Sponges and Their Microbiomes Show Similar Community Metrics Across Impacted and Well-Preserved Reefs.

Sponge diversity has been reported to decrease from well-preserved to polluted environments, but whether diversity and intra-species variation of their associated microbiomes also change as function of environmental quality remains unknown. Our study aimed to assess whether microbiome composition and structure are related to the proliferation of some sponges and not others under degraded conditions. We characterized the most frequent sponges and their associated bacteria in two close areas (impacted and well-preserved) of Nha Trang Bay (Indo-Pacific). Sponge assemblages were richer and more diverse in the well-preserved reefs, but more abundant (individuals/m. transect) in the impacted environments, where two species (Clathria reinwardti and Amphimedon paraviridis) dominated. Sponge microbiomes from the polluted zones had, in general, lower bacterial diversity and core size and consequently, higher intra-species dispersion than microbiomes of sponges from the well-preserved environments. Microbial communities reflect the reduction of diversity and richness shown by their host sponges. In this sense, sponges with less complex and more variable microbiomes proliferate under degraded environmental conditions, following the ecological paradigm that negatively correlates community diversity and environmental degradation. Thereby, the diversity and structure of sponge microbiomes might indirectly determine the presence and proliferation of sponge species in certain habitats.

Multipartner Symbiosis across Biological Domains: Looking at the Eukaryotic Associations from a Microbial Perspective.

Sponges establish tight associations with both micro- and macroorganisms. However, while studies on sponge microbiomes are numerous, nothing is currently known about the microbiomes of sponge-associated polychaetes and their relationships with those of their host sponges. We analyzed the bacterial communities of symbiotic polychaetes (Haplosyllis spp.) and their host sponges (Clathria reinwardti, Amphimedon paraviridis, Neofibularia hartmani, and Aaptos suberitoides) to assess the influence of the sponges on the polychaete microbiomes. We identified both eukaryote partners by molecular (16S and COI genes) and morphological features, and we identified their microbial communities by high-throughput sequencing of the 16S rRNA gene (V4 region). We unravel the existence of six Haplosyllis species (five likely undescribed) associated at very high densities with the study sponge species in Nha Trang Bay (central Vietnam). A single polychaete species inhabited A. paraviridis and was different from the single species that inhabited A. suberitoides Conversely, two different polychaete species were found in C. reinwardti and N. hartmani, depending on the two host locations. Regardless of the host sponge, polychaete microbiomes were species specific, which is a widespread feature in marine invertebrates. More than half of the polychaete bacteria were also found in the host sponge microbiome but at contrasting abundances. Thus, the associated polychaetes seemed to be able to select, incorporate, and enrich part of the sponge microbiome, a selection that appears to be polychaete species specific. Moreover, the bacterial diversity is similar in both eukaryotic partners, which additionally confirms the influence of food (host sponge) on the structure of the polychaete microbiome.IMPORTANCE The symbiotic lifestyle represents a fundamental cryptic contribution to the diversity of marine ecosystems. Sponges are ideal targets to improve understanding the symbiotic relationships from evolutionary and ecological points of view, because they are the most ancient metazoans on earth, are ubiquitous in the marine benthos, and establish complex symbiosis with both prokaryotes and animals, which in turn also harbor their own bacterial communities. Here, we study the microbiomes of sponge-polychaete associations and confirm that polychaetes feed on their host sponges. The study worms select and enrich part of the sponge microbiome to shape their own species-specific bacterial communities. Moreover, worm microbiome diversity runs parallel to that of its food host sponge. Considering our results on symbiotic polychaetes and previous studies on fishes and mammals, diet appears to be an important source of bacteria for animals to shape their species-specific microbiomes.

Biodiversity loss in a Mediterranean ecosystem due to an extreme warming event unveils the role of an engineering gorgonian species.

