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The aquaculture industry faces significant challenges due to bacterial infections caused by Edwardsiella tarda , Photobacterium damselae , and Vibrio harveyi . The extensive use of traditional antibiotics, has resulted in widespread antibiotic resistance. This study aimed to investigate the antibacterial potential of the brown seaweed Eisenia bicyclis , particularly its synergistic effects with antibiotics against these fish pathogenic bacteria. E. bicyclis were processed to obtain methanolic extracts and fractionated using different polar solvents. The antibacterial activities of these extracts and fractions were assessed through disc diffusion and minimum inhibitory concentration (MIC) assays. The study further evaluated the antibiotic susceptibility of the bacterial strains and the synergistic effects of the extracts combined with erythromycin and oxyteteracycline using the fractional inhibitory concentration index. Results showed that the ethyl acetate (EtOAc) fraction of E. bicyclis methanolic extract exhibited the highest antibacterial activity. The combination of the EtOAc fraction with erythromycin significantly enhanced its antibacterial efficacy against the tested strains. This synergistic effect was indicated by a notable reduction in MIC values, demonstrating the potential of E. bicyclis to enhance the effectiveness of traditional antibiotics. The findings suggest that E. bicyclis extracts, particularly the EtOAc fraction, could serve as a potent natural resource to counteract antibiotic resistance in aquaculture.

This study aimed to conduct a multidisciplinary investigation integrating detailed morphology, molecular characterization, water parameters, histopathology alteration, and the trials of treatment of Clinostomum spp. In this study, 300 Nile tilapia (Oreochromis niloticus) were collected from the farmed and wild Nile River at Al Bahr Al Aazam, Giza Governorate to assess Clinostomid infection prevalence. Fish and water samples were collected from private fish farms, and water drains at Dakahlia, and Giza, Egypt. Analysis of the water revealed inadequate water quality, particularly in the fish farms. Snails and piscivorous birds were abundant at fish collection sites. The recovered Clinostomid MCs morphological characteristics and COI gene sequence analysis identified them as Clinostomum complanatum, C. phalacrocoracis, and Euclinostomum heterostomum. Clinostomid MCs disturbed the fish's hematological and biochemical blood parameters. Bath treatment of parasitized fish with praziquantel (2 mg/L for 24 h) revealed a significant reduction in the number of vital MCs vs. infected fish (non-treated). Praziquantel (PZQ) is an effective and safe therapy for controlling Clinostomid infections affecting farmed Nile tilapia. The current findings indicate a link between poor environmental conditions and Clinostomum infections in tilapia. The study highlights the impacts of Clinostomid MCs on fish health and recommends bath treatment with PZQ as an efficient control method for these dangerous parasites to protect human and fish health., (© 2024. The Author(s).)

In the aquaculture industry, biocides are routinely used to treat parasitosis in fish, and researchers are continually developing sustainable alternatives that can replace these harsh chemicals. In this context, the objective of this study was to evaluate the effectiveness of a new natural compound, BiokosTM, for the treatment against Epistylis sp. in Carassius auratus fish. The infestation was identified by the presence of whitish plaques on the integument of five animals, and the diagnosis was confirmed through skin scrapings. BiokosTM is a lipopeptide derived from the bacteria Pseudomonas that can destroy the functionality of the cell membrane of ciliated protozoa. The action of BiokosTM does not harm animals and the environment because the compound degrades into amino acids and fatty acids within days. A 0.15 m3 (150 L) aquarium was treated with an Ich-AwayTM water conditioner manufactured by the Danish company Sundew ApS, which has BiokosTM as the active ingredient. Six tablets were added to the water daily for two days, and new skin scrapings were performed. The fish were clinically well and no longer possessed lesions or parasites. The results obtained indicate that BiokosTM can be an innovative and more sustainable alternative for controlling epistyliasis in ornamental fish.

Sarmentosin (SA) and Quercetin (QC) are two active components of Sedum Sarmentosum Bunge, which is a traditional Chinese herbal medicine. This study aimed to investigate the role and regulatory mechanism of SA and QC in fatty liver of Genetic Improvement of Farmed Tilapia (GIFT) tilapia. GIFT tilapia were randomly divided into two groups with three replicates per treatment (30 fish in each replicate): normal diet group (average weight 3.51±0.31 g) and high-fat diet group (average weight 3.44±0.09 g). After 8 weeks feeding trial, growth index, lipid deposition, and biochemical indexes were measured. Lipid deposition, and lipid and inflammation-related gene expression were detected in a primary hepatocyte model of fatty liver of GIFT tilapia treated with SA or QC. Our results showed that high-fat diet caused lipid deposition and peroxidative damage in the liver of GIFT tilapia. The cell counting kit-8 assay results indicated that 10 μM SA and 10 μM of QC both had the least effect on hepatocyte proliferation. Moreover, both 10 μM of SA and 10 μM of QC showed lipolytic effects and inhibited the expression of lipid-related genes (FAS, Leptin, SREBP-1c, and SREBP2) in fatty liver cells. Interestingly, QC induced autophagosome-like subcellular structure and increased the expression of IL-8 in fatty liver cells. In conclusion, this study confirmed that SA and QC improved fatty liver caused by high-fat diet, providing a novel therapeutic approach for fatty liver of GIFT tilapia., Competing Interests: NO authors have competing interests., (Copyright: © 2024 Guo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The global surge in multidrug-resistant bacteria owing to antibiotic misuse and overuse poses considerable risks to human and animal health. With existing antibiotics losing their effectiveness and the protracted process of developing new antibiotics, urgent alternatives are imperative to curb disease spread. Notably, improving the bactericidal effect of antibiotics by using non-antibiotic substances has emerged as a viable strategy. Although reduced nicotinamide adenine dinucleotide (NADH) may play a crucial role in regulating bacterial resistance, studies examining how the change of metabolic profile and bacterial resistance following by exogenous administration are scarce. Therefore, this study aimed to elucidate the metabolic changes that occur in Edwardsiella tarda ( E. tarda ), which exhibits resistance to various antibiotics, following the exogenous addition of NADH using metabolomics. The effects of these alterations on the bactericidal activity of neomycin were investigated. NADH enhanced the effectiveness of aminoglycoside antibiotics against E. tarda ATCC15947, achieving bacterial eradication at low doses. Metabolomic analysis revealed that NADH reprogrammed the ATCC15947 metabolic profile by promoting purine metabolism and energy metabolism, yielding increased adenosine triphosphate (ATP) levels. Increased ATP levels played a crucial role in enhancing the bactericidal effects of neomycin. Moreover, exogenous NADH promoted the bactericidal efficacy of tetracyclines and chloramphenicols. NADH in combination with neomycin was effective against other clinically resistant bacteria, including Aeromonas hydrophila, Vibrio parahaemolyticus , methicillin-resistant Staphylococcus aureus , and Listeria monocytogenes . These results may facilitate the development of effective approaches for preventing and managing E. tarda -induced infections and multidrug resistance in aquaculture and clinical settings.

