We report the first tetrodotoxin (TTX) poisoning cases from Van, Eastern Turkey, a city located more than 1000 km inland. Three male patients presented in December 2020 after eating Lagocephalus sceleratus, which a local vendor mistook for an edible fish. One patient, a 41-year-old man who consumed the fish liver, rapidly developed severe signs of poisoning (adverse event grade 4) with complete paralysis and respiratory failure. Arterial blood gas analysis showed severe acidosis (pH 7.179) and high lactate levels (5.3 mmol/L). Despite intensive care, including mechanical ventilation and high-dose vasopressors, the patient died on the fourth day of hospitalisation due to refractory cardiac arrest. The other two patients, who consumed less fish and no liver, had only mild symptoms (adverse event grades 1 and 2) and recovered completely within 24 h. This incident illustrates the extreme danger of consuming pufferfish and particularly its visceral organs, which typically contain substantially higher tetrodotoxin levels. Finding these cases so far inland reveals serious gaps in the Turkish food distribution system. The fish likely travelled from Mediterranean ports to Van in mixed fish crates without having been identified. Fish vendors need proper training to identify toxic species, and supply chains require better oversight to prevent such tragedies. Ovo je prvi prikaz niza slučajeva trovanja tetrodotoksinom (TTX) u gradu Vanu u istočnoj Turskoj, smještenom više od 1000 km od obale. Tri muškarca primljena su u hitnu službu u prosincu 2020. nakon konzumacije otrovne napuhače, vrste Lagocephalus sceleratus, koju je mjesni prodavač zamijenio s jestivom ribom. Jedan bolesnik, 41-godišnji muškarac koji je pojeo riblju jetru, ubrzo je dobio teške znakove trovanja (4. stupanj štetnih događaja) s potpunom paralizom i respiratornim zatajenjem. Analiza plinova arterijske krvi pokazala je tešku acidozu (pH 7,179) i povišene vrijednosti laktata (5,3 mmol/L). Unatoč intenzivnom liječenju, uključujući mehaničku ventilaciju i primjenu visokih doza vazopresora, bolesnik je preminuo četvrtoga dana hospitalizacije zbog refraktornoga srčanog zastoja. Druga dva bolesnika, koja su pojela manju količinu mesa ribe i nisu jela jetru, imali su samo blage simptome (1. i 2. stupanj štetnih događaja) te su se potpuno oporavili unutar 24 sata. Ovaj incident potvrđuje iznimnu opasnost konzumacije napuhača, osobito iznutrica, koje obično sadržavaju znatno više razine tetrodotoksina. Činjenica da je do trovanja došlo tako daleko od obale upozorava na ozbiljne propuste u turskom sustavu distribucije hrane. Riba je vjerojatno dopremljena iz mediteranskih luka u Van u sanducima s miješanom ribom, a da pritom nije bila identificirana. Prodavači ribe trebaju proći odgovarajuću edukaciju za prepoznavanje otrovnih vrsta, a opskrbni lanci morali bi imati bolji nadzor kako bi se spriječile ovakve tragedije.
This study assessed the comprehensive human risk exposure to metal(loid)s through consumption of fish and clams from the Nun River. Thirteen elements (Ag, As, Ba, Cd Co, Cr, Cu, Fe, Mn, Ni, Pb, V, and Zn) were determined in the aquatic samples using inductively coupled plasma-optical emission spectrometry (ICP-OES). Metal(loid) data were subjected to simple descriptive and inferential statistical analysis to examine variations among species and sampling periods. In fish, Fe was the dominant element, while Zn and Mn showed temporal increments. Toxic metal(loid)s such as Pb, Cr, Cd, Ni, and As were detected at concerning levels, with some concentrations exceeding the international safety thresholds of the joint Food and Agriculture/World Health Organization. Species differences were observed, with higher accumulation in carnivorous fish, while clams generally showed lower levels. Health risk assessment indicated that most hazard quotient (HQ) values were below 1.0, suggesting generally low noncarcinogenic risk under the exposure assumptions applied; however, elevated values were observed for Cd and As in some fish species. Cancer risk analysis identified Cd and As as the major contributors to lifetime carcinogenic risk. This study provides a comprehensive and integrated assessment of metal(loid) contamination in Nun River fish and clams, examining temporal variation, species-specific accumulation patterns, and associated human health risks. In doing so, it helps to address a significant regional data gap. The research findings highlight potential exposure concerns and emphasize the need for continued monitoring, pollution control, and public awareness to support food safety in the Niger Delta region.
