From feces to data: A metabarcoding method for analyzing consumed and available prey in a bird‐insect food web

Diets play a key role in understanding trophic interactions. Knowing the actual structure of food webs contributes greatly to our understanding of biodiversity and ecosystem functioning. The research of prey preferences of different predators requires knowledge not only of the prey consumed, but also of what is available. In this study, we applied DNA metabarcoding to analyze the diet of 4 bird species (willow tits Poecile montanus, Siberian tits Poecile cinctus, great tits Parus major and blue tits Cyanistes caeruleus) by using the feces of nestlings. The availability of their assumed prey (Lepidoptera) was determined from feces of larvae (frass) collected from the main foraging habitat, birch (Betula spp.) canopy. We identified 53 prey species from the nestling feces, of which 11 (21%) were also detected from the frass samples (eight lepidopterans). Approximately 80% of identified prey species in the nestling feces represented lepidopterans, which is in line with the earlier studies on the parids' diet. A subsequent laboratory experiment showed a threshold for fecal sample size and the barcoding success, suggesting that the smallest frass samples do not contain enough larval DNA to be detected by high‐throughput sequencing. To summarize, we apply metabarcoding for the first time in a combined approach to identify available prey (through frass) and consumed prey (via nestling feces), expanding the scope and precision for future dietary studies on insectivorous birds.     

Subfamily III of mammalian oxysterol-binding protein (OSBP) homologues: the expression and intracellular localization of ORP3, ORP6, and ORP7

The human OSBP related protein (ORP) family consists of 12 members, which can be divided into six subfamilies based on the genomic organization and amino acid homology. Here we performed basic characterization of subfamily III, which consists of three members: ORP3, ORP6, and ORP7. According to cDNA hybridization, the three genes are expressed in a tissue-specific manner. While ORP3 mRNA is most abundant in kidney, lymph nodes, and thymus, ORP6 shows highest expression in brain and skeletal muscle, and ORP7 in the gastrointestinal tract. Using monospecific peptide antibodies, we confirmed the presence of the three proteins in human and mouse tissues. ORP6 gene expression was induced upon differentiation of F9 embryonic carcinoma cells into parietal endoderm, while ORP3 and ORP7 mRNA levels were unchanged. In the F9 cells, endogenous ORP6 associated predominantly with the nuclear envelope. When expressed from the cDNA in cultured cells, the three proteins were distributed between the cytosol and endoplasmic reticulum (ER) membranes, with a minor portion found at the plasma membrane. Experiments with truncated constructs showed that the N-terminal portion of the proteins, containing a pleckstrin homology (PH) domain, has markedly strong plasma membrane targeting specificity, while the C-terminal half remains largely cytosolic. The expression data demonstrates that ORP3, -6, and -7 are not merely redundant gene products but show marked quantitative differences in tissue expression, suggesting tissue-specific aspects in their function. The dual targeting of the proteins indicates a putative role in communication between the ER and the plasma membrane.This study was supported by the Clinical Research Fund of Helsinki University Central Hospital (J.T.), the Academy of Finland (grant 51883 to M.L.; grants 49987, 50641, and 54301 to V.M.O.), the Sigrid Juselius Foundation, and the Finnish Cultural Foundation     

Keratinocyte growth factor stimulates migration and hyaluronan synthesis in the epidermis by activation of keratinocyte hyaluronan synthases 2 and 3

Keratinocyte growth factor (KGF) activates keratinocyte migration and stimulates wound healing. Hyaluronan, an extracellular matrix glycosaminoglycan that accumulates in wounded epidermis, is known to promote cell migration, suggesting that increased synthesis of hyaluronan might be associated with the KGF response in keratinocytes. Treatment of monolayer cultures of rat epidermal keratinocytes led to an elongated and lifted cell shape, increased filopodial protrusions, enhanced cell migration, accumulation of intermediate size hyaluronan in the culture medium and within keratinocytes, and a rapid increase of hyaluronan synthase 2 (Has2) mRNA, suggesting a direct influence on this gene. In stratified, organotypic cultures of the same cell line, both Has2 and Has3 with the hyaluronan receptor CD44 were up-regulated and hyaluronan accumulated in the epidermis, the spinous cell layer in particular. At the same time the expression of the early differentiation marker keratin 10 was inhibited, whereas filaggrin expression and epidermal permeability were less affected. The data indicate that Has2 and Has3 belong to the targets of KGF in keratinocytes, and support the idea that enhanced hyaluronan synthesis acts an effector for the migratory response of keratinocytes in wound healing, whereas it may delay keratinocyte terminal differentiation.     

Recovery of plant communities after ecological restoration of forestry‐drained peatlands

Ecological restoration is expected to reverse the loss of biodiversity and ecosystem services. Due to the low number of well‐replicated field studies, the extent to which restoration recovers plant communities, and the factors underlying possible shortcomings, are not well understood even in medium term. We compared the plant community composition of 38 sites comprising pristine, forestry‐drained, and 5 or 10 years ago restored peatlands in southern Finland, with special interest in understanding spatial variation within studied sites, as well as the development of the numbers and the abundances of target species. Our results indicated a recovery of community composition 5–10 years after restoration, but there was significant heterogeneity in recovery. Plant communities farthest away from ditches were very similar to their pristine reference already 10 years after restoration. In contrast, communities in the ditches were as far from the target as the drained communities. The recovery appears to be characterized by a decline in the number and abundance of species typical to degraded conditions, and increase in the abundance of characteristic peatland species. However, we found no increase above the drained state in the number of characteristic peatland species. Our results suggest that there is a risk of drawing premature conclusions on the efficiency of ecological restoration with the current practice of short‐term monitoring. Our results also illustrate fine‐scale within‐site spatial variability in the degradation and recovery of the plant communities that should be considered when evaluating the success of restoration. Overall, we find the heterogeneous outcome of restoration observed here promising. However, low recovery in the number of characteristic species demonstrates the importance of prioritizing restoration sites, and addressing the uncertainty of recovery when setting restoration targets. It appears that it is easier to eradicate unwanted species than regain characteristic species by restoration.     

