Symbiodinium diversity among host clionaid sponges from Caribbean and Pacific reefs: Evidence of heteroplasmy and putative host-specific symbiont lineages

Among the Porifera, symbiosis with Symbiodinium spp. (i.e., zooxanthellae) is largely restricted to members of the family Clionaidae. We surveyed the diversity of zooxanthellae associated with sponges from the Caribbean and greater Indo-Pacific regions using chloroplast large subunit (cp23S) domain V sequences. We provide the first report of Clade C Symbiodinium harbored by a sponge (Cliona caesia), and the first report of Clade A Symbiodinium from an Indo-Pacific sponge (C. jullieni). Clade A zooxanthellae were also identified in sponges from the Caribbean, which has been reported previously. Sponges that we examined from the Florida Keys all harbored Clade G Symbiodinium as did C. orientalis from the Indo-Pacific, which also supports earlier work with sponges. Two distinct Clade G lineages were identified in our phylogenetic analysis; Symbiodinium extracted from clionaid sponges formed a monophyletic group sister to Symbiodinium found in foraminiferans. Truncated and 'normal' length variants of 23S rDNA sequences were detected simultaneously in all three morphotypes of C. varians providing the first evidence of chloroplast-based heteroplasmy in a sponge. None of the other sponge species examined showed evidence of heteroplasmy. As in previous work, length variation in cp23S domain V sequences was found to correspond in a highly precise manner to finer resolution of phylogenetic topology among Symbiodinium clades. On a global scale, existing data indicate that members of the family Clionaidae that host zooxanthellae can form symbiotic associations with at least four Symbiodinium clades. The majority of sponge hosts appear to harbor only one cladal type of symbiont, but some species can harbor more than one clade of zooxanthellae concurrently. The observed differences in the number of partners harbored by sponges raise important questions about the degree of coevolutionary integration and specificity of these symbioses. Although our sample sizes are small, we propose that one of the Clade G lineages identified in this study is comprised of sponge-specialist zooxanthellae. These zooxanthellae are common in Caribbean sponges, but additional work in other geographic regions is necessary to test this idea. Sponges from the Indo-Pacific region harbor zooxanthellae from Clades A, C, and G, but more sponges from this region should be examined.     

Phylogenetic relationships among zooxanthellae (Symbiodinium) associated to excavating sponges (Cliona spp.) reveal an unexpected lineage in the Caribbean

Phylogenetic relationships of symbiotic dinoflagellate lineages, distributed in all tropical and subtropical seas, suggest strategies for long distance dispersal but at the same time strong host specialization. Zooxanthellae (Symbiodinium: Dinophyta), which are associated to diverse shallow-water cnidarians, also engage in symbioses with some sponge species of the genus Cliona. In the Caribbean, zooxanthellae-bearing Cliona has recently become abundant due to global warming, overfishing, and algae abundance. Using molecular techniques, the symbionts from five excavating species (Clionacaribbaea, C. tenuis, C. varians, C. aprica and C. laticavicola) from the southern and southwestern Caribbean were surveyed. Several DNA sequence regions were used in order to confirm zooxanthellae identity; 18S rDNA, domain V of chloroplast large subunit (cp23S), internal transcribed spacer 2 (ITS2), and ITS2 secondary structure. Sequence analyses corroborated the presence of three zooxanthellae clades: A, B, and G. Presence of clades A and B in common boring sponges of the Caribbean fit with the general pattern of the province. The discovery of clade G for the first time in any organism of the Atlantic Ocean leads us to consider this unusual finding as a phylogenetic relict through common ancestors of sponge clades or an invasion of the sponge from the Indo-Pacific.     

