Suchergebnisse

Marine heat waves (MHWs) are periods of extremely high seawater temperature that affect marine ecosystems in several ways. Anthozoans (corals and gorgonians) and Porifera (sponges) are usually among the taxa most affected by MHWs. Both are holobiont entities that form complex interactions with a wide range of microbes, which are an essential part of these organisms and play key roles in their health status. Here, we determine microbial community changes suffered in two corals (Cladocora caespitosa and Oculina patagonica), one gorgonian (Leptogorgia sarmentosa) and one sponge (Sarcotragus fasciculatus) during the 2015 MHW. The microbial communities were different among hosts and displayed shifts related to host health status, with a higher abundance in necrosed tissues of Ruegeria species or of potential pathogens like Vibrio. We also carry out a meta-analysis using 93 publicly accessible 16S rRNA gene libraries from O. patagonica, C. caespitosa and L. sarmentosa to establish a Mediterranean core microbiome in these species. We have identified one Ruegeria OTU that maintained a stable and consistent association with these species, which was also related to tissue necrosis in their hosts. Therefore, Ruegeria sp. could play an important and still underexplored role in the health status of its hosts. ; This work has been carried out within the CIESM project "Tropical Signals" and it was funded by the European Union's framework program Horizon 2020 (LEIT-BIO-2015-685474, Metafluidics, to JA).

The interactions between viruses and their microbial hosts play a central role in the control of microbial communities in nature. However, the study of such interactions within the uncultured majority is technically very challenging. Here, we review how microarray tools can be used to analyze the interactions between viruses and their microbial hosts in nature, away from laboratory pure culture-based models. We show examples of how DNA arrays have been used to study the expression of viral assemblages in natural samples, and to assign viruses to hosts within uncultured communities. Finally, we briefly discuss the possibilities of protein and glycan arrays to gain insight into the ways microbes interact with their viruses. ; Our current studies with viral microarrays are supported by projects CGL2012-39627-C03-01 (to Josefa Antón) and AYA2011-24803 (to Víctor Parro) of the Spanish Ministry of Science and Innovation, which are co-financed with FEDER support from the European Union.

Homologous recombination (HR) enables the exchange of genetic material between and within species. Recent studies suggest that this process plays a major role in the microevolution of microbial genomes, contributing to core genome homogenization and to the maintenance of cohesive population structures. However, we still have a very poor understanding of the possible adaptive roles of intraspecific HR and of the factors that determine its differential impact across clades and lifestyles. Here we used a unified methodological framework to assess HR in 338 complete genomes from 54 phylogenetically diverse and representative prokaryotic species, encompassing different lifestyles and a broad phylogenetic distribution. Our results indicate that lifestyle and presence of restriction-modification (RM) machineries are among the main factors shaping HR patterns, with symbionts and intracellular pathogens having the lowest HR levels. Similarly, the size of exchanged genomic fragments correlated with the presence of RM and competence machineries. Finally, genes exchanged by HR showed functional enrichments which could be related to adaptations to different environments and ecological strategies. Taken together, our results clarify the factors underlying HR impact and suggest important adaptive roles of genes exchanged through this mechanism. Our results also revealed that the extent of genetic exchange correlated with lifestyle and some genomic features. Moreover, the genes in exchanged regions were enriched for functions that reflected specific adaptations, supporting identification of HR as one of the main evolutionary mechanisms shaping prokaryotic core genomes. ; The group of J.A. is funded by grant CLG2015_66686-C3-03 from the Spanish Ministry of Economy and Competitiveness (MINECO), which is cofinanced with FEDER support from the European Union. P.G.-T. was an FPI-MINECO fellow associated with project CGL2012-39627-C03-01 (to J.A.). The T.G. group acknowledges support from the Spanish Ministry of Economy and Competitiveness (grants "Centro de Excelencia Severo Ochoa 2013-2017" SEV-2012-0208 and BFU2015-67107; cofounded by European Regional Development Fund [ERDF]); from the CERCA Program/Generalitat de Catalunya; from the Catalan Research Agency (AGAUR) (grant SGR857); and from the European Union's Horizon 2020 research and innovation program under grant agreement ERC-2016-724173 (Marie Sklodowska-Curie grant agreement no. H2020-MSCA-ITN-2014-642095).