Genomic structure of the human ezrin gene

The ERM proteins, ezrin, radixin, and moesin, act as linkers between the plasma membrane and actin cytoskeleton. They are involved in a variety of cellular functions, such as cell adhesion, migration, and the organization of cell surface structures, and are highly homologous, both in protein sequence and in functional activity, with merlin/schwannomin, a neurofibromatosis-2-associated tumor-suppressor protein. We report here the genomic structure and intron junction sequences of the human ezrin gene. Ezrin consists of 13 exons and spans approximately 24 kb genomic DNA. The coding parts of the exons range in size from 12 bp to 275 bp and the introns from 182 bp to 7 kb. The genomic structures of ezrin and moesin are highly conserved, suggesting their recent divergence. Radiation hybrid mapping has refined the location of ezrin to the interval between D6S442 and D6S281. Received: 1 June 1998 / Accepted: 25 August 1998     

Different metabolic responses of human brown adipose tissue to activation by cold and insulin

We investigated the metabolism of human brown adipose tissue (BAT) in healthy subjects by determining its cold-induced and insulin-stimulated glucose uptake and blood flow (perfusion) using positron emission tomography (PET) combined with computed tomography (CT). Second, we assessed gene expression in human BAT and white adipose tissue (WAT). Glucose uptake was induced 12-fold in BAT by cold, accompanied by doubling of perfusion. We found a positive association between whole-body energy expenditure and BAT perfusion. Insulin enhanced glucose uptake 5-fold in BAT independently of its perfusion, while the effect on WAT was weaker. The gene expression level of insulin-sensitive glucose transporter GLUT4 was also higher in BAT as compared to WAT. In conclusion, BAT appears to be differently activated by insulin and cold; in response to insulin, BAT displays high glucose uptake without increased perfusion, but when activated by cold, it dissipates energy in a perfusion-dependent manner.     

Use of Wishart Prior and Simple Extensions for Sparse Precision Matrix Estimation

A conjugate Wishart prior is used to present a simple and rapid procedure for computing the analytic posterior (mode and uncertainty) of the precision matrix elements of a Gaussian distribution. An interpretation of covariance estimates in terms of eigenvalues is presented, along with a simple decision-rule step to improve the performance of the estimation of sparse precision matrices and associated graphs. In this, elements of the estimated precision matrix that are zero or near zero can be detected and shrunk to zero. Simulated data sets are used to compare posterior estimation with decision-rule with two other Wishart-based approaches and with graphical lasso. Furthermore, an empirical Bayes procedure is used to select prior hyperparameters in high dimensional cases with extension to sparsity.     

Influence of growth temperature on the production of antibody Fab fragments in different microbes: a host comparative analysis

Microorganisms encounter diverse stress conditions in their native habitats but also during fermentation processes, which have an impact on industrial process performance. These environmental stresses and the physiological reactions they trigger, including changes in the protein folding/secretion machinery, are highly interrelated. Thus, the investigation of environmental factors, which influence protein expression and secretion is still of great importance. Among all the possible stresses, temperature appears particularly important for bioreactor cultivation of recombinant hosts, as reductions of growth temperature have been reported to increase recombinant protein production in various host organisms. Therefore, the impact of temperature on the secretion of proteins with therapeutic interest, exemplified by a model antibody Fab fragment, was analyzed in five different microbial protein production hosts growing under steady-state conditions in carbon-limited chemostat cultivations. Secretory expression of the heterodimeric antibody Fab fragment was successful in all five microbial host systems, namely Saccharomyces cerevisiae, Pichia pastoris, Trichoderma reesei, Escherichia coli and Pseudoalteromonas haloplanktis. In this comparative analysis we show that a reduction of cultivation temperature during growth at constant growth rate had a positive effect on Fab 3H6 production in three of four analyzed microorganisms, indicating common physiological responses, which favor recombinant protein production in prokaryotic as well as eukaryotic microbes.     

Cardiovascular, ventilatory and total activity responses of brown trout to handling stress

Changes in total activity, heart and ventilation rates were observed in 2-year-old brown trout, following handling stress, using non-contact bioelectronic monitoring equipment. Experiments were carried out in laboratory conditions at water temperatures below 4° C, Transfer between tanks as well as 5 min restraint stress increased the total activity of fish for 24 to 48 h, after which it declined to near the pre-stress level. The transfer and struggle both elevated the heart rate for 3 to 4 days. Ventilation rate was elevated to a maximum of about 30% above the nominal level and recovered within 3 to 4 days. Both heart and ventilation rates were higher in feeding fish relative to fasting fish after stress and rates remained higher throughout a 7 day period of recovery. A diel rhythm of lower rates during the night appeared in both heart and ventilation rates within 3 to 4 days after handling stress.