Evaluation of an ultrafiltration membrane for the removal of fish viruses and bacteria in aquaculture water

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Smoltification, seawater performance, and maturation in Atlantic salmon (Salmo salar) fed different fat levels

IntroductionThe use of recirculating aquaculture systems (RAS) to produce Atlantic salmon smolts has resulted in exceptionally high fish growth rates. However, there are potential negative trade-offs between fast growth and key physiological processes, such as inadequate smoltification and early sexual maturation, which can both be linked to body energy reserves.MethodsThis study determined the effect of i) dietary fat levels on Atlantic salmon whole-body fat and fatty acids composition, growth performance and smoltification and ii) a previous dietary regime on seawater growth performance and male early sexual maturation. In freshwater RAS, salmon parr (~19g) were fed 3 fat levels (20, 24, and 28%) over a 14-week period at 12°C. Subsequently, in seawater flow-through systems, smolt (~96g) were fed a control diet (26%) for 12 weeks at 2 temperatures (12 and 16°C).ResultsDietary fat levels resulted in differences in k-factor, fish whole-body fat and fatty acids composition; the low fat diet resulted in fish with a 23% lower whole-body fat when compared to the other two dietary groups at the end of the freshwater phase. These differences in whole-body fat faded at the end of the seawater phase. During the freshwater phase, all three feeds resulted in a comparable growth performance and smoltification indicator values, including k-factor, gill Na+, K+-ATPase, blood serum chloride and smolt index score. In contrast, water temperature resulted in fish body weight differences, where groups reared at 16°C were larger than those reared at 12°C. Nevertheless, both temperature regimes supported a similar thermal growth coefficient. Smolts grown at 16°C showed a higher level of the sex steroids androstenedione compared to those at 12°C. However, 11-ketotestosterone and testosterone levels did not differ despite a trend for higher levels at 16°C. Furthermore, testis histology at the final sampling indicated that some individuals showed initial signs of maturation (stage 3).DiscussionIn conclusion, varying dietary fat levels (20 - 28%) during the freshwater phase did not influence smoltification or male early sexual maturation during the subsequent grow-out phase. However, a temperature increase from 12 to 16°C resulted in larger fish and appeared to stimulate early male maturation in some fish individuals

Determination of the Oxidative Stress Biomarkers of 8-Hydroxydeoxyguanosine and Dityrosine in the Gills, Skin, Dorsal Fin, and Liver Tissue of Atlantic Salmon (Salmo salar) Parr

Oxidative stress is a condition caused by an imbalance in the occurrence of reactive oxygen species in the cells and tissues of organisms. An ultra-performance liquid chromatography–electrospray ionization tandem mass spectrometry (UPLC–ESI–MS/MS) method was developed for the simultaneous determination of two oxidative stress biomarkers, 8-hydroxydeoxyguanosine (8OHDG) and dityrosine (DIY), in the gills, skin, dorsal fin, and liver tissue of Atlantic salmon (Salmo salar) parr. The use of target analyte-specific 13C and 15N internal standards allowed quantification of each target analyte to be performed through the standard solvent calibration curve. The relative recoveries [mean ± (relative standard deviation%)] of 8OHDG and DIY were 101 ± 11 and 104 ± 13% at a fortified concentration of 10 ng/mL (8OHDG) and 500 ng/mL (DIY), respectively, ensuring the accuracy of the extraction and quantification. The chromatographic separation was carried out using a gradient elution program with a total run time of 5 min. The limits of detection (LODs) were 0.11 and 1.37 ng/g wet weight (w.w.) for 8OHDG and DIY, respectively. To demonstrate the applicability of the developed method, it was applied in 907 tissue samples that were collected from Atlantic salmon parr individuals reared in an experimental land-based recirculating aquaculture system (RAS) treated with peracetic acid. Moreover, the possibility of using the dorsal fin as an alternative matrix for the minimally invasive assessment of oxidative stress in Atlantic salmon parr was introduced. To our knowledge, 8OHDG and DIY were used for the first time as biomarkers for biomonitoring the fish health (oxidative stress) of Atlantic salmon parr in RAS.Determination of the Oxidative Stress Biomarkers of 8-Hydroxydeoxyguanosine and Dityrosine in the Gills, Skin, Dorsal Fin, and Liver Tissue of Atlantic Salmon (Salmo salar) ParrpublishedVersio

