Cultivating the macroalgal holobiont: effects of integrated multi-trophic aquaculture on the microbiome of Ulva rigida (chlorophyta)

Ulva is a ubiquitous macroalgal genus of commercial interest. Integrated Multi-Trophic Aquaculture (IMTA) systems promise large-scale production of macroalgae due to their high productivity and environmental sustainability. Complex host-microbiome interactions play a decisive role in macroalgal development, especially in Ulva spp. due to algal growth- and morphogenesis-promoting factors released by associated bacteria. However, our current understanding of the microbial community assembly and structure in cultivated macroalgae is scant. We aimed to determine (i) to what extent IMTA settings influence the microbiome associated with U. rigida and its rearing water, (ii) to explore the dynamics of beneficial microbes to algal growth and development under IMTA settings, and (iii) to improve current knowledge of host-microbiome interactions. We examined the diversity and taxonomic composition of the prokaryotic communities associated with wild versus IMTA-grown Ulva rigida and surrounding seawater by using 16S rRNA gene amplicon sequencing. With 3141 Amplicon Sequence Variants (ASVs), the prokaryotic richness was, overall, higher in water than in association with U. rigida. Bacterial ASVs were more abundant in aquaculture water samples than water collected from the lagoon. The beta diversity analysis revealed distinct prokaryotic communities associated with Ulva collected in both aquacultures and coastal waters. Aquaculture samples (water and algae) shared 22% of ASVs, whereas natural, coastal lagoon samples only 9%. While cultivated Ulva selected 239 (8%) host-specific ASVs, wild specimens possessed more than twice host-specific ASVs (17%). Cultivated U. rigida specimens enriched the phyla Cyanobacteria, Planctomycetes, Verrucomicrobia, and Proteobacteria. Within the Gammaproteobacteria, while Glaciecola mostly dominated the microbiome in cultivated algae, the genus Granulosicoccus characterized both Ulva microbiomes. In both wild and IMTA settings, the phylum Bacteroidetes was more abundant in the bacterioplankton than in direct association with U. rigida. However, we observed that the Saprospiraceae family within this phylum was barely present in lagoon water but very abundant in aquaculture water. Aquaculture promoted the presence of known morphogenesis-inducing bacteria in water samples. Our study suggests that IMTA significantly shaped the structure and composition of the microbial community of the rearing water and cultivated U. rigida. Detailed analysis revealed the presence of previously undetected taxa associated with Ulva, possessing potentially unknown functional traits.European Union (EU)642575; German Research Foundation (DFG) CRC 1127 ChemBioSys;COST Action "Phycomorph" FA1406info:eu-repo/semantics/publishedVersio

Draft genome sequence of vibrio sp. strain Evh12, a bacterium retrieved from the gorgonian coral eunicella verrucosa.

To shed light on the associations established between Vibrio species and soft corals in coastal ecosystems, we report here the draft genome sequence of Vibrio sp. strain Evh12, a bacterium that has been isolated from the gorgonian coral Eunicella verrucosa and that shows antagonistic activity against Escherichia coli

Resistance of seagrass habitats to ocean acidification via altered interactions in a tri-trophic chain

Despite the wide knowledge about prevalent effects of ocean acidification on single species, the consequences on species interactions that may promote or prevent habitat shifts are still poorly understood. Using natural CO2 vents, we investigated changes in a key tri-trophic chain embedded within all its natural complexity in seagrass systems. We found that seagrass habitats remain stable at vents despite the changes in their tri-trophic components. Under high pCO2, the feeding of a key herbivore (sea urchin) on a less palatable seagrass and its associated epiphytes decreased, whereas the feeding on higher-palatable green algae increased. We also observed a doubled density of a predatory wrasse under acidified conditions. Bottom-up CO2 effects interact with top-down control by predators to maintain the abundance of sea urchin populations under ambient and acidified conditions. The weakened urchin herbivory on a seagrass that was subjected to an intense fish herbivory at vents compensates the overall herbivory pressure on the habitat-forming seagrass. Overall plasticity of the studied system components may contribute to prevent habitat loss and to stabilize the system under acidified conditions. Thus, preserving the network of species interactions in seagrass ecosystems may help to minimize the impacts of ocean acidification in near-future oceans.The research was funded by an ASSEMBLE access project within the EU FP7/2007–2013 program (grant agreement n° 227799) hosted by MCG at the SZN. The Portuguese FCT– Fundação para a Ciência e a Tecnologia funded BMC in the ambit of the contract program DL57/2016/CP1361/CT0004 and CCMAR through the project UID/Multi/04326/2019.info:eu-repo/semantics/publishedVersio

Macroalgal–bacterial interactions: identification and role of thallusin in morphogenesis of the seaweed Ulva (Chlorophyta)