Posidonia oceanica is a long-living and very slow-growing marine seagrass endemic to the Mediterranean Sea. It produces large amounts of leaf material and rhizomes, which can reach the shore and build important banks known as "banquettes." In recent years, interest in the potential uses of these P. oceanica banquettes has increased, and it was demonstrated that biomass extracts showed antioxidant, antifungal, and antiviral activities. The discovery of new compounds through the culture of microorganisms is limited, and to overcome this limitation, we performed a metagenomic study to investigate the microbial community associated with P. oceanica banquettes. Our results showed that the microbial community associated with P. oceanica banquettes was dominated by Alphaproteobacteria, Gammaproteobacteria, Bacteroidetes, and Cyanobacteria. Pseudoalteromonas was the dominant genus, followed by Alteromonas, Labrenzia, and Aquimarina. The metagenome reads were binned and assembled into 23 nearly complete metagenome-assembled genomes (MAGs), which belonged to new families of Cyanobacteria, Myxococcota, and Granulosicoccaceae and also to the novel genus recently described as Gammaproteobacteria family UBA10353. A comparative analysis with 60 published metagenomes from different environments, including seawater, marine biofilms, soils, corals, sponges, and hydrothermal vents, indicated that banquettes have numbers of natural products and carbohydrate active enzymes (CAZymes) similar to those found for soils and were only surpassed by marine biofilms. New proteins assigned to cellulosome modules and lignocellulose-degrading enzymes were also found. These results unveiled the diverse microbial composition of P. oceanica banquettes and determined that banquettes are a potential source of bioactive compounds and novel enzymes. ; This work was carried out within the project Metafluidics, funded by the European Union's framework program Horizon 2020 (grant no. LEIT‐BIO‐2015‐685474 to J.A.).

Viruses control natural microbial communities. Identification of virus–host pairs relies either on their cultivation or on metagenomics and tentative assignment based on genomic signatures. Both approaches have severe drawbacks when aiming to target such pairs within the uncultured majority. Here we present an unambiguous way to assign viruses to hosts that does not rely on any previous information about either of them nor requires their cultivation. First, genomic contents of individual cells present in an environmental sample are retrieved by means of single-cell genomic technologies. Then, individual cell genomes are hybridized against a set of individual viral genomes from the same sample, previously immobilized on a microarray. Infected cells will yield positive hybridization as they carry viral genomes, which can be then sequenced and characterized. Using this method, we pinpoint viruses infecting the ubiquitous hyperhalophilic Nanohaloarchaeota, included in the so-called 'microbial dark matter' (the uncultured fraction of the microbial world). ; This work was supported by the projects CGL2012-39627-C03-01 (to J.A.) and AYA2011-24803 (to V.P.) of the Spanish Ministry of Science and Innovation, which are co-financed with FEDER support from the European Union.

The endemic Mediterranean zooxanthellate scleractinian reef-builder Cladocora caespitosa is among the organisms most affected by warming-related mass mortality events in the Mediterranean Sea. Corals are known to contain a diverse microbiota that plays a key role in their physiology and health. Here we report the first study that examines the microbiome and pathobiome associated with C. caespitosa in three different Mediterranean locations (i.e., Genova, Columbretes Islands, and Tabarca Island). The microbial communities associated with this species showed biogeographical differences, but shared a common core microbiome that probably plays a key role in the coral holobiont. The putatively pathogenic microbial assemblage (i.e., pathobiome) of C. caespitosa also seemed to depend on geographic location and the human footprint. In locations near the coast and with higher human influence, the pathobiome was entirely constituted by Vibrio species, including the well-known coral pathogens Vibrio coralliilyticus and V. mediterranei. However, in the Columbretes Islands, located off the coast and the most pristine of the analyzed locations, no changes among microbial communities associated to healthy and necrosed samples were detected. Hence, our results provide new insights into the microbiome of the temperate corals and its role in coral health status, highlighting its dependence on the local environmental conditions and the human footprint. ; This work was funded by the European Union's framework program Horizon 2020 (LEIT - BIO - 2015 - 685474, Metafluidics, to JA) and the Ministerio de Economía y Competitividad Smart project (CGL2012-32194, to CL).

