Planktonic foraminifera eDNA signature deposited on the seafloor remains preserved after burial in marine sediments.

Environmental DNA (eDNA) metabarcoding of marine sediments has revealed large amounts of sequences assigned to planktonic taxa. How this planktonic eDNA is delivered on the seafloor and preserved in the sediment is not well understood. We address these questions by comparing metabarcoding and microfossil foraminifera assemblages in sediment cores taken off Newfoundland across a strong ecological gradient. We detected planktonic foraminifera eDNA down to 30 cm and observed that the planktonic/benthic amplicon ratio changed with depth. The relative proportion of planktonic foraminiferal amplicons remained low from the surface down to 10 cm, likely due to the presence of DNA from living benthic foraminifera. Below 10 cm, the relative proportion of planktonic foraminifera amplicons rocketed, likely reflecting the higher proportion of planktonic eDNA in the DNA burial flux. In addition, the microfossil and metabarcoding assemblages showed a congruent pattern indicating that planktonic foraminifera eDNA is deposited without substantial lateral advection and preserves regional biogeographical patterns, indicating deposition by a similar mechanism as the foraminiferal shells. Our study shows that the planktonic eDNA preserved in marine sediments has the potential to record climatic and biotic changes in the pelagic community with the same spatial and temporal resolution as microfossils.

Parasitic Chytrids Upgrade and Convey Primary Produced Carbon During Inedible Algae Proliferation.

Microbial parasites have only recently been included in planktonic food web studies, but their functional role in conveying dietary energy still remains to be elucidated. Parasitic fungi (chytrids) infecting phytoplankton may constitute an alternative trophic link and promote organic matter transfer through the production of dissemination zoospores. Particularly, during proliferation of inedible or toxic algal species, such as large Cyanobacteria fostered by global warming, parasites can constitute an alternative trophic link providing essential dietary nutrients that support somatic growth and reproduction of consumers. Using phytoplankton-parasites associations grown under laboratory controlled conditions we assessed the fatty acids and biochemical composition of species with different nutritional quality and followed the metabolic pathway from the algal host and their parasites zoospores using compound-specific stable isotope analysis. This study demonstrated that chytrids are trophic upgraders able to retain essential nutrients that can be transferred to upper trophic levels both in terms of organic matter quantity and nutritional quality. Through the production of zoospores, nutritionally important long-chain polyunsaturated fatty acids that can be consequently assimilated by consumers. We conclude that parasitism at the base of aquatic food webs may represent a crucial trophic link for dietary nutrients and essential biomolecules alternative to herbivory or bacterivory, which can be particularly crucial during the proliferation of inedible or nutritionally inadequate algal species fostered by climate change.

The planktonic protist interactome: where do we stand after a century of research?

Microbial interactions are crucial for Earth ecosystem function, but our knowledge about them is limited and has so far mainly existed as scattered records. Here, we have surveyed the literature involving planktonic protist interactions and gathered the information in a manually curated Protist Interaction DAtabase (PIDA). In total, we have registered ~2500 ecological interactions from ~500 publications, spanning the last 150 years. All major protistan lineages were involved in interactions as hosts, symbionts (mutualists and commensalists), parasites, predators, and/or prey. Predation was the most common interaction (39% of all records), followed by symbiosis (29%), parasitism (18%), and 'unresolved interactions' (14%, where it is uncertain whether the interaction is beneficial or antagonistic). Using bipartite networks, we found that protist predators seem to be 'multivorous' while parasite-host and symbiont-host interactions appear to have moderate degrees of specialization. The SAR supergroup (i.e., Stramenopiles, Alveolata, and Rhizaria) heavily dominated PIDA, and comparisons against a global-ocean molecular survey (TARA Oceans) indicated that several SAR lineages, which are abundant and diverse in the marine realm, were underrepresented among the recorded interactions. Despite historical biases, our work not only unveils large-scale eco-evolutionary trends in the protist interactome, but it also constitutes an expandable resource to investigate protist interactions and to test hypotheses deriving from omics tools.

