Author

Peter L. Kahn

Date

11-2014

Document Type

Dissertation

Degree Name

Ph.D.

Institution

Oregon Health & Science University

Abstract

Heterotrophic protists, a diverse group of microbial eukaryotes characterized by great morphological variability and extensive taxonomic representation, play important ecological roles in aquatic food webs as prey and predators. Difficulty in heterotrophic protist identification has often resulted in lumping them into broad groups, but there is a crucial need to develop methods that increase the spatial and temporal resolution of observations applied to particular organisms in order to discover the drivers of population structure and ecological function. This research characterizes the spatiotemporal distribution of heterotrophic protist assemblages in the Columbia River coastal margin (including the tidal freshwater reaches of the river, the chemical estuary, and the river plume in the adjacent coastal ocean) using DNA sequence and morphological approaches. I analyzed partial small subunit (SSU) rRNA gene sequences of heterotrophic protists from the Columbia River estuary and plume during spring and summer using metagenomic next-generation sequencing (NGS) technology (Illumina HiSeq) and PCR (polymerase chain reaction)-based Sanger sequencing. Ciliates were the dominant heterotrophic protists in the estuary during both spring and summer according to analysis of the Illumina amplicon sequences, with a seasonal transition of abundant species occurring from spring to summer, while the heterotrophic flagellate Katablepharis sp. dominated spring protist assemblages estimated by the Sanger method. In the river plume, the assemblage transitioned from one dominated by ciliates in the spring to one dominated by heterotrophic dinoflagellates in the summer. Absolute cell abundances of ciliates and heterotrophic dinoflagellates were determined using light microscopy, and were strongly positively correlated with the relative proportions of sequences retrieved from the metagenomic dataset for both ciliates and heterotrophic dinoflagellates. However, correlations between the abundances of cells and SSU were weak when the PCR-based approach was used, suggesting that heterotrophic protist diversity is not adequately captured by this method. In contrast, metagenomics provided a reasonable estimate of protist diversity and abundance across a river-to-ocean continuum. Chapter Three details the discovery of a 332 base pair unique sequence element (USE) insertion in the large subunit rRNA gene of the Columbia River Estuary Katablepharis (Katablepharis CRE) that is not present in other katablepharids or any other organisms. Using USE-specific probes, I determined the spatial and temporal patterns of Katablepharis CRE in absolute abundance through quantitative approaches. The presence of USEs in several other protist taxa and the utility of these elements in tracking protist biogeography were determined in Chapter Four. USEs were detected in the parasitic dinoflagellate genus Euduboscquella and the flagellate groups Diplonemea and Cercozoa. The distributions of these taxa were tracked with USE-specific probes, and fine-scale genotypic differences were detected amongst closely related strains with putatively restricted biogeography. The USE presented here can be a useful tool for studying protist biogeography due to their highly specific nature, and has wide-reaching potential for a variety of environmental applications. Taken together, this dissertation provides the first detailed characterization of the seasonal distribution of heterotrophic protist assemblages in the Columbia River coastal margin, and presents new tools to track specific protist taxa distribution within a system and globally.

Identifier

doi:10.6083/M4N58K49

Division

Division of Environmental & Biomolecular Systems

School

School of Medicine

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