Cyrtocarpa edulis fruit and its immunostimulant effect on Almaco Jack Seriola rivoliana: in vitro, in vivo and ex vivo studies.

The present study investigates for the first time chemical, proximate analyses and immunostimulant effect of Cyrtocarpa edulis fruit (CeF). Three design experiments were carried out to evaluate immunostimulant effect of C. edulis fruit: in vitro, in vivo and ex vivo studies in juveniles Almaco jack Seriola rivoliana. In general, nutraceutical studies performed by gas chromatography/mass spectrometry (GC-MS) in CeF revealed a major quantity of the carbohydrate groups and phytosterols such as ß-sitosterol. Their phytochemical and antioxidant values exposed a significant content of total phenols, flavonoids, and tannins, showing an antioxidant capacity against hydroxyl and superoxide radical. The in vitro results confirm that CeF is edible and enhanced the innate immune response in head-kidney leukocytes after 24 h of immunostimulation. The in vivo results showed that myeloperoxidase, nitric oxide production, as well as antioxidant enzymes were enhanced in skin mucus of those fish fed with CeF. Interestingly in the intestine, IL-ß, TNF-α, MARCO and Piscidin gene expression were up-regulated in fish fed with C. edulis after 4 weeks. Finally, ex vivo experiments showed an important enhancement on cellular parameters (phagocytosis, respiratory burst, myeloperoxidase, and nitric oxide production) in head-kidney leukocytes of fish fed CeF and intraperitoneally infected with A. hydrophila. The results demonstrate that C. edulis fruit (0.5%) represents an available phytochemical and antioxidant rich alternative with great potential as fish immunostimulant additive.

Bisphenol A Induces Reactive Oxygen Species Production and Apoptosis-Related Gene Expression in Pacific Red Snapper Lutjanus peru Leukocytes.

Bisphenol A is one of the most used components of the polycarbonate plastic industry in the word. This contaminant has disrupting effect in cells in in vitro and in vivo in fish. This study evaluated for the first time the cytotoxicity, oxidative stress and apoptosis induced by bisphenol A (BPA) in head-kidney and spleen leukocytes isolated from Pacific red snapper Lutjanus peru. Head-kidney and spleen leukocytes were exposed to 100, 1000 and 10,000 µg/mL of BPA at 2 and 24 h. Results showed cytotoxicity of BPA at 1000 and 10,000 µg/mL. Cell viability > 80% was observed in leukocytes exposed to 100 µg/mL for 2 h; thus, this concentration was selected for the remainder of the study. Reactive oxygen species (ROS) production, analyzed by DCF-DA and NBT assays, significantly increased in those leukocytes exposed to BPA compared to controls after 2 or 24 h. Superoxide dismutase and catalase activities increased in head-kidney leukocytes after 24 h of BPA exposure. Apoptosis was inferred from caspase (casp-1 and casp-3), granzyme A (granz-A) and perforin 1 (perf-1) gene expression, which was significantly up-regulated, at 2 h BPA exposure in head-kidney leukocytes, and from granz-A and perf-1, which were up-regulated, after 24 h BPA exposure in spleen leukocytes. Short cytoplasmic prolongations and membrane blebs, suggestive of apoptosis, were observed by scanning electron microscopy. These data suggest that BPA at 100 µg/mL induces cytotoxicity, oxidative stress, apoptosis in Pacific red snapper head-kidney and spleen leukocytes.

In vivo and ex vivo studies support the immunostimulant and immunoprotective effect of Damiana (Turnera diffusa Willd) in Almaco Jack (Seriola rivoliana).

