Growth of brown trout in the wild predicted by embryo stress reaction in the laboratory.

Laboratory studies on embryos of salmonids, such as the brown trout (Salmo trutta), have been extensively used to study environmental stress and how responses vary within and between natural populations. These studies are based on the implicit assumption that early life-history traits are relevant for stress tolerance in the wild. Here we test this assumption by combining two data sets from studies on the same 60 families. These families had been experimentally produced from wild breeders to determine, in separate samples, (1) stress tolerances of singly kept embryos in the laboratory and (2) growth of juveniles during 6 months in the wild. We found that growth in the wild was well predicted by the larval size of their full sibs in the laboratory, especially if these siblings had been experimentally exposed to a pathogen. Exposure to the pathogen had not caused elevated mortality among the embryos but induced early hatching. The strength of this stress-induced change of life history was a significant predictor of juvenile growth in the wild: the stronger the response in the laboratory, the slower the growth in the wild. We conclude that embryo performance in controlled environments can be a useful predictor of juvenile performance in the wild.

Effect of Dietary Origanum onites on Growth, Non Specific Immunity and Resistance against Yersinia ruckeri of Rainbow Trout ( Oncorhynchus mykiss).

Natural substances has been identified to maintain health and improve growth performance in the aquaculture. The effect of Origanum onites on growth and immune response of rainbow trout was investigated. Experimental groups (A and B) of 70 fish were separated into 10 different treatments. A groups were fed with dietary administration of O. onites essential oil (0.5 mL kg-1 and 3.0 mL kg-1) and crude powder (1.0 g kg-1 and 10.0 g kg-1) for a period of 8 weeks. Other groups (B) were vaccinated against Yersinia ruckeri at the beginning of experiment and then fed the same diets described above. Results showed that feed conversion ratio in fish fed a combination of O. onites and vaccine was statistically better than the control. NBT-positive cells, phagocytic activity, serum lysozyme activity and immunoglobulin M level were stimulated in both non vaccinated and vaccinated fish (p<0.05). Cumulative mortality in fish fed O. onites was lower than controls following challenge with Y. ruckeri. No mortality was observed in vaccinated fish fed with 0.5 mL kg-1 of O. onites. These results indicated that dietary administration of O. onites could act as an enhanced non specific immune response, growth performance and resistance to Y. ruckeri.

Yersinia ruckeri infection activates local skin and gill B cell responses in rainbow trout.

Teleost fish lack organized structures in mucosal tissues such as those of mammals, but instead contain dispersed B and T cells with the capacity to respond to external stimuli. Nonetheless, there is still a great lack of knowledge regarding how B cells differentiate to plasmablasts/plasma cells in these mucosal surfaces. To contribute to a further understanding of the mechanisms through which fish mucosal B cells are activated, in the current study, we have studied the B cell responses in the skin and gills of rainbow trout (Oncorhynchus mykiss) exposed to Yersinia ruckeri. We have first analyzed the transcription levels of genes related to B cell function in both mucosal surfaces, and in spleen and kidney for comparative purposes. In a second experiment, we have evaluated how the infection affects the presence and size of B cells in both skin and gills, as well as the presence of plasmablasts secreting total or specific IgMs. The results obtained in both experiments support the local differentiation of B cells to plasmablasts/plasma cells in the skin and gills of rainbow trout in response to Y. ruckeri. Interestingly, these plasmablasts/plasma cells were shown to secrete specific IgMs as soon as 5 days after the exposure. These findings contribute to a further understanding of how B cells in the periphery respond to immune stimulation in teleost fish.

Effects of diets rich in Agaricus bisporus polysaccharides on the growth, antioxidant, immunity, and resistance to Yersinia ruckeri in channel catfish.