In aquaculture worldwide, most of the chemotherapeutic agents used for disease control and treatment are unregulated chemical products derived from agriculture. In this study, we investigated the efficacy of therapeutic baths with albendazole against the monogeneans Anacanthorus spathulatus, Notozothecium janauachensis and Mymarothecium boegeri, which infest the gills of Colossoma macropomum, and the hematological and histopathological effects of this anthelmintic agent on these fish. Albendazole at a concentration of 500 mg/L was used in three baths of 24 hours each, with intervals of 24 hours between these baths. Three replications of this treatment were used, and the control group consisted of water from the cultivation tank. Afterwards, hematological, histopathological and parasitological analyses were conducted. We found that the therapeutic baths with albendazole at 500 mg/L presented high efficacy (94.9%) against monogeneans de C. macropomum and caused few physiological or histopathological alterations. Therefore, baths with albendazole at 500 mg/L, as used in this strategy, can be recommended for controlling and treating infections by monogeneans in C. macropomum.

Tilapia is one of the most important farmed fish in the world and the most cultivated in Brazil. The increase of this farming favors the appearance of diseases, including bacterial diseases. Therefore, the aim of this study was to evaluate the bactericidal activity of copaiba oil, Copaifera duckei, against Streptococcus agalactiae and Flavobacterium columnare and the dietary effect of copaiba oil on zootechnical performance, hematological, biochemical, immunological, and histological analysis before and after an intraperitoneal infection (body cavity) with S. agalactiae in Nile tilapia. For this, fish were randomly distributed into 15 fiber tanks in five treatments (0, 0.25, 0.50, 0.75, and 1.0%) and fed with a commercial diet supplemented with copaiba oil for 30 days. After this period, the fish were randomly redistributed for the experimental challenge with S. agalactiae into six treatments (T0, T1, T2, T3, T4, and T5), the fish were anesthetized, and blood samples were collected to assess hematological, biochemical, immunological, and histological parameters. Copaiba oil showed bactericidal activity against Streptococcus spp. and Flavobacterium spp. in vitro. In addition, concentrations of 0.75 and 1.0% of copaiba oil have an anti-inflammatory effect and improve hematological and immunological parameters, increasing leukocyte numbers, albumin, and serum lytic activity. Furthermore, there is an increase in the intestinal villus length and tissue damage in groups at concentrations of 0.75 and 1.0% of copaiba oil. In conclusion, copaiba oil presented bactericidal activity against Streptococcus spp. and Flavobacterium spp. in vitro, and oral supplementation at concentrations of 0.75 and 1.0% compared to the control group enhanced non-specific immune parameters and digestibility in Nile Tilapia., (© 2024. The Author(s).)

Background: Gills monogenean infestation causes significant mortalities in cultured fishes as a result of respiratory manifestation. Medicinal plants are currently being heavily emphasized in aquaculture due to their great nutritional, therapeutic, antimicrobial activities, and financial value., Methods: The current study is designed to assess the effect of garlic (Allium sativum) and onion (Allium cepa) extracts as a water treatment on the hematological profile, innate immunity, and immune cytokines expression besides histopathological features of gills of Nile tilapia (Oreochromis niloticus L.) infected with gills monogenetic trematodes (Dactylogyrus sp.). Firstly, the 96-hour lethal concentration 50 (96 h-LC 50 ) of garlic extract (GE) and onion extract (OE) were estimated to be 0.4 g/ L and 3.54 g/ L for GE and OE, respectively. Moreover, the in-vitro anti-parasitic potential for (GE) was found between 0.02 and 0.18 mg/mL and 0.4 to 1.8 mg/mL for OE. For the therapeutic trial, fish (n = 120; body weight: 40-60 g) were randomly distributed into four groups in triplicates (30 fish/group, 10 fish/replicate) for 3 days. Group1 (G1) was not infected or treated and served as control. G2 was infected with Dactylogyrus spp. and not exposed to any treatment. G3, G4 were infected with Dactylogyrus sp. and treated with 1/ 10 and 1/ 5 of 96 h LC 50 of OE, respectively. G5, G6 were infected with Dactylogyrus sp. and treated with 1/ 10 and 1/ 5 of 96 h LC 50 of GE, respectively., Results: No apparent signs or behaviors were noted in the control group. Dactylogyrus spp. infected group suffered from clinical signs as Pale color and damaged tissue. Dactylogyrus spp. infection induced lowering of the hematological (HB, MCH, MCHC and WBCs), and immunological variables (lysozyme, nitric oxide, serum Anti- protease activities, and complement 3). the expression of cytokine genes IL-ß and TNF-α were modulated and improved by treatment with A. sativum and A. cepa extracts. The obtained histopathological alterations of the gills of fish infected with (Dactylogyrus spp.) were hyperplasia leading to fusion of the gill filament, lifting of epithelial tissue, aneurism and edema. The results indecated that G4 and G5 is more regenarated epithelium in compare with the control group., Conclusion: A. sativum and A. cepa extracts enhance the blood profile and nonspecific immune parameters, and down-regulated the expression level of (IL-1β and TNF-α)., (© 2024. The Author(s).)

