Made on 1997.6.25, Revised on 2005.7.17 (Author _ Dr. Iwao WATANABE, former Soil Microbiologist, International Rice Research Institute)

Azolla (Japanese here)


Azolla News, reporting new development Click here

Scenery of using Azolla as rice greenmanure at northern Vietnam in 1979 Click here


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A.microphylla Incorporation
Left: Azolla microphylla
Right: A Philippino farmer, incorporating azolla


Azolla and its use



   CONTENTS

1.1 What is Azolla

Azolla is an aquatic fern (pteridophyte), floating on water surface of flooded rice fields, small ponds, and canals.Its size is 1-5 cm except for a giant A. nilotica, In Japan, two species A. japonica and A. imbricata are indigenous.
Generally it multiplies vegetatively, and often sexually.
Seven extant Azolla species are recognized (some taxonomists recognize 6 species), are distribution widely from temperate to tropical regions. Their features are shown in the following Tables


A. nilotica
A giant Azolla, indigenous in central Africa, elongating as long as 15 cm.

 A. japonica 
Azolla indigenous in Japan, close to A. filiculoides.

Hybrid Hybrid MI4030 and its parents
Left of the photo: Female parent   A. microphylla   (tolerant of high temperature)
Center: Hybrid (tolerant of high temperature), grow better than parents, and higher in N content
Right: Male parent A. filiculoides (tolerant of low temperature)


Azolla species
Features of Azolla species
(Section) SpeciesOriginMegasporocarpsMicrosporocarpsOthers
RhizospermaA. niloticaCentral-East Africa9-Float Small glochidia of MassulaeA set of 4 sporocarps
RhizospermaA. pinnataAsia, Oceania, Africa 9-FloatNo hook-like tip in glochidia of Massulaea pair of sporocarps
AzollaA. filiculoidesLatin America3-floats Hook-like tip in glochidia of massulae with 0~2 septaa pair of sporocarps
single cell leaf trichome
AzollaA. rubraOceania3-floatsHook-like tip in glochidia of massulae a pair of sporocarps
less pronounced leaf trichome
AzollaA. cristata Latin America3-floats Hook-like tip in glochidia of massulae with more than 2 septaa pair of sporocarps
double cell leaf trichome

Remarks 1) A. nilotica's chromosome number is 2n = 52 unlike others, 2n=44, a new name, Tetrasporocarpia nilotica was proposed.
A. rubra can be a subspecies of A. filiculoides
2) A. pinnata is divided to A.p. subsp. asiatica (Origin-Asia)、A.p.subsp. pinnata (Australia, and New Caledonia origin)、A.p.subsp. africana(Africa origin.
3) Previously described three "species" ;A. microphylla, A. mexicana, A. caroliniana are replaced by A. cristata(2005)
4) A. filiculoides has one-cell leaf surface trichomes、and two cells in A. cristata.
5) A. japonica indigenous in Japan may be A. filiculoides or closely related one.


Azolla is useful as a "soybean plant in rice field", because it can assimilate atmospheric nitrogen gas owing to the nitrogen fixation by cyanobacteria (blue green alga) living in the cavities located at the lower side of upper (dorsal) lobes of leaf.
Azolla can, therefore, grow on the water deficient in nitrogen compounds, and is high in nitrogen and protein. It fixes nitrogen as high as 3-5 kg N per ha per day under the optimum condition. In the tropics, annual nitrogen gain amounts to 500-1000 kg per ha, when grown throughout the year. It doubles its biomass within 2-3 days. The maximum biomass contains 30-80 ton fresh weight, 1.5-4.0 ton dry weight, and 50-150 kg N per ha.
Generally, Azolla multiplies vegetatively. Sexual multiplication is also observed. Under certain conditions (not known well), it forms sexual organs (sporocarps) at the backside of leaf.
Symbiotic cyanobacteria is transferred to the pedigrees through mega(female)sporocarps.