珊瑚共生体碳代谢特征研究进展

珊瑚礁作为一种典型的海洋生态系统,具有巨大的固碳和储碳潜力。然而,目前对于珊瑚礁的净碳能力(碳释放与碳吸收)仍存在争议,主要归因于珊瑚共生体碳代谢的多样性和复杂性。珊瑚礁在生物钙化、呼吸过程中向大气释放二氧化碳(CO₂);但在生物合成和沉积过程中却可以将碳进行固定与埋藏;为此,珊瑚礁的碳源碳汇身份还有待明确。现有部分研究表明,共生体通过碳代谢可以促进珊瑚礁吸收大气中的CO₂。此外,珊瑚礁和海岸带蓝碳生态系统通常表现出很强的连通性,珊瑚共生体碳代谢能有效提高海岸带盐沼植被、海草床、海洋浮游植物等生物的碳汇功能。为了加深对珊瑚礁碳源-碳汇功能的理解,综述了珊瑚共生体的碳代谢特征,梳理了共生体中碳的关键生态过程(有机碳的迁移、无机碳的转化、两者的赋存状态),总结了细菌-虫黄藻-病毒在共生体碳代谢中的作用,评述了珊瑚礁碳源-碳汇特征及影响因子。旨在阐明珊瑚共生体碳代谢的关键过程,并基于此寻求有效的珊瑚礁碳增汇技术,形成以碳增量为主的珊瑚保护与修复技术,提升珊瑚礁在蓝碳生态系统中的贡献。     

Stability of Sponge-Associated Bacteria over Large Seasonal Shifts in Temperature and Irradiance

Complex microbiomes reside in marine sponges and consist of diverse microbial taxa, including functional guilds that may contribute to host metabolism and coastal marine nutrient cycles. Our understanding of these symbiotic systems is based primarily on static accounts of sponge microbiota, while their temporal dynamics across seasonal cycles remain largely unknown. Here, we investigated temporal variation in bacterial symbionts of three sympatric sponges (Ircinia spp.) over 1.5 years in the northwestern (NW) Mediterranean Sea, using replicated terminal restriction fragment length polymorphism (T-RFLP) and clone library analyses of bacterial 16S rRNA gene sequences. Bacterial symbionts in Ircinia spp. exhibited host species-specific structure and remarkable stability throughout the monitoring period, despite large fluctuations in temperature and irradiance. In contrast, seawater bacteria exhibited clear seasonal shifts in community structure, indicating that different ecological constraints act on free-living and on symbiotic marine bacteria. Symbiont profiles were dominated by persistent, sponge-specific bacterial taxa, notably affiliated with phylogenetic lineages capable of photosynthesis, nitrite oxidation, and sulfate reduction. Variability in the sponge microbiota was restricted to rare symbionts and occurred most prominently in warmer seasons, coincident with elevated thermal regimes. Seasonal stability of the sponge microbiota supports the hypothesis of host-specific, stable associations between bacteria and sponges. Further, the core symbiont profiles revealed in this study provide an empirical baseline for diagnosing abnormal shifts in symbiont communities. Considering that these sponges have suffered recent, episodic mass mortalities related to thermal stresses, this study contributes to the development of model sponge-microbe symbioses for assessing the link between symbiont fluctuations and host health.     

Metagenomic Analysis of Genes Encoding Nutrient Cycling Pathways in the Microbiota of Deep-Sea and Shallow-Water Sponges

Sponges host complex symbiotic communities, but to date, the whole picture of the metabolic potential of sponge microbiota remains unclear, particularly the difference between the shallow-water and deep-sea sponge holobionts. In this study, two completely different sponges, shallow-water sponge Theonella swinhoei from the South China Sea and deep-sea sponge Neamphius huxleyi from the Indian Ocean, were selected to compare their whole symbiotic communities and metabolic potential, particularly in element transformation. Phylogenetically diverse bacteria, archaea, fungi, and algae were detected in both shallow-water sponge T. swinhoei and deep-sea sponge N. huxleyi, and different microbial community structures were indicated between these two sponges. Metagenome-based gene abundance analysis indicated that, though the two sponge microbiota have similar core functions, they showed different potential strategies in detailed metabolic processes, e.g., in the transformation and utilization of carbon, nitrogen, phosphorus, and sulfur by corresponding microbial symbionts. This study provides insight into the putative metabolic potentials of the microbiota associated with the shallow-water and deep-sea sponges at the whole community level, extending our knowledge of the sponge microbiota’s functions, the association of sponge- microbes, as well as the adaption of sponge microbiota to the marine environment.     