Macroalgal microbiomes have core functions related to biofilm formation, growth, and morphogenesis of seaweeds. In particular, the growth and development of the sea lettuce Ulva spp. (Chlorophyta) depend on bacteria releasing morphogenetic compounds. Under axenic conditions, the macroalga Ulva mutabilis develops a callus-like phenotype with cell wall protrusions. However, co-culturing with Roseovarius sp. (MS2) and Maribacter sp. (MS6), which produce various stimulatory chemical mediators, completely recovers morphogenesis. This ecological reconstruction forms a tripartite community which can be further studied for its role in cross-kingdom interactions. Hence, our study sought to identify algal growth- and morphogenesis-promoting factors (AGMPFs) capable of phenocopying the activity of Maribacter spp. We performed bioassay-guided solid-phase extraction in water samples collected from U. mutabilis aquaculture systems. We uncovered novel ecophysiological functions of thallusin, a sesquiterpenoid morphogen, identified for the first time in algal aquaculture. Thallusin, released by Maribacter sp., induced rhizoid and cell wall formation at a concentration of 11 pmol l-1. We demonstrated that gametes acquired the iron complex of thallusin, thereby linking morphogenetic processes with intracellular iron homeostasis. Understanding macroalgae-bacteria interactions permits further elucidation of the evolution of multicellularity and cellular differentiation, and development of new applications in microbiome-mediated aquaculture systems.FCT: UID/Multi/04326/2019; UIDB/04326/2020; CCMAR/ID/16/2018; CEECINST/00114/2018. German Research Foundation (DFG) CRC1127; European Union (EU) 642575; ASSEMBLE 227799; COST Action 'Phycomorph' FA1406.info:eu-repo/semantics/publishedVersio

Insights into the Evolution of Multicellularity from the Sea Lettuce Genome

We report here the 98.5 Mbp haploid genome (12,924 protein coding genes) of Ulva mutabilis, a ubiquitous and iconic representative of the Ulvophyceae or green seaweeds. Ulva’s rapid and abundant growth makes it a key contributor to coastal biogeochemical cycles; its role in marine sulfur cycles is particularly important because it produces high levels of dimethylsulfoniopropionate (DMSP), the main precursor of volatile dimethyl sulfide (DMS). Rapid growth makes Ulva attractive biomass feedstock but also increasingly a driver of nuisance “green tides.” Ulvophytes are key to understanding the evolution of multicellularity in the green lineage, and Ulva morphogenesis is dependent on bacterial signals, making it an important species with which to study cross-kingdom communication. Our sequenced genome informs these aspects of ulvophyte cell biology, physiology, and ecology. Gene family expansions associated with multicellularity are distinct from those of freshwater algae. Candidate genes, including some that arose following horizontal gene transfer from chromalveolates, are present for the transport and metabolism of DMSP. The Ulva genome offers, therefore, new opportunities to understand coastal and marine ecosystems and the fundamental evolution of the green lineage

Inter-kingdom interactions shapes macroalga Ulva holobiont: studies on algal growth and morphogenesis promoting factors (phytohormones and thallusin) and algal associated microbial community

Ulva is a ubiquitous marine and freshwater macroalgae genus that belongs to the phylum Chlorophyta. Macroalgae and bacteria strongly interact through chemical queues. Ulva mutabilis forms with the bacteria, Roseovarius sp. and Maribacter sp. a robust community. Ulva attracts and supports bacterial growth, and, in return, the bacteria release algal growth and morphogenetic factors (AGMPFs). This work aims to investigate AGMPFs, and their effect on algal metabolism. The first step was characterized by the optimization of the protocol to obtain axenic Ulva germ cells and the standardization of the workflow for algal metabolomics. Phytohormones presence and concentrations were screened in different morphotypes of U. mutabilis and their activity was tested with axenic algae. Even though they did not affect Ulva's development, the concentration of salicylic acid was higher in the axenic than in presence of bacteria. The study of AGMPFs continued with their characterization in Ulva aquaculture and in coastal lagoon. With solid-phase extraction and targeted liquid chromatography it was possible to identify the morphogen thallusin in the phycosphere of Ulva. The final proof was obtained after the discovery that thallusin is produced and released into the growing medium by Maribacter sp. and the morphogen alone induces the differentiation of the primary rhizoid cell and a normal cell wall formation in Ulva. With transcriptomics and metabolomics analyses it was possible to suggest that in the early phase of Ulva's development, thallusin regulates the transcription factor CCCH-type zinc finger protein, phosphoglycerate kinase and enolase enzymes, which affect the algal metabolism and likely future morphology. Lastly, the characterization of the microbiome associated with Ulva rigida of an industrial facility set the bases to study further the biotechnological application of thallusin and a synthetic microbiome in aquaculture

The effect of live feeds bathed with the red seaweed Asparagopsis armata on the survival, growth and physiology status of Sparus aurata larvae

