Cells In Fish Research Articles

Characterization of ccl20a.3 and ccl20l as gene markers for Th17 cell in turbot

T-helper 17 lymphocytes (Th17) are the most common inflammatory cells identified in mammals. However, the identification of Th17 cells and the clarification of their function in teleost fish remain largely unknown. In this study, we took advantage of the single-cell RNA sequencing-based immune cell atlas that was identified in turbot (Scophthalmus maximus), and revealed two chemokine-related genes, ccl20a.3 and ccl20l, that were specifically expressed in Th17 cells. Moreover, through immuno-fluorescence analysis, we found that CCL20a.3 or CCL20l was co-expressed with the classical makers in Th17 cells, including IL17a/f1 and IL22. Furthermore, through a Th17 lineage-specific transcription factor RORc inhibitor GSK805 treatment, we found that the expression of ccl20a.3 and ccl20l was significantly impaired, compared with other T cell markers. Besides, we also found that ccl20a.3 and ccl20l exhibited the same dynamic response with the classical markers that were identified in Th17 cells during bacterial infection. Taken together, these results provide potential gene markers for better understanding of the dynamic immune responses of Th17 cells in teleost fish.

The RNA-binding protein Adad1 is necessary for germ cell maintenance and meiosis in zebrafish.

The double stranded RNA binding protein Adad1 (adenosine deaminase domain containing 1) is a member of the adenosine deaminase acting on RNAs (Adar) protein family with germ cell-specific expression. In mice, Adad1 is necessary for sperm differentiation, however its function outside of mammals has not been investigated. Here, through an N-ethyl-N-nitrosourea (ENU) based forward genetic screen, we identified an adad1 mutant zebrafish line that develops as sterile males. Further histological examination revealed complete lack of germ cells in adult mutant fish, however germ cells populated the gonad, proliferated, and entered meiosis in larval and juvenile fish. Although meiosis was initiated in adad1 mutant testes, the spermatocytes failed to progress beyond the zygotene stage. Thus, Adad1 is essential for meiosis and germline maintenance in zebrafish. We tested if spermatogonial stem cells were affected using nanos2 RNA FISH and a label retaining cell (LRC) assay, and found that the mutant testes had fewer LRCs and nanos2-expressing cells compared to wild-type siblings, suggesting that failure to maintain the spermatogonial stem cells resulted in germ cell loss by adulthood. To identify potential molecular processes regulated by Adad1, we sequenced bulk mRNA from mutants and wild-type testes and found mis-regulation of genes involved in RNA stability and modification, pointing to a potential broader role in post-transcriptional regulation. Our findings suggest that the RNA regulatory protein Adad1 is required for fertility through regulation of spermatogonial stem cell maintenance in zebrafish.

Assessment of Valerian (Valeriana officinalis) on Common Carp, Cyprinus carpio: Anesthesia

The anesthetic efficacy of Valerian (Valeriana officinalis) powders was evaluated on common carp (Cyprinus carpio L.) The following concentrations (250, 350, 450) mg/liter were used in three replicates. During the experiment, the period required for partial and total anesthesia and the time required for partial and total recovery was tested, as well as the number of red and white blood cells after and before Anesthesia in addition to blood serum enzymes represented (ALP, GOT, GPT, CK, and LDH in Ul / l). The results showed that the least period of anesthesia occurred in the fourth treatment, the treatment with a concentration of 450 mg / l, as well as the least period required for recovery, while the recovery period was the longest. And anesthesia in the first treatment exposed to a concentration of 250 mg/liter, and it was not noticed that there was a significant difference between each the number of red blood cells, white blood cells in the test fish before the experiment and after treatment with valerian plants, as well as each of (ALP, GOT, GPT, CK, and LDH in Ul / l) There were no significant differences between all treatments before and after exposure to valerian, and it is concluded from the study that valerian is a safe plant for use in anesthetizing fish.

Selection for disease resistance in Oncorhynchus mykissincreases the myeloid to lymphoid balance and is affected by rapamycin as measured by flow cytometry and scRNAseq.

