Jorge Galindo-Villegas | Nord University (original) (raw)

PAPERS by Jorge Galindo-Villegas

Aquaculture, Jun 1, 2024

This study aimed to deepen our understanding of the reproductive biology of male spotted wolffish... more This study aimed to deepen our understanding of the reproductive biology of male spotted wolffish (Anarhichas minor) using two different experimental approaches involving juvenile and mature broodstock fish. The first approach consisted of a detailed histological examination of the testes to identify the onset of gonadal maturation and characterize the spermatogenic stages in two- and three-year-old juvenile specimens. Light microscopy analysis revealed clear differences between the age groups. Two-year-old fish displayed well-defined interstitial tissue, Sertoli cells, and cysts housing spermatogonia stem cells in which meiosis had not yet begun. In contrast, three-year-old fish exhibited cysts containing spermatocytes, spermatids, and abundant spermatozoa, indicating the initiation of the spermatogenic cycle, albeit with asynchronous puberty. Histochemical staining revealed a significant presence of smooth myoid cells in the interstitial tissue of sexually mature fish. In contrast, electron microscopy further revealed synaptonemal complexes indicating the onset of meiosis and centriolar structures that gave rise to flagella. The second approach focused on optimizing semen freezing and cryopreservation procedures in mature broodstock individuals over the age of 10 years. Seven freezing extenders (KT, TS-2, OP, MT, MH, HBSS, or SR), with seawater (SW) as a control, were assessed along with two cryoprotectants dimethylsulfoxide (DMSO) or methanol to evaluate their impact on pre- and post-thaw semen quality. Results showed that the MT and HBSS extenders were superior in total sperm kinetics at 1:3 dilution and that DMSO showed optimal results in sperm motility and velocity variants. Moreover, the MT and HBSS groups demonstrated consistent sperm viability after cryopreservation, with values similar to fresh samples. Based on the viability results of the SYBR-green-14/PI assay comparing fresh and cryopreserved sperm using MT and HBSS, the MT extender emerged as the most effective freezing medium for cryopreservation of spotted wolffish broodstock sperm. In conclusion, this study provides a comprehensive understanding of the reproductive dynamics of male spotted wolffish, offering valuable insights for both scientific research and aquaculture management.

Aquaculture, Feb 1, 2024

Disease prevention is pivotal in aquaculture, and while vaccines offer protective immunity, chall... more Disease prevention is pivotal in aquaculture, and while vaccines offer protective immunity, challenges such as cost and low efficacy persist. The present study investigated the potential of plant-derived compounds, known as phytogenics, to bolster the effectiveness of vaccines against vibriosis in European seabass. Two phytogenic blends, namely PHYTO1 (terpenes) and PHYTO2 (terpenes and flavonoids) were supplemented to a commercial diet to obtain three experimental diets: a non-supplemented control diet, PHYTO1 (a 200-ppm blend of garlic and Lamiaceae oils with 87.5 mg kg − 1 terpenes), and PHYTO2 (a 1000 ppm blend containing citrus fruits, Asteraceae and Lamiaceae oils with 57 mg kg − 1 terpenes and 55 mg kg − 1 flavonoids). Following vaccination by bath immersion, juvenile European seabass were divided into groups and fed one of the three diets for 30 days. After this feeding period, fish were anesthetized and boosted with a single dose of vaccine through intraperitoneal injection. They continued to be fed their respective diets for another 30 days. At day 60, after the priming vaccination, fish were challenged with Vibrio anguillarum via intraperitoneal injection. Various parameters were measured at different time points post each vaccination, including total weight, circulating plasma cortisol and glucose levels, serum immunoglobulin M (IgM) titers, antioxidant power of leucocytes, and the expression of several antioxidant and immune-related genes. The results showed that fish fed with phytogenic supplements did not differ in weight compared to the control group. However, they exhibited lower plasma cortisol and glucose levels, increased IgM titers, and enhanced antioxidant protection and antioxidant power of head kidney leucocytes. In addition, phytogenics upregulated several immune-related genes in the gills and head kidney immediately after each vaccination. Notably, PHYTO2, enriched with flavonoids and terpenes, exhibited an even more pronounced positive effect on boosted fish by reducing vaccine-associated stress while improving antioxidant protection and modulating the vaccine-induced immune response. This synergistic effect of vaccination combined with phytogenics introduces new pathways for enhancing fish health in aquaculture.

