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Probiotics play an important role in growth increment, immune enhancement and stress mitigation in fish. Increasing temperature is a major concern in present aquaculture practices as it markedly deteriorates the health condition and reduces the growth in fish. In order to explore the possibilities of using probiotics as a counter measure for temperature associated problems, a 30 days feeding trial was conducted to study the hemato-immunological and apoptosis response of Labeo rohita (8.3±0.4 g) reared at different water temperatures, fed with or without dietary supplementation of a probiotic mixture (PM) consisting of Bacillus subtilis, Lactococcus lactis and Saccharomyces cerevisiae) (1011 cfu kg(-1)). Three hundred and sixty fish were randomly distributed into eight treatment groups in triplicates, namely, T1(28°C+BF(Basal feed)+PM), T2(31°C+BF+PM), T3(34°C+BF+PM), T4(37°C+BF+PM), T5(28°C+BF), T6(31°C+BF), T7(34°C+BF) and T8(37°C+BF). A significant increase (P<0.01) in weight gain percentage was observed in the probiotic fed fish even when reared at higher water temperature (34-37°C). Respiratory burst assay, blood glucose, erythrocyte count, total serum protein, albumin, alkaline phosphatase and acid phosphatase were significantly higher (P<0.01) in the probiotic fed groups compared to the non-probiotic fed groups. A significant (P<0.01) effect of rearing temperature and dietary probiotic mixture on serum myeloperoxidase activity, HSP70 level and immunoglobulin production was observed. Degree of apoptosis in different tissues was also significantly reduced in probiotic-supplemented groups. Hence, the present results show that a dietary PM could be beneficial in enhancing the immune status of the fish and also help in combating the stress caused to the organism by higher rearing water temperature.

This study was conducted to investigate the effects of selenium nanoparticle (Se-NP) supplementation on the growth performance, carcass composition, antioxidant status, hepatic enzyme activities, and immunity of Cirrhinus mrigala . For this purpose, fish with an average initial weight of 7.44 ± 0.04 g were fed five experimental diets containing 0 (control), 0.25, 0.5, 1, and 2 mg kg -1 Se-NPs diets for 90 days. The analysed selenium (Se) contents of the diets were 0.35, 0.64, 0.92, 1.43, and 2.39 mg kg -1 . Twenty five fish were randomly distributed in each of 5 aquarium (36 × 23.7 × 24.3 inches) in triplicate. The results showed that supplementation with Se up to 0.92 mg/kg significantly increased (p<0.05) weight gain, weight gain% (WG%), and specific growth rate (SGR) by 34%, 33%, and 16%, respectively, compared to the control diet. Dietary Se concentrations up to 0.92 mg/kg significantly increased the crude protein and crude fat and reduced (p<0.05) the moisture content as compared to the control group. Fish fed 0.92 mg kg -1 Se had significantly lower malondialdehyde (MDA) contents and higher activities of catalase, superoxide dismutase, and glutathione peroxidase in liver and serum as compared to other experimental diets. Moreover, a significant increase (p<0.05) in the level of serum immunoglobulin and lysozyme (LYZ) activity was recorded in fish fed 0.92 mg/kg Se diet. Moreover, the highest (p<0.05) values of aspartate transaminase (AST) and alanine transaminase (ALT) were recorded in fish fed 2.39 mg/kg Se level. However, serum alkaline phosphatase (ALP) activity remained unaffected by dietary treatment. Broken-line regression analysis indicated that 0.83 mg/kg Se is required for the optimum growth performance of C . mrigala .

