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I A PRIMARY

15JECT J Food production and nutrition AF25-0O00-GG50 CLASSI

SI0tfA~y0 OFFICATION Soil fertility fertilizers and plant nutrition--Tropics

2 TITLE AND SUBTITLE

Utilization of the Azolla-Anabaena complex as a nitrogen fertilizer for rice

3AUTHOR(S)

WatanabeI EspinasCR BerjaNS AlimagnoBV

4 DOCUMENT DATE 15 NUMBER OF 5 ARC NUMBERPAGES

1977 1 5P IG - J ARC

7 REFERENCE ORGANIZATION NAME AND ADDRESS

IRRI

0 SUPPLEMENTARY NOTES (Sponsorind Ordaniatlon Publlahers Availability)

(InIRRI respaper sernoll) (Presented at meeting of the Crop Science Socof the PhilippinesBaguio1977)

9 ABSTRACT

Because it contains nitrogen fiLinr blue-green algae Anabaoena asoZa in its fronds AzoZZa grows on a nitrogen-free solution doubles its mass in 3 or 5 days and accumulates 30 or 40 kg of Nha in 2 weeks

A calcium or phosphorus deficiency in the AzoiZa culture solution produces severe deterioration on growth The optimum solution pH for Azolla is 55 a higheL plHproduces iron deficiency AzoZZa is sensitive to a temperature higher than 31

0C (350 day - 270 night) which causes reddish-brown discoloration and reduces vigor

Nitrogen inAzoZZa is slowly mineralized in a submerged soil about

75 of total nitrogen mineralizes in 6 to 8 weeks Pot experiments reveal that nitrogen in dried AzoZLa increases rice growth but its availability to rice is 40 lower than that of ammonium-fertilizer nitrogen

In the rice paddy AzolZa is easily multiplied Five crops of AzoZZa (October to January) produced a total of 117 kg Nha in 106 days Phosphorus fertilizer was effective in promoting abmdant growth Inoculating AzoZZa with phosphorus at the time of transplanting rice and incorporaLing it after 40 days of growth it-reased the yield of dry-season rice (IR30) over that in plots with only midseason puddling and phosphorus

AxoZZa has a potential as a self-supplyi-ag nitrogen fertilizer source for farmers growing rice inbunded flooded soils

II PRICE OF DOCUMENT10 CONTROL NUMBER

13 PROJECT NUMBER12 DESCRIPTORS

Azolla-Anabaena Tropics Fertilizers 14 CONTRACT NUMBER

Nitrogen AIDta-G-1074 GTS Rice sTYPE OF DOCUMENT

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IRPS No 11 November 1977

COMPLEXrE UTILIZATION OF THE AZOLLA-ANABAENA 1 AS A NITROGEN FERTILIZER FOR RICE

ABSTRACT

Because it contains nitrogen fixing blue-green algae Anabaefa azota

in its fronds AzoZla grows on a nitrogen-free solution doubles its

mass in 3 or 5 days and accumulates 30 or 40 kg of Nha in 2 weeks

A calcium or phosphorus deficiency in the Azolla culture solution The optimum solution pH forproduces severe deterioration on growth

a higher pH produces iron deficiency Azolla isAzolia is 55 - 270 night)sensitive to a temperature higher than 31

0 C (350 day

which causes reddish-brown discoloration and reduces vigor

Nitrogen in Azolla is slowly mineralized in a submerged soil about Pot experiments75 of total nitrogen mineralizes in 6 to 8 weeks

reveal that nitrogen in dried Azolla increases rice growth but its

availability to rice is 40 lower than that of ammonium-fertilizer

nitrogen

In the rice paddy Azolla is easily multiplied Five crops of AzolUa

(October to January) produced a total of 117 kg Nha in 106 days-

Phosphorus fertilizer was effective in promoting abundant growth

Inoculating Azolla with phosphorus at the time of transplanting rice

and incorporating it after 40 days of growth increased the yield of

dry-season rice (IR30) over that in plots with only midseason puddling

and phosphorus

AzoLa has a potential as a self-supplying nitrogen fertilizer source

for farmers growing rice in bunded flooded soils

by I Watanabe soil microbiologist Corazon R Espinas research

assistant Nilda S Berja research assistant B V Alimagno senior

research assistant International Rice Research Institute Los Bafios

Philippines This paper was originally presented at a meeting of the

Crop Science Society of the Philippines 5-7 May 1977 Baguio Philippines

and revised and submitted to the IRRI Research Paper Series Committee

3 IRPS No 11 November 1977

THE UTILIZATION OF THE AZOLLA-ANBAENA COMPLEX

AS A NITROGEN FERTILIZER FOR RICE

AoZla is a genus of water fern that assimilates atmospheric nitrogen in association with nitrogen fixing blue-green algae Anabaena azollae that live in the cavities of AzoZlas upper lobes (Oes 1913 Saubert 1949 Moore 1969 Secking 1976 and Ashton and almsley 1976) The nitrogen fixing ability of the Azolla-Anabaena complex offers potentia for increasing rice yield at comparatively low cost in the face of possible shortages and rising prices of fertilizer nitrogen The AzolZa fern especially offers a source of organic nitrogelL fertilizers for rice farmers who cannot afford chemical nitrogen fertilizers

