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We performed several experiments using three strains of Virgibacillus salexigens, namely, P2, NT N53, and C-20MoT (DSM 11483T), which were isolated from completely different sources, in relation to bacteriocin production ability. Results of whole-genome sequencing analysis revealed that all strains have very similar sequences encoding class IId bacteriocin. Although a partial amino acid sequence of the purified bacteriocin produced by strain P2 isolated from fermented food was previously reported, whole-genome sequencing and the N-terminal sequencing results in this study showed that its complete amino acid sequence consisted of 48 residues, which corresponded to that of the hypothetical bacteriocin encoded by the gene in Virgibacillus massiliensis strain Vm-5T (DSM 28587T) isolated from the human gut. From the results of 16S rRNA gene sequencing and whole-genome sequencing analyses, we taxonomically confirmed Vm-5T to be a strain of V. salexigens, and its broth culture showed antibacterial activity. Strain NT N53 isolated from the deep-sea floor produced two bacteriocins, namely, NTN-A and NTN-B. The results of N-terminal sequencing, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and whole-genome sequencing analyses showed that their amino acid sequences differed in only one residue, and NTN-A showed the same sequence as the bacteriocin produced by strain P2. Although strain C-20MoT isolated from a solar saltern had the coding sequence very similar to that of NTN-A, its broth culture showed no antibacterial activity. This finding suggests that class IId bacteriocin-producing or bacteriocin-gene-encoding V. salexigens strains are widely distributed in distinct environment sources with different geographical and material properties.

Citrus essential oils are utilized as phytogenic feed additives. The current study investigated the effects of bergamot essential oil (BEO) supplementation in fish feed on growth performance, hematological and biochemical parameters, and antioxidant activities of juvenile common carp Cyprinus carpio . All groups (control, 0.2%, 0.4%, and 0.8% BEO) were designed in triplicate. The fish were fed these doses twice daily for 60 days. The 0.2% BEO concentration was the most effective in enhancing fish growth, as evidenced by the highest relative and specific growth rate. No significant differences were observed in the hematological parameters in the experimental groups ( p  > 0.05). Compared with the control group, positive effects of BEO feed supplementation on serum biochemical parameters were observed in all groups, especially in the 0.2% BEO group. Although there was no statistically significant difference in catalase enzyme activity, antioxidant activity as indicated by superoxide dismutase and glutathione peroxidase antioxidant enzyme activities significantly increased in the 0.2% and 0.4% BEO groups ( p  < 0.05). In conclusion, these findings suggest that in all BEO groups, especially in the 0.2% and 0.4% concentrations, BEO could enhance growth performance, health, and cellular defense mechanisms in common juvenile carp, making BEO a valuable additive in fish feed.

To test the effects of probiotic bacteria against crowding stress, juvenile Nile tilapia Oreochromis niloticus were fed commercial feed supplemented with 1 × 10 10  cfu/g pellets of Lactobacillus rhamnosus for 3 weeks. The fish were confined and subjected to crowding for 7 and 14 days after which their physiological condition was investigated. The administration of probiotic prophylaxis improved growth performance even in the stressed fish. After 7 days of stress exposure, fish receiving probiotic-supplemented feed showed proactive behavior and coping responses to the stressor, as evidenced by elevated plasma glucose levels and osmolality, stabilized plasma electrolytes, and a higher RNA:DNA ratio. Fish stressed for 7 days and fed normal commercial diet showed impairment of plasma electrolytes; after 14 days, the plasma osmolality, electrolytes, glucose, and RNA:DNA ratio decreased, indicating physiological maladaptation to the stressor. The growth rate was also found to be reduced, suggesting that energy demand exceeded the energy available from metabolism and dietary uptake. This latter effect was not observed in fish fed the probiotic-supplemented diet. The results suggest that probiotic bacteria prophylaxis may increase energy availability for metabolic support of the crowding stress response and improve the stress coping capacity of fish.