Abstract Heritable variation in disease resistance has been described in many species, but the effects of selection on innate immunity are unclear. Here, we compare the cellular response of two lines of rainbow trout that differ in survival due to five generations of selection for resistance to Flavobacterium psychrophilum(Fp), an important fish pathogen. We examined bacterial load, plasma biomarkers, whole blood cellular composition, and blood chemistry at days 1 and 5 after bacterial or PBS injection. We also examined the effect of 0.8 mg/kg rapamycin pretreatment, which results in immunosuppression and a ~100-fold reduction in the challenge LD 50. On day 1, bacterial load did not differ between genetic lines and treatment, but by day 5, the R-line exhibited no increase in Fpload, while rapamycin increased R- and S-line loads 33- and 506-fold, respectively. Biomarkers: cathelicidin 2 and C1q-LP3 significantly differed between genetic lines and by time, corresponding to Fpload. Analysis of whole blood by flow cytometry identified a response to infection and an ~8 percentage point higher frequency of myeloid-gated cells in R-line fish that was consistent in both naïve and challenged animals. S-line fish exhibited a higher percentage of IgM +B cells. Single-cell RNAseq was conducted on day 1 purified blood and identified 23 cell clusters including multiple B cell, neutrophil and macrophage subpopulations. In naïve animals, S-line fish harbored a cxcr4and mychexpressing B cell subpopulation absent in R-line fish. Rapamycin pretreatment caused profound loss of multiple B cell subpopulations in R- and S-line fish. In summary, selection of rainbow trout for resistance to Fpis correlated with a cellular shift in peripheral blood and is mTOR dependent. US Department of Agriculture, Agricultural Research Service; Project number 1930-32000-007

Identification of fish spermatogenic cells through high-throughput immunofluorescence against testis with an antibody set.

Image-based identification and quantification of different types of spermatogenic cells is of great importance, not only for reproductive studies but also for genetic breeding. Here, we have developed antibodies against spermatogenesis-related proteins in zebrafish (Danio rerio), including Ddx4, Piwil1, Sycp3, and Pcna, and a high-throughput method for immunofluorescence analysis of zebrafish testicular sections. By immunofluorescence analysis of zebrafish testes, our results demonstrate that the expression of Ddx4 decreases progressively during spermatogenesis, Piwil1 is strongly expressed in type A spermatogonia and moderately expressed in type B spermatogonia, and Sycp3 has distinct expression patterns in different subtypes of spermatocytes. Additionally, we observed polar expression of Sycp3 and Pcna in primary spermatocytes at the leptotene stage. By a triple staining of Ddx4, Sycp3, and Pcna, different types/subtypes of spermatogenic cells were easily characterized. We further demonstrated the practicality of our antibodies in other fish species, including Chinese rare minnow (Gobiocypris rarus), common carp (Cyprinus carpio), blunt snout bream (Megalobrama amblycephala), rice field eel (Monopterus albus) and grass carp (Ctenopharyngodon idella). Finally, we proposed an integrated criterion for identifying different types/subtypes of spermatogenic cells in zebrafish and other fishes using this high-throughput immunofluorescence approach based on these antibodies. Therefore, our study provides a simple, practical, and efficient tool for the study of spermatogenesis in fish species.

The primary culture and subculture of kidney-derived cells of the bony lip barb fish (Osteochilus vittatus) using different concentrations of serum

Cell and tissue culture have advanced and developed in a short time yet continuous improvement needs to be conducted for better results. This research was conducted to obtain suitable culture conditions of kidney-derived cell culture from bony lip barb kidney (Osteochilus vittatus). The development of primary cell culture from the kidney fragments was performed in a cell culture medium composed of DMEM supplemented with various concentrations (0%, 5%, 10%, and 15%) of Fetal Bovine Serum, Penicillin-Streptomycin, and 2Mm L-Glutamine, and incubated at 29°C with 0.8% CO2. The results showed that the primary cell culture achieved confluence on day four and subsequently subcultured. The addition of 10% serum increased cell density and cell survival. The cell morphological evaluation using a phase-contrast microscope showed two dominant cell types for both primary and subcultures, i.e. erythrocytes and small-rounded cells. The highest cell yield was obtained with the addition of 10% serum concentration with Population Doubling Time (PDT) and Population Doubling Level (PDL) for 69 h 23 min and 0.47 times, respectively. Statistical analysis indicated that serum addition significantly increased cell density of the primary culture (p<0.05). A better proportion of serum supplementation to culturing success is essential to establish an auspicious cell line.

In Amphioxus Embryos, Some Neural Tube Cells Resemble Differentiating Coronet Cells of Fishes and Tunicates.