This study aimed to evaluate the effect of adding liquid extract of algae (Hypnea musciformis, Grateloupia acuminata, and Sargassum muticum) (HGS) and Magnesium oxide nanoparticles (MgO NPs) using this extract to rear water of Oreochromis niloticus, on improving culture water indices, growth performance, digestive enzyme, hemato-biochemical characters, immune, antioxidative responses, and resistance after challenged by Aeromonas hydrophila with specific refer to the potential role of the mixture in vitro as resistance against three strains bacteria (Aeromonas sobria, Pseudomonas fluorescens, P. aeruginosa) and one parasite (Cichlidogyrus tilapia). The first group represented control, HGS0, whereas the other group, HGS5, HGS10, and HGS15 mL -1 of liquid extract, as well as all groups with 7.5 μg mL -1 MgO-NPs added to culture water of O. niloticus, for 60 days. Data showed that increasing levels at HGS 10 and HGS15 mL -1 in to-culture water significantly enhanced growth-stimulating digestive enzyme activity and a significantly improved survival rate of O. niloticus after being challenged with A. hydrophila than in the control group. The total viability, coliform, fecal coliform count, and heavy metal in muscle partially decreased at HGS 10 and HGS15 mL -1 than in the control group. Correspondingly, the highest positive effect on hemato-biochemical indices was noticed at levels HGS 10 and HGS15 mL -1 . Fish noticed an improvement in immune and antioxidant indices compared to control groups partially at HGS 10 and HGS15 mL -1 . Interestingly, fish cultured in rearing water with the mixture provided downregulated the related inflammatory genes (HSP70, TNF, IL-1β, and IL-8) partially at HGS15 mL -1 . In vitro, the mixture showed positive efficiency as an antibacterial and partially antiparasitic at HGS 10 and HGS15 mL -1 . This study proposes utilizing a mixture of (HGS) and (MgO-NPs) with optimum levels of 10-15 mL -1 in cultured water to improve water indices, growth, health status, and increased resistance of O. niloticus against bacterial and parasitic infection., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. The authors declare the following financial interests/personal relationships which may be considered as potential competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Aquatic fishes are threatened by the strong pathogenic bacterium Nocardia seriolae, which challenges the current prevention and treatment approaches. This study introduces luminogens with aggregation-induced emission (AIE) as an innovative and non-antibiotic therapy for N. seriolae. Specifically, the AIE photosensitizer, TTCPy-3 is employed against N. seriolae. We evaluated the antibacterial activity of TTCPy-3 and investigated the killing mechanism against N. seriolae, emphasizing its ability to aggregate within the bacterium and produce reactive oxygen species (ROS). TTCPy-3 could effectively aggregate in N. seriolae, generate ROS, and perform real-time imaging of the bacteria. A bactericidal efficiency of 100% was observed while concentrations exceeding 4 μM in the presence of white light irradiation for 10 min. In vivo, evaluation on zebrafish (Danio rerio) confirmed the superior therapeutic efficacy induced by TTCPy-3 to fight against N. seriolae infections. TTCPy-3 offers a promising strategy for treating nocardiosis of fish, paving the way for alternative treatments beyond traditional antibiotics and potentially addressing antibiotic resistance., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The misuse of antibiotics and the emergence of antimicrobial resistance (AMR) is a concern in the aquaculture industry because it contributes to global health risks and impacts the environment. This study analyzed the AMR of sentinel bacteria associated with striped catfish ( Pangasisanodon hypophthalmus ) and giant snakehead ( Channa micropeltes ), the two main fish species reared in the pond culture in Cambodia. Phenotypic and genotypic characterization of the recovered isolates from fish, water, and sediment samples revealed the presence of bacteria, such as 22 species belonging to families Aeromonadaceae , Enterobacteriaceae , and Pseudomonadaceae . Among 48 isolates, Aeromonas caviae (n = 2), Aeromonas hydrophila (n = 2), Aeromonas ichthiosmia (n = 1), Aeromonas salmonicida (n = 4) were detected. A. salmonicida and A. hydrophilla are known as fish pathogens that occur worldwide in both fresh and marine water aquaculture. Antibiotic susceptibility testing revealed antibiotic resistance patterns of 24 (50 %) isolates among 48 isolates with higher multiple antibiotic resistance index (> 0.2). All the isolates of Enterobacteriaceae were susceptible to ciprofloxacin. Ciprofloxacin is a frontline antibiotic that is not recommended to use in aquaculture. Therefore, its use has to be strictly controlled. This study expands our knowledge of the AMR status in aquaculture farms which is very limited in Cambodia., Competing Interests: The authors declare there is no conflict., (© 2024 The Authors This is an Open Access article distributed under the terms of the Creative Commons Attribution Licence (CC BY 4.0), which permits copying, adaptation and redistribution, provided the original work is properly cited (http://creativecommons.org/licenses/by/4.0/).)

Background: Oxygen deprivation (OD) is a critical condition that can lead to brain damage and even death. Current hypoxia management approaches are limited in effectiveness. Centella asiatica (CA), known for its neuroprotective properties, offers a potential alternative for OD treatment., Aims: This study aims to investigate the neuroprotective effects of CA on the expression of brain-derived neurotrophic factor (BDNF) and vesicular glutamate transporter 1 (VGLUT1) in zebrafish larvae under oxygen-deficient conditions., Methods: Zebrafish embryos were subjected to low oxygen levels (1.5 mg/l) 0-2 hours post-fertilization (hpf) until 3 days post-fertilization (dpf), simulating the early stages of OD. Subsequent treatment involved varying concentrations of CA (1.25-5 µg/ml) up to 9 days post-fertilization. The expression levels of BDNF and VGLUT1 were measured using PCR methods. Statistical analysis was conducted using a two-way analysis of variance to evaluate the impact of CA on the expression of BDNF and VGLUT1 in zebrafish larvae aged 3 and 9 dpf in oxygen-deprived conditions., Results: CA significantly influenced the expression of BDNF and VGLUT1 under OD ( p < 0.001). An increase in BDNF expression ( p < 0.001) and a decrease in VGLUT1 ( p < 0.01) were observed in zebrafish larvae experiencing OD and treated with CA. There was no significant difference in BDNF and VGLUT1 expression across age variations in zebrafish larvae at 3 dpf and 9 dpf in the treatment groups ( p > 0.05). CA concentration of 2.5 µg/ml effectively enhanced BDNF and reduced VGLUT1 in 3-9 dpf zebrafish larvae., Conclusion: CA demonstrates potential as a neuroprotective agent, modulating increased BDNF expression and reduced VGLUT1 under OD conditions. These findings lay a foundation for further research in developing therapies for oxygen deficiency., Competing Interests: The authors declare that there is no conflict of interest.

The present study aimed to determine the major cause of the high mortality affecting farmed gilthead seabream (Sparus aurata) and controlling this disease condition. Fifteen diseased S. aurata were sampled from a private fish farm located at Eldeba Triangle, Damietta, fish showed external skin hemorrhages, and ulceration. Bacterial isolates retrieved from the diseased fish were identified biochemically as Pseudomonas putida and then confirmed by phylogenetic analysis of the 16 S rRNA gene sequence. P. putida was also isolated from three batches of tilapia-trash feed given to S. aurata. Biofilm and hemolytic assay indicated that all P. putida isolates produced biofilm, but 61.11% can haemolyse red blood cells. Based on the antibiotic susceptibility test results, P. putida was sensitive to florfenicol with minimum inhibitory concentrations ranging between 0.25 and 1.0 µg mL - 1 , but all isolates were resistant to ampicillin and sulfamethoxazole-trimethoprim. Pathogenicity test revealed that P. putida isolate (recovered from the tilapia-trash feed) was virulent for S. aurata with LD 50 equal to 4.67 × 10 7 colony forming unit (CFU) fish - 1 . After intraperitoneal (IP) challenge, fish treated with 10 mg kg - 1 of florfenicol showed 16.7% mortality, while no mortality was recorded for the fish group that received 20 mg kg - 1 . The non-treated fish group showed 46.7% mortality after bacterial challenge. HPLC analysis of serum florfenicol levels reached 1.07 and 2.52 µg mL - 1 at the 5th -day post-drug administration in the fish groups received 10 and 20 mg kg - 1 , respectively. In conclusion, P. putida was responsible for the high mortality affecting cultured S. aurata, in-feed administration of florfenicol (20 mg kg - 1 ) effectively protected the challenged fish., (© 2024. The Author(s).)

