FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Issue No. 7
February 2007

Message from the Philippines
Dear Readers,This 7th issue of the FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Newsletter features the activities pertaining to research and development, industry, use and promotion and impact
of Biofertilizers in Philippine agriculture. It's my pleasure to share to you information herein thatyou may find interesting and valuable in line with the same activities in your respective areas.
The need to address the food requirement of the growing population in the Philippines made thefarmers adopt the latest technology in agriculture like the use of high yielding and resistant cropvarieties. These varieties however require high amount of nutrients making the farmers dependent
on agricultural input like inorganic fertilizers. It was reported that from 1966 to 1998 fertilizer
application in the Philippines increased from 20% to 80%. Nitrogen, considered as the most limiting factor in crop production, gains the highest mark of all the major nutrients from inorganic
sources applied by farmers. The 2001 FAO (Food and Agriculture Organization) Yearbook on fertilizers stated that from 1987 to 1998, Philippine consumption of nitrogenous fertilizers increased
from 371,487 MT to 546,499 MT. Such consumption resulted to high production for some time but
adverse soil and other environmental conditions were observed in the long run. This called for theuse of other alternative inputs. The use of biofertilizers has been determined as one of the main
options to address the rising concern on agricultural and environmental sustainability.
Biofertilizer industry in the Philippines grown considerably with the promotion of Government of
bioorganic farming and the active involvement of research institutions, private individuals andenterprises. Biofertilizers in various forms have been developed and more researches on rhizobia
legume symbiosis, azolla, mychorriza, sesbania, blue green algae and the living microorganism (Nfixing bacteria) were encouraged.
My sincere thanks to the contributors who graciously contributed some highlights of their researchworks and efforts. Without their contributions, this issue would not have materialized.
Best regards,
Richard M. Balog
FNCA Biofertilizer Project Leader, Philippines

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

FNCA (Forum for Nuclear Cooperation in Asia)
2005 Biofertilizer Workshop
The 2005 FNCA Biofertilizer Workshop washeld last 9-13 January 2006 at the PNRI(Philippine Nuclear Research Institute), Quezon
City, Manila jointly hosted by the DOST
(Department of Science and Technology) Philippines and MEXT (Ministry of Education,
Culture, Sport, Science and Technology) Japan. It was attended by 17 participants, primarily country project leaders from Indonesia,
Japan, Korea, Malaysia, Philippines, Thailand
and Viet Nam. Local participants included
PNRI collaborators from Biotech (National
Institute of Molecular Biology andBiotechnology) - UPLB (University ofPhilippines Los Banos) and DA (Department ofAgriculture) - BSWM (Bureau of Soils andWater Management). The workshop was highlighted by the presence and messages of key
persons of the host organizations in person of
DOST Secretary Dr. Estrella Alabastro, PNRI
Director Dr. Alumanda Dela Rosa and Director
for International Nuclear Cooperation of
MEXT Ms. Takiko Sano. The workshop activities comprise of presentation of country reportsby the respective project leaders, roundtablediscussions of project accomplishments/activities and plans, special lectures by invited lecturers and technical visits to biofertilizer facilities.

Visit to the Bio-N production facility
at Biotech-UPLB. Supervising thefacility were Dr. Mercedes U.
Garcia (standing 4th from right)
and her staff Ms. Juliet Anarna(standing center in front). Inset:
inside the facility.

Dr. Mercedes U. Garcia of the National
Institute of Biotechnology-UPLB lectured onthe "Role of Bio-N in Ginintuang Masaganang
Ani Program for corn in the Philippines". Dr.
Gina P. Nilo of DA-BSWM lectured on
Organic-Based Agriculture "Agri-Kalikasan"
Program/Tipid-Abono Program in thePhilippines.
The participants visited Biotech-UPLB and
Bio-N Mixing Plant at Barangay Lawy, Capas,
Tarlac. The participants were given overview
about the Biotech particularly pertaining to its
developed biofertilizers under the program on
agriculture and forestry. The participants also
visited the Bio-N production facility of theinstitute where they were informed about theprocesses of the Bio-N inoculant production.
The facility, supervised by the inventor of BioN Dr. Mercedes U. Garcia and her staff Juliet
A. Anarna, was established through the funds
provided by the DA in 2003 for the development and application of Bio-N on high value
crops.
The establishment of mixing plants is one of
the strategies in the promotion of biofertilizer
use by farmers which was a big help for sus-

The workshop participants with Dr. Alumanda Dela Rosa
(sitting 2nd from left), Dr. Estrella Alabastro (sitting 3rd
from left) and Ms. Takiko Sano (sitting 4th from left).

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
tainable crop production in the locality. The
mixing plant visited was operated by a farmer's
cooperative St. Philip PMPC (Primary MultiPurpose Cooperative) Inc. in collaboration with
the Biotech-UPLB. The Bio-N mixing plant
was also established in cooperation with the
DA-RFU (Regional Field Unit) 3-Regional
Soils Laboratory, Tarlac LGU and PDA-Tarlac.
Mr. Tolentino, chairman of the St. Philip PMPC talked tothe participants about the cooperative and the mixing
plant. (Inset: the participants in discussion about how themixing plant operates).

