Evaluation of Alternative High-Throughput Sequencing Methodologies for the Monitoring of Marine Picoplanktonic Biodiversity Based on rRNA Gene Amplicons

16 pages, 6 figures, 2 tablesSequencing of rRNA gene polymerase chain reaction amplicons (rRNA tags) is the most common approach for investigating microbial diversity. The recent development of high-throughput sequencing (HTS) technologies has enabled the exploration of microbial biodiversity at an unprecedented scale, greatly expanding our knowledge on the microbiomes of marine ecosystems. These approaches provide accurate, fast, and cost efficient observations of the marine communities, and thus, may be suitable tools in biodiversity monitoring programs. To reach this goal, consistent and comparable methodologies must be used over time and within sites. Here, we have performed a cross-platform study of the two most common HTS methodologies, i.e., 454-pyrosequencing and Illumina tags to evaluate their usefulness in biodiversity monitoring and assessment of environmental status. Picoplankton biodiversity has been compared through both methodologies by sequencing the 16 and 18 S rRNA genes of a set of samples collected in the coast of Barcelona (NW Mediterranean). The results show that, despite differences observed in the rare OTUs retrieved, both platforms provide a comparable view of the marine picoplankton communities. On a taxonomic level, there was an accurate overlap in the detected phyla between the two methods and the overall estimates of alpha- and betadiversity were comparable. In addition, we explored the concept of “indicator species” and found that certain taxa (i.e., members of the Gammaproteobacteria among others) as well as the ratio between some phylogenetic groups (i.e., the ratio of Alphaproteobacteria/Gammaproteobacteria, Alteromonas/SAR11, and Alteromonas + Oceanospirillales/SAR11) have potential for being useful indicators of environmental status. The data show that implementing new protocols and identifying indicators of environmental status based on rRNA amplicon sequencing is feasible, and that is worth exploring whether the identified indices are universally applicableThis manuscript is a result of DEVOTES (DEVelopment Of innovative Tools for understanding marine biodiversity and assessing GEnS) project, funded by the European Union (grant agreement no. 308392), and a MINECO Grant GRADIENTS Fine-scale structure of cross-shore GRADIENTS along the Mediterranean coast (CTM2012-39476-C02)Peer Reviewe

Marked changes in diversity and relative activity of picoeukaryotes with depth in the world ocean

Microbial eukaryotes are key components of the ocean plankton. Yet, our understanding of their community composition and activity in different water layers of the ocean is limited, particularly for picoeukaryotes (0.2–3 µm cell size). Here, we examined the picoeukaryotic communities inhabiting different vertical zones of the tropical and subtropical global ocean: surface, deep chlorophyll maximum, mesopelagic (including the deep scattering layer and oxygen minimum zones), and bathypelagic. Communities were analysed by high-tthroughput sequencing of the 18S rRNA gene (V4 region) as represented by DNA (community structure) and RNA (metabolism), followed by delineation of Operational Taxonomic Units (OTUs) at 99% similarity. We found a stratification of the picoeukaryotic communities along the water column, with assemblages corresponding to the sunlit and dark ocean. Specific taxonomic groups either increased (e.g., Chrysophyceae or Bicosoecida) or decreased (e.g., Dinoflagellata or MAST-3) in abundance with depth. We used the rRNA:rDNA ratio of each OTU as a proxy of metabolic activity. The highest relative activity was found in the mesopelagic layer for most taxonomic groups, and the lowest in the bathypelagic. Altogether, we characterize the change in community structure and metabolic activity of picoeukaryotes with depth in the global ocean, suggesting a hotspot of activity in the mesopelagic