To promote the application of Agaricus bisporus polysaccharides (ABPs) in channel catfish (Ictalurus punctatus) culture, we evaluated the effects of ABPs on the growth, immunity, antioxidant, and antibacterial activity of channel catfish. When the amount of ABPs was 250 mg/kg, channel catfish's weight gain and specific growth rates increased significantly while the feed coefficient decreased. We also found that adding ABPs in the feed effectively increased the activities of ACP, MDA, T-SOD, AKP, T-AOC, GSH, and CAT enzymes and immune-related genes such as IL-1ß, Hsp70, and IgM in the head kidney of channel catfish. Besides, long-term addition will not cause pathological damage to the head kidney. When the amount of ABPs was over 125 mg/kg, the protection rate of channel catfish was more than 60%. According to the intestinal transcriptome analysis, the addition of ABPs promoted the expression of intestinal immunity genes and growth metabolism-related genes and enriched multiple related KEEG pathways. When challenged by Yersinia ruckeri infection, the immune response of channel catfish fed with ABPs was intenser and quicker. Additionally, the 16S rRNA gene sequencing analysis showed that the composition of the intestinal microbial community of channel catfish treated with ABPs significantly changed, and the abundance of microorganisms beneficial to channel catfish growth, such as Firmicutes and Bacteroidota increased. In conclusion, feeding channel catfish with ABPs promoted growth, enhanced immunity and antioxidant, and improved resistance to bacterial infections. Our current results might promote the use of ABPs in channel catfish and even other aquacultured fish species.

Elucidating bacterial adhesion to mucosal surface by an original AFM approach.

BACKGROUND: Fish skin represents an ancient vertebrate mucosal surface, sharing characteristics with other mucosal surfaces including those of the intestine. The skin mucosa is continuously exposed to microbes in the surrounding water and is therefore important in the first line defense against environmental pathogens by preventing bacteria from accessing the underlying surfaces. Understanding the microbe-host interactions at the fish skin mucosa is highly relevant in order to understand and control infection, commensalism, colonization, persistence, infection, and disease. Here we investigate the interactions between the pathogenic bacteria Aeromonas salmonicida (A. salmonicida) and Yersinia ruckeri (Y. ruckeri), respectively, and the skin mucosal surface of Atlantic salmon fry using AFM force spectroscopy. RESULTS: The results obtained revealed that when retracting probes functionalized with bacteria from surfaces coated with immobilized mucins, isolated from salmon mucosal surfaces, rupture events reflecting the disruption of adhesive interactions were observed, with rupture strengths centered around 200 pN. However, when retracting probes functionalized with bacteria from the intact mucosal surface of salmon fish fry no adhesive interactions could be detected. Furthermore, rheological measurements revealed a near fluid-like behavior for the fish fry skin mucus. Taken together, the experimental data indicate that the adhesion between the mucin molecules within the mucous layer may be significantly weaker than the interaction between the bacteria and the mucin molecules. The bacteria, immobilized on the AFM probe, do bind to individual mucins in the mucosal layer, but are released from the near fluid mucus with little resistance upon retraction of the AFM probe, to which they are immobilized. CONCLUSION: The data provided in the current paper reveal that A. salmonicida and Y. ruckeri do bind to the immobilized mucins. However, when retracting the bacteria from intact mucosal surfaces, no adhesive interactions are detected. These observations suggest a mechanism underlying the protective function of the mucosal surface based on the clearing of potential threats by adhering them to loosely attached mucus that is subsequently released from the fish skin.

Participation of two sRNA RyhB homologs from the fish pathogen Yersinia ruckeri in bacterial physiology.

Small noncoding RNAs (sRNAs) are important regulators of gene expression and physiology in bacteria. RyhB is an iron-responsive sRNA well characterized in Escherichia coli and conserved in other Enterobacteriaceae. In this study, we identified and characterized two RyhB homologs (named RyhB-1 and RyhB-2) in the fish pathogen Yersinia ruckeri. We found that, as in other Enterobacteriaceae, both RyhB-1 and RyhB-2 are induced under iron starvation, repressed by the Fur regulator, and depend on Hfq for stability. Despite these similarities in expression, the mutant strains of Y. ruckeri lacking RyhB-1 (ΔryhB-1) or RyhB-2 (ΔryhB-2) exhibited differential phenotypes. In comparison with the wild type, the ΔryhB-1 strain showed a hypermotile phenotype, reduced biofilm formation, increased replication rate, faster growth, and increased ATP levels in bacterial cultures. By contrast, in salmon cell cultures, the ΔryhB-1 strain exhibited an increased survival. On the other hand, the ΔryhB-2 strain was non-motile and showed augmented biofilm formation as compared to the wild type. The expression of a subset of RyhB conserved targets, selected from different bacterial species, was analyzed by quantitative RT-PCR in wild type, ΔryhB-1, ΔryhB-2, and ΔryhB-1 ΔryhB-2 strains cultured in iron-depleted media. RyhB-1 negatively affected the expression of most analyzed genes (sodB, acnA, sdhC, bfr, fliF, among others), whose functions are related to metabolism and motility, involving iron-containing proteins. Among the genes analyzed, only sdhC and bfr appeared as targets for RyhB-2. Taken together, these results indicate that Y. ruckeri RyhB homologs participate in the modulation of the bacterial physiology with non-redundant roles.

