Original Symbiont Research Articles

Growth and Metabolic Response of Glycine max to the Plant Growth-Promoting Enterobacter Delta PSK and Bradyrhizobium japonicum Under Salinity Stress

This study was designed to test the ability of some plant growth-promoting bacteria to adverse the effects of salinity on soybean growth. The effect of Bacillus MAP3 and Enterobacter Delta PSK, along with Bradyrhizobium japonicum was studied on soybean at two levels of NaCl salinization (50 and 100 mM). The physical growth parameters of bacterized soybean (21 days old), particularly plants co-inoculated with Bradyrhizobium japonicum + Enterobacter Delta PSK, were significantly enhanced compared to control plants. The shoot length, leaf area, root length, and chlorophyll a content increased by 49.58%, 78.58%, 20.19%, and 57.35%, respectively, indicating the promoting activity of this bacterial combination. After 19 days following the onset of salinity stress, the retarded growth parameters in controls improved significantly due to bacterial treatments, especially by Bradyrhizobium japonicum + Enterobacter DeltaPSK, which increased the values of all growth parameters significantly regardless of the salinity level. Additionally, electrolyte leakage, the amounts of malondialdehyde and hydrogen peroxide decreased considerably due to this combined bacterial treatment. Overall, the combination treatment of Enterobacter Delta PSK and the original symbiont B. japonicum enhanced soybean growth under salt stress, indicating the ability of Enterobacter Delta PSK to mitigate osmotic stress. The effect of this strain on soybean yield should be further evaluated to pave the way for its use as a biofertilizer along with B. japonicum, especially under salt stress.

A tale of two lineages: how the strains of the earliest divergent symbiotic Frankia clade spread over the world

It is currently assumed that around 100 million years ago, the common ancestor to the Fabales, Fagales, Rosales and Cucurbitales in Gondwana, developed a root nodule symbiosis with a nitrogen-fixing bacterium. The symbiotic trait evolved first in Frankia cluster-2; thus, strains belonging to this cluster are the best extant representatives of this original symbiont. Most cluster-2 strains could not be cultured to date, except for Frankia coriariae, and therefore many aspects of the symbiosis are still elusive. Based on phylogenetics of cluster-2 metagenome-assembled genomes (MAGs), it has been shown that the genomes of strains originating in Eurasia are highly conserved. These MAGs are more closely related to Frankia cluster-2 in North America than to the single genome available thus far from the southern hemisphere, i.e., from Papua New Guinea.To unravel more biodiversity within Frankia cluster-2 and predict routes of dispersal from Gondwana, we sequenced and analysed the MAGs of Frankia cluster-2 from Coriaria japonica and Coriaria intermedia growing in Japan, Taiwan and the Philippines. Phylogenetic analyses indicate there is a clear split within Frankia cluster-2, separating a continental from an island lineage. Presumably, these lineages already diverged in Gondwana.Based on fossil data on the host plants, we propose that these two lineages dispersed via at least two routes. While the continental lineage reached Eurasia together with their host plants via the Indian subcontinent, the island lineage spread towards Japan with an unknown host plant.

Photoacclimation and induction of light-enhanced calcification in the mesophotic coral Euphyllia paradivisa.