The growing increase in the fish farming sector has favored the establishment of bacterial outbreaks caused by Aeromonas hydrophila in several species. The hexane extract of Hesperozygis ringens (HEHR) (Lamiaceae) leaves increased the survival rate of silver catfish (Rhamdia quelen) experimentally infected by A. hydrophila. However, it is noteworthy that no reports have been found on the possible mechanisms of action of this extract in infected fish. This study aimed to evaluate the effect of the HEHR, administered through single immersion bath, on lipid peroxidation and antioxidant defenses in muscle and liver tissue of silver catfish challenged with A. hydrophila. The results showed that the oxidative status of silver catfish was altered, although oxidative stress was not triggered during the experiment. HEHR at 30 mg/L (HEHR30) was not characterized as a pro-oxidant agent in the presence of infection, unlike florfenicol and HEHR at 15 mg/L treatments in some cases. In short, HEHR30 provided an important increase in hepatic catalase activity, characterizing one of the possible mechanisms involved in the greater survival of fish experimentally infected by A. hydrophila. Additionally, HEHR30 did not induce lipid peroxidation, nor reduced antioxidant defenses of silver catfish infected or not by A. hydrophila.

This study was divided into two parts. The first part involved the isolation, and detection of the prevalence and antimicrobial resistance profile of Aeromonas hydrophila, Pseudomonas aeruginosa, and Vibrio species from Nile tilapia fish and marine aquatic water. One hundred freshly dead Nile tilapia fish were collected from freshwater aquaculture fish farms located in Al-Abbassah district, Sharkia Governorate, and 100 samples of marine aquatic water were collected from fish farms in Port Said. The second part of the study focused on determining the in vitro inhibitory effect of dual-combination of AgNPs-H2O2 on bacterial growth and its down regulatory effect on crucial virulence factors using RT-PCR. The highest levels of A. hydrophila and P. aeruginosa were detected in 43%, and 34% of Nile tilapia fish samples, respectively. Meanwhile, the highest level of Vibrio species was found in 37% of marine water samples. Additionally, most of the isolated A. hydrophila, P. aeruginosa and Vibrio species exhibited a multi-drug resistance profile. The MIC and MBC results indicated a bactericidal effect of AgNPs-H2O2. Furthermore, a transcriptional modulation effect of AgNPs-H2O2 on the virulence-associated genes resulted in a significant down-regulation of aerA, exoU, and trh genes in A. hydrophila, P. aeruginosa, and Vibrio spp., respectively. The findings of this study suggest the effectiveness of AgNPs-H2O2 against drug resistant pathogens related to aquaculture., (© 2024. The Author(s).)

Background: Pseudomonas putida is a pathogenic bacterium that induces great losses in fishes, including Nile tilapia (Oreochromis niloticus). Currently, the application of nanomaterials in aquaculture practices has gained more success as it endows promising results in therapies compared to traditional protocols., Objective: Therefore, the current perspective is considered the first report to assess the anti-bacterial efficacy of titanium dioxide nanogel (TDNG) against Pseudomonas putida (P. putida) in Nile tilapia., Methods: The fish (n = 200; average body weight: 47.50±1.32 g) were allocated into four random groups (control, TDNG, P. putida, and TDNG + P. putida), where 0.9 mg/L of TDNG was applied as bath treatment for ten days., Results: Outcomes revealed that P. putida infection caused ethological alterations (surfacing, abnormal movement, and aggression) and depression of immune-antioxidant variables (complement 3, lysozyme activity, total antioxidant capacity, superoxide dismutase, and reduced glutathione content). Additionally, a substantial elevation in hepatorenal biomarkers (aspartate and alanine aminotransferases and creatinine) with clear histopathological changes and immuno-histochemical alterations (very weak BCL-2 and potent caspase-3 immuno-expressions) were seen. Surprisingly, treating P. putida-infected fish with TDNG improved these variables and obvious restoration of the tissue architectures., Conclusion: Overall, this report encompasses the key role of TDNG as an anti-bacterial agent for controlling P. putida infection and improving the health status of Nile tilapia., (© 2024. The Author(s).)

Infectious hematopoietic necrosis virus (IHNV) causes infectious hematopoietic necrosis and severe economic losses to salmon and trout aquaculture worldwide. Currently, the only commercial vaccine against IHNV is a DNA vaccine with some biosafety concerns. Hence, more effective vaccines and antiviral drugs are needed to prevent IHNV infection. In this study, 1,483 compounds were screened from a traditional Chinese medicine monomer library, and bufalin showed potential antiviral activity against IHNV. The 50% cytotoxic concentration of bufalin was >20 µM, and the 50% inhibitory concentration was 0.1223 µΜ against IHNV. Bufalin showed the inhibition of diverse IHNV strains in vitro , which confirmed that it had an inhibitory effect against all IHNV strains, rather than random activity against a single strain. The bufalin-mediated block of IHNV infection occurred at the viral attachment and RNA replication stages, but not internalization. Bufalin also inhibited IHNV infection in vivo and significantly increased the survival of rainbow trout compared with the mock drug-treated group, and this was confirmed by in vivo viral load monitoring. Our data showed that the anti-IHNV activity of bufalin was proportional to extracellular Na + concentration and inversely proportional to extracellular K + concentration, and bufalin may inhibit IHNV infection by targeting Na + /K + -ATPase. The in vitro and in vivo studies showed that bufalin significantly inhibited IHNV infection and may be a promising candidate drug against the disease in rainbow trout., Importance: Infectious hematopoietic necrosis virus (IHNV) is the pathogen of infectious hematopoietic necrosis (IHN) which outbreak often causes huge economic losses and hampers the healthy development of salmon and trout farming. Currently, there is only one approved DNA vaccine for IHN worldwide, but it faces some biosafety problems. Hence, more effective vaccines and antiviral drugs are needed to prevent IHNV infection. In this study, we report that bufalin, a traditional Chinese medicine, shows potential antiviral activity against IHNV both in vitro and in vivo . The bufalin-mediated block of IHNV infection occurred at the viral attachment and RNA replication stages, but not internalization, and bufalin inhibited IHNV infection by targeting Na + /K + -ATPase. The in vitro and in vivo studies showed that bufalin significantly inhibited IHNV infection and may be a promising candidate drug against the disease in rainbow trout., Competing Interests: The authors declare no conflict of interest.

