Preferential expulsion of dividing algal cells as a mechanism for regulating algal-cnidarian symbiosis

A wide range of both intrinsic and environmental factors can influence the population dynamics of algae in symbiosis with marine cnidarians. The present study shows that loss of algae by expulsion from cnidarian hosts is one of the primary regulators of symbiont population density. Because there is a significant linear correlation between the rate of algal expulsion and the rate of algal division, factors that increase division rates (e.g., elevated temperature) also increase expulsion rates. Additionally, 3H-thymidine is taken up to a greater extent by algae destined to be expelled than by algae retained in the host cnidarians. Taken together, data for rates of expulsion, rates of division at different temperatures, and uptake of 3H-thymidine suggest that dividing algal cells are preferentially expelled from their hosts. The preferential expulsion of dividing cells may be a mechanism for regulation of algal population density, where the rate of expulsion of algae may be an inverse function of the ability of host cells to accommodate new algal daughter cells. This kind of regulation is present in some cnidarian species (e.g., Aiptasia pulchella, Pocillopora damicornis), but not in all (e.g., Montipora verrucosa, Porites compressa, and Fungia scutaria).     

From extracellular to intracellular: the establishment of a symbiosis.

The colonization of host cells by modern symbionts is surveyed. The morphological distinction between extracellular and intracellular symbionts is not sharp, and the various kinds of association can be arranged in a graded series of increasing morphological integration of the symbiont into the host cell. Apart from some aggressive parasitic infections, the great majority of symbionts are enclosed by a host membrane in a vacuole. Those not enclosed in a host vacuole usually cannot be cultivated outside the cell. It is therefore surmised that encirclement by a vacuolar membrane would only disappear, if at all, in the later stages of the evolution of intracellular symbiosis. Recognition mechanisms between host and symbiont occur, but have been little studied. In some associations, recognition at surface contact occurs, and there is evidence for the involvement of lectins in certain cases. In other associations, recognition may occur wholly or in part after the entry of symbiont into host cells. After entry, special mechanisms for the biotrophic transfer of nutrients from symbiont to host develop. Both the symbiont population size and its rate of increase are strictly regulated by the host cell; symbiont metabolism may be controlled likewise. Rates of evolution of intracellular symbionts are probably very rapid, owing in part to responses of the host cell to its symbiont.     

Regulation of numbers of intracellular algae.

Members of three classes of unicellular algae have exploited an intracellular habitat and occur as endosymbionts in aquatic invertebrates, including Protozoa. Such associations manifest a range of host--symbiont cellular interactions and achieve stability through the regulation of symbiont numbers. The mechanism of regulation is poorly understood. Steady-state algae:host cell ratios might be achieved by expulsion, digestion, or inhibition of growth of algal symbionts. Digestion and expulsion have been observed directly in some associations but their role in regulating numbers is circumstantial. Inhibition of growth as a result of nutrient limitation or inhibitor secretion is an attractive, but inadequately tested, hypothesis. The relation between the host cell mitosis and algal proliferation is a potential focal point for further study.     

Host-symbiont associations of polycystine Radiolaria: epifluorescence microscopic observation of living Radiolaria

In all 29 polycystine radiolarian species were obtained from surface seawater on May 28, 1999, using a plankton-net at one station (Site 990528; 26°37′18″N, 127°47′35″E) approximately 5 km northwest of Okinawa Island, Japan. In most polycystine radiolarians of the orders Nassellarida and Spumellarida symbiotic algae were observed under light microscopy. The light microscopic (LM) images of the symbionts, however, varied in clarity among individuals because of the variations in microanatomy of the host radiolarian cells. On the other hand, epifluorescence microscopic (EFM) observation easily detected and confirmed the existence of the algal symbionts within the host cytoplasm even in radiolarians such as Dictyocoryne truncatum (Ehrenberg) that include algal symbionts in the depth of the cytoplasm. The chloroplasts of the algal symbionts emitted autofluorescence in ultraviolet irradiation and they appeared red. That is, the autofluorescence images of the chloroplasts can be used to recognize the existence of the algal symbionts within the host radiolarians. Moreover, staining of the symbiont cells with 4′,6-diamido-2-phenylindle permitted visualization of the nucleus in the center of the symbiont cell, confirming the existence of living endosymbiotic algae within the polycystine radiolarians. Both the LM and EFM observations of eight polycystine radiolarian species revealed the specific patterns of various host-symbiont associations. (1) The investigated polycystine radiolarians all possess algal symbionts, except for one species, i.e. Dictyocoryne profunda Ehrenberg. (2) The size of the algal symbionts depends on the radiolarian species. The symbionts are largely classified into two types based on the size of their diameters, i.e. about 8–10 μm for the larger group and about 5 μm for the smaller one. (3) The algal symbionts show a variety of locations within the host radiolarian cytoplasm. The types of distribution of algal symbionts may be a useful characteristic for radiolarian taxonomy.     

