Selenium alleviates cadmium-induced oxidative stress, endoplasmic reticulum stress, and apoptosis in L8824 cells.

Cadmium (Cd) is a toxic pollutant in industrial production that induces organ damage and apoptosis, While, selenium (Se) has the biological function of antagonizing Cd toxicity. Hence, to gain further insight into the protective mechanisms of selenium against Cd-induced damage in Ctenopharyngodon idella liver (L8824) cells, L8824 were exposed to 5 µM, 15 µM, 25 µM cadmium chloride for 24 h after pre-incubation with 25 µM sodium selenite for 9 h. Cell proliferation and morphological changes, the levels of reactive oxygen species (ROS) and antioxidant enzyme activity, mitochondrial membrane potential (MMP), endoplasmic reticulum stress (ERS)-related pathway genes expression, intracellular calcium levels and apoptosis were assessed to explore the protective effect of selenium in Cd-induced L8824 cell damage. The results showed that Cd caused decreased cell viability, ROS accumulation, reduced activity of antioxidant enzymes (SOD, CAT GPx and T-AOC) and apoptosis in L8824 cells. The incubation of Se prominently ameliorated cell proliferation, activated the Keap1-Nrf2 pathway, and restored antioxidant enzyme activity. Furthermore, the expression of grp78, perk, eif-2α, atf4, chop bax, jnk, caspase-3 and caspase-9 was significantly upregulated after Cd exposure, while the expression of bcl-2 was significantly downregulated. Se supplementation alleviated Cd-induced ERS and apoptosis. Moreover, Cd-induced elevation of intracellular Ca2+ levels were alleviated by dantrolene and 2-APB, suggesting that intracellular calcium disorders were caused by Ca2+ released by RyR and IP3R-mediated ER. The results of this study suggested that Cd could induce oxidative stress, ERS, mitochondrial damage and evoke apoptosis, whereas Se had protective effects in preventing Cd induced damage by inhibiting ERS, maintaining intracellular calcium homeostasis, enhancing the antioxidant capacity of L8824 cells and downregulating the Keap1/Nrf2 pathway.

A preliminary study of the cultivable microbiota on the plastic litter collected by commercial fishing trawlers in the south-eastern Adriatic Sea, with emphasis on Vibrio isolates and their antibiotic resistance.

Mediterranean Sea is the sixth largest area of marine litter accumulation in the world, and plastic pollution is a growing problem in its Adriatic sub-basin. The aim of the present study was to evaluate the cultivable microbiota associated with plastic litter collected by commercial fishing trawlers in the south-eastern Adriatic Sea in comparison with microbiota in seawater and sediment. Plastic litter in the sea contains an autochthonous microbiota that is different from that of the surrounding seawater and sediment. Vibrio abundance was higher on plastic litter than in surrounding seawater and sediment. All isolated Vibrio showing resistance to ampicillin and vancomycin, while resistance to other antibiotics depended on the isolated species. Overall, this study provides for the first time information on the cultivable microbiota associated with plastic litter collected by commercial fishing trawlers and provides a data base for further studies.

Limited population genetic variation but pronounced seascape genetic structuring in populations of the Mediterranean mussel (Mytilus galloprovincialis) from the eastern Adriatic Sea.

Population genetic analysis of variation at five neutral microsatellite loci for Mediterranean mussels (Mytilus galloprovincialis) from 18 sites along the eastern Adriatic Sea revealed little or no spatial variation. In contrast, seascape genetics analysis revealed a pronounced locus-specific gradient in allelic and genotypic frequencies across the study region. At a sixth locus, MGE7, the frequencies of two alleles, MGE7243 and MGE7249, were strongly associated, negatively and positively, respectively, with a single environmental variable - minimum salinity (minSAL). The frequency of the MGE7243/243 homozygous genotype was strongly negatively associated with minSAL, whereas the frequencies of the MGE7246/249 and the MGE7249/249 genotypes were strongly positively correlated with minSAL. Interpretation of these pronounced gradients is confounded by the fact that minSAL and another environmental variable, maximum sea surface temperature (maxSST), are highly correlated (R = -.911) and are therefore not necessarily acting independently. BLAST searches of the MGE7 locus against M. galloprovincialis whole genome shotgun sequence returned an alignment with contig mg10_S01094 (accession UYJE01010330.1) and 7 predicted M. galloprovincialis proteins VDI82194.1 - VDI82200.1. Conserved domain searches revealed a similar structure to the transcriptional regulator Msx2-interacting protein. The BLASTp search also returned significant alignments to Msx2-interacting proteins in Mytilus coruscus, Crassostrea virginica, and Haliotis rubra. The existence of the MGE7 gradient highlights the role that environmental variation may play in retarding gene flow among wild M. galloprovincialis populations, and also how the success of collection of young mussels (spat) from one site and their transfer to another site (the farm) may be influenced by a single factor such as minSAL or maxSST on a localized scale.