Aquaculture, Jan 31, 2024

The lumpfish, Cyclopterus lumpus, holds significant promise as a candidate for large-scale aquacu... more The lumpfish, Cyclopterus lumpus, holds significant promise as a candidate for large-scale aquaculture production, particularly in its role as a cleaner fish used to manage sea lice infestations in Atlantic salmon Salmo salar farming. Melanomacrophage centers (MMCs) represent polymorphic structures present in the hemolymphopoietic organs of various vertebrates, serving as a widely applicable histological indicator of the fish immune and health status. This study aims to investigate the histochemical characteristics of MMCs within lumpfish livers and to compare MMC density between hatchery-produced (farmed) and wild individuals. Liver samples were collected from 34 lumpfish and subjected to a range of staining techniques, including haematoxylin-eosin, Azan-Mallory's trichrome, Masson-Fontana, Perls-Van Geison, Mallory's hemofuscin, immunohistochemical detection of cytochrome P450 monooxygenase 1 A (CYP1A), and the terminal deoxynucleotidyl transferase-mediated d'UTP nick-end labelling (TUNEL) method. Hepatocytes from hatchery-produced males exhibited notably high lipid content. Additionally, cells showing positive staining with Masson-Fontana, likely associated with the monocyte/macrophage lineage, were identified. Furthermore, small MMCs containing melanin, lipofuscinceroids, and ferric ions were detected. While the density of single monocytes/macrophages was markedly higher in hatchery-produced males, no significant discrepancies in MMCs density were observed between wild and hatchery-produced fish, or between males and females of the same origin. The study also revealed the presence of necrotic foci, characterized by hypertrophic hepatocytes positive for both TUNEL and CYP1A staining. These hypertrophic hepatocytes displayed large lipid droplets and pycnotic nuclei, with hatcheryproduced males showing a higher numerical density of such foci. In contrast to findings in other fish species, the study found that MMCs did not appear to serve as reliable markers of health status in lumpfish. This conclusion was reached as MMCs density did not exhibit a correlation with necrotic foci or hepatocyte lipid content.

Aquaculture, 2024

BMC Bioinformatics, May 19, 2023

Background: Circular RNAs (circRNAs) are covalently closed-loop RNAs with critical regulatory rol... more Background: Circular RNAs (circRNAs) are covalently closed-loop RNAs with critical regulatory roles in cells. Tens of thousands of circRNAs have been unveiled due to the recent advances in high throughput RNA sequencing technologies and bioinformatic tools development. At the same time, polymerase chain reaction (PCR) cross-validation for circRNAs predicted by bioinformatic tools remains an essential part of any circRNA study before publication. Results: Here, we present the CircPrime web-based platform, providing a user-friendly solution for DNA primer design and thermocycling conditions for circRNA identification with routine PCR methods. Conclusions: User-friendly CircPrime web platform (http:// circp rime. elgene. net/) works with outputs of the most popular bioinformatic predictors of circRNAs to design specific circular RNA primers. CircPrime works with circRNA coordinates and any reference genome from the National Center for Biotechnology Information database).

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Fish & Shellfish Immunology, Apr 1, 2022

Scientific Reports

In the Introduction, "In addition to traditional host models, zebrafish (Danio rerio), an extreme... more In the Introduction, "In addition to traditional host models, zebrafish (Danio rerio), an extremely versatile and attractive organism, have gained prominence in research https:// doi. org/ 10. 3389/ fphar. 2020. 00680. " now reads: "In addition to traditional host models, zebrafish (Danio rerio), an extremely versatile and attractive organism, have gained prominence in research 20. " The reference has now been added to the Reference list as Reference 20. As a result, the subsequent references have been renumbered. The original Article has been corrected.

BackgroundCircular RNAs (circRNAs) are covalently closed-loop RNAs with critical regulatory roles... more BackgroundCircular RNAs (circRNAs) are covalently closed-loop RNAs with critical regulatory roles in cells. The tenth of thousands of circRNAs have been unveiled due to the recent advances in high throughput RNA sequencing technologies and bioinformatic tools development. At the same time, polymerase chain reaction (PCR) cross-validation for circRNAs predicted by bioinformatic tools remains an essential part of any circRNA study before publication.ResultsHere, we present the CircPrime web-based platform, providing a user-friendly solution for DNA primer design and thermocycling conditions for circRNA identification with routine PCR methods.ConclusionsUser-friendly CircPrime web platform (http://circprime.elgene.net/) works with outputs of the most popular bioinformatic predictors of circRNAs to design specific circular RNA primers. CircPrime works with circRNA coordinates and any reference genome from the National Center for Biotechnology Information database (NCBI).

