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Symbioses between aquatic invertebrates and algae
SYMBIOSES
BETWEEN
AQUATIC INVERTEBRATES
AND ALGAE
ROSALIND HINDE School of Biological Sciences, University of Sydney, N.S.W. 2006, Australia. INTRODUCTION Many aquatic invertebrates have symbiotic relationships with algae. Although these relationships are usually considered to be mutualistic symbioses, 1 will use the term symbiosis in its original, broadest and most neutral sense (“living together of differently named organisms”; de Bary, 1879, cited by Goff, 1983) throughout this paper. After outlining the nature of some of the symbioses which have been studied in reasonable detail, and which are generally treated as typical of such relationships, I will discuss their apparent place in the spectrum of symbiotic associations. Since algal symbioses have been the subject of a great deal of research and of a number of reviews in the last few years I will simply attempt to introduce the field, particularly those areas which are important in assessing the degree of mutualism involved, referring interested readers to other reviews and to the more recent research papers for more detail. CHARACTERISTICS
OF ALGAL/INVERTEBRATE
SYMBIOSES
The Organisms
Both the animals and the algae found in these associations are very diverse indeed. Smith, Muscatine & Lewis (1969), Taylor (1974), Trench (1979) and Pardy (1983) have listed many of the 200-odd genera of aquatic invertebrates which are known to harbour symbiotic algae, and the examples below are described in their papers, unless otherwise noted. Many protozoa, sponges, Cnidaria of all classes, turbellarian Platyhelminthes, molluscs and ascidians are associated with algae. The associated algae include both prokaryotic and eukaryotic plants. Blue-green algae (cyanobacteria) are found as symbionts in some protozoa and in many species of marine sponges, while the recently discovered Prochloron spp. (prokaryotic algae containing chlorophylls a and b) are associated with ascidians. The major eukaryotic algal groups involved in symbioses with animals include green algae (members of the divisions Chlorophyta and Prasinophyta), dinoflagellates (Pyrrophyta), diatoms (Bacillariophyta) and Cryptophyta. Unicellular green algae (Chlorella spp.) are found in freshwater protozoa, sponges and Cnidaria (eg. Hydra spp.). The marine turbellarian Convoluta roscoffensis is associated with the unicellular prasinophyte Platymonas convolutae. Symbiotic dinoflagellates, commonly referred to as zooxanthellae, occur in reef-building corals and in many other cnidarians, including soft corals, gorgonians, anemones and jellyfish, as well as in giant clams, nudibranchs (e.g. Rudman, 1982), sponges (Sara & Liaci, 1964), Foraminifera (Lee & McEnery, 1983) and Radiolaria (Anderson, 1983). For a long time the dinoflagellates found in all these hosts have been treated as belonging to one species, Symbiodinium microadriaticum Freudenthal. However, Blank & Trench (1985) have recently showwn that zooxanthellae from different hosts differ in many important characteristics, including chromosome number, and have therefore suggested that many of the so-called “strains” of zooxanthellae are separate species. Other types of dinoflagellate (Amphidinium spp.) are also found in associations with protozoa, cnidarians and platyhelminths. Rarer algal symbionts include diatoms, which are found in foraminiferans (Lee & McEnery, 1983), radiolarians (Anderson, 1983), sponges (Cox & Larkum, 1983), and flatworms, and cryptomonads, found in protozoa (Taylor, Blackbourn & Blackbourn, 1971) and sponges (Duclaux, 1973). Macroscopic, multicellular algae are occasionally found in symbioses, particularly in sponges (e.g. Price, Fricker & Wilkinson, 1984). Some species of sea slug retain functional chloroplasts from the macroscopic algae on which they feed.
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Transmission
of Algal
Symbionts
Where the animal host can reproduce asexually (e.g. by division in protozoans or budding in Cnidaria) algae are incorporated directly into the daughter individuals. If the symbionts of the parent are genetically homogenous, then the products of asexual reproduction will be identical with the parents, since both the animal and plant components will be clones of their parents. Trench (1981) has suggested that infection with a single alga may be enough to re-establish the symbiosis in a sexually produced individual, so that in some cases all the symbionts of an individual animal may be genetically identical. When an animal host reproduces sexually, algae may be passed on via the egg cytoplasm, but this appears to be rare. It has been shown to occur in the marine hydroid Myrionemu umboinense (Trench, 1981). Much more commonly, the eggs themselves do not contain the symbiotic algae, and it has been argued that the larval or juvenile stages must, therefore, pick up suitable algae from their environment. Interestingly, both Con\~o/utu roscoffensis and Cussiopeiu xumuchunu fall into this category, in spite of the vital roles of the algae in their life cycles (see below). However, evidence that the symbionts are carried in the egg cases or capsules of most hosts (including C’. rosaoflensis) is starting to accumulate (see Smith, 1981). The free-living stages of “S. microudridicum” and symbiotic Chlorella have not yet been cultured from, or recognized in, natural communities, so it is difficult to say whether they can live in the phytoplankton at all. Although the need for them may be rare under natural conditions, there may be specializations which promote the infection of the next generation of the animal with algae. For example, Douglas & Gooday (1982) found that free-living algae of the genera Plutgmonus and Tetruselmis (both of which can establish symbioses with C. roscogensis) stopped swimming when they encountered egg capsules of these worms, and subsequently adhered to the capsules. On the other hand they could not find evidence of chemical attraction of the algae to the egg capsules, or of extrusion of algae on to the capsules during the laying of the eggs, although both of these had previously been suggested as mechanisms to promote reinfection (see Douglas & Gooday, 1982). Cussiopeiu xumuchanu becomes infected with zooxanthellae during the scyphistoma stage, presumably by free-living algae of appropriate strains (Trench, Colley & Fitt, 1981); in laboratory experiments Cu. xumuchanu (and other hosts of zooxanthellae) discriminate between various strains of zooxanthellae, favouring those isolated from members of their own species (Trench et al., 1981). Little is known of the transmission of algal symbionts in marine sponges. Some ascidian species release larvae which carry frochloron in specialised sacs (Eldredge, 1965, cited by Pardy, 1983). Obligate
and Fucultutive
Associations
Some of the symbioses between algae and invertebrates which have been studied in detail are definitely obligate for the animal partner. For example, Convolutu roscoffensis will not reach sexual maturity if it does not become infected with a suitable symbiont (see Holligan & Gooday, 1975). The scyphistoma of Cassiopeia xumuchunu cannot strobilate (produce adult medusae) unless it contains zooxanthellae (Trench et al., 1981). In other associations the situation is not so clear. Although in many cases it is possible to produce symbiont-free (aposymbiotic) animals in the laboratory, Pardy (1983) has pointed out that it is doubtful whether aposymbiotic members of these species occur naturally. Thus in most of the associations between algae and invertebrates the animals probably require the presence of the algae for long-term survival and normal growth. There are, however, a few species in which symbiotic and aposymbiotic animals occur naturally, and in the same habitat. For example, the temperate coral Astrungiu dunae grows well whether or not it has zooxanthellae, but cannot survive by photosynthesis alone if denied particulate food; colonies with algae probably fare better when food is scarce (SzmantFroelich & Pilson, 1980). Tropical corals often lose their zooxanthellae when subjected to stresses such as increased temperature or decreased salinity. However, these “bleached” corals suffer from increased mortality while lacking symbionts, and the survivors become populated with zooxanthellae again, rather than becoming permanently aposymbiotic (Harriott, 1985). It is hard to determine whether these associations are obligate for the algae. It is not certain which symbionts are passed on directly during sexual reproduction, and which are acquired, by larvae or juveniles, from the environment (see above). Those acquired from the environment may be taken up as free-living algae, or as undigested cells in zooplankton eaten by the host or in pellets of algae extruded by mature host animals (Trench et ul., 1981), so even evidence of this mode of transmission does not offer proof that the algae can exist as stable free-living populations. There are some associations between flagellate protozoans and blue-green algae in which the algae are so heavily modified that they have been regarded as chloroplasts by some authors (e.g. Cyunophoru puradoxu; see Smith, 1979). These associations may be obligate for both partners; however, in such cases the free-living, aposymbiotic flagellates and blue-green algae might differ so much from the symbiotic forms as to be unrecognizable except by biochemical, immunological or genetic tests.