Stochastic perturbations can trigger major ecosystem shifts. Marine systems have been severely affected in recent years by mass mortality events related to positive thermal anomalies. Although the immediate effects in the species demography affected by mortality events are well known, information on the mid- to long-term effects at the community level is much less documented. Here, we show how an extreme warming event replaces a structurally complex habitat, dominated by long-lived species, by a simplified habitat (lower species diversity and richness) dominated by turf-forming species. On the basis of a study involving the experimental manipulation of the presence of the gorgonian Paramuricea clavata, we observed that its presence mitigated the effects of warming by maintaining the original assemblage dominated by macroinvertebrates and delaying the proliferation and spread of the invasive alga Caulerpa cylindracea. However, due to the increase of sediment and turf-forming species after the mortality event we hypothesize a further degradation of the whole assemblage as both factors decrease the recruitment of P.clavata, decrease the survival of encrusting coralligenous-dwelling macroinvertebrates and facilitate the spreading of C. cylindracea.

Description of two new genera (Antarctotetilla, Levantiniella) and a new species of Tetillidae.

Tetillidae is a sponge family distributed all over the world but with some genera apparently endemic from the Antarctic and Subantarctic (the "Antarctic clade"). Species identification results tricky due to the similarities of their morphological characters. However, molecular phylogenies have helped to resolve the family taxonomy. The last phylogenetic study on Tetillidae suggested the creation of two new genera: Levantiniella and Antarctotetilla. Lenvantiniella, from Middle East Mediterranean Sea, was previously classified within Cinachyrella, from which it differs in the small rounded surface cavities, distinctive from true porocalices. Antarctotetilla has up to now an Antarctic distribution, and harbors species wrongly classified within Tethya, Craniella, or Tetilla. The main differences of Antarctotetilla to other Tetillidae genera are the presence of pores grouped in small areas, and a poorly-defined cortex (pseudocortex). This study aims to re-describe in detail the species of Tetillidae that belong in the two above mentioned new genera, and to highlight that molecular phylogenies should be combined with morphological analyses to improve taxonomical decisions. We also describe a new Tetillidae species with a hair-like hispidation, which we name Antarctotetilla pilosa nov. sp. Furthermore, the types of Tethya coactifera and T. crassispicula (Lendenfeld, 1907) were reexamined because of some morphological similarities with Antarctotetilla. The minibarcode sequences (a small COI fragment) placed them within the Antarctic clade harboring Antarctotetilla and Cinachyra, but did not resolve their genus position. A morphological revision, however, suggests placing T. coactifera in Antarctotetilla, while T. crassispicula, which owns porocalices and a spicule-reinforced cortex, appeared to belong in Cinachyra.

Showcasing the role of seawater in bacteria recruitment and microbiome stability in sponges.

We studied the core bacterial communities of 19 sponge species from Nha Trang Bay (Central Vietnam), with particular emphasis on the contribution of planktonic seawater bacteria to the sponge core microbiomes. To ensure consistent sponge-microbe associations and accurate identification of planktonic bacteria transmitted from seawater, we were very restrictive with the definition of the sponge core microbiomes (present in all the replicates), and with the identification of valid biological 16S rRNA gene sequences (100% sequence identity) that belonged to potentially different bacterial taxa. We found a high overlap (>50% relative abundance) between the sponge species core microbiome and the seawater bacterial core in ca. a half of the studied species, including representatives of both, HMA and LMA sponges. From our restrictive analysis, we point to horizontal transmission as a relevant way of symbiont acquisition in sponges. Some species-specific recognition mechanisms may act in sponges to enrich specific seawater bacteria in their tissues. These mechanisms would allow the maintenance of bacterial communities in a species across geographical ranges. Moreover, besides contrasting preferences in bacteria selection from seawater, divergent physiological traits may also account for the different microbiomes in species of HMA and LMA sponges.

-Reproductive strategies of two common sympatric Mediterranean sponges: Dysidea avara (Dictyoceratida) and Phorbas tenacior (Poecilosclerida).

Despite their abundance in benthic ecosystems, life cycles and reproductive features of most sponge species remain unknown. We have studied the main reproductive features of two demosponges, Dysidea avara and Phorbas tenacior, belonging to phylogenetically distant groups: Orders Dictyoceratida and Poecilosclerida, respectively. Both sponges are abundant and share habitat in the Mediterranean rocky sublittoral. They brood parenchymella larvae with different morphology and behaviour. Sampling was conducted monthly over a two-year period in a locality where both species coexist. The two species reproduced in spring-summer, and presented species-specific reproductive features despite being subject to the same environmental conditions. D. avara has a shorter reproductive period than P. tenacior, ending before the peak of temperature in summer, while the reproductive period of P. tenacior lasts until beginning of autumn. Brooding larvae were present in June-July in D. avara, and in August-October in P. tenacior. Larval size, reproductive effort and number of larvae produced (measured the month with the maximum production) were significantly higher in D. avara than in P. tenacior. A higher reproductive effort and larval traits point to a more opportunistic life strategy in D. avara than in P. tenacior. A lack of overlap in the timing of larval release, as well as different reproductive traits, may reduce competition and facilitate the coexistence of these two sympatric and abundant sponges.