AbstractFor neurula embryos of amphioxus (chordate subphylum Cephalochordata), the anterior region of the neural tube was studied with transmission electron microscopy. This survey demonstrated previously unreported cells, each characterized by a cilium bearing on its shaft a protruding lateral bubble packed with vesicles. Such cilia resemble those known from immature coronet cells in other chordates-namely, fishes in the Vertebrata and ascidians and appendicularians in the Tunicata. This wide occurrence of coronet-like cells raises questions about their possible homologies within the phylum Chordata. When considered at the level of the whole cell, such homology is not well supported. For example, the fish cells are generally thought to be glia, while the tunicate cells are considered to be neurons; moreover, cytoplasmic smooth endoplasmic reticulum, which is predominant in the former, is undetectable in the latter. In contrast, a more convincing case for homology can be made by limiting comparisons to the cell apices with their modified cilia. In addition to the fine-structural similarities between fishes and tunicates already mentioned, nonvisual opsins have been found associated with the vesicles in the modified cilia of both groups. Such opsins are thought to link photoreception to endocrine output controlling behavior. Further work would be needed to test the idea that the amphioxus diencephalic cells with lateral bubble cilia might similarly be opsin rich and could provide insights into the evolutionary history of the coronet cells within the phylum Chordata.

Density, Distribution and Staining Properties of Eosinophilic Granular Cells in Oscar Fish (Astronotus ocellatus) Intestine

Fish eosinophilic granular cells are found in the connective tissues of structures associated with the external environment. These cells are similar to mammalian mast cells in terms of structural and functional characteristics. The cytoplasmic granules of these cells give different staining reactions depending on fixative type. This study aimed to determine the staining properties and densities of eosinophilic granular cells in Oscar fish (Astronotus ocellatus Agassiz, 1831) intestine using different fixatives and histochemical techniques. Formalin and basic lead acetate fixation-giemsa staining indicated that eosinophilic granular cells were abundant in anterior intestine, localizing around especially the blood vessels and submucosa. Giemsa staining of Bouin’s fixed rather than other fixatives showed that eosinophilic granular cells were higher in posterior intestine. No reaction was observed in eosinophilic granular cells in Thionin and Toluidine Blue staining in any fixative. Eosinophilic granular cells in samples fixed with different fixatives had metachromatic Alcian Blue staining. In conclusion, this study shows that fixatives may have different effects on the distribution and staining properties of eosinophilic granular cells in Oscar fish intestinal regions. Abundance in anterior intestine and around submucosal blood vessels shows that eosinophilic granular cells could play an active role in mucosal immunity against food-borne pathogens.

Interspecific Variation in the Inner Ear Maculae of Sharks.

There is well-documented diversity in the organization of inner ear hair cells in fishes; this variation is thought to reflect the differing functional requirements of species across a range of ecological niches. However, relatively little is known about interspecific variation (and its potential ecological implications) in the number and density of inner ear hair cells in elasmobranchs (sharks, skates, and rays). In this study, we quantified inner ear hair cells in the saccule, lagena, utricle, and macula neglecta of 9 taxonomically and ecologically distinct shark species. Using phylogenetically informed comparative approaches, sharks that feed in the water column had significantly greater hair cell density and total number of hair cells in the lagena and macula neglecta (i.e., vertically oriented maculae) compared to species that feed primarily on the seafloor. In addition, sharks within Carcharhinidae seemingly possess a specialized macula neglecta compared to other shark species. Overall, findings suggest that, similar to bony fishes, there is considerable variation in hair cell organization of shark inner ears, which may be tied to variation in ecology and/or specialized behaviors between different species.

Growth performance and blood figures of African catfish (Clarias sp.) fed with marine fungal fermented feed

This study examined the impact of marine fungal fermented feed at various concentrations on the growth and blood profiles of African catfish Clarias sp.. Catfish with an average body weight of 0.74 ± 0.01 g were raised in an aquarium that measured 30 cm x 45 cm x 30 cm and contained 25 L of water. The feed was not fermented in this study (control), and the feed was fermented with marine fungi at various concentrations (using 0.5%, 1%, 2%, and 4% extract marine fungi). At 8:00 am, 12:00 pm, and 16:00 pm, fish were fed three times a day until they were completely satisfied. The results showed that feeding juvenile catfish marine fungal fermented diet resulted in significantly higher growth performance. The larger final biomass, faster growth, and lower feed conversion ratio were used to examine this. The generation of red blood cells in fish fed with fermented diets was higher and better than the control treatment, according to the results of the blood profile of catfish (P 0.05).