:  The physiology of yellowtail fed a non‐fishmeal diet was examined, with a specific interest in the role of taurine in disease resistance and cholesterol metabolism. Decrease of disease resistance in fish fed a non‐fishmeal diet was confirmed by mortality due to natural infection with pseudotuberculosis and artificial infection with Lactococcus garvieae. It is suggested that the most important symptoms related to decrease of disease resistance in fish fed a non‐fishmeal diet is anemia. Anemia was improved by supplementation with taurine. Significant elevation of relative expression of HMG‐CoA reductase mRNA in fish fed a non‐fishmeal diet suggests that cholesterol synthesis would be activated and not dysfunctional. Plasma cholesterol of these fish was elevated to the levels of control fish by supplementation of both cholesterol and taurine. These results suggest that hypocholesterolemia observed in fish fed a non‐fishmeal diet compared with a fishmeal diet would be caused by insufficient dietary cholesterol and decrease of endogenous cholesterol due to the lack of dietary taurine.

The microbiological properties of Myanmar traditional fermented products were characterized using four different brands of bottled shrimp sauce products sold frequently at a large grocery market. Viable cells of these products were detected at various levels (102–105 cfu/ml) using de Man, Rogosa and Sharpe (MRS) agar medium containing 10% NaCl, and all isolates from the product samples were identified as the halophilic lactic acid bacterium Tetragenococcus muriaticus by 16S rRNA gene analysis. On the other hand, culture-independent bacterial analysis using a combination of clone library analysis and terminal restriction fragment length polymorphism (T-RFLP) analysis targeting the 16S rRNA gene demonstrated that these bacterial communities can be classified into three types. In two shrimp sauce products, a strictly anaerobic non-spore-forming halophile belonging to the genus Halanaerobium was detected as the dominant bacterium. In one shrimp sauce product, Tetragenococcus was the dominant genus. However, in the remaining shrimp sauce product, a number of bacterial species typically found in various environments, including the genera Geosporobacter, Sporohalobacter, Lysinibacillus, Soehngenia, and Tepidiphilus, were detected. These findings suggest that typical fermentation bacterial species found in traditional fermented fishery products, such as those belonging to the genera Tetragenococcus and Halanaerobium, are also frequently found in Myanmar shrimp sauce products.

The Birt-Hogg-Dubé (BHD) disease is a genetic cancer syndrome. The responsible gene, BHD, has been identified by positional cloning and thought to be a novel tumor suppressor gene. BHD mutations cause many types of diseases including renal cell carcinomas, fibrofolliculomas, spontaneous pneumothorax, lung cysts, and colonic polyps/cancers. By combining Gateway Technology with the Ksp-Cre gene knockout system, we have developed a kidney-specific BHD knockout mouse model. BHD(flox/flox)/Ksp-Cre mice developed enlarged kidneys characterized by polycystic kidneys, hyperplasia, and cystic renal cell carcinoma. The affected BHD(flox/flox)/Ksp-Cre mice died of renal failure at approximate three weeks of age, having blood urea nitrogen levels over tenfold higher than those of BHD (flox/+)/Ksp-Cre and wild-type littermate controls. We further demonstrated that these phenotypes were caused by inactivation of BHD and subsequent activation of the mTOR pathway. Application of rapamycin, which inhibits mTOR activity, to the affected mice led to extended survival and inhibited further progression of cystogenesis. These results provide a correlation of kidney-targeted gene inactivation with renal carcinoma, and they suggest that the BHD product FLCN, functioning as a cyst and tumor suppressor, like other hamartoma syndrome-related proteins such as PTEN, LKB1, and TSC1/2, is a component of the mTOR pathway, constituting a novel FLCN-mTOR signaling branch that regulates cell growth/proliferation.

Cell lines derived from fish tissues are resistant to premature senescence under typical culture conditions. Previously, we demonstrated that fish genomes do not have a p16 INK4a / Arf locus and that the absence of this locus underlies the lack of senescence in cultured fish cells. However, other factors may also contribute to this resistance. In amniotes, promyelocytic leukemia (PML)-IV proteins are involved in the generation of PML nuclear bodies (PML NBs), which are connected with premature senescence. The lack of a pml gene in fish genomes may be involved in the mechanism of resistance to cellular senescence. Heterologous expression of human PML-IV in an Epithelioma papulosum cyprini cell line induced the formation of PML NB-like speckled structures. The cells displayed characteristic features of cellular senescence, namely, growth suppression, a large, flattened morphology, and increased SA-β-gal activity. Additionally, the levels of proinflammatory senescence-associated secretory phenotype (SASP) factors increased in the cells, suggesting a link between the absence of PML NBs and cellular resistance to senescence. Expression of the CCAT enhancer binding protein beta gene, which encodes a transcription factor of proinflammatory SASPs, was not increased, nor was there any elevation in the activity of NF-κB, a transcription factor for proinflammatory SASP factors and C/EBPβ. Epigenetic regulatory mechanisms may contribute to the induction of proinflammatory SASP factors by PML NBs.