Azolla is widely distributed in the rice-growing regions of the tropic and the temperate zones and grows on the water of irrigated rice fields It quickly covers the surface of the floodwater but does not interfere with the normal cultivation practices for the rice crop Azolla is

extensively used as a green-manure crop in Vietnam (Tran Quang Thuyet

and Dao The Tuan 1973) and southern China but it has had less attention

in the rest of -Southeast Asia

Experiments started at IRRI in 1975 to determine the problems End the

applicability of Azolla culture for rice production This paper details some preliminary results that may be of interest and use to others wishing to study Azolia as a source of organic nitrogen for the rice crop

AZOLLA GROWTH IN WATER CULTURE

AzoZla for the IRRI studies was collected from a rice field at Santo Domingo Albay province Philippines The collected plants resemble Azolla pinnata (Fig 1) Because the plants failed to form sporocarps

the species has not been identified The laboratory and greenhouse studies determined growth rates mineral requirements and temperature responses

Growth rate in culture solution

Initially Azolla was grown in a nitrogen-free nutrient solution containitig 20 ppm P (NaH2PO42H20) 40 ppm K (K2S04) 40 ppm Ca (CaC12) 40 ppm Mg

(MgS047H20) 05 ppm Mn (MnCl2 4H20) 01 ppm Mo (Na2MoO4 2H20) 02 ppm B (H3B03) 001 ppm Zn (ZnS04) 001 ppm Cu (CuS04 5H20) and 2 ppm Fe (Fe-citrate) The pH of the solution was adjusted to 55 Preparation of the solution was similar to that described for rice culture by Yoshida

et al in 1971 Growth was in 3- or 5-cm deep culture solution The

4 IRPS No 11 Novembe 1977

lopI

Fig 1 Azolla species used in IRRI experiments 1975-77

new solution everyAzolla was maintained in the greenhouse by transfer to a

2 weeks

een 3 to In the nitrogen-free solution Azolla doubled its weight bet

In 2 weeks the equivalent of 22 and bull 5 days during the first week

36 kg Nha was accnnulated in the Azolla mass grown in shallow trays

Azolla contains between 3 and 5 nitrogen on a dry-Vigorously growing weight basis or 01 to 02 nitrogen on a fresh-weight

basis Assuming

3 nitrogen content and 3 days doubling time the nitrogen fixing rate

of AzolZa is 78 mg Ndry weight daily

The ammonium concentration in the nitrogen-free solution was less than

1 ppm at the saturated population level of AzolZa growth

Mineral requirement studies

The mineral requirements of Azolla were studied in solutions deficient

in phosphorus potassium calcium or magnesium and in a nitrogen-

Each medium amended solution with 40 ppm nitrogen as ammonium

nitrate

was tested with 2 g of Azolla placed in a tray filled to a depth of 33

The pH of the solution cm nutrient solution and grown in a gieenhouse Fresh weight of AzolZla was adjusted to 55 daily and changed weekly

was recorded and after 2 weeks a sample was taken for total nitrogen

analysis acetylene-reduction assay and microscopic observation to

check the presence of blue-green algae in the fronds

Table 1 shows Azolla growth in phosphorus- potassium- calcium- and

magnesium-deficient solutions in a nitrogen-amended solution and in

Phosphorus and calcium the complete nitrogen-free solution as control Fresh weight nitrogendeficiencies affected Azolta growth severely

IRPS NolINovember 1977

Table 1 Fresh weight totalnitrogen and acetylene-reduction activity

of AzoLa grown in P- K- Ca- and Hg-deficient solutions and in a nitrogen-amended solution Harvest was 2 weeks after inoculation IRRI 1975

Fresh weight Total N content Acetylene-reduction assay Treatment - a

(gplate) (kgha) (nmolesg fresh weighth)

~b 340 + 102- N controls 368 + 147 446 + 014

-N-P 82+ 04 73+011 12+0 - N -K 116 10 109 T 10 830 120 - N -Ca 34 02 22 + 007 0 - N-g 30 3 + 08c 343 + 04 440 + 277 + N 125 +05- 193 + 10 289 + 75

a Plate size was 314cmz b Average + standard deviation of the average

(2 replicates) E Contaminating green-algae were not completely removed

content and acetylene-reduction activity in the phosphorus- and calciumshydeficient solutions were generally far less than those for the other treatments