Long-term patterns of an interconnected core marine microbiota

Background Ocean microbes constitute ~ 70% of the marine biomass, are responsible for ~ 50% of the Earth’s primary production and are crucial for global biogeochemical cycles. Marine microbiotas include core taxa that are usually key for ecosystem function. Despite their importance, core marine microbes are relatively unknown, which reflects the lack of consensus on how to identify them. So far, most core microbiotas have been defined based on species occurrence and abundance. Yet, species interactions are also important to identify core microbes, as communities include interacting species. Here, we investigate interconnected bacteria and small protists of the core pelagic microbiota populating a long-term marine-coastal observatory in the Mediterranean Sea over a decade. Results Core microbes were defined as those present in \u3e 30% of the monthly samples over 10 years, with the strongest associations. The core microbiota included 259 Operational Taxonomic Units (OTUs) including 182 bacteria, 77 protists, and 1411 strong and mostly positive (~ 95%) associations. Core bacteria tended to be associated with other bacteria, while core protists tended to be associated with bacteria. The richness and abundance of core OTUs varied annually, decreasing in stratified warmers waters and increasing in colder mixed waters. Most core OTUs had a preference for one season, mostly winter, which featured subnetworks with the highest connectivity. Groups of highly associated taxa tended to include protists and bacteria with predominance in the same season, particularly winter. A group of 13 highly-connected hub-OTUs, with potentially important ecological roles dominated in winter and spring. Similarly, 18 connector OTUs with a low degree but high centrality were mostly associated with summer or autumn and may represent transitions between seasonal communities. Conclusions We found a relatively small and dynamic interconnected core microbiota in a model temperate marine-coastal site, with potential interactions being more deterministic in winter than in other seasons. These core microbes would be essential for the functioning of this ecosystem over the year. Other non-core taxa may also carry out important functions but would be redundant and non-essential. Our work contributes to the understanding of the dynamics and potential interactions of core microbes possibly sustaining ocean ecosystem function

Long-term patterns of an interconnected core marine microbiota

Background Ocean microbes constitute ∼70% of the marine biomass, are responsible for ∼50% of the Earth’s primary production, and are crucial for global biogeochemical cycles. Marine microbiotas include core taxa that are usually key for ecosystem function. Despite their importance, core marine microbes are relatively unknown, which reflects the lack of consensus on how to identify them. So far, most core microbiotas have been defined based on species occurrence and abundance. Yet, species interactions are also important to identify core microbes, as communities include interacting species. Here, we investigate interconnected bacteria and small protists of the core pelagic microbiota populating a long-term marine-coastal observatory in the Mediterranean Sea over a decade. Results Core microbes were defined as those present in >30% of the monthly samples over 10 years, with the strongest associations. The core microbiota included 259 Operational Taxonomic Units (OTUs) including 182 bacteria, 77 protists, and 1,411 strong and mostly positive (∼95%) associations. Core bacteria tended to be associated with other bacteria, while core protists tended to be associated with bacteria. The richness and abundance of core OTUs varied annually, decreasing in stratified warmers waters and increasing in colder mixed waters. Most core OTUs had a preference for one season, mostly winter, which featured subnetworks with the highest connectivity. Groups of highly associated taxa tended to include protists and bacteria with predominance in the same season, particularly winter. A group of 13 highly-connected hub-OTUs, with potentially important ecological roles dominated in winter and spring. Similarly, 18 connector OTUs with a low degree but high centrality were mostly associated with summer or autumn and may represent transitions between seasonal communities. Conclusions We found a relatively small and dynamic interconnected core microbiota in a model temperate marine-coastal site, with potential interactions being more deterministic in winter than in other seasons. These core microbes would be essential for the functioning of this ecosystem over the year. Other non-core taxa may also carry out important functions but would be redundant and non-essential. Our work contributes to the understanding of the dynamics and potential interactions of core microbes possibly sustaining ocean ecosystem function.Preprin