Contrasting biological features in morphologically cryptic Mediterranean sponges.

Sponges are key organisms in the marine benthos where they play essential roles in ecological processes such as creating new niches, competition for resources, and organic matter recycling. Despite the increasing number of taxonomical studies, many sponge species remain hidden, whether unnoticed or cryptic. The occurrence of cryptic species may confound ecological studies by underestimating biodiversity. In this study, we monitored photographically growth, fusions, fissions, and survival of two morphologically cryptic species Hemimycale mediterranea Uriz, Garate & Agell, 2017 and H. columella (Bowerbank, 1874). Additionally, we characterized the main environmental factors of the corresponding species habitats, trying to ascertain whether some abiotic factors were correlated with the distribution of these species. Sponge monitoring was performed monthly. Seawater samples were collected the same monitoring days in the vicinity of the target sponges. Results showed contrasting growth and survival patterns for each species: H. mediterranea totally disappeared after larval release while 64% of individuals of H. columella survived the entire two years we monitored. The species also differed in the number of fissions and fusions. These events were evenly distributed throughout the year in the H. mediterranea population but concentrated in cold months in H. columella. No measured environmental factor correlated with H. mediterranea growth rates, while temperature and dissolved organic nitrogen were negatively correlated with H. columella growth rates. The strong differences in depth distribution, survival, growth, fusions, and fissions found between these two cryptic species, highlights the importance of untangling cryptic species before ecological studies are performed in particular when these species share geographical distribution.

Redescription and establishment of a holotype and three paratypes for the species Hemimycale mediterranea sp. nov.

BACKGROUND: In a recent paper, we described a new sponge species named Hemimycale mediterranea Uriz, Garate & Agell, 2017. However, we failed to designate a holotype and a type locality, as required by the International Commission on Zoological Nomenclature (ICZN). Although the validity of the previous conclusions remains unchanged, the species name cannot be considered available according to ICZN regulations until a holotype is designated. RESULTS: The present work fulfills the requirements of the ICZN by designating a holotype, three paratypes and the type locality for the new species Hemimycale mediterranea and has been registered in ZooBank.

Endosymbiotic calcifying bacteria across sponge species and oceans.

From an evolutionary point of view, sponges are ideal targets to study marine symbioses as they are the most ancient living metazoans and harbour highly diverse microbial communities. A recently discovered association between the sponge Hemimycale columella and an intracellular bacterium that generates large amounts of calcite spherules has prompted speculation on the possible role of intracellular bacteria in the evolution of the skeleton in early animals. To gain insight into this purportedly ancestral symbiosis, we investigated the presence of symbiotic bacteria in Mediterranean and Caribbean sponges. We found four new calcibacteria OTUs belonging to the SAR116 in two orders (Poecilosclerida and Clionaida) and three families of Demospongiae, two additional OTUs in cnidarians and one more in seawater (at 98.5% similarity). Using a calcibacteria targeted probe and CARD-FISH, we also found calcibacteria in Spirophorida and Suberitida and proved that the calcifying bacteria accumulated at the sponge periphery, forming a skeletal cortex, analogous to that of siliceous microscleres in other demosponges. Bacteria-mediated skeletonization is spread in a range of phylogenetically distant species and thus the purported implication of bacteria in skeleton formation and evolution of early animals gains relevance.

Molecular phylogenies confirm the presence of two cryptic Hemimycale species in the Mediterranean and reveal the polyphyly of the genera Crella and Hemimycale (Demospongiae: Poecilosclerida).

