the Azolla Superorganism
The Azolla-Anabaena symbiosis has been called a superorganism that combines the individual talents of two very different organisms. The cyanobacterium Anabaena evolved during the early history of the Earth more than three billion years ago when the planet’s atmosphere was devoid of oxygen. The other organism is the fern Azolla.
Azolla’s floating leaves contain cavities filled with nitrogen that replicate the Earth’s ancient atmosphere. These provide a microenvironment for Anabaena which draws down up to 1000 kg of atmospheric nitrogen per acre per year. The nitrogen provides a natural fertilizer for Azolla’s growth, freeing the plant from its reliance on soil and enabling it to grow free floating on freshwater bodies.
The Azolla Superorganism: A unique biological system
In 2010 Francisco Carrapiço proposed that Azolla-Anabaena is a superorganism “because of its unique symbiosis in which the two partners have successful co-evolved into a system that makes important contributions to ecology, biofertilization and biotechnology” (Carrapiço, 2010). (A pdf of this publication (AzollaSuperorg_2010.pdf), and more than 30 of Dr Carrapiço’s other papers, can be can be downloaded for free from http://azolla.fc.ul.pt/publicacoes.html.)
The Azolla-Anabaenasuperorganism is unique. We know of no other symbiotic relationship in which a cyanobacterium and plant pass down together during reproduction from generation to generation. A few other plants have a symbiotic relationship with cyanobacteria, including some cycads and the anthophyte Gunnera, but the relationship has to be renewed each generation; it is broken after the plant dies and new cyanobacteria must re-colonize the plants in order to continue the relationship.
In contrast, Azolla and Anabaena are never apart; they have not been separated for almost a hundred million of years. They have evolved together continuously during this immense period of time as the Earth’s climate changed from a greenhouse world to the present phase of glacial-interglacial cycles.
Azolla’s morphology is unlike that of other ferns and, in particular, its leaf structure has evolved to provide an environment that is ideal for Anabaena. Azolla’s life cycle also makes it possible for Anabaena to pass uninterrupted from one generation of Azolla to the next. This has enabled the two organisms to evolve continuously together for more than 90 million years, a relationship unknown elsewhere on the planet.
Azolla’s leaves occur in two rows along each side of the plant’s stem. Each leaf has a very thin ventral lobe and a thick, greenish or reddish photosynthetic dorsal lobe containing a cavity that is the key to the Azolla and Anabaena symbiosis.
The cavity is a highly specialized structure that is formed during Azolla’s growth by part of the leaf epidermis folding inwards during Azolla’s development (Peters & Meeks, 1989). The cavity measures approximately 0.15 x 0.3 mm and opens to the external environment through a pore that is surrounded by two cell layers.
Azolla’s leaf cavities provide an oxygen-free home for Anabaena. Figure modified from Carrapiço (2001).
The centre of the cavity is filled with a gas or liquid and the bacteria are immobilized in the peripheral area of the cavity by a mucilaginous fibrillar network (Carrapiço 1991, 2002, 2010).
Azolla’s leaf cavities provide an ideal micro-environment for the heterocyst-forming nitrogen-fixing filamentous bacterium Anabaena azollae.
This is the key to Azolla’s ability to sequester enormous amounts of atmospheric CO2.
Azolla’s life-cycle
Unlike plant-cyanobacterial symbioses in vascular plants, the Azolla-Anabaena symbiotic system is sustained throughout the fern’s life cycle, where the cyanobacterium and bacteria are always present (Carrapiço, 2010), either in the dorsal lobe leaf cavities or in the sexual structures (sporocarps) (Carrapiço, 1991, 2002). The Azolla plants are never infected de novo, since Anabaena is transferred between generations as akinete inocula (Carrapiço, 2010). This maintains continuity of the symbiosis during sexual reproduction, summarized in Peters & Meeks (1989).
Anabaena is transmitted directly to successive generations via the Azolla’s reproductive spores. Figure modified from Carrapiço (2001).
Other bacterial symbionts
Azolla’s leaf cavity provides an ideal micro-environment for a bacterial community that includes various strains of the genera Anthrobacter, Corynobacterium and Agrobacterium plus the heterocyst-forming nitrogen-fixing filamentous bacterium Anabaena azollae.
