High-resolution monitoring of marine protists based on an observation strategy integrating automated on-board ship filtration and molecular analyses

Information on recent biomass distribution and biogeography of photosynthetic marine protists with adequate temporal and spatial resolution is urgently needed to better understand consequences of environmental change for marine ecosystems. Here we introduce and review a molecular-based observation strategy for high resolution assessment of these protists in space and time. It is the result of extensive technology developments, adaptations and evaluations which are documented in a number of different publications and the results of recently accomplished field testing, which are introduced in this review. The observation strategy is organized at four different levels. At level 1, samples are collected at high spatio-temporal resolution using the remote-controlled automated filtration system AUTOFIM. Resulting samples can either be preserved for later laboratory analyses, or directly subjected to molecular surveillance of key species aboard the ship via an automated biosensor system or quantitative polymerase chain reaction (level 2). Preserved samples are analyzed at the next observational levels in the laboratory (level 3 and 4). This involves at level 3 molecular fingerprinting methods for a quick and reliable overview of differences in protist community composition. Finally, selected samples can be used to generate a detailed analysis of taxonomic protist composition via the latest Next Generation Sequencing Technology (NGS) at level 4. An overall integrated dataset of the results based on the different analyses provides comprehensive information on the diversity and biogeography of protists, including all related size classes. At the same time the cost effort of the observation is optimized in respect to analysis effort and time

A molecular survey of protist diversity through the central Arctic Ocean

The protist assemblage in the central Arctic Ocean is scarcely surveyed despite them being the major primary producers. Elucidating their response to changing environmental variables requires an a priori analysis of their current diversity, including abundant and rare species. In late summer 2011, samples were collected during the ARK-XXVI/3 expedition (RV Polarstern) to study Arctic protist community structures, by implementation of automated ribosomal intergenic spacer analysis (ARISA) and 454-pyrosequencing. Protist assemblages were related to the hydrology and environmental variables (temperature, salinity, ice coverage, nitrate, phosphate, and silicate). The abundant (a parts per thousand yen1 %) biosphere and rare (< 1 %) biosphere were considered separately in the diversity analysis in order to reveal their mutual relationships. A relation between hydrology and protist community structure was highly supported by ARISA and partially by 454-pyrosequencing. Sea ice showed a stronger influence on the local community structure than nutrient availability, making statements on the water mass influence more difficult. Dinoflagellates (Syndiniales), chlorophytes (Micromonas spp.), and haptophytes (Phaeocystis spp.) were important contributors to the abundant biosphere, while other dinoflagellates and stramenopiles dominated the rare biosphere. No significant correlation was found between the abundant and rare biosphere. However, relative contributions of major taxonomic groups revealed an unexpected stable community structure within the rare biosphere, indicating a potential constant protist reservoir. This study provides a first molecular survey of protist diversity in the central Arctic Ocean, focusing on the diversity and distribution of abundant and rare protists according to the environmental conditions, and can serve as baseline for future analysis

Phytoplankton diversity and distribution in the Pacific sector of the Southern Ocean

Rising water temperatures and ocean acidification are the major threats for polar marine ecosystems and will affect phytoplankton communities. Phytoplankton plays a major role in primary production and biogeochemical cycles and forms the basis of marine food webs. Changes in the composition and distribution of phytoplankton will affect the whole marine ecosystem. To assess the effects of changing environmental conditions on phytoplankton communities we have to know their current diversity and distribution. There is a lack of Phytoplankton diversity studies in the Pacific sector of the Southern Ocean, especially in the Amundsen Sea. To resolve this gap this study will deliver basic data of phytoplankton diversity and distribution, which will help identifying the dominant phytoplankton phyla and provide information on the rare biosphere in that area. Environmental samples, taken on the RV Polarstern cruise ANT XXVI/3, were analyzed with molecular approaches, including ARISA (automated ribosomal intergenic spacer analysis) and 454-pyrosequencing. Furthermore pigment analysis and flow cytometry were conducted. First results indicate a clustering of the samples according to the different water masses and regions with comparable environmental conditions. The sequencing will deliver more detailed information about the structure and diversity of phytoplankton in the Pacific sector of the Southern Ocean

Assessment of eukaryotic communities in environmental samples: A workflow comparison for next-generation sequencing data

To understand function and stability of ecosystems it is crucial to gain insights into their species composition, particulary in the face of global warming. Next Generation Sequencing (NGS) is the method of choice for getting fast overviews of species diversity in a high number of samples. Currently, there are lively discussions about bioinformatic techniques to enhance the quality of sequencing outputs and how to post process these data in order to estimate the “real” diversity as precisely as possible. In this study, we analyzed the protist composition of three water samples, collected in the Fram Strait in 2010. We compared different potential sequencing error corrected and uncorrected datasets, which were generated with widely used open-source software: QIIME, mothur and PhyloAssigner. Relative abundance of protist phyla was hardly affected by the choice of the software, quality filtering and error correction. However, the outputs differed strongly in relative abundance of diatom genera and were not comparable to dominant diatoms observed with light microscopy. Our main findings are beneficial for the enhancement of study design, data preparation and interpretation and gives insights into the optimization potential of NGS experiments in general