Corals and their photosymbionts experience inherent changes in light along depth gradients, leading them to have evolved several well-investigated photoacclimation strategies. As coral calcification is influenced by light (a process described as LEC—‘light-enhanced calcification’), studies have sought to determine the link between photosynthesis and calcification, but many puzzling aspects still persist. Here, we examine the physiology of Euphyllia paradivisa, a coral species found at a wide range of depths but that is strictly mesophotic in the Red Sea; and also examines the coupling between photosynthesis and LEC by investigating the response of the coral under several controlled light regimes during a long-term experiment. E. paradivisa specimens were collected from 40 to 50 m depth and incubated under three light conditions for a period of 1 year: full-spectrum shallow-water light (approx. 3 m, e.g. shallow-light treatment); blue deep-water light (approx. 40 m, e.g. mesophotic-light treatment) or total darkness (e.g. dark treatment). Net photosynthesis remained similar in the shallow-light-treated corals compared to the mesophotic-light-treated corals, under both low and high light. However, calcification increased dramatically with increasing light intensity in the shallow-light-treated corals, suggesting a decoupling between these processes. Photoacclimation to shallow-water conditions was indicated by enhanced respiration, a higher density of zooxanthellae per polyp and lower chlorophyll a content per cell. The dark-treated corals became completely bleached but did not lower their metabolism below that of the mesophotic-light-treated corals. No Symbiodinium clade shift was found following the year-long light treatments. We conclude that E. paradivisa, and its original symbiont clade, can adapt to various light conditions by controlling its metabolic rate and growth energy investment, and consequently induce LEC.

Thermal Tolerance in Green Hydra: Identifying the Roles of Algal Endosymbionts and Hosts in a Freshwater Holobiont Under Stress.

It has been proposed that holobionts (host-symbiont units) could swap endosymbionts, rapidly alter the hologenome (host plus symbiont genome), and increase their stress tolerance. However, experimental tests of individual and combined contributions of hosts and endosymbionts to holobiont stress tolerance are needed to test this hypothesis. Here, we used six green hydra (Hydra viridissima) strains to tease apart host (hydra) and symbiont (algae) contributions to thermal tolerance. Heat shock experiments with (1) hydra with their original symbionts, (2) aposymbiotic hydra (algae removed), (3) novel associations (a single hydra strain hosting different algae individually), and (4) control hydra (aposymbiotic hydra re-associated with their original algae) showed high variation in thermal tolerance in each group. Relative tolerances of strains were the same within original, aposymbiotic, and control treatments, but reversed in the novel associations group. Aposymbiotic hydra had similar or higher thermal tolerance than hydra with algal symbionts. Selection on the holobiont appears to be stronger than on either partner alone, suggesting endosymbiosis could become an evolutionary trap under climate change. Our results suggest that green hydra thermal tolerance is strongly determined by the host, with a smaller, non-positive role for the algal symbiont. Once temperatures exceed host tolerance limits, swapping symbionts is unlikely to allow these holobionts to persist. Rather, increases in host tolerance through in situ adaptation or migration of pre-adapted host strains appear more likely to increase local thermal tolerance. Overall, our results indicate green hydra is a valuable system for studying aquatic endosymbiosis under changing environmental conditions, and demonstrate how the host and the endosymbiont contribute to holobiont stress tolerance.

Symbiotic Origin of Aging.

Normally aging cells are characterized by an unbalanced mitochondrial dynamic skewed toward punctate mitochondria. Genetic and pharmacological manipulation of mitochondrial fission/fusion cycles can contribute to both accelerated and decelerated cellular or organismal aging. In this work, we connect these experimental data with the symbiotic theory of mitochondrial origin to generate new insight into the evolutionary origin of aging. Mitochondria originated from autotrophic α-proteobacteria during an ancient endosymbiotic event early in eukaryote evolution. To expand beyond individual host cells, dividing α-proteobacteria initiated host cell lysis; apoptosis is a product of this original symbiont cell lytic exit program. Over the course of evolution, the host eukaryotic cell attenuated the harmful effect of symbiotic proto-mitochondria, and modern mitochondria are now functionally interdependent with eukaryotic cells; they retain their own circular genomes and independent replication timing. In nondividing differentiated or multipotent eukaryotic cells, intracellular mitochondria undergo repeated fission/fusion cycles, favoring fission as organisms age. The discordance between cellular quiescence and mitochondrial proliferation generates intracellular stress, eventually leading to a gradual decline in host cell performance and age-related pathology. Hence, aging evolved from a conflict between maintenance of a quiescent, nonproliferative state and the evolutionarily conserved propagation program driving the life cycle of former symbiotic organisms: mitochondria.