BACKGROUND: Sponges are particularly prone to hiding cryptic species as their paradigmatic plasticity often favors species phenotypic convergence as a result of adaptation to similar habitat conditions. Hemimycale is a sponge genus (Family Hymedesmiidae, Order Poecilosclerida) with four formally described species, from which only Hemimycale columella has been recorded in the Atlanto-Mediterranean basin, on shallow to 80 m deep bottoms. Contrasting biological features between shallow and deep individuals of Hemimycale columella suggested larger genetic differences than those expected between sponge populations. To assess whether shallow and deep populations indeed belong to different species, we performed a phylogenetic study of Hemimycale columella across the Mediterranean. We also included other Hemimycale and Crella species from the Red Sea, with the additional aim of clarifying the relationships of the genus Hemimycale. METHODS: Hemimycale columella was sampled across the Mediterranean, and Adriatic Seas. Hemimycale arabica and Crella cyathophora were collected from the Red Sea and Pacific. From two to three specimens per species and locality were extracted, amplified for Cytochrome C Oxidase I (COI) (M1-M6 partition), 18S rRNA, and 28S (D3-D5 partition) and sequenced. Sequences were aligned using Clustal W v.1.81. Phylogenetic trees were constructed under neighbor joining (NJ), Bayesian inference (BI), and maximum likelihood (ML) criteria as implemented in Geneious software 9.01. Moreover, spicules of the target species were observed through a Scanning Electron microscope. RESULTS: The several phylogenetic reconstructions retrieved both Crella and Hemimycale polyphyletic. Strong differences in COI sequences indicated that C. cyathophora from the Red Sea might belong in a different genus, closer to Hemimycale arabica than to the Atlanto-Mediterranean Crella spp. Molecular and external morphological differences between Hemimycale arabica and the Atlanto-Mediterranean Hemimycale also suggest that Hemimycale arabica fit in a separate genus. On the other hand, the Atlanto-Mediterranean Crellidae appeared in 18S and 28S phylogenies as a sister group of the Atlanto-Mediterranean Hemimycale. Moreover, what was known up to now as Hemimycale columella, is formed by two cryptic species with contrasting bathymetric distributions. Some small but consistent morphological differences allow species distinction. CONCLUSIONS: A new family (Hemimycalidae) including the genus Hemimycale and the two purported new genera receiving C. cyathophora and Hemimycale arabica might be proposed according to our phylogenetic results. However, the inclusion of additional Operational Taxonomic Unit (OTUs) appears convenient before taking definite taxonomical decisions. A new cryptic species (Hemimycale mediterranea sp. nov.) is described. Morphologically undifferentiated species with contrasting biological traits, as those here reported, confirm that unidentified cryptic species may confound ecological studies.

Correction: Phylogenetic Reassessment of Antarctic Tetillidae (Demospongiae, Tetractinellida) Reveals New Genera and Genetic Similarity among Morphologically Distinct Species.

[This corrects the article DOI: 10.1371/journal.pone.0160718.].

Phylogenetic Reassessment of Antarctic Tetillidae (Demospongiae, Tetractinellida) Reveals New Genera and Genetic Similarity among Morphologically Distinct Species.

Species of Tetillidae are distributed worldwide. However, some genera are unresolved and only a few genera and species of this family have been described from the Antarctic. The incorporation of 25 new COI and 18S sequences of Antarctic Tetillidae to those used recently for assessing the genera phylogeny, has allowed us to improve the resolution of some poorly resolved nodes and to confirm the monophyly of previously identified clades. Classical genera such as Craniella recovered their traditional diagnosis by moving the Antarctic Tetilla from Craniella, where they were placed in the previous family phylogeny, to Antarctotetilla gen. nov. The morphological re-examination of specimens used in the previous phylogeny and their comparison to the type material revealed misidentifications. The proposed monotypic new genus Levantiniella had uncertain phylogenetic relationships depending on the gene partition used [corrected]. Two more clades would require the inclusion of additional species to be formally established as new genera. The parsimony tree based on morphological characters and the secondary structure of the 18S (V4 region) almost completely matched the COI M1-M6 and the COI+18S concatenated phylogenies. Morphological synapomorphies have been identified for the genera proposed. New 15 28S (D3-D5) and 11 COI I3-M11 partitions were exclusively sequenced for the Antarctic species subset. Remarkably, species within the Antarctic genera Cinachyra (C. barbata and C. antarctica) and Antarctotetilla (A. leptoderma, A. grandis, and A. sagitta), which are clearly distinguishable morphologically, were not genetically differentiated with any of the markers assayed. Thus, as it has been reported for other Antarctic sponges, both the mitochondrial and nuclear partitions used did not differentiate species that were well characterized morphologically. Antarctic Tetillidae offers a rare example of genetically cryptic (with the traditional markers used for sponges), morphologically distinct species.

