High throughput sequencing technology has great promise for biodiversity studies. However, an underlying assumption is that the primers used in these studies are universal for the prokaryotic or eukaryotic groups of interest. Full primer universality is difficult or impossible to achieve and studies using different primer sets make biodiversity comparisons problematic. The aim of this study was to design and optimize universal eukaryotic primers that could be used as a standard in future biodiversity studies. Using the alignment of all eukaryotic sequences from the publicly available SILVA database, we generated a full characterization of variable versus conserved regions in the 18S rRNA gene. All variable regions within this gene were analyzed and our results suggested that the V2, V4 and V9 regions were best suited for biodiversity assessments. Previously published universal eukaryotic primers as well as a number of self-designed primers were mapped to the alignment. Primer selection will depend on sequencing technology used, and this study focused on the 454 pyrosequencing GS FLX Titanium platform. The results generated a primer pair yielding theoretical matches to 80% of the eukaryotic and 0% of the prokaryotic sequences in the SILVA database. An empirical test of marine sediments using the AmpliconNoise pipeline for analysis of the high throughput sequencing data yielded amplification of sequences for 71% of all eukaryotic phyla with no isolation of prokaryotic sequences. To our knowledge this is the first characterization of the complete 18S rRNA gene using all eukaryotes present in the SILVA database, providing a robust test for universal eukaryotic primers. Since both in silico and empirical tests using high throughput sequencing retained high inclusion of eukaryotic phyla and exclusion of prokaryotes, we conclude that these primers are well suited for assessing eukaryote diversity, and can be used as a standard in biodiversity studies.

The use of DNA as a marker for prey inside the gut of predators has been instrumental in further understanding of known and unknown interactions. Molecular approaches are in particular useful in unavailable environments like the deep sea. Trophic interactions in the deep sea are difficult to observe in situ, correct deep-sea experimental laboratory conditions are difficult to obtain, animals rarely survive the sampling, or the study organisms feed during the sampling due to long hauls. Preliminary studies of vent and seep systems in the Nordic Seas have identified the temperate-cold-water pelagic amphipod Themisto abyssorum as a potentially important predator in these chemosynthetic habitats. However, the prey of this deep-sea predator is poorly known, and we applied denaturing high performance liquid chromatography (DHPLC) to investigate the predator-prey interactions of T. abyssorum in deep-water vent and seep systems. Two deep-water hydrothermally active localities (The Jan Mayen and Loki's Castle vent fields) and one cold seep locality (The Håkon Mosby mud volcano) in the Nordic Seas were sampled, genomic DNA of the stomachs of T. abyssorum was extracted, and 18S rDNA gene was amplified and used to map the stomach content. We found a wide range of organisms including micro-eukaryotes, metazoans and detritus. Themisto abyssorum specimens from Loki's Castle had the highest diversity of prey. The wide range of prey items found suggests that T. abyssorum might be involved in more than one trophic level and should be regarded as an omnivore and not a strict carnivore as have previously been suggested.

Acute myeloid leukemia (AML) frequently comprises mutations in genes that cause perturbation in intracellular signaling pathways, thereby altering normal responses to growth factors and cytokines. Such oncogenic cellular signal transduction may be therapeutic if targeted directly or through epigenetic regulation. We treated 24 selected elderly AML patients with all-trans retinoic acid for 2 days before adding theophylline and the histone deacetylase inhibitor valproic acid (ClinicalTrials.gov NCT00175812; EudraCT no. 2004-001663-22), and sampled 11 patients for peripheral blood at day 0, 2 and 7 for single-cell analysis of basal level and signal-transduction responses to relevant myeloid growth factors (granulocyte-colony-stimulating factor, granulocyte/macrophage-colony-stimulating factor, interleukin-3, Flt3L, stem cell factor, erythropoietin, CXCL-12) on 10 signaling molecules (CREB, STAT1/3/5, p38, Erk1/2, Akt, c-Cbl, ZAP70/Syk and rpS6). Pretreatment analysis by unsupervised clustering and principal component analysis divided the patients into three distinguishable signaling clusters (non-potentiated, potentiated basal and potentiated signaling). Signal-transduction pathways were modulated during therapy and patients moved between the clusters. Patients with multiple leukemic clones demonstrated distinct stimulation responses and therapy-induced modulation. Individual signaling profiles together with clinical and hematological information may be used to early identify AML patients in whom epigenetic and signal-transduction targeted therapy is beneficial.

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