Diversity estimates of microeukaryotes below the chemocline of the anoxic Mariager Fjord, Denmark.

Microbial communities of extreme environments have often been assumed to have low species richness. We analysed 18S rRNA gene signatures in a sample collected below the chemocline of the anoxic Mariager Fjord in Denmark, and from these data we computed novel parametric and standard nonparametric estimates of protistan phylotype richness. Our results indicate unexpectedly high richness in this environment: at the 99.5% phylotype definition, our most conservative estimate was 568 phylotypes (+/-114, standard error). Phylogenetic analyses revealed that the sequences collected cover the majority of described lineages in the eukaryotic domain. Out of 384 sequences analysed, 307 were identified as protistan targets, none of which was identical to known sequences. However, based on what is known about species that are phylogenetically related to the Mariager sequences, most of the latter seem to belong to strictly or facultative anaerobe organisms. We also found signatures that together with other environmental 18S rRNA gene sequences represent environmental clades of possibly high taxonomic levels (class to kingdom level). One of these clades, consisting exclusively of sequences from anoxic sampling sites, branches at the base of the eukaryotic evolutionary tree among the earliest eukaryotic lineages. Assuming eukaryotic evolution under oxygen-depleted conditions, these sequences may represent immediate descendants of early eukaryotic ancestors.

Morphology, ultrastructure, molecular phylogeny, and autecology of Euplotes elegans Kahl, 1932 (Hypotrichida; Euplotidae) isolated from the anoxic Mariager Fjord, Denmark.

The morphology, autecology, and molecular phylogeny of an euryhaline Euplotes isolate collected from the anoxic water column of the Mariager Fjord in Denmark were investigated. The isolate matches the original description of Euplotes elegans Kahl, 1932 very well. However, its dorsal silverline system is clearly distinct from the redescription of this species by Tuffrau. Thus, a neotypification is proposed for E. elegans Kahl, 1932. The oval-shaped cell has a mean size of 107 x 51 microm and is characterized by 9.4 dorsolateral kineties, seven prominent dorsal ridges, large elongated ampullae, which encircle the dorsal kinetids, 18 kinetids in the middorsal row, nine frontoventral cirri, five transversal cirri, and three caudal cirri (two right caudal cirri and one left marginal cirrus). The dorsal silverline system is of the double type with the narrow polygons located on the right side of the dorsal kinetids. The ecological tolerances of this species to pH, salinity, temperature, and oxygen match the ambient environmental conditions of the sampling site. Molecular phylogeny was studied using small subunit rRNA (SSU rRNA) gene sequences. The molecular data cluster E. elegans with Euplotes raikovi, a member of the Euplotopsis group. The data suggest that the E. elegans-E. raikovi clade represents an isolated and deep branch at the base of the Euplotes tree.

A molecular approach to identify active microbes in environmental eukaryote clone libraries.

A rapid method for the simultaneous extraction of RNA and DNA from eukaryote plankton samples was developed in order to discriminate between indigenous active cells and signals from inactive or even dead organisms. The method was tested using samples from below the chemocline of an anoxic Danish fjord. The simple protocol yielded RNA and DNA of a purity suitable for amplification by reverse transcription-polymerase chain reaction (RT-PCR) and PCR, respectively. We constructed an rRNA-derived and an rDNA-derived clone library to assess the composition of the microeukaryote assemblage under study and to identify physiologically active constituents of the community. We retrieved nearly 600 protistan target clones, which grouped into 84 different phylotypes (98% sequence similarity). Of these phylotypes, 27% occurred in both libraries, 25% exclusively in the rRNA library, and 48% exclusively in the rDNA library. Both libraries revealed good correspondence of the general community composition in terms of higher taxonomic ranks. They were dominated by anaerobic ciliates and heterotrophic stramenopile flagellates thriving below the fjord's chemocline. The high abundance of these bacterivore organisms points out their role as a major trophic link in anoxic marine systems. A comparison of the two libraries identified phototrophic dinoflagellates, "uncultured marine alveolates group I," and different parasites, which were exclusively detected with the rDNA-derived library, as nonindigenous members of the anoxic microeukaryote community under study.