Accelerated Solvent Extraction and Pulsed Electric Fields for Valorization of Rainbow Trout (Oncorhynchus mykiss) and Sole (Dover sole) By-Products: Protein Content,Molecular Weight Distribution and Antioxidant Potential of the Extracts

Fishery by-products are rich in biologically active substances and the use of green and efficient extraction methods to recover these high-added-value compounds is of particular importance. In this study, head, skin and viscera of rainbow trout and sole were used as the target matrices and accelerated solvent extraction (ASE) (45–55 °C, 15 min, pH 5.2–6.8, 103.4 bars) and pulsed electric fields (PEF) (1–3 kV/cm, 123–300 kJ/kg, 15–24 h) were applied as extraction technologies. The results showed that ASE and PEF significantly increased the protein extract efficiency of the fish by-products (p < 0.05) by up to 80%. SDS-PAGE results showed that ASE and PEF treatments changed the molecular size distribution of the protein in the extracts, which was specifically expressed as the change in the area or number of bands between 5 and 250 kDa. The antioxidant capacity of the extracts was evaluated by oxygen radical absorbance capacity (ORAC) and total antioxidant capacity (ABTS) assays. The results showed that both ASE and PEF treatments significantly increased the antioxidant capacity of rainbow trout and sole skin and head extracts (p < 0.05). ASE and PEF extraction processes can be used as new technologies to extract high-added-value compounds from fish by-products.     

Tissue distribution of Red Spotted Grouper Nervous Necrosis Virus (RGNNV) genome in experimentally infected juvenile European seabass (Dicentrarchus labrax)

The distribution of viral genome in the tissues of juvenile European seabass (Dicentrarchus labrax) during the course of a Red Spotted Grouper Nervous Necrosis Virus (RGNNV) infection has not yet been described. The present study addresses this and indicates which target organs may be involved in viral replication. This information should enable more accurate detection of virus in asymptomatic carriers, and in turn help to control the spread of the disease. The aim of this study was to examine the pattern of expression of viral genomic segments RNA1 and RNA2, using two absolute real-time PCRs (RT-qPCR), over the course of a RGNNV infection after administering the virus by intramuscular injection. In situ hybridization was also used to locate the RNA2 viral segment in different organs throughout the infection. The experimental challenge provoked an acute form of viral nervous necrosis (VNN), with a resulting cumulative mortality of 37%. The RT-qPCRs designed allowed the detection of both genomic segments in all the organs tested (nervous and non-nervous tissues) at all sampling times examined. The highest viral RNA copy number was found in eyes, although viral replication appeared to begin in the brain. Viral replication was also recorded in pooled internal organs and in caudal fin. However, the increase in the viral RNA copy number in these organs did not result in an increased viral titre, which may indicate that a productive infection does not take place in non-nervous tissues, possibly due to a failure in a viral post-replication step.     

Progress towards clonal propagation of Camellia japonica cv. Alba Plena by tissue culture techniques

Summary

Optimal rooting conditions have been determined for shoot cultures of Camellia japonica cv. Alba Plena derived from a 50 year old tree: dipping the base of shoots in 1 g l^?l solution for 15 min, followed by 12 days’ darkness, induced 87% rooting in shoots cultured with Woody Plant Medium (WPM) macronutrients. Halving the auxin concentration or exposure time considerably reduced this rate, and root formation was severely inhibited if an initial dark period for the entire shoot was not used. The type of support (agar or paper bridges) did not significantly affect the rooting percentage or number of roots per rooted shoot, but liquid media induced greater root elongation. No significant differences were observed between the use of WPM and a modified Heller’s medium as regards the rooting percentages achieved, but the number of roots formed was considerably greater with WPM, as was the survival rate after transfer to soil (70% for transfer as against 35% with the modified Heller’s medium). Best survival rates were achieved when shoots were transferred to the acclimatization soil 12 days after auxin treatment, i.e. immediately after the dark period.     