珊瑚共生体中“细菌-虫黄藻-宿主”三角关系的通讯交流

“细菌-虫黄藻-珊瑚”是生态系统中一对经典的三角关系,其中包含着复杂的物质流、信息流和能量流,三者的平衡与稳定是维护珊瑚礁生态系统健康的重要保障。过去20年里针对共生体交互关系进行了大量研究,并取得了一些重要成果,明确了“细菌-虫黄藻-宿主”三者之间的物质代谢、营养交换以及与环境的交互关系。然而,基于共生系统的复杂性,一些现象背后的机制仍然未被充分揭示,尤其是共生体之间的通讯交流。信号分子介导的相互作用是珊瑚共生体稳态维持和高效运转的内在驱动力。本文以珊瑚共生体系中化学信号为重点,尝试梳理最新的研究进展,包括细菌与细菌、细菌与珊瑚、细菌与虫黄藻以及虫黄藻与珊瑚之间的通讯方式,重点关注了群体感应信号(QS)、二甲基巯基丙酸盐(DMSP)、糖类信号、脂类信号以及非编码RNA。选择性例举了QS信号介导的微生物协作和竞争、DMSP调节下的细菌和宿主的相互作用,以及环境胁迫下珊瑚和虫黄藻对非编码RNA的响应过程,强调了它们在共生体中的作用模式和生态意义。并对今后的研究重点和可能方向进行了提炼,包括研究维度的扩充、新技术-新方法的应用以及生态模型的构建等,旨在提升对三角关系互作方式的认识,增进对珊瑚共生体的理解,探索基于通讯语言的操纵方式为珊瑚礁生态系统的恢复和保护提供新思路。     

Sponge erosion under acidification and warming scenarios: differential impacts on living and dead coral

Ocean acidification will disproportionately impact the growth of calcifying organisms in coral reef ecosystems. Simultaneously, sponge bioerosion rates have been shown to increase as seawater pH decreases. We conducted a 20‐week experiment that included a 4‐week acclimation period with a high number of replicate tanks and a fully orthogonal design with two levels of temperature (ambient and +1 °C), three levels of pH (8.1, 7.8, and 7.6), and two levels of boring sponge (Cliona varians, present and absent) to account for differences in sponge attachment and carbonate change for both living and dead coral substrate (Porites furcata). Net coral calcification, net dissolution/bioerosion, coral and sponge survival, sponge attachment, and sponge symbiont health were evaluated. Additionally, we used the empirical data from the experiment to develop a stochastic simulation of carbonate change for small coral clusters (i.e., simulated reefs). Our findings suggest differential impacts of temperature, pH and sponge presence for living and dead corals. Net coral calcification (mg CaCO₃ cm^?2 day^?1) was significantly reduced in treatments with increased temperature (+1 °C) and when sponges were present; acidification had no significant effect on coral calcification. Net dissolution of dead coral was primarily driven by pH, regardless of sponge presence or seawater temperature. A reevaluation of the current paradigm of coral carbonate change under future acidification and warming scenarios should include ecologically relevant timescales, species interactions, and community organization to more accurately predict ecosystem‐level response to future conditions.     

Assessment of the relative contribution of asexual propagation in a population of the coral-excavating sponge Cliona delitrix from the Bahamas

Cliona delitrix is a very destructive coral-excavating sponge in Caribbean coral reef systems, particularly for Montastraea species. Little is known about how these excavating sponges propagate across coral reefs. In this study a hypothesis was tested that coral breakage caused by the bioeroding activity facilitates the asexual propagation of this sponge and in turn favors the spread of the most aggressive sponge genotypes. An allozyme analysis, involving 12 loci systems of 52 sponge individuals from a total of 13 Montastraea heads, found that no two sponges possessed identical multi-locus genotypes. Contrary to the pattern expected for fragmenting species, the incidence of clonality and asexual propagation at the population level was minimal. The lack of correlation between genetic and physical distances for the studied sponges also suggests that population maintenance appears to derive from larval dispersal, with a spatial range of dispersal larger than the average distance between the coral heads (10–10² m).     