Background: Salinomycin, an antibiotic, have potential as a veterinary drug for fish due to its anti-parasitic activity against several fish parasites. Thus the residual levels of salinomycin in muscles of two significant aquaculture species in Korea, olive flounder and black rockfish, were analyzed using HPLC-MS-MS., Results: The proper method to analyze the residual salinomycin in fish muscles using LC-MS-MS was settled and the method was validated according to CODEX guidelines. The residues in three distinct groups for two fish species were analyzed using the matrix match calibration curves at points of five different times following oral administration. After oral administration, salinomycin rapidly breaks down in both olive flounder and black rockfish. After 7 th days, the average residue in all groups of two fish spp. decreased below limit of quantitation (LOQ)., Conclusion: Due to low residue levels in fish muscles, salinomycin may therefore be a treatment that is safe for both fish and humans. This result could contribute to establishment of MRL (minimal residual limit) for approval of salinomycin for use in aquaculture., (© 2024. The Author(s).)

This study investigated for the first time the effectiveness of therapeutic baths with essential oil (EO) of Piper hispidum against monogeneans Anacanthorus spathulatus, Notozothecium janauachensis, Mymarothecium boegeri and Linguadactyloides brinkmanni from the gills of Colossoma macropomum, as well as the hematological and histological effects on this fish. In therapeutic baths, 100 mg/L of P. hispidum essential oil and two control groups (water from the culture tank and water from the culture tank with 70% alcohol) were exposed for 1 h/day, with intervals of 48 hours for 3 days, and three replicates each were used. Therapeutic baths with 100 mg/L of P. hispidum essential oil had an efficacy of 78.6% against monogeneans. The toxicity of this essential oil was low, since there were a few physiological and histopathological changes that did not compromise the functioning of the gills of the fish. Therefore, 100 mg/L of P. hispidum essential oil was effective for controlling monogeneans in C. macropomum, when short therapeutic baths were used without compromising the health of the exposed fish.

Singapore grouper iridovirus (SGIV) is a virus with high fatality rate in the grouper culture industry. The outbreak of SGIV is often accompanied by a large number of grouper deaths, which has a great impact on the economy. Therefore, it is of great significance to find effective drugs against SGIV. It has been reported that edaravone is a broad-spectrum antiviral drug, most widely used clinically in recent years, but no report has been found exploring the effect of edaravone on SGIV infections. In this study, we evaluated the antiviral effect of edaravone against SGIV, and the anti-SGIV mechanism of edaravone was also explored. It was found that the safe concentration of edaravone on grouper spleen (GS) cells was 50 µg/mL, and it possessed antiviral activity against SGIV infection in a dose-dependent manner. Furthermore, edaravone could significantly disrupt SGIV particles and interference with SGIV binding to host cells, as well as SGIV replication in host cells. However, edaravone was not effective during the SGIV invasion into host cells. This study was the first time that it was determined that edaravone could exert antiviral effects in response to SGIV infection by directly interfering with the processes of SGIV infecting cells, aiming to provide a theoretical basis for the control of grouper virus disease.

Tilapia Lake Virus (TiLV) is a disease that affects tilapia fish, causing a high rate of sudden death at any stage in their life cycle. Unfortunately, there are currently no effective antiviral drugs or vaccines to prevent or control the progression of this disease. Researchers have discovered that the CRM1 protein plays a critical function in the development and spreading of animal viruses. By inhibiting CRM1, the virus's spread in commercial fish farms can be suppressed. With this in mind, this study intended to identify potential antiviral drugs from two different tropical mangrove plants from tropical regions: Heritiera fomes and Ceriops candolleana. To identify promising compounds that target the CRM1 protein, a computer-aided drug discovery approach is employed containing molecular docking, ADME (absorption, distribution, metabolism and excretion) analysis, toxicity assessment as well as molecular dynamics (MD) simulation. To estimate binding affinities of all phytochemicals, molecular docking is used and the top three candidate compounds with the highest docking scores were selected, which are CID107876 (-8.3 Kcal/mol), CID12795736 (-8.2 Kcal/mol), and CID12303662 (-7.9 Kcal/mol). We also evaluated the ADME and toxicity properties of these compounds. Finally, MD simulation was conducted to analyze the stability of the protein-ligand complex structures and confirm the suitability of these compounds. The computational study demonstrated that the phytochemicals found in H. fomes and C. candolleana could potentially serve as important inhibitors of TiLV, offering practical utility. However, further in vivo investigations are necessary to investigate and potentially confirm the effectiveness of these compounds as antiviral drugs against the virus TiLV., Competing Interests: There are no competing interests., (Copyright: © 2023 Sumon et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The aquaculture industry is constantly increasing its fish production to provide enough products to maintain fish consumption worldwide. However, the increased production generates susceptibility to infectious diseases that cause losses of millions of dollars to the industry. Conventional treatments are based on antibiotics and antivirals to reduce the incidence of pathogens, but they have disadvantages, such as antibiotic resistance generation, antibiotic residues in fish, and environmental damage. Instead, functional foods with active compounds, especially antimicrobial peptides that allow the generation of prophylaxis against infections, provide an interesting alternative, but protection against gastric degradation is challenging. In this study, we evaluated a new immunomodulatory recombinant peptide, CATH-FLA, which is encapsulated in chitosan microparticles to avoid gastric degradation. The microparticles were prepared using a spray drying method. The peptide release from the microparticles was evaluated at gastric and intestinal pH, both in vitro and in vivo. Finally, the biological activity of the formulation was evaluated by measuring the expression of il-1β , il-8 , ifn-γ , Ifn-α , and mx1 in the head kidney and intestinal tissues of rainbow trout ( Oncorhynchus mykiss ). The results showed that the chitosan microparticles protect the CATH-FLA recombinant peptide from gastric degradation, allowing its release in the intestinal portion of rainbow trout. The microparticle-protected CATH-FLA recombinant peptide increased the expression of il-1β , il-8 , ifn-γ , ifn-α , and mx1 in the head kidney and intestine and improved the antiprotease activity in rainbow trout. These results suggest that the chitosan microparticle/CATH-FLA recombinant peptide could be a potential prophylactic alternative to conventional antibiotics for the treatment of infectious diseases in aquaculture.

