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Identification of Associations between Bacterioplankton and Photosynthetic Picoeukaryotes in Coastal Waters.

Farnelid HM, Turk-Kubo KA, Zehr JP - Front Microbiol (2016)

Bottom Line: Photosynthetic picoeukaryotes are significant contributors to marine primary productivity.The results show that diverse bacterial phylotypes are found in association with photosynthetic picoeukaryotes.Taxonomic identification of these associations is a prerequisite for further characterizing and to elucidate their metabolic pathways and ecological functions.

View Article: PubMed Central - PubMed

Affiliation: Ocean Sciences Department, University of California at Santa CruzSanta Cruz, CA, USA; Centre for Ecology and Evolution in Microbial Model Systems, Linnaeus UniversityKalmar, Sweden.

ABSTRACT
Photosynthetic picoeukaryotes are significant contributors to marine primary productivity. Associations between marine bacterioplankton and picoeukaryotes frequently occur and can have large biogeochemical impacts. We used flow cytometry to sort cells from seawater to identify non-eukaryotic phylotypes that are associated with photosynthetic picoeukaryotes. Samples were collected at the Santa Cruz wharf on Monterey Bay, CA, USA during summer and fall, 2014. The phylogeny of associated microbes was assessed through 16S rRNA gene amplicon clone and Illumina MiSeq libraries. The most frequently detected bacterioplankton phyla within the photosynthetic picoeukaryote sorts were Proteobacteria (Alphaproteobacteria and Gammaproteobacteria) and Bacteroidetes. Intriguingly, the presence of free-living bacterial genera in the photosynthetic picoeukaryote sorts could suggest that some of the photosynthetic picoeukaryotes were mixotrophs. However, the occurrence of bacterial sequences, which were not prevalent in the corresponding bulk seawater samples, indicates that there was also a selection for specific OTUs in association with photosynthetic picoeukaryotes suggesting specific functional associations. The results show that diverse bacterial phylotypes are found in association with photosynthetic picoeukaryotes. Taxonomic identification of these associations is a prerequisite for further characterizing and to elucidate their metabolic pathways and ecological functions.

No MeSH data available.


Example of a flow cytogram with the cell populations that were targeted in this study. Red fluorescence was used as a proxy for chlorophyll a content and forward scatter (FSC) as a proxy for size. The indicated cell populations are Syn (Synechococcus) as defined by phycoerythrin content (Orange fluorescence), and two size fractions of photosynthetic picoeukaryotes, small (P1, ∼ <3 μm beads) and large (P2, ∼ >3 μm beads).
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Figure 1: Example of a flow cytogram with the cell populations that were targeted in this study. Red fluorescence was used as a proxy for chlorophyll a content and forward scatter (FSC) as a proxy for size. The indicated cell populations are Syn (Synechococcus) as defined by phycoerythrin content (Orange fluorescence), and two size fractions of photosynthetic picoeukaryotes, small (P1, ∼ <3 μm beads) and large (P2, ∼ >3 μm beads).

Mentions: Populations of photosynthetic picoeukaryotes were sorted using a BD Biosciences Influx Cell Sorter equipped with a small particle detector (BD Biosciences, San Jose, CA, USA) and a 488 nm Sapphire laser (Coherent, Santa Clara, CA, USA). The sorting was done using sterile filtered BioSure Sheath fluid diluted in ultrapure water to 1x concentration. Prior to flow cytometric sorting, the seawater samples were prefiltered using a 30 μm filter (Partec CellTrics, Swedesboro, NJ, USA) to prevent clogging of the 70 μm diameter nozzle. To minimize the risk of dislodging associations there was no concentration step and only fresh samples were processed. Data collection and sorting was triggered in the forward scatter (FSC) channel. The sorting gates were constructed in the BD FACS software using FSC as a proxy for cell size and red fluorescence (692–740 nm) as a proxy for chlorophyll a content. Photosynthetic picoeukaryotes were distinguished from Synechococcus populations using a NOT gate, deselecting cells with orange fluorescence (572–627 nm; i.e., phycoerythrin containing cells). For photosynthetic picoeukaryotes a smaller (P1, ∼ <3 μm beads) and a larger (P2, ∼ >3 μm beads) population was sorted (see Figure 1 and Supplementary Figure 1). All sorts were collected in “1.0 Drop Purity” mode, and in combination with the small particle detection capabilities and triggering sorts on the FSC signal, this configuration is the optimal set-up for ensuring a low probability of co-sorting of unattached particles. Cell counts and cytograms were processed in FlowJo v10.0.7 (Tree Star). To minimize the risk of contamination, fluidic lines were regularly decontaminated with 1% bleach.