See details of biological nitrogen fixation here

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1.2 Conditions for growth

(Water)Azolla is sensitive to desiccation. When water depth over soil is a few cm, it grows well. It can grow adhering on moist soil. Therefore, it should be kept in a small pond during dry season or non-irrigated period.
(Wind) Wind pushes azolla to one side of a field plot, accumulating a dense mass, leading often to its death. Strong wind makes azolla fragmented, leading to poor growth or death.
Azolla growth (% per day) as affected by daily maximum temperature, FI:A. filiculoides, MI:A. microphylla, HYB: Hybrids of A. microphylla with A. filiculoides (Exp. in Mie, Japan)

(Temperature)Azolla multiplies at the daily mean temperature of 15-30oC. Optimum temperature is about 25oC. A. filiculoides, A. rubra and A. japonica require lower temperature than A. microphylla and A. pinnata do. Above 30-33oC or below 10oC, the growth was sharply retarded. Azolla can tolerate (survive) up to -5oC.

(Light) Under nutrient deficient and strong light conditions, Azolla becomes red. During hot summer or cold winter, it also turns red or brownish red. Under shaded conditions or nutrient-rich conditions, it remains green.
(Mineral nutrition)Azolla absorbs nutrient from water, when it is floating on water. Because phosphorus diffusion from soil to water is slow, field population of floating Azolla is generally deficient in phosphorus. The application of phosphorus fertilizer is effective to enhance its growth. Phosphorus fertilizer should be water soluble, and be applied on top of Azolla mat.
(pH)Azolla prefers slightly acidic media up to pH 4.
(Insect damage)Lepidoptera, Pyralidae larva are most harmful insect pests to azolla. In the tropics, Webworm -Elophyla and Case worm - Epheosispsis are major pests. At higher temperature around 30oC, the damage is severe, because azolla growth speed slows down while insect growth is active. Insect damage is one of the major problems in the tropics. In extreme cases, azolla mat in the field is eliminated within a few days by Pyralidae. Chironomus is also considered to give damage.

2 Use of azolla

2.1 History of azolla use

Until middle of the 1980's, azolla was used for greenmanure to spring rice in northern Vietnam. Phot shows farmers collecting azolla from inoculum producing pond. (1979, Jan)
Because of symbiotic nitrogen fixation, and consequently, high nitrogen content, Azolla has been used as a green manure for wetland rice in northern Vietnam, and central to southern China for centuries. In the early 1980's, the use in South Cotabato, Philippines had spread. However, since the late 1980's, the socio-economic situation in these countries has drastically changed with the introduction of market economy system. Increased use of chemical fertilizers, the search for a high income crop during azolla growth period, and the collapse of farmers' organizations who provided inoculum for rice cultivation are factors to explain rapid decline of azolla use.

2.2 New development in azolla use

In response to the decline of its use, multiple purposes of using azolla has been expored.
Multiple benefits of azolla are as follows. (Biological herbicide) Azolla covering water surface reduce light penetration to soil surface, resulting in the depression the germination of weeds. Thus, growth of azolla reduces occurrence aquatic weeds in flooded rice fields.
(Nutrient accumulation) Nutrients in floodwater cannot be directly absorbed by rice. Azolla can accumulate nutrients from floodwater, and provides these after azolla's decomposition. For example, one third of potassium absorbed by rice is once accumulated in azolla body. Under the mat of azolla, floodwater PH does not turn alkaline. Preventing alkaline reaction reduces ammonia loss.
(Animal feed) Azolla has been used as feed for pig, duck, and fish. It has high content of protein (20-30% on dry weight basis). Because its protein lacks in methionine and cystein, combination with cereals is needed. Nutritional values of azolla to animals vary greatly on azolla species. A. microphylla is the best, and palatability by fish is better than other species. On dry weight basis, azolla can be mixed up to 10% of the purchased animal feed.
(Is azolla weed?--supplemental comment) Azolla has been described as a weed to wetland rice. When floodwater level is high, and azolla is pushed into one side of a field, azolla often covers the top of young rice seedlings, leading to the depression of rice growth or death. Azolla cover reduces water temperature to 0.5-1℃. When azolla grows actively above 20 ℃, the reduction of water temperature may not be detrimental to rice growth. However,the surrounding farmers are often ignorant of beneficial effects of azolla and the way to control its growth. Distribution from the fields to the surounding fields should be minimized.
Many benefits of using azolla are recognized. As a result, the integrated use of azolla with rice and fish farming has been developed at Fujian Academy of Agricultural Sciences, China. The integrated approach can increase a farmer's income, while reducing the use of pesticides and fertilizers, and, consequently, environmental pollution.
Integrated use was also developed by a Filipino farmer, Mr. Fantilanan. He combined azolla culture with rice and vegetables culture, and rearing pigs and ducks. The excreta from pigs and ducks were introduced to a house scale biogas plant, And the effluent from biogas plant was returned to wetland rice fields with azolla.