Multi‐temporal Digital Photogrammetric Analysis for Quantitative Assessment of Soil Erosion Rates in the Landola Catchment of the Upper Orcia Valley (Tuscany,Italy)

This paper focuses on the analysis of the slope and catchment erosion dynamics in a typical Mediterranean context and its sensitivity to recent climatic and socio‐economic changes. The main objective of the present study is to test the reliability of digital photogrammetric analysis to evaluate the time and space evolution of erosion processes mainly triggered by surface running waters and landslides for about the last 60 years. The selected test area is the Landola catchment, a minor tributary of the Upper Orcia River Valley (Southern Tuscany). The Upper Orcia valley is a key site for the comprehension of denudation processes typically acting in Mediterranean badland areas thanks to the following: (i) the availability of long‐lasting erosion monitoring datasets (20 years' long direct measurements at erosion ‘hot spots’); (ii) its representativeness as a sub‐humid Mediterranean badland area; and (iii) the rapidity of development of erosion processes, which makes it suitable as an open‐air laboratory for the study of badland dynamics. The outputs of this work highlight that the application of high‐resolution digital photogrammetric methodologies can represent a powerful and low‐cost tool to evaluate rates and spatial–temporal distribution of denudation processes, as confirmed by the validation through field point monitoring in areas close to the study area. The results obtained for the study area confirm that high erosion rates are a persistent environmental problem for the Upper Orcia Valley, which is not solved up to now, despite various land conservation interventions. The performed multi‐temporal analysis shows a slight decrease in the average water erosion rate during the last 60 years and a parallel increase in the frequency of mass wasting events. These variations are most likely related to a complex interplay between land use changes that have affected the study area during the studied period, revealing a very delicate equilibrium between farming activities and erosion processes. Copyright © 2014 John Wiley & Sons, Ltd.     

Mediation of poly(ADP-ribose) polymerase-1-dependent cell death by apoptosis-inducing factor

Poly(ADP-ribose) polymerase-1 (PARP-1) protects the genome by functioning in the DNA damage surveillance network. PARP-1 is also a mediator of cell death after ischemia-reperfusion injury, glutamate excitotoxicity, and various inflammatory processes. We show that PARP-1 activation is required for translocation of apoptosis-inducing factor (AIF) from the mitochondria to the nucleus and that AIF is necessary for PARP-1-dependent cell death. N-methyl-N'-nitro-N-nitrosoguanidine, H2O2, and N-methyl-d-aspartate induce AIF translocation and cell death, which is prevented by PARP inhibitors or genetic knockout of PARP-1, but is caspase independent. Microinjection of an antibody to AIF protects against PARP-1-dependent cytotoxicity. These data support a model in which PARP-1 activation signals AIF release from mitochondria, resulting in a caspase-independent pathway of programmed cell death.     

Distribution of red-spotted grouper nervous necrosis virus (RGNNV) antigens in nervous and non-nervous organs of European seabass (Dicentrarchus labrax) during the course of an experimental challenge

The distribution of red-spotted grouper nervous necrosis virus (RGNNV) antigens was examined by immunohistochemistry in the nervous and non-nervous organs of juvenile European seabass (Dicentrarchus labrax) during the course of an intramuscular infection. Histological changes resulting from the infection were evaluated from 3 days to 2 months post-infection. The specific antibody response was also studied 2 months post-challenge. Viral proteins were present throughout the experimental period in the retina (inner nuclear layer, ganglion layer, outer limiting membrane, and outer plexiform layer), brain (cerebellum and tectum opticum), and liver (hepatocytes and endothelial cells). These proteins were also observed in the renal tubular cells, white pulp of spleen, and in fibroblasts and cartilage of caudal fin. This is the first report of RGNNV proteins appearing in these organs, where the immunostaining was only detected at certain sampling times after the onset of mortality. Brain and retina of virus-exposed fish showed high levels of vacuolation, while accumulation of fat vacuoles was observed in the liver. RGNNV infection also induced a specific antibody response as measured by an ELISA. In summary, this is the first study demonstrating the presence of viral proteins in cells of caudal fin, kidney and spleen of European seabass.     

Phytotoxic polyketides produced by <Emphasis Type="Italic">Phomopsis foeniculi</Emphasis>, a strain isolated from diseased Bulgarian fennel

Recently, a new fungal disease caused by Diaporthe angelicae (anamorph Phomopsis foeniculi) has been found with increasingly frequency on fennel (Foeniculum vulgare) in Bulgaria. Using a bioassay-guided isolation and purification procedure, different metabolites were isolated from the fungal culture filtrates. They were identified by spectroscopic methods as nectriapyrone, a pentaketide monoterpenoid, and altersolanols A and J and macrosporin, three octaketides anthracenones. Leaf puncture bioassay was applied on detached tomato leaves to prove the phytotoxic activity of the fractions and of pure compounds. Nectriapyrone and altersolanols A and J showed a modulated phytotoxicity, while macrosporin was not toxic. Altersolanol A was the most active compound.     