HAL (Le Centre pour la Communication Scientifique Directe), Oct 5, 2010

SummaryEstablishing new experimental animal models to assess the safety and immune response to th... more SummaryEstablishing new experimental animal models to assess the safety and immune response to the antigen used in the development of COVID-19 vaccine is an imperative issue. Based on the advantages of using zebrafish as a model in research, herein we suggest doing this to test the safety of the putative vaccine candidates and to study immune response against the virus. We produced a recombinant N-terminal fraction of the Spike SARS-CoV-2 protein and injected it into adult female zebrafish. The specimens generated humoral immunity and passed the antibodies to the eggs. However, they presented adverse reactions and inflammatory responses similar to severe cases of human COVID-19. The analysis of the structure and function of zebrafish and human Angiotensin-converting enzyme 2, the main human receptor for virus infection, presented remarkable sequence similarities. Moreover, bioinformatic analysis predicted protein-protein interaction of the Spike SARS-CoV-2 fragment and the Toll-like...

Aquaculture, 2023

The Journal of Immunology

Vertebrate olfactory sensory neurons (OSNs) are located in direct contact with the external envir... more Vertebrate olfactory sensory neurons (OSNs) are located in direct contact with the external environment and therefore they directly exposed to microorganisms such as viruses. Teleost fish have four types of OSNs, including crypt neurons, a cell type with enigmatic function. Crypt neurons express tyrosine receptor kinase A (TrkA)-like molecule and ORA4. In this study, we hypothesized that TrkA expressing crypt neurons are fast sensors of viruses in the olfactory mucosa of fish serving as an ultra-rapid mechanism to elicit local antiviral immune responses. Our results show that, within minutes, nasal delivery of rhadboviruses induces caspase dependent apoptosis in crypt neurons along with neuronal activation in rainbow trout olfactory organ (OO) and olfactory bulb (OB). Induction of chemokine CCL19 and prostaglandin synthase expression was detected 15 minutes after nasal delivery of virus in OO; whereas, in parallel, significant downregulation of CCL19 expression was measured in the O...

This work presents the first study showing how photobiomodulation (PBM) significantly increases c... more This work presents the first study showing how photobiomodulation (PBM) significantly increases cellular and tissue repair and elucidating the role of PBM with low-level laser as a possible new therapy in pathologies in COVID-19-associated cytokine storm syndrome from a zebrafish model. Our results demonstrate new strategies for treating SARS-COV-2 using PBM to modulate the expression of the genes and metabolites involved in inflammatory processes. These metabolic alterations show that the r-Spike led to disturbance in the energetic and inflammatory system, corroborating with the severe clinical conditions of human patients. Furthermore, PBM decreased the gene expression levels of pro-inflammatory cytokines such as il1b, il6, tnfa, and nfkbiab, and of factors involved in oxidative stress (romo1) and energy metabolism (slc2a1a, coa1), in various tissues, promoting an anti-inflammatory response. In summary, our study suggests that PBM may have a positive role in treating cytokine stor...

Despite all efforts to combat the pandemic of COVID-19, we are still living with high numbers of ... more Despite all efforts to combat the pandemic of COVID-19, we are still living with high numbers of infected persons, an overburdened health care system, and the lack of an effective and definitive treatment. Understanding the pathophysiology of the disease is crucial for the development of new technologies and therapies for the best clinical management of patients. Since the manipulation of the whole virus requires a structure with an adequate level of biosafety, the development of alternative technologies, such as the synthesis of peptides from viral proteins, is a possible solution to circumvent this problem. In addition, the use and validation of animal models is of extreme importance to screen new drugs and to compress the organism's response to the disease. Peptides derived from recombinant S protein from SARS-CoV-2 were synthesized and validated by in silico, in vitro and in vivo methodologies. Macrophages and neutrophils were challenged with the peptides and the production ...