The potential of azooxanthellate poriferan hosts to assess the fundamental and realized <Emphasis Type="Italic">Symbiodinium</Emphasis> niche: evaluating a novel method to initiate <Emphasis Type="Italic">Symbiodinium</Emphasis> associations

On coral reefs, Symbiodinium spp. are found in most cnidarian species, but reside in only a small number of sponge species. Of the sponges that do harbor Symbiodinium, most are found in the family Clionaidae, which represents a minor fraction of the poriferan diversity on a reef. Our goal was to determine whether Symbiodinium can be taken up by sponge hosts that do not typically harbor these algal symbionts, and then to follow the fate of any Symbiodinium that enter the intracellular space. We used the filter-feeding capacity of sponges to initiate intracellular interactions between sponge-specialist clade G Symbiodinium and six sponge species that do not associate with Symbiodinium. Using a pulse-chase experimental design, we determined that all of the species we examined captured Symbiodinium, and undamaged intracellular algae were found up to 1 h after inoculation. In a longer-term experiment, Symbiodinium populations in Amphimedon erina persisted in sponge cells for at least 5 d post-inoculation. While no evidence of digestion was detected, the population decreased exponentially after inoculation. We contrast these data with the characteristics of symbiont acquisition and establishment in Cliona varians, which normally harbors Symbiodinium. Explants from experimentally derived aposymbiotic sponges were placed in the field where they acquired Symbiodinium from ambient sources (i.e., we did not inoculate them as in the pulse-chase experiments). We began to detect Symbiodinium cells in C. varians after 12 d, and the algal population increased exponentially until densities approached those typically found in this host (after ~128 d). We discuss the implications of this work in light of growing interest in the evolution of specificity between hosts and symbionts, and the fundamental and realized niche of Symbiodinium.     

Bleaching and stress in coral reef ecosystems: hsp70 expression by the giant barrel sponge Xestospongia muta

Sponges are a prominent component of coral reef ecosystems. Like reef-building corals, some sponges have been reported to bleach and die. The giant barrel sponge Xestospongia muta is one of the largest and most important components of Caribbean coral reef communities. Tissues of X. muta contain cyanobacterial symbionts of the Synechococcus group. Two types of bleaching have been described: cyclic bleaching, from which sponges recover, and fatal bleaching, which usually results in sponge death. We quantified hsp70 gene expression as an indicator of stress in X. muta undergoing cyclic and fatal bleaching and in response to thermal and salinity variability in both field and laboratory settings. Chlorophyll a content of sponge tissue was estimated to determine whether hsp70 expression was related to cyanobacterial abundance. We found that fatally bleached sponge tissue presented significantly higher hsp70 gene expression, but cyclically bleached tissue did not, yet both cyclic and fatally bleached tissues had lower chlorophyll a concentrations than nonbleached tissue. These results corroborate field observations suggesting that cyclic bleaching is a temporary, nonstressful state, while fatal bleaching causes significant levels of stress, leading to mortality. Our results support the hypothesis that Synechococcus symbionts are commensals that provide no clear advantage to their sponge host. In laboratory experiments, sponge pieces incubated at 30 °C exhibited significantly higher hsp70 expression than control pieces after 1.5 h, with sponge mortality after less than 15 h. In contrast, sponges at different salinities were not significantly stressed after the same period of time. Stress associated with increasing seawater temperatures may result in declining sponge populations in coral reef ecosystems.     

Host-symbiont specificity during onset of symbiosis between the dinoflagellates Symbiodinium spp. and planula larvae of the scleractinian coral Fungia scutaria

Many corals which engage in symbioses with dinoflagellates from the genus Symbiodinium (zooxanthellae) produce offspring which initially lack zooxanthellae. These species must choose their symbionts from numerous genetically distinct strains of zooxanthellae co-occurring in the environment. In most cases, symbiosis onset results in an association between a specific host coral and a specific strain of algal symbiont. This is the first study to examine host-symbiont specificity during symbiosis onset in a larval cnidarian, and the first to examine such events in a scleractinian of any life stage. We infected planula larvae of the solitary Hawaiian scleractinian Fungia scutaria with both homologous zooxanthellae, freshly isolated from F. scutaria adults, and heterologous zooxanthellae, isolated from Montipora verrucosa, Porites compressa, and Pocillopora damicornis, three species of scleractinians which co-occur with F. scutaria. We found that homologous zooxanthellae were better able to establish symbioses with larval hosts than were heterologous isolates, by two separate measures: percent of a larval population infected, and densities of zooxanthellae per larva. We also measured algal densities in larvae over a 4-day period until the onset of settlement and metamorphosis. We found no changes in zooxanthella population densities, regardless of zooxanthella type or the light environment in which they were incubated. Strong infection of host larvae with homologous algae compared to heterologous algae suggests that there is a specificity process which occurs sometime during the early stages of infection between the partners, and which results in the establishment of a specific symbiosis.     