Biological Concepts for the Control of Aquatic Zoosporic Diseases.

Aquatic zoosporic diseases are threatening global biodiversity and ecosystem services, as well as economic activities. Current means of controlling zoosporic diseases are restricted primarily to chemical treatments, which are usually harmful or likely to be ineffective in the long term. Furthermore, some of these chemicals have been banned due to adverse effects. As a result, there is a need for alternative methods with minimal side-effects on the ecosystem or environment. Here, we integrate existing knowledge of three poorly interconnected areas of disease research - amphibian conservation, aquaculture, and plankton ecology - and arrange it into seven biological concepts to control zoosporic diseases. These strategies may be less harmful and more sustainable than chemical approaches. However, more research is needed before safe application is possible.

A de novo approach to disentangle partner identity and function in holobiont systems.

BACKGROUND: Study of meta-transcriptomic datasets involving non-model organisms represents bioinformatic challenges. The production of chimeric sequences and our inability to distinguish the taxonomic origins of the sequences produced are inherent and recurrent difficulties in de novo assembly analyses. As the study of holobiont meta-transcriptomes is affected by challenges invoked above, we propose an innovative bioinformatic approach to tackle such difficulties and tested it on marine models as a proof of concept. RESULTS: We considered three holobiont models, of which two transcriptomes were previously published and a yet unpublished transcriptome, to analyze and sort their raw reads using Short Read Connector, a k-mer based similarity method. Before assembly, we thus defined four distinct categories for each holobiont meta-transcriptome: host reads, symbiont reads, shared reads, and unassigned reads. Afterwards, we observed that independent de novo assemblies for each category led to a diminution of the number of chimeras compared to classical assembly methods. Moreover, the separation of each partner's transcriptome offered the independent and comparative exploration of their functional diversity in the holobiont. Finally, our strategy allowed to propose new functional annotations for two well-studied holobionts (a Cnidaria-Dinophyta, a Porifera-Bacteria) and a first meta-transcriptome from a planktonic Radiolaria-Dinophyta system forming widespread symbiotic association for which our knowledge is considerably limited. CONCLUSIONS: In contrast to classical assembly approaches, our bioinformatic strategy generates less de novo assembled chimera and allows biologists to study separately host and symbiont data from a holobiont mixture. The pre-assembly separation of reads using an efficient tool as Short Read Connector is an effective way to tackle meta-transcriptomic challenges and offers bright perpectives to study holobiont systems composed of either well-studied or poorly characterized symbiotic lineages and ultimately expand our knowledge about these associations.

Integrating dimensions of biodiversity in choreotrichs and oligotrichs of marine plankton.

Choreotrichs and oligotrichs are the main ciliate groups in marine plankton, where they play major roles as trophic intermediaries. We have studied these groups with a variety of approaches to combine the three dimensions of biodiversity-taxonomy, genetics and function. Here we revisit our findings with an integrative perspective, and highlight future directions. In our studies, the correspondence between classical taxonomy (mostly based on morphology) and the increasingly available genetic data (DNA sequences) is examined at the individual, population, species, and assemblage levels. We use a combination of single-cell and environmental sequencing to quantify diversity, track distribution patterns, and explain biogeography processes. Comparativelly, we know little about how the morphological and genetic estimates of diversity relate to function, but we expect to better link these aspects by incorporating modern -omics approaches. For example, we have pioneered functional transcriptomic analyses in these groups by contrasting a heterotrophic choreotrich and a mixotrophic oligotrich. These data provide a tremendous resource to start building reference databases needed to measure differential expression of key functional genes, either experimentally or directly in the environment.

Chromium isotopic composition of core-top planktonic foraminifera.