Metabolic interactions between algal symbionts and invertebrate hosts

Some invertebrates have enlisted autotrophic unicellular algae to provide a competitive metabolic advantage in nutritionally demanding habitats. These symbioses exist primarily but not exclusively in shallow tropical oceanic waters where clear water and low nutrient levels provide maximal advantage to the association. Mostly, the endosymbiotic algae are localized in host cells surrounded by a host-derived membrane (symbiosome). This anatomy has required adaptation of the host biochemistry to allow transport of the normally excreted inorganic nutrients (CO₂, NH₃ and PO₄³⁻) to the alga. In return, the symbiont supplies photosynthetic products to the host to meet its energy demands. Most attention has focused on the metabolism of CO₂ and nitrogen sources. Carbon-concentrating mechanisms are a feature of all algae, but the products exported to the host following photosynthetic CO₂ fixation vary. Identification of the stimulus for release of algal photosynthate in hospite remains elusive. Nitrogen assimilation within the symbiosis is an essential element in the host's control over the alga. Recent studies have concentrated on cnidarians because of the impact of global climate change resulting in coral bleaching. The loss of the algal symbiont and its metabolic contribution to the host has the potential to result in the transition from a coral-dominated to an algal-dominated ecosystem.     

Azolla-Anabaena Relationships: VII. Distribution of Ammonia-assimilating Enzymes, Protein, and Chlorophyll between Host and Symbiont

The N₂-fixing Azolla-Anabaena symbiotic association is characterized in regard to individual host and symbiont contributions to its total chlorophyll, protein, and levels of ammonia-assimilating enzymes. The phycocyanin content of the association and the isolated blue-green algal symbiont was used as a standard for this characterization. Phycocyanin was measured by absorption and fluorescence emission spectroscopy. The phycocyanin content and total phycobilin complement of the symbiotic algae were distinct from those of Anabaena cylindrica and a free-living isolate of the Azolla endophyte. The algal symbiont accounted for less than 20% of the association's chlorophyll and protein. Acetylene reduction rates in the association (based solely on the amount of algal chlorophyll) were 30 to 50% higher than those attained when the symbiont was isolated directly from the fern. More than 75% of the association's glutamate dehydrogenase and glutamine synthetase activities are contributed by the host plant. The specific activity of glutamate dehydrogenase is greater than that of glutamine synthetase in the association and individual partners. Both the host and symbiont have glutamate synthase activity. The net distribution of these enzymes is discussed in regard to the probable roles of the host and symbiont in the assimilation of ammonia resulting from N₂ fixation by the symbiont.     

Cellular growth of host and symbiont in a cnidarian-zooxanthellar symbiosis

The hydroid Myrionema ambionense, a fast-growing cnidarian (doubling time = 8 days) found in shallow water on tropical back-reefs, lives in symbiosis with symbiotic dinoflagellates of the genus Symbiodinium (hereafter also referred to as zooxanthellae). The symbionts live in vacuoles near the base of host digestive cells, whereas unhealthy looking zooxanthellae are generally located closer to the apical end of the host cell. Cytokinesis of zooxanthellae occurred at night, with a peak in number of symbionts with division furrows (mitotic index, MI = 12%-20%) observed at dawn. The MI of zooxanthellae decreased to near zero by the middle of the afternoon and remained there until the middle of the next night. Densities of live zooxanthellae living inside of host digestive cells peaked following cytokinesis, whereas densities of unhealthy looking symbionts were highest just before the division peak. Mitosis of host digestive cells was highest in the evening, also preceding the peak in zooxanthellar MI. This is the first study relating phased host cell division to diel zooxanthellar division in marine cnidarians. Food vacuoles were prevalent inside of digestive cells of field-collected hydroids within a few hours after sunset and throughout the night, coinciding with digestion of captured demersal plankton. Laboratory experiments showed that food vacuoles appeared in digestive cell cytoplasm within 2 h of feeding with nauplii of Artemia. The number and size of food vacuoles per digestive cell and the percentage of digestive cells with food vacuoles all decreased 5-7 h following feeding in laboratory experiments, and by mid-day in field-collected hydroids. Light and external food supply were important in maintaining phased division of the symbionts, with a lag in response time to both parameters of 11-36 h. Altering light and feeding during the night did not influence the level of the peak MI the next morning, though in one experiment the absence of light slowed final separation of daughter cells at the end of cytokinesis. In another experiment, hydroids starved for 3-7 d and "pulse-fed" Artemia nauplii for 1 h at the beginning of the dark period showed continued low symbiont division (< 5%) after 11 h, whether maintained in constant light or darkness, implying that most algal division is set more than 24 h prior to actual cytokinesis. Transferred to a 14:10 h light:dark cycle for another 24 h (36 h after feeding), the same hydroids exhibited a "normal" peak MI (ca. 15%) at dawn, but zooxanthellae from hydroids kept in constant darkness still showed a low MI. These results show that mitosis of symbiotic dinoflagellates requires three factors: external food; a minimum period of time following feeding (11-36 h), presumably for digestion; and a period of light following feeding, presumably to provide carbon skeletons necessary for completing cytokinesis.     

