Collecting and Culturing Kamptozoans for Regenerative Studies.

Kamptozoa, also known as Entoprocta, are small aquatic filter-feeders that belong to the Lophotrochozoan superphylum, which also contains other acoelomate phyla including Annelida, Nemertea, and Mollusca. The study of Kamptozoa is thus of great interest to understand the early Lophotrochozoan evolution. Moreover, many kamptozoans have been shown to possess great regeneration capacities, including whole-body regeneration. In addition, and in particular for colonial cosmopolitan species such as Barentsia benedeni, kamptozoans are highly suitable as laboratory model organisms because of their simple culture, low space requirements, transparency and rapid life cycle. This chapter provides a brief introduction into field collection, culturing techniques for both the animals as well as the algae required for their feeding, fixation, staining, and sequencing.

The rheology and foamability of crystal-melt suspensions composed of triacylglycerols.

The rheology of triacylglycerol (TAG) crystal-melt suspensions (CMSs) consisting of anhydrous milk fat (AMF), cocoa butter (CB), and palm kernel oil (PKO) as function of crystallization shear rate cryst and crystal volume fraction ΦSFC is investigated by in-line ultrasound velocity profiling - pressure difference (UVP-PD) rheometry. Measurements up to ΦSFC = 8.8% are presented. Below the percolation threshold Φc, no yield stress τ0 is observed and the viscosity η scales linearly with ΦSFC. Above Φc, a non-linear dependency of both τ0 and η as function of ΦSFC is apparent. For AMF and CB, the increase in cryst leads to a decrease in η and τ0 as function of ΦSFC, whereas for PKO based CMSs the opposite is the case. Scanning electron microscopy (SEM) and polarized light microscopy (PLM) relate these rheological findings to the microstructure of the investigated CMSs by taking the effective aspect ratio aeff and the concept of the effective crystal volume fraction ΦeffSFC into account. Foam formation by dynamically enhanced membrane foaming (DEMF) is performed directly after crystallization and reveals that depending on the CMS rheology and crystallite-, crystallite cluster- and crystal floc microstructure, a wide range of gas volume fractions between 0.05-0.6 are achievable.

Bilirubin-A Possible Prognostic Mortality Marker for Patients with ECLS.

Extracorporeal life support (ECLS) is a promising therapeutic option for patients with refractory cardiogenic shock. However, as the mortality rate still remains high, there is a need for early outcome parameters reflecting therapy success or futility. Therefore, we investigated whether liver enzyme levels could serve as prognostic mortality markers for patients with ECLS. The present study is a retrospective single-center cohort study. Adult patients >18 years of age who received ECLS therapy between 2011 and 2018 were included. Bilirubin, glutamic-oxaloacetic transaminase (GOT), and glutamic-pyruvic-transaminase (GPT) serum levels were analyzed at day 5 after the start of the ECLS therapy. The primary endpoint of this study was all-cause in-hospital mortality. A total of 438 patients received ECLS during the observation period. Based on the inclusion criteria, 298 patients were selected for the statistical analysis. The overall mortality rate was 42.6% (n = 127). The area under the curve (AUC) in the receiver operating characteristic curve (ROC) for bilirubin on day 5 was 0.72 (95% confidence interval (CI): 0.66-0.78). Cox regression with multivariable adjustment revealed a significant association between bilirubin on day 5 and mortality, with a hazard ratio (HR) of 2.24 (95% CI: 1.53-3.30). Based on the results of this study, an increase in serum bilirubin on day 5 of ECLS therapy correlates independently with mortality.

Phylogenomics of Lophotrochozoa with Consideration of Systematic Error.