Aquaculture, Jun 1, 2024

Aquaculture, Feb 1, 2024

Aquaculture, Jan 31, 2024

Aquaculture, 2024

BMC Bioinformatics, May 19, 2023

Fish & Shellfish Immunology, Apr 1, 2022

Scientific Reports

HAL (Le Centre pour la Communication Scientifique Directe), Oct 5, 2010

Aquaculture, 2023

The Journal of Immunology

Ediotrial on the Research Topic Overcoming challenges in mucosal immunity: 2022 The mucous-produc... more Ediotrial on the Research Topic Overcoming challenges in mucosal immunity: 2022 The mucous-producing membranes house the highest concentration of lymphoid tissue in the vertebrate body due to their extensive interaction with the external environment. Although considerable research effort has been devoted to their study, certain crucial aspects necessitate innovative investigative approaches. Preserving mucosal integrity and regulating immune responses, both locally and systemically, are critical for maintaining homeostasis. This Research Topic is dedicated to addressing fundamental questions pertaining to mucosal immunity and its associated challenges. These challenges encompass aspects such as the role of the epithelium in governing both mucosal and systemic immune responses. Contributions to this Research Topic shed light on various topics, for example, the preservation of mucosal barrier integrity, the influence of nutritional elements, namely amino acids, short-chain fatty acid (SCFA) metabolism, and the unfolded protein response (UPR) within intestinal epithelial cells (IEC). Additionally, this Research Topic delves into immunological senescence and its relevance to mucosal immunity, particularly in the context of aging populations. The study by Ouyang et al. focused on patients with HIV infection undergoing antiretroviral therapy. Despite the reduction in morbidity and mortality rates in treated patients, their immune systems do not appear to be fully restored, leading to chronic inflammatory infections and damage to the intestinal epithelial barrier. This review explored the feasibility of using liquid blood biomarkers as indicators of intestinal barrier damage in HIV patients, as invasive intestinal biopsies are impractical for this purpose. Damage to the intestinal mucosal barrier allows microbial products to translocate into the systemic circulation, contributing to chronic inflammation and elevating the risk of non-AIDS-related health issues. Therefore, the study provided insight into plasma biomarkers of intestinal permeability and envisions their future application in therapeutic strategies to address leaky gut in individuals living with HIV. Martıń-Adrados et al. investigated the unfolded protein response (UPR) in intestinal epithelial cells, a response associated with inflammatory bowel disease. Inflammatory triggers prompt intestinal epithelial cells to produce excessive amounts of proteins, rendering them susceptible to the UPR, which, in turn, leads to severe inflammatory conditions. In this study, the researchers identified several UPR-regulated genes in IECs, including the enzyme 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2), which Frontiers in Immunology frontiersin.org 01

Frontiers Media SA., 2022

05.- Editorial: 10.- Diet, Immunity, and Microbiota Interactions: An Integrative Analysis of the ... more 05.- Editorial: 10.- Diet, Immunity, and Microbiota Interactions: An Integrative Analysis of the Intestine Transcriptional Response and Microbiota Modulation in Gilthead Seabream (Sparus aurata) Fed an Essential Oils-Based Functional Diet 29.- Carvacrol, Thymol, and Garlic Essential Oil Promote Skin Innate Immunity in Gilthead Seabream (Sparus aurata) Through the Multifactorial Modulation of the Secretory Pathway and Enhancement of Mucus Protective Capacity 48.- Methionine and Tryptophan Play Different Modulatory Roles in the European Seabass (Dicentrarchus labrax) Innate Immune Response and Apoptosis Signaling—An In Vitro Study 65.- Dietary Phytogenics and Galactomannan Oligosaccharides in Low Fish Meal and Fish Oil-Based Diets for European Sea Bass (Dicentrarchus labrax) Juveniles: Effects on Gill Structure and Health and Implications on Oxidative Stress Status 80.- Oral Administration of Lactococcus lactis Producing Interferon Type II, Enhances the Immune Response Against Bacterial Pathogens in Rainbow Trout. 92.- GAS1: A New β-Glucan Immunostimulant Candidate to Increase Rainbow Trout (Oncorhynchus mykiss) Resistance to Bacterial Infections With Aeromonas salmonicida achromogenes 108.- Lactococcus lactis Expressing Type I Interferon From Atlantic Salmon Enhances the Innate Antiviral Immune Response In Vivo and In Vitro 124.- The Spleen as a Target to Characterize Immunomodulatory Effects of Down-Stream Processed Cyberlindnera jadinii Yeasts in Atlantic Salmon Exposed to a Dietary Soybean Meal Challenge 136.- Vaccination of Gilthead Seabream After Continuous Xenoestrogen Oral Exposure Enhances the Gut Endobolome and Immune Status via GPER1 153.- Essential Oils Improve the Survival of Gnotobiotic Brine Shrimp (Artemia franciscana) Challenged With Vibrio campbellii 171.- Sinomenine Hydrochloride Ameliorates Fish Foodborne Enteritis via α7nAchR-Mediated Anti-Inflammatory Effect Whilst Altering Microbiota Composition.