Fine Structure of Blue-Green Algae and the Cells Lined along the Endophyte Cavity in the Coralloid Root of Cycas

The ultrastructure of vegetative cells of blue-green alga, Anabaena cycadae, in the coralloid root of Cycas revoluta has the general characteristics of the cyanophycean cells. Their heterocysts are characterized by heavy envelope deposition, well developed pore channel with its plug, absence of large granules as inclusions and reduced and flattened photosynthetic thylakoids. By these characteristical features, the frequency of heterocysts occurring in this algal population of the coralloid root may be estimated to ca. 40%. This high heterocyst frequency is a sign of relatively high activity of nitrogen fixation in this symbiont. The ultrastructure of the cells lined along the endophyte cavity in the coralloid root shows that they have the function to maintain vigorous nutritional transport in short distance. These cells are especially characterized by the presence of numerious outgrowths on the cell wall into the endophyte cavity. Correspondingly, there are abundant mitochondria, dictyosomes and numerious vesicles in the cytoplasm. The plasma membrane becomes tortuous along the cell wall and many secretory granules are present between the plasma membrane and cell wall in the cytoplasm amyloplasts and starch granules also occur constantly. The ultrastructure observed above indicates the fact that there is sound structural basis for the metabolic relationship between the host cells and the symbiont.     

Nutritional role of two algal symbionts in the temperate sea anemone Anthopleura elegantissima brandt

The intertidal sea anemone Anthopleura elegantissima in the Pacific Northwest may host a single type of algal symbiont or two different algal symbionts simultaneously: zooxanthellae (Symbiodinium muscatinei) and zoochlorellae (green algae; Trebouxiophyceae, Chlorophyta). A seasonal comparison of zooxanthellate and zoochlorellate anemones showed stable symbiont population densities in summer and winter, with densities of zoochlorellae about 4 times those of zooxanthellae. Photosynthesis-irradiance curves of freshly isolated symbionts show that the productivity (P(max) cell) of freshly isolated zooxanthellae was about 2.5 times that of zoochlorellae during July; comparable rates were obtained in other months. Models of algal carbon flux show that zoochlorellae may supply the host with more photosynthetic carbon per unit anemone biomass than zooxanthellae supply. Zooxanthellate anemone tissue was 2 per thousand ((13)C) and 5 per thousand ((15)N) enriched and zoochlorellate anemone tissue was 6 per thousand ((13)C) and 8 per thousand ((15)N) enriched over their respective symbionts, suggesting that zoochlorellate anemones receive less nutrition from their symbionts than do zooxanthellate individuals. The disparity between predicted contributions from the algal carbon budgets and the stable isotopic composition suggests that short-term measures of algal contributions may not reflect actual nutritional inputs to the host. Isotopic data support the hypothesis of substantial reliance on external food sources. This additional nutrition may allow both algae to persist in this temperate intertidal anemone in spite of differences in seasonal photosynthetic carbon contributions.     