Damiana (Turnera diffusa Willd) was evaluated in vitro for antioxidant and antibacterial activities against Staphylococcus aureus and Streptococcus pyogenes (as a preliminary screening assessment) by high-performance thin-layer chromatography (HPTLC)-Direct bioautography. A study was performed in vivo to evaluate the effects of Damiana enriched diets at 0.5 % on immune parameters in mucus and serum and gene expression in Almaco Jack (Seriola rivoliana) intestine after two and four weeks; an infection with Aeromonas hydrophila at 1x107 colony forming units (CFU) followed and an ex vivo study was carried out using head-kidney leukocytes. Ferric reducing ability of plasma (FRAP) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) assays showed high antioxidant activities in Damiana leaves; even in the ABTS assay, Damiana at 300 µg/mL showed similar activity to ascorbic acid - the standard control. Damiana exhibited strong in vitro antimicrobial activity against S. aureus and S. pyogenes. In vivo studies showed a strong enhancement of myeloperoxidase, nitric oxide, superoxide dismutase, and catalase activities in mucus and serum of S. rivoliana supplemented with Damiana; their immunological response enhanced after infection with A. hydrophila. IL-1ß, TNF-α, and IL-10 gene expressions upregulated in the fish intestine challenged with the bacterium. Piscidin and macrophage (MARCO) receptor gene expression up-regulated at week 4 and down-regulated after infection. Intestinal histology results confirm that Damiana not cause inflammation or damage. Finally, the ex vivo study confirmed the immunostimulant and protective effects of Damiana through increased phagocytic, respiratory burst, myeloperoxidase activities and nitric oxide generation before and upon the bacterial encounter. These results support the idea that Damiana has the potential as an immunostimulant additive for diets in aquaculture by enhancing immune parameters and protecting Almaco Jack against A. hydrophila infections upon four weeks of supplementation.

Immunostimulant effects of diet supplementation with yellow (Pouteria campechiana), white (Casimiroa edulis), and black (Diospyros digyna) sapote nanocapsules on laying hens: in vitro and in vivo study.

Poultry is commonly infected by different bacteria and parasites in the environment, resulting in increased morbidity and mortality, but immunostimulants have been enhancing non-specific defense mechanisms conferring laying hens' protection. For this purpose, the pulp of yellow (Pouteria campechiana), white (Casimiroa edulis), and black (Diospyros digyna) sapotes were nanoencapsulated (YWB-SN) and evaluated in laying hens' peripheral blood leukocytes to test their addition to the experimental diets at a concentration of 0.5% (5g/kg of dry food) for 1 month (with two samples at days 15 and 30). The YWB-SN were safe when exposed to peripheral blood leukocytes (PBLs). The in vitro experiment showed that these nanocapsules enhanced reactive oxygen species production, and B-SN stimulated phagocytosis activity. Concerning the proinflammatory cytokine (TNF-α) transcription, this gene was upregulated after W-SN stimulation, while B-SN upregulated the IgG gene expression significantly. IgM was upregulated with any YBW-SN in PBLs after 24 h of stimulation. The in vivo study showed a notable B-SN immunostimulation in serum and an upregulation of TNF-α, IgM, and IgG mRNA transcription. Therefore, this study provides a new result of the yellow, white, and black sapote nanocapsules as a functional food for the poultry industry, highlighting the black sapote Diospyros digyna immunostimulant effect.

Development, characterization, and immunomodulation performance of spray-dried Moringa oleifera seed extract in Longfin yellowtail Seriola rivoliana.

Moringa oleifera is one of the most promising plants in aquaculture because it improves the health status, zootechnical parameters and resistance against diseases. This research evaluates the physicochemical, antioxidant values of spray-dried Moringa oleifera seed extract microencapsulates obtained at 140 and 180 °C with whey protein concentrate (WPC) and maltodextrin (MD) as wall materials in two different proportions: WPC 100% and WPC-MD (3:1). Also, immune response of peripheral blood leukocytes (PBL) of Longfin yellowtail Seriola rivoliana stimulated with spray-dried Moringa oleifera seed for 24 h was assessed. The physicochemical parameters show that the recovery yield for all the treatments was of 65% and microencapsulates demonstrated to be stable in the physicochemical tests with low solubilization times and protection against humidity. For WPC-MD (3:1)/140 °C, bioactive compound retention and antioxidant potential were higher than in other combinations. The immunological test show that any treatments was non-cytotoxic against peripheral blood leukocytes. WPC-MD (3:1)/140 °C treatment enhanced immune parameters as phagocytosis, respiratory burst, myeloperoxidase activities and nitric oxide production. Immune related genes as IL-1ß and TNF-α were up-regulated in those stimulated leukocytes with WPC-MD (3:1)/140 °C. The results suggest that this combination may be a good alternative for animal health as a medicinal and immunostimulant additive.

