Metabolic turnover rate, digestive enzyme activities, and bacterial communities in the white shrimp Litopenaeus vannamei under compensatory growth.

The present work aimed to evaluate the effects promoted by a phase of compensatory growth on metabolic turnover rate, digestive enzyme activity, and bacterial biota of the Pacific white shrimp Litopenaeus vannamei kept under different feeding regimes. Three treatments were evaluated as follows: 70% feed restriction during 3 (T3) and 6 (T6) days, followed by a period of feeding to satiety, and a control treatment without restriction periods. The results showed a full compensatory growth in treatments T3 and T6 by day 35 of the bioassay. A significant increase in trypsin and lipase (T6) activities was observed during compensatory growth, whereas specific amylase activity was significantly lower in treatment T6 compared to T3 but not significantly different from the control group. To determine the metabolic turnover rate of nitrogen in muscle tissue, an analysis of nitrogen isotope values (δ15N) at natural abundance levels was performed. At the end of the experimental period, shrimp under feed restriction had lower metabolic turnover rates and longer nitrogen residence times (t 50) in muscle tissue, as compared to individuals in the control treatment. Regarding the changes in the bacterial communities in shrimp gut, no significant differences were observed at the phylum level, with Proteobacteria being the most abundant bacteria, followed by Actinobacteria. At family taxa level, Rhodobacteraceae presented the highest relative abundance in all treatments, whereas a decrease in Vibrionaceae was observed in treatments T3 and T6 when compared to control shrimps during compensatory growth. At the genus level, a decrease in Celeribacter, Catenococcus, and Epibacterium, and an increase in Ruegeria and Shimia, were identified in shrimp subjected to feed restriction when compared to control organisms during compensatory growth (day 14). At the end of the experimental period, the evaluated parameters showed similar results as those observed in the control treatment, suggesting a normalization of the metabolism and the physiological state. The present findings contribute to a better understanding on the physiological effects produced during compensatory growth in shrimp, which in turn could assist in the development of improved feeding strategies in benefit of the aquaculture industry.

Expression of ion transport proteins and routine metabolism in juveniles of tropical gar (Atractosteus tropicus) exposed to ammonia.

Tropical gar (Atractosteus tropicus) thrives in aquatic habitats with high levels of total nitrogen (TAN) and unionized ammonia (NH3). However, the tolerance of TAN and NH3, the excretion mechanisms involved, and the effects of these chemicals on routine metabolism are still unknown. Therefore, our objectives were to assess the acute toxicity of TAN and NH3 in A. tropicus juveniles after a 96-h exposure (LC50-96 h) to NH4Cl and after chronic exposure to two concentrations (15% and 30% of LC50-96 h TAN) for 12 days, as well as to evaluate the transcriptional effects associated with Rhesus proteins (rhag, rhbg, rhcg) and ion transporters (NHE, NKA, NKCC, and CFTR) in gills and skin; and to determine the effects of TAN and NH3 on routine metabolism through oxygen consumption (µM g-1 h-1) and gill ventilation frequency (beats min-1). LC50-96 h values were 100.20 ± 11.21 mg/L for TAN and 3.756 ± 0.259 mg/L for NH3. The genes encoding Rhesus proteins and ion transporters in gills and skin showed a differential expression according to TAN concentrations and exposure time. Oxygen consumption on day 12 showed significant differences between treatments with 15% and 30% TAN. Gill ventilation frequency on day 12 was higher in fish exposed to 30% TAN. In conclusion, A. tropicus juveniles are highly tolerant to TAN, showing upregulation of the genes involved in TAN excretion through gills and skin, which affects routine oxygen consumption and energetic cost. These findings are relevant for understanding adaptations in the physiological response of a tropical ancestral air-breathing fish.

In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins.