High degree of specificity in the association between symbiotic betaproteobacteria and the host Euplotes (Ciliophora, Euplotia)

The Betaproteobacteria-Euplotes association is an obligatory symbiotic system involving a monophyletic group of ciliate species and two betaproteobacteria species which can be alternatively present. Recent data showed that this relationship has been established more than once and that several symbiont-substitution events took place, revealing a complex and intriguing evolutionary path. Due to the different evolutionary pathways followed by the different symbionts, each bacterial strain could have differentially evolved and/or lost functional traits. Therefore, we performed re-infection experiments, both by phagocytosis and by microinjection, to test the possible functional role of the different bacteria towards the ciliates. Our results confirm that the growth capacity of the host is indissolubly linked to the presence of its original symbionts. Results of the attempts of re-infection by phagocytosis showed that none of the bacteria is able to successfully colonize the host cytoplasm in this way, even if regularly ingested. Re-infection by microinjection succeed only in one case. Such results point to a high degree of specificity in the interactions between bacteria and Euplotes even after the invasion step. Due to a co-evolutive pathway of reciprocal adaptation, different degree of re-colonization ability could have been conserved by the different species and strains of the symbionts.

The Structure and Evolution of Beta-Rhizobial Symbiotic Genes Deduced from Their Complete Genomes

Abstract Nitrogen-fixing Rhizobia were discovered more than 100 years ago. They are classified into two clades, Alphaand Beta-rhizobia. Their symbiotic function is remarkable, but its origin and evolution has been confusing from a phylogenetic perspective. In this study, we make use of 33 publicly available complete genome sequences downloaded from NCBI, which consist of bacteria and archaea, and focus on 10 strains, constructing symbiotic structural maps for them based on their genomes and previous gene annotations. Phylogenies of the symbiosisessential genes nodA and nifH were examined. Although large incongruities with some hypotheses from previous studies were detected by the present study, we support the general concept that Beta-rhizobia were the original symbionts of legumes, but that their symbotic genes originated from a common ancestor to the Alpha-rhizobia. We also confirm that the spread and maintenance of symbiotic genes occurred mainly through vertical transmission, with lateral transfer playing a significant, albeit supporting, role.

Coral symbioses under prolonged environmental change: living near tolerance range limits.

As climate change progresses, understanding the long-term response of corals and their endosymbionts (Symbiodinium) to prolonged environmental change is of immediate importance. Here, a total of 1152 fragments from 72 colonies of three common coral species (Stylophora pistillata, Pocillopora damicornis, Seriatopora hystrix) underwent a 32-month reciprocal depth transplantation. Genetic analysis showed that while S. hystrix maintained its generalist symbiont, some S. pistillata and P. damicornis underwent temporary changes in resident symbionts immediately after stress (transplantation; natural bleaching). These temporary changes were phylogenetically constrained to ‘host-compatible’ symbionts only and reversion to original symbionts occurred within 7 to 12 months, indicating long-term fidelity and stability of adult symbioses. Measurements of symbiont photo-physiology (dark adapted yield, pressure over photosystem II) and coral health (host protein, bleaching status, mortality) indicated a broad acclimatory capacity. However, this came at an apparent energetic expense as disproportionate mortality amongst symbioses that persisted outside their distribution range was observed following a natural bleaching event. As environmental changes due to climate change become more continuous in nature, sub-lethal effects linked to the existence near tolerance range limits coupled with the inability of adult coral colonies to change resident symbionts makes corals particularly susceptible to additional environmental fluctuations or stress events and reduces the resilience of coral populations.