Adaptation of organisms to extreme environments requires dramatic phenotypic changes. Studying these changes can elucidate mechanisms underlying phenotypic differences in the context of both evolution and human disease. The Mexican tetra, Astyanax mexicanus, is a powerful model of extreme adaptation over a short evolutionary time scale. This fish species includes surface- and cave-dwelling ecotypes, with cavefish displaying many adaptations to subterranean life, including behavioral changes such as sleep loss, increased appetite, and reduced aggression. Unraveling the mechanisms underlying these changes has been challenging, presumably because they are complex traits that required coordinated changes across multiple cell types to evolve. Here, we present a spatially integrated comparative cell atlas of whole adult brains of surface and cavefish. After establishing the molecular signatures of 35 cell types, we show that cave colonization drove canalized regulatory changes to gene expression across diverse cell types. Cavefish brains show shifts in cell-type composition compared to their surface counterparts, as well as complex regulatory changes to pathways governing hypoxia response and circadian rhythm. Microglia in the cavefish brain underwent extensive transcriptional remodelling, including changes in senescence and AMPK pathways. Further, cell-cell communication analysis identified a cave-enriched ligand-receptor communication pattern centered on signals sent from glial cells to diverse populations of neurons. This atlas identifies genetic changes associated with neural and behavioral evolution and provides a resource for mechanistic studies examining brain evolution.
The large-scale construction of cascade dams worldwide has profoundly altered river hydrological regimes and habitats, producing significant threats to fish diversity. This study integrated environmental DNA (eDNA) and traditional fishing data to construct a fish dataset from the middle and upper reaches of the Jinsha River (2003-2024). It was used to systematically reveal the spatiotemporal patterns of fish diversity and analyze key environmental drivers. Additionally, the Isolation Forest algorithm was employed to remove 29 outliers from the historical data. Distance-based redundancy analysis showed that flow velocity was the strongest environmental driver for species composition (R2 = 0.51, p = 0.005) and functional composition (R2 = 0.41, p = 0.018). This suggests that flow velocity reduction, closely associated with cascade dam construction, may act as an important agent of environmental filtering. β-diversity analysis revealed that local-scale functional nestedness (β-nes: 37.4%) and a directional functional shift toward resident and limnophilic groups suggest incipient functional homogenization in the cascade dam system, although temporal evidence was lacking due to staggered dam construction. Notably, although species richness increased, functional evenness (FEve) declined significantly, revealing an imbalanced functional diversity despite higher taxonomic diversity. These findings enhance our understanding of the long-term ecological impacts of cascade dams and provide a scientific basis for mitigating incipient biodiversity homogenization induced by hydropower development.
The flavor quality of traditional fermented fish in Guizhou exhibits significant regional variations, yet its microbial and metabolic mechanisms remain unclear. This study integrated sensory evaluation, high-throughput sequencing, and untargeted metabolomics to compare the quality, microbial communities, and metabolite profiles of fermented fish from Jinping, Liping, and Tianzhu. Results showed that Liping samples demonstrated optimal performance in flavor harmony, acidity perception, and texture. Tianzhu samples exhibited the lowest microbial α-diversity, with Staphylococcus accounting for over 99% of bacterial communities, whereas Jinping and Liping samples were enriched with Weissella, Saccharomyces, and Debaryomyces. Metabolomic analysis identified a total of 1,372 metabolites. PLS-DA revealed significant separation of metabolic profiles among the three groups, with differential metabolites primarily enriched in amino acid metabolism and the citric acid cycle. Liping samples exhibited significant upregulation of L-glutamine, citric acid, and L-glutamate. Spearman correlation analysis indicated that Weissella and Saccharomyces were positively correlated with the aforementioned flavor metabolites. This multi-omics study demonstrates that specific microbial community structures and their metabolic activities are core factors driving regional flavor differences in Guizhou traditional fermented fish, providing a theoretical basis for improving product quality through targeted microbial community regulation.