The hybrid offspring of barbel chub Squaliobarbus curriculus and grass carp Ctenopharyngodon idella exhibit stronger resistance to the grass carp reovirus (GCRV) infection than grass carp. Toll-like receptors (TLRs) play indispensable roles in the antiviral immunity of fish. In this study, the structures and antiviral immune functions of barbel chub TLR19 (ScTLR19) and grass carp TLR19 (CiTLR19) were compared. The amino acid sequence of ScTLR19 shared high similarity (97.4%) and identity (94.0%) with that of CiTLR19, and a phylogenetic tree revealed the close evolutionary relationship between ScTLR19 and CiTLR19. Protein domain composition analyses showed that ScTLR19 possessed an additional leucine-rich repeat (designated as LRR9) located at amino acid positions 654–677 in the extracellular region, which was absent in CiTLR19. Multiple sequence alignment and three-dimensional structure comparison also indicated that the extracellular regions of ScTLR19 and CiTLR19 exhibited greater differences compared to their intracellular regions. Molecular docking revealed that the extracellular region of ScTLR19 (docking score = −512.31) showed a stronger tendency for binding with polyI:C, compared to the extracellular region of CiTLR19 (docking score = −474.90). Replacing LRR9 in ScTLR19 with the corresponding amino acid sequence from CiTLR19 reduced the binding activity of ScTLR19 to polyI:C, as confirmed by an ELISA. Moreover, overexpression experiments suggested that ScTLR19 could regulate both the IRF3–TRIF and IRF3–MyD88 signaling pathways during GCRV infection, while CiTLR19 only regulated the IRF3–MyD88 signaling pathway. Importantly, replacing LRR9 in ScTLR19 with the corresponding amino acid sequence from CiTLR19 altered the expression regulation on IRF3, MyD88, and TRIF during GCRV infection. These findings collectively reveal the structural and functional differences between ScTLR19 and CiTLR19, and they may provide data to support a deeper understanding of the molecular mechanisms underlying the differences in GCRV resistance between barbel chub and grass carp, as well as the genetic basis for the heterosis of GCRV resistance in their hybrid offspring.

Mammalian interleukin-22 (IL-22) attenuates organismal injury by inhibiting reactive oxygen species (ROS) and impeding the NLRP3 inflammasome activation. However, the role of fish IL-22 in this process remains unclear. We characterized MaIL-22, an IL-22 homolog in blunt snout bream ( Megalobrama amblycephala ). Despite its low sequence identity, it shares conserved structures and close evolutionary relationships with other teleost IL-22s. Furthermore, Aeromonas hydrophila ( A. hydrophila ) infection leads to tissue injury in M. amblycephala immune organs and concomitantly altered Mail-22 mRNA expression, suggesting that MaIL-22 was involved in the antimicrobial immune response. To explore MaIL-22’s biological functions, we produced recombinant MaIL-22 (rMaIL-22) protein and demonstrated it significantly enhanced the survival of M. amblycephala post- A. hydrophila infection. To unravel its protective mechanisms, we explored the ROS/NLRP3 inflammasome axis and its downstream signaling responses. The results showed that rMaIL-22 treatment significantly elevated antioxidant enzyme (T-SOD, CAT and GSH-PX) activities to inhibit MDA activity and scavenge ROS in visceral tissues. Meanwhile, rMaIL-22 impeded the activation of NLRP3 inflammasome by suppressing NLRP3 protein and mRNA expression. This indicated that rMaIL-22 contributed to inhibit A. hydrophila -induced activation of the ROS/NLRP3 inflammasome axis. Consistent with these findings, rMaIL-22 treatment attenuated the expression of proinflammatory cytokines ( il-1β , tnf-α and il-6 ) and proapoptotic genes ( caspase-3 and caspase-8 ) while promoting antiapoptotic genes ( bcl-2b and mcl-1a ) expression, ultimately mitigating tissue injury in visceral tissues. In conclusion, our research underscores MaIL-22’s key role in microbial immune regulation, offering insights for developing IL-22-targeted therapies and breeding programs.

The effect of chrysophanic acid (CA) (2, 4, and 8 mg kg−1) on the immunity and immune-related gene profile of Catla catla against Aeromonas hydrophila is reported. In both control and treated groups fed with 2 mg kg−1 (2 CA), the phagocytosis, hemolytic, myeloperoxidase content, and superoxide anion production decreased significantly between 6th and 8th weeks, whereas when fed with 4 mg kg−1 CA (4 CA) the H2O2 production and nitric oxide synthase increased significantly between 4th and 8th week. When fed with 2 CA and 4 CA diets, the total protein, bactericidal, and antibody titer increased significantly from the 4th week onwards. When fed with 2 CA, the IL-1β and IL-10 mRNA expression of head kidney leucocytes were significant between weeks 6 and 8. The expressions of toll-like receptors significantly increased when fed with a 4 CA diet from 4th week onwards. The 4 CA group significantly increased in TNF-α, TNF receptor-associated factor 6 (NOD), which influences protein expression, after the 4th week. The mRNA transcription of MHCI, lysozyme-chicken and goose type expressions significantly increased in 4 CA group within the 4th week. In summary, the dietary administration of 4 mg kg−1 of CA (4 CA) provides better immunity and enhances the up-regulation of immune-related genes in Catla against A. hydrophila.