Biofertilizer studies conducted by PNRI under FNCA framework
The biofertilizer used in the study conductedby the PNRI under the FNCA framework was
Bio-N, developed by Dr. Mercedes U. Garciaof the Biotech-UPLB. Though various forms of
biofertilizer have been developed in the country, Bio-N got the nod from the government as
the most promising technology in crop production particularly in reducing the use of chemical fertilizers. The government, in line with its
program on sustainable agriculture allocatedfunds for the mass production of Bio-N as thiswas foreseen to greatly benefit the farmers.
Bio-N is an inoculant in solid powder form
that contains two species of the nitrogen fixing
bacteria Azospirillum isolated from he roots ofRoot development of rice as affected by
Bio-N inoculation

a local grass talahib (Saccharum spontaneum
L.) which have been screened for their effectiveness to variety of agricultural crops. It can
fix atmospheric nitrogen and transform it into
form usable by plants or crops. Beneficial
effects of Bio-N includes enhancement ofshoot growth and root development, makes
plant resistant to drought and pest attack,reduces incidence of rice tungro and corn earworm infestation and increases yield andmilling recovery of rice and corn. It wasobserved to provide approximately 30%-50%of the nitrogen required by crops.

Bio-N Biofertilizer

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
field or on prepared beds. For transplanted
rice, Bio-N is mixed with water to form slurry.
Roots of seedlings are pruned into uniform
length and dipped into the slurry for at least 30
minutes to an hour before transplanting.

Method of Bio-N application on crops

Field experiment and yield trials
Since the development of Bio-N in 1985, fielddemonstration trials have been conducted totest its efficacy to various crops.
The Biotech, as the primary institute for thedevelopment of Bio-N conducted and supervised several field trials and supervised farmer
managed field trials for assessment on effect of
Bio-N to crops. In the study conducted on theeffect of Bio-N and inorganic fertilizer on theyield of corn (Super sweet var.) in aBio-N
application has higher yield than application of
inorganic fertilizer and was comparable to theyield when application of inorganic fertilizer
was cut into half plus Bio-N. This further
showed that the use of Bio-N has the least
value of input (in peso) per yield (kg) whichindicates that farmers can save big amount of
fertilizer expenses through the use of Bio-N.

Bio-N application on corn as seed coating

Bio-N application on rice as root dip

Bio-N is generally applied as seed coating forcorn or as suspension for use in watering seed
beds or root dips for rice. The use of 5-6 packets (200g per packet) of Bio-N per hectare isrecommended to inoculate rice or corn.
For corn, seeds are moistened with waterand mixed with sufficient amount of inoculants
until seeds are evenly coated. A sticker could
be use to enhance adsorption of Bio-N to theseeds. Coated seeds are sown immediately andexposure of the inoculated seeds from direct
sunlight should be avoided.

The study conducted by the PNRI on theuse of nuclear techniques in the assessment of
Bio-N fertilizer as seed inoculant for corn on
low fertility area in the province of Isabela(around 400km north of Manila) showed that
corn yield was only 208 kg/ha (kilogram perhectare) when not inoculated with Bio-N andwithout fertilizer application. Corn yield wasincreased tremendously to 2,880kg/ha when

For direct-seeded rice, germinated seeds(with embryonic roots sprouted) are coatedwith the inoculant and sown directly to theTable 1. Effect of Bio-N and inorganic fertilizer on the yield of corn (Super sweet var.) in farmers field demo, dry season 2001.

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
inoculated with Bio-N plus application of P
and K fertilizer. This resulted to an increase inincome from PhP1,456.00 to PhP14,341.50.
Bio-N inoculation plus application of half andfull recommended rate of N fertilizer yielded
3,460kg/ha and 2,960kg/ha, resulting inincrease in income of PhP16,230 and
PhP10,558 respectively. The experiment also
used N-15 to quantify biological nitrogen fixing capacity of Bio-N. N-15 data indicated that
19-24% of the nitrogen taken by the plants isnitrogen from the atmosphere that is fixed by
the Bio-N. (Please also see related article by
Biotech on biofertilizers)
Field experiment on corn (with yield levels) in low fertility
area at Isabela province.

Promotion of biofertilizer use by farmers
continuous endeavor of promoting the use of
biofertilizers. Such means includes conduct of
exhibitions and trade fair, consultations, symposia, trainings, field demonstrations, and tapping broadcast and print media for information
dissemination.
One of the recent and remarkable efforts by
the government in promoting the use of thebiofertilizer Bio-N was the establishments ofTransfers of technology on use of Biofertilizers
in line with various initiatives and efforts inpromoting organic agriculture still remains anenormous challenge in Philippine agriculture.
Despite of the numerous discoveries and technologies on biofertilizer, bringing it to the end
users particularly the farmers is a difficult task.
Convincing farmers is costly as it entails
unlearning and getting rid of their old habits ofusing chemical fertilizers. Different means of
has been undertaken in the Philippines in its

A pose with the farmers during"Farmers Field Day" in
Isabela province. (Inset:
explaining to farmers about theexperiment.)