Cytokine networks provide sufficient evidence for the differentiation of CD4+ T cells in teleost fish

Cytokines, a class of small molecular proteins with a wide range of biological activities, are secreted mainly by immune cells and function by binding to the corresponding receptors to regulate cell growth, differentiation and effects. CD4+ T cells can be defined into different lineages based on the unique set of signature cytokines and transcription factors, including helper T cells (Th1, Th2, Th17) and regulatory T cells (Treg). In teleost, CD4+ T cells have been identified in a variety of fish species, thought to play roles as Th cells, and shown to be involved in the immune response following specific antigen stimulation. With the update of sequencing technologies, a variety of cytokines and transcription factors capable of characterizing CD4+ T cell subsets also have been described in fish, including hallmark cytokines such as IFN-γ, TNF-α, IL-4, IL-17, IL-10, TGF-β and unique transcription factors such as T-bet, GATA3, RORγt, and Foxp3. Hence, there is increasing evidence that the subpopulation of Th and Treg cells present in mammals may also exist in teleost fish. However, the differentiation, plasticity and precise roles of Th cell subsets in mammals remain controversial. Research on the identification and differentiation of fish Th cells is still in its infancy and requires more significant effort. Here we will review recent research advances in characterizing the differentiation of fish CD4+ T cells by cytokines and transcription factors, mainly including the identification of Th and Treg cell hallmark cytokines and transcription factors, the regulatory role of cytokines on Th cell differentiation, and the function of Th and Treg cells in the immune response. The primary purpose of this review is to deepen our understanding of cytokine networks in characterizing the differentiation of CD4+ T cells in teleost.

Ontogenetic development of inner ear hair cell organization in the New Zealand carpet shark Cephaloscyllium isabellum

IntroductionThe inner ear hair cells of fishes can provide insight into the early evolution of vertebrate inner ear structure. Fishes represent some of the first vertebrates to evolve auditory capacity, and the same basic structure, the sensory hair cell, provides the fundament for auditory and vestibular function in jawed vertebrates. Despite holding critical basal position in the evolutionary tree of gnathostomes, relatively little is known about inner ear hair cells in elasmobranchs. Specifically, the extent of plasticity in hair cell organization throughout ontogeny among different sensory epithelia and the degree of variation between species is unknown.MethodsIn this study, we characterized the inner ear hair cells of the New Zealand carpet shark Cephaloscyllium isabellum throughout ontogeny by quantifying macular area, number of hair cells, hair cell density, and hair cell orientations in the inner ear maculae from a range of body sizes.ResultsSimilar to other elasmobranchs and bony fishes, macular area and the number of hair cells increased throughout ontogeny in the otolith organs. The orientations of hair cells within each maculae also was consistent with the limited data on other elasmobranchs. However, contrary to expectation, the macula neglecta did not increase in area or hair cell number throughout ontogeny, and hair cell density did not change with body size in any maculae.DiscussionThese findings suggest there may be variation between elasmobranch species in ontogenetic development of hair cell organization that may be related to hearing capabilities throughout life.

A System-Level Study Reveals That a Defective Thymic Niche Accelerates the T-ALL Initiation

The ontogeny of T-cell acute lymphoblastic leukemia (T-ALL) is unknown. In humans and mice, T-ALL is diagnosed when malignant T cells are detectable in the peripheral blood, i.e., at a stage when disease development is complete. Therefore, the malignant processes in the thymus - where the T cells develop - are an unexplored "black box". We took an interdisciplinary approach combining in vivo experiments with computational modeling to investigate the role of the thymic niche in disease manifestation. We focused on the role of interleukin 7 (IL-7) in the thymus, as approximately 70-80% of T-ALL patients have either increased IL-7 receptor (IL-7R) expression or gain-of-function mutations in this gene. In addition, autocrine production of IL-7 by T-ALL blasts has been detected in some patients. Using our recently developed "virtual thymus" model (Aghaallaei et al., Science Advances, 2021), we first simulated more than 200 scenarios to find out the conditions that lead to massive expansion of a single clone in the thymus. We then genetically engineered thymocytes and thymic epithelial cells (TECs) in medaka fish based on in silico predictions. Consistent with our computational model, we found that manipulation of thymocytes or TECs alone resulted in only 25% (N=36) and 10% (N=29) T-ALL development, respectively, in the thymus of Medaka larvae after five days; however, this frequency was accelerated to 54% (N=45) when both cell types were manipulated simultaneously. Our results strongly suggest that defective thymic niche has a strong impact on T-ALL development. Taken together, we propose a previously unknown mechanism that may trigger T-ALL initiation in the thymus. In addition, we introduce the "virtual thymus" as a new versatile in silico model that can predict the outcome of conditions for the development or treatment of T-ALL.