Effects of Greek juniper (Juniperus excelsa) extract on immune responses and disease resistance against Yersinia ruckeri in rainbow trout (Oncorhynchus mykiss).

This study investigated the effects of Greek juniper extract on immune responses of rainbow trout. In this experiment, 4 doses [0 (Control), 1 (J1), 4 (J4) and 8 (J8) mg/kg] of the extract were administered orally using an oral gavage twice a day for 14 days. Immune responses were measured on 7th and 14th days. On 14th day, Yersinia ruckeri was injected intraperitoneally to all fish of all groups. On 14th day, ORP in fish of J1 group increased significantly. Lysozyme activity (LA) was increased in J8 group on 7th day (p < .05). On 14th day, a significant decrease was determined in J1 and J4 treatments in LA. Myeloperoxidase activity was significantly decreased in all groups irrespective of sampling times (p < .05). Interleukin (IL)-1ß was significantly elevated in fish of J8 group on 7th day. IL-8 increased in fish of J8 and J4 groups on 7th day of the study. IL-12 gene expression was significantly up-regulated in J8 fish group on 7th day, and in J4 fish group on 14th day. Survival rate was higher in J8 treatment compared to the control and other treatments (p < .05). The results suggest that Juniperus excelsa provides protection against Y. ruckeri in rainbow trout.

Synbiotic feed supplementation significantly improves lipid utilization and shows discrete effects on disease resistance in rainbow trout (Oncorhynchus mykiss).

Enteric redmouth disease caused by the bacterial pathogen Yersinia ruckeri is the main reason for antimicrobial prescription, and a cause of substantial economic losses and decreased animal welfare in aquaculture. Given the importance of the intestinal microbiota in digestion and disease, our aim was to investigate whether synbiotic feed supplementation strategies could improve feed performance and disease resistance. Four experimental synbiotic feeds formulated with pre- and probiotics were tested against a commercially available probiotic control feed. Each experimental feed was evaluated for feed performance, effects on gross as well as intestinal morphometrics, and finally their effect on resistance against a waterborne experimental infection with Yersinia ruckeri serotype O1, biotype 2. While co-supplementing Pediococcus acidilactici with citrus flavonoids or bacterial paraprobiotics significantly improved utilization of feed lipid content relative to the control group, a decrease in lipid utilization was observed for feeds that combined P. acidilactici with yeast paraprobiotics. No significant improvements on disease resistance were observed. Still, synbiotic formulations including P. acidilactici led to reduced risks relative to that of the control group, while an increased relative risk was observed for a Bacillus-based formulation. In conclusion, two of the synbiotic supplements significantly improved lipid utilization and contributed to minor increases in disease resistance.

Immune gene expression and genome-wide association analysis in rainbow trout with different resistance to Yersinia ruckeri infection.