Diversity and expression of nitrogen fixation genes in bacterial symbionts of marine sponges

Marine sponges contain complex assemblages of bacterial symbionts, the roles of which remain largely unknown. We identified diverse bacterial nifH genes within sponges and found that nifH genes are expressed in sponges. This is the first demonstration of the expression of any protein-coding bacterial gene within a sponge. Two sponges Ircinia strobilina and Mycale laxissima were collected from Key Largo, Florida and had delta(15)N values of c. 0-1 per thousand and 3-4 per thousand respectively. The potential for nitrogen fixation by symbionts was assessed by amplification of nifH genes. Diverse nifH genes affiliated with Proteobacteria and Cyanobacteria were detected, and expression of nifH genes affiliated with those from cyanobacteria was detected. The nifH genes from surrounding seawater were similar to those of Trichodesmium and clearly different from the cyanobacterial nifH genes detected in the two sponges. This study advances understanding of the role of bacterial symbionts in sponges and suggests that provision of fixed nitrogen is a means whereby symbionts benefit sponges in nutrient-limited reef environments. Nitrogen fixation by sponge symbionts is possibly an important source of new nitrogen to the reef environment that heretofore has been neglected and warrants further investigation.     

Uric acid deposits in symbiotic marine algae

The symbiosis between cnidarians and dinoflagellate algae is not understood at the cell or molecular level, yet this relationship is responsible for the formation of thousands of square kilometres of coral reefs. We have investigated the nature of crystalline material prominent within marine algal symbionts of Aiptasia sp. anemones. This material, which has historically been considered to be calcium oxalate, is shown to be uric acid. We demonstrate that these abundant uric acid stores can be mobilized rapidly, thereby allowing the algal symbionts to flourish in an otherwise N-poor environment. This is the first report of uric acid accumulation by symbiotic marine algae. These data provide new insight and considerations for understanding the physiological basis of algal symbioses, and represent a new and previously unconsidered aspect of N metabolism in cnidarian, and a variety of other, marine symbioses.     

Metabolic reconstruction of the near complete microbiome of the model sponge Ianthella basta

Many marine sponges host highly diverse microbiomes that contribute to various aspects of host health. Although the putative function of individual groups of sponge symbionts has been increasingly described, the extreme diversity has generally precluded in-depth characterization of entire microbiomes, including identification of syntrophic partnerships. The Indo-Pacific sponge Ianthella basta is emerging as a model organism for symbiosis research, hosting only three dominant symbionts: a Thaumarchaeotum, a Gammaproteobacterium, and an Alphaproteobacterium and a range of other low abundance or transitory taxa. Here, we retrieved metagenome assembled genomes (MAGs) representing >90% of I. basta's microbial community, facilitating the metabolic reconstruction of the sponge's near complete microbiome. Through this analysis, we identified metabolic complementarity between microbes, including vitamin sharing, described the importance of low abundance symbionts, and characterized a novel microbe–host attachment mechanism in the Alphaproteobacterium. We further identified putative viral sequences, highlighting the role viruses can play in maintaining symbioses in I. basta through the horizontal transfer of eukaryotic-like proteins, and complemented this data with metaproteomics to identify active metabolic pathways in bacteria, archaea, and viruses. This data provide the framework to adopt I. basta as a model organism for studying host–microbe interactions and provide a basis for in-depth physiological experiments.     