The present study histologically examined the effects of glucan-containing and Lactobacillus rhamnosus GG (LGG)-containing diets on intestinal damages inflicted on Nile tilapia by Aeromonas challenges. Tilapia were fed control, glucan, and LGG diets for 2 weeks and were subsequently challenged with Aeromonas . The intestines were then histologically examined at 1, 7, 14, and 21 days post-infection. Mortality following the challenge was lower for the fish fed the glucan and LGG diets. The intestines of these groups also showed increased inflammatory cell infiltration and reduced intestinal damage from Aeromonas . Moreover, inflammatory cell infiltration occurred more rapidly in the glucan-fed than in the LGG-fed fish following the challenge. Before the challenge, the dominant mucous cell was the acid type in all the tests. After the challenge, the main mucus cell type in the proximal intestine of the glucan-fed fish shifted to AB-PAS double-staining cells, while in the LGG-fed fish, it remained the acid type throughout the test period, and the number of double-staining cells was smaller than in the control fish after the challenge. Thus, the different mucous cell and inflammatory cell responses show that glucan and LGG might have different immunostimulative effects, although they both reduced the intestinal damage following Aeromonas challenges.

Furazolidone, an antibacterial drug that was once widely used in the livestock industry and aquaculture, is now prohibited in numerous countries. It is difficult to detect residual furazolidone because it is readily metabolized in animal tissues but, by using and liquid chromatography coupled with tandem mass spectrometry, its metabolite, 3-amino-2-oxazolidinone (AOZ) can be detected. Here we describe the validity of an enzyme-linked immunosorbent assay (ELISA) kit to detect AOZ in Japanese eel Anguilla japonica tissue. ELISA is capable of detecting AOZ at 1.0 μg/kg in an eel sample with excellent accuracy and precision. Our results show that ELISA is suitable for regulatory purposes and for studying the fate of AOZ residues in eel treated with furazolidone. To measure the persistence of AOZ in eel tissues, eels (1.4–6.5g) were immersed in tanks containing 2 and 10 mg furazolidone/L for 3 h, and then maintained in a tank supplying well water for the next 160 days. The half-lives of AOZ, calculated from the linear terminal part of the excretion curve, were 25.0 days in muscle and 21.6 days in liver from fish exposed to 2 mg/L furazolidone. In the eels treated with 10 mg/L furazolidone, by contrast, high levels of AOZ were detected in liver and muscle, but the half-lives of AOZ were similar to those in fish treated with 2 mg/L furazolidone. The half-lives of AOZ in eel tissues were prolonged by the condition of low water temperature.

Ammonia is one of the most common and pervasive pollutants in the aquatic habitat affecting the health of aquatic animals. It affects the number and morphology of mucus producing cells, resulting in excessive and irregular secretion and alterations in quantity of mucus, which can lead to complications in fish health. A toxicity test on the mucus cells localized on the gills (primary lamellae) and inner operculum epithelial lining of zebrafish Danio rerio at different periods of exposure (24, 48 and 72 h) followed by recovery periods was studied using histopathology, scanning electron microscopy and anterior gradient 2 homologue ( agr2 ) gene expression techniques. Fish samples subjected to low, medium and high external toxic ammonia concentrations (L: 17.21 mg/l, M: 25.81 mg/l and H: 38.91 mg/l NH 3 -N) were observed to display a higher mucus layer production and active secretion compared with the control. Gill cellular alterations were more severe at 48 and 72 h. A high expression of agr2 was detected at 48 h (L and M) recovery periods and a (H) exposure and recovery period indicating an increase in quantity in newly proliferated alcian blue stained mucus cells and excessive secretion observed by histopathology. Such expression decreased at 72 h resulting in a decrease in mucus cell density and secretion.