In this study, a composite dressing, based on fish scale collagen, Eclipta prostrata (L.) L. (EA) and Gardenia jasminoides J. Ellis (GJE) extracts, was prepared using a 3D printing method. The optimization of technological factors for the 3D printing process of the composite thin films includes the number of printing layers, the concentration of GJE extract, and the concentration of EA extract. The response surface methodology (RSM) combined with a Box-Behnken design and quadratic order model were applied for optimization of technological factors. Based on the objective function values - the in vitro blood clotting time, the optimal parameters for the 3D printing process of the thin films were six printing layers, 2.20% of GJE extract, and 10.74% of EA extract. After being prepared under optimal conditions, the collagen-based film was evaluated for characteristics and properties using methods and techniques such as infrared spectroscopy, scanning electron microscopy, water contact angle, color change, and mechanical properties. The variation in mechanical and surface properties of the collagen-based film after 60 days of storage under room temperature has also been assessed. Additionally, the anti-inflammatory activity through the inhibition of NO production, cell viability, and biocompatibility of this composite film with simulated human body fluids were also assessed. The obtained results indicate that the fish scale collagen exhibits excellent bioactivity when combined with plant extracts, suitable for biomedicine applications. The 3D printed fish scale collagen/Eclipta prostrata L. extract thin films have potential to be used as a new-generation hemostatic dressing with significant cost savings.
Collagenases can hydrolyze collagen to produce bioactive peptides, but their low activity limits industrial application. This study aimed to identify an efficient collagenase and construct a high-level expression system for fish scale biovalorization. A new collagenase (LsCol) from Lysinibacillus sp. was identified using deep-learning-based computational tools CLEAN and DLKcat, and its high-level expression in Bacillus subtilis was achieved through a combination of strain engineering strategies. The engineered strain produced an extracellular collagenase activity of 21,080.6 U/mL in fed-batch fermentation. LsCol hydrolyzed gelatin, Pz peptide and insoluble type I collagen. LsCol preferentially cleaved Hyp-Gly peptide bonds within Gly-Pro-Hyp repeat units of collagen. LsCol efficiently degraded fish scales collagen into low molecular weight peptides (< 500 Da, accounting for 73.0%) and produced collagen tripeptides (CTP) and Gly-Pro-Hyp (GPH) with yields of 26.5% and 5.75%, respectively. This study provides an effective way for collagenase production and biovalorization of fish scales.
Insulin-like growth factor 1 (IGF1) is a key somatotropic hormone that controls growth and metabolism, thereby conveying growth status to the reproductive endocrine system. The regulation of gonadotropin-releasing hormone (GnRH) to synthesize and release both the gonadotropin hormones (FSH and LH) is well documented. Numerous attempts have been made to understand the interaction of gonadotropic (reproductive) and somatotropic (growth) axis, but the dynamics of their interaction are still not clear at the pituitary level among teleost. The present study bioinformatically demonstrated protein-protein interaction of IGF1 with the proteins of both somatotropic and gonadotropic axes suggesting a cross-talk between them. In vitro, the effects of IGF1 alone or in combination with GnRH on pituitary transcripts for follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were examined. Experiments were conducted with dispersed pituitary cells of the catfish, Heteropneustes fossilis, to determine the time- and dose-dependent response of IGF1 and GnRH alone or in combination to assess fshβ and lhβ. The results showed a time-dependent effect with maximum transcripts level at 1 h of exposure. IGF1 and GnRH showed dose-dependent effects on differential expression of both transcripts when administered alone, with maximum fshβ and minimal lhβ levels at the higher dose. A synergistic impact on both the transcripts was observed in the presence of IGF1 and GnRH. Collectively, the study suggests that IGF1 plays a crucial role in regulating the synthesis of hormones in the catfish by interacting between the gonadotrophs and somatotrophs at the pituitary level.