One of the Bo-N mixing plants
established in province of
Cotabato. (inset:: Giving technical assistance to the recipient)

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Projectsame time served as distribution channels thatmade Bio-N accessible to the local consumers
especially the farmers who were sourcing itfrom UPLB. There have been 32 mixing plants
strategically established throughout the country
that produces and sold Bio-N at price affordable to farmers. (Please also see related article
by Biotech on promotion and technology transfer of Biotech biofertilizers.)

mixing plants. The establishments of mixing
plants is one of the projects being undertakenby the DA Ginintuang Masaganang Ani Corn
Program, in collaboration with the BiotechUPLB, LGU's (Local Government Units) andfarmers organization/cooperatives in respond to
the farmers increasing demand of the Bio-N inlate 1996. The mixing plants are consideredsatellite production and sales centers and at the

Application of gamma irradiation for sterilization of biofertilizer carriers
tion process poses some disadvantages such as
incomplete sterilization, may result to somechanges in physical and chemical properties ofcarriers, costly and suitable only to small scale
treatments. Alternative means of sterilization
process has been found in the utilization of thekilling effect of gamma rays to microorganism.
This process has been undergone in thePhilippines in line with its activities in
improvement of the quality of biofertilizers.
The PNRI operates a multi purpose Co-60
gamma irradiation facility, which is the only
one in the country. The application of gamma
irradiation for sterilization of biofertilizer carrier however is still in its experimental stage.
Bio-N for instance, whose carrier is composeof soil and charcoal, gamma irradiation showeddecrease of microbial load at increasing doseand is sterilized at the dose 30kGy. Different
biofertilizers however have different carriers so
further experimentation is needed and sufficient data should be acquired especially on theadvantages of the gamma irradiation compared
to conventional methods to convince thebiofertilizer producers to adopt the applicationof gamma irradiation for sterilization of biofertilizer carriers.

Biofertilizer primarily contains microbial inoculants usually proliferated in a nutrient source
medium called as carriers. In the production of
biofertilizers, carriers undergo a process ofsterilization to create a favorable environment
for the growth of desired inoculants free fromantagonistic microbes. Conventional method ofsterilization of carriers for biofertilizers is done
through the use heat-autoclave. This steriliza-

Co-60 irradiation facility at
PNRI (inset: the gamma
source)6

FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Researches on BIOFERTILIZERS in the Bureau of Soils & Water Management,
Deapartment of Agriculture, Dilliman, Quezon City, Phillippines
ENHANCEMENT OF BIOLOGICAL NITROGEN FIXATION IN LEGUMESTO INCREASE PRODUCTIVITY OF ACID UPLAND SOILS
M. J. Palis, A. O. Yambot and J. S. Rojalesof legumes and to determine the increase inproductivity with the application of these
inputs.

Rationale:
Marginal soils occupy a large area in thePhilippines. The acid upland soil is estimatedto be 17 million hectares or roughly equivalent
to 58 % of the total land area of the country(Atienza, 1991). These soils are generally characterized as highly weathered soils with low
productivity (IBSRAM, 1987).

Methodology:
The experiment was established in Kapatalan,Siniloan, Laguna, around 106 kilometers southof Manila. The soil in the experimental area
was taxonomically classified as Fine
Isohypothermic, Typic, Kanhapludult (Evangelista, pers.com) with a pH of 4.4 and12.6 ppm available P. Peanut (cv. UPL-Pn2)
and mungbean (cv. MG-50 L Yellow) wereused as test crops.

Despite the widespread belief that acidupland soils cannot support intensive and sustained agriculture, there are ample evidence
that they can be continuously cultivated andintensely managed for growing annual andperennial crops. To sustain crop production inthese soils, inclusion of legumes in the production systems had been advocated. Theselegumes can serve as nitrogen sources and theiruse by farmers can reduce the need for nitrogen
fertilizers. However, growth and nitrogen fixation of legumes in acid soils are constrained bya number of soil factors which include toxicities of Al, Mn and other metal ions (Zn, Cu,
Cd), low soil pH, deficiencies of Ca, Mg, P,
Mo and other elements including N and K. To
successfully grow legumes in acid soils
requires soil management strategies such as
choosing legume species that show tolerance tosoil acidity, liming and addition of fertilizers
and organic matter.

The study had the following treatments:
a. Lime application- with lime (5 tons CaCO3/ha)- without limeb. Micronutrient application- with micronutrient (50 kg fused traceelements/ha)- without micronutrientc. Phosphorus application (kg/ha)- 0,40,80,120,160
A split-plot design was used with liming as
the main plot, micronutrient application as the
sub-plot and phosphorus levels as the sub-subplot. The treatments were replicated threetimes. Nitrogen and potassium fertilizers wereapplied at the rate of 20 kg N/ha and 30 kg
K20 /ha. Rhizobium inoculants for peanut andmungbean were inoculated onto the seed prior
to planting.

This study aims to determine the effects oflime, phosphorus and micronutrient applications on the growth, nitrogen fixation and yield
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Salient Findings:
1. The application of lime at the rate of 5 tons
CaCO3 /ha gave a marked increase on thedry matter, nitrogenase activity, nodule
weight, and pod and seed weight of both
peanut and mungbean as shown in Figures 36.
Figure 1. General view of the experiment.

Figure 2. Researchers preparing samples for acetylenereduction assay.