The AzoUla grown in the phosphorus-deficient solution had a red-brown discoloration that spread from the center of the frond to the tip of

the body and frond size was smaller than that for AzolZa grown in the

control solution Compared with those in the other deficient solutions

the roots of Azolla grown in a phosphorus-deficient solution were also

red-brown longer and tended to separate easily from the Azolla body Calcium-deficient Azolla showed more browning than the phosphorusshy

deficient Azolla and had the smallest frond size among treatments

Fresh weight and nitrogen content of the Azolla in the potassiumshy

deficient solution were less than those in the control but the acetyleneshy

reduction activity was more than twice that of the Azolla in the control

solutions The acetylene-reduction activity in the control solution may

have been lower because the AzoZZa had reached a saturated population density at harvest and its nitrogen fixing activity could have been lower

than that of the actively growing one Potassium-deficient Azolla

did not have any discoloration but its frond size was relatively smaller

than that of Azolla in the control

Fresh weight and nitrogen content of Azolla in the magnesium-deficient The magnesiumshysolutions were slightly lower than those in the control

deficient Azolla did not visually differ in color and frond size from

the control Azolla

Azo la in the nitrogen-amended medium had lower fresh weight and nitrogen assimilation than that in the control solution but retained

Because the growth conditionsasimilar acetylene-reduction activity

6 IRPS No 11November 1977

were not sterile green algae covered the solution surface witUn 2

That would accountdays and allowed little area for Azolla to spread The Azotla samples were not completelyfor the low fresh weight

free of green algae at harvest

In the calcium-deficient Azola no blue-green algae were visible under

the microscope In phosphorus-deficient Azolla blue-green algae were

but were less green than those in normal Azola In thevisible AzolZa deficient in potassium and magnesium the number of

the blue-green

algae was smaller and the color was less green than that in the control

The addition of mineral nitrogen to the solution did not completely

exclude blue-green algae

pH and iron avaiZability

AxoZa was grown in the nitrogen-free nutrient solutioYn in the greennouge

with two levels of iron concentration (2 and 4 ppm as iron citrate) and

The pH was adjusted daily Thefour pH levels (45 55 65 75)

solutions with 2 ppm Fe at pH 55 and without iron at pH 75 served as

control Water depth was kept at 33 cm

Five grams (fresh weight) of Azolla were inoculated into each solution

and grown for 2 weeks with weekly change of solution Weekly fresh

weight of Azolla per tray was measured and a 2-g sample was taken for

After harvest an acetylene-reduction assaytotal nitrogen analysis was made

The effect of different levels of pH and iron after 2 weeks growth is

Azolla shown in Table 2 Increasing the pH level caused a decrease in

The growth and nitrogen accumulationgrowth at low iron concentration At a pH higher than were lowest in the iron-deficient medium at pH 75

65 Azolla fronds turned yellowish The nitrogen fixing activity was

These results indicate that availability of iron for lowest at pH 75 Such an effect is common in water culture of Azolla is affected by pH

higher plants The optimum pH of water-culture solution is 55 but the

optimum pH for Azolla growth in floodwater may be different and should

be determined separately

Response to temperature

AzolZas response to temperature was checked with 2 g fresh Azoia

samples inoculated into each of three 20-cm diameter trays with

1 liter of the medium and kept in the greenhouse at four temperature

Night (00O-1800 hours) temperature was 80 lower than daylevels temperature Fresh weight was measured and the culture solution

After 2 weeks the Azoila was harvested to was changed weekly determine fresh weight and total nitrogen

The results are in Table 3 The fresh-weight yield and nitrogen accumulation

were not significantly different among treatments kept at 220 250 and 280 C

IRPS No11 November 1977

Table 2 The fresh weight total nitrogen and acetylene-reductionactivity of AzoZa grown in culture solutions of 2 ppm iron and 4 ppm ironwith 4 pH levels IRRI 1975

ppm Fe pH 45 pH 65pH 55 pH75

STotal fresh weight (gplate)

U b -6 - 261+ 11 262+ 02 260 + 26 213 + 02 2 - 235+ 02 4 -210+ 06- 3331+ 16 227 +06 226+-02-

Increase in nitrogen (kgha

0 84 +16 1 252-+16 270 +23 171 + 18 i50 +31 2 208+14 4 185 +_03 218+ 15 162 +09 154+ 06

Acetylene reduction (nmoles C2H4g fresh weighth)

0 77+8 1 605 +48 915+ 40 386 + 14 151 +4 1 1000 1 44t 4 895+100 945 + 23 821 + 183 259 + 17

2a Plate size is 314 cm b Average + standard deviation of the average

zruc weignc ana uora nicrogen ot AOagrownatfourdifferenttemperatures

a -- NitrogenTemperature PC Fresh weight (gtray)- (kgha) at

Daynight Average 1 wk 2 wk 2 wk

2618 22- 88 + 0 276 + 03 320 + 042921 25 102 + 01 266 + 11 311 + 08 3224 28 108 + 02 273 T 08 308 T 18 3527 31 154 + 28 1949 + 08 146 +

a Plate size was 314 cm2 b Average + standard deviation of the average (3 replicates)

