Forum, inc. (JAIF)
Tel:+81-3-5777-0753 Fax:+81-3-5777-0757 E-mail: yuko@jaif.or.jp* This Biofertilizer Newsletter is issued by Japan Atomic Industrial Forum, Inc. (JAIF)
under the contract of the Ministry of Education, Culture, Sport, Science and Technology (MEXT)

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