Microbial opsin (i.e. retinal-binding protein) dynamics has been studied along a salinity gradient in Santa Pola solar salterns (Alicante, Spain) by using culture-independent approaches and statistical analyses. Five ponds of salinities ranging from 18% to above 40% were sampled nine times along a year. Forty-three opsin-like sequences were retrieved by denaturing gradient gel electrophoresis and clustered into 18 different phylogroups, indicating that their diversity was higher than expected according to previous data. Moreover, the statistical correlation between environmental factors controlling microbial community structure and dynamics of environmental rhodopsin proteins indicated almost identical temporal fluctuations between the opsin-related sequences and their corresponding putative 'producers' in nature. Although most sequences were related to others previously detected in hypersaline environments, some pond-specific opsins putatively belonged to previously uncharacterized hosts. Furthermore, we propose that subtle changes in the bacteriorhodopsin 'retinal proton binding pocket', which is key in the photocycle function, could be the molecular basis behind a fine 'photocycle-tuning' mechanism to avoid inter/intraspecies light-competition in hypersaline environments. ; This work was supported by project CGL2012-39627-C03-01 of the Spanish Ministry of Economy and Competitiveness, which was also co-funded with FEDER support from the European Union.

Comparative genomics, metagenomics, and single-cell technologies have shown that populations of microbial species encompass assemblages of closely related strains. This raises the question of whether individual bacterial lineages respond to the presence of their close relatives by modifying their gene expression or, instead, whether assemblages simply act as the arithmetic addition of their individual components. Here, we took advantage of transcriptome sequencing to address this question. For this, we analyzed the transcriptomes of two closely related strains of the extremely halophilic bacterium Salinibacter ruber grown axenically and in coculture. These organisms dominate bacterial assemblages in hypersaline environments worldwide. The strains used here cooccurred in the natural environment and are 100% identical in their 16S rRNA genes, and each strain harbors an accessory genome representing 10% of its complete genome. Overall, transcriptomic patterns from pure cultures were very similar for both strains. Expression was detected along practically the whole genome albeit with some genes at low levels. A subset of genes was very highly expressed in both strains, including genes coding for the light-driven proton pump xanthorhodopsin, genes involved in the stress response, and genes coding for transcriptional regulators. Expression differences between pure cultures affected mainly genes involved in environmental sensing. When the strains were grown in coculture, there was a modest but significant change in their individual transcription patterns compared to those in pure culture. Each strain sensed the presence of the other and responded in a specific manner, which points to fine intraspecific transcriptomic modulation. ; The group of J.A. is funded by grant CGL2012-39627-C03-01 from the Spanish Ministry of Economy and Competitiveness (MINECO), which is cofinanced with FEDER support from the European Union. P.G.-T. was an FPI-MINECO fellow. Research by the group of T.G. is funded in part by a grant from the ...

The culturable fraction of aerobic, heterotrophic and extremely halophilic microbiota retrieved from sediment and brine samples of eight sampling sites in the Mediterranean, Canary Islands and Chile was studied by means of a tandem approach combining large-scale cultivation, MALDI-TOF MS targeting whole cell biomass, and phylogenetic reconstruction based on 16S rRNA gene analysis. The approach allowed the identification of more than 4200 strains and a comparison between different sampling sites. The results indicated that the method constituted an excellent tool for the discovery of taxonomic novelty. Four new genera and nine new species could be identified within the archaeal family Halobacteriaceae, as well as one new bacterial species, and a representative of Salinibacter ruber phylotype II, a group that had been refractory to isolation for the last fifteen years. Altogether, the results indicated that in order to provide better yields for the retrieval of novel taxa from the environment, performance of non-redundant environment sampling is recommended together with the screening of large sets of strains. ; The current study was funded with the scientific support given by the Spanish Ministry of Economy through the projectsCGL2012-39627-C03-01 and CGL2012-39627-C03-03, which were also supported with European Regional Development Fund (FEDER) funds, and the preparatory phase of the Microbial Resource Research Infrastructure (MIRRI) funded by the EU (grant number 312251). In addition, the funding from competitive research groups (Microbiology) of the Government of the Balearic Islands (also co-supported with FEDER funds), is also acknowledged. TVP acknowledges the predoctoral fellowship of the Ministerio de Economía y Competitividad of the Spanish Government for the FPI fellowship (Nr BES-2013-064420) supporting his research activities.