The chromium isotope system (53 Cr/52 Cr expressed as δ53 Cr relative to NIST SRM 979) is potentially a powerful proxy for the redox state of the ocean-atmosphere system, but a lack of temporally continuous, well-calibrated archives has limited its application to date. Marine carbonates could potentially serve as a common and continuous Cr isotope archive. Here, we present the first evaluation of planktonic foraminiferal calcite as an archive of seawater δ53 Cr. We show that single foraminiferal species from globally distributed core tops yielded variable δ53 Cr, ranging from 0.1‰ to 2.5‰. These values do not match with the existing measurements of seawater δ53 Cr. Further, within a single core-top, species with similar water column distributions (i.e., depth habitats) yielded variable δ53 Cr values. In addition, mixed layer and thermocline species do not consistently exhibit decreasing trends in δ53 Cr as expected based on current understanding of Cr cycling in the ocean. These observations suggest that either seawater δ53 Cr is more heterogeneous than previously thought or that there is significant and species-dependent Cr isotope fractionation during foraminiferal calcification. Given that the δ53 Cr variability is comparable to that observed in geological samples throughout Earth's history, interpreting planktonic foraminiferal δ53 Cr without calibrating modern foraminifera further, and without additional seawater measurements, would lead to erroneous conclusions. Our core-top survey clearly indicates that planktonic foraminifera are not a straightforward δ53 Cr archive and should not be used to study marine redox evolution without additional study. It likewise cautions against the use of δ53 Cr in bulk carbonate or other biogenic archives pending further work on vital effects and the geographic heterogeneity of the Cr isotope composition of seawater.

Planktonic ciliate community structure in shallow lakes of lowland Western Europe.

Temperate shallow meso- to eutrophic lakes can exist in one of two alternative states with contrasting foodwebs, referred to as the clear-water and the turbid state. We describe the planktonic ciliate communities of such lakes based on a survey of 66 northwestern European lakes. Ciliates were enumerated and identified to species level according to the quantitative protargol staining technique. Ciliate biomass was on average twice as high in the turbid than in the clear-water lakes. The ciliate communities were dominated by oligotrichs and protostomatids, and no differences in functional composition or α-diversity could be detected between turbid and clear-water lakes, although ß-diversity tended to be higher in the latter. At the species level, however, community structure strongly differed between turbid and clear-water lakes, and several indicator species could be identified for the different lake categories. Variation partitioning showed that nutrient status did not explain ciliate community structure independent of the alternative states, while lake area was identified as an additional structuring factor for the ciliate communities. These results stress the importance of the ecosystem structure in shaping ciliate communities in temperate shallow lakes and suggest that nutrient status has little direct effect on ciliate community structure in such lakes.

Environmental selection of protistan plankton communities in hypersaline anoxic deep-sea basins, Eastern Mediterranean Sea.

High salt concentrations, absence of light, anoxia, and high hydrostatic pressure make deep hypersaline anoxic basins (DHABs) in the Eastern Mediterranean Sea one of the most polyextreme habitats on Earth. Taking advantage of the unique chemical characteristics of these basins, we tested the effect of environmental selection and geographic distance on the structure of protistan communities. Terminal restriction fragment length polymorphism (T-RFLP) analyses were performed on water samples from the brines and seawater/brine interfaces of five basins: Discovery, Urania, Thetis, Tyro, and Medee. Using statistical analyses, we calculated the partitioning of diversity among the ten individual terminal restriction fragment (T-RF) profiles, based on peak abundance and peak incidence. While a significant distance effect on spatial protistan patterns was not detected, hydrochemical gradients emerged as strong dispersal barriers that likely lead to environmental selection in the DHAB protistan plankton communities. We identified sodium, magnesium, sulfate, and oxygen playing in concerto as dominant environmental drivers for the structuring of protistan plankton communities in the Eastern Mediterranean DHABs.

Southern ocean biogeography of tintinnid ciliates of the marine plankton.