Larval rearing is affected by a wide range of microorganisms that thrive in larviculture systems. Some seaweed species have metabolites capable of reducing the bacterial load. However, no studies have yet tested whether including seaweed metabolites on larval rearing systems has any effects on the larvae development. This work assessed the development of Sparus aurata larvae fed preys treated with an Asparagopsis armata product. Live prey, Brachionus spp. and Artemia sp., were immersed in a solution containing 0.5% of a commercial extract of A. armata (Ysaline 100, YSA) for 30 min, before being fed to seabream larvae (n = 4 each). In the control, the live feed was immersed in clear water. Larval parameters such as growth, survival, digestive capacity (structural-histology and functional-enzymatic activity), stress level (cortisol content), non-specific immune response (lysozyme activity), anti-bacterial activity (disc-diffusion assay) and microbiota quantification (fish larvae gut and rearing water) were monitored. Fish larvae digestive capacity, stress level and non-specific immune response were not affected by the use of YSA. The number of Vibrionaceae was significantly reduced both in water and larval gut when using YSA. Growth was enhanced for YSA treatment, but higher mortality was also observed, especially until 10 days after hatching (DAH). The mortality peak observed at 8 DAH for both treatments, but higher for YSA, indicates larval higher susceptibility at this development stage, suggesting that lower concentrations of YSA should be used until 10 DAH. The application of YSA after 10 DAH onwards promotes a safer rearing environment.Portuguese Foundation for Science and TechnologySATA project [PTDC/MAR/112792/2009]info:eu-repo/semantics/publishedVersio

Altered epiphyte community and sea urchin diet in Posidonia oceanica meadows in the vicinity of volcanic CO2 vents

Ocean acidification (OA) predicted for 2100 is expected to shift seagrass epiphyte communities towards the dominance of more tolerant non-calcifying taxa. However, little is known about the indirect effects of such changes on food provision to key seagrass consumers. We found that epiphyte communities of the seagrass Posidonia oceanica in two naturally acidified sites (i.e. north and south sides of a volcanic CO2 vent) and in a control site away from the vent at the Ischia Island (NW Mediterranean Sea) significantly differed in composition and abundance. Such differences involved a higher abundance of non-calcareous crustose brown algae and a decline of calcifying polychaetes in both acidified sites. A lower epiphytic abundance of crustose coralline algae occurred only in the south side of the vents, thus suggesting that OA may alter epiphyte assemblages in different ways due to interaction with local factors such as differential fish herbivory or hydrodynamics. The OA effects on food items (seagrass, epiphytes, and algae) indirectly propagated into food provision to the sea urchin Paracentrotus lividus, as reflected by a reduced P. oceanica exploitation (i.e. less seagrass and calcareous epiphytes in the diet) in favour of non-calcareous green algae in both vent sites. In contrast, we detected no difference close and outside the vents neither in the composition of sea urchin diet nor in the total abundance of calcareous versus non-calcareous taxa. More research, under realistic scenarios of predicted pH reduction (i.e. <= 0.32 units of pH by 2100), is still necessary to better understand cascading effects of this altered urchin exploitation of food resources under acidified conditions on ecosystem diversity and function. (C) 2017 Elsevier Ltd. All rights reserved.ASSEMBLE Access project within the EU [227799]Portuguese Foundation for Science and Technology (FCT) [SFRH/BPD/75307/2010]info:eu-repo/semantics/publishedVersio

Effects of live feed manipulation with algal‐derived antimicrobial metabolites on fish larvae microbiome assembly: a molecular‐based assessment

Opportunistic microorganisms acquired through rearing water or live feed ingestion are believed to underpin high mortality rates of fish larvae, constituting a production bottleneck for the aquaculture industry. We employed 16S rRNA gene sequencing to determine whether treatment of live feed (rotifers and Artemia) with algal-derived, antibacterial metabolites could alter bacterial community structure of gilthead seabream (Sparus aurata) larvae in a larviculture facility. Owing to a large degree of sample-to-sample variation, pronounced 'legacy effects' of live feed manipulation on the total fish larvae bacterial community could not be verified. Notwithstanding, the approach induced shifts in relative abundance of specific bacterial phylotypes in both the live feed and fish larvae. Some phylotypes representing opportunistic taxa such as Stenotrophomonas, Pseudomonas and Klebsiella displayed reduced abundances in the bacterial community of fish larvae fed metabolite-treated vs. control live feed. Conversely, potentially beneficial phylotypes in the Alphaproteobacteria clade were consistently-although not significantly-promoted in the treated larval samples. These outcomes encourage future microbiome manipulation attempts to improve fish larviculture. However, successful host colonization and competition with resident symbionts are primary barriers that need to be overcome if live feeds are to be used as effective delivery systems of beneficial bacteria to fish larvae.info:eu-repo/semantics/publishedVersio