8 IRPS No 11November 1977-

At 310C the growth of Azolla decreased Althou~gh no difference in fresh

weight at 2-week growth was observed Azola showed smaller fronds and more

yellowish-brown color as temperature increased above 22degC

ReZation of acetyZene-reduction rate to nitrogen increase in Azolla

AaoZZa was grown in a nitrogen-free solution in the greenhouse une

gram (fresh weight) of Azolla was analyzed for total nitrogen content

at the start Samples were taken 14 19 and 22 days after inoculation

The solution was replaced after each sampling

Azolla samples were placed in 50-ml Erlenmeyer flasks with 10 ml nitrogenshy

free culture solution Half of the flasks had 10 acetylene added in the

gas phase the other half received no acetylene All flasks were plugged

with serum stoppers After 24 h incubation under 4 klux illumination

by fluorescent light at 320 C the samples with acetylene were

analyzed for ethylene formation and those without acetylene for total

nitrogen content by the Kjeldahl method The increase in nitrogen was

determined by measuring the differences in total nitrogen during incubation

Three samples of Azolla were used for nitrogen and ethylene determination

Ethylene formation was almost linear during 24-h illumination

For AzolZa samples taken 14 19 and 22 days after inoculation total

nitrogen increase for 24-h incubation showed 028 + 009 (SD) 045 + 10

(SD) 024 + 010 mgg fresh weight respectively whereas acetylene

reduction rate was 343 + 125 253 + 245 and 202 + 14 pmole C2H4g

fresh weight respectively Thus the ratio of C2H2 reduction to N2 reduction was 34 16 and 24 respectively The values are close to

the theoretical ratio value 3

AVAILABILITY OF AZOLLA NITROGEN TO THE RICE CROP

Laboratory greenhouse and field experiments with flooded soils measured the availability of AzolZa nitrogen to the rice plant The nitrogen accumulated in the Azol~a bodyis considered available to rice plants only after Azol~a decomposition Fresh and air-dried Azolla were tested in the laboratory Azotla was placed on half of a sample of air-dried Maahas clay soil in a test tube (2-cm diameter) and the other half of the soil placed on top of the Azolla The whole was flooded to a 5-cm depth with distilled water The procedure kept AzoZla from floating The added Azolla was adjusted to give about 5 mg nitrogentube The tubes were incubated in the dark at 300C and 2N KCl-extractable ammonia was determined periodically

The results are in Table 4 Ammonia release was more rapid from fresh AzolZ than from dried Azolla After 6 weeks 62-75 of total nitrogen in Azol~a had been released as ammonia The results indicate that the- nitrogen inAzolZa becomes available slowly


Table 4 Mineralizationi of niroienofrom dried- and fresh AzolZa in flodedsoi

Weeks NH4 -N Percentage of after Control With from ammonification of flooding soil AzoZZa AzoZZa AzoZia nitrogen

pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731

a Total nitrogen of AaofZa is 497 pgg soil h Total nitrogen of AzoUa is 273 igg soil

Pot experiments to deteinine avaiZabiZity of AzoZla nitrogen

The availability of the nitrogenin Azolla was measured in freshMaahas clay soil from a field where no nitrogen fertilizer had been applied The soil was put in pots (200-cmz surface) and nitrogen fertilizers at 05 g Npot and 10 g Npot either as ammonium sulfate or as dried AzoiUa were mixed with the surface 10 cm of soil and the pots were flooded with deionized water Two-week-old seedlings of IR26 were transplanted and the pots were kept flooded in the greenhouse At harvest (110 days after transplanting) the grain and straw of rice plants that received no fertilizer contained 159 mg Npot those of plants that received 05 and 10 g Npot as ammonium sulfate contained 601 and 936 mg Npot respectively In the pots with dried Azoltla at the 05 and 10 g N level nitrogen uptake was 411 and 694 mg Npot respectively Applied nitrogen absorbed by plants was 884 and 777 for ammonium sulfate and 504 and 533 for Azolla at low and high levels of nitrogen application respectively As with the laboratory


incubation study the pot experiment showed that nitrogen 4n the shyreleased quite slowly It is also slightlylessayailablei

Azolla body is than mineral nitrogen

Azolla growth trial in paddy field

Forlfield trials Azolla was multiplied in a 18m2 plastic-lined pond

The pond was flooded to a 10-cm depth with tap water adjusted to pH

55 and 10 g superphosphate 08 g ferric sulfate 0552 g citric acid

and 118 g carbofuran (0354 g active ingredient) About 100 g (fresh

weight) of AzolZa inocilum was added to the pond and allowed to grow

until Azolta covered the surface

Field triaZ For Azolla growth trials in an IRRI paddy a 5-m x 5-m

plot was prepared and enclosed with a metal fence to avoid overflow of Azolta A plastic net prevented loss of Azolla at the outlet

and inlet of irrigation water The plot was divided into four

quadrants by two pieces of bamboo floated perpendicular to each

Water depth was kept at about 5 cm and Azolla inoculumother obtained from the plastic-lined pond was added at about 4 kgplot