Many governments have approved the use of oxytetracycline as an antibiotic additive to food fish, with oxytetracycline now routinely used in many nations. However, oxytetracycline is known to have immunosuppression impacts. We, therefore, evaluated the immunological, antioxidative, and histopathological status of Nile tilapia fed a diet containing silymarin (100 mg/kg fish feed) for 0, 2, 4, 6, and 8 weeks. The protective effects of silymarin against Aeromonas hydrophila (A. hydrophila) infection and oxytetracycline treatment were evaluated. Blood parameters (erythrocyte count, white blood cell count, hemoglobin, and packed cell volume) improved over time in fish fed on dietary silymarin. Serum levels of alanine aminotransferase (ALT) were lower in fish fed on dietary silymarin, whereas serum levels of aspartate transferase (AST)and alkaline phosphatase (ALK) were unchanged. Dietary silymarin affected serum lipid profiles as decreases in serum triglyceride and low-density lipoprotein cholesterol levels and a trend toward lower cholesterol levels, whereas serum high-density lipoprotein cholesterol levels were increased compared to fish fed on the control diet. Dietary silymarin resulted in an increase of serum total protein levels and globulin fractions. Significant and progressive increases in catalase and glutathione peroxidase levels were observed after six weeks of feeding on a dietary silymarin before decreasing to control levels at the end of the experimental period. Fish fed on dietary silymarin, interleukin-1 and fish tumor necrosis factor-alpha were upregulated in hepatic tissues; however, interleukin-10 levels decreased to comparable levels to controls after eight weeks. Fish infected with A. hydrophila displayed septicemia (opaque eye, hemorrhagic ulcers, dentated fins, hepatomegaly, and splenomegaly). Reduced mortality was observed in Nile tilapia infected with A. hydrophila and fed a diet containing silymarin, indicating that silymarin improves fish responses to oxytetracycline with a 37% reduction in mortality., (© 2023. The Author(s).)

The vitamin D receptor (VDR) plays a pivotal role in the biological actions of vitamin D (VitD). However, little is known about the functions of VDR in the production of viral inclusion bodies (VIBs). Using a representative strain of grass carp reovirus (GCRV) genotype I, GCRV-873, we show that GCRV-873 recruits grass carp Vdrs for promoting the production of VIBs in the absence of VitD. Inhibition of cholesterol synthesis by lovastatin impairs the production of VIBs and blocks the effects of grass carp Vdrs in promoting the production of VIBs in the absence of VitD. Furthermore, grass carp Vdrs are found to form the Vdra-Vdrb heterodimer, which is vital for 3-hydroxy-3-methylglutaryl-coenzyme A reductase ( hmgcr )-dependent cholesterol synthesis and GCRV replication. Intriguingly in the presence of VitD, grass carp Vdra but not Vdrb forms the heterodimer with the retinoid X receptor beta b (Rxrbb), which induces the transcription of those genes involved in the RIG-I-like receptor (RLR) antiviral signaling pathway for inhibiting GCRV infection. Furthermore, the VitD-activated Vdra-Vdrb heterodimer attenuates the transcription of the RLR antiviral signaling pathway induced by VitD. In the presence of VitD, a balance between the Vdra-Rxrbb heterodimers as coactivators and Vdra-Vdrb heterodimers as corepressors in affecting the transcriptional regulation of the RLR antiviral signaling pathway may eventually determine the outcome of GCRV infection. Transfection with VitD can abolish the effect of grass carp Vdrs in promoting GCRV replication in a dose-dependent manner. Taken together, these findings demonstrate that GCRV utilizes host Vdrs to increase hmgcr -dependent cholesterol synthesis for promoting its replication, which can be prevented by VitD treatment. IMPORTANCE Grass carp reovirus (GCRV) is the causative agent of grass carp hemorrhagic disease, which seriously harms freshwater fish. Although many positive or negative regulators of GCRV infection have been identified in teleosts, little is known about the molecular mechanisms by which GCRV utilizes host factors to generate its infectious compartments beneficial for viral replication and infection. Here, we show that in the absence of VitD, the GCRV-873 strain utilizes host vitamin D receptors Vdra/Vdrb to increase hmgcr -dependent cholesterol synthesis for promoting the production of VIBs, which are important functional sites for aquareovirus replication and assembly. The negative regulation of Vdrs during viral infection can be prevented by VitD treatment. Thus, this present work broadens understanding of the pivotal roles of Vdrs in the interaction between the host and GCRV in the absence or presence of VitD, which might provide a rational basis for developing novel anti-GCRV strategies., Competing Interests: The authors declare no conflict of interest.

Aporocotylid blood flukes Cardicola forsteri and C. orientalis are an ongoing health concern for Southern Bluefin Tuna (SBT), Thunnus maccoyii , ranched in Australia. Therapeutic application of praziquantel (PZQ) has reduced SBT mortalities, however PZQ is not a residual treatment therefore reinfection can occur after the single treatment application. This study documents the epidemiology of Cardicola spp. infection in ranched SBT post treatment over three ranching seasons (2018, 2019 and 2021). Infection prevalence (percentage of SBT affected) and intensity (parasite load) was determined by adult fluke counts from heart, egg counts from gill filaments and the use of specific quantitative polymerase chain reaction (qPCR) for detection of C. forsteri and C. orientalis ITS-2 DNA in SBT hearts and gills. SBT Condition Index decreased as intensity of Cardicola spp. DNA in SBT gills increased, suggesting blood fluke infection had a negative effect on SBT growth (Spearman's r = -0.2426, d.f. = 138, p = 0.0041). Prevalence and intensity of infection indicated PZQ remained highly effective at controlling Cardicola spp. infection in ranched SBT, 10 years after PZQ administration began in this industry. Company A had the highest prevalence and intensity of Cardicola spp. infection in 2018, and Company G had the highest in 2019. No consistent pattern was seen in 2021. Overall, intensity of infection did not increase as ranching duration increased post treatment. Results from this study improve our knowledge of the biology of blood flukes and helps the SBT industry to modify or design new blood fluke management strategies to reduce health risks and improve performance of SBT., Competing Interests: Barbara F. Nowak is an Academic Editor for PeerJ. Kirsten Rough is an employee of the Australian Southern Bluefin Tuna Industry Association., (© 2023 Power et al.)