This study presents the most comprehensive phylogenetic analysis to date of the high-Andean fish genus Orestias, a diverse and largely endemic group, using Genotype-by-Sequencing (GBS) and extensive sampling across the full range of the genus, from northern Peru to northern Chile. In particular, the evolutionary relationships and population structure within the O. agassii species complex were reconstructed. Phylogenetic analyses consistently recovered three major clades: a newly identified Northern clade, the Lake Titicaca radiation, and the O. agassii species complex. The genus originated around 4 Ma, with a deep divergence between clades (4-3 Ma) and more recent diversification within each clade (0.3-1.15 Ma). This pattern suggests historical extinctions linked to climatic shifts and the survival of lineages in large Andean lakes acting as refuges. The O. agassii species complex is the most widespread clade, with a dispersal route from north (Mantaro basin) to south (Chilean Altiplano). Population genomic analyses revealed strong geographic structuring within the O. agassii species complex, with nine distinct lineages, corresponding to major basins such as Chillon, Coastal Arequipa, Mantaro, Apurímac, Urubamba and Titicaca. The O. agassii populations show an isolation-by-distance pattern attributed to ecological preference and low vagility. Our results provide a robust phylogenetic framework and emphasize the need for further taxonomic and conservation efforts, as Orestias species are particularly vulnerable to anthropogenic pressures such as pollution, habitat loss, climate-driven lake shrinkage, and predation by introduced trout.
Intensive aquaculture serves as the cornerstone of global edible aquatic protein supply, yet pesticide residues from agricultural non-point source pollution have become a major environmental threat to teleost health, compromising both aquaculture sustainability and food safety. Pesticide exposure induces acute and chronic toxicity through multiple pathological pathways, including oxidative stress, intestinal microbiota dysbiosis, immunosuppression, endocrine and metabolic disruption, as well as genetic and tissue damage. As environmentally benign feed additives, probiotics demonstrate considerable promise in mitigating pesticide-induced toxicity in teleosts owing to their multi-target regulatory activities and innate biodegradability. Here, we critically review the core toxic effects of pesticides on teleost fish and synthesize current understanding of the molecular mechanisms underlying probiotic-mediated protection. We particularly emphasize the key unresolved challenges that hinder industrial translation-including pronounced strain specificity, insufficient mechanistic depth, lack of standardized dosing protocols, severe research gaps under realistic co-exposure scenarios, incomplete safety evaluation systems, and fragmented regulatory frameworks. Moreover, we propose future directions with specific emphasis on multi-omics integrated approaches, high-throughput strain screening platforms, combined pollution research, industrial formulation technologies, and harmonized safety assessment standards. This review aims to provide a theoretical and practical framework for the green prevention and control of pesticide pollution in aquaculture and to guide the rational development of targeted probiotic products for sustainable teleost farming.
One of the most serious pollution problems in aquatic lake systems is contamination by heavy metals (HMs) and microplastics (MPs). These pollutants cause serious harm to aquatic organisms. The research focused on studying HMs (Mn, Cd, Pb, Ni, and Cr) and MPs in organs of Clarias gariepinus inhabitant of Burullus Lake. Water and live fish samples were collected from the northwestern part of the lake, preserved and then transferred for laboratory analyses. HMs were analyzed using digestion methods with nitric acid and hydrogen peroxide. For MPs, 10% potassium hydroxide was used. The metal ions followed these orders Mn > Cr > Cd > Ni > Pb and Mn > Cr > Cd > Pb > Ni in the liver and muscle, respectively. In water, this order was Mn > Cr > Pb > Ni > Cd. The highest bioconcentration factor (BCF) was for Mn in both tissues; the lowest was Pb in the liver and Cd in the muscle. The metal concentrations exceeded the limits reported by FAO/WHO. The estimated daily and weekly intakes (EDI and EWI) were higher than 1 in both adults and children. The hazard index (HI) was within the limit of 1. The values were higher for Cd, Pb, and Ni than the tolerable value of lifetime carcinogenic risk according to USEPA of (10-5). The most abundant microplastics (MPs) were transparent lines, followed by colored fibers and then colored fragments. The polymeric composition of microplastics was 333 T Teflon (polytetrafluoroethylene [PTFE]) in the intestine and 118 T (low-density polyethylene [LDPE]) in the muscle. The pollution load index for MPs was more than 1 in both tissues. The histopathological changes indicated a loss of detoxification ability and intestinal integrity, as manifested by hepatocellular necrosis and vacuolar degeneration. Intestinal lesions exhibited villous sloughing along with necrobiotic alterations and iron deposition. It is concluded that high contamination ratios of MPs were observed in the tested samples and these ratios may increase within time if water and sediments are not treated effectively.