Regulation of the Endosymbiotic Algae in Hydra by Digestive Cells and Tissue Growth

Endosymbiotic algae in hydra release photosynthetically fixedcarbon to the host which augments the animals nutrition duringstarvation. The algae may acquire metabolites from hydra asa source of nutrition when the animals are maintained in thedark. The kind of algae acquired by aposymbiotic hydra is determinedby a recognition phenomenon involving the hosts digestive cellsand potential endosymbionts. The size of the algal populationis related to the necessity of light for maximum algal multiplication,the rate of cell division in hydra, and the potential for heterotrophyby the algae.     

Nucleus- and nucleomorph-targeted histone proteins in a chlorarachniophyte alga

The plastid of chlorarachniophytes is distinguished by the retention of a relict nucleus (nucleomorph) derived from a green algal endosymbiont, which is located in the periplastidal compartment (PPC). The nucleomorph genome of a chlorarachniophyte, Bigelowiella natans, encodes several plastid-targeted proteins and hundreds of housekeeping proteins, but it lacks many fundamental genes to maintain itself. Here we report the first two host nucleus-encoded genes for proteins targeted to the nucleomorph, histone H2A and H2B. We identified 20 histone genes from the host nuclear genome, and based on phylogenetic analyses predicted that most of these are derived from the host, but that two histone genes are symbiont-derived. The genes both encode N-terminal extensions resembling PPC targeting signals, further suggesting they function in the nucleomorph. Using green fluorescent protein (GFP) fusion proteins expressed in transformed cells, we confirmed that the putative symbiont H2A and H2B were targeted into the nucleomorph, whereas putative host proteins were localized to the host nucleus. Furthermore, we have developed a method to temporarily synchronize B. natans cells, and confirmed that both host and symbiont histone expression is controlled during the cell cycle. Our findings provide the first evidence of how the nucleomorph may be regulated by host-encoded gene products.     

Timing of perialgal vacuole membrane differentiation from digestive vacuole membrane in infection of symbiotic algae Chlorella vulgaris of the ciliate Paramecium bursaria

Each symbiotic Chlorella of the ciliate Paramecium bursaria is enclosed in a perialgal vacuole derived from the host digestive vacuole to protect from lysosomal fusion. To understand the timing of differentiation of the perialgal vacuole from the host digestive vacuole, algae-free P. bursaria cells were fed symbiotic C. vulgaris cells for 1.5min, washed, chased and fixed at various times after mixing. Acid phosphatase activity in the vacuoles enclosing the algae was detected by Gomori's staining. This activity appeared in 3-min-old vacuoles, and all algae-containing vacuoles demonstrated activity at 30min. Algal escape from these digestive vacuoles began at 30min by budding of the digestive vacuole membrane into the cytoplasm. In the budded membrane, each alga was surrounded by a Gomori's thin positive staining layer. The vacuoles containing a single algal cell moved quickly to and attached just beneath the host cell surface. Such vacuoles were Gomori's staining negative, indicating that the perialgal vacuole membrane differentiates soon after the algal escape from the host digestive vacuole. This is the first report demonstrating the timing of differentiation of the perialgal vacuole membrane during infection of P. bursaria with symbiotic Chlorella.     

Role of host nutrition in symbiont regulation: impact of dietary nitrogen on proliferation of obligate and facultative bacterial endosymbionts of the pea aphid Acyrthosiphon pisum

The impact of host nutrition on symbiont regulation in the pea aphid Acyrthosiphon pisum was investigated. The population density of the obligate symbiont Buchnera aphidicola positively correlated with dietary nitrogen levels. In contrast, the population density of the facultative symbiont Serratia symbiotica increased in aphids reared on low-nitrogen diets, indicating distinct regulatory mechanisms in the same insect host.     

Hybridization in Endophyte Symbionts Alters Host Response to Moisture and Nutrient Treatments

When a host organism is infected by a symbiont, the resulting symbiotum has a phenotype distinct from uninfected hosts. Genotypic interactions between the partners may increase phenotypic variation of the host at the population level. Neotyphodium is an asexual, vertically transmitted endophytic symbiont of grasses often existing in hybrid form. Hybridization in Neotyphodium rapidly increases the symbiotum’s genomic content and is likely to increase the phenotypic variation of the host. This phenotypic variation is predicted to enhance host performance, especially in stressful environments. We tested this hypothesis by comparing the growth, survival, and resource allocation of hybrid and nonhybrid infected host plants exposed to controlled variation in soil moisture and nutrients. Infection by a hybrid endophyte did not fit our predictions of comparatively higher root and total biomass production under low moisture/low nutrient treatments. Regardless of whether the host was infected by a hybrid or nonhybrid endophyte, both produced significantly higher root/total biomass when both nutrient and moisture were high compared to limited nutrient/moisture treatments. However, infection by hybrid Neotyphodium did result in significantly higher total biomass and host survival compared to nonhybrid infected hosts, regardless of treatment. Endophyte hybridization alters host strategies in response to stress by increasing survival in depauperate habitats and thus, potentially increasing the relative long-term host fitness.     