Determining the Photoelectrical Behavior and Photocatalytic Activity of an h-YMnO3 New Type of Obelisk-like Perovskite in the Degradation of Malachite Green Dye.

YMnO3 is a P-type semiconductor with a perovskite-type structure (ABO3). It presents two crystalline systems: rhombohedral and hexagonal, the latter being the most stable and studied. In the hexagonal system, Mn3+ ions are coordinated by five oxygen ions forming a trigonal bipyramid, and the Y3+ ions are coordinated by five oxygen ions. This arrangement favors its ferroelectric and ferromagnetic properties, which have been widely studied since 1963. However, applications based on their optical properties have yet to be explored. This work evaluates the photoelectric response and the photocatalytic activity of yttrium manganite in visible spectrum wavelengths. To conduct this, a rod-obelisk-shaped yttrium manganite with a reduced indirect bandgap value of 1.43 eV in its hexagonal phase was synthesized through the precipitation method. The synthesized yttrium manganite was elucidated by solid-state techniques, such as DRX, XPS, and UV-vis. It was non-toxic as shown by the 100% leukocyte viability of mice BALB/c.

Probiotic Debaryomyces hansenii CBS 8339 yeast enhanced immune responses in mice.

This study aimed to examine the effect of Debaryomyces hansenii CBS 8339 on innate immune responses in mice. Thirty BALB/c mice were randomly treated with phosphate buffered saline (PBS) (control) and two D. hansenii (Dh) doses: Dh 10ˆ6 CFU (colony forming units) and Dh 10ˆ8 CFU daily for 15 days. Spleen, blood, and gut samples were taken on days 7 and 15. Mouse splenocytes were isolated and challenged with Escherichia coli. Immunological assays and immune-related gene expressions were performed. Serum was obtained from blood for total IgA and IgG antibody titer determination. Gut samples were taken for yeast colonization assessment. Phagocytosis, respiratory burst activity, and nitric oxide production in mice were mainly enhanced (p < 0.05) upon 7 days of D. hansenii intake at a concentration of 10ˆ8 CFU before and after bacterial challenge. Moreover, oral D. hansenii in mice upregulated (p < 0.05) gene expression of pro-inflammatory cytokines (INF-γ, IL-6 and IL-1ß) before or after E. coli challenge on day 7 but downregulated (p < 0.05) on day 15. Furthermore, total serum IgG and IgA titers were higher (p < 0.05) in Dh 10ˆ8 CFU at days 7 and 15, and only at day 7, respectively, than that in the other dose and control groups. Finally, D. hansenii was detected in the gut of mice that received the treatments, suggesting that yeast survived gastrointestinal transit. Altogether, a short period (7 days) of D. hansenii CBS 8339 oral delivery improved immune innate response on mice.

Cylindropuntia cholla aqueous root rich in phytosterols enhanced immune response and antimicrobial activity in tilapia Oreochromis niloticus leukocytes.

"Cacti" are rich sources of phytochemicals with antioxidant activity, and their use is mainly focused on infusions in traditional medicine in Mexico. This study characterizes the chemical compounds found in Cylindropuntia cholla root by gas chromatography coupled to mass spectrometry (GC-MS) and determines the total content of polyphenols and flavonoids, as well as their antioxidant capacity. The immunostimulatory effect of aqueous C. cholla root extract (ACcr) was evaluated at concentrations of 50, 250, 500, and 1000 µg/mL in Tilapia peripheral blood leukocytes. The results obtained by the GC-MS analysis revealed the presence of phenolic acids, flavonoid and phytosterol derivatives as ß-sitosterol and campesterol. The determination of the total polyphenol and flavonoid contents indicated that ACcr is abundant in polyphenols, showing an anti-radical capacity of scavenging free radicals, such as those of hydroxyl and superoxide, as well as an increase in lipid peroxidation inhibition capacity. Stimulation of tilapia leukocytes resulted in the increase of its phagocytic activity, respiratory burst, nitric oxide production, and superoxide dismutase activity. Finally, the results obtained for the first time allowed establishing the chemical profile of ACcr and its antimicrobial activity against three important pathogenic bacteria. The potential of this root is indicated as an additive in formulating antioxidant and immunostimulant supplements for the aquaculture and pharmaceutical industry.

