Estimation of Fish Biomass Using Environmental DNA

Environmental DNA (eDNA) from aquatic vertebrates has recently been used to estimate the presence of a species. We hypothesized that fish release DNA into the water at a rate commensurate with their biomass. Thus, the concentration of eDNA of a target species may be used to estimate the species biomass. We developed an eDNA method to estimate the biomass of common carp (Cyprinus carpio L.) using laboratory and field experiments. In the aquarium, the concentration of eDNA changed initially, but reached an equilibrium after 6 days. Temperature had no effect on eDNA concentrations in aquaria. The concentration of eDNA was positively correlated with carp biomass in both aquaria and experimental ponds. We used this method to estimate the biomass and distribution of carp in a natural freshwater lagoon. We demonstrated that the distribution of carp eDNA concentration was explained by water temperature. Our results suggest that biomass data estimated from eDNA concentration reflects the potential distribution of common carp in the natural environment. Measuring eDNA concentration offers a non-invasive, simple, and rapid method for estimating biomass. This method could inform management plans for the conservation of ecosystems

Cloning and characterization of a 9-lipoxygenase gene induced by pathogen attack from Nicotiana benthamiana for biotechnological application

<p>Abstract</p> <p>Background</p> <p>Plant lipoxygenases (LOXs) have been proposed to form biologically active compounds both during normal developmental stages such as germination or growth as well as during responses to environmental stress such as wounding or pathogen attack. In our previous study, we found that enzyme activity of endogenous 9-LOX in <it>Nicotiana benthamiana </it>was highly induced by agroinfiltration using a tobacco mosaic virus (TMV) based vector system.</p> <p>Results</p> <p>A <it>LOX </it>gene which is expressed after treatment of the viral vectors was isolated from <it>Nicotiana benthamiana</it>. As the encoded LOX has a high amino acid identity to other 9-LOX proteins, the gene was named as <it>Nb-9-LOX</it>. It was heterologously expressed in yeast cells and its enzymatic activity was characterized. The yeast cells expressed large quantities of stable 9-LOX (0.9 U ml^-1cell cultures) which can oxygenate linoleic acid resulting in high yields (18 μmol ml^-1cell cultures) of hydroperoxy fatty acid. The product specificity of Nb-9-LOX was examined by incubation of linoleic acid and Nb-9-LOX in combination with a 13-hydroperoxide lyase from watermelon (Cl-13-HPL) or a 9/13-hydroperoxide lyase from melon (Cm-9/13-HPL) and by LC-MS analysis. The result showed that Nb-9-LOX possesses both 9- and 13-LOX specificity, with high predominance for the 9-LOX function. The combination of recombinant Nb-9-LOX and recombinant Cm-9/13-HPL produced large amounts of C₉-aldehydes (3.3 μmol mg^-1crude protein). The yield of C₉-aldehydes from linoleic acid was 64%.</p> <p>Conclusion</p> <p>The yeast expressed Nb-9-LOX can be used to produce C₉-aldehydes on a large scale in combination with a <it>HPL </it>gene with 9-HPL function, or to effectively produce 9-hydroxy-10(<it>E</it>),12(<it>Z</it>)-octadecadienoic acid in a biocatalytic process in combination with cysteine as a mild reducing agent.</p

Useful pharmacodynamic endpoints in children: selection, measurement, and next steps.

Pharmacodynamic (PD) endpoints are essential for establishing the benefit-to-risk ratio for therapeutic interventions in children and neonates. This article discusses the selection of an appropriate measure of response, the PD endpoint, which is a critical methodological step in designing pediatric efficacy and safety studies. We provide an overview of existing guidance on the choice of PD endpoints in pediatric clinical research. We identified several considerations relevant to the selection and measurement of PD endpoints in pediatric clinical trials, including the use of biomarkers, modeling, compliance, scoring systems, and validated measurement tools. To be useful, PD endpoints in children need to be clinically relevant, responsive to both treatment and/or disease progression, reproducible, and reliable. In most pediatric disease areas, this requires significant validation efforts. We propose a minimal set of criteria for useful PD endpoint selection and measurement. We conclude that, given the current heterogeneity of pediatric PD endpoint definitions and measurements, both across and within defined disease areas, there is an acute need for internationally agreed, validated, and condition-specific pediatric PD endpoints that consider the needs of all stakeholders, including healthcare providers, policy makers, patients, and families.Pediatric Research advance online publication, 11 April 2018; doi:10.1038/pr.2018.38

Overexpression of Akt1 Enhances Adipogenesis and Leads to Lipoma Formation in Zebrafish

<div><h3>Background</h3><p>Obesity is a complex, multifactorial disorder influenced by the interaction of genetic, epigenetic, and environmental factors. Obesity increases the risk of contracting many chronic diseases or metabolic syndrome. Researchers have established several mammalian models of obesity to study its underlying mechanism. However, a lower vertebrate model for conveniently performing drug screening against obesity remains elusive. The specific aim of this study was to create a zebrafish obesity model by over expressing the insulin signaling hub of the <em>Akt1</em> gene.</p> <h3>Methodology/Principal Findings</h3><p><em>Skin oncogenic transformation screening shows that a stable zebrafish transgenic of Tg(krt4Hsa.myrAkt1</em>)^cy18 displays severely obese phenotypes at the adult stage. In Tg(<em>krt4:Hsa.myrAkt1</em>)^cy18, the expression of exogenous human constitutively active Akt1 (myrAkt1) can activate endogenous downstream targets of mTOR, GSK-3α/β, and 70S6K. During the embryonic to larval transitory phase, the specific over expression of myrAkt1 in skin can promote hypertrophic and hyperplastic growth. From 21 hour post-fertilization (hpf) onwards, myrAkt1 transgene was ectopically expressed in several mesenchymal derived tissues. This may be the result of the integration position effect. Tg(<em>krt4:Hsa.myrAkt1</em>)^cy18 caused a rapid increase of body weight, hyperplastic growth of adipocytes, abnormal accumulation of fat tissues, and blood glucose intolerance at the adult stage. Real-time RT-PCR analysis showed the majority of key genes on regulating adipogenesis, adipocytokine, and inflammation are highly upregulated in Tg(<em>krt4:Hsa.myrAkt1</em>)^cy18. In contrast, the myogenesis- and skeletogenesis-related gene transcripts are significantly downregulated in Tg(<em>krt4:Hsa.myrAkt1</em>)^cy18, suggesting that excess adipocyte differentiation occurs at the expense of other mesenchymal derived tissues.</p> <h3>Conclusion/Significance</h3><p>Collectively, the findings of this study provide direct evidence that Akt1 signaling plays an important role in balancing normal levels of fat tissue in vivo. The obese zebrafish examined in this study could be a new powerful model to screen novel drugs for the treatment of human obesity.</p> </div