Interspecific differences and ecological correlations of ultraviolet radiation tolerance in low- and high-altitude fishes

Ultraviolet radiation (UVR) is a ubiquitous environmental factor and with complex and diverse effects on organisms, and the UVR tolerance of species varies due to selection, adaptation and evolution. This study aimed to improve our understanding of the interspecific differences in UVR tolerance of fishes. First, we compared skin histology between high-altitude fishes and their low-altitude relatives. Second, we assessed the skin histological changes of 22 fish species after UVR exposure and quantified their damage levels. Finally, the factors affecting the difference in UVR tolerance among fishes were studied. Thin or absent scales was a common characteristic of high-altitude fishes in different taxonomic groups, but other skin structure parameters (the thickness of the dermis and epidermis, the size and number of mucous and club cells) did not show convergence. After UVR exposure, the size and number of mucous cells and club cells in the experimental fishes were generally reduced, but the changes in epidermal and dermal thickness varied among species. The change rata of melanin area was strong negatively correlated with the level of damage, indicating that increased melanin was helpful to reduce UVR damage. On the whole, the skin histological changes of high-altitude fishes after UVR exposure did not differ from those of low-altitude fishes. However, the epidermis and dermis of Gymnocypris eckloni and Oncorhynchus mykiss, which had the highest UVR tolerance, were nonpathologically thickened, indicating that they may contribute to UVR tolerance. There were significant interspecific differences in UVR tolerance among the experimental fishes, with high-altitude and cold water fishes showing stronger UVR tolerance. Interestingly, some low-altitude cold water fish (e.g., O. mykiss) also showed high UVR tolerance, suggesting that besides high-altitude adaptation, low-temperature adaptation also may play an important role in organismal adaptation to UVR.

Turbidity derived from palm oil mill effluent alters feeding ability of male siamese fighting fish (Betta splendens, Regan 1910)

To date, several harmful effects of Palm Oil Mill Effluent (POME) have been reported on aquatic organisms, including fish. However, the turbidity effects of POME on the feeding ability of fish are still unexplored. We first assessed the turbidity effects of POME on the feeding ability of male siamese fighting fish (Betta splendens). Siamese fighting fish were exposed to < 1 NTU (control) and five concentrations of POME turbidity levels, namely 20 NTU (Treatment A), 40 NTU (Treatment B), 60 NTU (Treatment C), 80 NTU (Treatment D), and 100 NTU (Treatment E). Each of the control and treatment groups was maintained in five replicates. Preference tests were carried out in a 10-L aquarium (35 ×21×25 cm3). The feeding rate, time to capture first prey, aggressive behaviour, foraging area, and retinal light adaptation rate and retina histopathology were analyzed. The exposure time for the determination of feeding rate, foraging area, and retinal light adaptation rate and retina histopathology was set for 15 min, while for aggressive behaviour test, the exposure duration was 5 min. Results showed that the feeding rate, aggressive behaviour and foraging area significantly decreased with increasing turbidity levels of POME. In contrast, the time to capture first prey significantly increased. No significant changes in retinal light adaptation rate was recorded between treatments. Furthermore, results showed several retinal degenerations, including reduced granular spherical cells (GL) and detachment of bipolar cells from photoreceptor cells (DBPC) in siamese fighting fish exposed to high-level turbidity of POME. This study provides valuable information to support POME remediation management, especially turbidity parameters.

Mycn regulates intestinal development through ribosomal biogenesis in a zebrafish model of Feingold syndrome 1.