Snapshot of a Bacterial Microbiome Shift during the Early Symptoms of a Massive Sponge Die-Off in the Western Mediterranean.

Ocean warming is affecting marine benthic ecosystems through mass mortality events that involve marine invertebrates, in particular bivalves, corals, and sponges. Among these events, extensive die-offs of Ircinia fasciculata sponges have been recurrently reported in western Mediterranean. The goal of our study was to test whether the temperature-related mass sponge die-offs were associated with or preceded by an early unbalanced bacterial microbiome in the sponge tissues. We took advantage of the early detection of disease and compared the microbiomes of healthy vs. early diseased I. fasciculata tissues. Our results showed a microbiome shift in early diseased tissues. The abundance of Gammaproteobacteria and Acidobacteria increased and that of Deltaproteobacteria decreased in diseased vs. healthy tissues. The change in community composition was also noticeable at the operational taxonomic unit (OTU) level. Diseased tissues contained more bacterial sequences previously identified in injured or stressed sponges and corals than healthy tissues. Bacterial diversity increased significantly in diseased tissues, which contained a higher number of low abundance OTUs. Our results do not support the hypothesis of one particular pathogen, whether a Vibrio or any other bacteria, triggering the Northwestern Mediterranean mass mortalities of I. fasciculata. Our data rather suggest an early disruption of the bacterial microbiome balance in healthy sponges through a shift in OTU abundances, and the purported consequent decline of the sponge fitness and resistance to infections. Opportunistic bacteria could colonize the sponge tissues, taking benefit of the sponge weakness, before one or more virulent pathogens might proliferate ending in the mass sponge die-off.

High genetic diversity, phenotypic plasticity, and invasive potential of a recently introduced calcareous sponge, fast spreading across the Atlanto-Mediterranean basin.

Sponges are considered poor invaders, and no genetic studies on introduced sponges have been performed up to now. Paraleucilla magna is the first calcareous sponge introduced to the Mediterranean and Northeastern Atlantic. The study aimed at investigating the genetic makeup and connectivity of the introduced populations of P. magna and at exploring signs of local phenotypic adaptation, to gain insight on the species invasive potential. Ten populations along the species introduction range (Brazil, Açores, Madeira, and continental Europe) were genetically characterized by using nine microsatellite markers. Most populations were genetically structured as suggested by significant Dst and Fst values, significant differences among populations (AMOVA) and the presence of private alleles. The analyzed populations belonged to three genetically homogeneous groups (K) according to the Bayesian algorithm (structure software) and the UPGMA dendrogram. Genetic diversity within populations was higher than expected. Recurrent introductions of non-randomly selected individuals from the native sources may have contributed to the heterozygote deficit found in all populations by forming pedigree structures with mating among relatives. Moreover, the species biological cycle was monitored in a population established on native Mediterranean assemblages (41°40'27″N, 2°47'25″E) and compared with the species cycle in other habitats. Contrasting life spans, growth habits, and reproduction cycles, depending on the habitat conditions, were recorded. To summarize, high genetic diversity, phenotypic local adaptation, and high reproduction rates altogether allow predicting the fast proliferation of P. magna in newly colonized regions and point to its strong invasive potential.

Similar sponge-associated bacteria can be acquired via both vertical and horizontal transmission.

Marine sponges host diverse communities of microorganisms that are often vertically transmitted from mother to oocyte or embryo. Horizontal transmission has often been proposed to co-occur in marine sponges, but the mechanism is poorly understood. To assess the impact of the mode of transmission on the microbial assemblages of sponges, we analysed the microbiota in sympatric sponges that have previously been reported to acquire bacteria via either vertical (Corticium candelabrum and Crambe crambe) or horizontal transmission (Petrosia ficiformis). The comparative study was performed by polymerase chain reaction-denaturing gradient gel electrophoresis and pyrosequencing of barcoded PCR-amplified 16S rRNA gene fragments. We found that P. ficiformis and C. candelabrum each harbour their own species-specific bacteria, but they are similar to other high-microbial-abundance sponges, while the low-microbial-abundance sponge C. crambe hosts microbiota of a very different phylogenetic signature. In addition, nearly 50% of the reads obtained from P. ficiformis were most closely related to bacteria that were previously reported to be vertically transmitted in other sponges and comprised vertical-horizontal transmission phylogenetic clusters (VHT clusters). Therefore, our results provide evidence for the hypothesis that similar sponge-associated bacteria can be acquired via both vertical and horizontal transmission.