The plants were grown to maturity. At nearblooming stage (7 weeks after planting), 10
representative plants from the sampling area
were obtained for determination of dry matteryield, acetylene-reduction activity and nodule
weight. The general view of the experiment at
maximum vegetative stage is shown in Figure
1. Preparation of samples for acetylene-reduction assay is shown in Figure 2.
Harvesting of the plants was done at physiological maturity. All pods from the plants in theharvest area were obtained. The pods were sundried, shelled and the weight of seeds was
taken.
A simple cost and return analysis was alsodone after harvest to determine whether growing legumes in acid soils is profitable for themarginal farmers.

Figures 3 and 4. Effect of liming on the dry matter andacetylene-reduction activity of peanut and mungbean
grown in an Ultisol.

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Figures 7 and 8. Effect of rate of phosphorus fertilizationon the growth, nitrogen fixation and yield of peanut andmungbeans grown in an Inceptisol.

3. Based on simple cost and return analysis,
growing of mungbean in acid soils could not
be recommended to farmers. A net incomeof P21,563.00/ha (USD 431.26) can beobtained with planting of peanut in acid soils
with the application of 5 tons lime, micronutrient, and 40 kg of P2O5 per hectare.
4. Peanut seemed to be more tolerant to soil
acidity when compared to mungbean.

Figures 5 and 6. Effect of liming on the nodule weightand pod and seed yield of peanut and mungbean grownin an Ultisol.

<References>
1. Atienza, R.N. 1991. Research Status of Acid
Soil Management and Utilization in thePhilippines. In. Management and Utilizationof Acid Soils. PCARRD. Los Banos,
Laguna, Philippines.
2. IBSRAM (International Board for Soil
Research and Management). Management ofacid tropical soils for sustainable agriculture.Bangkok.
3. Evangelista, P.P. . Bureau of Soils & Water
Management, Diliman, Quezon City,
Philippines (pers.com.)

2. Phosphorus application likewise increased
the growth, nitrogen fixation and yield of
peanut, but the increase was not as pronounced as when compared to the effect oflime (Figures 7 and 8).

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

SELECTION OF COMPETITIVE STRAINS OF RHIZOBIAFOR MUNGBEANS IN MARGINAL SOILS
M. J. Palis, A. O. Yambot, J. S. Rojales, V. F. Naboa, S. Arai,
H. Ito and M. Nakamura
This method is very sensitive and reliable indetecting and quantifying the inoculated strainsin the nodules of plants.
The intrinsic antibiotic resistance of four
rhizobial strain (MCO, CY 44, TAL 441, TAL209) were developed by exposure of the rhizobial strains to different concentrations of theantibiotics like streptomycin, kanamycin,
erythromycin and spectinomycin. Retention orloss of the symbiotic effectiveness of the four(4) antibiotic-resistant rhizobial strains was
assayed by using the plant-infection test.

Introduction:
A commonly used technique to improve
legume growth and seed yield is inoculation
with useful Rhizobium strain with a highpotential for nitrogen-fixation. These strains
however often fail to nodulate because they areunable to overcome environmental stresses.
The inoculant strains also have to compete with
the indigenous rhizobia which are often unable
to fix N in symbiosis. In order to compete
effectively with indigenous strains, inocula
have to be applied in a titer several times higher than the native rhizobia in the soils.
Testing the competitive ability of inocula
strain is very important to enhance the effect of
inoculation. There are no easy and accurate
methods for determining competitiveness of
rhizobia in soils and identifying the strains
responsible for nodulation. However, with thedevelopment of a technique using the resistance of rhizobial strains to antibiotics, strain
identification can be carried out without too
much difficulty. The method therefore wasadopted in this study to select effective andcompetitive strains of rhizobia for mungbean
in marginal soils.

b. Field Experiments (Tanay National Soil andWater Resources Research Center)
Two subsequent field trials were done in
Tanay which was previously classified as an
Ultisol. The treatments in the experiments were
the following:
1. Liming:- no lime- with lime (3 tons dolomite)
2. Rhizobium strains- no inoculation- MCO- CY 44- TAL 209
- TAL 441
- Mixture of the strains

Methodology:
a. Development of antibiotic resistant strains of
rhizobia for mungbean

The treatments were replicated 3 times andwere arranged following the Randomized
Complete Block Design (RCBD). In bothexperiments, the mungbean cultivar planted

To investigate the successful establishment
and survival of applied rhizobia in the field, theantibiotic resistance marker was employed.
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
was PSB Mg-6. Fertilization was based on soil
analysis. (29-60-60 kg NPK/ha). At full blooming stage, 10 representative plants from thesampling area in each plot were taken for dry
weight determination and nodule occupancy
analysis. The plants were grown to maturity for
determination of yield and yield components.
The general view of the experiment is shown in
Figure 1.
Fig.2. Plant-infection test showing infectiveness ofantibiotic-resistant strains for mungbean as indicated by
the presence of nodules on the test crop.

was the most competitive among the strainstested during the first field trial (82 % nodule occupancy in unlimed soil and 100 % in
limed soil) as shown in Table 1. The otherstrains were not as competitive as strain
MCO. However, during the second fieldtrial, recovery of the strains from the nodules
was lower. Strain MCO still occupied mostof the nodules of plants in soils not amended
with lime (75.3 %) while strain TAL 209
occupied 65.4 % of the nodules of plants insoils applied with lime.

Fig. 1. General view of the experiment during the first
field trial.