Other bacteria are present in Azolla’s leaf cavities and include Arthrobacter which commonly occurs in soils. Like Anabaena, these bacteria also occur throughout Azolla’s life cycle and have a developmental pattern that is identical to that of Anabaena (Forni et al., 1990; Van Hove, 1989; Carrapiço, 1991). Their rolein the symbiosis is not yet fully understood, but the present data indicate that Anabaena is the only bacterial symbiont of Azolla that fixes nitrogen.
The free-living species Arthrobacter chlorophenolicus is capable of degrading high concentrations of 4-chlorophenol, indicating its potential use for bioremediation (Westerberg et al., 2000).
Marriage Between A Fern & Cyanobacterium
Source Of Prokaryotic Cyanobacteria
Introduction
See Desert Varnish & Lichen Crust On Boulders |
| Ponds along the San Dieguito River (San Diego County, California) are covered with a reddish carpet of Azolla filiculoides during the fall months. Photo also shows clump of cattails (Typha latifolia) and naturalized Australian Eucalyptus camaldulensis. The Eucalyptus trees were introduced into California near the turn of the century for fast-growing hardwood lumber for railroad ties, but proved inadequate because the spikes would not hold in the badly checked wood. Now these trees have literally taken over parts of San Diego County. |
| A pond along the San Dieguito River (San Diego County, California) covered with a reddish carpet of Azolla filiculoides. The bright green areas are masses of the filamentous green algae, Mougeotia and Spirogyra. |
It has been estimated that there are at least 10,000 different species of ferns in the world, from large tree ferns of tropical rain forests to small rock ferns of desert canyons and alpine crevices. Fossil evidence indicates that many additional species of ferns flourished on earth during the Carboniferous period, some 300 million years ago. But of all the great diversity of ferns, relatively few kinds have colonized the water. Azolla belongs to the Salvinia Family (Salviniaceae), although some authorities now place it in the monotypic family, Azollaceae. Six species are distributed worldwide, three of which occur in the United States: A. filiculoides, A. mexicana and A. caroliniana.
Individual Azolla plants have slender, branched stems with minute, overlapping scalelike leaves only one millimeter long. Each plant resembles a little floating moss with slender, pendulous roots on its underside. The plants tend to clump together and often form compact mats on the water surface. Azolla is sometimes called "duckweed fern" and commonly grows with one or more species of duckweeds (Lemnaceae), including Lemna, Spirodela, Wolffia and Wolffiella. When growing in full sunlight, particularly in late summer and fall, Azolla may produce reddish anthocyanin in the leaves, in contrast with the bright green carpets of duckweed and filamentous green algae.
| Several water fern plants (Azolla filiculoides) floating on the water surface. The plant in upper right with oval fronds is a duckweed (Lemna minuta). The minute plants which resemble tiny green bubbles are Wolffia borealis and another interesting species W. columbiana, two of the world's smallest flowering plants. |
A related African water fern (Salvinia rotundifolia), also listed as S. auriculata in some floras, is mentioned in the Guinness Book of World Records (1985 UK Edition) as the "most intransigent weed." This mat-forming aquatic fern was detected on the filling of Kariba Lake in May 1959. Within 11 months it covered an area of 77 square miles (199 km2), and by 1963 it covered 387 square miles (1002 km2).