Algal invertebrate symbioses
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Morph~lo~~~ai Aspects
The algal symbionts of invertebrates may be extracellular or intracellular in their hosts. The algae may show extreme morphological adaptations to symbiosis or be little different from, or even identical with, their free-living or cultured counterparts. Neither the degree of morphological adaptation of the algae nor their position in the host (within or between its cells) is correlated with the degree of interdependence between the partners. Piatymonas convolutae in Convofuta rosco&nsis differ greatly from free-living ones. In symbiosis the algae lose their flagella, eyespots and pectin cell covering (theta) and become irregular in shape (and thus have a greater surface area than the free-living form) (Douglas, 1983; Trench, 1979). The blue-green algae found as intracellular symbionts in the flagellate Cyanophora have much thinner cell walls than do free-living blue-green algae (Trench, 1979). Free-living dinoflagellates have a complex cell covering, the amphiesma, consisting of a number of layers of simple membranes and at least one layer of vesicles. The amphiesmas of zooxanthellae in their hosts are always thinner, with fewer layers of membrane, than those of free-living dinoflagellates; the amphiesmas of cultured zooxanthellae are similar to those of free-living dinoflagellates (Trench et al., 1981). Similarly, zooxanthellae in sitar lack flagella and the characteristic surface grooves (girdle and sulcus) of dinoflagellates, but in culture they alternate between a non-motile coccoid phase and a motile phase similar in form to free-living dinoflagellates (Trench et at., 1981). All the various types of algae which inhabit foraminiferans (Lee & McEnery, 1983) and the dinoffagellate symbionts of radiolarians (Anderson, 1983) lose their flagella and have modified cell walls. It has generally been supposed that the thinning of the cell walls of algae when they are in symbiosis allows improved flow of inorganic nutrients and organic metabolites between the symbionts. In contrast, the symbiotic Chlorella of Hydru and other freshwater invertebrates do not seem to change morphologically in any way when they are in symbiosis. There is no loss of cell wall material. Vegetative stages of all Chlorella spp. are non-motile, so there is no loss of motility associated with becoming symbiotic. There may be changes in the internal structure as well as in surface structures of algae when they become symbiotic in animals. For example blue-green algae in the sponge Dysidea herbacea contain distinctive, complex inclusions (the stellar bodies) of unknown function (Berthold, Borowitzka & Mackay, 1982). In invertebrate hosts the symbiotic algae are taken up via the digestive system (Smith, 1981). Since in most of the hosts of such algae digestion is partly or wholly intracellular, entry through the digestive system implies phagocytosis of the algae by the digestive cells. The algae may then remain restricted to cells normally involved in digestion, or they may migrate from the digestive system to other tissues or sites. In C. roscoflensis the algae are usually described as lying between the cells of the animal, rather than within them; however there is some evidence that they are actually enclosed in vacuoles within the cells of the worms (see Douglas, 1983; Trench, 1979). Most of the algae are found in the epidermal and sub-epidermal layers of C. rosco$ensis (Holligan & Gooday, 1975). Symbiotic dinoflagellates occur inside the cells of most of their hosts. In Cnidaria they are usually found only in the endoderm, but in a few species they may also occur in the mesogloea and ectoderm. In the giant clam, Triducnu sp., they are extracellular, being found mainly in the haemal sinuses (Trench, 1979). In nudibranchs, the zooxanthellae are usually inside digestive gland cells, but they may migrate into connective tissue cells or be intercellular in other tissues (Rudman, 1982). Where the symbiotic algae inhabit the cells of their animal hosts, they are normally found in a vacuole surrounded by membrane of animal origin. Since they are taken up by phagocytosis, this membrane is presumably derived from the membrane of the phagocytic vacuole. It may help to control the movement of metabolites between the symbionts. In contrast, most of the retained chloroplasts of sacoglossan molluscs are free in the cytoplasm (see Hinde, 1983a), and thus their relationship to the cytoplasm is the same as it was in the plant. Specificity
There is evidence that associations between algae and invertebrates are specific. Although in many cases aposymbiotic animals can be infected with algae from other hosts, these artificial symbioses are not usually fully “effective”. For example, algae not closely related to Plutymonus convolutae may enter, but do not persist in, Convolutu rosco$ensis; algae closely related to P. convolutae do persist and support growth to varying degrees, but they will be replaced by P. convolutae if the worms are exposed to it (Douglas, 1983; Provasoli, Yamasu & Manton, 1968). Similarly, various strains of Chforellu can infect Paramecium bursuria. Those isolated from Pm. bursuria form effective, stable symbioses with a range of strains of the ciliate, but infections with free-living strains of Chforelfa do not support good growth, and are less stable (~~akashian, 1975). Some hosts will expel “foreign” strains of algae after a period; in Hydra viridis this may occur within 1 to 2 days of infection (Muscatine, Cook, Pardy & Fool, 197.5). The means by which specificity is monitored and maintained are not fully understood in any one
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symbiosis, but there has been-research on this aspect of a number of associations. The secretion of organic metabolites by symbionts does not seem to be an important trigger for their recognition by the host (Smith, 1981). Surface charge and the ability to bind lectins affect the likelihood that algae will be phagocytosed, but many non-symbiotic organisms and particles can be phagocytosed by host cells, so these characteristics are not sufficient to guarantee specificity (Smith, 1981). Trench et al. (1981) found that both surface properties of the algae and the occurrence of translocation of organic compounds from the algae increased the rate of infection of Cassiopeia xamachana by zooxanthellae isolated from this host, but pointed out that these stimulating factors were unlikely to be important in the sea, as they both depended on the presence of host-derived material around the freshly isolated algae. Finally, it should be remembered that the need for infection by truly free-living algae is probably rare in natural populations (Smith, 1981) (see above), so that recognition mechanisms may not be very important most of the time. Because of this Smith (1981) has suggested that mechanisms promoting specificity may be more important in preventing unsuitable organisms from entering than in promoting infection with the “correct” symbiont. Resistance of algae to host defences
For a symbiont to survive in its host, it must be able to resist the defensive mechanisms of that host; as algal symbionts enter via the digestive system, they must also be resistant to the host’s digestive enzymes. Many of the invertebrates which have symbiotic algae are carnivores, and may lack the enzymes which would enable them to break down the cell walls of algae (e.g. all cnidarians; nudibranchs). Other hosts can digest algae, but do not digest their symbionts or algae of related strains. For example, if Paramecium bursaria is exposed to a mixture of live and dead symbiotic algae, both are phagocytosed. The living algae are not digested but the dead ones are, unless they happen to be in a phagocytic vacuole which also contains live symbiotic algae. In this case, none of the contents of the vacuole are digested (Karakashian, 1975). In both Pm. bursaria (Karakashian, 1975) and hydra (Pool, 1981) the live algae appear to inhibit fusion of lysosomes with the phagocytic vacuoles which contain them. There are a number of pathogenic organisms which can also inhibit lysosomeiphagosome fusion (Moulder, 1985). In sea slugs which retain functional chloroplasts the slugs apparently digest the cytoplasm of the algae, and all the organelles apart from the chloroplasts, most of which are eventually released from vacuoles into the animals’ cytoplasm (see above). This may protect them from attack by lysosomes, which must fuse with a vacuole to release their enzymes. Certainly the slugs’ cells can digest chloroplasts, as they do when the animals are starved: the remains of digested chloroplasts are always found in vacuoles (Hinde, 1983a). There have been no detailed studies of the defensive mechanisms of invertebrates in relation to their potential effects on algal (or bacterial) symbionts. Metabolic and Other Interactions in Algal/Invertebrate Organic Metabolites In all the associations
Symbioses
which have been investigated, the algae photosynthesize at normal rates, and fix COZ into organic compounds by normal pathways. If the association is exposed to CO:! labelled with carbon-14, labelled products of photosynthesis can be detected in animal cells soon after the start of photosynthesis. For example, a large proportion of the products of photosynthesis can be recovered from animal tissue within 1 hour of the start of photosynthesis in various associations between cnidarians and zooxanthellae; the rate of translocation is usually 20-40% of the total fixed carbon during the first hour or so after fixation (Smith, 1974), but values between !2% and 82% have been recorded (see Hinde, 1983b). However, the work of Muscatine and other (see Muscatine, Falkowski, Porter & Dubinsky, 1984) suggests that, in corals at least, a much higher proportion of the products of photosynthesis eventually finds its way into the animal (more than 95% of total fixed carbon in Sty/ophora pistillata-Muscatine et a[., 1984). Two types of translocation have been observed in associations between corals and zooxanthellae. Both lipids (mainly triglycerides) and low molecular weight water soluble compounds pass from the algae to the animals (Battey & Patton, 1984); the latter make up that 2060% of total fix carbon which moves from algae to animal in the first hour or so after fixation (Smith, 1974; Hinde, 1983b). In these associations the major water-soluble compound translocated from the algae to the host is glycerol, with small amounts of glucose, amino acids and organic acids (see Hinde, 1983; Smith, 1974). Similar specific translocation of small molecules has been observed in all investigations of symbioses between algae and invertebrates, and in each case the rates of translocation have fallen in the same range as in corals (i.e. 20 to 60% in the first hour). For example, the symbiotic Chlorella of Hydra and Paramecium translocate maltose (with small amounts of other compounds) (Smith, 1974). In Convoluta rosco$ensis the major mobile compounds are amino acids, mostly alanine, but short-term release rates are lower than in other associations (Smith, 1974). In Elysia viridis, which retains chloroplasts from Codium fragile, glucose and glycollic acid are translocated (Hinde, 1983b).