Type I interferons (IFNs) are central antiviral cytokines in vertebrates, yet the mechanisms underlying their functional diversification in early vertebrates remain unclear. Teleost fish, whose IFN repertoires expanded through whole-genome duplication, provide a powerful model to address this question. Here, we systematically characterize grass carp (Ctenopharyngodon idella) IFNd to elucidate the mechanisms underlying its functional divergence from IFNa. IFNd used the same receptors of IFNa to activate JAK-STAT pathway, albeit with significantly lower antiviral potency than IFNa. Structural analysis revealed significant differences in F helix length and key receptor-binding residues between IFNa and IFNd. Furthermore, single-cell RNA sequencing demonstrated high heterogeneity among immune cell subpopulations responding to IFNa and IFNd, Notably, the signaling pathways enriched by differentially expressed genes in monocytes and macrophages were distinct for the 2 cytokines. Together, our findings link ligand structural conformation and receptor-binding composition to IFN signaling strength and immune cell specificity, providing mechanistic insight into the functional diversification of type I IFNs in lower vertebrates.
Vaccination is crucial for Atlantic salmon farming, protecting against bacterial and viral infections, such as infectious salmon anemia (ISA) caused by infectious salmon anemia virus (ISAV). Salmon immune responses are highly temperature-dependent and optimal water temperatures are expected to impact vaccine efficacy, while those out of the optimal range may weaken immunity and compromise protection. Additionally, vaccination regime impacts the length of protection against most common pathogens. To assess the impact of temperature and dosing regime on commercial vaccination, we evaluated two commercial multivalent vaccines in salmon reared at 8°C, 12°C, and 15°C. Fish were intraperitoneally injected with 100 μL of either vaccine 1 (V1) or vaccine 2 (V2). Half of each group received a booster after, between 700-750-degree days (dd), including groups that were boosted with the opposite vaccination (V1/V2; V2/V1). After an additional ∼650 dd post boost (∼1400 dd post first vaccination), ten fish from each group were sampled for serum IgM detection. One week later, donor fish previously infected with ISAv (ISAV-HPR4 at TCID50 of 1x105/ml) were introduced to cohabitation tanks at a 6.5:1 ratio of cohabitants to donors. Post-infection, survival rate was recorded, and serum samples were collected for specific-IgM detection, as well as head kidney to determine ISAv load. The results show that elevated rearing temperatures (12-15°C) consistently enhanced specific antibody responses against ISAv, A. salmonicida, and V. anguillarum, whereas cold conditions (8°C) limited or delayed antibody-mediated immune responses in pre-exposed fish. Booster vaccinations and higher temperatures effectively increased and maintained IgM levels in pre-exposed Atlantic salmon, compensating for low-temperature suppression. Survival and viral load data further highlight the interaction between temperature and immune protection, with fish hold at warmer temperatures during vaccination exhibiting higher survival and more efficient ISAv clearance. These findings demonstrate that water temperature and vaccination strategy, including regimen and formulation, critically influence adaptive immunity in Atlantic salmon. Aligning vaccination protocols with seasonal and environmental conditions can maximize protection and limit pathogen persistence.