Hatena arenicola gen. et sp. nov., a katablepharid undergoing probable plastid acquisition

Hatena arenicola gen. et sp. nov., an enigmatic flagellate of the katablepharids, is described. It shows ultrastructural affinities to the katablepharids, including large and small ejectisomes, cell covering, and a feeding apparatus. Although molecular phylogenies of the 18S ribosomal DNA support its classification into the katablepharids, the cell is characterized by a dorsiventrally compressed cell shape and a crawling motion, both of which are unusual within this group. The most distinctive feature of Hatena arenicola is that it harbors a Nephroselmis symbiont. This symbiosis is distinct from previously reported cases of ongoing symbiosis in that the symbiont plastid is selectively enlarged, while other structures such as the mitochondria, Golgi body, cytoskeleton, and endomembrane system are degraded; the host and symbiont have developed a morphological association, i.e., the eyespot of the symbiont is always at the cell apex of Hatena arenicola; and only one daughter cell inherits the symbiont during cell division, resulting in a symbiont-bearing green cell and a symbiont-lacking colorless cell. Interestingly, the colorless cells have a feeding apparatus that corresponds to the location of the eyespot in symbiont-bearing cells, and they are able to feed on prey cells. This indicates that the morphology of the host depends on the presence or absence of the symbiont. These observations suggest that Hatena arenicola has a unique "half-plant, half-predator" life cycle; one cell divides into an autotrophic cell possessing a symbiotic Nephroselmis species, and a symbiont-lacking colorless cell, which later develops a feeding apparatus de novo. The evolutionary implications of Hatena arenicola as an intermediate step in plastid acquisition are discussed in the context of other examples of ongoing endosymbioses in dinoflagellates.     

Molecular identification of Rab7 (ApRab7) in Aiptasia pulchella and its exclusion from phagosomes harboring zooxanthellae

The establishment and maintenance of the intracellular association between marine cnidarians and their symbiotic microalgae is essential to the well being of coral reef ecosystems; however, little is known concerning its underlying molecular mechanisms. In light of the critical roles of the small GTPase, Rab7, as a key regulator of vesicular trafficking, we cloned and characterized the Rab7 protein in the endosymbiosis system between the sea anemone, Aiptasia pulchella and its algal symbiont, Symbiodinium spp. The Aiptasia homologue of Rab7 proteins, ApRab7 is 88% identical to human Rab7 protein and contains all Rab-specific signature motifs. Results of EGFP reporter analysis, protein fractionation, and immunocytochemistry support that ApRab7 is located in late endocytic and phagocytic compartments and is able to promote their fusion. Significantly, the majority of phagosomes containing live symbionts that either have taken long residency in, or were newly internalized by Aiptasia digestive cells did not contain detectable levels of ApRab7, while most phagosomes containing either heat-killed or photosynthesis-impaired symbionts were positive for ApRab7 staining. Overall, our data suggest that live algal symbionts persist inside their host cells by actively excluding ApRab7 from their phagosomes, and thereby, establish and/or maintain an endosymbiotic relationship with their cnidarian hosts.     

Characterization of the population of the sulfur-oxidizing symbiont of Codakia orbicularis (Bivalvia, Lucinidae) by single-cell analyses

We investigated the characteristics of the sulfur-oxidizing symbiont hosted in the gills of Codakia orbicularis, a bivalve living in shallow marine tropical environments. Special attention was paid to describing the heterogeneity of the population by using single-cell approaches including flow cytometry (FCM) and different microscopic techniques and by analyzing a cell size fractionation experiment. Up to seven different subpopulations were distinguished by FCM based on nucleic acid content and light side scattering of the cells. The cell size analysis of symbionts showed that the symbiotic population was very heterogeneous in size, i.e., ranging from 0.5 to 5 mum in length, with variable amounts of intracellular sulfur. The side-scatter signal analyzed by FCM, which is often taken as a proxy of cell size, was greatly influenced by the sulfur content of the symbionts. FCM revealed an important heterogeneity in the relative nucleic acid content among the subclasses. The larger cells contained exceptionally high levels of nucleic acids, suggesting that these cells contained multiple copies of their genome, i.e., ranging from one copy for the smaller cells to more than four copies for the larger cells. The proportion of respiring symbionts (5-cyano-2,3-ditolyl-terazolium chloride positive) in the bacteriocytes of Codakia revealed that around 80% of the symbionts hosted by Codakia maintain respiratory activity throughout the year. These data allowed us to gain insight into the functioning of the symbionts within the host and to propose some hypotheses on how the growth of the symbionts is controlled by the host.     