Programmed cell death (PCD) is an essential process for the immune system's development and homeostasis, enabling the remotion of infected or unnecessary cells. There are several PCD's types, depending on the molecular mechanisms, such as non-inflammatory or pro-inflammatory. Hemocytes are the main component of cellular immunity in bivalve mollusks. Numerous infectious microorganisms produce toxins that impair hemocytes functions, but there is little knowledge on the role of PCD in these cells. This study aims to evaluate in vitro whether marine toxins induce a particular type of PCD in hemocytes of the bivalve mollusk Crassostrea gigas during 4 h at 25°C. Hemocytes were incubated with two types of marine toxins: non-proteinaceous toxins from microalgae (saxitoxin, STX; gonyautoxins 2 and 3, GTX2/3; okadaic acid/dynophysistoxin-1, OA/DTX-1; brevetoxins 2 and 3, PbTx-2,-3; brevetoxin 2, PbTx-2), and proteinaceous extracts from bacteria (Vibrio parahaemolyticus, Vp; V. campbellii, Vc). Also, we used the apoptosis inducers, staurosporine (STP), and camptothecin (CPT). STP, CPT, STX, and GTX 2/3, provoked high hemocyte mortality characterized by apoptosis hallmarks such as phosphatidylserine translocation into the outer leaflet of the cell membrane, exacerbated chromatin condensation, DNA oligonucleosomal fragments, and variation in gene expression levels of apoptotic caspases 2, 3, 7, and 8. The mixture of PbTx-2,-3 also showed many apoptosis features; however, they did not show apoptotic DNA oligonucleosomal fragments. Likewise, PbTx-2, OA/DTX-1, and proteinaceous extracts from bacteria Vp, and Vc, induced a minor degree of cell death with high gene expression of the pro-inflammatory initiator caspase-1, which could indicate a process of pyroptosis-like PCD. Hemocytes could carry out both PCD types simultaneously. Therefore, marine toxins trigger PCD's signaling pathways in C. gigas hemocytes, depending on the toxin's nature, which appears to be highly conserved both structurally and functionally.

Assessment of bacteriophage vB_Pd_PDCC-1 on bacterial dynamics during ontogenetic development of the longfin yellowtail (Seriola rivoliana).

The Seriola genus includes species of worldwide commercial importance due to its rapid growth and easy adaptability to confinement conditions. However, like other fish species, large mortalities occur during their early life stages, where the main problems are caused by opportunistic bacteria. Disease control strategies are thus urgently needed. The present study aimed to evaluate the efficacy of phage vB_Pd_PDCC-1 during the early development of longfin yellowtail (Seriola rivoliana), as well as its effect on microbial communities. This broad-host-range phage was added to the culture every 3 days starting from the egg-stage until 12 days after hatching (DAH) at a concentration of 1.41×1010 plaque-forming units (PFU) per mL and at a multiplicity of infection (MOI) of 1. The results showed positive effects (p<0.05) on egg hatching, survival, growth, and pigmentation area in treated larvae. Moreover, high-throughput sequencing analysis of 16S rRNA genes showed that phage administration did not produce significant changes (p>0.05) in the composition and structure of the associated microbiota. However, sequences affiliated to the Gammaproteobacteria class were displaced by those belonging to the Alphaproteobacteria class over time regardless of the treatment received. At the family level, there was a decrease in Rhodobacteraceae, Pseudoalteromonadaceae, and Flavobacteriaceae in both groups over time. To our best knowledge, this study represents the first attempt to evaluate the effect of a phage as a biological control agent during ontogenetic development of longfin yellowtail larvae. KEY POINTS: • Phages can be used against proliferation of Vibrio in fish cultures. • Seriola includes several important commercial fish species due to its rapid growth. • Phages do not cause significant changes in the associated microbiota.

Comparative ontogenetic development of two marine teleosts, gilthead seabream and European sea bass: New insights into nutrition and immunity.