Salient Findings:
1. All the strains used in the study developed
their resistance to antibiotics after three (3)sub-culturing. Result of the confirmatory
trial to test retention or loss of infectivenessof the antibiotic resistant strains showed that
all the strains retained their infectiveness ontheir host as shown by the presence of nodules in plants inoculated with the four strains(Fig. 2).
2. For single strain inoculation, strain MCO

In mixed inoculation, the most competitive
were strains MCO and CY 44, but the indigenous rhizobia occupied most of the nodules
especially during the 2nd field trial (Table 2).
This phenomenon showed the very formidable
advantage of the indigenous rhizobia in forming nodules. The introduced rhizobia were notable to completely overcome the resident rhi-

Table 1. Percent nodule occupancy of Rhizobium strains in nodules of mungbean plants grown in limed and unlimed
soil in Tanay (Ultisol) during the 1st and 2nd field trials.

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Table 2. Competitiveness of Rhizobium strains in mixed inoculation.

<References>
1. Somesagaran, P. H. and H. J. Hoben. 1994.
Handbook for Rhizobia. Springer-Vorlag.
New York. 450 pp.
2. Trinick M. J. 1990. The ecology of
Rhizobium. Interactions between Rhizobium
and other soil organisms. Plant and Soil.
108: 137-141.
3. Triplett, E. W. 1992. Genetics of competition
for nodulation of legumes. Annual Review
of Microbiology. 46: 399-428.

zobia which formed a bulk of the total nodules
analyzed.
3. Response of mungbeans to inoculation of thedifferent strains showed that on the average,
a slight increase on the seed yield of theplant was observed with inoculation. Liming
seemed to have a negative effect on the yieldof the plants during the first field trial, however, during the second trial, the applicationof lime increased the seed yield irrespectiveof inoculation treatment (Figure 3.)

Fig. 3. Inoculation and liming effects on the seed yield of mungbean grown in an Ultisol.

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Philsurin developed Biofertilizer for sugarcane
the tillering capacity of the plants. There wasno significant difference on the number oftillers between the inoculated plants grown at
suboptimal level of nitrogen and the plants
grown at optimum level of nitrogen indicatingthat bacterial inoculation can replace needed
nitrogen to meet the optimum level of growth.

The Philsurin (Philippine Sugar ResearchInstitute Foundation, Inc.), a non-profit organization created in 1995 has developed biofertilizer for sugarcane in line with its functions toundertake sugarcane research, development,
and extension. Sugarcane production requires
high fertilizer inputs, thus development of
biofertilizer for sugarcane entails reduction of
production cost. Sugarcane was found to havethe capability of obtaining nitrogen from air for
its development through the association established with nitrogen fixing bacteria called asdiazotrophic endophytes.

Pot experiment was also conducted to evaluate the effects of inoculation at different developmental stage of sugarcane. Seven bacterial
isolates were inoculated on four sugarcane
varieties (VMC 86-550, VMC 87-599, PS3,
and Phil 80-13). Agronomic parameters of sugarcane were evaluated as affected by the inoculation of different bacterial isolates. No result
however was reported for this aspect.

Philsurin microbiologists, Dr. Lucille C.
Villegas has conducted a research to develop
microbial inoculant for sugarcane by finding
superior strains of diazotrophic endophytes todevelop viable nitrogen fixing in association
with sugarcane. Promising isolates have been
selected and identified and the selected strains
can be successfully introduced to sugarcane invitro. Philsurin reported promising results.
Inoculation on micro propagated sugarcane
indicates that four isolates positively affected

Inoculated plants growing under natural conditions<Source>Dr. Lucille C. Villegas, Ph.D. Developmentof Microbial Inoculant for Sugarcane. Philsurin
Annual Report 2002 - 2003. Pg 39-41.

Diazotrophic endophytes on different media
A) thick pellicle in semisolid N free medium, B) orange
colonies on N free agar, C) chocolate brown growth on
glucose yeast extract CACO3 agar D) greenish browngrown growth on potato agar.

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BIOTECH Fertilizers:
the approach to improve crop productivity and quality

orrhiza, plant growth promoters and microorganisms that can degrade organic wastes.

Over at the National Institute of Molecular
Biology and Biotechnology (BIOTECH) in the
University of the Philippines Los Banos, themood is upbeat as scientists work their way inproducing agricultural biotechnology breakthroughs that would combat global hunger biofertilizers which they hope would solve thenever-ending problem on food security.
Biotechnology, a set of scientific tools thatmakes use of living organisms, such as
microorganisms to generate products and services, has been doing wonders to agriculture andfood sector. At BIOTECH, the first wave of
products paid off in the form of environmentfriendly yet affordable microbial-based biofertilizers and inoculants that can substantiallydouble the yield of rice, corn and other important crops and substitute for chemical-based
farm inputs. These products, generated by the
Biotechnology for Agriculture and Forestry
Program, underwent a series of trials - fromgreenhouse to larger field testing - to determine
if they are ready for commercialization and fortransfer to private entrepreneurs. These areclassified as: biological nitrogen fixers, myc-