Alga/ invertebrate symbioses
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In spite of the similarities in rates of transfer of low molecular weight metabolites between alga and animal in a broad range of symbioses, the mechanisms by which translocation is stimulated are probably diverse. In many hosts with zooxanthellae (e.g. hard corals, anemones) the animal tissue contains one or more substances (“host factors”) which stimulate the flow of glycerol (and the other compounds which move in the intact association) from isolated zooxanthellae. The isolated zooxanthellae do not “leak” products of photosynthesis unless homogenized, alga-free host tissue is added to the suspension, when they translocate the same photosynthetic products into the water, at about the same rates, as they do in the intact association (see review by Hinde, 1983b). There is some disagreement in the literature about the control of translocation in Hydra. While host homogenates seem to be effective in Hydra, translocation by the isolated algae is also stimulated by low pH, being fastest at pH 4.0-5.0 (Hinde, 1983b). In C. roscofensis translocation is slightly stimulated by host homogenates (Hinde, 1983b). Since in these associations the rates of photosynthesis (per mg chlorophyll) are similar to those of other microscopic algae, and large proportions of the organic compounds produced are lost to the host animals, it follows that the growth rates of the symbiotic algae must be lower than those of otherwise similar free-living algae. Indeed, the measured growth rates of zooxanthellae are relatively low (Muscatine et al., 1984) except when the host’s tissues are being populated rapidly (personal observations). There has been very little work on the movement of organic compounds from animals to symbiotic algae, although it has been shown that this does occur in several associations (Trench, 1979). lnorgank nutrients Plants require sources of combined nitrogen, phosphorus and other elements; these are usually taken up as inorganic compounds, although many microalgae can also use organic sources of nitrogen and phosphorus. In the oceans, nitrogen is the nutrient which is most likely to limit plant growth; in fresh waters phosphorus is most likely to be limiting. It has been proposed that one of the advantages of the symbiotic condition for the algae is an abundant supply of nutrients, compared to that available to free-living microalgae in the same waters. There are two possible ways in which this supply might be ensured. Firstly, the animals absorb inorganic nutrients which then diffuse to the algae (Muscatine, 1980). However, where there is a large algal population, the total surface area of the algae must be much greater than that of the animal which harbours them. Although the animals can draw a current of water over their surfaces, it seems unlikely that this would make up for the relatively small surface area available for absorption of nutrients. Thus living in the animals probably provides no net advantage to the algae in the uptake of inorganic nutrients. Secondly, the algae are known to take up and use various excretory products of their hosts. For example, corals can use ammonia and amino acids (Muscatine, 1980) and C. roscoflensis uses uric acid produced by its host (Holligan & Gooday, 1975). These nutrients are used to synthesize organic compounds, some of which will eventually be translocated to, and catabolized by, the host. This internal recycling of nutrients between the partners may be highly efficient; for example, aposymbiotic corals excrete more nitrogen compounds than do corals with zooxanthellae (Muscatine, 1980; Szmant-Froelich & Pilson, 1984). Such recycling obviously decreases the requirment of the association as a whole for nutrients from external sources. Enhancement of Calcification In the true corals (Scleractinia) the calcium carbonate skeleton is laid down much faster in the light than in the dark (Taylor, 1983). Experiments using inhibitors of photosynthesis show that it is the occurrence of photosynthesis, not some other effect of light, which causes this enhancement of calcification. The relationship between the two processes is not yet understood (Taylor, 1983). Shading There is evidence that the thick layer of algae found in many species of sponge provides shade for the relatively delicate underlying cells of the animal. Sponges which live in shallow waters tend either to be heavily pigmented or to have a thick layer of symbiotic algae (Wilkinson, 1980). Other interactions Except for studies of nutrient supply (see above) there has been almost no work on the possible benefits of these symbioses to the algae. Many species of microalgae, including most dinoflagellates and some green algae, show absolute requirements for various vitamins (particularly B-group vitamins) when they are grown in culture, and the growth of other species is improved by addition of vitamins (Provasoli & Carlucci, 1974). The animal hosts may supply appropriate vitamins to their symbionts. Similarly, many microscopic algae can use organic compounds to support heterotrophic growth; the possible advantages to the algae of a steady supply of such compounds from the host have never been investigated. Another possible benefit to the algae is protection from herbivores. Planktonic algae are often heavily grazed by zooplankton, which would not attack host animals of the symbiotic algae. These host animals do have their own predators, and since there have been no relevant studies, it is not possible to compare the relative risks of being eaten in the plankton or in a host. However, since most of the hosts’ predators are probably specialized carnivores, in many cases the symbiotic algae would pass through the gut of the
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predator unharmed, and be released into the water column (where they would, of course, be exposed to the herbivores). Finally, it is probably advantageous to the algae to be supported in a matrix (the host animal) which remains in a constant position in the water column; microscopic algae normally have to expend energy to remain buoyant and thus stay within the photic zone (i.e. that part of the water column where there is enough light to support plant growth). SIGNIFICANCE
AND NATURE OF ALGAL/INVERTEBRATE
SYMBIOSES
As noted in the Introduction, symbioses between algae and invertebrates are usually regarded as mutualistic. Yet, as can be seen from the outline above, almost all research has centred on the advantages of the associations to the larger partner, the animal. This has been, at least in part, due to the fact that it is relatively easy to follow the fate of products of photosynthesis, using ‘*CO2as a tracer, and relatively difficult to divise means of investigating the transfer of metabolites (particularly organic compounds) from the animals to the algae. The animals do indeed seem to benefit from these associations. In every case studied they gain products of photosynthesis, usually in large quantities. For instance, Muscatine et al. (1984) found that when the coral Stylophoru pistillata grows in well-lit habitats, it receives almost one-and-a-half times the amount of organic carbon it requires for respiration-i.e. the algae supply enough substrates to provide for all respiratory needs and to contribute to the growth of the animal. In shade-adapted S. pisti/[utu translocated products of the algae are sufficient for 58% of daily respiratory needs (Muscatine et al., 1984). Few species have been studied in as much detail as S. pistihtu, but there is no reason to suppose that it is unique. There may also be other benefits to the animal host: for example the algae may provide essential compounds not available in its diet (amino acids, fatty acids, etc.); they may provide oxygen and remove COZ during photosynthesis; they may remove toxic metabolic wastes; they may enhance calcification (where there is a calcareous skeleton) and protect animal cells from excess light or ultraviolet radiation. There has been little or no investigation of these or other potential benefits to the animals. The costs of symbiosis to the animals have not been investigated either. It is likely that there is some cost-i.e. some energy expended-in maintaining the symbionts, but its nature and