Phylogenomic studies have improved understanding of deep metazoan phylogeny and show promise for resolving incongruences among analyses based on limited numbers of loci. One region of the animal tree that has been especially difficult to resolve, even with phylogenomic approaches, is relationships within Lophotrochozoa (the animal clade that includes molluscs, annelids, and flatworms among others). Lack of resolution in phylogenomic analyses could be due to insufficient phylogenetic signal, limitations in taxon and/or gene sampling, or systematic error. Here, we investigated why lophotrochozoan phylogeny has been such a difficult question to answer by identifying and reducing sources of systematic error. We supplemented existing data with 32 new transcriptomes spanning the diversity of Lophotrochozoa and constructed a new set of Lophotrochozoa-specific core orthologs. Of these, 638 orthologous groups (OGs) passed strict screening for paralogy using a tree-based approach. In order to reduce possible sources of systematic error, we calculated branch-length heterogeneity, evolutionary rate, percent missing data, compositional bias, and saturation for each OG and analyzed increasingly stricter subsets of only the most stringent (best) OGs for these five variables. Principal component analysis of the values for each factor examined for each OG revealed that compositional heterogeneity and average patristic distance contributed most to the variance observed along the first principal component while branch-length heterogeneity and, to a lesser extent, saturation contributed most to the variance observed along the second. Missing data did not strongly contribute to either. Additional sensitivity analyses examined effects of removing taxa with heterogeneous branch lengths, large amounts of missing data, and compositional heterogeneity. Although our analyses do not unambiguously resolve lophotrochozoan phylogeny, we advance the field by reducing the list of viable hypotheses. Moreover, our systematic approach for dissection of phylogenomic data can be applied to explore sources of incongruence and poor support in any phylogenomic data set. [Annelida; Brachiopoda; Bryozoa; Entoprocta; Mollusca; Nemertea; Phoronida; Platyzoa; Polyzoa; Spiralia; Trochozoa.].

Muscular anatomy of an entoproct creeping-type larva reveals extraordinary high complexity and potential shared characters with mollusks.

BACKGROUND: Entoprocta (Kamptozoa) is an enigmatic, acoelomate, tentacle-bearing phylum with indirect development, either via a swimming- or a creeping-type larva and still debated phylogenetic position within Lophotrochozoa. Recent morphological and neuro-anatomical studies on the creeping-type larva support a close relationship of Entoprocta and Mollusca, with a number of shared apomorphies including a tetraneurous nervous system and a complex serotonin-expressing apical organ. However, many morphological traits of entoproct larvae, in particular of the putative basal creeping-type larva, remain elusive. RESULTS: Applying fluorescent markers and 3D modeling, we found that this larval type has the most complex musculature hitherto described for any lophotrochozoan larva. The muscle systems identified include numerous novel and most likely creeping-type larva-specific structures such as frontal organ retractors, several other muscle fibers originating from the frontal organ, and longitudinal prototroch muscles. Interestingly, we found distinct muscle sets that are also present in several mollusks. These include paired sets of dorso-ventral muscles that intercross ventrally above the foot sole and a paired enrolling muscle that is distinct from the musculature of the body wall. CONCLUSION: Our data add further morphological support for an entoproct-mollusk relationship (Tetraneuralia) and strongly argue for the presence of an enrolling musculature as well as seriality (but not segmentation) in the last common tetraneuralian ancestor. The evolutionary driving forces that have led to the emergence of the extraordinarily complex muscular architecture in this short-lived, non-feeding entoproct larval type remain unknown, as are the processes that give rise to the highly different and much simpler muscular bodyplan of the adult entoproct during metamorphosis.

Spiral cleavage and early embryology of a loxosomatid entoproct and the usefulness of spiralian apical cross patterns for phylogenetic inferences.

BACKGROUND: Among the four major bilaterian clades, Deuterostomia, Acoelomorpha, Ecdysozoa, and Lophotrochozoa, the latter shows an astonishing diversity of bodyplans. While the largest lophotrochozoan assemblage, the Spiralia, which at least comprises Annelida, Mollusca, Entoprocta, Platyhelminthes, and Nemertea, show a spiral cleavage pattern, Ectoprocta, Brachiopoda and Phoronida (the Lophophorata) cleave radially. Despite a vast amount of recent molecular phylogenetic analyses, the interrelationships of lophotrochozoan phyla remain largely unresolved. Thereby, Entoprocta play a key role, because they have frequently been assigned to the Ectoprocta, despite their differently cleaving embryos. However, developmental data on entoprocts employing modern methods are virtually non-existent and the data available rely exclusively on sketch drawings, thus calling for thorough re-investigation. RESULTS: By applying fluorescence staining in combination with confocal microscopy and 3D-imaging techniques, we analyzed early embryonic development of a basal loxosomatid entoproct. We found that cleavage is asynchronous, equal, and spiral. An apical rosette, typical for most spiralian embryos, is formed. We also identified two cross-like cellular arrangements that bear similarities to both, a "molluscan-like" as well as an "annelid-like" cross, respectively. CONCLUSIONS: A broad comparison of cleavage types and apical cross patterns across Lophotrochozoa shows high plasticity of these character sets and we therefore argue that these developmental traits should be treated and interpreted carefully when used for phylogenetic inferences.