Symbiosis-specific changes in dimethylsulphoniopropionate concentrations in Stylophora pistillata along a depth gradient

Scleractinian corals are prolific producers of dimethylsulphoniopropionate (DMSP), but ecophysiological mechanisms influencing cellular concentrations are uncertain. While DMSP is often proposed to function as an antioxidant, interactions between specific host–symbiont genotype associations, plasticity in DMSP concentrations and environmental conditions that can either exert or alleviate oxidative stress are unclear. We used long-term (6 months) reciprocal transplantation of Stylophora pistillata hosting two distinct symbiont phylotypes along a depth gradient, clades A ( 20 m), to assess the effect of change in depth (light intensity) on DMSP concentrations in relation to symbiont genotype and photoacclimation in corals between 3 and 50 m in the Gulf of Aqaba. Bathymetric distribution of total DMSP (DMSPt) per cell varied significantly while particulate DMSP (DMSPp) appeared to be unaffected by depth. Highest DMSPt concentrations in control corals occurred at 20 m. While 3-m transplants showed a significant increase in DMSPt concentration at 20 m and became affiliated with an additional genotype (C72), 50-m transplants largely persisted with their original genotype and exhibited no significant changes in DMSPt concentrations. DMSPt concentrations in transplants at both 3 and 50 m, on the other hand, increased significantly while all corals maintained their original symbiont genotypes. Photoacclimation differed significantly with transplantation direction relative to the controls. Symbionts in 3-m transplants at 20 m exhibited no changes in chlorophyll a (chl a) concentration, cell density or cell diameter while symbiont densities decreased and chl a concentrations increased significantly at 50 m. In contrast, symbiont densities in 50-m transplants remained unaffected across depths while symbiont diameters decreased. Chl a concentrations decreased at 20 m and increased at 3 m. Our results indicate that DMSPt concentrations following changes in depth are not only a function of symbiont genotype but result from different acclimation abilities of both symbiotic partners.

Rhizobium leguminosarumis the symbiont of lentils in the Middle East and Europe but not in Bangladesh

Lentil is the oldest of the crops that have been domesticated in the Fertile Crescent and spread to other regions during the Bronze Age, making it an ideal model to study the evolution of rhizobia associated with crop legumes. Housekeeping and nodulation genes of lentil-nodulating rhizobia from the region where lentil originated (Turkey and Syria) and regions to which lentil was introduced later (Germany and Bangladesh) were analyzed to determine their genetic diversity, population structure, and taxonomic position. There are four different lineages of rhizobia associated with lentil nodulation, of which three are new and endemic to Bangladesh, while Mediterranean and Central European lentil symbionts belong to the Rhizobium leguminosarum lineage. The endemic lentil grex pilosae may have played a significant role in the origin of these new lineages in Bangladesh. The presence of R. leguminosarum with lentil at the center of origin and in countries where lentil was introduced later suggests that R. leguminosarum is the original symbiont of lentil. Lentil seeds may have played a significant role in the initial dispersal of this Rhizobium species within the Middle East and on to other countries. Nodulation gene sequences revealed a high similarity to those of symbiovar viciae.

Diversification of endosymbiosis: replacements, co-speciation and promiscuity of bacteriocyte symbionts in weevils

The processes and mechanisms underlying the diversification of host-microbe endosymbiotic associations are of evolutionary interest. Here we investigated the bacteriocyte-associated primary symbionts of weevils wherein the ancient symbiont Nardonella has experienced two independent replacement events: once by Curculioniphilus symbiont in the lineage of Curculio and allied weevils of the tribe Curculionini, and once by Sodalis-allied symbiont in the lineage of grain weevils of the genus Sitophilus. The Curculioniphilus symbiont was detected from 27 of 36 Curculionini species examined, the symbiont phylogeny was congruent with the host weevil phylogeny, and the symbiont gene sequences exhibited AT-biased nucleotide compositions and accelerated molecular evolution. These results suggest that the Curculioniphilus symbiont was acquired by an ancestor of the tribe Curculionini, replaced the original symbiont Nardonella, and has co-speciated with the host weevils over evolutionary time, but has been occasionally lost in several host lineages. By contrast, the Sodalis-allied symbiont of Sitophilus weevils exhibited no host-symbiont co-speciation, no AT-biased nucleotide compositions and only moderately accelerated molecular evolution. These results suggest that the Sodalis-allied symbiont was certainly acquired by an ancestor of the Sitophilus weevils and replaced the original Nardonella symbiont, but the symbiotic association must have experienced occasional re-associations such as new acquisitions, horizontal transfers, replacements and/or losses. We detected Sodalis-allied facultative symbionts in populations of the Curculionini weevils, which might represent potential evolutionary sources of the Sodalis-allied primary symbionts. Comparison of these newcomer bacteriocyte-associated symbiont lineages highlights potential evolutionary trajectories and consequences of novel symbionts after independent replacements of the same ancient symbiont.