Biological nitrogen fixers
The use of microorganisms to harness elemental nitrogen through nitrogen fixation is one ofthe promising alternative technologies toaddress the high cost of chemical nitrogen fertilizers and the increasing environmental problems attributed to their use. These microbes,
known as biological nitrogen fixers (BNF), candirectly fix nitrogen gas from the atmosphereto make it usable by the plants.
An example of this microorganism is therhizobia, which reside inside the nodules ofleguminous plants. The mixture of rhizobia,
sterile soil and charcoal led to the developmentof an inoculant known as NitroPlus. It is abiofertilizer specific for soybean, mungbean,
cowpea, peanut, garden pea, pole sitao andother food legumes. NitroPlus comes in powder form in 100-gram packet which is used tocoat seeds before sowing. One hectare requires
4-5 packets which is a lot cheaper than 2-3
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
and longer roots. It was also found effective inthe rooting of blackpepper cuttings and hardto-root ornamentals Eugenia and Bushida.
BioGroe also promoted leaf bud formation andplant survival of coffee stem cuttings andenhanced seed germination of the herbs marjoram and thyme. In the efficacy test conductedin Tarlac and Cavite, BioGroe was found toenhance seedling growth and lowered farmers'
production cost.
Cocogro is a mixture of plant growth hormones derived from coconut water. This natural product can be used as a fertilizer supplement to enhance and promote growth and yieldof plants, especially tissue cultured orchids andorchid seedlings. Researchers have proven thatorchid seedlings, when sprayed with Cocogro,
increased in growth by 100% within a month.
Field trials showed that Cocogro can increase
yield of beans by 64%, peanuts by 15%, sweet
pepper by 100% and Chinese cabbage and by
20-30%. Cocogro is also good for ornamentals,
cereal crops, fruit trees and flowering plants. It
increases root proliferation, shoot development, bud formation and early flowering.

bags of ammonium sulfate. Field trials done incollaboration with various agencies showed anincrease in soybean yield by 124%, mungbean
by 29% and peanut by 39%. It was also found
to replace 30-50% of the nitrogen requirementsof plants and has a fertilizing value equal to 30
kg N/ha. Presently, NitroPlus is being tested on
non-legumes.
BIOTECH is also successful in producing
BIO-N, a biofertilizer that contains
Azospirillum, a bacterium isolated from thegrass Saccharum spontaneum or talahib. BION enhances root development, growth andyield of plants (rice, corn and other agricultural
crops) and provides 30-50% of their nitrogenrequirements. Efficacy testing of Bio-N on rice
variety RC 18 showed improvement in the
average number of tillers, straw and grain
yields. Five packs of Bio-N are needed to inoculate a hectare of rice or corn. It was also
found to increase yield of vegetables (tomatoes, okra, eggplant, lettuce and ampalaya).
BIO-N is available in powder form in 200-g
pack, which is meant either for seed inoculation, direct broadcasting over seeds or mixed
with water as root dip.

Mycorrhiza
Plant Growth Promoters
Certain microorganisms also provide phosphorus and other nutrients. Mycorrhiza is a symbiotic association between the roots of plants anda fungus. The association provides many benefits to plants such as increased absorption ofnutrients and water. It likewise serves as a biological control agent against root infection and
can improve soil properties. BIOTECH has
produced four mycorrhizal products:
Mycogroe, Mykovam, VAM Root Inoculant
and BrownMagic.
Mycogroe is a biofertilizer in tablet form
containing spores of ectomycorrhizal fungi.
The fungi, when inoculated to reforestation
seedlings, will infect the roots and help absorb
water and nutrients particularly phosphorus.
They can also prevent root infection bySome microorganisms produce plant growth
regulators such as auxins, gibberilins andcytokinins which can be used in plant propagation.
BioGroe is a biological fertilizer containing plant growth-promoting bacteria (PGPB)
that influence root growth by producing plant
hormones and provide nutrients in soluble
form. It is packaged as a powdered inoculantthat can be used for almost all types of plants vegetables, ornamentals, flowering plants andherbs. Experiments done at BIOTECH showedthat it increased ampalaya yield by 90% andlettuce by 40%. Dendrobium, on the other
hand, had early flowering while Mussaenda
and Hibiscus (gumamela) produced heavier
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
VAM Root Inoculant is composed ofdried chopped roots infected with VAM fungi.
These fungi assist plant roots in absorbing
water and nutrients and thus reduces chemicalfertilizer requirement of crops. VAM Root
Inoculant also acts as biocontrol agent against
root pathogens and produces certain substancesthat promote plant growth and improve soil
structure. Experiments done at BIOTECH
showed improved growth and fruiting capacityof papaya, increased yield of peanut andenhanced phosphorus uptake of bitter gourd. It
is also an effective growth promoter of agricultural (banana, sugarcane, corn, root crops) andhorticultural crops, vegetables (eggplant, okra,
tomatoes, onion), fruit trees and ornamentals.