Larval Fish Production,, 2018

Executive Summary Background Factors such as feed composition and water temperature during the ea... more Executive Summary Background Factors such as feed composition and water temperature during the early larval stages, as well as the fish genetic line, have major impact on fish larvae and juveniles quality. Some of these effects are only easily noticeable later in the juvenile stage. Therefore it is important to develop appropriate criteria / indices for the description and prediction of fish quality. Larval and juvenile external appearance, histology and analysis of the skeleton by X-ray have been widely used as such quality indices. Sorting of fish with severe anomalies has been successfully applied at the hatchery level, but only when juveniles reach a few grams. Still this causes significant economic losses, and not all fish with anomalies are identified. Moreover, external morphology is only part of fish quality. Other aspects such as gamete quality, nutritional condition, growth potential at later stages, immune competence and gut microbiology are paramount in defining quality of juvenile and adult fish. Still, little progress has been so far accomplished on identification and validation of early predictors/descriptors of fish quality. Such indices should ideally estimate quality as early as possible in larvae life, with high accuracy and precision, with fast and simple methodologies, and at low cost.

Principal Findings and Conclusions Currently, available precise indices are few and limited to the prediction of fish quality in terms of some aspects of morphology alone. Existing literature clearly suggests that the development of indices for the prediction of fish quality in terms of normal larval development, growth potential, immune and nutritional condition will soon be possible, by exploiting the rapidly growing knowledge on different aspects of fish larvae development. Still, this quality assessment needs to be relatively simple and at a reasonable cost so it can be routinely applied in fish farms. Probably such a set of practical quality indices with predicting All rights reserved. power should include a combination of morphology characteristics and expression of selected genes.

Scientific Significance This review provides a framework for the future development of a science-based set of practical quality indices that can predict larval and juvenile quality. Promising candidates include: morphology characteristics that establish a clear correlation of larval/early juvenile and market-size fish external appearance; expression of genes for bone development, i.e., TRAP, osteocalcin and alkaline phosphatase; nutrition condition descriptors such as gut histology, as well as gene expression genes of genes coding for digestive enzymes; indices of growth potential, such as muscle fibre number and size, and expression of genes related to muscle growth, e.g, MRFs, GH, IGFs, MAPK, and growth-related QTLs; gamete quality descriptors such as sperm motility/viability/ATP levels, early egg cleavage pattern, blastomere morphology and maternal gene transcripts; descriptors of immune condition such as immunoglobulin M and IgT/Z levels for systemic and mucosal responses respectively, and disease challenge tests.

Practical Application It is expected that indices for the prediction of fish quality in terms of high survival, normal appearance, growth potential, the immune and nutritional condition will be available in the future. However, this will require a concerted R&D effort from researchers and producers, focusing on the validation of such indices at the commercial scale.