Protist community composition in the Pacific sector of the Southern Ocean during austral summer 2010

Knowledge about the protist diversity of the Pacific sector of the Southern Ocean is scarce. We tested the hypothesis that distinct protist community assemblages characterize large-scale water masses. Therefore, we determined the composition and biogeography of late summer protist assemblages along a transect from the coast of New Zealand to the eastern Ross Sea. We used state of the art molecular approaches, such as automated ribosomal intergenic spacer analysis and 454-pyrosequencing, combined with high-performance liquid chromatography pigment analysis to study the protist assemblage. We found distinct biogeographic patterns defined by the environmental conditions in the particular region. Different water masses harbored different microbial communities. In contrast to the Arctic Ocean, picoeukaryotes had minor importance throughout the investigated transect and showed very low contribution south of the Polar Front. Dinoflagellates, Syndiniales, and small stramenopiles were dominating the sequence assemblage in the Subantarctic Zone, whereas the relative abundance of diatoms increased southwards, in the Polar Frontal Zone and Antarctic Zone. South of the Polar Front, most sequences belonged to haptophytes. This study delivers a comprehensive and taxon detailed overview of the protist composition in the investigated area during the austral summer 2010

Material Science Lab operations onboard the International Space Station

The Materials Science Laboratory (MSL) onboard the International Space Station is designed for the research on solidification processes under microgravity conditions. It is equipped with two exchangeable furnace inserts of Bridgman-type allowing temperatures of up to 1400 ◦C. MSL is operated under ESA contract by the Microgravity User Support Center (MUSC) at DLR in Cologne in collaboration with Marshal Space Flight Center at Huntsville which is responsible for the Materials Science Research Rack (MSRR) which hosts MSL and provides services. MSL was launched in 2009 and installed in the US Destiny laboratory module. Since then a number of experiments by different project teams have been performed and research is still ongoing. Since 2018, a new type of cartridges developed by NASA allows investigation of sintering processes within MSL. The paper will give a survey on MSL operations over the last decade and provide an outlook for future MSL planning

Regional variability in eukaryotic protist communities in the Amundsen Sea

We determined the composition and structure of late summer eukaryotic protist assemblages along a west–east transect in the Amundsen Sea. We used state-of-the-art molecular approaches, such as automated ribosomal intergenic spacer analysis (ARISA) and 454-pyrosequencing, combined with pigment measurements via high performance liquid chromatography (HPLC) to study the protist assemblage. We found characteristic offshore and inshore communities. In general, total chlorophyll a and microeukaryotic contribution were higher in inshore samples. Diatoms were the dominant group across the entire area, of which Eucampia sp. and Pseudo-nitzschia sp. were dominant inshore and Chaetoceros sp. was dominant offshore. At the most eastern station, the assemblage was dominated by Phaeocystis sp. Under the ice, ciliates showed their highest and haptophytes their lowest abundance. This study delivers a taxon detailed overview of the eukaryotic protist composition in the Amundsen Sea during the summer 2010

Monitoring of Arctic marine protists via an observation strategy integrating automated under-way filtration, long-term sediment traps and molecular analyses

nformation on recent diversity and biogeography of Arctic marine protists with adequate temporal and spatial resolution is urgently needed to better understand consequences of environmental change for marine ecosystems. Here, we introduce a molecular-based observation strategy for high resolution assessment of marine protists in space and time, even in remote areas such as the Arctic Ocean. The observation strategy involves molecular analyses (e.g. Next Generation Sequencing (NGS) or quantitative PCR) of samples, collected with a set of complementary methods such as a newly developed automated under-way sampling device, CTD-casts and moored sediment traps. This integrated approach allows generating detailed information on marine protist community composition or abundance with adequate resolution. Currently, the observation strategy is organized at four major levels. At level 1, samples are collected at high spatial and temporal resolution based on under-way sampling with the remote-controlled automated filtration system AUTOFIM (developed in the COSYNA-project), and sampling at fixed stations based on CTD-casts and moored sediment traps. Resulting samples can either be preserved for later laboratory analyses, or directly subjected to molecular surveillance of key species aboard the ship, e.g. via quantitative polymerase chain reaction (level 2). Preserved samples are analyzed at the next observational levels in the laboratory (level 3 and 4). This involves at level 3 molecular fingerprinting methods for a quick and reliable overview of differences in protist community composition. Finally, selected samples can be used to generate a detailed analysis of taxonomic protist composition via the latest Next Generation Sequencing Technology (NGS) at level 4. An overall integrated dataset of all results provides comprehensive information on the diversity and biogeography of protists, including all related size classes. In the future, the observation strategy for Arctic marine protists will be part of the Molecular Microbial Observatory envisioned for the Arctic observatory FRAM (Frontiers in Arctic Monitoring)