2.3 Rice-duck-azolla system developed by a Japanese farmer

A Japanese farmer in Kyushu, Japan, Mr. T.Furuno has been practicing rice and hybrid duck culture. He tried hard not to use pesticide in rice cultivation. The most difficult task was weeding. He introduced hybrid duck primarily for weeding purpose. The duck effectively made a weeding job by disturbing soil surface. He found the duck contributed a lot to rice cultivation. Now, rice-duck culture is widely practiced in organic rice farming. Primarily for providing nitrogen nutrient, azolla was introduced to this system. Azolla provided nitrogen nutrient for rice and protein for duck, and contributed to the suppression of weed. Duck, on the other hand, contributed to azolla by eradicating azolla insect pest, and spreading azolla by its movement. Duck's excreta may supply phosphorus to azolla. This rice-duck-azolla system is now being adopted by organic farming farmers.

Azolla-effect

Multiple effects of Duck-Azolla-Rice Farming

Components Effects Artificials
Duck Disturbing, and Tillage Cultivator
Eating young weeds Herbicides
Making water turbid(Depressing ammonia loss?) Urease inhubitor
Eating rice pests(brownhoppers etc.) Insecticides
Eating azolla pests(Pyraids Insecticides
Providing nutrients from excreta(to rice and azolla) Fertilizers
Azolla Nitrogen fixation Nitrogen fertilizers
Feeds to duck Artificial feeds
Depressing weeds(Surface cover) Herbicides

3 Azolla Germplasm Collections

Duplicate living azolla germplasm collections are maintained at the International Rice Research Institute (IRRI), Philippines, and at Universite Catholique de Louvain, Belgium (UCL, c/o Prof. C. Van Hove). As of 1997, 562 accessions are maintained, covering all extant species collected worldwide in cooperation with many researchers. Details of the IRRI collection, including maintenance, are described in the following monograph.
Watanabe, I. Roger PA, Ladha J.K. And Van Hove C. (1992). Biofertilizer Germplasm Collection at IRRI, The International Rice Research Institute, Los Banos,Philippines, 66 pp.
The IRRI collection is now maintained as shoot-tip agar cultures, with transfer every 3-6 months. The breakdown of accessions in the IRRI collection is as follows:
The International Rice Research Institute Azolla germplasm Collection
speciesCode numbernumber
A. pinnata var. imbricataPI001-93
A. filiculoidesFI1001-135
A. mexicanaME2001-56
A. carolinianaCA3001-72
A. microphylla MI4001--129
A. niloticaNI5001-3
A. pinnata var. pinnataPP7001--53
Unknown8001-17
Groundtotal562


Remarks 1.Accessions categorized as A. pinnata var. pinnata include their subsp. pinnata and africana, and those as A. pinnata var. imbricata, although consisting mainly of the subsp. asiatica, also contain subsp. pinnata.
2. Code number starts from two Roman characters, identifying species, followed by numerals. 3. Taxonomic identity is based primarily on that given by the collectors; some of these identifications were later revised. Sexual hybrids are classified by their female parents.