Biology of European Sea Bass (Immune system) , 2015

A potential concern of fish culturists is to reduce any economic loss. In European sea bass (Dice... more A potential concern of fish culturists is to reduce any economic loss. In European sea bass (Dicentrarchus labrax L.) aquaculture rearing facilities, mostly at those located in southern European countries, infectious diseases are a leading cause of mortality. Bacterial and viral diseases of sea bass (the most recurrent, namely vibriosis, pasteurellosis, and viral erythrocytic infection) are now a major challenge for biological studies on this species and further intensive fish production. To fight against diseases, preventive strategies should be implemented in every single farm. Best management
practices, which may include improved sanitary conditions, clean water supplies, and vector control are by far the most effective measures to reduce the incidence of infectious diseases. However, the development of specific target strategies like vaccines, disease-resistant breeds and therapeutics are much more important in achieving protection, but this requires a sufficient understanding of the fish immune system. The sea bass belongs to the Teleostei which is one of three infraclasses in the class Actinopterygii. This diverse group, which arose later in evolution during the Triassic period, roughly makes up half of the extant vertebrate species and occupies a key evolutionary position in the development of immune system evolution. Teleostei is the earliest class of vertebrates sharing similar immune system organization with other vertebrates, and thus, possesses the elements of both innate and adaptive immunity. Therefore, the sea bass has become very attractive as a fish model in regards to developmental, anatomical and functional comparative studies of the immune system. So far, it has been recognized that the fish immune system is characterized by a multi-layered organization that provides immunity to infectious organisms and each layer can be considered to have increasing complexity. In this way the central challenge of fish immunology, which includes the sea bass model, which is now to fully characterize the occurring immune responses in as many tissues as possible, and determine the functionality of each layer. Foundations of present knowledge on innate and adaptive immunity in sea bass have been established during the two previous decades. Remarkable milestones were provided during the early 90s by several workers. Such studies included the isolation of sea bass innate immune cells and the description of the ultrastructural characterization of the main features displayed by each cell group. And, extensive reports on the role of macrophages, granulocytes, lymphocytes, plasmocytes and fibroblast-like cells after myxosporean parasites invasion on host testes. Some years later, Mulero et al. conducted an interesting experiment in which functional properties of host leucocytes were shown. The authors of this paper characterized the changes followed by effector sea bass leucocytes incubated in vitro with HeLa or B16 melanoma cell targets. They observed that effector cells established spot contacts with targets and characterized morphological cell changes like smooth surfaces, the appearance of cell processes or surface blebs, suggesting that changes in targets are similar to those described to
be mediated by mammalian cytotoxic cells. On the adaptive capacity of sea bass immune system, two findings made great improvements in the study of fish immunology. These contributions were provided by Scapigliati et al., who produced and characterized specific markers for T and B lymphocytes respectively—the monoclonal antibody DLT15 against thymocytes and the DLIg3 antibody directed against the immunoglobulin (Ig)-bearing B cells. In this regard, Picchietti and colleagues, applying both antibodies, showed through functional analyses that gut-associated lymphoid tissue (GALT) developed earlier than other lymphoid compartments. This was a highly remarkable finding which provides evidence of the abundance and predominance of T cells in the gut. Together, these studies along with several more conducted in carp brought to light the importance of the gut as an immunocompetent organ in fish defense against invader bacteria. Recent studies have complemented the basic sea bass immune research with steadily increasing work conducted mainly at the cellular and molecular level. However, the former global vision on the study of vertebrate immunology has just recently changed and new players from the environment, the microbes, have begun to be recognized as a vital key modulator of the immune response. The sea bass model of immunity is not the exception and new efforts have been directed towards discovering the mechanisms of microbial pathogenesis and fish-microbe symbiosis through the characterization of gut microbiota in conventional or gnotobiotically raised fish. Indeed, the diverse and unique immune tools available for European sea bass, its taxonomic position, and commercial relevance have turned this species into an excellent model to study developmental and comparative immunological processes, including basic and functional aspects. Therefore, in this chapter, we briefly summarize and update the emergence, phylogenetic distribution, and functionality of D. labrax innate and adaptive immunity, as well as the most relevant interactions between them towards host defense against pathogens.

Marine aquaculture has grown into a very significant industry in the world. Particularly, in the... more Marine aquaculture has grown into a very significant industry in the world. Particularly, in the developed countries, marine fish farming is intensively conducted at super-high population densities. Thus, infectious diseases pose a constant threat to successful fish husbandry. Despite the antibiotics provide a useful means in the control of many bacterial diseases, the associated antibiotic resistance produced and the recurrent outbreaks require further costly treatments. Immunostimulants administered through the diet have been perceived as a potential and important role player in aquaculture. Here, the efficiencies and strategic use of immunostimulation methods are presented, and the dose-effects between species analyzed. In addition, the effects reported by several authors in the use of nutritional and non-nutritional factors as immunostimulants in marine fish are discussed. While the recent findings on the wide range of humoral and cellular innate immune responses affected by immunostimulation are summarized. The actual knowledge of potential immunostimulants is still obscure in several aspects, including the pathways and mechanisms. Nevertheless, immunostimulants as diet supplements, particularly those of non-nutritional origin have a high potential to induce disease resistance enhancement in marine fish. Although, the particular methods immune responses selected for their evaluation should be standardized among researchers. This will improve the dissection of the disease resistance mechanisms making easier the development of these therapies until the production scale and not just leave them as mare laboratory trials.