Characterization of Vibrio Populations from Cultured European Seabass and the Surrounding Marine Environment with Emphasis on V. anguillarum.

Vibrio species are widely distributed and can be potentially pathogenic to aquatic organisms. In this study, we isolated Vibrio spp. from environmental samples (seawater, sediment, and fish swabs) collected over a three-year period from a fish farm in Mali Ston Bay in the Adriatic Sea, Croatia, and assess their distribution. A total of 48 seawater samples and 12 sediment samples, as well as gill and skin swabs from 110 farmed European seabass, were analysed for the presence of Vibrio. Vibrio strains were identified to the species level by MALDI TOF MS. The analysis revealed that V. alginolyticus was the predominant species in European seabass, followed by V. anguillarum. V. alginolyticus was isolated from the sediments, along with V. gigantis and V. pomeroyi, while V. chagasii, V. cyclitrophicus, V. fortis, V. gigantis, V. harveyi, V. pelagius, and V. pomeroyi were isolated from seawater. V. anguillarum was isolated only twice during two different spring seasons, once from a diseased sea bass and the second time from a healthy sea bass. We analysed these two isolates and found that they differ both genetically and in terms of resistance to antibiotics. Our results confirm the seasonality of vibriosis incidence and the presence of the pathogenic V. anguillarum, which increases the risk of vibriosis.

Investigating the Ability of Growth Models to Predict In Situ Vibrio spp. Abundances.

Vibrio spp. have an important role in biogeochemical cycles; some species are disease agents for aquatic animals and/or humans. Predicting population dynamics of Vibrio spp. in natural environments is crucial to predicting how the future conditions will affect the dynamics of these bacteria. The majority of existing Vibrio spp. population growth models were developed in controlled environments, and their applicability to natural environments is unknown. We collected all available functional models from the literature, and distilled them into 28 variants using unified nomenclature. Next, we assessed their ability to predict Vibrio spp. abundance using two new and five already published longitudinal datasets on Vibrio abundance in four different habitat types. Results demonstrate that, while the models were able to predict Vibrio spp. abundance to an extent, the predictions were not reliable. Models often underperformed, especially in environments under significant anthropogenic influence such as aquaculture and urban coastal habitats. We discuss implications and limitations of our analysis, and suggest research priorities; in particular, we advocate for measuring and modeling organic matter.

High stocking density alters growth performance, blood biochemical profiles, and hepatic antioxidative capacity in gibel carp (Carassius gibelio).

This study investigated the effect of stocking density on growth performance, blood biochemical profiles, antioxidative capacity, and muscle quality of gibel carp (Carassius gibelio). Gibel carps (initial body weight 57.04 ± 1.89 g) were reared at high stocking density (HSD, 10.85 kg m-3), medium stocking density (MSD, 5.06 kg m-3), and low stocking density (LSD, 1.47 kg m-3) for 60 days. The LSD group exhibited the highest growth rate, while HSD significantly inhibited fish growth. The muscular compositions of crude fat, crude ash, and moisture were significantly changed by stocking density, but crude protein content did not differ significantly. The stocking density altered the muscular texture profiles of gibel carp. Compared to either the HSD group or the MSD group, the highest levels of resilience and springiness occurred in the LSD group. Significant differences were observed in the levels of plasma glucose, alanine aminotransferase, aspartate aminotransferase, cholesterol, and creatinine among three groups. The fish exhibited the highest level of plasma cortisol as well as the lowest levels of triiodothyronine and thyroxine in the HSD group. The fish stocked in the LSD group showed the highest activities of superoxide dismutase, glutathione peroxidase, and catalase as well as the highest content of glutathione in liver. The significant highest total antioxidant capacity occurred in the fish stocked in the LSD group. The results showed that HSD resulted in chronic crowding stress, and exerted negative impact on growth performance, muscle quality, and antioxidative capacity of gibel carp.

Sugar transporter genes in grass carp (Ctenopharyngodon idellus): molecular cloning, characterization, and expression in response to different stocking densities.