Immediately before inoculation Azolla was mixed with 210 g supershy

phosphate (15 kg P205ha) and 25 g carbofuran (075 g ai) The

Azolla was spread on two quadrlts in a plot After 7 to 10 days

when the surface of the inoculated area was covered by Azolla half

of the Azolla was c6llected and transferred to the two empty quadrants

Azolla had covered the plot it was collected by net FreshWhen weight dry weight and total nitrogen were measured

The data of five harvests from October to January are presented in

Table 5 In 106 days 57 t Azollaha (fresh weight) was obtained

from five crops The sum of nitrogen increase during this period was

35 kgha for each crop Doubling117 kgha ranging from 17 kgha to

days fluctuated from 55 days to 10 days Growth rate was lower than

that obtained in the culture solution in the greenhouse

the effect of Azolla inoculation on rice yieldField experiment on

rice growthA field experiment examined the effect of Azolla on

AzolZaa relation to phosphorus addition and the effect of midseason

Three factors at twopuddling to incorporate Azolla into the soil with and without Azollalevels were combined in the layout of plots

inoculation with and without phosphorus addition and with and without

midseason puddling Eight treatments were set at random on each of

three 5-m x 5-m blocks Azolla inoculum was obtained from the growth

trial plot Two-week-old seedlings of IR30 were transplanted 20 cm x was spread20 cm One day after transplanting 125 kg fresh Azolla

the surface of each Azolla-treated plot and superphosphateuniformly on

at 30 kg P205 ha was immediately broadcast on the phosphorus-treated

Plots were kept floodedplots No other fertilizer was applied


Table 5 Growth and nitrogen accumulatiI ohof AolZIinlthe field n1ocul piot (25 IR197C6 in2 )

Crop nmber 1 2 1 3 4

Date inoculated Oct 4 Oct -25 Nov 15 Dec--8 Decs- 28 Date transferreda Oct15 Nov-3 Nov 22 Dec 21- Jan 3 Date harvested Oct 25 Nov 15 Dec 8 IDec 28 Jan -18 Period of growth (days) 21 21 23 20 21 Amount of inoculated

(kgha) 1700 1200 2300 1900 1600 Amount harvested

(kgha) 10300- 15200 13700 12800 4800

Increased amount (kgha) 8600 14000 11000 10900 2400

Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen

I(dry basis) -ND D l 410 411 500 (fresh basis) 20lt i- 020s 0166 0169 0267

Nitrogen kg Nna

Initial u 46 32 27 Final 206 304 27 216 374 Increase 172 280 L81 184 347

Accumulated nitrogen

Increase (kg Nha) 453 634 r 818 1165

Doubling time of nitrogen increase (days) 80 57 100 sd 72

Temperature 330270C 320260C 29024 0C 30026degC lt29deg24dC

aDate of transfer to half of plot bNot determined cEstimation _dMaximum daily mean temperatureminimum daily mean temperature


Thirty-nine days after Azoica was added to the plots it was To do that floodwater wasincorporated during midseason puddling

drained until the surface soil was exposed and a hand weeder was passed

twice between rows of rice in passes perpendicular to each other

One day after puddling the plot was reflooded Azolla coverage of the

surface of the plots was estimated visually

Azofla growth in plots that received phosphorus fertilizer was more

vigorous than that in nonphosphorus plots The areas covered by

AzoZla at 21 days and 39 days after inoculation were 85 and 96 in

phosphorus plots but 62 and 76 in nonphosphorus plots After

AzoZla incorporation a small portion of it remained visible but

disintegrated at a later stage of rice growth

Straw yield and grain yield are given in Table 6 Among the plots

without phosphorus where Azolla growth was poorer inoculation

increased grain yield only in the plots without puddling In the

puddled plots puddling alone increased rice growth and overshadowed Among the plots that received phosphorusthe effectiveness of inoculation

fertilizer Azolla growth was abundant and straw and grain yields in

the plots with Azolla incorporation were significantly higher than those

in plots with puddling but no Azolla Azolla incorporation increased

straw weight more than it did grain yield The grain yield increase

by Azolla inoculation was about 12 when phosphorus was added and

midseason puddling was done

Table 6 Straw yield andgrain yield of IR30 field experiment on

AsoZa inoculation IRRI 1976 dry season

A Straw yield_ Grain yield

P W (kgha) (kh)Index i i

- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112

a P - phosphorus W = midseason puddling A = AzoVia inoculation

bIn each column means fqllowed by a common letter are not significantly

different at 5 level c- Grain yield of the corresponding treatment

without AzolZa inoculation is taken as 100


In phosphorus-treated plots where the surface of the field was fully covered by Azola the nitrogen content was estimated as 25 kg Nha As mentioned earlier availability of AzoZla in nitrogen is 40 less than that from ammoniacal fertilizer Consequently it is estimated that a field fully covered with Azolta might give nitrogen equivalent to 15 kg Nha from ammoniacal fertilizer