The increase in seawater temperature associated with global warming is a significant threat to coral health and is linked to increasing mass mortality events and Vibrio-related coral diseases. In the Mediterranean Sea, the endemic Cladocora caespitosa and the invasive species Oculina patagonica are the main scleractinian corals affected by mass mortalities. In this study, culturable Vibrio spp. assemblages associated with healthy and unhealthy colonies of these two shallow coral species were characterized to assess the presence of Vibrio pathogens in tissue necrosis. Vibrio communities associated with O. patagonica and C. caespitosa showed geographical differences, although these became more homogeneous in unhealthy specimens of both species. Furthermore, the number of recovered Vibrio specimens was more than five times higher in unhealthy than in healthy corals. Within these culturable vibrios, the known pathogens Vibrio mediterranei and Vibrio coralliilyticus were present in unhealthy colonies of both coral species in the two localities, suggesting that they could play a role in the health status of C. caespitosa and thus act as generalist pathogens in Mediterranean corals. Nonetheless, a clonal type of V. coralliilyticus detected in C. caespitosa was not associated with disease signs, suggesting that this species could encompass assemblages with different levels of virulence. ; This work was funded by the European Union's Horizon 2020 framework program (LEIT-BIO-2015-685474, Metafluidics, to JA) and the grant CLG2015 66686-C3-3 (to JA) of the Spanish Ministry of Economy and Competitiveness, which was also co-financed with FEDER support from the European Union.

The microbiota of multi-pond solar salterns around the world has been analyzed using a variety of culture-dependent and molecular techniques. However, studies addressing the dynamic nature of these systems are very scarce. Here we have characterized the temporal variation during 1 year of the microbiota of five ponds with increasing salinity (from 18% to >40%), by means of CARD-FISH and DGGE. Microbial community structure was statistically correlated with several environmental parameters, including ionic composition and meteorological factors, indicating that the microbial community was dynamic as specific phylotypes appeared only at certain times of the year. In addition to total salinity, microbial composition was strongly influenced by temperature and specific ionic composition. Remarkably, DGGE analyses unveiled the presence of most phylotypes previously detected in hypersaline systems using metagenomics and other molecular techniques, such as the very abundant Haloquadratum and Salinibacter representatives or the recently described low GC Actinobacteria and Nanohaloarchaeota. In addition, an uncultured group of Bacteroidetes was present along the whole range of salinity. Database searches indicated a previously unrecognized widespread distribution of this phylotype. Single-cell genome analysis of five members of this group suggested a set of metabolic characteristics that could provide competitive advantages in hypersaline environments, such as polymer degradation capabilities, the presence of retinal-binding light-activated proton pumps and arsenate reduction potential. In addition, the fairly high metagenomic fragment recruitment obtained for these single cells in both the intermediate and hypersaline ponds further confirm the DGGE data and point to the generalist lifestyle of this new Bacteroidetes group. ; This work was supported by the projects CGL2012-39627-C03-01 and 02 of the Spanish Ministry of Economy and Competitiveness, which were also co-financed with FEDER support from the European Union. ...

Corals are known to contain a diverse microbiota that plays a paramount role in the physiology and health of holobiont. However, few studies have addressed the variability of bacterial communities within the coral host. In this study, bacterial community composition from the mucus, tissue and skeleton of the scleractinian coral Oculina patagonica were investigated seasonally at two locations in the Western Mediterranean Sea, to further understand how environmental conditions and the coral microbiome structure are related. We used denaturing gradient gel electrophoresis in combination with next-generation sequencing and electron microscopy to characterize the bacterial community. The bacterial communities were significantly different among coral compartments, and coral tissue displayed the greatest changes related to environmental conditions and coral health status. Species belonging to the Rhodobacteraceae and Vibrionaceae families form part of O. patagonica tissues core microbiome and may play significant roles in the nitrogen cycle. Furthermore, sequences related to the coral pathogens, Vibrio mediterranei and Vibrio coralliilyticus, were detected not only in bleached corals but also in healthy ones, even during cold months. This fact opens a new view onto unveiling the role of pathogens in the development of coral diseases in the future. ; This work was supported by the projects CGL2012-39627-C03-01 and CGL2015-66686-C3-3-P (to JA) of the Spanish Ministry of Economy and Competitiveness, that include FEDER funds from the European Union.