extent are unknown. However, it is probable that in most of the symbioses between invertebrates and algae, the benefits to the animals are large, and outweigh the costs. The success of these associations, which dominate the highly productive communities of coral reefs and are found in many other habitats, and which add greatly to the total productivity of these communities, is evidence of the advantages to the animals of maintaining photosynthetic symbionts. Difficult as it is to fully interpret the balance of benefits and costs to the animals, there is even less to go on in trying to decide whether the algae benefit from the associations, or whether they are harmed. The only reasonable way of measuring this seems to be to compare the Darwinian fitness of the algae (i.e. their success in passing their genes on to the next generation) in symbiosis and when free-living. It is clear that the growth rates of symbiotic algae are very low. For example, Muscatine et al. (1984) calculate mean doubling times of 53 and 74 days for zooxanthellae in light- and shade-adapted S. pistillutu respectively. Free-living microalgae commonly have doubling times of 1 to 2 days. However, free-living algae are heavily grazed in most waters, and dense populations are rare and short-lived. As pointed out above, the free-living algae also face other hazards, such as sinking below the photic zone or being limited by lack of inorganic nutrients or vitamins. For symbiotic algae these problems may be prevented or minimized by the contributions of the host. Of course, it is not yet known whether most of the symbiotic strains of algae are capable of surviving outside their hosts. Until we know this, and can compare the growth rates and survival of free-living and symbiotic algae, it is not possible to define their symbioses with invertebrates as beneficial or harmful to the algae. The algae do not appear to be able to escape actively from the animals’ cells. It seems very likely that they are parasitized by the animals rather than being mutualistic symbionts. Many host species appear to regulate the numbers of their symbiotic algae (Smith, 1981). The extensive removal of photosynthetic products by the animal host might be enough to ensure that the animal controls the growth and reproduction of the algae in some associations. Animal hosts may also control the supply of nutrients to the algae. The “host factors” which promote translocation of photosynthetic products may not be the only specific compounds produced by the animals which can control their symbionts; algal growth and reproduction may also be controlled by the animal by means of specific regulators. Until all these aspects of the physiology of algal/invertebrate associations, and of the algae in their independent form (where it exists) are understood, it is not possible to make a useful assessment of the place of any of these associations in the spectrum of symbioses.
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SUMMARY The characteristics of symbiotic associations between algae and invertebrates are outlined, using a number of the better-studied examples. The common assumption that these symbioses are mutualistic ones is discussed. In the present state of knowledge, it is impossible to determine the nature of these associations, but the evidence favours the view that the animal hosts benefit markedly from the presence of the algae, but that the possibility that the algae may be harmed (i.e. parasitized) by the animals cannot be dismissed. Some general lines of investigation which would help to define these relationships are suggested. REFERENCES ANDERSONO.R. 1983. The radiolarian symbiosis. In: Algal Symbiosis+ continuum of interaction strategies pp 6939 (Edited by Goff L.J.) Cambridge University Press, Cambridge and New York. BAITEY J.F. & PATTON J.S. 1984. A re-evaluation of the role of glycerol in carbon translocation in zooxanthellaecoelenterate symbiosis. Marine Biology 79: 27-38. BERTHOLD R.J., BOROWITZKAM.A. & MACKAY M.A. 1982. The ultrastructure of Oscillatoria spongeliue, the blue-green algal endosymbiont of the sponge Dysidea herbaceu. Phycologiu 21: 327-335. BLANK R.J. & TRENCHR.K. 1985. Speciation and symbiotic dinoflagellates. Science 229: 656458. Cox G. & LARKUM A.W.D. 1983. A diatom apparently living in symbiosis with a sponge. Bulletin of Marine Science 33: 943-94s. DOUGLAS A.E. 1983. Establishment of the symbiosis in Convoluta roscofensis. Journal of the Murine Biological Association of the U.K. 63: 419-434. DOUGLASA.E. & GOODAYG.W. 1982. The behaviour of algal cells towards egg capsules of Con,,oluta roscoflensisand its role in the persistence of the Convoluta-alga symbiosis. British Phyco/ogical Journull7: 383-388. DUCLALJXG. 1973. Presence de cryptophycCes symbiontiques dans trois Cponges des &es franqaises. Bulletin de lu Sock% de Phycologie de la Frunce 18: 54-57. GOFF L.J. 1983. Introduction. In: Algul Symbiosis--a continuum of interaction strutegies pp l-4 (Edited by Goff L.J.) Cambridge University Press, Cambridge and New York. HARRIOTTV.J. 1985. Mortality rates of icleractinian corals before and during a mass bleaching event. Marine Ecology Progress Series 21: 81-88. HINDE R. 1983a. Retention of algal chloroplasts by molluscs. In: A/gal Symbiosis--a continuum of interacfion strutegies pp 97-107 (Edited by Goff L.J.) Cambridge University Press, Cambridge and New York. HINDE R. 1983b. Host release factors in symbioses between algae and invertebrates. In: Endocytobio/ogy 11. Intracellular Space us Oligogenetic System pp 709-726 (Edited by Schenk H. & Schwemmler W.) Walter de Gryter, Berlin & New York. HOLLIGANP.M. & GOODAYG.W. 1975. Symbiosis in Convolutu roscoffensis. In: Symbiosis, Symposium of the Society for Experimental Biology 29: 205-227 (Edited by Jennings D.H. & Lee D.L.) CambrIdge University Press, Cambridge. KARAKASHIANM.W. 1975. Symbiosis in Paramecium bursaria. In: Symbiosis, Symposium of the Society for Experimental Biology 29: 145-173 (Edited by Jennings D.H. & Lee D.L.) Cambridge University Press, Cambridge. LEE J.J. & MCENERYM.E. 1983. Symbiosis in foraminifera. In: Algal Symbiosis4 continuum of interaction strategies pp 3768 (Edited by Goff L.J.) Cambridge University Press, Cambridge and New York. MOULDERJ.W. 1985. Comparative biology of intracellular parasitism. Microbiologicul Reviebcs 49: 298-337. MUSCATINE L. 1980. Uptake, retention, and release of dissolved inorganic nutrients by marine algae-invertebrate associations. In: Cellular Interactions in Symbiosis und Parasitism pp 229-244 (Edited by Cook C.B., Pappas P.W. & Rudolph E.D.) Ohio State University Press, Columbus. MUSCATIN; L., COOK C.B.. PARDY R.-L. & POOL R.R. 1975. Uptake, recognition and maintenance of symbiotic Chlorellu by Hydru viridis. In: Symbiosis, Symposium of the Society for E.uperimenta/ Biology 29: 175-203 (Edited by Jennings D.H. & Lee D.L.) Cambridge University Press, Cambridge. MUSCA~INEL., FALKOWSKIP.G., PORTERJ.W. & DUBINSKYZ. 1984. Fate of photosynthetic fixed carbon in light- and shade-adapted colonies of the symbiotic coral Stylophorcl pistillata. Proceedings I of_ the Royal Society_ of. London, series B, i22: 181-202. PARDY R.L. 1983. Phycozoans, phycozoology, phycozoologists? In: Algal Symbiosis--a continuum of interuction strugeties pp 5-17 (Edited by Goff L.J.) Cambridge University Press, Cambridge and New York. POOLR.R. 1981. The establishment of the Hydra-Chlorella symbiosis: recognition of potential algal symbionts. Berichte Deutsches Botanisches Gesellscahft 94: 565-569. PRICE I.R., FRICKLERR.L. & WILKINSONC.R. 1984. Cerutodictyon spongiosum (Rhodophyta), the macroalgal partner in an algae-sponge symbiosis, grown in unialgal culture. Journal of Phycology 20: 156-158. PROVASOLIL., YAMASUT. & MANTONI. 1968. Experiments on the resynthesis of symbiosis in Convoluta roscofensis with different flagellate cultures. Journal of the Marine Biological Association of the U.K. 48: 465-479. PROVASOLIL. & CARLUCCI A.F. 1974. Vitamins and growth regulators. In: Algal Physiology and Biochemistry PP 741-787 (Edited bv Stewart W.D.P.) Blackwell Scientific Publications. Oxford. R;;MAN W.B. 1982. The taxonomy and biology of further aeolidacean and arminacean nudibranch molluscs with symbiotic zooxanthellae. Zoologicul Journal of the Linnean Society 74: 147-196.