Applications of hyperspectral imaging for documenting smoltification status and welfare in Atlantic salmon

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Evaluation of an ultrafiltration membrane for the removal of fish viruses and bacteria in aquaculture water

Ultrafiltration (UF) membranes are used to successfully remove waterborne virus and bacteria from wastewater and drinking water. However, UF membrane application in aquaculture water treatment is limited. In this study we evaluate the performance of a capillary polyethersulfone UF membrane to remove two benchmark waterborne fish pathogens: i) the infectious pancreatic necrosis virus - IPNV, which is an unenveloped icosahedral virus, and ii) the bacterium Aeromonas salmonicida, which is a Gram-negative, facultative anaerobic bacilli. Moreover, the UF membrane bench-scale unit was tested at two temperatures according to salmonid aquaculture: low (4 - 7°C) and high (16 - 19°C). Sterilised natural seawater was spiked with laboratory cultured pathogens, the water was filtrated, and the membrane permeate collected. Both pathogen solution and permeate were evaluated using a cell culture method to estimate the colony-forming units (CFU/ml) for bacteria presence, a median tissue culture infectious dose (TCID50/ml) assay for virus presence, and real-time quantitative polymerase chain reaction (RT-qPCR) for both bacteria and virus presence. The membrane permeate was negative for both virus and bacteria for all analysis and for both low and high temperatures. The results from this bench-scale study are encouraging for the application of UF membrane technology in aquaculture water treatment to prevent virus and bacteria outbreaks. Further studies should validate this UF membrane technology results in commercial aquaculture conditions

Polyurea Dendrimer Folate-Targeted Nanodelivery of l-Buthionine sulfoximine as a Tool to Tackle Ovarian Cancer Chemoresistance

The research was funded by iNOVA4Health—UID/Multi/04462/, a program financially supported by the Fundação para a Ciência e a Tecnologia—Ministério da Educação e Ciência (FCT-MCTES), through nationalfunds and co-funded by FEDER under the PT2020 Partnership Agreement. We also acknowledge funding from FCT-MCTES through the project DREAM—PTDC/MEC-ONC/29327/2017 and FAI2017 from IPOLFG internal funding.: Ovarian cancer is a highly lethal disease, mainly due to chemoresistance. Our previous studies on metabolic remodeling in ovarian cancer have supported that the reliance on glutathione (GSH) bioavailability is a main adaptive metabolic mechanism, also accounting for chemoresistance to conventional therapy based on platinum salts. In this study, we tested the effects of the in vitro inhibition of GSH synthesis on the restoration of ovarian cancer cells sensitivity to carboplatin. GSH synthesis was inhibited by exposing cells to l-buthionine sulfoximine (l-BSO), an inhibitor of -glutamylcysteine ligase (GCL). Given the systemic toxicity of l-BSO, we developed a new formulation using polyurea (PURE) dendrimers nanoparticles (l-BSO@PUREG4-FA2), targeting l-BSO delivery in a folate functionalized nanoparticle.publishersversionpublishe

Gradually decreasing daylength after smoltification induced by “winter signal“ reduced sexual maturation in male Atlantic salmon

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Targeting Glutathione and Cystathionine β-Synthase in Ovarian Cancer Treatment by Selenium-Chrysin Polyurea Dendrimer Nanoformulation