Use of the scrotum as a transposition flap for closure of surgical wounds in three dogs

OBJECTIVE: To describe use of the scrotum as a pedicle flap to cover defects created by tumor excision in the perineum or caudal and medial aspect of the thigh. STUDY DESIGN: Clinical study. ANIMALS: Three dogs. METHODS: After tumor excision and prescrotal castration, an incision was made around the base of the scrotum, leaving a pedicle on the side opposite the skin defect. After removal of the tunica dartos and abdominal fascia, the scrotal skin was stretched and shaped to the defect, then sutured in position with a simple interrupted pattern. Bandages that were changed every 48 hours compressed the flaps, and healing was observed until sutures were removed. RESULTS: Two dogs had first intention healing in 15 days, whereas 1 dog developed necrosis of 10% of the flap, and this area healed by second intention. CONCLUSION: In male dogs, the scrotal skin can be used as a pedicle flap for reconstructive surgery of wounds in the perineum and the proximomedial and caudal aspect of the thigh. CLINICAL RELEVANCE: The perineal region remains a surgical challenge because of the lack of the available skin for reconstruction of surgical wounds. The scrotal skin should be considered for use as a transposition flap to cover skin defects in this region.     

Induction of an antioxidant enzyme system and other oxidative stress markers associated with compatible and incompatible interactions between chickpea (Cicer arietinum L.) and Fusarium oxysporum f. sp.ciceris

To ascertain if active oxygen species play a role in fusarium wilt of chickpea caused by Fusarium oxysporum f. sp. ciceris, the degree of lipid peroxidation (malondialdehyde formation) and the activity levels of diamine oxidase (DAO), an apoplastic H₂O₂-forming oxidase, and several antioxidant enzymes, namely ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR), guaiacol-dependent peroxidase (GPX) and superoxide dismutase (SOD), were determined spectrophotometrically in roots and stems of ‘WR315’ (resistant) and ‘JG62’ (susceptible) chickpea cultivars inoculated with the highly virulent race 5 of the pathogen. Moreover, APX, CAT, GPX and SOD were also analysed in roots and stems by gel electrophoresis and activity staining; and the protein levels of APX and SOD in roots were determined by Western blotting. In roots, infection by the pathogen increased lipid peroxidation and CAT and SOD activities, although such responses occurred earlier in the incompatible compared with the compatible interactions. APX, GPX and GR activities were also increased in infected roots, but only in the compatible interaction. In stems, infection by the pathogen increased lipid peroxidation and APX, CAT, SOD and GPX activities only in the compatible interaction, and DAO activity only in the incompatible one. In general, electrophoregrams agreed with the activity levels determined spectrophotometrically and did not reveal any differences in isoenzyme patterns between cultivars or between infected and non-infected plants. Further, Western blots revealed an increase in the root protein levels of APX in the compatible interaction and in those of SOD in both compatible and incompatible interactions. In conclusion, whereas enhanced DAO activity in stems, and earlier increases in lipid peroxidation and CAT and SOD activities in roots, can be associated with resistance to fusarium wilt in chickpea, the induction of the latter three parameters in roots and stems along with that of APX, GR (only in roots) and GPX (only in stems) activities are rather more associated with the establishment of the compatible interaction.     

Feeding behavior of feedlot lambs fed diets containing levels of cassava wastewater

In this study, we evaluated the effects of including cassava wastewater in the diet on the feeding behavior of feedlot lambs in 35 male uncastrated Santa Inês × Dorper crossbred lambs at an approximate age of 3 months, with an average live weight of 20.0?±?3.4 kg. Diets were formulated with hay of cassava shoots (roughage) and a concentrate based on corn and soybean, with a roughage:concentrate ratio of 50:50, plus inclusion of cassava wastewater at the levels of 0, 12, 24, 36, or 48 g/kg of the total diet. Feeding behavior was evaluated between the 46th and 52nd days of the experiment. Increasing cassava wastewater levels in the diet reduced (P?<?0.05) the intakes (kg/day) of dry matter and neutral detergent fiber as well as the efficiency of rumination (g/cud and g/h) of dry matter and neutral detergent fiber. The other behavioral parameters were not affected by wastewater inclusion in the diet. Therefore, the inclusion of up to 48 g/kg of cassava wastewater on fresh matter of diets is not recommended for feedlot lambs.