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S~ti M. & LIACI L. 1964. Symbiotic association between zooxanthellae and two marine sponges of the genus C/ions. Nature London 203: 321. SMITH D.C. 1974. Transport from symbiotic algae and symbiotic chloroplasts to host cells. Symposia ofthe Societyfor Experimental Biology 28: 485-520. SMITH D.C. 1979. From extracellular to intracellular: the establishment of a symbiosis. Proceedings of the Royal Society of London, series B, 204: 115-130. SMITH D.C. (1981). The role of nutrient exchange in recognition between symbionts. Berichte Deutsches Botanisches Gesellschaft 94: 517-528. SMITH D.C., MLJSCATINEL. & LEWIS D. 1969. Carbohydrate movement from autotrophs to heterotrophs in parasitic and mutalistic symbiosis. Biological Reviews 44: 17-90. SZMANT-FROELICHA. & PILSONM.E.Q. 1980. The effects of feeding frequency and symbiosis with zooxanthellae on the biochemical composition of Astrangia danae Milne Edwards and Haime 1849. Journal of Experimental Marine Biology and Ecology 48: 85-97. SZMANT-FROELICHA. & PILSON M.E.Q. 1984. Effects of feeding frequency and symbiosis with zooxanthellae on nitrogen metabolism and respiration of the coral Astragia danae. Marine Biology 81: 153-162. TAYLOR D.L. 1974. Symbiotic marine algae: taxonomy and biological fitness. In: Symbiosis in the Sea pp 245-262 (Edited by Vernberg, W.B.) University of,South Carolina Press, Columbia, SC. TAYLOR D.L. 1983. The coral-algal symbiosis. In: Atgal Symbiosis--a continuum of interacton strategies pp 19-35 (Edited by Goff L.J.) Cambridge University Press, New York. TAYLOR F.J.R., BLACKBOURND.J. & BLACKBOURNJ. 1971. The red water ciliate Mesodinium rubrum and its “incomplete symbionts”: a review including new ultrastructural observations. Journal o.f the Fisheries Reseurch Board of Canada 28: 391-407. TRENCHR.K. 1979. The cell biology of plant-animal symbiosis. Annual Review of Plant Physiology 30: 485-531. TRENCHR.K. 1981. Cellular and molecular interactions in symbioses between dinoflagellates and marine invertebrates. Pure and Applied Chemistry 53: 819835. TRENCH R.K., COLLEYN.J. & FITT W.K. 1981. Recognition phenomena in symbioses between marine invertebrates uptake, sequestration and persistence. Berichte Deutsches Botanisches Gesellschaft and “zooxanthellae”: 94: 529-545. WILKINSON C.R. 1980. Cyanobacteria symbiotic in marine sponges In: Endocytobiology: endosymbiosis and cell biology pp 553-563 (Edited by Schwemmler, W. & Schenk, H.) Walter de Gruyter, Berlin & New York.
BETWEEN
AQUATIC INVERTEBRATES
AND ALGAE
ROSALIND HINDE School of Biological Sciences, University of Sydney, N.S.W. 2006, Australia. INTRODUCTION Many aquatic invertebrates have symbiotic relationships with algae. Although these relationships are usually considered to be mutualistic symbioses, 1 will use the term symbiosis in its original, broadest and most neutral sense (“living together of differently named organisms”; de Bary, 1879, cited by Goff, 1983) throughout this paper. After outlining the nature of some of the symbioses which have been studied in reasonable detail, and which are generally treated as typical of such relationships, I will discuss their apparent place in the spectrum of symbiotic associations. Since algal symbioses have been the subject of a great deal of research and of a number of reviews in the last few years I will simply attempt to introduce the field, particularly those areas which are important in assessing the degree of mutualism involved, referring interested readers to other reviews and to the more recent research papers for more detail. CHARACTERISTICS
OF ALGAL/INVERTEBRATE
SYMBIOSES
The Organisms
Both the animals and the algae found in these associations are very diverse indeed. Smith, Muscatine & Lewis (1969), Taylor (1974), Trench (1979) and Pardy (1983) have listed many of the 200-odd genera of aquatic invertebrates which are known to harbour symbiotic algae, and the examples below are described in their papers, unless otherwise noted. Many protozoa, sponges, Cnidaria of all classes, turbellarian Platyhelminthes, molluscs and ascidians are associated with algae. The associated algae include both prokaryotic and eukaryotic plants. Blue-green algae (cyanobacteria) are found as symbionts in some protozoa and in many species of marine sponges, while the recently discovered Prochloron spp. (prokaryotic algae containing chlorophylls a and b) are associated with ascidians. The major eukaryotic algal groups involved in symbioses with animals include green algae (members of the divisions Chlorophyta and Prasinophyta), dinoflagellates (Pyrrophyta), diatoms (Bacillariophyta) and Cryptophyta. Unicellular green algae (Chlorella spp.) are found in freshwater protozoa, sponges and Cnidaria (eg. Hydra spp.). The marine turbellarian Convoluta roscoffensis is associated with the unicellular prasinophyte Platymonas convolutae. Symbiotic dinoflagellates, commonly referred to as zooxanthellae, occur in reef-building corals and in many other cnidarians, including soft corals, gorgonians, anemones and jellyfish, as well as in giant clams, nudibranchs (e.g. Rudman, 1982), sponges (Sara & Liaci, 1964), Foraminifera (Lee & McEnery, 1983) and Radiolaria (Anderson, 1983). For a long time the dinoflagellates found in all these hosts have been treated as belonging to one species, Symbiodinium microadriaticum Freudenthal. However, Blank & Trench (1985) have recently showwn that zooxanthellae from different hosts differ in many important characteristics, including chromosome number, and have therefore suggested that many of the so-called “strains” of zooxanthellae are separate species. Other types of dinoflagellate (Amphidinium spp.) are also found in associations with protozoa, cnidarians and platyhelminths. Rarer algal symbionts include diatoms, which are found in foraminiferans (Lee & McEnery, 1983), radiolarians (Anderson, 1983), sponges (Cox & Larkum, 1983), and flatworms, and cryptomonads, found in protozoa (Taylor, Blackbourn & Blackbourn, 1971) and sponges (Duclaux, 1973). Macroscopic, multicellular algae are occasionally found in symbioses, particularly in sponges (e.g. Price, Fricker & Wilkinson, 1984). Some species of sea slug retain functional chloroplasts from the macroscopic algae on which they feed.