Yeast ß-Glucans as Fish Immunomodulators: A Review.

Administration of immunostimulants in fish is a preventive method to combat infections. A wide variety of these biological molecules exist, among which one of the yeast wall compounds stands out for its different biological activities. The ß-glucan that forms the structural part of yeast is capable of generating immune activity in fish by cell receptor recognition. The most frequently used ß-glucans for the study of mechanisms of action are those of commercial origin, with doses recommended by the manufacturer. Nevertheless, their immune activity is inefficient in some fish species, and increasing the dose may show adverse effects, including immunosuppression. Conversely, experimental ß-glucans from other yeast species show different activities, such as antibacterial, antioxidant, healing, and stress tolerance properties. Therefore, this review analyses the most recent scientific reports on the use of yeast ß-glucans in freshwater and marine fish.

Oral organic nanovaccines against bacterial and viral diseases.

Vaccines have saved millions of humans and animals from deadly diseases. Many vaccines are still under development to fight against lethal diseases. Indeed, subunit vaccines are a versatile approach with several advantageous attributes, but they lack strong immunogenicity. Nanotechnology is an avenue to vaccine development because nanoparticles may serve as nanocarriers and adjuvants, which are critical aspects for oral vaccines. This review provides an update of oral organic nanovaccines, describing suitable nanomaterials for oral vaccine design and recent (last five-year view) oral nanovaccine developments to fight against those principal pathogens causing human and animal diseases.

LptD-antigen system on gold nanoparticles: an innovative strategy in the nanovaccine development.

Nanovaccine development is a growing research field in which the development of new carriers and bioconjugation approaches is a priority. In this sense, this report describes for the first time, the development of a novel conjugate that consists of gold nanoparticles (AuNPs) obtained by a one-step synthesis using an immunogenic peptide of the Lipopolysaccharide-assembly protein LptD fromVibrio parahaemolyticusbacteria as a reducing and capping agent. The resultingLptD@AuNPscompounds were fully characterized and the results showed the high capacity of the peptide to form complexes and reduce gold ions. The reaction yield estimated was higher than 83% and the chemical integrity of the peptide on the NP surface revealed a tyrosine amino acid bonding on the AuNP surface. Furthermore, theLptD@AuNPsystem showed high colloidal stability in a wide pH range (3-11 pH values), where the hydrodynamic diameter and Zeta potential behavior were strongly influenced by the functional groups of the antigenic peptide. The cytotoxicity assays showed that the obtained system is safe for mouse leukocytes, while immunized mice withLptD@AuNPsproduced specific IgG antibodies. These encouraging results revealed the efficacy of some antigenic peptides as reducers and capping agents, in addition, opening the path to determine immunogenicity and immunoprotective efficacy of theLptD@AuNPsystem against the disease induced byVibrio parahaemolyticus.

ß-Glucan bioactivities from Cystobasidium benthicum in Totoaba macdonaldi thymus cells.

The overuse of antibiotics in aquaculture has led to serious concerns on microbial resistance and chemical residues. Novel sources of immunostimulants could help to solve this problem by stimulating the immune system to fight against pathogens. Therefore, this study aims to explore the immunostimulant potential of Cystobacidium benthicum-ß-glucans (Cb-ßG) using thymus cells from Totoaba (Totoaba macdonaldi), a recently farmed fish species in Mexico. The Cb-ßG was characterized and tested for its own antioxidant capacity. Then, a Cb-ßG safety experiment was carried out in thymus cells by evaluating the effects on immune parameters and immune-related genes. Cb-ßG had a molecular weight of 2.32 kDa, comprised of ß-1,3-1,6-glucan (53.4%), and showed strong antioxidant capacity compared to that of the positive antioxidant control. Cb-ßG had no toxic effects of thymus cells and enhanced phagocytic, respiratory burst, myeloperoxidase and superoxide activities. Additionally, immune-related genes implicated in recognition and effector functions of yeast glucans were up-regulated (Toll like receptor 2, C-type lectin family 17 member A, colony-stimulating factor 1 receptor 2, macrophage mannose receptor 1, and Interleukin-1ß). In conclusion, the glucan -characterized physically-chemically from the yeast C. benthicum (Cb-ßG)- was safe, had strong antioxidant capacity to scavenge free radicals, and stimulated immune parameters and immune-related gene expressions on thymus cells from Totoaba macdonaldi.