Large-scale ocean connectivity and planktonic body size

Villarino, Ernesto ... et al.-- 13 pages, 5 figures, 5 tables, supplementary material https://dx.doi.org/10.1038/s41467-017-02535-8Global patterns of planktonic diversity are mainly determined by the dispersal of propagules with ocean currents. However, the role that abundance and body size play in determining spatial patterns of diversity remains unclear. Here we analyse spatial community structure - β-diversity - for several planktonic and nektonic organisms from prokaryotes to small mesopelagic fishes collected during the Malaspina 2010 Expedition. β-diversity was compared to surface ocean transit times derived from a global circulation model, revealing a significant negative relationship that is stronger than environmental differences. Estimated dispersal scales for different groups show a negative correlation with body size, where less abundant large-bodied communities have significantly shorter dispersal scales and larger species spatial turnover rates than more abundant small-bodied plankton. Our results confirm that the dispersal scale of planktonic and micro-nektonic organisms is determined by local abundance, which scales with body size, ultimately setting global spatial patterns of diversityThis research was funded by the project Malaspina 2010 Circumnavigation Expedition (Consolider-Ingenio 2010, CSD2008-00077) and cofounded by the Basque Government (Department Deputy of Agriculture, Fishing and Food Policy). [...] E.V. was supported by a PhD Scholarship granted by the Iñaki Goenaga−Technology Centres FoundationPeer Reviewe

Disentangling the mechanisms shaping the surface ocean microbiota

BACKGROUND: The ocean microbiota modulates global biogeochemical cycles and changes in its configuration may have large-scale consequences. Yet, the underlying ecological mechanisms structuring it are unclear. Here, we investigate how fundamental ecological mechanisms (selection, dispersal and ecological drift) shape the smallest members of the tropical and subtropical surface-ocean microbiota: prokaryotes and minute eukaryotes (picoeukaryotes). Furthermore, we investigate the agents exerting abiotic selection on this assemblage as well as the spatial patterns emerging from the action of ecological mechanisms. To explore this, we analysed the composition of surface-ocean prokaryotic and picoeukaryotic communities using DNA-sequence data (16S- and 18S-rRNA genes) collected during the circumglobal expeditions Malaspina-2010 and TARA-Oceans. RESULTS: We found that the two main components of the tropical and subtropical surface-ocean microbiota, prokaryotes and picoeukaryotes, appear to be structured by different ecological mechanisms. Picoeukaryotic communities were predominantly structured by dispersal-limitation, while prokaryotic counterparts appeared to be shaped by the combined action of dispersal-limitation, selection and drift. Temperature-driven selection appeared as a major factor, out of a few selected factors, influencing species co-occurrence networks in prokaryotes but not in picoeukaryotes, indicating that association patterns may contribute to understand ocean microbiota structure and response to selection. Other measured abiotic variables seemed to have limited selective effects on community structure in the tropical and subtropical ocean. Picoeukaryotes displayed a higher spatial differentiation between communities and a higher distance decay when compared to prokaryotes, consistent with a scenario of higher dispersal limitation in the former after considering environmental heterogeneity. Lastly, random dynamics or drift seemed to have a more important role in structuring prokaryotic communities than picoeukaryotic counterparts. CONCLUSIONS: The differential action of ecological mechanisms seems to cause contrasting biogeography, in the tropical and subtropical ocean, among the smallest surface plankton, prokaryotes and picoeukaryotes. This suggests that the idiosyncrasy of the main constituents of the ocean microbiota should be considered in order to understand its current and future configuration, which is especially relevant in a context of global change, where the reaction of surface ocean plankton to temperature increase is still unclear. Video Abstract