Ciliate microzooplankton are important grazers in most pelagic ecosystems and among them, tintinnids, with their largely species-specific loricas, allow relatively easy assessment of questions of diversity and distributions. Herein, we present the results of a survey of species records of tintinnids from the Southern Ocean (locations below 40°S) reported in 56 publications yielding 2,047 species records (synonyms included) from 402 locations. The 192 species reported can be parsed into two main groups: 32 endemic Southern Ocean species, known only from 40°S and further south, and a second group of 181 widespread species, forms with extensive geographic ranges extending into the Southern Ocean. Widespread species reported from the Southern Ocean can be further divided into a group of 81 species, each recorded multiple times in the Southern Ocean waters and 70 apparent "stray" species which have only been found but once. The endemic and widespread species of the Southern Ocean show both distinct distributional patterns and morphological differences. The assemblage of Southern Ocean endemics is found mostly within the Antarctic zone delimited by the average location of the Polar Front and contains a relatively large portion of wide-mouthed forms. We give suggestions for future study.

Development of single-nucleotide polymorphism-based phylum-specific PCR amplification technique: application to the community analysis using ciliates as a reference organism.

Despite recent advance in mass sequencing technologies such as pyrosequencing, assessment of culture-independent microbial eukaryote community structures using universal primers remains very difficult due to the tremendous richness and complexity of organisms in these communities. Use of a specific PCR marker targeting a particular group would provide enhanced sensitivity and more in-depth evaluation of microbial eukaryote communities compared to what can be achieved with universal primers. We discovered that many phylum- or group-specific single-nucleotide polymorphisms (SNPs) exist in small subunit ribosomal RNA (SSU rRNA) genes from diverse eukaryote groups. By applying this discovery to a known simple allele-discriminating (SAP) PCR method, we developed a technique that enables the identification of organisms belonging to a specific higher taxonomic group (or phylum) among diverse types of eukaryotes. We performed an assay using two complementary methods, pyrosequencing and clone library screening. In doing this, specificities for the group (ciliates) targeted in this study in bulked environmental samples were 94.6% for the clone library and 99.2% for pyrosequencing, respectively. In particular, our novel technique showed high selectivity for rare species, a feature that may be more important than the ability to identify quantitatively predominant species in community structure analyses. Additionally, our data revealed that a target-specific library (or ciliate-specific one for the present study) can better explain the ecological features of a sampling locality than a universal library.

Intraclass evolution and classification of the Colpodea (Ciliophora).

Using nine new taxa and statistical inferences based on morphological and molecular data, we analyze the evolution within the class Colpodea. The molecular and cladistic analyses show four well-supported clades: platyophryids, bursariomorphids, cyrtolophosidids, and colpodids. There is a widespread occurrence of homoplasies, affecting even conspicuous morphological characteristics, e.g. the inclusion of the micronucleus in the perinuclear space of the macronucleus. The most distinct changes in the morphological classification are the lack of a basal divergence into two subclasses and the split of the cyrtolophosidids into two main clades, differing mainly by the presence vs. absence of an oral cavity. The most complex clade is that of the colpodids. We partially reconcile the morphological and molecular data using evolutionary systematics, providing a scenario in which the colpodids evolved from a Bardeliella-like ancestor and the genus Colpoda performed an intense adaptive radiation, giving rise to three main clades: Colpodina n. subord., Grossglockneriina, and Bryophryina. Three new taxa are established: Colpodina n. subord., Tillinidae n. fam., and Ottowphryidae n. fam. Colpodean evolution and classification are far from being understood because sequences are lacking for most species and half of their diversity is possibly undescribed.

Diversity of oligotrichia and choreotrichia ciliates in coastal marine sediments and in overlying plankton.