| The water fern (Salvinia rotundifolia), a ubiquitous floating fern in quiet waters of streams and ponds throughout tropical America, Africa and Florida. The small duckweed in photo is Landoltia punctata. |
Sexual Reproduction In Azolla
| Close-up view of Azolla filiculoides showing scalelike, overlapping leaves and several globose reproductive structures called sporocarps. The male sporocarp (middle right) contains microsporangia that resemble eggs inside an egg sac. One sporocarp has broken open releasing many spore clusters or massulae. The minute ovoid plants in center are Wolffia borealis, a minute flowering seed plant. |
| Closeup view of Azolla filiculoides showing a small female sporocarp flanked by two larger, globose male sporocarps. These tiny structures are so small that they could easily fit on the head of an ordinary straight pin. |
| Highly magnified view (400X) of one spore mass (massula) of Azolla filiculoides with unique barbed projections called glochidia. In a related species of water fern (A. mexicana), the glochidia are septate with several distinct partitions. |
According to some references, the universal occurrence of Anabaena azollae inside the leaves of Azolla suggests that reproduction of this water fern may be chiefly vegetative; however, it has been shown that this cyanobacterium and fern may develop in synchrony. Short filaments of Anabaena, called hormogonia, often survive under the "indusium cap," on top of the germinating megaspore. Hormogonia may be entrapped by the embryo Azolla plant during differentiation of the shoot apex and dorsal lobe primordia of the first leaves. It is fascinating to speculate on just how and when these two diverse organisms formed such an intimate association. Although filamentous cyanobacteria (with cells resembling heterocysts) date back more than two billion years, fossil Azolla plants are known only from late Cretaceous deposits less than 80 million years ago.
Anabaena and Nitrogen Fixation
| Modern-day filamentous cyanobacteria (Anabaena azollae) from cavities within the leaves of the ubiquitous water fern (Azolla filiculoides). The larger, oval cells are heterocysts (red arrow), the site of nitrogen-fixation where atmospheric nitrogen (N2) is converted into ammonia (NH3). Polar nodules are visible in some of the heterocysts. The water fern benefits from its bacterial partner by an "in house" supply of usable nitrogen. The cellular structure of these bacteria has changed very little in the past one billion years. |
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Note: Here is a more accurate update for the above equation: N2 + 8 H+ + 8e- +16 ATP + 16 H2O = 2 NH3 + H2 + 16 ADP +16 Pi
The fowing explanation is from Jim Deacon of the Institute of Cell and Molecular Biology, The University of Edinburgh.Two molecules of ammonia are produced from one molecule of nitrogen gas. The reaction requires 16 molecules of ATP and a supply of electrons and protons (hydrogen ions) plus the enzyme nitrogenase. Nitrogenase consists of two proteins, an iron protein and a molybdenum-iron protein. The reaction occurs while N2 is bound to the nitogenase enzyme complex. The Fe protein is first reduced by electrons donated by ferredoxin. Then the reduced Fe protein binds ATP and reduces the molybdenum-iron protein, which donates electrons to N2, producing HN=NH. In two futher cycles of this process (each requiring electrons donated by ferredoxin) HN=NH is reduced to H2N-NH2, and this in turn is reduced to 2 NH3. Depending on the type of microorganism, the reduced ferredoxin which supplies electrons for this process in generated by photosynthesis, respiration or fermentation.
Recent studies have shown that the actual site of nitrogen fixation occurs within the thick-walled heterocysts. As the heterocyst matures, the photosynthetic membranes (thylakoid membranes) become contorted or reticulate compared to regular photosynthetic cells of Anabaena, and they become non-photosynthetic (and do not produce oxygen). This fact is especially noteworthy because nitrogen fixation requires the essential enzyme nitrogenase, and the activity of nitrogenase is greatly inhibited by the presence of oxygen.
Azolla and Rice Productivity
In addition to nitrogen fixation, Azolla has a number of other uses. Apparently fish and shrimp relish the Azolla. In fact, Azolla was grown for fish food and water purification at the Biospere II project in Arizona (a 2.5 acre glass enclosure simulating an outer space greenhouse). Fresh Azolla and duckweed (Wolffia) can also be used in salads and sandwiches, just as alfalfa and bean sprouts are used. Dried, powdered Wolffia and Azolla make a nutritious, high protein powder similar to the popular alga (cyanobacterium) Spirulina that is sold in natural food stores. Azolla has also proved useful in the biological control of mosquitos. The mosquito larvae are unable to come up for air because of the dense layer of Azolla on the water surface. Azolla grows very quickly in ponds and buckets, and in makes an excellent fertilizer (green manure) and garden mulch.
| A male tree frog (Hyla regilla) floating in a pond of Azolla filiculoides. On a warm summer night, the amazing chorus of dozens of these small frogs can be almost deafening. |
References
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