Gilthead seabream and European sea bass are two of the most commonly farmed fish species. Larval development is critical to ensure high survival rates and thus avoid unacceptable economic losses, while nutrition and immunity are also important factors. For this reason this paper evaluates the ontogenetic development of seabream and sea bass digestive and immune systems from eggs to 73 days post-fertilisation (dpf) by assessing the expression levels of some nutrition-relevant (tryp, amya, alp and pept1) and immune-relevant (il1b, il6, il8, tnfa, cox2, casp1, tf, nccrp1, ighm and ight) genes. The results point to similar ontogenetic development trends for both species as regard nutrition and differences in some immunity related genes.

Dietary administration of ß-1,3/1,6-glucan and probiotic strain Shewanella putrefaciens, single or combined, on gilthead seabream growth, immune responses and gene expression.

It is widely known that ß-glucans and probiotic bacteria are good immunostimulants for fish. In the present work we have evaluated the dietary effect of ß-1,3/1,6-glucan (isolated from Laminarina digitata) and Pdp 11 (Shewanella putrefaciens, probiotic isolated from gilthead seabream skin), single or combined, on growth, humoural (seric level of total IgM antibodies and peroxidase and antiprotease activities) and cellular innate immune response (peroxidase and phagocytic activities of head-kidney leucocytes), as well as the expression of immune-related genes in gilthead seabream (Sparus aurata). Four treatment groups were established: control (non-supplemented diet), Pdp 11 (10(9) cfu g(-1)), ß-1,3/1,6-glucan (0.1%) and ß-1,3/1,6-glucan + Pdp 11 (0.1% and 10(9) cfu g(-1), respectively). Fish were sampled after 1, 2 and 4 weeks of feeding. Interestingly, all supplemented diets produced increments in the seabream growth rates, mainly the Pdp 11-suplemented diet. Overall, Pdp 11 dietary administration resulted in decreased serum IgM levels and peroxidase activity. However, the seric antiprotease activity was increased in fish fed with both supplements together. Furthermore, ß-1,3/1,6-glucan and combined diet increased phagocytic activity after 2 or 4 weeks. At gene level, IL-1ß and INFγ transcripts were always up-regulated in HK but only the interleukin reached significance after 4 weeks in the group fed with ß-glucan. On the contrary, IgM gene expression tended to be down-regulated being significant after 1 week in seabream specimens fed with ß-glucan or ß-glucan plus Pdp 11. These results suggest that ß-1,3/1,6-glucan and Pdp 11 modulate the immune response and stimulates growth of the gilthead seabream, one of the species with the highest rate of production in Mediterranean aquaculture.

Effects of dietary ß-1,3/1,6-glucan on the antioxidant and digestive enzyme activities of Pacific red snapper (Lutjanus peru) after exposure to lipopolysaccharides.

The effect of ß-1,3/1,6-glucan, derived from yeast, on growth, antioxidant, and digestive enzyme performance of Pacific red snapper Lutjanus peru before and after exposure to lipopolysaccharides (LPS) was investigated. The ß-1,3/1,6-glucan was added to the basal diet at two concentrations (0.1 and 0.2 %). The treatment lasted 6 weeks, with sampling at regular intervals (0, 2, 4, and 6 weeks). At the end of this period, the remaining fish from either control or ß-glucan-fed fish were injected intraperitoneally with LPS (3 mg kg(-1)) or with sterile physiological saline solution (SS) and then sampled at 0, 24, and 72 h. The results showed a significant increase (P < 0.05) in growth performance after 6 weeks of feeding with ß-glucan. Superoxide dismutase (SOD) activity in liver was significantly higher in diets containing 0.1 % ß-glucan in weeks 4 and 6, compared to the control group. ß-Glucan supplementation at 0.1 and 0.2 % significantly increased aminopeptidase, trypsin, and chymotrypsin activity. At 72 h after injection of LPS, we observed a significant increase in catalase activity in liver from fish fed diets supplemented with 0.1 and 0.2 % ß-glucan; SOD activity increased in fish fed with 0.1 % ß-glucan in relation to those injected with SS. Feed supplemented with ß-1,3/1,6-glucan increased growth, antioxidant activity, and digestive enzyme activity in Pacific red snapper.