Identification of Associations between Bacterioplankton and Photosynthetic Picoeukaryotes in Coastal Waters.

Farnelid HM, Turk-Kubo KA, Zehr JP - Front Microbiol (2016)

Example of a flow cytogram with the cell populations that were targeted in this study. Red fluorescence was used as a proxy for chlorophyll a content and forward scatter (FSC) as a proxy for size. The indicated cell populations are Syn (Synechococcus) as defined by phycoerythrin content (Orange fluorescence), and two size fractions of photosynthetic picoeukaryotes, small (P1, ∼ <3 μm beads) and large (P2, ∼ >3 μm beads).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4834442&req=5

Figure 1: Example of a flow cytogram with the cell populations that were targeted in this study. Red fluorescence was used as a proxy for chlorophyll a content and forward scatter (FSC) as a proxy for size. The indicated cell populations are Syn (Synechococcus) as defined by phycoerythrin content (Orange fluorescence), and two size fractions of photosynthetic picoeukaryotes, small (P1, ∼ <3 μm beads) and large (P2, ∼ >3 μm beads).
Mentions: Populations of photosynthetic picoeukaryotes were sorted using a BD Biosciences Influx Cell Sorter equipped with a small particle detector (BD Biosciences, San Jose, CA, USA) and a 488 nm Sapphire laser (Coherent, Santa Clara, CA, USA). The sorting was done using sterile filtered BioSure Sheath fluid diluted in ultrapure water to 1x concentration. Prior to flow cytometric sorting, the seawater samples were prefiltered using a 30 μm filter (Partec CellTrics, Swedesboro, NJ, USA) to prevent clogging of the 70 μm diameter nozzle. To minimize the risk of dislodging associations there was no concentration step and only fresh samples were processed. Data collection and sorting was triggered in the forward scatter (FSC) channel. The sorting gates were constructed in the BD FACS software using FSC as a proxy for cell size and red fluorescence (692–740 nm) as a proxy for chlorophyll a content. Photosynthetic picoeukaryotes were distinguished from Synechococcus populations using a NOT gate, deselecting cells with orange fluorescence (572–627 nm; i.e., phycoerythrin containing cells). For photosynthetic picoeukaryotes a smaller (P1, ∼ <3 μm beads) and a larger (P2, ∼ >3 μm beads) population was sorted (see Figure 1 and Supplementary Figure 1). All sorts were collected in “1.0 Drop Purity” mode, and in combination with the small particle detection capabilities and triggering sorts on the FSC signal, this configuration is the optimal set-up for ensuring a low probability of co-sorting of unattached particles. Cell counts and cytograms were processed in FlowJo v10.0.7 (Tree Star). To minimize the risk of contamination, fluidic lines were regularly decontaminated with 1% bleach.

Bottom Line: Photosynthetic picoeukaryotes are significant contributors to marine primary productivity.The results show that diverse bacterial phylotypes are found in association with photosynthetic picoeukaryotes.Taxonomic identification of these associations is a prerequisite for further characterizing and to elucidate their metabolic pathways and ecological functions.

View Article: PubMed Central - PubMed

Affiliation: Ocean Sciences Department, University of California at Santa CruzSanta Cruz, CA, USA; Centre for Ecology and Evolution in Microbial Model Systems, Linnaeus UniversityKalmar, Sweden.

ABSTRACT
Photosynthetic picoeukaryotes are significant contributors to marine primary productivity. Associations between marine bacterioplankton and picoeukaryotes frequently occur and can have large biogeochemical impacts. We used flow cytometry to sort cells from seawater to identify non-eukaryotic phylotypes that are associated with photosynthetic picoeukaryotes. Samples were collected at the Santa Cruz wharf on Monterey Bay, CA, USA during summer and fall, 2014. The phylogeny of associated microbes was assessed through 16S rRNA gene amplicon clone and Illumina MiSeq libraries. The most frequently detected bacterioplankton phyla within the photosynthetic picoeukaryote sorts were Proteobacteria (Alphaproteobacteria and Gammaproteobacteria) and Bacteroidetes. Intriguingly, the presence of free-living bacterial genera in the photosynthetic picoeukaryote sorts could suggest that some of the photosynthetic picoeukaryotes were mixotrophs. However, the occurrence of bacterial sequences, which were not prevalent in the corresponding bulk seawater samples, indicates that there was also a selection for specific OTUs in association with photosynthetic picoeukaryotes suggesting specific functional associations. The results show that diverse bacterial phylotypes are found in association with photosynthetic picoeukaryotes. Taxonomic identification of these associations is a prerequisite for further characterizing and to elucidate their metabolic pathways and ecological functions.

No MeSH data available.