Elucidating the relationship between ciliate communities in the benthos and the plankton is critical to understanding ciliate diversity in marine systems. Although data for many lineages are sparse, at least some members of the dominant marine ciliate clades Oligotrichia and Choreotrichia can be found in both plankton and benthos, in the latter either as cysts or active forms. In this study, we developed a molecular approach to address the relationship between the diversity of ciliates in the plankton and those of the underlying benthos in the same locations. Samples from plankton and sediments were compared across three sites along the New England coast, and additional subsamples were analyzed to assess reproducibility of methods. We found that sediment and plankton subsamples differed in their robustness to repeated subsampling. Sediment subsamples (i.e., 1-g aliquots from a single approximately 20-g sample) gave variable estimates of diversity, while plankton subsamples produced consistent results. These results indicate the need for additional study to determine the spatial scale over which diversity varies in marine sediments. Clustering of phylogenetic types indicates that benthic assemblages of oligotrichs and choreotrichs appear to be more like those from spatially remote benthic communities than the ciliate communities sampled in the water above them.

Spatial and temporal patterns of ciliate species composition (Protozoa: Ciliophora) in the plankton of the Upper Paraná River floodplain.

Spatial and temporal patterns of plankton ciliates species composition in the Paraná River floodplain were investigated. Samplings were carried out in twelve environments in two distinct hydrological periods (limnophase and potamophase). A total of 61 species of ciliates were recorded, and among them 21 are classified as pelagic while 40 are considered preferentially as littoral species. The registered species belong to eleven orders, and among them, Prostomatida was the most specious followed by Hymenostomatida and Peritrichida. The ciliate species composition was significantly distinct between periods, but not among environments. In this way, typically pelagic species characterized the ciliate community during the limnophase period, while the littoral species were predominant during the potamophase period. Our results strongly support the idea of the flood pulse as the main factor driving the composition pattern of the planktonic ciliates community in the Paraná River floodplain.

Spatial and temporal patterns of ciliate species composition (Protozoa: Ciliophora) in the plankton of the Upper Paraná River floodplain / Padrões espaciais e temporais da composição de espécies de ciliados (Protozoa: Ciliophora) no plâncton da planície de inundação do Alto Rio Paraná

Spatial and temporal patterns of plankton ciliates species composition in the Paraná River floodplain were investigated. Samplings were carried out in twelve environments in two distinct hydrological periods (limnophase and potamophase). A total of 61 species of ciliates were recorded, and among them 21 are classified as pelagic while 40 are considered preferentially as littoral species. The registered species belong to eleven orders, and among them, Prostomatida was the most specious followed by Hymenostomatida and Peritrichida. The ciliate species composition was significantly distinct between periods, but not among environments. In this way, typically pelagic species characterized the ciliate community during the limnophase period, while the littoral species were predominant during the potamophase period. Our results strongly support the idea of the flood pulse as the main factor driving the composition pattern of the planktonic ciliates community in the Paraná River floodplain.

No presente estudo foram investigados os padrões espaciais e temporais da composição de espécies de ciliados planctônicos na planície de inundação do Alto Rio Paraná. As amostras foram obtidas em 12 ambientes, em dois períodos hidrológicos distintos (limnofase e potamofase). Foram registradas 61 espécies de ciliados entre as quais 21 foram classificadas como pelágicas enquanto 40 foram consideradas preferencialmente litorâneas. As espécies registradas pertencem a 11 ordens, sendo Prostomatida foi a mais especiosa, seguida por Hymenostomatida e Peritrichida. A composição de espécies de ciliados foi significativamente distinta entre os períodos hidrológicos, enquanto que entre os ambientes, diferenças na composição não foram evidenciadas. Dessa forma, espécies tipicamente pelágicas caracterizaram a comunidade de ciliados na limnofase, enquanto que espécies litorâneas foram preponderantes para a composição de ciliados na potamofase. Os resultados suportam a idéia do pulso de inundação como principal fator controlador dos padrões de composição da comunidade de ciliados planctônicos na planície de inundação do Alto Rio Paraná.

Easy visualization of the protist Oxyrrhis marina grazing on a live fluorescently labelled heterotrophic nanoflagellate.