The variated environmental stimuli interacting at the mucosal surfaces on vertebrates are critica... more The variated environmental stimuli interacting at the mucosal surfaces on vertebrates are critical threats affecting their health status. However, the long-term effects and the possible heritability of the same have not been approached. This study sought to investigate whether a lack of bacterial priming in newly hatched zebrafish (ZF) embryos affects development, survival, life span, and immune competence. To address this hypothesis, we raised wild-type ZF in either germ-free (GF) or conventional (CONR) conditions until 7 days post fertilization (dpf). After that, GF and CONR larvae were housed in our general ZF facility and reared indefinitely in similar diets and environmental conditions. Starting from 10 dpf, growth, survival, and histological changes in the gut were analyzed and recorded for several months. Surprisingly, morphometric measurements revealed that ex-GF animals present a greater size and show reduced natural mortality than their CONR siblings. Moreover, gut morphology observations through histochemical preparations of 90 dpf ZF revealed a significantly reduced number of goblet cells, suggesting a reduced mucus hydrogel layer in ex-GF ZF. Strikingly, the offspring of ex-GF animals were more susceptible to a bacterial challenge. Studies in progress revealed that some immune mechanisms involved in commensal priming at the early developmental stages have a long impact along with development, providing enhanced physiological characteristics to the adults that further transmitted enhanced disease resistance to their offspring. The preliminary results point toward epigenetic changes as mediators of the response, but to test the hypothesis studies are in progress.

International Society of Fish Nutrition, 2006

A feeding trial was conducted to evaluate potential immunomodulatory effects of dietary arginine ... more A feeding trial was conducted to evaluate potential immunomodulatory effects of dietary arginine dosages in Japanese flounder. A basal diet was formulated to contain 60 % crude protein, 9 % crude lipid, 4 % crude sugar, and digestible energy of 3.2 kcal g-1. Four experimental diets based on the common basal were prepared by mixing 0 (control), 0.25, 0.5, or 1.0 g kg-1 of dl-arginine. Each diet was fed to replicate groups of juvenile Japanese flounder twice daily at a total rate of 2.5 % biomass day-1 for 45 days. In addition, after 2, 4, and 6 weeks of arginine treatment Edwardsiella tarda intraperitoneally challenges were executed with 0.1 ml of 2.5_10 6 cells ml-1 per fish, 10 fish in duplication per treatment at 22°C average temperature, to test the effect of diet on disease resistance. No significant (P>0.05) differences in weight gain, hematological parameters, C-reactive protein and serum peroxidase activity were generally observed along the trial in fish fed the diets supplemented with arginine compared to the control. However, leucocytes oxidative radical production from specimens treated with diets containing 0.5 and 1.0 g kg-1 supplemental arginine after 6 weeks had a significant effect (P<0.05). Macrophage activator factor (MAF) production was significantly enhanced by the 0.25 and 0.5 g kg-1 supplemental arginine after 4 weeks of treatment and only by the 0.25 g kg-1 supplemental arginine after 6 weeks. Nitric oxide (NO) production in serum and in leucocytes became significantly enhanced in fish treated with the diet containing 0.5 g kg-1 supplemental arginine after 4 or 6 weeks. In the present experiment, all arginine-treated specimens after 2 weeks did not present any significant difference. And, animals treated with the highest arginine supplementation tested level for a period longer than 4 weeks presented inert effect. In addition, arginine-treatment resulted in a positive correlation among immune parameters and disease resistance after exposure to E. tarda in controlled challenges. The relative percent survival (RPS) in fish arginine-treated with 0.25, 0.5 or 1.0 g kg-1 after 2, 4 and 6 weeks were (0:12:0); (29:57:43) and (50:50:25) respectively. Based on the results of this study, it is concluded that dietary arginine supplemented with a fish meal-based diet in a dose close to 0.5 mg kg-1 dry diet positively influences the immune response of Japanese flounder and increases resistance to E. tarda infection when administered for a period of 4 to 6 weeks. Observations suggest that NO and MAF increased production have a determinant role in disease resistance enhancement. Further work should be needed to determine the kinetics of reactions.