The procedures and methodologies employed to study microbial eukaryotic plankton have been thoroughly discussed. Two main schools exist—one insisting on classic microscopy methodologies and the other ...supporting modern high-throughput sequencing (DNA metabarcoding). However, few studies have attempted to combine both these approaches; most studies implement one method while ignoring the other. This work aims to contribute to this discussion and examine the advantages and disadvantages of each methodology by comparing marine plankton community results from microscopy and DNA metabarcoding. The results obtained by the two methodologies do not vary significantly for Bacillariophyta, although they do for Dinoflagellata and Ciliophora. The lower the taxonomic level, the higher the inconsistency between the two methodologies for all the studied groups. Considering the different characteristics of microscopy-based identification and DNA metabarcoding, this work underlines that each method should be chosen depending on the aims of the study. DNA metabarcoding provides a better estimate of the taxonomic richness of an ecosystem while microscopy provides more accurate quantitative results regarding abundance and biomass. In any case, the combined use of the two methods, if properly standardized, can provide much more reliable and accurate results for the study of marine microbial eukaryotes.
It remains unclear as to how mixoplankton (coupled phototrophy and phagotrophy in one cell) affects the estimation of grazing rates obtained from the widely used dilution grazing technique. To ...address this issue, we prepared laboratory-controlled dilution experiments with known mixtures of phyto-, protozoo-, and mixoplankton, operated under different light regimes and species combinations. Our results evidenced that chlorophyll is an inadequate proxy for phytoplankton when mixoplankton are present. Conversely, species-specific cellular counts could assist (although not fully solve) in the integration of mixoplanktonic activity in a dilution experiment. Moreover, cell counts can expose prey selectivity patterns and intraguild interactions among grazers. Our results also demonstrated that whole community approaches mimic reality better than single-species laboratory experiments. We also confirmed that light is required for protozoo- and mixoplankton to correctly express their feeding activity, and that overall diurnal grazing is higher than nocturnal. Thus, we recommend that a detailed examination of initial and final plankton communities should become routine in dilution experiments, and that incubations should preferably be started at the beginning of both day and night periods. Finally, we hypothesize that in silico approaches may help disentangle the contribution of mixoplankton to the community grazing of a given system.
Ciliates have been recognized as one of the major components of the microbial food web, especially in ultra-oligotrophic waters, such as the Eastern Mediterranean Sea, where nutrients are scarce and ...the microbial community is dominated by pico- and nano-sized organisms. For this reason, ciliates play an important role in these ecosystems since they are the main planktonic grazers. Regardless the importance of these organisms, little is known about the community structure of heterotrophic and mixotrophic ciliates and how they are associated to their potential prey. In this study, we used 18S V4 rRNA gene metabarcoding to analyze ciliate community dynamics and how the relationship with potential prey changes according to different seasons and depths. Samples were collected seasonally at two stations of the Eastern Mediterranean Sea (HCB: coastal, M3A: offshore) from the surface and deep chlorophyll maximum (DCM) layers. The ciliate community structure varied across depths in HCB and across seasons in M3A, and the network analysis showed that in both stations, mixotrophic oligotrichs were positively associated with diatoms and showed few negative associations with ASVs annotated as marine Stramenopiles (MAST). On the other hand, heterotrophic tintinnids showed negative relationships in both HCB and M3A stations, mostly with Ochrophyta and Chlorophyta. These results showed, in first place that, although the two stations are close to each other, the ciliate dynamics differed between them. Moreover, mixotrophic and heterotrophic ciliates may have different ecological niches since mixotrophic ciliates may be more selective compared to heterotrophic species regarding their prey. These findings are the first glimpse into an understanding of the dynamics between heterotrophic and mixotrophic ciliates and their role in microbial assemblages and dynamics of ultra-oligotrophic environments.
Abstract
Mesocosm experiments have been fundamental to investigate the effects of elevated CO
2
and ocean acidification (OA) on planktic communities. However, few of these experiments have been ...conducted using naturally nutrient-limited waters and/or considering the combined effects of OA and ocean warming (OW). Coccolithophores are a group of calcifying phytoplankton that can reach high abundances in the Mediterranean Sea, and whose responses to OA are modulated by temperature and nutrients. We present the results of the first land-based mesocosm experiment testing the effects of combined OA and OW on an oligotrophic Eastern Mediterranean coccolithophore community. Coccolithophore cell abundance drastically decreased under OW and combined OA and OW (greenhouse, GH) conditions.
Emiliania huxleyi
calcite mass decreased consistently only in the GH treatment; moreover, anomalous calcifications (i.e. coccolith malformations) were particularly common in the perturbed treatments, especially under OA. Overall, these data suggest that the projected increase in sea surface temperatures, including marine heatwaves, will cause rapid changes in Eastern Mediterranean coccolithophore communities, and that these effects will be exacerbated by OA.