Planktonic heterotrophic flagellates are ubiquitous eukaryotic microorganisms that play a crucial role in carbon and nutrient fluxes through pelagic food webs. Here we illustrate for the first time a grazing model of planktonic dinoflagellate, Oxyrrhis marina, on the heterotrophic nanoflagellate Goniomonas amphinema, using the DNA-binding fluorescent dye Hoechst 33342. A solution of 1 microg/mL of the fluorochrome allowed viability of the prey for at least 48 hours, provided low fluorescence quenching, and labelled the flagellate without masking the cytoplasm. After 2 hours of contact between the fluorescent prey and the predator, O. marina population had preyed on live G. amphinema at an ingestion rate of 2.2 prey Oxyrrhis (-1) h(-1). Results show that this model is a time-effective and inexpensive approach for the direct observation of heterotrophic flagellate grazing. The fact that prey remain alive while predation occurs, as well as the low rate of quenching, could be of help in studying the fate of real-time trophic interactions between protists in microbial webs.

Detection of Schistosoma mansoni cercariae in plankton samples by PCR.

A PCR assay on the basis of a tandemly repeated DNA sequence was employed for the detection of Schistosoma mansoni in artificial plankton samples. It was highly specific, since as few as 1fg DNA from this species were sufficient to obtain a clear signal, while 10pg DNA of Schistosoma rodhaini were required and no PCR products were obtained with even 10ng DNA of planktonic organisms and any other trematode species tested. In areas with transmission of different Schistosoma species 10pg DNA should be used for amplification, which would allow detection of 20 S. mansoni cercariae in 0.05g plankton without interference caused by DNA of other Schistosoma species. In other areas 10ng DNA from plankton samples can be amplified, detecting less than one S. mansoni cercaria specifically in 0.05g plankton. This assay might help to identify S. mansoni in samples from field studies, where a multitude of different organisms hinder a correct species identification.

Description of a tandem repeated DNA sequence of Echinostoma caproni and methods for its detection in snail and plankton samples.

Echinostome larval stages in the snail have a great potential as effective competitors for the control of schistosomes and adult worms can cause painful intestinal diseases in humans. Ecology and transmission of the larval stages of trematodes are poorly understood, especially because their identification in field-collected samples by microscopy is difficult. We cloned, sequenced and analysed a 192 bp tandem repreated DNA sequence of Echinostoma caproni (EcSau3A), an often discussed antagonist of Schistosoma mansoni in Biomphalaria snails. PCR primers against this sequence can detect less than 10 fg of E. caproni DNA, 2 miracidia in snails 1 day p.i., 1 metacercaria in 50 mg snail tissue and 1 cercaria in 50 mg plankton with high specificity. Methods described in this study can support the discovery of fundamental ecological principles on distribution, host specificity and epidemiology of E. caproni larvae under field conditions.

Multiple strains of the parasitic dinoflagellate Amoebophrya exist in Chesapeake Bay.

Small subunit rRNA sequences were amplified from Amoebophrya strains infecting Karlodinium micrum, Gymnodinium instriatum and an unidentified Scrippsiella species in Chesapeake Bay. The alignable parts of the sequences differed from each other and from the previously reported rRNA sequence of the Amoebophrya strain infecting Akashiwo sanguinea in Chesapeake Bay by 4 to 10%. This is a greater degree of difference than sometimes found between sequences from separate genera of free-living dinoflagellates. These sequence differences indicate that the Amoebophrya strains parasitizing dinoflagellates in Chesapeake Bay do not all belong to the same species. In spite of their relative dissimilarity, the sequences do group together into a single clade with high bootstrap support in phylogenetic trees constructed from the sequences.

A new species of Acusicola Cressey (Copepoda: Ergasilidae) from northeastern Brazil.

A new species of Acusicola is described based on adults of both sexes taken from plankton samples collected in the upper reaches of the Piauí River estuary, in the northeast of Brazil. Ovigerous females were present in the plankton. The new species, Acusicola minuta n. sp., can be distinguished from its congeners by its small body size, female antennal morphology and leg setation. The male described here as A. minuta n. sp. is the first known male attributed to the genus.