Functional remodeling of RNA processing in replacement chloroplasts by pathways retained from their predecessors.

Chloroplasts originate through the endosymbiotic integration of a host and a photosynthetic symbiont, with processes established within the host for the biogenesis and maintenance of the nascent chloroplast. It is thought that several photosynthetic eukaryotes have replaced their original chloroplasts with others derived from different source organisms in a process termed "serial endosymbiosis of chloroplasts." However, it is not known whether replacement chloroplasts are affected by the biogenesis and maintenance pathways established to support their predecessors. Here, we investigate whether pathways established during a previous chloroplast symbiosis function in the replacement chloroplasts of the dinoflagellate alga Karenia mikimotoi. We show that chloroplast transcripts in K. mikimotoi are subject to 3' polyuridylylation and extensive sequence editing. We confirm that these processes do not occur in free-living relatives of the replacement chloroplast lineage, but are otherwise found only in the ancestral, red algal-derived chloroplasts of dinoflagellates and their closest relatives. This indicates that these unusual RNA-processing pathways have been retained from the original symbiont lineage and made use of by the replacement chloroplast. Our results constitute an addition to current theories of chloroplast evolution in which chloroplast biogenesis may be radically remodeled by pathways remaining from previous symbioses.

Host–symbiont recombination versus natural selection in the response of coral–dinoflagellate symbioses to environmental disturbance

Mutualisms between reef-building corals and endosymbiotic dinoflagellates are particularly sensitive to environmental stress, yet the ecosystems they construct have endured major oscillations in global climate. During the winter of 2008, an extreme cold-water event occurred in the Gulf of California that bleached corals in the genus Pocillopora harbouring a thermally 'sensitive' symbiont, designated Symbiodinium C1b-c, while colonies possessing Symbiodinium D1 were mostly unaffected. Certain bleached colonies recovered quickly while others suffered partial or complete mortality. In most colonies, no appreciable change was observed in the identity of the original symbiont, indicating that these partnerships are stable. During the initial phases of recovery, a third species of symbiont B1(Aiptasia), genetically identical to that harboured by the invasive anemone, Aiptasia sp., grew opportunistically and was visible as light-yellow patches on the branch tips of several colonies. However, this symbiont did not persist and was displaced in all cases by C1b-c several months later. Colonies with D1 were abundant at inshore habitats along the continental eastern Pacific, where seasonal turbidity is high relative to offshore islands. Environmental conditions of the central and southern coasts of Mexico were not sufficient to explain the exclusivity of D1 Pocillopora in these regions. It is possible that mass mortalities associated with major thermal disturbances during the 1997-1998 El Niño Southern Oscillation eliminated C1b-c holobionts from these locations. The differential loss of Pocillopora holobionts in response to thermal stress suggests that natural selection on existing variation can cause rapid and significant shifts in the frequency of particular coral-algal partnerships. However, coral populations may take decades to recover following episodes of severe selection, thereby raising considerable uncertainty about the long-term viability of these communities.