Microbes are the most significant ubiquitous pathogens that cause serious infections in freshwater fish, leading to tremendous economic losses. The present study was designed to investigate the extent of changes in cytokine expression, hemato-biochemical parameters, and tissue histology of Cirrhinus mrigala (C. mrigala) challenged with Pseudomonas aeruginosa (P. aeruginosa) and Fusarium oxysporum (F. oxysporum). Fish were divided into three major groups: control, P. aeruginosa-challenged, and F. oxysporum-challenged. The infection in both challenge assays was allowed to progress until 7 days post infection. Upregulated expression of TNF-α and IL-1β was found in blood, gills, livers, and kidneys of the challenged fish. Significant differences were noted in hematological parameters of challenged fish. Alanine aminotransferase, aspartate aminotransferase, and alkaline aminotransferase levels also showed significant differences in infected and control groups. An increase in serum albumin and globulin and a decrease in total protein were noted in infected groups as compared to the control group. Severe histological alterations were noted in gill, liver, and kidney tissues of the infected groups as compared to control. The order of histological alteration index for P. aeruginosa challenge was liver > kidney > gills, and for F. oxysporum challenge it was kidney > liver > gills. These changes in fish infected by P. aeruginosa and F. oxysporum can be used as an effective and subtle index to monitor the physiological and pathological conditions of fish.

Aphanomyces invadans , the causative agent of epizootic ulcerative syndrome, is one of the most destructive pathogens of freshwater fishes. To date, the disease has been reported from over 160 fish species in 20 countries and notably, this is the first non-salmonid disease that has resulted in major impacts globally. In particular, Indian major carps (IMCs) are highly susceptible to this disease. To increase our knowledge particularly with regards to host immune response against A. invadans infection in a susceptible host, the gene expression profile in head kidney of A. invadans -infected and control rohu, Labeo rohita was investigated using RNA sequencing. Time course analysis of RNA-Seq data revealed 5608 differentially expressed genes, involved among others in Antigen processing and presentation, Leukocyte transendothelial migration, IL-17 signaling, Chemokine signaling, C-type lectin receptor signaling and Toll-like receptor signaling pathways. In the affected pathways, a number of immune genes were found to be downregulated, suggesting an immune evasion strategy of A. invadans in establishing the infection. The information generated in this study offers first systematic mechanistic understanding of the host–pathogen interaction that might underpin the development of new management strategies for this economically devastating fish-pathogenic oomycete A. invadans .

An effective alternative approach to combat aquaculture challenges is the strategic application of bioresources, which not only mitigate disease ailment but also optimize fish growth. Hence, current research was undertaken to highlight the synergic role of bioresources such as plant immunostimulant Ocimum sanctum along with potent gut-derived probiotic Bacillus cereus strain SL1 (Gen Bank Accession Number: FJ627945.1) on mrigal ( Cirrhinus mrigala) growth, antioxidant status, gut histopathology, and immune response. For 90 days, fingerlings (average weight 6.8 ± 0.5 g) were fed on diets having varying concentrations of O. sanctum and B. cereus . After the completion of the feeding trial, various growth, immunity, and histological and antioxidant metrics were evaluated according to standard procedures. In comparison to the control and other treatment groups, T3 group showed a significant ( P  < 0.05) increase in growth parameters, antioxidant enzymatic activity, and hematological and immunological parameters. In addition to it, supplementation of both B. cereus and O. sanctum also upregulated the antioxidant-related gene expressions, such as hepatic catalase gene by 1.89–3.00 folds, hepatic SOD-1 by 4.46–7.52 folds, and GPx-1of the liver by 1.56–1.95 folds. For 10 days, fingerlings were challenged with the pathogenic bacterium Aeromonas hydrophila ( MTCC-1739), and maximum survival rate (77.77%) was reported in fingerlings of T3 treatment. Further histopathological studies of gut tissues affirm that O. sanctum and B. cereus play a synergic role in the protection of digestive organs from the pathogenic bacterium A. hydrophila. These results suggest that O. sanctum and B. cereus synergically improved the growth performance, immunity, antioxidant status, and gut histology of C. mrigala leading to its sustainable culture.