Selective breeding programmes involving marker assisted selection of innately pathogen resistant strains of rainbow trout rely on reliable controlled infection studies, extensive DNA typing of individual fish and recording of expression of relevant genes. We exposed juvenile rainbow trout (6 h bath to 2.6 × 105 CFU mL-1) to the fish pathogen Yersinia ruckeri serotype O1, biotype 2, eliciting Enteric Red Mouth Disease ERM, and followed the disease progression over 21 days. Cumulative mortality reached 42% at 12 days post challenge (dpc) after which no disease signs were recorded. All fish were sampled for DNA-typing (50 k SNP chip, Affymetrix®) throughout the course of infection when they showed clinical signs of disease (susceptible fish) or at day 21 when fish showed no clinical signs of disease (survivors - resistant fish). Genome-wide association analyses of 1027 trout applying single nucleotide polymorphisms (SNPs) as markers revealed an association between traits (susceptible/resistant) and certain regions of the trout genome. It was indicated that multiple genes are involved in rainbow trout resistance towards ERM whereby it is considered a polygenic trait. A corresponding trout group was kept as non-exposed controls and a comparative expression analysis of central innate and adaptive immune genes in gills, spleen and liver was performed for three fish groups: 1) moribund trout exhibiting clinical signs 7 dpc (CS), 2) exposed fish without clinical signs at the same sampling point (NCS) and 3) surviving fish at 21 dpc (survivors). Immune genes encoding inflammatory cytokines (IL-1ß, IL-2A, IL-6A, IL-8, IL-10A, IL-12, IL-17A/F2A, IL-17C1, IL-17C2, IL-22, IFNγ, TNFα), acute phase reactants (SAA, C3, cathelicidins, lysozyme) were expressed differently in CS and NCS fish. Correlation (negative or positive) between expression of genes and bacterial load suggested involvement of immune genes in protection. Down-regulation of adaptive immune genes including IgDm, IgDs, IgT and TCR-ß was seen primarily in CS and NCS fish whereas survivors showed up-regulation of effector molecule genes such as cathelicidins, complement and lysozyme suggesting their role in clearing the infection. In conclusion, SNP analyses indicated that ERM resistance in rainbow trout is a multi-locus trait. The gene expression in surviving fish suggested that several immune genes are associated with the trait conferring resistance.

Effects of Yersinia ruckeri invasion on the proteome of the Chinook salmon cell line CHSE-214.

Yersinia ruckeri is an important bacterial pathogen of fish, in particular salmonids, it has been associated with systemic infections worldwide and, like many enteric bacteria, it is a facultative intracellular pathogen. However, the effect of Y. ruckeri's interactions with the host at the cellular level have received little investigation. In the present study, a culture of Chinook Salmon Embryo (CHSE) cell line was exposed to Y. ruckeri. Afterwards, the proteins were investigated and identified by mass spectrometry and compared to the content of unexposed cultures. The results of this comparison showed that 4.7% of the identified proteins were found at significantly altered concentrations following infection. Interestingly, infection with Y. ruckeri was associated with significant changes in the concentration of surface adhesion proteins, including a significantly decreased presence of ß-integrins. These surface adhesion molecules are known to be the target for several adhesion molecules of Yersiniaceae. The concentration of several anti-apoptotic regulators (HSP90 and two DNAj molecules) appeared similarly downregulated. Taken together, these findings suggest that Y. ruckeri affects the proteome of infected cells in a notable manner and our results shed some light on the interaction between this important bacterial pathogen and its host.

Resistant and susceptible rainbow trout (Oncorhynchus mykiss) lines show distinctive immune response to Lactococcus garvieae.

Lactococcosis is one of the main bacterial diseases affecting rainbow trout (Oncorhynchus mykiss), with significant economic and sanitary repercussion. Vaccination and antibiotic treatments are commonly used to prevent and control the infection outbreaks; however, these strategies have some drawbacks including limited coverage, handling costs, induction of antibiotic resistance and chemical residues in the environment. Selective breeding programs represent a promising complementary approach for increasing fish disease resistance in commercial farms and some immunological parameters may be tentatively used as indirect indicators for this purpose. The present study investigated for the first time some innate and adaptive immune responses in two groups of rainbow trout derived from selected lines (susceptible and resistant) showing a different "in field" phenotypical resistance to Yersinia ruckeri, Flavobacterium branchiophilum, F. psychrophilum, and Ichthyophthirius multifiliis, after an immersion-dilution based exposure to Lactococcus garvieae carried out in controlled experimental conditions. Twenty-six resistant and twenty-six susceptible female rainbow trout (mean body weight 80 g, 9 months aged, F5 generation) were obtained from an intensive farm considered L. garvieae free and were exposed to the pathogen. Moreover, 10 resistant and 10 susceptible fish were used as uninfected controls. After 5 days, blood and tissue samples were collected for immunological analyses. A significantly higher serum and mucus lysozyme activity was recorded in resistant rainbow trout compared to susceptible fish (P ≤ 0.05), both before and after exposure to L. garvieae. Similarly, respiratory burst activity of head kidney leukocytes resulted more intense in resistant fish (P ≤ 0.05), suggesting that phagocytes could more quickly activate their microbicidal mechanisms to counteract the bacterial spread. Resistant group displayed also an up-regulation of immunoglobulins M (IgM), major histocompatibility complex II (MHC-II) and interleukin 8 (IL-8) gene expression (P ≤ 0.05) and a significantly higher blood lymphocytes count (P ≤ 0.05), highlighting their potential better ability to trigger the recruitment of defensive cells and the initiation of specific immune processes such as antigen presentation to CD4+ T lymphocytes and IgM synthesis. The results herein presented might be useful for the identification of immunological markers to be used as indirect indicators in rainbow trout selective breeding programs.