Feingold syndrome type 1, caused by loss-of-function of MYCN, is characterized by varied phenotypes including esophageal and duodenal atresia. However, no adequate model exists for studying the syndrome's pathological or molecular mechanisms, nor is there a treatment strategy. Here, we developed a zebrafish Feingold syndrome type 1 model with nonfunctional mycn, which had severe intestinal atresia. Single-cell RNA-seq identified a subcluster of intestinal cells that were highly sensitive to Mycn, and impaired cell proliferation decreased the overall number of intestinal cells in the mycn mutant fish. Bulk RNA-seq and metabolomic analysis showed that expression of ribosomal genes was down-regulated and that amino acid metabolism was abnormal. Northern blot and ribosomal profiling analysis showed abnormal rRNA processing and decreases in free 40S, 60S, and 80S ribosome particles, which led to impaired translation in the mutant. Besides, both Ribo-seq and western blot analysis showed that mTOR pathway was impaired in mycn mutant, and blocking mTOR pathway by rapamycin treatment can mimic the intestinal defect, and both L-leucine and Rheb, which can elevate translation via activating TOR pathway, could rescue the intestinal phenotype of mycn mutant. In summary, by this zebrafish Feingold syndrome type 1 model, we found that disturbance of ribosomal biogenesis and blockage of protein synthesis during development are primary causes of the intestinal defect in Feingold syndrome type 1. Importantly, our work suggests that leucine supplementation may be a feasible and easy treatment option for this disease.

Effects of Jerusalem Artichoke (Helianthus tuberosus) as a Prebiotic Supplement in the Diet of Red Tilapia (Oreochromis spp.).

Simple SummaryFactors such as changes in water quality, the transport and handling of the fish, the presence of pollutants, and the high densities used in intensive farming of red tilapia, among others, can cause oxidative stress in fish, which can lead to immune system suppression and increase the risk of opportunistic infections. The use of Jerusalem artichoke (JA) as an oligofructose-rich prebiotic dietary supplement in fish feed could enhance growth performance, hematological and immunological parameters, as well as disease resistance. However, the effect of JA in diets for red tilapia is poorly reported. In the present study, we evaluated the effects of a Jerusalem artichoke-supplemented diet on the blood chemistry, growth performance, intestinal morphology, expression of antioxidant-related genes, and disease resistance against Aeromonas veronii challenge in juvenile red tilapia. The results of JA-supplemented (JA5 and JA10) diets showed beneficial effects in terms of growth performance, blood chemistry, and intestinal morphology. Importantly, the prebiotic diets (JA5 and JA10) were associated with a significant increase in the expression of gpx1 and gst antioxidant-related genes and disease resistance against A. veronii. In conclusion, prebiotic diets have the potential to be employed as prospective supplements in sustainable red tilapia farming.The aim of the present study was to evaluate the effects of a Jerusalem artichoke-supplemented diet on the blood chemistry, growth performance, intestinal morphology, expression of antioxidant-related genes, and disease resistance against Aeromonas veronii challenge in juvenile red tilapia. A completely randomized design (CRD) was followed to feed red tilapias with three experimental diets: control, 5.0 g/kg JA-supplemented (JA5), or 10.0 g/kg JA-supplemented (JA10) diets in triplicates for 4 weeks. The results revealed that the growth performance, weight gain (WG), specific growth rate (SGR), and average daily gain (ADG) of fish fed diets JA5 and JA10 were significantly higher (p < 0.05) than those of fish fed the control diet. Fish fed the control diet had significantly higher T-bilirubin, D-bilirubin, and ALT in blood serum than fish fed JA5 and JA10, as well as higher BUN than fish fed JA5. The number of goblet cells in the proximal and distal parts of the intestine revealed that the number of acid, neutral, and double-staining mucous cells of fish fed diets JA5 and JA10 was significantly higher (p < 0.05) than in fish fed the control diet. The diets including the prebiotic (JA5 and JA10) were associated with a significant increase in the expression of gpx1 and gst antioxidant-related genes and disease resistance against A. veronii in juvenile red tilapia. Therefore, JA5 and JA10 can be employed as promising prebiotics for sustainable red tilapia farming.

Myo-inositol supplement helps the performance of seawater-acclimated Nile tilapia, Oreochromis niloticus