Arranging organisms into functional groups aids ecological research by grouping organisms (irrespective of phylogenetic origin) that interact with environmental factors in similar ways. Planktonic ...protists traditionally have been split between photoautotrophic “phytoplankton” and phagotrophic “microzooplankton”. However, there is a growing recognition of the importance of mixotrophy in euphotic aquatic systems, where many protists often combine photoautotrophic and phagotrophic modes of nutrition. Such organisms do not align with the traditional dichotomy of phytoplankton and microzooplankton. To reflect this understanding, we propose a new functional grouping of planktonic protists in an eco-physiological context: (i) phagoheterotrophs lacking phototrophic capacity, (ii) photoautotrophs lacking phagotrophic capacity, (iii) constitutive mixotrophs (CMs) as phagotrophs with an inherent capacity for phototrophy, and (iv) non-constitutive mixotrophs (NCMs) that acquire their phototrophic capacity by ingesting specific (SNCM) or general non-specific (GNCM) prey. For the first time, we incorporate these functional groups within a foodweb structure and show, using model outputs, that there is scope for significant changes in trophic dynamics depending on the protist functional type description. Accordingly, to better reflect the role of mixotrophy, we recommend that as important tools for explanatory and predictive research, aquatic food-web and biogeochemical models need to redefine the protist groups within their frameworks.
Microplastics are a major environmental challenge, being ubiquitous and persistent as to represent a new component in all marine environments. As any biogenic particle, microplastics provide surfaces ...for microbial growth and biofilm production, which largely consists of carbohydrates and proteins. Biofilms influence microbial activity and modify particle buoyancy, and therefore control the fate of microplastics at sea. In a simulated 'plastic ocean', three mesocosms containing oligotrophic seawater were amended with polystyrene microbeads and compared to three control mesocosms. The evolution of organic matter, microbial communities and nutrient concentrations was monitored over 12 days. The results indicated that microplastics increased the production of organic carbon and its aggregation into gel particulates. The observed increase of gel-like organics has implications on the marine biological pump as well as the transport of microplastics in the ocean.
Plankton metabarcoding is increasingly implemented in marine ecosystem assessments and is more cost-efficient and less time-consuming than monitoring based on microscopy (morphological). 18S rRNA ...gene is the most widely used marker for groups’ and species’ detection and classification within marine eukaryotic microorganisms. These datasets have commonly relied on the acquisition of organismal abundances directly from the number of DNA sequences (i.e. reads). Besides the inherent technical biases in metabarcoding, the largely varying 18S rRNA gene copy numbers (GCN) among marine protists (ranging from tens to thousands) is one of the most important biological biases for species quantification. In this work, we present a gene copy number correction factor (CF) for four marine planktonic groups: Bacillariophyta, Dinoflagellata, Ciliophora miscellaneous and flagellated cells. On the basis of the theoretical assumption that ‘1 read’ is equivalent to ‘1 GCN’, we used the GCN median values per plankton group to calculate the corrected cell number and biomass relative abundances. The species-specific absolute GCN per cell were obtained from various studies published in the literature. We contributed to the development of a species-specific 18S rRNA GCN database proposed by previous authors. To assess the efficiency of the correction factor we compared the metabarcoding, morphological and corrected relative abundances (in cell number and biomass) of 15 surface water samples collected in the Belgian Coastal Zone. Results showed that the application of the correction factor over metabarcoding results enables us to significantly improve the estimates of cell abundances for Dinoflagellata, Ciliophora and flagellated cells, but not for Bacillariophyta. This is likely to due to large biovolume plasticity in diatoms not corresponding to genome size and gene copy numbers. C-biomass relative abundance estimations directly from amplicon reads were only improved for Dinoflagellata and Ciliophora. The method is still facing biases related to the low number of species GCN assessed. Nevertheless, the increase of species in the GCN database may lead to the refinement of the proposed correction factor.