In mammals, insulin-like growth factor (IGF)-1 and IGF-2 regulate postnatal growth and embryonic development, respectively. In fish, as in mammals, IGF-1 promotes somatic growth, whereas evidence suggests the possible involvement of IGF-2 in the regulation of somatic growth. However, the measurement of circulating IGF-2 in fish has been hampered, possibly because of interference from IGF-binding proteins (IGFBPs). Extraction of IGFs from IGFBPs is essential before measuring them using immunoassays. We optimized acid-ethanol (AE) extraction method for salmonid IGF-2 and developed a time-resolved fluoroimmunoassay (TR-FIA) using recombinant IGF-2 as the assay component. In TR-FIA, the standard AE-extraction of IGF-1 was not validated for quantifying serum IGF-2 because of the presence of ethanol. Three treatments-vacuum drying, dialysis, and ultrafiltration-were compared after AE extraction, and ultrafiltration by centrifugation was found to be effective. With this extraction method, circulating IGF-2 levels were measured by TR-FIA in fed, fasted, and refed rainbow trout. Serum IGF-2 levels were similar or slightly lower in fed fish than in fasted fish, and refeeding had no positive effect. Serum IGF-2 levels were not correlated with individual growth rates. GH treatment tended to increase circulating IGF-2 levels albeit not statistically significant. Smolting masu salmon had similar but slightly lower serum IGF-2 levels than non-smolting fish, while exhibiting high serum IGF-1 levels. These results suggest that circulating IGF-2 levels were relatively stable. This newly developed assay should facilitate further exploration of the physiological responses and roles of circulating IGF-2 in salmonids.
In fish, early puberty can lead to divergent growth trajectories and increased size dispersion within populations. In European sea bass, this pattern is evident by the end of the second year, with females of greater body weight generally tending to reach puberty earlier, although this association is not always consistent. This study investigates the factors influencing the relationship between growth and reproductive maturation in this teleost fish. Four female groups were defined based on body weight (small, S; large, L) and gonadal development (immature, I; advanced, A): SI, SA, LI, and LA. Results showed that larger fish size did not always correspond with a more advanced gonadal stage (late vitellogenesis). Furthermore, females with a higher condition factor during the spring-summer period preceeding putative gonadal development (PGD; autumn-winter months), exhibited elevated plasma levels of follicle stimulating hormone and insulin-like growth factor 1 (Igf-1). These females showed a higher gonadosomatic index (GSI), as well as elevated plasma levels of 17β-estradiol and vitellogenin during the PGD period. This suggests that body condition before PGD may be critical for fish to subsequently meet reproductive demands, as females likely with insufficient energy reserves were unable to sustain the costs associated with maturation later on (i.e., SI, LI vs. SA, LA). Levels of igf expression were linked to ovarian development and were positively correlated with intraovarian lipid gene expression. Of note, SA and LA females displayed increased ovarian expression of igf-2 and lipoprotein lipase (lpl) in comparison to that of SI and LI females. The ovarian igf-2 correlated with GSI and ovarian expression of lpl, suggesting that igf-2 might potentially promote lipid uptake. These findings indicate that body size alone does not predict ovarian development in European sea bass, whereas lipid accumulation appears critical for triggering gonadal development, emphasizing the role of energy status in the relationship between growth and reproduction in this species.
Intensive farming of Nile tilapia (Oreochromis niloticus) heightens susceptibility to infectious diseases, including motile aeromonad septicemia caused by Aeromonas hydrophila, thereby necessitating sustainable alternatives to antibiotics. This study investigated the effects of dietary supplementation with Ocimum basilicum (sweet basil) essential oil (1% w/w) and Thymus vulgaris (thyme) essential oil (0.5% w/w) on growth performance, hematological parameters, serum biochemistry, innate immune responses, cytokine profiles, and resistance to experimental A. hydrophila challenge. A total of 180 juvenile Nile tilapia (initial weight 13.21 ± 0.16 g) were randomly assigned to three dietary treatment groups (three replicates of 20 fish per treatment) and fed for 60 days: (1) basal control diet, (2) basal diet supplemented with 1% O. basilicum essential oil, and (3) basal diet supplemented with 0.5% T. vulgaris essential oil. Fish then received intraperitoneal A. hydrophila challenge (0.1 mL of 1.5 × 108 CFU mL⁻1). Growth, hematological, serum biochemical, and innate immune parameters (lysozyme, nitric oxide, phagocytic index) were assessed at days 30, 60, and 75 (15 days post-challenge). Differential leukocyte counts and cytokine concentrations (TNF-α, IL-1β, IL-6, IL-2, IL-10) were assessed at day 75, and histopathological analysis of hepatic and cephalic kidney tissues was performed to evaluate the protective effects of dietary supplementation against A. hydrophila-induced tissue damage. Both essential oil supplements significantly enhanced growth performance (final body weight and weight gain), hematological indices (hemoglobin