Probiotic and Immunomodulatory Activity of Marine Yeast Yarrowia lipolytica Strains and Response Against Vibrio parahaemolyticus in Fish.

Yarrowia lipolytica has been widely used in food industry but scarcely explored as probiotics. Thus, the aims of this study were to characterize in vitro the probiotic potential, antioxidant capacity, and antimicrobial activity of the marine yeast Y. lipolytica D-1 and N-6 strains. Dietary administration effect was evaluated in vivo on immunological parameters in serum, skin-mucus, intestine, and fish leukocytes upon challenge with Vibrio parahaemolyticus. The results showed that Y. lipolytica D-1 and N-6 strains grew with NaCl or bile salts but were sensitive to low pH. Each of the Y. lipolytica strains had a distinctive antioxidant capacity and fatty acid profile, but their antimicrobial activity was similar against fish bacterial pathogens. Fish (Lutjanus peru) supplemented with Y. lipolytica strains showed normal intestinal morphology, high IgM levels, and antioxidant enzyme activities. Immune-related genes were modulated in fish fed Y. lipolytica in a strain-dependent fashion. In addition, leucocytes from fish fed Y. lipolytica challenged with V. parahaemolyticus increased innate immune and antioxidant parameters compared with the control groups. In conclusion, the marine yeast Y. lipolytica D-1 and N-6 strains may be potential probiotics for fish by exerting free-radical scavenging, antimicrobial activity, and improved immune-protective responses against V. parahaemolyticus infection.

Yarrowia lipolytica N6-glucan protects goat leukocytes against Escherichia coli by enhancing phagocytosis and immune signaling pathway genes.

Immunostimulant and protective effects of Yarrowia lipolytica glucans against important pathogens, such as Escherichia coli, have not been investigated in goats and other ruminants. This study aimed to characterize Y. lipolytica N6-glucan (Yl-glucan) and its possible role in immunological signaling pathway activation and immunoprotection against E. coli in goat leukocytes. Characterization analyses showed that Y. lipolytica content had a mix of ß and α-D-glucans, molecular weight of 3301.53 kDa and low solubility after the heat treatment. The stimulation of goat leukocytes with Yl-glucan induced protection against E. coli challenge. Remarkably, Yl-glucan and E. coli interaction increased gene expression of dectin-1 and TLR-2 receptors, signaling pathway Syk/NFκB, and cytokines, such as TNF-α and IL-10. As a consequence of signaling activation, phagocytosis, and nitric oxide production enhanced killing of pathogens. Altogether, Y. lipolytica-glucan demonstrated to possess an immunoprotective potential against E. coli through innate immune response modulation in goat leukocytes.

Composition, antioxidant capacity, intestinal, and immunobiological effects of oregano (Lippia palmeri Watts) in goats: preliminary in vitro and in vivo studies.