El poder dels productors primaris unicel·lulars

3 pages, 1 figure[EN] Marine phytoplankton, including cyanobacteria and microalgae, dominates primary production across two thirds of the earth’s surface, sustaining virtually all marine life and exerting a fundamental control over global climate through carbon sequestration into the deep ocean. These unicellular photoautotrophs are responsible for roughly 50% of global net primary production, which is equivalent to producing 50 gigatons of organic carbon (C) per year (about 140 million t per day). […][ES] El fitoplancton marino, que incluye tanto a las cianobacterias como a las microalgas, domina la producción primaria en dos tercios de la superficie de la Tierra, sustentando prácticamente toda la vida marina y ejerciendo un control fundamental sobre el clima global mediante el secuestro de carbono en las profundidades del océano. Estos productores primarios unicelulares son responsables de aproximadamente el 50% de la producción primaria neta mundial, lo que equivale a producir 50 gigatoneladas de carbono orgánico (C) al año (alrededor de 140 millones de toneladas al día). […][CAT] El fitoplàncton marí, que inclou tant als cianobacteris com a les microalgues, domina la producció primària en dos terços de la superfície de la Terra, sustentant pràcticament tota la vida marina i exercint un control fonamental sobre el clima global mitjançant el segrest de carboni en les profunditats de l’oceà. Aquests productors primaris unicel·lulars són responsables d’aproximadament el 50% de la producció primària neta mundial, la qual cosa equival a produir 50 gigatones de carboni orgànic (C) l’any (al voltant de 140 milions de tones al dia). […]The ideas embodied in this essay are part of the objectives of the PRODIGIO project “Developing early warning systems for improved microalgae PROduction and anaerobic DIGgestIOn”. The PRODIGIO project has received funding from the European Union’s Horizon 2020 Research and Innovation programme under grant agreement no. 101007006Peer reviewe

Global beta diversity patterns of microbial communities in the surface and deep ocean

This is contribution 1112 from AZTI Marine Research Division.-- 14 pages, 4 figures, 3 tables, supporting information https://doi.org/10.1111/geb.13572.-- Data Availability Statement: DNA sequences for surface prokaryotes are publicly available at the European Nucleotide Archive [http://www.ebi.ac.uk/ena; accession number PRJEB25224 (16S rRNA genes)], for deep prokaryotes at the National Center for Biotechnology Information (NCBI) Sequence Read Archive (http://www.ncbi.nlm.nih.gov/Traces/sra) under accession ID SRP031469, and for surface and deep picoeukaryotes at the European Nucleotide Archive with accession number PRJEB23771 (http://www.ebi.ac.uk/ena). Environmental data used in this study are available from https://github.com/ramalok/malaspina.surface.metabacoding, Giner et al. (2020) and Salazar et al. (2015). The code to analyze the data and produce the figures of this research is available from the corresponding author upon request.-- This is the pre-peer reviewed version of the following article: Ernesto Villarino, James R. Watson, Guillem Chust ,A. John Woodill, Benjamin Klempay, Bror Jonsson, Josep M. Gasol, Ramiro Logares, Ramon Massana, Caterina R. Giner, Guillem Salazar, X. Anton Alvarez-Salgado, Teresa S. Catala, Carlos M. Duarte, Susana Agusti, Francisco Mauro, Xabier Irigoien, Andrew D. Barton; Global beta diversity patterns of microbial communities in the surface and deep ocean; Global Ecology and Biogeography 31(11): 2323-2336 (2022), which has been published in final form at https://doi.org/10.1111/geb.13572. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Use of Self-Archived VersionsAim: Dispersal and environmental gradients shape marine microbial communities, yet the relative importance of these factors across taxa with distinct sizes and dispersal capacity in different ocean layers is unknown. Here, we report a comparative analysis of surface and deep ocean microbial beta diversity and examine how these patterns are tied to oceanic distance and environmental gradients. Location: Tropical and subtropical oceans (30°N–40°S). Time period: 2010-2011. Major taxa studied: Prokaryotes and picoeukaryotes (eukaryotes between 0.2 and 3 μm). Methods: Beta diversity was calculated from metabarcoding data on prokaryotic and picoeukaryotic microbes collected during the Malaspina expedition across the tropical and subtropical oceans. Mantel correlations were used to determine the relative contribution of environment and oceanic distance driving community beta diversity. Results: Mean community similarity across all sites for prokaryotes was 38.9% in the surface and 51.4% in the deep ocean, compared to mean similarity of 25.8 and 12.1% in the surface and deep ocean, respectively, for picoeukaryotes. Higher dispersal rates and smaller body sizes of prokaryotes relative to picoeukaryotes likely contributed to the significantly higher community similarity for prokaryotes compared with picoeukaryotes. The ecological mechanisms determining the biogeography of microbes varied across depth. In the surface ocean, the environmental differences in space were a more important factor driving microbial distribution compared with the oceanic distance, defined as the shortest path between two sites avoiding land. In the deep ocean, picoeukaryote communities were slightly more structured by the oceanic distance, while prokaryotes were shaped by the combined action of oceanic distance and environmental filtering. Main conclusions: Horizontal gradients in microbial community assembly differed across ocean depths, as did mechanisms shaping them. In the deep ocean, the oceanic distance and environment played significant roles driving microbial spatial distribution, while in the surface the influence of the environment was stronger than oceanic distanceData collection was funded by the Malaspina 2010 Circumnavigation Expedition project (Consolider-Ingenio 2010, CSD2008-00077) and cofunded by the Basque Government (Department Deputy of Agriculture, Fishing and Food Policy). We acknowledge funding from the Spanish Government through the “Severo Ochoa Center of Excelence” accreditation CEX2019-000928-S. [...] We also acknowledge H2020 Mission Atlantic project (Ref. Grant Agreement Number 862428). EV was supported by an international exchange post-doc scholarship to Scripps Institution of Oceanography and Oregon State University granted by the Education Department of the Basque GovernmentPeer reviewe