A novel antibacterial immunostimulant using Platinum nanoparticles (PtNPs) and lectin from Metapenaeus dobsoni (Md-Lec) was developed. The Md-Lec and PtNPs (Pt-lec) hybrid formed through non-covalent interaction exhibits antimicrobial activity against fish specific pathogens by affecting membrane integrity and producing excess reactive oxygen species. The therapeutic efficacy of Pt-lec was demonstrated through rescuing Aeromonas hydrophila infected Nile Tilapia. Pt-lec prevents the infection spreading and reduces the bacterial bioburden in less than 12 h, and as a result of this the fish were restored to normalcy. To assess immunostimulation, we studied the expression of three different immune related genes, namely LEC, Myd88 and COX-2 in the gills, liver, spleen and kidney of fish under various experimental conditions. Our results showed that Pt-lec treatment appeared to be better when compared to lectin alone in enhancing the expression of Myd88 and COX-2, but LEC was not as expected. These results suggest that Pt-lec has the ability to protect Nile Tilapia against bacterial infection by restricting bacterial bioburden through their direct effects on the bacterial membrane and indirectly through their effects on host immune-related gene expression. This hybrid could have potential "green" application in fish farming in rescuing infected animals when compared to widely and unregulated antibiotics., (© 2023. The Author(s).)

Cyprinid herpesvirus 3 (CyHV-3) can cause severe disease in koi and common carp ( Cyprinus carpio ). Currently, no effective treatment is available against CyHV-3 infection in koi. Both LSD1 and JMJD2 are histone demethylases (HD) and are critical for immediate-early (IE) gene activation essential for lytic herpesvirus replication. OG-L002 and ML324 are newly discovered specific inhibitors of LSD1 and JMJD2, respectively. Here, HD inhibitors were compared with acyclovir (ACV) against CyHV-3 infection in vitro and in vivo. ML324, at 20-50 µM, can completely block ~1 × 10 3 PFU CyHV-3 replication in vitro, while OG-L002 at 20 µM and 50 µM can produce 96% and 98% inhibition, respectively. Only about 94% inhibition of ~1 × 10 3 PFU CyHV-3 replication was observed in cells treated with ACV at 50 µM. As expected, CyHV-3 IE gene transcription of ORF139 and ORF155 was blocked within 72 h post-infection (hpi) in the presence of 20 µM ML324. No detectable cytotoxicity was observed in KF-1 or CCB cells treated for 24 h with 1 to 50 µM ML324. A significant reduction of CyHV-3 replication was observed in ~6-month-old infected koi treated with 20 µM ML324 in an immersion bath for 3-4 h at 1-, 3-, and 5-days post-infection compared to the control and ACV treatments. Under heat stress, 50-70% of 3-4-month-old koi survived CyHV-3 infection when they were treated daily with 20 µM ML324 in an immersion bath for 3-4 h within the first 5 d post-infection (dpi), compared to 11-19% and 22-27% of koi in the control and ACV treatments, respectively. Our study demonstrates that ML324 has the potential to be used against CyHV-3 infection in koi.

Monogenean infestations can cause high mortality in farmed fish and therefore significant economic losses. The present study investigated the efficacy of albendazole in therapeutic baths against monogeneans of Piaractus brachypomus Cuvier, 1818 and Megaleporinus macrocephalus (Garavello & Britski, 1988). For both fish, a 24 hours therapeutic bath with albendazole concentrations of 150, 300 and 500 mg/L were tested against monogeneans from the gills. The baths had an efficacy from 61.4 ± 32.9 (95%CI=64.5) against monogeneans of P. brachypomus, and from 95.4 ± 5.6 (95%CI=10.9) against monogeneans of M. macrocephalus. In P. brachypomus, the hepatosomatic index (HSI) in fish exposed to 150 mg/L of albendazole was higher than in fish exposed to 300 mg/L. The splenosomatic index (SSI) values in fish exposed to 150 mg/L of albendazole were lower than in fish exposed to 300 mg/L. In M. macrocephalus, the HSI and SSI values decreased in treatments with 150, 300 and 500 mg/L of albendazole to control and treat infestations by monogeneans. For M. macrocephalus, 150 mg/L of albendazole can be used to control and treat infestations by monogeneans, while for P. brachypomus 500 mg/L of albendazole can be used in a 24 hours bath.

Objective: To evaluate the effect of a multidose acyclovir protocol on koi herpesvirus (KHV) viral load and mortality in a cohabitation challenge., Animals: 180 koi fish., Procedures: Forty fish (shedders) were immersed in a 0.5 KHV plaque-forming units/mL static bath for 8 hours. Mock shedders were treated similarly but exposed to cell culture media. KHV shedders were then transferred into 8 tanks (5 shedders per tank) containing 10 naïve fish (cohabitants) each. Fish in the acyclovir group (AT) received a 10 mg/kg acyclovir intracoelomic injection 1, 3, and 6 days after the first confirmed KHV mortality. Positive controls (PC) were treated similarly but received sterile saline injections. Negative controls (NC) were exposed to mock shedders. Morbidity and mortality were evaluated daily for 50 days post-challenge. Quantitative PCR was used to determine viral load in the gill biopsies of shedders and cohabitants collected at days 19 (T1), 22 (T2), 25 (T3), 34 (T4), and 50 (T5) post-challenge., Results: Survival curves analyzed by the Gehan-Breslow-Wilcoxon method revealed a delayed onset of mortalities and a significantly lower KHV load at T2 and T3 detected in AT cohabitant fish (P = .042) compared to PC group. However, there were no significant differences in overall mortality or viral loads at T5., Clinical Relevance: The acyclovir protocol used in this study did not control viral infection or mortality at the end of the 50-day trial. Shorter intervals between injections could improve outcomes, but the additional stress inflicted by handling should be considered. Exploring other therapeutic alternatives and doses is warranted.

The use of phytogenic extracts is considered a sustainable strategy for the prevention of fish diseases, including Alliaceae as a potential option due to their variety of bioactive compounds. In this study, we analyzed the antibacterial and antiparasitic potential of propyl-propane-thiosulfinate (PTS) and propyl-propane-thiosulfonate (PTSO) from onions. The in vitro activity against Pseudomonas anguilliseptica , Tenacibaculum maritimum , and Photobacterium damselae of both compounds was tested. In addition, the viability of Sparicotyle chrysophrii larvae was evaluated. Moreover, a diet that consisted of a blend of PTS/PTSO (ALLIUM) was used. A total of 90 gilthead sea bream juveniles were tested against P. damselae subsp. Piscicida after 12 weeks of dietary administration. Furthermore, 150 fish with a rate of 10-15 parasites/fish were fed for 21 days and the number of gill parasites was recorded. All strains were sensitive to both compounds. PTSO showed the highest inhibitory effect against all target strains, while PTS showed higher effectiveness against S. chrysophrii. Fish from ALLIUM group presented the highest probability of survival, increasing up to 91.1%, whereas in the control group, the probability of survival was 66.7%. The number of parasites in the gilthead sea bream decreased in the ALLIUM group over time. These results suggest the inclusion of PTS and PTSO in feed as a natural strategy to prevent antibacterial and antiparasitic fish diseases.