Effect of a multi-citrus extract-based feed additive on the survival of rainbow trout (Oncorhynchus mykiss) following challenge with Lactococcus garvieae.

Growing global concerns about antibiotic resistance have generated a considerable interest in the search for alternative environmental-friendly approaches. This study was aimed to assess the antimicrobial activity of a multi-citrus extract-based feed additive (Biocitro®) against some fish pathogens, as well as evaluate its capacity to protect rainbow trout (Oncorhynchus mykiss) to lactococcosis. A broth dilution method was used to determine the minimum inhibitory concentration (MIC) of Biocitro®, and the results showed a strong antibacterial activity against Aeromonas salmonicida, Lactococcus garvieae and Yersinia ruckeri with MIC values of 2.0 µg/mL. Afterwards, rainbow trout juveniles were fed a Biocitro®-enriched diet (750 mg/kg feed) at a daily rate of 1.5% body weight for 4 weeks, then they were challenged with L. garvieae by the cohabitation method. At the end of the experimental period, fish treated with Biocitro® showed significantly (P < 0.001) improved protection against L. garvieae compared to control fish. Although further studies are needed to understand how Biocitro® increases rainbow trout resistance to L. garvieae, this feed additive could be considered as a useful alternative to chemotherapeutic treatment in aquaculture.

Effective isolation of GALT cells: Insights into the intestine immune response of rainbow trout (Oncorhynchus mykiss) to different bacterin vaccine preparations.

The teleost gut is a multifunction complex structure that plays a pivotal immunological role in homeostasis and the maintenance of health, in addition to digestion of food and/or nutrient absorption. In vitro examination of the intestine leucocyte repertoire has the potential to aid our understanding of gut immune competence and allows a rapid screen of host-microorganism interactions in different immunological contexts. To explore this possibility, in the present study we investigated the response of isolated gut leucocytes to 4 bacterins of Aeromonas salmonicida, prepared from different strains, combinations and strains grown in different environments, in comparison to a Yersinia ruckeri bacterin for which a commercial/effective oral booster vaccine has been developed. To aid this study we also optimized further our method of GALT cell isolation from rainbow trout, so as to avoid mechanical clearance of the intestine contents. This drastically increased the cell yield from ~12 × 106 to ~210 × 106/fish with no change in the percent cell viability over time or presence of transcripts typical of the key leucocyte types needed for the study of immune modulation (i.e. T- and B-cells, dendritic cells and macrophages). A wide array of immune transcripts were modulated by the bacterins, demonstrating the diversity of GALT cell responses to bacterial stimulation. Indeed, the GALT leucocyte responses were sensitive enough to distinguish the different bacterial species, strains and membrane proteins, as seen by distinct kinetics of immune gene expression. However, the response of the GALT cells was often relatively slow and of a low magnitude compared to those of PBL. These results enhance our knowledge of the gut biocapacity and help validate the use of this model for screening of oral vaccine candidates.

Toll/IL-1 receptor-containing proteins STIR-1, STIR-2 and STIR-3 synergistically assist Yersinia ruckeri SC09 immune escape.

Immune escape is a common feature of bacteria, viruses, parasites and even cancer cells. Our earlier work on an integrative and conjugative element (ICEr2) of Yersinia ruckeri SC09 demonstrated contributory roles of stir-1, stir-2 and stir-3 in bacterial toxicity and ability to code for immune evasion. Here, we further examined the ability of stir-4 in ICE (r2) and its encoded STIR-4 protein to mediate immune evasion using comparative genomic analysis. Additionally, the mechanisms underlying the synergistic activities of STIR-1, STIR-2, STIR-3 and STIR-4 in immune evasion were examined. Our results showed that STIR-4 did not contribute to bacterial toxicity, either in vivo nor in vitro, or show the ability to assist in bacterial immune escape. STIR-1, STIR-2, and STIR-3 formed heterotrimers in bacteria while facilitating immune evasion, which we speculate may be essential to maintain their stability. This discovery also partially explains the previous finding that a single gene can mediate immune evasion. Our data provide further knowledge on the distribution of ICE (r2)-like elements in bacteria, validating the prevalence of large-scale gene transfer in pathogens and its potential for enhancing virulence levels. Further studies are necessary to establish the biological significance of the ICE (r2) component.