The temporal variability of planktonic ciliates was studied in an extreme oligotrophic environment with special focus on trophic modes and size classes. Abundance, biomass, size classes, mixotrophy ...vs. heterotrophy, and species composition of planktonic ciliates were investigated focusing on temporal (samples collected on a monthly basis during 2019) and vertical (7 depth layers in the euphotic zone, from surface to 120 m) distribution at a coastal station in the oligotrophic Eastern Mediterranean. Abundance was in general very low (20 to 1150 cells L
–1
), except for September, which presented the highest abundance and biomass. Aloricate species dominated the ciliate community in all months and depths (% contribution from 77% in September to 99% in April). In general, oligotrichs presented maximum abundance at 2–10 m (except for June, July, and November: 100–120 m) whereas choreotrichs were more homogeneously distributed and showed maxima at deep chlorophyll maximum (DCM). Small heterotrophs dominated the ciliate community at all depths and months, on average by 76% (they were 3 times more abundant than mixotrophs in terms of abundance and 2.5 times in terms of biomass). They were equally distributed both vertically and seasonally (and also in terms of size classes). In contrast, mixotrophs were found mainly at the surface layer to 20 m throughout the year, except for June and July (max at 100, 50 m). On average, 63% of integrated aloricate abundance was species <30 μm, of which 25% were mixotrophs. During the stratification period of May to November, the very small (<18 μm) and small (18–30 μm) mixotrophic species were distributed throughout the water column whereas the >50 μm mixotrophic species were found only above the DCM. In contrast, during the mixing period of December to May, mixotrophic ciliates were very few and were dominated by small and medium-sized species. It seems that mixotrophic and heterotrophic ciliates, on one hand, and the four size classes, on the other, are very distinct groups characterized by different distributions both vertically and seasonally. This may have important consequences for the structure and function of the microbial food web of which they are part, as well as for the carbon flow to the higher trophic levels of this oligotrophic environment in which ciliates are the main grazers.
This study investigates the dynamic processes affecting silver (Ag) nanoparticles that have been spiked into seawater at environmentally relevant concentrations (200 and 2000ngAgL−1). Seawater ...samples were taken at regular time intervals from multiple microcosm tanks and analysed rapidly, without any sample preparation, using a recently developed flow injection on-line dilution single particle inductively coupled plasma mass spectrometry method. Dissolution was found to be the predominant process of Ag nanoparticle transformation, with its rate being influenced by the type and thickness of the nanoparticle organic coating. More specifically the branched poly(ethyleneimine) coating provided additional stability to the 40 and 60nmAg nanoparticles that were tested, compared to the poly(vinylpyrrolidone) coated ones. At high Ag nanoparticle spiking levels and after 24h of exposure an extra Ag-containing nanoparticle peak appeared at the low range of the NP size distribution histogram. This peak corresponds to Ag-containing particles that contain Ag mass equivalent to 25–30nm Ag nanoparticles (assuming spherical shape). However, the composition and the “real” size of these particles remains unknown as the particles may have formed from the in-situ reduction of dissolved silver or they originate from other processes involving nanocrystal formation, as has been shown to occur in sewage sludge, or interaction with natural organic matter. Overall, this study provides additional insight into the physicochemical mechanisms behind Ag nanoparticle behavior in marine media.
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•spICP-MS was used to investigate the transformation of Ag NP in seawater medium.•Dissolution was the predominant process of Ag NP transformation.•Higher stability of BPEI-coated Ag NPs was observed for all investigated NP sizes.•An extra Ag NP peak was observed in the low size range at high spiking levels.
Introduction
Marine viruses regulate microbial population dynamics and biogeochemical cycling in the oceans. The ability of viruses to manipulate hosts’ metabolism through the expression of viral ...auxiliary metabolic genes (AMGs) was recently highlighted, having important implications in energy production and flow in various aquatic environments. Up to now, the presence and diversity of viral AMGs is studied using -omics data, and rarely using quantitative measures of viral activity alongside.
Methods
In the present study, four depth layers (5, 50, 75, and 1,000 m) with discrete hydrographic features were sampled in the Eastern Mediterranean Sea; we studied lytic viral community composition and AMG content through metagenomics, and lytic production rates through the viral reduction approach in the ultra-oligotrophic Levantine basin where knowledge regarding viral actions is rather limited.
Results and Discussion
Our results demonstrate depth-dependent patterns in viral diversity and AMG content, related to differences in temperature, nutrients availability, and host bacterial productivity and abundance. Although lytic viral production rates were similar along the water column, the virus-to-bacteria ratio was higher and the particular set of AMGs was more diverse in the bathypelagic (1,000 m) than the shallow epipelagic (5, 50, and 75 m) layers, revealing that the quantitative effect of viruses on their hosts may be the same along the water column through the intervention of different AMGs. In the resource- and energy-limited bathypelagic waters of the Eastern Mediterranean, the detected AMGs could divert hosts’ metabolism toward energy production, through a boost in gluconeogenesis, fatty-acid and glycan biosynthesis and metabolism, and sulfur relay. Near the deep-chlorophyll maximum depth, an exceptionally high percentage of AMGs related to photosynthesis was noticed. Taken together our findings suggest that the roles of viruses in the deep sea might be even more important than previously thought as they seem to orchestrate energy acquisition and microbial community dynamics, and thus, biogeochemical turnover in the oceans.
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