Stability of coral–endosymbiont associations during and after a thermal stress event in the southern Great Barrier Reef

Shifts in the community of symbiotic dinoflagellates to those that are better suited to the prevailing environmental condition may provide reef-building corals with a rapid mechanism by which to adapt to changes in the environment. In this study, the dominant Symbiodinium in 10 coral species in the southern Great Barrier Reef was monitored over a 1-year period in 2002 that coincided with a thermal stress event. Molecular genetic profiling of Symbiodinium communities using single strand conformational polymorphism of the large subunit rDNA and denaturing gradient gel electrophoresis of the internal transcribed spacer 2 region did not detect any changes in the communities during and after this thermal-stress event. Coral colonies of seven species bleached but recovered with their original symbionts. This study suggests that the shuffling or switching of symbionts in response to thermal stress may be restricted to certain coral species and is probably not a universal feature of the coral-symbiont relationship.

EPHEMERAL WINDOWS OF OPPORTUNITY FOR HORIZONTAL TRANSMISSION OF FUNGAL SYMBIONTS IN LEAF-CUTTING ANTS

Evolutionary theory predicts that hosts are selected to prevent mixing of genetically different symbionts when competition among lineages reduces the productivity of a mutualism. The symbionts themselves may also defend their interests: recent studies of Acromyrmex leaf-cutting ants showed that somatic incompatibility enforces single-clone gardens within mature colonies, thereby constraining horizontal transmission of fungal symbionts. However, phylogenetic analyses indicate that symbiont switches occur frequently enough to remove most signs of host-symbiont cocladogenesis. Here we resolve this paradox by showing that transmission among newly founded Acromyrmex colonies is not constrained. All tested queens of sympatric A. octospinosus and A. echinatior offered a novel fragment of fungus garden accepted the new symbiont. The outcome was unaffected by genetic distance between the novel and the original symbiont, and by the ant species the novel symbiont came from. The colony founding stage may thus provide an efficient but transient window for horizontal transmission, in which the fungus is unable to actively defend its partnership position before the host feeds on it, so that host fecal droplets remain compatible with alternative strains during the early stage of colony founding. We discuss how brief stages of low commitment between partners may increase the evolutionary stability of ancient coevolved mutualisms.

Bleaching susceptibility and mortality of corals are determined by fine-scale differences in symbiont type.

Coral bleaching has been identified as one of the major contributors to coral reef decline, and the occurrence of different symbionts determined by broad genetic groupings (clades A-H) is commonly used to explain thermal responses of reef-building corals. By using Stylophora pistillata as a model, we monitored individual tagged colonies in situ over a two-year period and show that fine level genetic variability within clade C is correlated to differences in bleaching susceptibility. Based on denaturing gradient gel electrophoresis of the internal transcribed spacer region 2, visual bleaching assessments, symbiont densities, host protein, and pulse amplitude modulated fluorometry, we show that subcladal types C78 and C8/a are more thermally tolerant than C79 and C35/a, which suffered significant bleaching and postbleaching mortality. Although additional symbiont types were detected during bleaching in colonies harboring types C79 and C35/a, all colonies reverted back to their original symbionts postbleaching. Most importantly, the data propose that the differential mortality of hosts harboring thermally sensitive versus resistant symbionts rather than symbiont shuffling/switching within a single host is responsible for the observed symbiont composition changes of coral communities after bleaching. This study therefore highlights that the use of broad cladal designations may not be suitable to describe differences in bleaching susceptibility, and that differential mortality results in a loss of both symbiont and host genetic diversity and therefore represents an important mechanism in explaining how coral reef communities may respond to changing conditions.