Growth kinetics of algal populations exsymbiotic from Paramecium bursaria by flow cytometry measurements.

BACKGROUND: The ciliate Paramecium bursaria normally exists as a green paramecium system because each animal cell carries several hundred, unicellular, green, algal cells in its cytoplasm. One of the remarkable and poorly understood pecularities of this system is the steady state in the number of algae per protozoan cell. A major point in the study of mechanisms governing the persistence of symbiont numbers is adequate understanding of the algal life cycle. METHODS: Asynchronously growing cell populations of several algal strains (SA-1, SA-3, and SA-9) exsymbiotic from P. bursaria were characterized by flow cytometry. Algal endogenous chlorophyll and DNA contents were monitored to analyze cell growth kinetics at logarithmic and stationary culture phases. Cell sorting visualized the morphology of algae corresponding to the hyperhaploid (2C and 4C) DNA peaks. RESULTS: Cell-division cycle-dependent changes in chlorophyll and DNA content distributions were most dramatic in logarithmically growing algal populations (an increase in the number of S-phase cells and cells with more chlorophyll), which are thought to be associated with accelerated DNA and chlorophyll metabolism in log-phase algal cultures. Upon reaching the stationary phase of growth, algal populations distinctly showed, in addition to one haploid (1C) DNA peak, two hyperhaploid peaks (2C and 4C) corresponding mainly to cells with two and four nuclei, respectively. CONCLUSIONS: Growth characteristics of algae exsymbiotic from P. bursaria monitored by flow cytometry provide valuable information for the analysis of the algal life cycle, which is important for understanding the regulation mechanisms of symbiont numbers.     

Role of Host Nutrition in Symbiont Regulation: Impact of Dietary Nitrogen on Proliferation of Obligate and Facultative Bacterial Endosymbionts of the Pea Aphid Acyrthosiphon pisum

The impact of host nutrition on symbiont regulation in the pea aphid Acyrthosiphon pisum was investigated. The population density of the obligate symbiont Buchnera aphidicola positively correlated with dietary nitrogen levels. In contrast, the population density of the facultative symbiont Serratia symbiotica increased in aphids reared on low-nitrogen diets, indicating distinct regulatory mechanisms in the same insect host.     

Phagosome-lysosome fusion inhibited by algal symbionts of Hydra viridis

Certain species of Chlorella live within the digestive cells of the fresh water cnidarian Hydra viridis. When introduced into the hydra gut, these symbiotic algae are phagocytized by digestive cells but avoid host digestion and persist at relatively constant numbers within host cells. In contrast, heat-killed symbionts are rapidly degraded after phagocytosis. Live symbionts appear to persist because host lysosomes fail to fuse with phagosomes containing live symbionts. Neither acid phosphatase nor ferritin was delivered via lysosomes into phagosomes containing live symbionts, whereas these lysosomal markers were found in 50% of the vacuoles containing heat-killed symbionts 1 h after phagocytosis. Treatment of symbiotic algae before phagocytosis with polycationic polypeptides abolishes algal persistence and perturbs the ability of these algae to control the release of photosynthate in vitro. Similarly, inhibition of photosynthesis and hence of the release of photosynthetic products as a result of prolonged darkness and 3-(3,4- dichlorophenyl)-1,1-dimethyl urea (DCMU) treatment also abolishes persistence. Symbiotic algae are not only protected from host digestive attack but are also selectively transported within host cells, moving from the apical site of phagocytosis to a basal position of permanent residence. This process too is disrupted by polycationic polypeptides, DCMU and darkness. Both algal persistence and transport may, therefore, be a function of the release of products from living, photosynthesizing symbionts. Vinblastine treatment of host animals blocked the movement of algae within host cells but did not perturb algal persistence: algal persistence and the transport of algae may be initiated by the same signal, but they are not interdependent processes.