APPLICATION OF DIGITAL INLINE HOLOGRAPHY FOR THE STUDY OF PARTICLES AND MICROPLANKTON IN THE OCEAN
Bochdansky, A. B.1and Herndl, G.J.2
1Old Dominion University, Norfolk, Virginia, USA, 2University of Vienna, Austria
In situ microscopy in aquatic systems is typically limited by the depth of field and consequently by a low image volume. This makes traditional microscopy unsuit-able for concentrations at which most aquatic microorganisms occur. Digital Inline Holographic Microscopy (DIHM) overcomes this problem by utilizing large conical fields of view containing up to several milliliters of water in one image.
Resolution (limited by the wavelength of the laser, the size of the point source and the resolution of the chip of the lensless camera) can reach into the micrometer range under ideal conditions. Here we present for the first time the application of DIHM on a basin-scale oceanographic expedition to the tropical and subtropical Atlantic Ocean, from the surface to a maximum depth of 6000 m. The DIHM was mounted on the frame of a CTD rosette, and thus became part of routine casts without the requirement of separate deployments. Even at speeds of 1 m per sec through the water, our instrument yielded clearly identifiable particles such as Trichodesmium colonies, radiolarians and tintinnids. Approximately 7 images with sizes of 2048x2048 pixels each were taken for every meter, each image represen-ting a volume of 1.3 ml. An additional important advantage of holographic images is that they allow for the precise determination of sizes and the locations of particles in three-dimensional space (i.e.,X,Y,Z coordinates). In this presentation we will show the principles of DIHM, our adaptations for oceanographic environ-ments, and show in live demonstrations how images of microplankton and amorphous particles are reconstructed from the original holograms.
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Novel Technologies and Methods
OT-6
DIEL GENE EXPRESSION PROFILES OF A PHOSPHORUS LIMITED HIGH-ALTITUDE
LAKE THROUGH METATRANSCRIPTOMICS
Vila-Costa, M.1, Sharma, S.1, Moran, MA.1, Casamayor, EO.1
1Centre Estudis Avançats de Blanes (CEAB-CSIC)
Contrastingly to the increasing advances in marine ecosystems, bacterial func-tionality of freshwater systems remains largely unexplored despite their relevance in the biogeochemical cycles. In this work, we used metatranscriptomic sequenc-ing to analyze day and night gene expression profiles of the bacterial assemblages from the oligotrophic high altitude Lake Llebreta (1620 m above sea level) in the Limnological Observatory of the Pyrenees (LOOP, Central Pyrenees, 42º 33’3”N, 0º 53’25”E). The goal of the study was to obtain clues about the main biogeo-chemical processes carried out by bacteria in a high altitude lake and explore unique biogeochemical features of this freshwater system. An average of 37871 unique reads were obtained per treatment after using 454 pyrosequencing technology to sequence the extracted and amplified messenger RNA (mRNA).
Bacteroidetes and betaproteobacteria were the most actively transcribing phyla of the community and they showed different strategies to process limiting nutrients like phosphorus (P). Thus, whereas bacteroidetes mostly obtained P by hydro-lization of stored polyphosphates and pyrophosphates generally at night and used proteorhodopsines as extra source of energy during the day, betaproteobacteria accounted for most of the phosphate membrane uptake systems and generated additional ATP molecules after CO oxidation during the day.Compared to marine diel studies, lake Llebreta showed an overabundance of transcripts related to lipid metabolism and specifically, an overrepresentation of transcripts related to degradation of polycyclic aromatic hydrocarbons (PAHs), which was over-represented at night. This study represents the first attempt to characterize the bacterial functionality of a freshwater system.
OT-7
Novel Technologies and Methods
METAGENOMIC AND METATRANSCRIPTOMIC ANALYSIS OF SULFIDIC WATERS WITHIN THE OXYGEN MINIMUM ZONE OFF PERU
Schunck, H.1, Desai, D.1, Großkopf, T.1, Loescher, C.2, Schmitz-Streit, R.2, Lavik, G.3, Kuypers, M.3, and LaRoche, J.1
1Leibniz Institute of Marine Sciences (IFM-GEOMAR), Kiel, Germany,
2Institute for General Microbiology, Christian-Albrechts University Kiel, Germany,
3Max-Planck-Institute of Marine Microbiology, Bremen, Germany
Oxygen minimum zones are hotspots of biogeochemical turnover processes. Nitrate and nitrite are usually thought to be the most favourable terminal electron accep-tors in the absence of oxygen. But also the cycling of sulfur species can contribute largely to the energy flux in oxygen depleted environments. So far, only few studies have investigated the microbial community in sulfidic waters. Hence, the knowledge of many bacterial and archaeal taxa and their contribution to biogeo-chemical cycles is still very limited.
Here we present sequencing data from a sulfidic water column on the continental shelf off Peru within the oxygen minimum zone in the Eastern Tropical South Pacific. Approximately 3.5 million DNA and cDNA sequences were obtained from six depths using the Roche GS-FLX sequencing technology.
The sequence data shows a diverse microbial community with (in some depths) almost 40% of the sequences belonging to organisms which actively contribute to the sulfur cycle. Although the organisms able to oxidize sulfur were dominating, a considerable number of sulfate reducers were identified in the water column as well. The most abundant microorganisms in the sulfidic waters could be identified as gammaproteobacteria affiliated with uncultured chemoautotrophic gill sym-bionts of deep-sea clams, in some depths accounting for up to 30% of the whole microbial community. Recent studies have postulated that these organisms can couple sulfide oxidation with nitrate reduction and thus potentially contribute largely to the loss of fixed nitrogen.
The comparison of our data to those of non-sulfidic oxygen minimum zone waters will deepen the understanding of the metabolic activity of microorganisms and their contribution to biogeochemical transformations within the oxygen deficient waters.
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Novel Technologies and Methods
OT-8
MICROBIAL DIVERSITY ALONG AN OXYGEN GRADIENT –COMMUNITY GENOMES FROM LANDSORT DEEP IN THE BALTIC SEA
Lundin, D.1
1SciLifeLab, KTH Royal School of Technology, Stockholm, Sweden
The Landsort Deep in the Baltic Sea south east of Stockholm, has a steep oxygen gradient from fully oxic surface waters, via an hypoxic zone at the halocline between 60 and 80m, down to anoxic conditions further down. We have charac-terised the microbial communities at three depths along the water column (10m, 75m and 400m) plus the sediment, using community genome shotgun 454 sequencing, yielding a total of 350Mb DNA. Bacteria are dominating all samples except for high frequencies of potentially ammonium oxidising Thaumarchaea in the 75m and 400m samples. Along the depth profile not only salinity and oxygen content varies, but also concentrations of e.g. phosphate, nitrate and sulfide. This is reflected in the taxonomical profile of our samples, with e.g. high frequencies of sulfate reducers in the anaerobic 400m sample. Furthermore, the degree of simi-larity, both in terms of taxonomical composition and functional coding capacity, between the 10m and 75m samples compared with the anaerobic samples, suggest that the ranges of many bacterial populations stretch down to the halocline. We also find a striking similarity between Landsort Deep 400m and sediment samples and other marine sediment and terrestrial soil community genomes.