Advancing the fathead minnow (Pimephales promelas) as a model for immunotoxicity testing: Characterization of the renal transcriptome following Yersinia ruckeri infection.

Recent studies have utilized the fathead minnow (Pimephales promelas) to explore the immunotoxic effects associated with a variety of environmental contaminants in the absence of immunological stimuli. Though this approach allows for alterations in the resting immune system to be detected, previous evidence suggests that many immunotoxic effects may only manifest in the activated immune system. However, basic immune responses to pathogens have not been well described in this species. To expand the utility of the fathead minnow as a model for immunotoxicity testing, a more comprehensive understanding of the activated immune system is required. As such, the main goal of this study was to characterize the transcriptomic response to pathogen infection in the fathead minnow using RNA sequencing. To achieve this goal, female fathead minnows were intraperitoneally injected with either Hank's Balanced Salt Solution (sham-injected) or Yersinia ruckeri (pathogen-injected). Eight hours following injection, fish were sacrificed for the assessment of general morphological (i.e., mass, length, condition factor, hepatic index) and immunological (i.e., leukocyte counts, spleen index) endpoints. To assess the molecular immune response to Y. ruckeri, kidney tissue was collected for transcriptomic analysis. A comparison of sham- and pathogen-injected fish revealed that >1800 genes and >500 gene networks were differentially expressed.Gene networks associated with inflammation, innate immunity, complement, hemorrhaging and iron absorption are highlighted and their utility within the context of immunotoxicity is discussed. These data reveal pathogen-related molecular endpoints to improve data interpretation of future studies utilizing the fathead minnow as a model for immunotoxicity.

Effects of siRNA silencing on the susceptibility of the fish cell line CHSE-214 to Yersinia ruckeri.

Yersinia ruckeri is a facultative intracellular enterobacterium mostly known as the causative agent of enteric redmouth disease in salmonid fish. In the present study, we applied RNA inhibition to silence twenty pre-selected genes on the genome of a fish cell line (CHSE-214) followed by a gentamicin assay to quantify the effect of silencing on the cells' susceptibility to infection and found that silencing of 18 out of 20 genes significantly reduced the number of Y. ruckeri recovered. These findings improve our understanding of the infection process by Y. ruckeri and of the interactions between this bacterial pathogen and host cells.

Effects of dietary chitosan and nano-chitosan loaded clinoptilolite on growth and immune responses of rainbow trout (Oncorhynchus mykiss).

In this study, rainbow trout Oncorhynchus mykiss weighing 27.75 ± 0.34 g were orally subjected to eight experimental diets each in three replicates containing varying amounts of chitosan and nano-chitosan (0.05, 0.5 and 5 g kg-1) loaded in clinoptilolite (14.28 g kg-1) for 70 days; and the growth and immune responses were evaluated. Results showed that growth parameters in fish fed diets chit + clin2, chit + clin3, nchit + clin1, nchit + clin2 and nchit + clin3 were significantly higher than in fish fed the control diet. All feeds, except chit + clin3, and nchit + clin3, significantly increased the total protein level. Feeds containing chit + clin2, nchit + clin1, and nchit + clin2 significantly elevated serum lysozyme activity compared with the control group. All treatments, except chit + clin3, and nchit + clin3 exhibited higher serum immunoglobulin (Ig) level than control one. In contrast, diet nchit + clin1 significantly unregulated the expression of Ig M gene in fish head-kidney compared to other groups. Additionally, all feeds, except clinoptilolite, and nchit + clin3, significantly improved the serum complement activity. Diets chit + clin2, nchit + clin1, and nchit + clin2 also significantly elevated antibacterial activity against Yersinia ruckeri compared with the control diet. Expression of inducible nitric oxide synthase (iNOS) gene in fish fed diets clinoptilolite, chit + clin1, chit + clin3, nchit + clin1, nchit + clin2, and nchit + clin3 was significantly higher than the control diet. All diets, except clinoptilolite, increased IL-1ß gene expression compared to the control group. Present results suggest that diets supplemented with nchit + clin, especially at 0.05 g kg-1 nano-chitosan inclusion, could improve growth performance and immune parameters of rainbow trout.