383
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Transmission
of Algal
Symbionts
Where the animal host can reproduce asexually (e.g. by division in protozoans or budding in Cnidaria) algae are incorporated directly into the daughter individuals. If the symbionts of the parent are genetically homogenous, then the products of asexual reproduction will be identical with the parents, since both the animal and plant components will be clones of their parents. Trench (1981) has suggested that infection with a single alga may be enough to re-establish the symbiosis in a sexually produced individual, so that in some cases all the symbionts of an individual animal may be genetically identical. When an animal host reproduces sexually, algae may be passed on via the egg cytoplasm, but this appears to be rare. It has been shown to occur in the marine hydroid Myrionemu umboinense (Trench, 1981). Much more commonly, the eggs themselves do not contain the symbiotic algae, and it has been argued that the larval or juvenile stages must, therefore, pick up suitable algae from their environment. Interestingly, both Con\~o/utu roscoffensis and Cussiopeiu xumuchunu fall into this category, in spite of the vital roles of the algae in their life cycles (see below). However, evidence that the symbionts are carried in the egg cases or capsules of most hosts (including C’. rosaoflensis) is starting to accumulate (see Smith, 1981). The free-living stages of “S. microudridicum” and symbiotic Chlorella have not yet been cultured from, or recognized in, natural communities, so it is difficult to say whether they can live in the phytoplankton at all. Although the need for them may be rare under natural conditions, there may be specializations which promote the infection of the next generation of the animal with algae. For example, Douglas & Gooday (1982) found that free-living algae of the genera Plutgmonus and Tetruselmis (both of which can establish symbioses with C. roscogensis) stopped swimming when they encountered egg capsules of these worms, and subsequently adhered to the capsules. On the other hand they could not find evidence of chemical attraction of the algae to the egg capsules, or of extrusion of algae on to the capsules during the laying of the eggs, although both of these had previously been suggested as mechanisms to promote reinfection (see Douglas & Gooday, 1982). Cussiopeiu xumuchanu becomes infected with zooxanthellae during the scyphistoma stage, presumably by free-living algae of appropriate strains (Trench, Colley & Fitt, 1981); in laboratory experiments Cu. xumuchanu (and other hosts of zooxanthellae) discriminate between various strains of zooxanthellae, favouring those isolated from members of their own species (Trench et al., 1981). Little is known of the transmission of algal symbionts in marine sponges. Some ascidian species release larvae which carry frochloron in specialised sacs (Eldredge, 1965, cited by Pardy, 1983). Obligate
and Fucultutive
Associations
Some of the symbioses between algae and invertebrates which have been studied in detail are definitely obligate for the animal partner. For example, Convolutu roscoffensis will not reach sexual maturity if it does not become infected with a suitable symbiont (see Holligan & Gooday, 1975). The scyphistoma of Cassiopeia xumuchunu cannot strobilate (produce adult medusae) unless it contains zooxanthellae (Trench et al., 1981). In other associations the situation is not so clear. Although in many cases it is possible to produce symbiont-free (aposymbiotic) animals in the laboratory, Pardy (1983) has pointed out that it is doubtful whether aposymbiotic members of these species occur naturally. Thus in most of the associations between algae and invertebrates the animals probably require the presence of the algae for long-term survival and normal growth. There are, however, a few species in which symbiotic and aposymbiotic animals occur naturally, and in the same habitat. For example, the temperate coral Astrungiu dunae grows well whether or not it has zooxanthellae, but cannot survive by photosynthesis alone if denied particulate food; colonies with algae probably fare better when food is scarce (SzmantFroelich & Pilson, 1980). Tropical corals often lose their zooxanthellae when subjected to stresses such as increased temperature or decreased salinity. However, these “bleached” corals suffer from increased mortality while lacking symbionts, and the survivors become populated with zooxanthellae again, rather than becoming permanently aposymbiotic (Harriott, 1985). It is hard to determine whether these associations are obligate for the algae. It is not certain which symbionts are passed on directly during sexual reproduction, and which are acquired, by larvae or juveniles, from the environment (see above). Those acquired from the environment may be taken up as free-living algae, or as undigested cells in zooplankton eaten by the host or in pellets of algae extruded by mature host animals (Trench et ul., 1981), so even evidence of this mode of transmission does not offer proof that the algae can exist as stable free-living populations. There are some associations between flagellate protozoans and blue-green algae in which the algae are so heavily modified that they have been regarded as chloroplasts by some authors (e.g. Cyunophoru puradoxu; see Smith, 1979). These associations may be obligate for both partners; however, in such cases the free-living, aposymbiotic flagellates and blue-green algae might differ so much from the symbiotic forms as to be unrecognizable except by biochemical, immunological or genetic tests.
Algal invertebrate symbioses
385
Morph~lo~~~ai Aspects
The algal symbionts of invertebrates may be extracellular or intracellular in their hosts. The algae may show extreme morphological adaptations to symbiosis or be little different from, or even identical with, their free-living or cultured counterparts. Neither the degree of morphological adaptation of the algae nor their position in the host (within or between its cells) is correlated with the degree of interdependence between the partners. Piatymonas convolutae in Convofuta rosco&nsis differ greatly from free-living ones. In symbiosis the algae lose their flagella, eyespots and pectin cell covering (theta) and become irregular in shape (and thus have a greater surface area than the free-living form) (Douglas, 1983; Trench, 1979). The blue-green algae found as intracellular symbionts in the flagellate Cyanophora have much thinner cell walls than do free-living blue-green algae (Trench, 1979). Free-living dinoflagellates have a complex cell covering, the amphiesma, consisting of a number of layers of simple membranes and at least one layer of vesicles. The amphiesmas of zooxanthellae in their hosts are always thinner, with fewer layers of membrane, than those of free-living dinoflagellates; the amphiesmas of cultured zooxanthellae are similar to those of free-living dinoflagellates (Trench et al., 1981). Similarly, zooxanthellae in sitar lack flagella and the characteristic surface grooves (girdle and sulcus) of dinoflagellates, but in culture they alternate between a non-motile coccoid phase and a motile phase similar in form to free-living dinoflagellates (Trench et at., 1981). All the various types of algae which inhabit foraminiferans (Lee & McEnery, 1983) and the dinoffagellate symbionts of radiolarians (Anderson, 1983) lose their flagella and have modified cell walls. It has generally been supposed that the thinning of the cell walls of algae when they are in symbiosis allows improved flow of inorganic nutrients and organic metabolites between the symbionts. In contrast, the symbiotic Chlorella of Hydru and other freshwater invertebrates do not seem to change morphologically in any way when they are in symbiosis. There is no loss of cell wall material. Vegetative stages of all Chlorella spp. are non-motile, so there is no loss of motility associated with becoming symbiotic. There may be changes in the internal structure as well as in surface structures of algae when they become symbiotic in animals. For example blue-green algae in the sponge Dysidea herbacea contain distinctive, complex inclusions (the stellar bodies) of unknown function (Berthold, Borowitzka & Mackay, 1982). In invertebrate hosts the symbiotic algae are taken up via the digestive system (Smith, 1981). Since in most of the hosts of such algae digestion is partly or wholly intracellular, entry through the digestive system implies phagocytosis of the algae by the digestive cells. The algae may then remain restricted to cells normally involved in digestion, or they may migrate from the digestive system to other tissues or sites. In C. roscoflensis the algae are usually described as lying between the cells of the animal, rather than within them; however there is some evidence that they are actually enclosed in vacuoles within the cells of the worms (see Douglas, 1983; Trench, 1979). Most of the algae are found in the epidermal and sub-epidermal layers of C. rosco$ensis (Holligan & Gooday, 1975). Symbiotic dinoflagellates occur inside the cells of most of their hosts. In Cnidaria they are usually found only in the endoderm, but in a few species they may also occur in the mesogloea and ectoderm. In the giant clam, Triducnu sp., they are extracellular, being found mainly in the haemal sinuses (Trench, 1979). In nudibranchs, the zooxanthellae are usually inside digestive gland cells, but they may migrate into connective tissue cells or be intercellular in other tissues (Rudman, 1982). Where the symbiotic algae inhabit the cells of their animal hosts, they are normally found in a vacuole surrounded by membrane of animal origin. Since they are taken up by phagocytosis, this membrane is presumably derived from the membrane of the phagocytic vacuole. It may help to control the movement of metabolites between the symbionts. In contrast, most of the retained chloroplasts of sacoglossan molluscs are free in the cytoplasm (see Hinde, 1983a), and thus their relationship to the cytoplasm is the same as it was in the plant. Specificity