Multidrug-resistant (MDR) bacterial pathogens represent an escalating challenge to sustainable aquaculture, particularly in high-value freshwater species such as Labeo rohita , a cornerstone of South Asian aquaculture. This study provides the first comprehensive integration of genomic, immunological, and microbiome analyses to characterize Proteus penneri as an emerging MDR pathogen associated with severe disease manifestations in L. rohita , including exophthalmia, ulceration, and hemorrhage. Robust identification through biochemical assays, 16S rRNA sequencing, and phylogenetic analysis confirms the clinical relevance of this isolate. Functional assays demonstrated pronounced virulence, evidenced by hemolysin activity, extensive histopathological damage, and dose-dependent mortality, underscoring its pathogenic capacity in vivo . The observed resistance to multiple frontline antibiotic classes, including tetracyclines, macrolides, and carbapenems, highlights a critical therapeutic limitation in aquaculture settings. Genomic analysis further revealed a diverse repertoire of antimicrobial resistance genes, virulence determinants (notably biofilm formation and secretion systems), and mobile genetic elements, suggesting a strong potential for persistence, adaptability, and horizontal gene transfer. Infection-associated gut microbiome disruption, marked by elevated MAR indices and enrichment of virulence-associated taxa, indicates that P. penneri not only exploits host tissues but also reshapes the microbial ecosystem in ways that may exacerbate disease severity and resistance dissemination. Concurrently, heightened serum cortisol, C3 , and Hsp70 levels, along with transcriptional upregulation of key immune and stress-related genes ( hsp70, nod, il6, sod, c3 , and myd88 ), reflect an intense pro-inflammatory and physiological stress response. In silico docking analyses implicating myd88 –lipopolysaccharide interactions provide mechanistic insight into potential immune-modulatory strategies employed by the pathogen. Collectively, these findings delineate a multifactorial basis for P. penneri virulence and MDR, emphasizing its significance as an emerging aquaculture pathogen. Future research should prioritize functional validation of key virulence and resistance genes, longitudinal surveillance to assess transmission dynamics and AMR spread, and experimental evaluation of alternative disease mitigation strategies, including probiotics, phage therapy, and immune-modulating interventions, to reduce antibiotic reliance and enhance fish health resilience in aquaculture systems.

To explore the feasibility of Musa acuminata (banana) peels as a feed additive, effects of banana peel flour (BPF) on the growth and immune functions of Labeo rohita were evaluated. Diets containing five different concentrations of BPF (0% [basal diet], 1% [B1], 3% [B3], 5% [B5], and 7% [B7]) were fed to the fish (average weight: 15.3 g) for 60 days. The final weight gain and specific growth rate were higher ( P < 0.05 ) in the B5 group. The most significant improvements in immune parameters such as lysozyme, alternative complement pathway, leukocyte phagocytic, superoxide dismutase, and catalase activities were observed in the B5 group. However, the B5 group exhibited the lowest malondialdehyde activity. IgM and glutathione peroxidise activities were significantly elevated in the treatment groups, except in B1, after only 30 days of feeding. Of the examined cytokine-related genes, IL-1β, TNF-α, and HSP70 were upregulated in the head kidney and hepatopancreas, and expressions were generally higher in the B3 and B5 groups. Moreover, B5 group challenged with Aeromonas hydrophila 60 days after feeding exhibited the highest survival rate (70%; P < 0.05 ). These results suggest that dietary BPF at 5% could promote growth performance and strengthen immunity in L. rohita.