Seawater (SW)-acclimated Nile tilapia, Oreochromis niloticus, reared under a salinity 30 environment had lower growth and survival than the freshwater (FW)-acclimated fish. It was hypothesized that cells of the SW-acclimated fish had not been able to synthesize an adequate level of a compatible osmolyte, myo-inositol (MI), in adjusting to the salinity 30 environment. In this study, MI supplements, at 250, 500, and 750 mg/kg pellets, were provided to the fish through top-dressing. After the 30-day feeding trial, the following parameters were determined: final body weights; survival; biomass increase; feed conversion ratio (FCR); plasma osmolality and ions; and two transcripts in the gills mips250 and mipa1 encoding enzymes responsible for MI biosynthesis. The SW-acclimated O. niloticus receiving 500-mg MI supplement had significantly higher survival, biomass increase, and lower FCR than those of the SW-acclimated fish receiving no supplement. At 500-mg MI supplemental level, the increasing values of plasma osmolality and Na+ observed in SW-acclimated fish were significantly attenuated. The transcript mipa1, but not mips250, was markedly up-regulated in the SW-acclimated O. niloticus, compared with that of the FW-acclimated fish. Again, MI at 500-mg supplement attenuated the up-regulation significantly. This study suggests that MI supplement at the optimum level enhanced the performance of SW-acclimated O. niloticus, and through yet unknown mechanisms, attenuated some of their physiological responses to the osmotic stress.

Nuclear and Morphological Alterations in Erythrocytes, Antioxidant Enzymes, and Genetic Disparities Induced by Brackish Water in Mrigal Carp (Cirrhinus mrigala).

Salinization of aquatic ecosystem, abrupt climate change, and anthropogenic activities cause adverse impact on agricultural land/soil as well as the aquaculture industry. This experimental study was designed to evaluate different biomarkers of oxidative stress, antioxidant enzymes, and genotoxic potential of diverse salinities of brackish water on freshwater fish. A total of 84 fresh water mrigal carp (Cirrhinus mrigala) were randomly segregated and maintained in four groups (T0, T1, T2, and T3) in a glass aquarium under similar laboratory conditions at various salinity levels (0, 3, 5, and 7 parts per thousand) to determine the pathological influence of brackish water. All the fish in groups T1, T2, and T3 were exposed to various salinity levels of brackish water for a period of 90 days while the fish of group T0 served as the control group. The experimental fish reared in different groups T1, T2, and T3 displayed various physical and behavioral ailments. The results revealed significantly augmented quantity of different oxidative stress indicators including reactive oxygen species (ROS) and thiobarbituric acid reactive substance (TBARS) in different visceral tissues (kidneys, liver, and gills) of exposed fish. Different antioxidant enzymes such as reduced glutathione (GSH), peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT) along with total proteins were remarkably reduced in the kidneys, gills, and liver tissues. Results showed significantly increased values of different nuclear abnormalities (erythrocyte with micronucleus, erythrocyte with condensed nucleus, and erythrocyte with lobed nucleus) and morphological changes (pear shaped erythrocyte, spindle-shaped erythrocytes, and spherocyte) in red blood cells of experimental fish. The results on genotoxic effects exhibited significantly increased DNA damage in isolated cells of liver, kidneys, and gills of exposed fish. The findings of our experimental research suggested that brackish water causes adverse toxicological impacts on different visceral tissues of fresh water fish at higher salinity level through disruption and disorder of physiological and biochemical markers.

Effects of ultrasound–assisted freezing on the quality of large yellow croaker (Pseudosciaena crocea) subjected to multiple freeze–thaw cycles

Repeated freezing and thawing due to temperature fluctuations irreversibly damage the muscle tissue cells of fish, thereby reducing their economic quality. In this study, the effects of ultrasound-assisted immersed freezing (UIF) technology on the changes in the quality of large yellow croaker (Pseudosciaena crocea) subjected to 0 to 5 freeze–thaw cycles were investigated. The results showed that the quality deterioration inevitably occurred after repeated freeze–thaw cycles. However, UIF significantly delayed the changes in the water holding capacity (WHC), immobilized water content, color and texture properties of fish. Compared to the control group (air freezing, AF), the thawing loss in the UIF group was reduced by 1.09 % to 4.54 % (P < 0.05), the centrifuging loss was reduced by 0.39 % to 1.86 % (P < 0.05), the migration of immobilized water content was reduced by 4 % to 5 % (P < 0.05). Moreover, SEM and LM images illustrated that the microstructures of muscle tissue in UIF group were more uniform and denser than that of the AF group after freeze–thaw cycles, and that the ice crystal size from UIF group were smaller and more regular than that of AF group. Furthermore, UIF did not caused more excessive protein oxidation of myofibrillary protein, but significantly delayed the lipid oxidation of fish muscle. The results indicated that UIF technology effectively inhibits the deterioration of fish quality affected by multiple freeze–thaw cycles, thus providing a reference for controlling the deterioration of aquatic products due to temperature fluctuations in the industry.