concentration and erythrocyte and total leukocyte counts), serum protein profiles (total protein, albumin, and globulin), and innate immune function (lysozyme activity, nitric oxide production, and phagocytic index) compared to the control group (P ≤ 0.05). Notably, the diet supplemented with 1% O. basilicum essential oil produced substantial responses than the diet supplemented with 0.5% T. vulgaris essential oil across several assessed parameters (P ≤ 0.05). Fish fed the O. basilicum-supplemented diet had the highest levels of both pro-inflammatory (TNF-α, IL-1β, IL-6) and anti-inflammatory (IL-10) cytokines, and the lowest overall mortality rate (2.2 ± 2.2%; relative percent survival [RPS] = 96.7 ± 3.3%). The T. vulgaris group had a mortality rate of 13.3 ± 3.9%; RPS = 82.1 ± 6.4%, and the non-supplemented control group had a mortality rate of 77.8 ± 5.9%. Histopathology confirmed markedly diminished renal and hepatic lesions, coupled with increased activation of melanomacrophage centers in both essential oil-supplemented groups. These results suggest that adding 1% O. basilicum essential oil to the diet of Nile tilapia promotes growth, boosts the immune system, and provides greater protection against A. hydrophila infection than adding 0.5% T. vulgaris essential oil. The findings support the potential use of these phytogenic additives as components of health-management strategies in Nile tilapia culture. However, additional studies under commercial production conditions, including dose-optimization, long-term safety assessments, and economic evaluations, are required before practical application can be recommended.
Aeromonas hydrophila, as an important zoonotic conditional pathogen, can cause various diseases such as bacterial septicemia in fish, as well as acute gastroenteritis in humans. Bacteriophages, due to their significant advantage in combating antibiotic-resistant bacteria, have become a research hotspot for novel biological control tools. This study focused on A. hydrophila phage 003s1, which maintained high activity under a wide range of conditions from pH 3 to 11 and at temperatures from 30 to 60°C. After 60 min of ultraviolet irradiation, 63.97% of the original activity remained. To develop a novel antimicrobial agent against A. hydrophila, the phage 003s1 was loaded into different protective agents and subjected to freeze-drying. It was found that using 1% whey protein (WPI) as the protective agent resulted in the optimal viability of the phage after freeze-drying. When stored at 4°C for 6 months, the titer decreased by only 0.86 log10 PFU/mL. In the zebrafish embryo infection model, after treatment with phage powder at a MOI of 0.1, the survival rate of embryos infected with the minimum lethal dose of A. hydrophila significantly increased to 56.67%. In summary, the phage powder developed in this study not only exhibits excellent storage stability but also demonstrates significant protective effects in the zebrafish infection model, providing an innovative antimicrobial agent for the biological control of A. hydrophila.
Temperature is a key environmental factor regulating physiological, biochemical, and immunological processes in ectothermic fish such as Labeo rohita. Exposure to low-temperature conditions during winter disrupts metabolism, suppresses immunity, and induces oxidative stress, thereby compromising fish health and productivity. Labeo rohita fingerlings (initial weight: 8.35 ± 0.50 g) were randomly distributed into four dietary treatments with three replicates each and fed isonitrogenous (25% crude protein) and isocaloric (16.5 MJ kg-1 gross energy) diets supplemented with graded levels of Wolffia extract (WGE; 0, 0.5, 1.5, and 3.0 g kg-1) for 75 days. The experiment was conducted in outdoor cemented tanks during the winter season (December-March) under natural low-temperature conditions (18-22 °C). Dietary WGE significantly enhanced (P< 0.05) non-specific immune responses, with anti-protease activity increasing by 8.22-12.32%, myeloperoxidase by ~200-300%, respiratory burst by 76.92%, and lysozyme by up to 146.94% compared to the control. Carcass composition improved significantly, with crude protein reaching 18.01% and lipid 4.63%. Serum glucose decreased (15.78-49.26%), while total protein and albumin increased. Liver function markers, including SGOT, SGPT, ALP, and CK-MB, declined significantly, indicating improved hepatic integrity and reduced physiological stress. Antioxidant status improved, with elevated SOD and CAT activities and reduced MDA levels by up to 43.97% (liver) and 46.69% (serum). Gene expression analysis revealed downregulation of stress-related genes (hsp70: 0.042-fold; hsp90: 0.147-fold) and upregulation of immune-related genes (c3: 5.51-fold; tnf-α: 3.37-fold; ifn-γ: 7.73-fold). This study evaluated the protective effects of Wolffia globosa extract on innate immunity, antioxidant defense, and stress- and immune-related gene expression in Labeo rohita. Dietary WGE effectively mitigated cold-induced stress by improving immune function, antioxidant capacity, and metabolic performance. Among the tested levels, 1.5 g kg-1 WGE provided the most balanced physiological benefits, while higher inclusion levels induced stronger immune and antioxidant responses. These findings highlight the potential of Wolffia globosa as a sustainable functional feed additive for enhancing fish resilience under low-temperature conditions.