Glucose and fructose play a central role in the metabolism and cellular homeostasis of organisms. Their absorption is co-mediated by two families of glucose transporters, Na+-coupled glucose co-transporters (SGLTs) and facilitative Na+-independent sugar carriers (GLUTs), in the intestine. However, limited information has been available on these transporters in fish. Therefore, we studied glut2, sglt1, and sglt4 genes in grass carp (Ctenopharyngodon idellus). The full-length cDNAs of glut2 was 2308 bp, with an open reading frame (ORF) of 503 amino acids (AAs). The full-length cDNAs of sglt1 was 2890 bp, with an ORF of 658 AAs. Additionally, the full-length cDNAs of sglt4 was 2090 bp, with an ORF encoding 659 AAs. The three deduced AA sequences showed high homology between grass carp and other cyprinid fish species. Based on homology modeling, three-dimensional models of GLUT2, SGLT1, and SGLT4 proteins were created and transmembrane domains were noted. glut2, sglt1, and sglt4 were abundantly expressed in the anterior and mid intestine. In particular, glut2 was markedly expressed in liver (P < 0.05). Additionally, the results indicated that different stocking densities (0.9 or 5.9 kg m-2) did not alter intestinal section-dependent expression patterns of the three transporter genes. However, high stocking density impacted segmental mRNA expression levels. This work demonstrated that mRNA expression of sugar transporter genes in the fish intestine was segment specific, and crowding stress may affect the activity of intestinal sugar transporters. These results provided new insights into the relationship between crowding stress and intestinal sugar transporters in fish.

Bacterial community analysis of marine recirculating aquaculture system bioreactors for complete nitrogen removal established from a commercial inoculum.

An experimental recirculating aquaculture system was constructed under ambient seawater conditions to compare microbial community diversity of nitrifying and denitrifying biofilters that were derived from a commercial inoculum used for aquarium applications. Next generation sequencing revealed distinct and diverse microbial communities in samples analyzed from the commercial inoculant and the denitrification and nitrification biofilters. In all samples, communities were represented by a few dominant operational taxonomic units (OTUs). Bacteria having the capacity to carry out ammonia and nitrite oxidation were more abundant in the nitrification biofilter. Similarly, the proportion of the bacterial taxa known to carry out heterotrophic and autotrophic denitrification and participate in sulfur cycling were found in the denitrification bioreactor, and likely originated from the ambient environmental water source. Our results indicated that environmental seawater can be a favorable enhancement to the bacterial consortium of recirculating aquaculture systems biofilters.

Long-term analysis of fish assemblage structure in the middle section of the Sava River - The impact of pollution, flood protection and dam construction.

At the beginning of the 20th century, the middle section of the Sava River in Croatia was unaffected by major human activities and rich in ichthyofauna. The Sava River was important for commercial and recreational fishing for the local population, which still remains today. However, the 1920s mining industry was established in Slovenia, which emitted carbon dust into the Sava River. At the same time, the construction of embankments to mitigate flooding started in the middle section. Furthermore, in the 1980s, the Krsko nuclear power plant (NPP), and in the 2010s, the Krsko hydropower plant (HPP) were built in Slovenia. These activities could have an impact on the composition of fish communities downstream from the major sources of disturbances. Therefore, the main aim of this paper were to analyze the changes in fish assemblages of the Sava River from 1978 to 2017, prior to and after the construction of Krsko NPP and HPP at the Medsave site on the Sava River, 20 km downstream from the major construction operations. Collected data were divided into four sampling periods (SP): SP1, from 1978 to 1980; SP2, from 1991 to 1994; SP3, from 2001 to 2006, and SP4 from 2011 to 2017. Besides alien fish species, water quality and hydromorphological modifications were identified as significant stressors. In SP1 and SP2 limnophilic and eurytopic fish groups were predominant, and 26 different fish species were identified, but in SP3 and SP4 rheophilic fish groups become dominant, and the diversity has declined to 21 species. Threatened species blageon, Telestes souffia seems to be missing from the main course of the Sava River in last 20 years. It can be concluded that disturbances in the fish assemblage pattern have coincided with the presence of multiple stressors of human origin.

First evidence of establishment of the rayed pearl oyster, Pinctada imbricata radiata (Leach, 1814), in the eastern Adriatic Sea.

The Mediterranean Sea is increasingly under threat from invasive species that may negatively affect biodiversity and/or modify ecosystem structure and function. The bivalve mollusc Pinctada imbricata radiata is listed among the 100 most invasive species in the Mediterranean. A first finding of an established population of P. imbricata radiata in the coastal waters of the eastern Adriatic Sea, is presented in this paper. Six and then 30 live specimens were collected in 2015 and in 2017, respectively, at depths of 5 to 15m from the island of Mljet, Croatia. DNA sequencing of the mitochondrial cytochrome c oxidase I gene (COI) revealed three different haplotypes. All samples showed greatest similarity (98 to >99%) to P. radiata COI sequence records in GenBank (=P. imbricata radiata as used in this paper). A Neighbour Joining tree placed all Croatian samples within the 100% bootstrap supported clade for P. imbricata radiata.

WITHDRAWN: First record and distribution of the black pygmy mussel, Limnoperna securis (Lamarck, 1819) in the eastern Adriatic and Ionian seas.

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