There is evidence that associations between algae and invertebrates are specific. Although in many cases aposymbiotic animals can be infected with algae from other hosts, these artificial symbioses are not usually fully “effective”. For example, algae not closely related to Plutymonus convolutae may enter, but do not persist in, Convolutu rosco$ensis; algae closely related to P. convolutae do persist and support growth to varying degrees, but they will be replaced by P. convolutae if the worms are exposed to it (Douglas, 1983; Provasoli, Yamasu & Manton, 1968). Similarly, various strains of Chforellu can infect Paramecium bursuria. Those isolated from Pm. bursuria form effective, stable symbioses with a range of strains of the ciliate, but infections with free-living strains of Chforelfa do not support good growth, and are less stable (~~akashian, 1975). Some hosts will expel “foreign” strains of algae after a period; in Hydra viridis this may occur within 1 to 2 days of infection (Muscatine, Cook, Pardy & Fool, 197.5). The means by which specificity is monitored and maintained are not fully understood in any one
386
R. Hinde
symbiosis, but there has been-research on this aspect of a number of associations. The secretion of organic metabolites by symbionts does not seem to be an important trigger for their recognition by the host (Smith, 1981). Surface charge and the ability to bind lectins affect the likelihood that algae will be phagocytosed, but many non-symbiotic organisms and particles can be phagocytosed by host cells, so these characteristics are not sufficient to guarantee specificity (Smith, 1981). Trench et al. (1981) found that both surface properties of the algae and the occurrence of translocation of organic compounds from the algae increased the rate of infection of Cassiopeia xamachana by zooxanthellae isolated from this host, but pointed out that these stimulating factors were unlikely to be important in the sea, as they both depended on the presence of host-derived material around the freshly isolated algae. Finally, it should be remembered that the need for infection by truly free-living algae is probably rare in natural populations (Smith, 1981) (see above), so that recognition mechanisms may not be very important most of the time. Because of this Smith (1981) has suggested that mechanisms promoting specificity may be more important in preventing unsuitable organisms from entering than in promoting infection with the “correct” symbiont. Resistance of algae to host defences
For a symbiont to survive in its host, it must be able to resist the defensive mechanisms of that host; as algal symbionts enter via the digestive system, they must also be resistant to the host’s digestive enzymes. Many of the invertebrates which have symbiotic algae are carnivores, and may lack the enzymes which would enable them to break down the cell walls of algae (e.g. all cnidarians; nudibranchs). Other hosts can digest algae, but do not digest their symbionts or algae of related strains. For example, if Paramecium bursaria is exposed to a mixture of live and dead symbiotic algae, both are phagocytosed. The living algae are not digested but the dead ones are, unless they happen to be in a phagocytic vacuole which also contains live symbiotic algae. In this case, none of the contents of the vacuole are digested (Karakashian, 1975). In both Pm. bursaria (Karakashian, 1975) and hydra (Pool, 1981) the live algae appear to inhibit fusion of lysosomes with the phagocytic vacuoles which contain them. There are a number of pathogenic organisms which can also inhibit lysosomeiphagosome fusion (Moulder, 1985). In sea slugs which retain functional chloroplasts the slugs apparently digest the cytoplasm of the algae, and all the organelles apart from the chloroplasts, most of which are eventually released from vacuoles into the animals’ cytoplasm (see above). This may protect them from attack by lysosomes, which must fuse with a vacuole to release their enzymes. Certainly the slugs’ cells can digest chloroplasts, as they do when the animals are starved: the remains of digested chloroplasts are always found in vacuoles (Hinde, 1983a). There have been no detailed studies of the defensive mechanisms of invertebrates in relation to their potential effects on algal (or bacterial) symbionts. Metabolic and Other Interactions in Algal/Invertebrate Organic Metabolites In all the associations
Symbioses
which have been investigated, the algae photosynthesize at normal rates, and fix COZ into organic compounds by normal pathways. If the association is exposed to CO:! labelled with carbon-14, labelled products of photosynthesis can be detected in animal cells soon after the start of photosynthesis. For example, a large proportion of the products of photosynthesis can be recovered from animal tissue within 1 hour of the start of photosynthesis in various associations between cnidarians and zooxanthellae; the rate of translocation is usually 20-40% of the total fixed carbon during the first hour or so after fixation (Smith, 1974), but values between !2% and 82% have been recorded (see Hinde, 1983b). However, the work of Muscatine and other (see Muscatine, Falkowski, Porter & Dubinsky, 1984) suggests that, in corals at least, a much higher proportion of the products of photosynthesis eventually finds its way into the animal (more than 95% of total fixed carbon in Sty/ophora pistillata-Muscatine et a[., 1984). Two types of translocation have been observed in associations between corals and zooxanthellae. Both lipids (mainly triglycerides) and low molecular weight water soluble compounds pass from the algae to the animals (Battey & Patton, 1984); the latter make up that 2060% of total fix carbon which moves from algae to animal in the first hour or so after fixation (Smith, 1974; Hinde, 1983b). In these associations the major water-soluble compound translocated from the algae to the host is glycerol, with small amounts of glucose, amino acids and organic acids (see Hinde, 1983; Smith, 1974). Similar specific translocation of small molecules has been observed in all investigations of symbioses between algae and invertebrates, and in each case the rates of translocation have fallen in the same range as in corals (i.e. 20 to 60% in the first hour). For example, the symbiotic Chlorella of Hydra and Paramecium translocate maltose (with small amounts of other compounds) (Smith, 1974). In Convoluta rosco$ensis the major mobile compounds are amino acids, mostly alanine, but short-term release rates are lower than in other associations (Smith, 1974). In Elysia viridis, which retains chloroplasts from Codium fragile, glucose and glycollic acid are translocated (Hinde, 1983b).
Alga/ invertebrate symbioses
387
In spite of the similarities in rates of transfer of low molecular weight metabolites between alga and animal in a broad range of symbioses, the mechanisms by which translocation is stimulated are probably diverse. In many hosts with zooxanthellae (e.g. hard corals, anemones) the animal tissue contains one or more substances (“host factors”) which stimulate the flow of glycerol (and the other compounds which move in the intact association) from isolated zooxanthellae. The isolated zooxanthellae do not “leak” products of photosynthesis unless homogenized, alga-free host tissue is added to the suspension, when they translocate the same photosynthetic products into the water, at about the same rates, as they do in the intact association (see review by Hinde, 1983b). There is some disagreement in the literature about the control of translocation in Hydra. While host homogenates seem to be effective in Hydra, translocation by the isolated algae is also stimulated by low pH, being fastest at pH 4.0-5.0 (Hinde, 1983b). In C. roscofensis translocation is slightly stimulated by host homogenates (Hinde, 1983b). Since in these associations the rates of photosynthesis (per mg chlorophyll) are similar to those of other microscopic algae, and large proportions of the organic compounds produced are lost to the host animals, it follows that the growth rates of the symbiotic algae must be lower than those of otherwise similar free-living algae. Indeed, the measured growth rates of zooxanthellae are relatively low (Muscatine et al., 1984) except when the host’s tissues are being populated rapidly (personal observations). There has been very little work on the movement of organic compounds from animals to symbiotic algae, although it has been shown that this does occur in several associations (Trench, 1979). lnorgank nutrients Plants require sources of combined nitrogen, phosphorus and other elements; these are usually taken up as inorganic compounds, although many microalgae can also use organic sources of nitrogen and phosphorus. In the oceans, nitrogen is the nutrient which is most likely to limit plant growth; in fresh waters phosphorus is most likely to be limiting. It has been proposed that one of the advantages of the symbiotic condition for the algae is an abundant supply of nutrients, compared to that available to free-living microalgae in the same waters. There are two possible ways in which this supply might be ensured. Firstly, the animals absorb inorganic nutrients which then diffuse to the algae (Muscatine, 1980). However, where there is a large algal population, the total surface area of the algae must be much greater than that of the animal which harbours them. Although the animals can draw a current of water over their surfaces, it seems unlikely that this would make up for the relatively small surface area available for absorption of nutrients. Thus living in the animals probably provides no net advantage to the algae in the uptake of inorganic nutrients. Secondly, the algae are known to take up and use various excretory products of their hosts. For example, corals can use ammonia and amino acids (Muscatine, 1980) and C. roscoflensis uses uric acid produced by its host (Holligan & Gooday, 1975). These nutrients are used to synthesize organic compounds, some of which will eventually be translocated to, and catabolized by, the host. This internal recycling of nutrients between the partners may be highly efficient; for example, aposymbiotic corals excrete more nitrogen compounds than do corals with zooxanthellae (Muscatine, 1980; Szmant-Froelich & Pilson, 1984). Such recycling obviously decreases the requirment of the association as a whole for nutrients from external sources. Enhancement of Calcification In the true corals (Scleractinia) the calcium carbonate skeleton is laid down much faster in the light than in the dark (Taylor, 1983). Experiments using inhibitors of photosynthesis show that it is the occurrence of photosynthesis, not some other effect of light, which causes this enhancement of calcification. The relationship between the two processes is not yet understood (Taylor, 1983). Shading There is evidence that the thick layer of algae found in many species of sponge provides shade for the relatively delicate underlying cells of the animal. Sponges which live in shallow waters tend either to be heavily pigmented or to have a thick layer of symbiotic algae (Wilkinson, 1980). Other interactions Except for studies of nutrient supply (see above) there has been almost no work on the possible benefits of these symbioses to the algae. Many species of microalgae, including most dinoflagellates and some green algae, show absolute requirements for various vitamins (particularly B-group vitamins) when they are grown in culture, and the growth of other species is improved by addition of vitamins (Provasoli & Carlucci, 1974). The animal hosts may supply appropriate vitamins to their symbionts. Similarly, many microscopic algae can use organic compounds to support heterotrophic growth; the possible advantages to the algae of a steady supply of such compounds from the host have never been investigated. Another possible benefit to the algae is protection from herbivores. Planktonic algae are often heavily grazed by zooplankton, which would not attack host animals of the symbiotic algae. These host animals do have their own predators, and since there have been no relevant studies, it is not possible to compare the relative risks of being eaten in the plankton or in a host. However, since most of the hosts’ predators are probably specialized carnivores, in many cases the symbiotic algae would pass through the gut of the
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predator unharmed, and be released into the water column (where they would, of course, be exposed to the herbivores). Finally, it is probably advantageous to the algae to be supported in a matrix (the host animal) which remains in a constant position in the water column; microscopic algae normally have to expend energy to remain buoyant and thus stay within the photic zone (i.e. that part of the water column where there is enough light to support plant growth). SIGNIFICANCE
AND NATURE OF ALGAL/INVERTEBRATE
SYMBIOSES
As noted in the Introduction, symbioses between algae and invertebrates are usually regarded as mutualistic. Yet, as can be seen from the outline above, almost all research has centred on the advantages of the associations to the larger partner, the animal. This has been, at least in part, due to the fact that it is relatively easy to follow the fate of products of photosynthesis, using ‘*CO2as a tracer, and relatively difficult to divise means of investigating the transfer of metabolites (particularly organic compounds) from the animals to the algae. The animals do indeed seem to benefit from these associations. In every case studied they gain products of photosynthesis, usually in large quantities. For instance, Muscatine et al. (1984) found that when the coral Stylophoru pistillata grows in well-lit habitats, it receives almost one-and-a-half times the amount of organic carbon it requires for respiration-i.e. the algae supply enough substrates to provide for all respiratory needs and to contribute to the growth of the animal. In shade-adapted S. pisti/[utu translocated products of the algae are sufficient for 58% of daily respiratory needs (Muscatine et al., 1984). Few species have been studied in as much detail as S. pistihtu, but there is no reason to suppose that it is unique. There may also be other benefits to the animal host: for example the algae may provide essential compounds not available in its diet (amino acids, fatty acids, etc.); they may provide oxygen and remove COZ during photosynthesis; they may remove toxic metabolic wastes; they may enhance calcification (where there is a calcareous skeleton) and protect animal cells from excess light or ultraviolet radiation. There has been little or no investigation of these or other potential benefits to the animals. The costs of symbiosis to the animals have not been investigated either. It is likely that there is some cost-i.e. some energy expended-in maintaining the symbionts, but its nature and extent are unknown. However, it is probable that in most of the symbioses between invertebrates and algae, the benefits to the animals are large, and outweigh the costs. The success of these associations, which dominate the highly productive communities of coral reefs and are found in many other habitats, and which add greatly to the total productivity of these communities, is evidence of the advantages to the animals of maintaining photosynthetic symbionts. Difficult as it is to fully interpret the balance of benefits and costs to the animals, there is even less to go on in trying to decide whether the algae benefit from the associations, or whether they are harmed. The only reasonable way of measuring this seems to be to compare the Darwinian fitness of the algae (i.e. their success in passing their genes on to the next generation) in symbiosis and when free-living. It is clear that the growth rates of symbiotic algae are very low. For example, Muscatine et al. (1984) calculate mean doubling times of 53 and 74 days for zooxanthellae in light- and shade-adapted S. pistillutu respectively. Free-living microalgae commonly have doubling times of 1 to 2 days. However, free-living algae are heavily grazed in most waters, and dense populations are rare and short-lived. As pointed out above, the free-living algae also face other hazards, such as sinking below the photic zone or being limited by lack of inorganic nutrients or vitamins. For symbiotic algae these problems may be prevented or minimized by the contributions of the host. Of course, it is not yet known whether most of the symbiotic strains of algae are capable of surviving outside their hosts. Until we know this, and can compare the growth rates and survival of free-living and symbiotic algae, it is not possible to define their symbioses with invertebrates as beneficial or harmful to the algae. The algae do not appear to be able to escape actively from the animals’ cells. It seems very likely that they are parasitized by the animals rather than being mutualistic symbionts. Many host species appear to regulate the numbers of their symbiotic algae (Smith, 1981). The extensive removal of photosynthetic products by the animal host might be enough to ensure that the animal controls the growth and reproduction of the algae in some associations. Animal hosts may also control the supply of nutrients to the algae. The “host factors” which promote translocation of photosynthetic products may not be the only specific compounds produced by the animals which can control their symbionts; algal growth and reproduction may also be controlled by the animal by means of specific regulators. Until all these aspects of the physiology of algal/invertebrate associations, and of the algae in their independent form (where it exists) are understood, it is not possible to make a useful assessment of the place of any of these associations in the spectrum of symbioses.
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SUMMARY The characteristics of symbiotic associations between algae and invertebrates are outlined, using a number of the better-studied examples. The common assumption that these symbioses are mutualistic ones is discussed. In the present state of knowledge, it is impossible to determine the nature of these associations, but the evidence favours the view that the animal hosts benefit markedly from the presence of the algae, but that the possibility that the algae may be harmed (i.e. parasitized) by the animals cannot be dismissed. Some general lines of investigation which would help to define these relationships are suggested. REFERENCES ANDERSONO.R. 1983. The radiolarian symbiosis. In: Algal Symbiosis+ continuum of interaction strategies pp 6939 (Edited by Goff L.J.) Cambridge University Press, Cambridge and New York. BAITEY J.F. & PATTON J.S. 1984. A re-evaluation of the role of glycerol in carbon translocation in zooxanthellaecoelenterate symbiosis. Marine Biology 79: 27-38. 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In: Symbiosis, Symposium of the Society for E.uperimenta/ Biology 29: 175-203 (Edited by Jennings D.H. & Lee D.L.) Cambridge University Press, Cambridge. MUSCA~INEL., FALKOWSKIP.G., PORTERJ.W. & DUBINSKYZ. 1984. Fate of photosynthetic fixed carbon in light- and shade-adapted colonies of the symbiotic coral Stylophorcl pistillata. Proceedings I of_ the Royal Society_ of. London, series B, i22: 181-202. PARDY R.L. 1983. Phycozoans, phycozoology, phycozoologists? In: Algal Symbiosis--a continuum of interuction strugeties pp 5-17 (Edited by Goff L.J.) Cambridge University Press, Cambridge and New York. POOLR.R. 1981. The establishment of the Hydra-Chlorella symbiosis: recognition of potential algal symbionts. Berichte Deutsches Botanisches Gesellscahft 94: 565-569. PRICE I.R., FRICKLERR.L. & WILKINSONC.R. 1984. Cerutodictyon spongiosum (Rhodophyta), the macroalgal partner in an algae-sponge symbiosis, grown in unialgal culture. Journal of Phycology 20: 156-158. 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