Experimental study of differentially rotating supersonic plasma flows produced by aluminium wire array Z-pinches

A novel approach to cylindrical wire array z-pinches has been developed in order to create a rotating plasma flow analogous to astrophysical accretion discs. The method involves subjecting the wire array to a cusp magnetic field (B_r) to create converging off axis ablation streams to form a rotating flow. The rotation is sustained by the ram pressure of the ablation streams in a quasi-equilibrium state for approximately 150 ns. This corresponds to one full rotation of the plasma about the axis. The rotating plasma is supersonic with Mach number ~2 and a radially constant rotation velocity between 60 and 75 km/s; the angular velocity therefore has an r^-1 dependence and the flow is differential. A Thomson scattering diagnostic is used to measure the electron and ion temperatures as Te ~30 eV and Ti >55 eV and the ionisation of the plasma (Z) between 6 and 8. These parameters are used to calculate the Reynolds number (10^5 to 10^6) and magnetic Reynolds numbers (20 to 100) which are large enough for viscous and resistive effects to be negligible on the large scale of the flow. These are of sufficient magnitude for the experiment to be scalable to astrophysical accretion discs. Further more the Reynolds number for the experiment is large enough for shear instabilities to manifest in the plasma. Some evidence for this can be seen in XUV images and Thomson spectra which indicate the development of perturbations and vorticity within the flow. Predictions for the growth rate of the Kelvin Helmholtz instability, 12 to 40 ns, agree reasonably well with the observed perturbation growth of ~30 ns. It is also possible that shear instabilities are driving hydrodynamic turbulence. Turbulent heating of the plasma could explain the approximately 500 eV increase in the ion temperature observed from some Thomson spectra. Further work is required however to prove the existence of shear flows and turbulence within the experiments.Open Acces

Seasonal diversity patterns of marine picoeukaryotes from a Mediterranean Coastal site

Aquatic Sciences Meeting, Aquatic Sciences: Global And Regional Perspectives - North Meets South, 22-27 February 2015, Granada, SpainPicoeukaryotes are the most abundant eukaryotes in the sea and are recognized as fundamental components of marine ecosystems, contributing to phytoplankton biomass, primary production and food web interactions. The study of their diversity requires molecular surveys, which have been lately expanding with the emergence of High Throughput Sequencing (HTS). Hitherto, many studies have focused in describing the diversity present in different sites but not along time. Here we performed a time series study in order to find out the seasonal patterns in the diversity of picoeukaryotes. We analyzed a sample dataset taken monthly during several years in a Northwestern Mediterranean coastal site and processed by HTS of the 18S rDNA. Preliminary results showed that only 1% of the OTUs were present in all samples. A few taxonomic groups were the most abundant in the community yearlong, while different groups were the most diverse. Interestingly, we found that the OTUs presenting the highest number of reads generally did not show a seasonal pattern, whereas some less abundant OTUs might exhibit a very marked temporal distributionPeer Reviewe