AZOLLA AS FERTILIZER SUBSTITUTE

The AzoZ a experiments reported here indicate the potential of-AzoZZa as a partial substitute for nitrogen fertilizer AzofZl has promise because of ease of its culture its high nitrogen fixing capacity the availability of its nitrogen to the current rice crop and its simultaneous growth with the rice crop

Ease of culture

Greenhouse and pond studies indicate the ease of growing Azol la

in a nitrogen-free culture solution if the pH of the solution is kept

slightly acid Itis easy to cover the rice field with ar AzolZa mat in

20 days (Table 5) Phosphorus and insecticides improve AzolZa growth

High nitrogen fixing activity

Our greenhouse study showed that the nitrogen fixing activity of the AzoUZa-Anabaena complex was as high as 78 mg Ndry weight per day The value is about 25 that of the nitrogen fixing activity of soybean nodules (Tuzimura 1950) In the field Azol~a fixed about 120 kg Nha in 106 days (daily nitrogen fixation rate of 11 kg Nha) Extrashypolating zhat value to an annual rate gives a potential 412 kg Nha

That makes the annual nitrogen fixing rate of Azolla comparable to

that of most tropical and subtropical nodulated leguminous plants

but inferior to that of Desnvdiwn intortwn (897 kg Nha per year) and

Sesbania cannabina (542 kg Nha per year) (Nutman 1976)

AvailabiZity of Azolla nitrogen

Our experiments reveal that the availability of nitrogen from dried AzolZa to a single crop of rice is slightly lower than that from ammoniacal fertilizer The availability of nitrogen in fresh AzolZa appears higher because mineralization of nitrogen is faster than that in dried AzoZla Tuzimu~as (1957) incubation studies also showed that Azofla as nitrogen fertilizer is as effective as defatted soybean cakes

Simultaneous growth with rice

Our experiments showed that Azol4 inoculated at lthe time of itransplanting


When grown under the canopy ofwill cover the surface of a rice field In

rice plants it also influences the rice yield of the current crop

Vietnam for example AzoZia is usually grown once or twice before

transplanting and the established mass is incorporated into the rice field

This procedurr may be more effective than growingbefore transplanting Azolla after transplanting rice

In a rainfed area or in an area where irrigation is not available long

before transplanting growing Azolla after transplanting rice is recommended

The use of phosphorus fertilizer is an important factor for good growth of

Further study is needed on ways to minimize phosphate fertilizerAzoZZa use while making Azolla use more efficient

Despite its promise growing Azolla in the tropics must overcome some

Azolla growth is depressed at adifficulties As shown in Table 3

temperature higher than 310 Experiences in Vietnam and China also indicate

AzolZas low tolerance for high temperature Azolla strains that are

tolerant of high temperature should be sought

Le Van Kan and Sobachkin (1963) reported from 6 to 29 increase in

rice yield where A pinnata was grown in combination with rice in

four fields in North Vietnam Talley et al (1977) reported that

A filicuioides and A mexicana grown with rice in California

increased rice yield 23 and 67 respectively

In contrast the IRRI trial of growing Azolla with rice gave less Our trialeffective results perhaps because of less mass of AzolZa

_yielded 14 t fresh weight of Azollaha (66 g dry weight of AzoZazm 2 )

g of Talley et al (1977) reported a maximum biomass

of 182 gm2

A filicuZoides dry weight

To overcome this shortcoming of our trials the selection of Azolla

clones more suitable to the tropics improvement of phosphorus

fertilizer application and the optimum kinds and amounts of other

nutrients will be studied


LITERATURE CITED

Ashton P J and R D Walmsley 1976 The aquatic fern Azolla and its Anabaen symiiont Endeavor 3539

Becking J H 1976 Contribution of plant algal associations In Proc of the lst International Symp on Nitrogen Fixation Vol 2 556 W E Newton and C J Nyman ed Washington State University Press

Le Van Kan and A A Sobachkin 1963 To the problem of the use of AzoZla as fertilizer in the Democratic Republic of Vietnam Timiryazev Moskwa Agricultural Academy 9493 (InRussian English translation in IRRI Library)

Moore A W 1959 Azolla biology and agronomic significance Bot Review 3517

Nutman P S 1976 IBP field experiments on nitrogen fixation by nodulated legumes In Symbiotic Nitrogen Fixation ed by P S Nutman Cambridge University Press 211 p

OeesA-1913 tber die Aqsimilation des Freien Stickstoff durch AzoZLa Zeitschr fur Botany 5145