pathogens and increase plant tolerance todrought and heavy metals. Mycogroe wasfound effective in promoting the survival andgrowth of trees and can replace 60-85% of
their fertilizer requirement. Application is done
only once throughout the entire life of the tree.
Mycogroe is applicable only for trees such aspines, dipterocarps, Eucalyptus, Alnus, Acacia
and Casuarina. One tablet is inoculated into
each seedling during transplanting in the nursery.
On the other hand, Mykovam is a powdered biofertilizer which has effective speciesof vesicular arbuscular mycorrhizal (VAM)
fungi. Basically, its functions are similar tothat of Mycogroe. The fungi infect the roots,
form a close association with the plant andassist in absorbing water and nutrients such as
phosphorus, zinc and nitrogen. Mykovam canbe applied to almost all plants: agricultural
crops (upland rice, corn, tomato, eggplant,
onion, garlic, pepper, cassava, and sweet potato), fruit trees (guava, rambutan, papaya, citrus,
banana, lanzones, coffee, guyabano, coconut
and mango), ornamentals and reforestation
species (acacia, gmelina and mahogany).
Mykovam even fights destructive pests and diseases and increases plants tolerance to drought
and heavy metals. Use of this product reduces
60 -85% of the fertilizer requirements of mostplants. It is economical to use since a kilo of
Mykovam can fertilize 400 seedlings.
Brown Magic consists of finely chopped
and powdered fruiting bodies of endomycorrhizal fungi. It is especially effective for invitro cultured orchids and provides the nutrients necessary for their growth. It also serves
as a biological control agent of root infection
and other harmful pests and diseases.
Experiments done at BIOTECH showed
healthier, greener and more vigorous
Dendrobium, Vanda and Cattleya seedlings.
Plants had more suckers and spikes and produced flowers earlier and at a longer period oftime.

Microorganisms that decompose
agricultural residues
BIOTECH has produced an organic fertilizer
technology through composting which can significantly enhance crop growth and yield.
Chicken, swine or cattle manure can be combined with farm wastes such as rice straw, coffee hulls and corn stover or with agro-industrial
wastes such as sugarcane bagasse and mudpress and processed into bio-organic fertilizer.
The technology also involves the use of two
inoculants: Bio-Quick which contains
Trichoderma sp. that can degrade organic
waste materials in only 3 to 4 weeks instead ofalmost 6 weeks, and Bio-Fix, the enrichment
inocula containing Azotobacter sp. which isadded to the compost heap to increase thenitrogen content of the final product. The endproduct called Bio-Green can be applied to allkinds of plants and can increase yield by 2030%. It also serves as soil conditioner,
enhances the nutrient uptake of plants in soil
and thus makes plant healthier. Bio-Green
can be applied basally or directly depending on
the crop. If applied at 10 to 20 bags/ha, it canequal the effects of chemical fertilizer at 60-3030.
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
However, Bio-N is just one of the biofertilizers which is being aggressively promoted by
the Institute. Media exposure for NitroPlus,
Mykovam, Mycogroe, BrownMagic and
BioGroe were also sought to intensify our
information campaign on biofertilizers. As aresult, researchers were interviewed in various
radio and television programs while featurearticles of their technologies were published innewspapers, magazines and newsletters.
Demonstration trials done both at BIOTECH
and in farmers' field validated the efficacy ofthe biofertilizers, much to the satisfaction of
farmers.
To facilitate the efficient technology transferfrom the laboratory to target industries and/or
beneficiaries, BIOTECH prepares pre-feasibility studies, industry analyses and businessplans. Applications for experimental use permit, product registration, technology licensing,
joint venture are likewise done. Bio-N for rice
and NitroPlus for soybeans are now registeredwith the Fertilizer and Pesticide Authority, as aresult of our techno-demo farms utilizing theidle lands around BIOTECH.
Judging from the increasing demand fromfarmers and plantation owners, BIOTECH isindeed fulfilling its mission of developing fertilizers that do more than just provide morebountiful yield. The plethora of interest generated by our technologies inspires ourresearchers to continue the research, development and extension programs they have started.
Concomitantly, they hope to further widen thepartnership with different public and private
agencies, biotech firms in particular, to facilitate the commercialization process of
BIOTECH biofertilizers to make them readily
available to farmers in different regions of thePhilippines and in other countries as well.
(Imelda V. Garcia, Program Leader,Communication and Technology Utilization
Program of Biotech)

Promotion and Technology Transfer
of BIOTECH Biofertilizers
Research does not end with the discovery of atechnology or process. More important is theutilization of the technology or process byintended clients. Thus, at BIOTECH,
researchers are not just keen on producing highquality fertilizers but also in reaching the farmers, even those in marginal areas of the country.
They go beyond the confines of laboratories togenerate and sustain a high level of awarenessand acceptance of biofertilizers. They do thesein tandem with the extension and communication specialists of the Institute. One promotional approach used is the production of an integrated information package through print andmultimedia materials. Biofertilizer fact sheetsor leaflets are given to farmers and extension
workers as supplementary materials duringseminars, forums or trainings, where
researchers serve as resource speakers. Anotherapproach is to facilitate institutional collaboration and establish links with different agencies,
including local government units, in the transfer or commercialization of BIOTECH biofertilizers. They also collaborate with setting-up
field demonstration areas to see the efficacy ofour biofertilizers.
Case in point: to create awareness andencourage extensive support and patronage of
farmers, the Bio-N staff work hand in handwith the Department of Agriculture in implementing the transfer of the technology in threedifferent phases. Training workshops on Bio-N
application were held in different regions of thecountry complemented by presentations,posters, handouts and product samples for
actual testing. Bio-N mixing plants were likewise established to address the problem on theunavailability of supply and/or accessibility to
distribution centers. BIOTECH also partners
with the Technology and Livelihood Resource
Center in marketing Bio-N.
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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