Cotylorhiza tuberculata is an important scyphozoan jellyfish producing population blooms in the Mediterranean probably due to pelagic ecosystem's decay. Its gastric cavity can serve as a simple model of microbial–animal digestive associations, yet poorly characterized. Using state-of-the-art metagenomic population binning and catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH), we show that only four novel clonal phylotypes were consistently associated with multiple jellyfish adults. Two affiliated close to Spiroplasma and Mycoplasma genera, one to chlamydial 'Candidatus Syngnamydia', and one to bacteroidetal Tenacibaculum, and were at least one order of magnitude more abundant than any other bacteria detected. Metabolic modelling predicted an aerobic heterotrophic lifestyle for the chlamydia, which were found intracellularly in Onychodromopsis-like ciliates. The Spiroplasma-like organism was predicted to be an anaerobic fermenter associated to some jellyfish cells, whereas the Tenacibaculum-like as free-living aerobic heterotroph, densely colonizing the mesogleal axis inside the gastric filaments. The association between the jellyfish and its reduced microbiome was close and temporally stable, and possibly related to food digestion and protection from pathogens. Based on the genomic and microscopic data, we propose three candidate taxa: 'Candidatus Syngnamydia medusae', 'Candidatus Medusoplasma mediterranei' and 'Candidatus Tenacibaculum medusae'. ; This research from RRM's group was partially supported by the Spanish Ministry of Economy projects CGL2012-39627-C03-03 and CLG2015_66686-C3-1-P, which were also supported with European Regional Development Fund (FEDER) funds. KTK's research was supported, in part, by the U.S. National Science Foundation (Award No. 1241046). RRM acknowledges the economic support of grant PR2015-00008 included in the program Salvador de Madariaga of the Ministry of Education, Culture and Sports in order to undertake a research stay at the MPI-MM in Bremen. TVP acknowledges the predoctoral fellowship of the Ministerio de Economía y Competitividad of the Spanish Government for the FPI fellowship (Nr BES-2013-064420) supporting his research activities.

The application of tandem MALDI-TOF MS screening with 16S rRNA gene sequencing of selected isolates has been demonstrated to be an excellent approach for retrieving novelty from large-scale culturing. The application of such methodologies in different hypersaline samples allowed the isolation of the culture-recalcitrant Salinibacter ruber second phylotype (EHB-2) for the first time, as well as a new species recently isolated from the Argentinian Altiplano hypersaline lakes. In this study, the genome sequences of the different species of the phylum Rhodothermaeota were compared and the genetic repertoire along the evolutionary gradient was analyzed together with each intraspecific variability. Altogether, the results indicated an open pan-genome for the family Salinibacteraceae, as well as the codification of relevant traits such as diverse rhodopsin genes, CRISPR-Cas systems and spacers, and one T6SS secretion system that could give ecological advantages to an EHB-2 isolate. For the new Salinibacter species, we propose the name Salinibacter altiplanensis sp. nov. (the designated type strain is AN15T = CECT 9105T = IBRC-M 11031T). ; This study was funded by the Spanish Ministry of Economy projects CGL2012-39627-C03-03 and CLG2015-66686-C3-1-P (to RRM), CLG2015-66686-C3-3-P (to JA), which were also both supported with European Regional Development Fund (FEDER) funds. RA was financed by the Max Planck Society. KTK's research was supported, in part, by the U.S. National Science Foundation (Award No. 1241046). TVP acknowledges a pre-doctoral fellowship (Nr BES-2013-064420) from the Ministerio de Economía y Competitividad of the Spanish Government. The authors also acknowledge the economic support and unconditional scientific interest of Lipotrue SL and Deep Blue Sea SL.