The nuclear factor of activated T cells (NFAT) proteins have crucial roles in the development and function of the immune system since they not only regulate activation of T cells but are also ...involved in the control of thymocyte development and T-cell differentiation. In this study, NFATc3 was characterized from the Pacific red snapper, Lutjanus peru. LpNFAtc3, which contains an open reading of 3300 bp frame coding for a protein of 1100 aa with a predicted molecular weight of 118.52 kDa. The predicted protein showed a conserved NFAT family structure with signature motifs and domains, sharing high identity (up to 76%) compared to other fish sequences. NFATc3 gene expression was analyzed by real time-PCR in head-kidney cells (leukocytes and lymphocytes) following yeast, zymosan and Vibrio parahaemolyticus stimulation along with the expression of upstream (ILF2, ILF3 and CaN) and downstream (CD3, TCRβ, IL-6 and IL-12) molecules. This study revealed a broad expression of NFATc3 with a relative strong expression in intestine and lymphocytes. The expression of NFATc3 was differentially up-regulated after stimulation with yeast in head-kidney leukocytes and after bacterial infection in lymphocytes at 24 h. Interestingly, the yeast and zymosan were able to activate ILF2, ILF3 and CaN mRNA gene expression in both kinds of cells. On the other hand, NFAT downstream genes such as CD3, TCRβ, IL-6 and IL-12 were significantly up-regulated in leukocytes stimulated with yeast or zymosan at 12 h; however in lymphocytes, this up-regulation was detected when cells reacted to V. parahaemolyticus stimuli at 24 h. Stimulating Pacific red snapper leukocytes with immunostimulants as yeast significantly up-regulated the expression of NFATc3, and up- and down-stream molecular genes and NFATc3 lymphocytes expression were potentially involved in responses to invasion of bacterial pathogens in an early immune response.
•NFATc3 functional architecture from Lutjanus peru was characterized in silico.•NFATc3 was highly expressed in intestine and isolated lymphocytes.•NFATc3 was up-regulated in HK leukocytes (yeast) and lymphocytes (bacteria).•Vibrio parahaemolyticus strongly activates NFAT downstream expression in lymphocytes.•NFAT-related genes are differentially modulated by yeast and bacteria.
Juvenile Pacific giant lions-paw scallops Nodipecten subnodosus were fed the toxic dinoflagellate Gymnodinium catenatum, a producer of paralytic shellfish poison (PSP), supplied with Isochrysis ...galbana (a nontoxic microalgae). Short-term (<24 h) experiments were performed to determine clearance and ingestion rates of G. catenatum. Kinetics of PSP was examined in longer-term experiments (>2 days). At high food concentrations, juvenile scallops showed production of pseudofeces, partial shell valve closure, and reduction in feeding. According to HPLC analysis, the only toxins present in the dinoflagellate G. catenatum and in the scallops were the gonyautoxins (GTXs), except in the labial palps and digestive gland, where trace amounts of saxitoxin (STX) were present in scallops. These tissues could play an important role in toxin biotransformation. The ranking of toxin concentration in tissues was: digestive gland > labial palps > intestine > gills > mantle > adductor muscle, where the total contribution of viscera was more than 80% of the total toxin body burden. Juvenile scallops exhibited no apparent detrimental physiological responses during the long-term feeding experiment. The dinoflagellate may contribute nutrients to the scallop, in addition to the microalgae I. galbana. The dinoflagellate may enhance cell uptake and byssus production. Once PSP accumulated during the first 12 days, it was slowly eliminated. The limited capacity for accumulating toxins in the adductor muscle favors domestic marketing of scallops. PUBLICATION ABSTRACT
The effect of dietary β-Ο-glucan, vitamin E, and β-carotene supplements in juvenile brown shrimp, Farfantepenaeus californiensis, inoculated with white spot syndrome virus (WSSV) was evaluated. ...Groups of 30 organisms (weighing 1 ± 0.5 g) were cultured in 60 L fiberglass tanks and fed daily with β-1.3-glucan (0.1%), vitamin E (0.01%), and β-carotene (0.01%) for 23 days; the specimens were then inoculated with WSSV. The antioxidant activity of the enzymes superoxide dismutase (SOD) and catalase (CAT) were determined in the hepatopancreas and muscle at 0, 1, 6, 12, 24, and 48 h after inoculation. Shrimp fed with β-1.3-glucan, vitamin E, and β-carotene significantly increased SOD activity in the hepatopancreas and muscle at 12 and 24 h post-infection, respectively. Shrimp fed with vitamin E and β-1.3-glucan registered an increment in SOD activity from 12 to 48 h post-infection. Shrimp fed with β-carotene increased SOD activity before infection with WSSV, and shrimp fed with β-1.3-glucan and vitamin E increased CAT activity, also before infection. The CAT activity response in shrimp muscle increased with respect to the control group for all treatments tested from 1 to 6 h after inoculation with WSSV. The highest antioxidant response was registered in shrimp fed with vitamin E. Juvenile shrimp fed with vitamin E and later inoculated with WSSV registered 100% mortality at 72 h, but shrimp fed with β-Ο-glucan and β-carotene showed greater resistance to WSSV, with mortality at 144 h post-infection. This study demonstrated the capacity of juvenile Farfantepenaeus californiensis fed β-Ο-glucan, vitamin E, or β-carotene to increase the antioxidant response before and after viral infection.