Saubert G G P 1949 Provisional communication on the fixation of elementary nitrogen by a floating fern Annal of R6yal Botanical Garden Buitenzorg 51177

Talley S N B J Talley and R W Rains 1977 Nitrogen fixation by AzolZa in rice field Pages 259-281 in Genetic engineering for nitrogen fixation ed D A Hollaender et al Plenum Press New York

Tran Quang Thuyet and Dao The Tuan 1973 AzoiZa a green manure Agricultural Problems Vol 4 Vietnamese Studies No 38119

Tuzimura K 1950 The growth of leguminous crop and nitrogen fixing activity (InJapanese) J Sci Soil and Manure 21181

Tuzimura K N Ikeda and K Tukamoto 1957 AzoZla as a green manure in paddy field J Sci Soil and Manure 28275 (In Japanese English translation in IRRILibrary)

Yoshida S D A Forno and J H Cock 1971 Laboratory manual for physiological studies of rice IRRI p 61

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ABSTRACT






0 C (350 day





nitrogen







and phosphorus



























lopI



2 weeks





basis Assuming









nitrate














~b 340 + 102- N controls 368 + 147 446 + 014

-N-P 82+ 04 73+011 12+0 - N -K 116 10 109 T 10 830 120 - N -Ca 34 02 22 + 007 0 - N-g 30 3 + 08c 343 + 04 440 + 277 + N 125 +05- 193 + 10 289 + 75




















the control Azolla










the blue-green


































U b -6 - 261+ 11 262+ 02 260 + 26 213 + 02 2 - 235+ 02 4 -210+ 06- 3331+ 16 227 +06 226+-02-


0 84 +16 1 252-+16 270 +23 171 + 18 i50 +31 2 208+14 4 185 +_03 218+ 15 162 +09 154+ 06


0 77+8 1 605 +48 915+ 40 386 + 14 151 +4 1 1000 1 44t 4 895+100 945 + 23 821 + 183 259 + 17





2618 22- 88 + 0 276 + 03 320 + 042921 25 102 + 01 266 + 11 311 + 08 3224 28 108 + 02 273 T 08 308 T 18 3527 31 154 + 28 1949 + 08 146 +

































pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED















ABSTRACT






0 C (350 day





nitrogen







and phosphorus



























lopI



2 weeks





basis Assuming









nitrate














~b 340 + 102- N controls 368 + 147 446 + 014

-N-P 82+ 04 73+011 12+0 - N -K 116 10 109 T 10 830 120 - N -Ca 34 02 22 + 007 0 - N-g 30 3 + 08c 343 + 04 440 + 277 + N 125 +05- 193 + 10 289 + 75




















the control Azolla










the blue-green


































U b -6 - 261+ 11 262+ 02 260 + 26 213 + 02 2 - 235+ 02 4 -210+ 06- 3331+ 16 227 +06 226+-02-


0 84 +16 1 252-+16 270 +23 171 + 18 i50 +31 2 208+14 4 185 +_03 218+ 15 162 +09 154+ 06


0 77+8 1 605 +48 915+ 40 386 + 14 151 +4 1 1000 1 44t 4 895+100 945 + 23 821 + 183 259 + 17





2618 22- 88 + 0 276 + 03 320 + 042921 25 102 + 01 266 + 11 311 + 08 3224 28 108 + 02 273 T 08 308 T 18 3527 31 154 + 28 1949 + 08 146 +

































pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED































lopI



2 weeks





basis Assuming









nitrate














~b 340 + 102- N controls 368 + 147 446 + 014

-N-P 82+ 04 73+011 12+0 - N -K 116 10 109 T 10 830 120 - N -Ca 34 02 22 + 007 0 - N-g 30 3 + 08c 343 + 04 440 + 277 + N 125 +05- 193 + 10 289 + 75




















the control Azolla










the blue-green


































U b -6 - 261+ 11 262+ 02 260 + 26 213 + 02 2 - 235+ 02 4 -210+ 06- 3331+ 16 227 +06 226+-02-


0 84 +16 1 252-+16 270 +23 171 + 18 i50 +31 2 208+14 4 185 +_03 218+ 15 162 +09 154+ 06


0 77+8 1 605 +48 915+ 40 386 + 14 151 +4 1 1000 1 44t 4 895+100 945 + 23 821 + 183 259 + 17





2618 22- 88 + 0 276 + 03 320 + 042921 25 102 + 01 266 + 11 311 + 08 3224 28 108 + 02 273 T 08 308 T 18 3527 31 154 + 28 1949 + 08 146 +

































pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED














lopI



2 weeks





basis Assuming









nitrate














~b 340 + 102- N controls 368 + 147 446 + 014

-N-P 82+ 04 73+011 12+0 - N -K 116 10 109 T 10 830 120 - N -Ca 34 02 22 + 007 0 - N-g 30 3 + 08c 343 + 04 440 + 277 + N 125 +05- 193 + 10 289 + 75




















the control Azolla










the blue-green


































U b -6 - 261+ 11 262+ 02 260 + 26 213 + 02 2 - 235+ 02 4 -210+ 06- 3331+ 16 227 +06 226+-02-