Warming waters can affect fish community dynamics, potentially exacerbating predator-prey asymmetries in behavioural and physiological metrics of thermal tolerance. Further, differing responses of prey fish species to concurrent predation risk and warming highlight the need to understand these impacts in various contexts, especially with at-risk species. This study explored variation in thermal tolerance between the Threatened pugnose shiner (Miniellus anogenus) and non-threatened congener blackchin shiner (Miniellus heterodon) given concurrent exposure to warming and predation risk via chemical cues. Using a series of three channel-side experiments, we (1) explored whether predator odours from centrarchids affect the thermal tolerance of pugnose shiner and blackchin shiner, (2) explored how conspecific disturbance cues impacted blackchin shiner thermal tolerance and (3) quantified the thermal tolerance of two centrarchid species (largemouth bass (Micropterus nigricans) and pumpkinseed (Lepomis gibbosus)) predators and qualitatively compared this to the two shiner species. We found that blackchin shiner had a lower critical thermal maximum (CTMax) when given predator odours compared to a water control, demonstrating a plastic upper thermal limit in response to risk. In contrast, the CTMax of pugnose shiner did not differ between treatments, indicating that they may lack the capacity to respond to these concurrent stressors. The CTMax of blackchin shiner and pugnose shiner increased over the summer coinciding with increasing water temperature in their home habitat. However, when blackchin shiner received disturbance cue relative to the water control in a separate assay, the effect of seasonal acclimatization on CTMax was reduced. Finally, largemouth bass qualitatively exhibited a CTMax that was higher than the shiners, but pumpkinseed did not. Continuing to explore how predator-prey relationships change with warming waters may strengthen our predictive capacity when examining future climate-change scenarios.
Interleukin-20 (il20) is known to participate in inflammation and host defense in mammals, whereas its immunological role in teleost fish needs to be further explored. In this study, a black carp (Mylopharyngodon piceus) il20 homolog, designated bcil20, was cloned and characterized. The full-length coding sequence of bcil20 encodes a putative protein of 179 amino acids with a predicted signal peptide and a conserved IL-10 family domain. Phylogenetic and structural analyses indicated that bcil20 is evolutionarily conserved. Expression analysis showed that bcil20 was constitutively expressed in multiple tissues and was markedly induced by spring viremia of carp virus (SVCV) and grass carp reovirus (GCRV). Western blot analysis confirmed that bcIl20 was present in both cell lysates and culture supernatants of transfected HEK293T cells, indicating that bcil20 is a secreted cytokine. Functional assays demonstrated that both bcil20 overexpression and treatment with bcIl20-conditioned medium significantly upregulated the expression of inflammatory and antiviral genes in EPC cells. Moreover, bcil20 reduced viral replication and cytopathic effects caused by both SVCV and GCRV. Besides, extracellular bcIl20 conferred dose-dependent antiviral protection. Collectively, these findings identify bcil-20 as a virus-responsive secreted cytokine that links innate immune activation to antiviral defense in black carp, and provide new insight into the function of il20 in teleost fish.