•Caspase-1, -3 and 8 functional architectures from Totoaba macdonaldi were characterized in silico.•Apoptosis-mediated signaling by caspase genes is activated after bacterial challenge in HK and ...spleen leukocytes.•Expression of CAT, GPX1 and GPX4 genes in leukocytes exposed to V. parahaemolyticus is promoted.
Caspases are a family of proteases involved in many important biological processes including apoptosis and inflammation. In order to get insights into the caspase gene family and antioxidant enzymes in Totoaba macdonaldi during bacterial infection, an in vitro assay was performed involving three different types of caspases (Casp-1, Casp-3 and Casp-8) and antioxidant enzymes (catalase, gluthathione peroxidase 1 and 4) after Vibrio parahaemolyticus and Aeromonas veronii infection, using head-kidney and spleen leukocytes from the teleost fish totoaba at 12 and 24 h post-exposure. Characterization of caspases by bioinformatics analyses showed that TmCas-1, TmCas-3 and TmCas-8 shared overall sequence identities of 82-61%, 85–97% and 77-63%, respectively, with other teleost fish. Caspase-1, -3 and -8 proteins revealed a conserved penta-peptide sequence at the catalytic site and three amino acid residues involved in the catalysis (H, G and C), as well as two conserved domains. The expression levels of the three caspases were detected in a wide range of fish tissues; however, they varied among tissues and caspases, which were highly up-regulated in immune organs, such as head-kidney, liver and/or spleen. The pathogen-induced gene expression pattern revealed two interesting facts; first, that the expression of all the caspase genes and antioxidant enzyme genes evaluated in this study were strongly induced following V. parahaemolyticus infection; second, these up-regulations reached a maximum level at 24 h post-infection in head-kidney whereas in spleen leukocytes, it was observed at 6-h post-infection. In conclusion, based on these observations, the acute toxic effects of V. parahaemolyticus are associated to cell death and release of free radicals. This information provides a better understanding of the effects and nature of early immune response against common bacterial infections in totoaba leukocytes.
Helicobacter pylori have colonized the gastric mucosa of half of the population worldwide. This bacterium is classified as a definitive type I carcinogen by the World Health Organization and no ...effective vaccine has been found against it yet. Thus, a logical and rational vaccine design against H. pylori is necessary. Because of its tremendous complexity and elicited immune responses, the vaccine design should considered multiple antigens to enhance immune-protection, involved in the different stages of pathogenesis besides inducing a specific immune response by B- and T-cell multi-epitopes. In this study, emphasis was placed on the design of a new unique vaccine named CTB-multiHp. In silico techniques were used to design a chimeric construct consisting of cholera toxin B subunit fused to multi-epitope of urease B (residue 148–158, 188–198), cytotoxin-associated gene A (residue 584–602), neutrophil activating protein (residue 4–28), vacuolating cytotoxin gene A (residue 63–81), H. pylori adhesine A (residue77–99), heat shock protein A (residue 32–54) and gamma glutamyl transpeptidase (residue 271–293). The tertiary structure and features of the vaccine were analyzed. The chimeric protein was expressed in Escherichia coli BL21 and the serology analyses indicated that the CTB-multiHp protein produced exhibit immune-reactivity. The results showed that CTB-multiHp could be a good vaccine candidate against H. pylori. Ongoing studies will evaluate the effects of CTB-multiHp against H. pylori infection.
•A novel chimeric vaccine against H. pylori was designed.•T and B cell epitopes for H. pylori antigens were predicted by in silico approach.•The CTB-multiHp designed vaccine exhibit immunoreactivity using serology analysis.
Debaryomyces hansenii has been described to be effective probiotic and immunostimulatory marine yeast in fish. Nonetheless, to the best of our knowledge, it has been not assayed in ruminants. This ...study attempts to describe the immunostimulatory effects of its β-glucan content through in vitro assays using goat peripheral blood leukocytes at 24 h of stimulation. The structural characterization of yeast glucans by proton nuclear magnetic resonance indicated structures containing (1-6)-branched (1-3)-β-D-glucan. In vitro assays using peripheral blood leukocytes stimulated with β-glucans derived from three D. hansenii strains and zymosan revealed that β-glucans significantly increased cell immune parameters, such as phagocytic ability, reactive oxygen species production (respiratory burst), peroxidase activity and nitric oxide production. Antioxidant enzymes revealed an increase in superoxide dismutase and catalase activities in leukocytes stimulated with yeast β-glucans. This study revealed that yeast β-glucans were able to activate dectin-1 mRNA gene expression in leukocytes. The TLR4 gene expression was up-regulated in leukocytes after stimulation with yeast β-glucans. In conclusion, β-glucans were able to modulate the immune system by promoting cell viability, phagocytic activity, antioxidant immune response and immune-related gene expression in leukocytes. Therefore, β-glucans derived from Debaryomyces hansenii should be considered a potential immunostimulant for goat production systems.