The research was funded by iNOVA4Health UID/Multi/04462, a program financially supported by Fundação para a Ciência e Tecnologia/Ministério da Educação e Ciência (FCT-MCTES), through national funds, and co-funded by FEDER under the PT2020 Partnership Agreement. We also acknowledge funding from FCT-MCTES through project DREAM PTDC/MEC-ONC/29327/2017.Ovarian cancer is the main cause of death from gynecological cancer, with its poor prognosis mainly related to late diagnosis and chemoresistance (acquired or intrinsic) to conventional alkylating and reactive oxygen species (ROS)-generating drugs. We and others reported that the availability of cysteine and glutathione (GSH) impacts the mechanisms of resistance to carboplatin in ovarian cancer. Different players in cysteine metabolism can be crucial in chemoresistance, such as the cystine/glutamate antiporter system Xc (xCT) and the H2S-synthesizing enzyme cystathionine β-synthase (CBS) in the pathway of cysteine catabolism. We hypothesized that, by disrupting cysteine metabolic flux, chemoresistance would be reverted. Since the xCT transporter is also able to take up selenium, we used selenium-containing chrysin (SeChry) as a plausible competitive inhibitor of xCT. For that, we tested the effects of SeChry on three different ovarian cancer cell lines (ES2, OVCAR3, and OVCAR8) and in two non-malignant cell lines (HaCaT and HK2). Results showed that, in addition to being highly cytotoxic, SeChry does not affect the uptake of cysteine, although it increases GSH depletion, indicating that SeChry might induce oxidative stress. However, enzymatic assays revealed an inhibitory effect of SeChry toward CBS, thus preventing production of the antioxidant H2S. Notably, our data showed that SeChry and folate-targeted polyurea dendrimer generation four (SeChry@PUREG4-FA) nanoparticles increased the specificity for SeChry delivery to ovarian cancer cells, reducing significantly the toxicity against non-malignant cells. Collectively, our data support SeChry@PUREG4-FA nanoparticles as a targeted strategy to improve ovarian cancer treatment, where GSH depletion and CBS inhibition underlie SeChry cytotoxicity.publishersversionpublishe

In Vivo Approaches Reveal a Key Role for DCs in CD4+ T Cell Activation and Parasite Clearance during the Acute Phase of Experimental Blood-Stage Malaria

Dendritic cells (DCs) are phagocytes that are highly specialized for antigen presentation. Heterogeneous populations of macrophages and DCs form a phagocyte network inside the red pulp (RP) of the spleen, which is a major site for the control of blood-borne infections such as malaria. However, the dynamics of splenic DCs during Plasmodium infections are poorly understood, limiting our knowledge regarding their protective role in malaria. Here, we used in vivo experimental approaches that enabled us to deplete or visualize DCs in order to clarify these issues. To elucidate the roles of DCs and marginal zone macrophages in the protection against blood-stage malaria, we infected DTx (diphtheria toxin)-treated C57BL/6.CD11c-DTR mice, as well as C57BL/6 mice treated with low doses of clodronate liposomes (ClLip), with Plasmodium chabaudi AS (Pc) parasites. The first evidence suggesting that DCs could contribute directly to parasite clearance was an early effect of the DTx treatment, but not of the ClLip treatment, in parasitemia control. DCs were also required for CD4+ T cell responses during infection. The phagocytosis of infected red blood cells (iRBCs) by splenic DCs was analyzed by confocal intravital microscopy, as well as by flow cytometry and immunofluorescence, at three distinct phases of Pc malaria: at the first encounter, at pre-crisis concomitant with parasitemia growth and at crisis when the parasitemia decline coincides with spleen closure. In vivo and ex vivo imaging of the spleen revealed that DCs actively phagocytize iRBCs and interact with CD4+ T cells both in T cell-rich areas and in the RP. Subcapsular RP DCs were highly efficient in the recognition and capture of iRBCs during pre-crisis, while complete DC maturation was only achieved during crisis. These findings indicate that, beyond their classical role in antigen presentation, DCs also contribute to the direct elimination of iRBCs during acute Plasmodium infection.São Paulo Research Foundation grants: (2011/24038-1 [MRDL], 2009/08559-1 [HBdS], CAPES/IGC 04/ 2012 [MRDL, CET])