Removing environmental sources of variation to gain insight on symbionts vs. transient microbes in high and low microbial abundance sponges.

In this study, we pursue unravelling the bacterial communities of 26 sponges, belonging to several taxonomical orders, and comprising low microbial abundance (LMA) and high microbial abundance (HMA) representatives. Particularly, we searched for species-specific bacteria, which could be considered as symbionts. To reduce temporal and spatial environmentally caused differences between host species, we sampled all the sponge species present in an isolated small rocky area in a single dive. The bacterial communities identified by pyrosequencing the 16S rRNA gene showed that all HMA species clustered separated from LMA sponges and seawater. HMA sponges often had highest diversity, but some LMA sponges had also very diverse bacterial communities. Network analyses indicated that no core bacterial community seemed to exist for the studied sponges, not even for such a space and time-restricted sampling. Most sequences, particularly the most abundant ones in each species, were species-specific for both HMA and LMA sponges. The bacterial sequences retrieved from LMA sponges, despite being phylogenetically more similar to seawater, did not represent transient seawater bacteria. We conclude that sponge bacterial communities depend more on the host affiliation to the HMA or LMA groups than on host phylogeny.

Endosymbiotic calcifying bacteria: a new cue to the origin of calcification in metazoa?

Sponges show the highest diversity of associated bacteria among marine invertebrates. Immunological evidence traces the origin of the sponge bacterial symbioses to the Precambrian era. Hence, sponges appear to be ideally suited for studying the evolutionary origins of prokaryote-metazoan associations. Sponges produce either calcareous or siliceous skeletons, which only coexist in a relict group of demosponges, the sclerosponges. We report here, for the first time, intensive calcification in nonsclerosponge siliceous demosponges. Calcification is mediated by endosymbiotic bacteria (calcibacteria) located in archeocyte-like sponge cells. These calcibacteria are devoid of bacterial walls and divide within sponge cells until they became surrounded by a calcitic sheet, being subsequently extruded to the sponge subectosomal (subepithelial) zone. Thousands of bacteria-produced calcitic spherules cover the surface of the host sponges, forming a cortex-like structure that mimics a rudimentary peripheral skeleton. Calcibacteria are vertically transferred to the sponge larvae during embryogenesis. Calcium detoxification may have generated this symbiotic association, with some additional benefits for the sponges, such as skeletal formation and deterrence from predation. This unique symbiosis holds implications for sponge biology and may advance discussions on the role of bacteria in early biocalcification processes in metazoans.

Sponge ecology in the molecular era.

Knowledge of the functioning, health state, and capacity for recovery of marine benthic organisms and assemblages has become essential to adequately manage and preserve marine biodiversity. Molecular tools have allowed an entirely new way to tackle old and new questions in conservation biology and ecology, and sponge science is following this lead. In this review, we discuss the biological and ecological studies of sponges that have used molecular markers during the past 20 years and present an outlook for expected trends in the molecular ecology of sponges in the near future. We go from (1) the interface between inter- and intraspecies studies, to (2) phylogeography and population level analyses, (3) intra-population features such as clonality and chimerism, and (4) environmentally modulated gene expression. A range of molecular markers has been assayed with contrasting success to reveal cryptic species and to assess the genetic diversity and connectivity of sponge populations, as well as their capacity to respond to environmental changes. We discuss the pros and cons of the molecular gene partitions used to date and the prospects of a plentiful supply of new markers for sponge ecological studies in the near future, in light of recently available molecular technologies. We predict that molecular ecology studies of sponges will move from genetics (the use of one or some genes) to genomics (extensive genome or transcriptome sequencing) in the forthcoming years and that sponge ecologists will take advantage of this research trend to answer ecological and biological questions that would have been impossible to address a few years ago.