•First report on the potential marine D. hansenii-derived β-glucans as immunostimulatory compounds in ruminants.•D. hansenii derived-β-glucans contain (1-6)-branched (1-3)-β-D-glucans.•β-glucans are no toxic and safe molecules for goat leukocytes.•β-glucans stimulate the immune parameters by promoting phagocytic activity, NO and ROS production in goat leukocytes.•β-glucans increase antioxidant activities in caprine peripheral blood leukocytes.
This study evaluates the effects of the dietary administration of the live yeast Debaryomyces hansenii strain L2 on the immune responses of gilthead seabream for 4 weeks. Cellular immune parameters ...were measured from serum and head‐kidney leucocytes respectively. The expression levels of immune‐associated genes were quantified by real‐time polymerase chain reaction. In addition, the profile of intestinal microbiota was studied by denaturing gradient gel electrophoresis. The results showed that seabream fed a diet containing D. hansenii had significantly increased cellular immune parameters. The yeast‐supplemented diet up‐regulated the expression of most seabream genes at week 2 and down‐regulated all of them at week 4, except in the head‐kidney. Finally, a reduction in the diversity of the intestinal microbiota was detected in those specimens receiving the yeast‐supplemented diet. These results support the idea that the live yeast D. hansenii strain L2 stimulates the immune system of gilthead seabream.
Although Litopenaeus vannamei is a widely studied species, the information on how the organisms respond to natural daily variations of environmental conditions such as temperature and dissolved ...oxygen, and how such conditions alter the physiological responses, is scarce. In the present work, the strategies used by shrimps to cope with temperature and dissolved oxygen fluctuations during 24 days were investigated through the evaluation of oxygen consumption and heat shock proteins (HSP) gene expression. During daily fluctuations, no change in oxygen consumption in the short-term, but a significant increase in the long-term during hyperthermia conditions was registered, whereas a significant decrease during hypoxia was observed during all the bioassay. On the other hand, HSP70 and HSP90 gene expression increased in gills under thermal stress but was down-regulated under hypoxia, in both the short- and the long-term. This study highlights that to counteract environmental variations of temperature and dissolved oxygen, the shrimps use molecular compensatory mechanisms (HSP gene expression) that are different to those used under constant hypoxic conditions, suggesting that hypoxia can compromise physiological cytoprotection.
•Shrimp's oxygen consumption responses to cyclic hyperthermia and hypoxia are different between short and long-term exposure.•Under oscillating conditions, HSP gene expression in gills is compromised during hypoxia.•Oxygen consumption and HSP gene expression responses during cyclic conditions are different from those at constant hypoxia.
The effectiveness of the yeast Debaryomyces hansenii against Pénicillium citrinum in Persian lime was evaluated. Postharvest decay is a serious problem in the storage of many fresh fruits and ...vegetables. P. citrinum was isolated from fruits of a packinghouse in San Pedro Lagunillas, Nayarit, Mexico and turned out to be an endemic pathogen against Persian lime. This is the first report that involves this strain as a pathogen for citrus. The control of Persian lime decay caused by P. citrinum in postharvest and yeast population dynamics were evaluated. In order to enhance yeast establishment on the fruit carposphere Dextrose and Tween 80 were added as adjuvants to the D. hansenii suspension. The use dextrose and Tween 80 promoted a better establishment of the yeast (3.2x10^sup 8^ UFC/fruit - 25 days storage), however the infection by P. citrinum on Persian lime was higher (20 to 60% of infected fruits). The application of yeast suspension without adjuvants strongly inhibited fungal decay (100%) even though yeast population counts were minimal (3.3x10^sup 5^ CFU/fruit) at the end of the storage period (25th day) at 25°C and 60-75% of relative humidity. Application of D. hansenii can be an alternative of postharvest disease management.
Una proteína de membrana externa de 48 kDa (omp48) de Aeromonas veronii (A. veronii), que tiene afinidad por secreciones de la mucosa (tal como mucina, lactoferrina, inmunoglobulinas o colágeno), ...induce una respuesta inmune estadísticamente significativa al ser administrada intragástricamente a conejos blancos de Nueva Zelanda. Estudios posteriores mostraron una correlación positiva entre las inmunoglobulinas y la respuesta de células productoras de anticuerpo (CPA). La administración intragástrica del antígeno induce una respuesta vigorosa, tanto local como sistémica, principalmente de inmunoglobulinas A y G contra la omp48 de A. veronii. Estas se evidenciaron por la presencia de (i) células productoras de anticuerpos contra omp48 en el tracto gastrointestinal, bazo y sangre, y (ii) elevados niveles de anticuerpos circulantes.La estimulación del sistema inmune de mucosa mediante antígenos bacterianos inmunogénicos, puede ser una importante alternativa para el control y prevención de enfermedades asociadas a Aeromonas.
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