OXYGEN MICROSENSOR STUDIES ON ZOOXANTHELLATE CLIONAID SPONGES FROM THE COSTA BRAVA,MEDITERRANEAN SEA1

Photosynthetic properties of two symbiotic demosponges were compared using Clark‐type oxygen microsensors. The putatively distinct sponge species, Cliona viridis (Schmidt, 1862) and Cliona nigricans (Schmidt, 1862) were discriminated by their mean megasclere lengths of 296 and 387 μm, respectively. Photosynthetic behavior was used to generate additional taxonomic information. Sponge–dinoflagellate symbioses were well adapted to low light due to the hosts' endolithic lifestyle. Both sponges reached light compensation and saturation at similar light levels with means close to 10 and 30 μmol photons·m^?2·s^?1, respectively. The gross photosynthetic activity was closely related to symbiont cell density in the sponge surface tissue. Mean symbiont densities, chl a content, and gross photosynthesis were about six times higher in C. viridis than in C. nigricans, with respective values of 3000 and 440 symbiont·mm^?2, 1.3 and 0.2 μg chl a·g^?1, and 5.4 and 1.0 μmol O₂·cm^?3·s^?1 gross photosynthesis. Net photosynthesis and respiration could not be calculated accurately from the oxygen gradients, because significant gas exchange occurs through the pumping activity. Thus, assumptions of diffusional oxygen exchange via the surface do not hold for sponges. Combined data of this study indicate that the metabolic activity of C. viridis depends on photosynthetic activity of its symbionts, whereas C. nigricans appears to have a higher pumping intensity and is more actively filter feeding. The difference in photosynthetic activities is not caused by different light adaptations but provides new evidence against the conspecifity of C. viridis and C. nigricans.     

Abundance and reproductive patterns of the excavating sponge Cliona vermifera: a threat to Pacific coral reefs?

Cliona vermifera is a common excavating sponge in coral reefs from the East Pacific. Abundance and reproductive patterns of the sponge in a Mexican Pacific coral reef over a 4-year period are herein described. Sponge abundance was estimated along three transects 50 m long which were randomly placed on the reef, and along each one, a piece of coral rubble and a branch of a live coral from the Pocillopora spp. coral colony closest to the transect were collected at random, approximately every 2 m, yielding 25 pieces of each category per transect (and 75 pieces total of each category). A 2-way ANOVA revealed that invasion was significantly higher in living coral colonies (34.8 %) than in rubble (13.7 %). It also indicated that the abundance in both coralline substrates showed a temporal variation without a clear pattern of increase over the years. It was estimated that 60–85 % of sponges in the population reproduced sexually every year. The sponge proved gonochoristic, with a sex ratio strongly departing from parity (1 male: 3 females). Over the 4-year study period, at least two cohorts of oocytes with densities of up to 3.5 oocytes per mm² tissue were observed. Spermatogenesis lasted about a month, but often producing more than a pulse from July to November, coupled with peaks of oocyte maturation. Fertilization occurred internally to produce encapsulated zygotes that were released in one or more spawning events from July to November. In the following months (December to February), which were the periods of lowest temperature (~18.5–20 °C), no gametic activity occurred in the sponges. Because anomalous temperature rises that are detrimental to corals do not appear to negatively affect the reproduction and abundance of C. vermifera, it is likely that the excavating activity of this sponge may be compromising the health of those coral reefs that are recurrently affected by episodes of thermal stress.     

The magnesium inhibition and arrested phagosome hypotheses: new perspectives on the evolution and ecology of Symbiodinium symbioses

Zooxanthella symbioses are arguably the most important ecological interaction on coral reefs because they energetically subsidize the entire community, and enhance the calcification process that provides structure for all other organisms. While we have developed a detailed understanding of the diversity among and within the Symbiodinium clades, we currently lack a mechanistic explanation for which factors favoured zooxanthella invasion of the intracellular habitat in heterotrophic hosts, and for what molecular mechanisms permit residence within the cell. We propose two hypotheses that explain important evolutionary and ecological features of zooxanthella symbioses. The magnesium inhibition hypothesis (MIH) states that increases in the Mg/Ca ratio in sea water that occurred over the last 100 million years created a situation where Mg(2+) inhibited Ca(2+) transport to zooxanthellae. The MIH predicts, among other things, that the intracellular niche was invaded as a response to this abiotic stressor. The arrested phagosome hypothesis (APH) states that Symbiodinium spp. mimic host cell endosomal digestive machinery via the symbiosome to appear like digesting prey through perpetual release of zooxanthella-derived compounds. The APH represents a subtle but important distinction from previous hypotheses regarding interactions between symbiont and host at the cellular level. The APH predicts that symbionts tune rates of material release to match expectations of host cellular machinery. An outcome of the APH is that intra-host residence time becomes a vital parameter to consider. Both hypotheses shift control of the symbiosis away from the host, and instead focus attention on the niche requirements of Symbiodinium spp.