Extract of common mallow (Malvae sylvestris) enhances growth, immunity, and resistance of rainbow trout (Oncorhynchus mykiss) fingerlings against Yersinia ruckeri infection.

The dietary effects of a native medicinal plant from Iran, common mallow (Malvae sylvestris), was evaluated on growth performance, innate immune parameters, mucosal immune parameters, and resistance of rainbow trout (Oncorhynchus mykiss) against Yersinia ruckeri. Therefore, 360 fish (initial weight 10.42 ± 0.09 g) were randomly distributed into 12 fiberglass tanks. Experimental diets supplemented with 0 (as control- C), 1% (M1), 3% (M2) and 5% (M3) levels of M. sylvestris flowers extract were fed to the fish based on 3% of body weight for 8 weeks. At the terminal sampling, growth performance, liver and digestive enzymes activities, blood and mucosal immune responses were determined. Results showed that M2 and M3 had greater final weight, weight gain, SGR, survival rate and lower FCR; higher levels of total protein, albumin, globulin, and lower cortisol levels in comparison to control; 5% extract also lowered cholesterol and glucose levels as well as Lactate Dehydrogenase (LDH) activity. We reported higher values of hematocrit, hemoglobin, Mean Corpuscular Hemoglobin (MCH), Mean Corpuscular Volume (MCV), White Blood Cell (WBC), Red Blood Cell (RBC) and lymphocytes for treated groups. Innate immune responses (Alternative complement activity (ACH50) in M2 and M3 group, total Immunoglobulin (Ig) and lysozyme in M3), mucosal immune parameters (ACH50, total Ig for M2 and M3 group and lysozyme in all treated groups) were enhanced. Activities of digestive enzymes (protease in all treated groups, amylase for 3 and 5%, while lipase only for 5%) and lower activity of liver ALT enzyme in individuals treated with highest dose was observed. Overall results indicated that the extract can positively affect growth performance and immune responses of rainbow trout.

Growth performance, haematological changes, immune response, antioxidant activity and disease resistance in rainbow trout (Oncorhynchus mykiss) fed diet supplemented with ellagic acid.

The present study was performed to investigate the effects of various levels of dietary ellagic acid (EA) on growth performance, haematological values, immune response, protection against Yersinia ruckeri infection, and oxidant/antioxidant status in rainbow trout, Oncorhynchus mykiss. Fish were fed with the control diet and three different experimental diets containing three graded levels of EA (50, 100 and 200 mg kg-1 diet) for 8 weeks. At the end of the experiment, the growth performance [weight gain (WG), specific growth rate (SGR) and feed conversion ratio (FCR)], haematological values [the red blood cell (RBC) count, haemoglobin (Hb) concentration, haematocrit (Ht) level and erythrocyte indices: mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH) and mean corpuscular haemoglobin concentration (MCHC)], immune response [white blood cell (WBC) count, oxidative radical production (nitroblue tetrazolium (NBT) assay), phagocytic activity (PA) and phagocytic index (PI), total protein (TP) and immunoglobulin M (IgM) levels, serum bactericidal activity (BA), lysozyme (LYZ) and myeloperoxidase (MPO) activities] and oxidant/antioxidant status [tissue malondialdehyde (MDA) levels and superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) activities] were analysed. In addition, fish were challenged by Y. ruckeri and survival rate was recorded for 14 days. In the groups fed with EA the growth parameters such as WG, SGR, and FCR did not vary significantly. The RBC count, Hb concentration, and Ht level increased in the groups fed with EA when compared with the control group. However, there were no significant differences in the MCV, MCH and MCHC values among the groups. The results demonstrated enhancement in all the immunological parameters in the groups fed with EA compared to the control group. The results obtained from challenge with Y. ruckeri revealed reduction in the mortalities in the groups fed with EA. The dietary EA stimulated the SOD, CAT and GSH-Px activities in liver, head kidney and spleen as compared to the control group; however, a reverse trend was observed in the MDA levels of tissues. The present study suggest that EA can effectively enhance the haematological values, immune response, antioxidant capacity, and disease resistance in rainbow trout.