This study investigated Lippia palmeri Watt (oregano) phytochemical compounds, their antioxidant capacity, and immunological effects on goat peripheral blood leukocytes (PBL), and on the presence of intermediate polar compounds in goat feces fed dietary oregano. The polar and nonpolar fractions of L. palmeri W. were characterized and phytochemical contents and antioxidant capacity were determined. Twelve healthy Anglo-Nubian goats were used for the in vivo trials, which were randomly assigned to control fed with basal diet, or oregano group fed with basal diet + 2.6% (DM basis) dried oregano leaves. Goat peripheral blood leukocytes (PBL) were isolated for the in vitro study, and PBL were stimulated with oregano extracts at 100 and 150 µg/mL after 24 h. For the in vivo trial, dietary oregano (2.6% on DM basis) was evaluated in the goats for 90 days. Relatively high abundance of carvacrol and thymol phytochemical compounds was found in oregano. The highest antioxidant capacity of oregano extracts was detected at 100 and 150 µg/mL. Nitric oxide production, phagocytosis, and superoxide dismutase activities increased (p < 0.05) in stimulated PBL with oregano extracts, whereas the pro-inflammatory (TNF-α and IL-1ß) transcription and antioxidant (CAT and GPX-4) genes downregulated. In the in vivo experiment, dietary oregano enabled the detection of nine compounds found in goat feces, from which caproic (C6) was in a high relative quantity compared with the control group. Oregano has phytochemical compounds with strong antioxidant capacity that protect cells against oxidative stress damage and could modulate immune response and feces composition in goats.

Probiotic and nutritional effects of Debaryomyces hansenii on animals.

Debaryomyces hansenii comes of age as a new potential probiotic for terrestrial and aquatic animals. Probiotic properties, including inmunostimulatory effects, gut microbiota modulation, enhanced cell proliferation and differentiation, and digestive function improvements have been related to the oral delivery of D. hansenii. Its functional compounds, such as cell wall components and polyamines, have been identified and implicated in its immunomodulatory activity. In addition, in vitro studies using immune cells have shown standpoints on the possible recognition, regulation, and effector immune mechanisms stimulated by this yeast. This review describes historic, cutting-edge research findings, implications, and perspectives on the use of D. hansenii as a promising probiotic for animals. KEY POINTS: • Debaryomyces hansenii has probiotic effects in terrestrial and aquatic animals. • Nutritional effects could be associated to probiotic D. hansenii strains. • ß-D-Glucan and polyamines from D. hansenii are associated to probiotic properties. • Adoption by the industry is expected in the next years.

Chemical and biological protection of food grade nisin through their partial intercalation in laminar hydroxide salts.

The use of antimicrobial agents within a matrix, specifically layered compounds, is of growing interest for reducing contamination due to food borne pathogens and deteriorative microorganisms, one of the main health problems worldwide. In this study, zinc layered hydroxide nanoparticles were synthesized as a matrix for nisin immobilization. Layered materials were characterized by X-ray diffraction, Fourier-Transform Infrared and Ultra Violet-Visible spectra, Scanning Electron Microscopy, and by Thermogravimetric Analysis. Thermal, chemical, enzymatic, and biological stabilities were assessed against Lactobacillus brevis as control strain. Free and immobilized nisin in solution were previously subjected to 25 and 121 °C, pH (7, 9) and inactivation with protease before antimicrobial tests that lasted 21 days. Immobilized nisin was found to maintain the activity levels after the protease action while the pure nisin solution lost its activity gradually. Furthermore, immobilized nisin treated at 121 °C and pH 7 showed higher activity than pure nisin after 21 days. These results may support that immobilizing nisin in zinc layered hydroxide salts promoted extended nisin inhibitory activity in solution after thermal, chemical or enzymatic treatments. This research provides an alternative to nisin application that could be used in processes where such operating conditions take place, as in dairy products.

ß-D-glucan from marine yeast Debaryomyces hansenii BCS004 enhanced intestinal health and glucan-expressed receptor genes in Pacific red snapper Lutjanus peru.

Previous studies have shown that marine yeast Debaryomyces hansenii BCS004 (also known as Dh004) has a potential biotechnological application. The aim of this study was to investigate the structural characterization, antioxidant properties and possible health inductor of dietary ß-D-glucan BCS004. In this study, a glucan BCS004 was obtained containing (1-6)-branched (1-3)-ß-D-glucan with low molecular weight and a high purity of 90 and 91.7% for one and 4 h, respectively. ß-D-glucan BCS004 showed higher antioxidant activity, including DPPH radical and superoxide anion scavenging, ß-carotene bleaching inhibition, and iron chelation activity. An in vitro study showed that ß-D-glucan BCS004 was safe for peripheral blood leukocytes inducing proliferative effects. Moreover, in an in vivo study using ß-D-glucan BCS004 no histopathological damages or intestinal inflammation were observed in fish. The gene expression analysis highlighted that dietary ß-D-glucan BCS004 could also up-regulate glucan and macrophage receptor genes in intestine, such as C-type lectin (CTL) and macrophage mannose receptors (MMR). Overall, the results demonstrated that ß-D-glucan from D. hansenii BCS004 could be an immunostimulant with antioxidant properties and beneficial effects on intestinal health in fish.