This collection also includes unique material which cannot be obtained from natural habitats, e.g.sexual hybrids (79 accessions), Anabaena-free (20), azolla with heterologous symbiotic cyanobacteria (6), and putative mutants (16).
To avoid loss, most of section Azolla in IRRI collection are duplicated at the Azolla Research Center, Fujian Academy of Agricultural Science (Fuzhou, Fujian, China, c/o Liu C.C).
The IRRI and UCL accessions are open to distribution on request.
For IRRI collection, contact with T. Ventura ((E mail: Tventura@cgnet.com or J.K.Ladha@cgnet.com )

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4 Major references

Lumpkin T.A. and Plucknett D.L. (1982) Azolla as a Green Manure: Use and Management in Crop Production, Westview Tropical Agriculture Series, No.5. Westview Press Inc., Colorado, 230 pp.
Anonym. (1987) Azolla Utilization, IRRI, Philippines, 296pp
Watanabe, I. and Liu C.C. (1992) Improving nitrogen fixing systems and integrating them into sustainable rice farming. Plant Soil, 141, 57-67
Watanabe I (1994) Genetic Enhancement and Azolla Collection- Problems in Applying Azolla Anabaena Symbiosis.In: Nitrogen Fixation with Non-Legumes, (Eds. Hegazi N.A., Fayez, M., and Monib M.) ,The American University in Cairo Press, Cairo, PP. 437-450
Watanabe I. and Van Hove C. (1996) Phylogenetic, molecular, and breeding aspects of Azolla-Anabaena symbiosis.In: J.M. Camus, M.Gibby, and R.J.Jones Eds. Pteridology in Perspective pp.611-619, Royal Botanic Gardens, Kew
Wagner, G.M.(1997), Azolla: A review of its biology and utilization, Botanical Review, 63(1), 1-26.
Lejeune, A., A Cagauan, C Van Hove (1999) Azolla research and development: Recent trends and priorities: Symbiosis, 27(3-4): 333-351--NEWEST
(FAO Publication)
Anonym (1988) Bio and Organic Fertilizers: Prospects and Progress in Asia, RAPA publication Vol. 10, FAO, Regional Office for Asia and Pacific, Bangkok, 78 pp.
Anonym (1988) The" Rice-Azolla-Fish" System, RAPA publication Vol. 4, FAO, Regional Office for Asia and Pacific, Bangkok, 35 pp.
Van Hove C.(1989) Azolla and its multiple uses with emphasis on Africa. FAO,Rome, 52 pp. Cagauan A., and Pullin S.V. (1994) Azolla in Aquaculture:Past, present and future. In:Recent Advances in Aquaculture V. (Eds. Muir J.T., and Roberts R.J.) Blackwell Science, Cambridge, pp. 104-130


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About Cyanobacteria

Anabaena azollae
Symbiotic cyanobacteria inside the leaf cavities

Nitrogen fixing cyanobacteria


Cyanobacteria (blue green algae)are single or multi-cell photosynthetic organisms. They are often called "algae", but belong to "bacteria". Most of them can assimilate atmospheric nitrogen under reduced oxygen pressure. This reaction is called Nitrogen fixation. Some can do nitrogen fixation under ordinary oxygen pressure. and most of them have large cells, calledHeterocyst, specialized for nitrogen fixation. Heterocyst is indicated as an arrow in the above figure.
Cyanobacteria are either free-living or in symbiosiswith plants.
Symbiotic cyanobacteria provide ammonia to the host plants, and the frequency of heterocysts is higher than in free-living ones. Some plants among moss, ferns, and cycads have these symbiotic cyanobacteria. In angiosperms, only Gunnera are such plants.

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About Sexual multiplication


Megasporocarp of A. filiculoides attached with massulae.

LEFT:Croos section of megasporocarps. Cyanobacteria cells are present beneath the surface wall(called indusium)., and transfered to the newly emerging sporophytes.
Antherozoids (male sperms) are released from massulae and fertilize with oospore, and new sporophytes emerge from megasporocarps.


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Iwao Watanabe E mail: gan2@asahi.email.ne.jp