0 84 +16 1 252-+16 270 +23 171 + 18 i50 +31 2 208+14 4 185 +_03 218+ 15 162 +09 154+ 06


0 77+8 1 605 +48 915+ 40 386 + 14 151 +4 1 1000 1 44t 4 895+100 945 + 23 821 + 183 259 + 17





2618 22- 88 + 0 276 + 03 320 + 042921 25 102 + 01 266 + 11 311 + 08 3224 28 108 + 02 273 T 08 308 T 18 3527 31 154 + 28 1949 + 08 146 +

































pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED


















~b 340 + 102- N controls 368 + 147 446 + 014

-N-P 82+ 04 73+011 12+0 - N -K 116 10 109 T 10 830 120 - N -Ca 34 02 22 + 007 0 - N-g 30 3 + 08c 343 + 04 440 + 277 + N 125 +05- 193 + 10 289 + 75




















the control Azolla










the blue-green


































U b -6 - 261+ 11 262+ 02 260 + 26 213 + 02 2 - 235+ 02 4 -210+ 06- 3331+ 16 227 +06 226+-02-


0 84 +16 1 252-+16 270 +23 171 + 18 i50 +31 2 208+14 4 185 +_03 218+ 15 162 +09 154+ 06


0 77+8 1 605 +48 915+ 40 386 + 14 151 +4 1 1000 1 44t 4 895+100 945 + 23 821 + 183 259 + 17





2618 22- 88 + 0 276 + 03 320 + 042921 25 102 + 01 266 + 11 311 + 08 3224 28 108 + 02 273 T 08 308 T 18 3527 31 154 + 28 1949 + 08 146 +

































pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED




















the blue-green


































U b -6 - 261+ 11 262+ 02 260 + 26 213 + 02 2 - 235+ 02 4 -210+ 06- 3331+ 16 227 +06 226+-02-


0 84 +16 1 252-+16 270 +23 171 + 18 i50 +31 2 208+14 4 185 +_03 218+ 15 162 +09 154+ 06


0 77+8 1 605 +48 915+ 40 386 + 14 151 +4 1 1000 1 44t 4 895+100 945 + 23 821 + 183 259 + 17





2618 22- 88 + 0 276 + 03 320 + 042921 25 102 + 01 266 + 11 311 + 08 3224 28 108 + 02 273 T 08 308 T 18 3527 31 154 + 28 1949 + 08 146 +

































pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED

















U b -6 - 261+ 11 262+ 02 260 + 26 213 + 02 2 - 235+ 02 4 -210+ 06- 3331+ 16 227 +06 226+-02-


0 84 +16 1 252-+16 270 +23 171 + 18 i50 +31 2 208+14 4 185 +_03 218+ 15 162 +09 154+ 06


0 77+8 1 605 +48 915+ 40 386 + 14 151 +4 1 1000 1 44t 4 895+100 945 + 23 821 + 183 259 + 17





2618 22- 88 + 0 276 + 03 320 + 042921 25 102 + 01 266 + 11 311 + 08 3224 28 108 + 02 273 T 08 308 T 18 3527 31 154 + 28 1949 + 08 146 +

































pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED












































pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED
















pg Ng soil

Dried AzoZZa

0 67 1 233 878 645 131 2 330 1286 956 195 3 z351 1465 1114 227 4 8118 3095 2277 465 6 9201 4596 3675 750

Fresh Azofli

0 81 2 906 2014 -1108 40o6 4 997 1 619 615 5921 66 1007 2707I 170-- 622

8 969 966 1997 731















































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED
























































(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED


















(kgha) 10300- 15200 13700 12800 4800


Increased amount

(kg dry wtha) 344 560 453 447 657

nitrogen


Nitrogen kg Nna



Increase (kg Nha) 453 634 r 818 1165






























- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED







































- - - 1020 d 1480 d 100

1540 ab 1850 c 125

- -- 1360 bc 2360 ab 100

- + 1320 bc 2160 abc 91

-+ - 1240 cd 1860 c 100

+ - + 1380 bc 2020ab 109

+ + 1310 bc 2250 ab 100 + + + 1640 a 2530 a 112









Ease of culture






































of 182 gm2







LITERATURE CITED

















Ease of culture






































of 182 gm2







LITERATURE CITED




































of 182 gm2







LITERATURE CITED














LITERATURE CITED













Documents

I AF25-0O00-GG50 1.5.JECT Food production and nutrition ...eprints.icrisat.ac.in/8591/1/RP_00998_Utilization_of_the..........pdf · Soil fertility, fertilizers, and plant nutrition--Tropics