VITAL N R : A Revolutionary Biofertilizer Technology with a Fastgrowing Market
Dr. Ponciano M. Halos
Chairman of the Board / Arnichem Corporation
Pleasant Village College, Laguna 4031 Philippineswww.vitaln.com
ity by at least 15%, and in some cases morethan 100%. Tests conducted by independentscientists and farmers in rice and corn show
that, VITAL N R increases yields by at least one
ton per hectare. In onion, additional yields of
15 tons per hectare can be realized by using the
product. With its efficacy and the low cost of
the product, it is well suited for the majority ofsmall farmers who can not afford the full costof recommended fertilizer requirement andwho badly need a harvest booster.
The following tables (Tables 1-3) were partof the technical data obtained by Dr. Nora

A relatively young
but revolutionary
biotech productsuch as VITAL N R
is now getting a lotof attention. The
demand for costeffective and innovative VITAL N R biofertilizer is rapidly growing. In fact, VITAL N R now has a fast-growing
list of distributors/dealers all over the country.
VITAL N R is a wettable powder formulationthat is available in a very convenient 100-gram
pack. This small package packs a lot. It acts
as a virtual nitrogen fertilizer factory at theroots of the plants. It protects plants from rootand leaf diseases. It also makes plants
drought-resistant. Not only is it convenient andpowerful, it is also easy to store. It can be
stored at room temperature for at least 3 years.
VITAL N R is a breakthrough and sustainableagriculture technology for increasing farmers'
incomes and food sufficiency. Furthermore, itis proudly 100% Filipino! The product is theresult of my years of research together with myscientist wife, Dr. Saturnina C. Halos.
Together with our children and in-laws, weformed a progressive biotech company,
Arnichem Corporation, to make the productavailable to the average Filipino farmer. With
this product, we hope to re-VITALize thenation.
VITAL N R is an FPA-registered organic
biofertilizer that replaces as much as half of therecommended fertilizer. This translates to asavings of 4 bags of fertilizer per hectare (ha)or about P3,000 for rice and corn and 6-8 bagsof fertilizer/ha (about P5-6,000/ha savings) for
vegetables. It also increases farmer's profitabil-

Table 1. The effect of VITAL N R inoculation in
OPV corn (white corn) yields

Table 2. The effect of VITAL N R on yellow hybrid
corn (DK818Y) under late drought
conditions.

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project
Table 3. The effect of VITAL N R on rice yields

Kinchay

Inciong, Consultant, Bureau of Soils and Water
Management, Dept. of Agriculture submitted to
the Fertilizer and Pesticide Authority in supportof the registration of VITAL N R as a biofertilizer. (RIF-Recommended Inorganic Fertilizationrate).
The following are results obtained by undergraduate thesis students of Dr. Betty Malab,Professor of Soil Science, College ofAgriculture, Mariano Marcos University,
Batac, Ilocos Norte. The results show that 1/2
RIF plus VITAL NR give similar yields as
those of RIF.
Farmer scientists participating in the program of Dr. Romulo Davide in Bantayan, Cebu
obtained more than 100% increase in yields
when they applied VITAL N R (3.86 MT/ha)
compared to their practice of applying nothing(1.76 MT/ha). This yield was comparable towhat they obtained when they applied complete fertilizer (14-14-14) also at 3.86MT/ha.
In Toledo City, using IPB 929, an increase of
62.5% of corn treated with VITAL N R (3.77
MT/ha) was obtained over that without any
treatment (2.32MT/ha).

rice

without VITAL N R

with VITAL N R
Corn

Table 4. The yield of pepper (Capsicum anuum L) as affected by varying levels of fertilizer and VITAL N R

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FNCA (Forum for Nuclear Cooperation in Asia) Biofertilizer Project

Project leader of The Philippines

Richard M. Balog
Science Research Specialist I
Soils Unit
Agricultural Research Group
Atomic Research Division
Philippine Nuclear Research Institute (PNRI)
Commonwealth Ave., Diliman, Quezon City, 1100 Philippines
Tel. No.: (+632) 929-6011 to 19 local 237
Fax No.: (+632) 920-8765
E.Mail: rmbalog@pnri.dost.gov.phResearch Interest□ Use of nuclear techniques in agricultural research● Use of 15N isotope dilution technique in fertilizer use efficiency and biological nitrogen
fixation studies● Use of fallout radionuclide 137Cs in soil erosion and sedimentation studies
Positions held
□ Project Leader, Forum for Nuclear Cooperation in Asia Biofertilizer Project□ Project Leader, Department of Agriculture-Bureau of Agricultural Research funded project on
the Use of Cs-137 in Estimating Soil Erosion and Sedimentation□ Member of the National Project Team for International Atomic Energy Agency RAS/5/043
project on Sustainable Land Use Management and Strategies for Controlling Soil Erosion and
Improving Soil and Water Quality

Check FNCA Web Site!
http://www.fnca.jp/english/

FNCA Biofertilizer News Letter No. 7
Editor:Mr. Richard M. Balog
Leader of FNCA Biofertilizer Project of the Philippines
Secretariat: Ms. Yuko Wada
Asia Cooperation Center (ACC)Japan Atomic Industrial Forum, Inc. (JAIF)
Tel:+81-3-6812-7104 Fax:+81-3-6812-7110 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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