Assessment of chemical, biological and immunological properties of "Damiana de California" Turnera diffusa Willd extracts in Longfin yellowtail (Seriola rivoliana) leukocytes.

In Mexican herbal medicines or natural remedies, Turnera diffusa (Turneraceae) known as "Damiana de California", has ethnopharmacological relevance, including aphrodisiac, diuretic, and antimicrobial activities. To explore the immunological effect of infusion and methanolic extracts from Damiana de California, this study investigated its chemical, biological, antimicrobial and immunological properties in Longfin yellowtail Seriola rivoliana leukocytes. The analysis of chemical compounds revealed a considerable level of total phenolic and flavonoid contents in the infusion compared with methanolic extract. Furthermore, the antioxidant activity showed high hydroxyl radical scavenging activity in infusion extract compared with BHT positive control. Superoxide radical scavenging activity and ion chelation were higher in methanolic extract followed by infusion treatment. Interestingly, notable antimicrobial activity was observed in both extracts of T. diffusa against Vibrio parahaemolyticus. An in vitro study was performed using leukocytes of S. rivoliana treated with infusion or methanolic extracts at 12.5, 25 and 50 µg/mL for 24 h. Remarkably, infusion extract induced proliferation at any concentration but not the methanolic extract, which was diminished in a dose-dependent fashion. The immunostimulation study demonstrated that the phagocytosis activity increased in those leukocytes stimulated with methanolic extract but diminished the respiratory burst activity, in contrast to the activity observed in those leukocytes stimulated with infusion treatment. Finally, leukocytes incubated with the extracts and confronted with V.parahaemolyticus up-regulated the transcription of proinflammatory cytokine IL-1ß gene in a dose response relationship. These findings suggest that the infusion treatment has potential therapeutic properties, promoting the antioxidant capacity and enhancing immune parameters in Longfin yellowtail S. rivoliana.

First screening report of immune and protective effect of non-toxic Jatropha vernicosa stem bark against Vibrio parahaemolyticus in Longfin yellowtail Seriola rivoliana leukocytes.

In México, the infusion of Jatropha vernicosa stem bark has been used in folk medicine for many clinical situations, but no reports were available about this particular species of Jatropha in fish of mammals. In this first screening report, the phytochemical, antioxidant profile and antimicrobial properties of aqueous J. vernicosa stem bark extract were explored against Vibrio parahaemolyticus, an opportunist fish pathogen. To evaluate the cytotoxicity and immunological effect for the possible application of aqueous J. vernicosa stem bark in aquaculture, this study assessed it by using Longfin yellowtail Seriola rivoliana leukocytes. The results showed that phytochemical composition of the J. vernicosa extract was rich in phenol, flavonoid, saponin, and coumarin compounds. The antioxidant capacity of hydroxyl radical and superoxide anion scavenging activities, iron-chelation activity and ß-carotene bleaching coupled to linoleic acid showed that J. vernicosa extracts had a moderate antioxidant effect compared with synthetic antioxidants (BHT, BHA and EDTA). No adverse effects were observed on spleen leukocytes (viability > 98%). Interestingly, J. vernicosa stem bark extract has immunostimulant and antioxidant effects, increasing phagocytosis, respiratory burns activity, and nitric oxide production, as well as superoxide dismutase and catalase activities. Additionally, J. vernicosa extract increased pro-inflammatory cytokine IL-1ß and suppressed anti-inflammatory IL-10 gene expression upon stimuli and V. parahaemolyticus challenge. Finally, the data confirms that J. vernicosa stem bark extract is non-cytotoxic, rich in bioactive compounds with antioxidant effects, capable of enhancing the immune system in leukocytes and with great potential to fight against opportunistic diseases, such as vibriosis in fish.