Persistence of non-caribbean algal symbionts in Indo-Pacific mushroom corals released to Jamaica 35�years ago

Mushroom corals endemic to the Indo-Pacific were collected and brought to Discovery Bay, Jamaica, and released on the fore-reef in the mid to late 1960’s by Thomas F. Goreau, an early pioneer in coral research (cf. Goreau et al. 1969 p 180; R. K. Trench, personal communication). Subsequently, the potential problems arising from introduction of an exotic species to the Caribbean were realized. Several attempts to eradicate these non-Caribbean species were then made between 1970 and 1980 and more than 25 adult individuals were removed (cf. Bush et al. 2004; J. Woodley, personal communication). However, re-discovery of two Fungia scutaria individuals in March of 2003 showed that a remnant population of this particular mushroom coral has persisted for over 35 years in the Caribbean (Bush et al. 2004). Since being introduced to Atlantic waters, these corals have displayed remarkable resilience, enduring two destructive hurricanes (Woodley 1992), environmental degradation resulting in collapse of native coral populations (Hughes 1994), and several bleaching events related to thermal stress (Goreau 1990). Understanding the stability and/or flexibility of coralalgal symbioses is of great interest to those concerned with the long-term effects of climate change on reef building corals (Baker 2003). Whether the original symbiont population of F. scutaria was replaced by one of the many local Symbiodinium spp. found in the Caribbean (LaJeunesse 2002; LaJeunesse et al. 2003) is addressed in this report through use of genetic analyses. Tissue from a F. scutaria collected on the fore-reef (15 m) off the north shore of Jamaica was preserved in 95% ethanol, homogenized, and the DNA isolated as described previously (LaJeunesse et al. 2003). PCRdenaturing gradient gel electrophoresis (DGGE) fingerprinting of the ribosomal internal transcribed spacer region (ITS) 2 was performed and the two lowest and most prominent DNA bands were excised and sequenced (Fig. 1). The genome of this Symbiodinium sp. contains two co-dominant sequence variants of the ITS 2. They appear to reflect a phylogenetic transition from C1 (ancestral) to the C1b (derived) sequence. When the DGGE fingerprints and sequences are compared with data from hundreds of Caribbean and Indo-Pacific host taxa, an identical match is made with a symbiont of Pacific origin, type C1b (Fig. 1). That member of Symbiodinium clade C was recently identified in the coral Pavona varians (collected at 3 m) from the southern Great Barrier Reef (GBR) (LaJeunesse et al. 2003) and in Leptastrea purpurea (15 m) and the organ pipe octocoral, Tubipora musica (15 m), from mid-shelf reefs on the central GBR (LaJeunesse et al., in press). It is also found in P. superficialis (6 m) from the Pacific coast of Panama (A.C. Baker and T.C. LaJeunesse, unpublished data) and appears to possess a wide, albeit uncommon, geographic distribution in the Pacific. The identical match of the ITS 2 fingerprint, C1b, from the Jamaican F. scutaria to samples taken from various Pacific hosts indicates that this Symbiodinium sp. is most probably a product of the Indo-Pacific diversification of clade C. Presently, over 50 genetically distinguishable symbiont types, mostly from Symbiodinium clades B and C, have been classified from extensive host collections at several sites in the Caribbean (LaJeunesse 2002; LaJeunesse, in press). They predominantly have limited geographic, host-specific and/or well-defined depth distributions within the Caribbean Sea. Only two types from clade C, namely C1 (S. goreaui) and C3, are known to also occur within a wide diversity of host taxa T. C. LaJeunesse (&) Department of Plant Biology and Institute of Ecology, University of Georgia, Plant Sciences Building, Athens, GA 30602, USA E-mail: lajeunes@fiu.edu Tel.: +1-305-3481317 Fax: +1-305-3481986

The chloroplasts of Euglena may have evolved from symbiotic green algae

It is proposed that the chloroplasts of Euglena have arisen from the progressive reduction of endosymbiotic green algae. The theory is supported by the presence of a third membrane around the chloroplasts of Euglena which is not endoplasmic reticulum (ER) in origin and may be derived from the plasmalemma of the original symbiont. In addition, Euglena is the only organism in which chloroplast loss can be induced experimentally. Dinoflagellate chloroplasts are also surrounded by three membranes, and it is proposed that they too evolved from symbiotic eucaryotic algae.