A. hydrophila is an important pathogen that mainly harms aquatic animals and has exhibited resistance to a variety of antibiotics. Here, to seek an effective alternative for antibiotics, the effects of umbelliferone (UM) at sub-MICs on A. hydrophila virulence factors and the quorum-sensing system were studied. Subsequently, RNA sequencing was employed to explore the potential mechanisms for the antivirulence activity of umbelliferone. Meanwhile, the protective effect of umbelliferone on grass carp infected with A. hydrophila was studied in vivo. Our results indicated that umbelliferone could significantly inhibit A. hydrophila virulence such as hemolysis, biofilm formation, swimming and swarming motility, and their quorum-sensing signals AHL and AI-2. Transcriptomic analysis showed that umbelliferone downregulated expression levels of genes related to exotoxin, the secretory system (T2SS and T6SS), iron uptake, etc. Animal studies demonstrated that umbelliferone could significantly improve the survival of grass carps infected with A. hydrophila , reduce the bacterial load in the various tissues, and ameliorate cardiac, splenic, and hepatopancreas injury. Collectively, umbelliferone can reduce the pathogenicity of A. hydrophila and is a potential drug for treating A. hydrophila infection.

Both infectious hematopoietic necrosis virus (IHNV) and infectious pancreatic necrosis virus (IPNV) are the causative agents of acute and highly contagious diseases of juvenile salmonids, resulting in severe economic losses to these cold-water fish globally. There is an urgent need to explore antiviral agents against IHNV and IPNV due to the lack of commercially available vaccines and antiviral drugs. More importantly, the co-infection of IHNV and IPNV is prevalent in nature, which not only aggravates extensive damage to the salmonids but also poses challenges to its prevention and control. The antiviral effects of a crude polysaccharide derived from seaweed (CSP) on IHNV and IPNV were evaluated in this study separately. Furthermore, the underlying antiviral mechanisms of CSP to IHNV and IPNV were analyzed, respectively. The results showed that CSP possessed excellent safety and good ability to inhibit IHNV, IPNV, and their co-infection. CSP preferred to act at the early stage of viral infection. The antiviral mechanism of CSP on IHNV is possibly involved in preventing viral attachment and release, while in IPNV, it is involved in suppressing viral attachment, entry, and release. Taken together, the results of this study shed new light on developing novel agents against viral infection in salmonid fish.

Micropterus salmoides rhabdovirus (MSRV) is a primary viral pathogen in largemouth bass aquaculture, which leads to tremendous economic losses yearly. Currently, there are no approved drugs for the treatment and control of this virus. Our previous studies screened the herb Magnolia officinalis from many traditional Chinese medicines, and we isolated and identified magnolol as its main active compound against multiple rhabdoviruses, including MSRV. On the basis of the structure-activity relationship and pharmacophore model of magnolol, two new magnolol derivatives, namely, hydrogenated magnolol and 2,2'-dimethoxy-magnolol, were designed and synthesized. Their anti-MSRV activities were systematically investigated both in vitro and in vivo. By comparing the half-maximal inhibitory concentration (IC 50 ), it was found that hydrogenated magnolol possessed a higher anti-MSRV activity than magnolol and 2,2'-dimethoxy-magnolol, with an IC 50 of 13.37 μM. Furthermore, hydrogenated magnolol exhibited a protective effect on the grass carp ovary (GCO) cell line by reducing the cytopathic effect induced by MSRV. Further studies revealed that hydrogenated magnolol did not directly impact virions or interfere with MSRV adsorption. It worked within the 6-8 h of the phase of virus replication. In vivo treatment of MSRV infection with magnolol and hydrogenated magnolol showed that they significantly improved the survival rate by 44.6% and 62.7%, respectively, compared to MSRV-infected groups. The viral load measured by the expression of viral glycoprotein in the organs including the liver, spleen, and kidney also significantly decreased when fish were intraperitoneally injected at a dose of 20 mg/kg. Altogether, the structural optimization of magnolol via hydrogenation of the propylene groups increased its anti-MSRV activity both in vitro and in vivo. These results may provide a valuable reference for anti-MSRV drug discovery and development in aquaculture.

Tissue factor pathway inhibitor-2 (TFPI-2) is a Kunitz-type serine protease inhibitor. Previous reports have shown that TFPI-2 plays an important role in innate immunity, and the C-terminal region of TFPI-2 proved to be active against a broad-spectrum of microorganisms. In this study, the TFPI-2 homologue ( SsTFPI-2 ) of black rockfish ( Sebastods schegelii ) was analyzed and characterized, and the biological functions of its C-terminal derived peptide TS40 (FVSRQSCMDVCAKGAKQHTSRGNVRRARRNRKNRITYLQA, corresponding to the amino acid sequence of 187-226) was investigated. The qRT-PCR (quantitative real-time reverse transcription-PCR) analysis showed that the expression of SsTFPI-2 was higher in the spleen and liver. The expression of SsTFPI-2 increased significantly under the stimulation of Listonella anguillarum . TS40 had a strong bactericidal effect on L . anguillarum and Staphylococcus aureus . Further studies found that TS40 can destroy the cell structure and enter the cytoplasm to interact with nucleic acids to exert its antibacterial activity. The in vivo study showed that TS40 treatment could significantly reduce the transmission of L. anguillarum and the viral evasion in fish. Finally, TS40 enhanced the respiratory burst ability, reactive oxygen species production and the expression of immune-related genes in macrophages, as well as promoted the proliferation of peripheral blood leukocytes. These results provide new insights into the role of teleost TFPI-2.

Since the anti-inflammatory effect of hydrogen has been widely known, it was supposed that hydrogen could suppress tissue damage by inhibiting virus-related inflammatory reactions. However, hydrogen is slightly soluble in water, which leads to poor effect of oral hydrogen-rich water therapy. In this study, the nano-bubble hydrogen water (nano-HW) (about 0.7 ​ppm) was prepared and its therapeutic effect against viral infection was investigated by utilizing spring viraemia of carp virus (SVCV)-infected zebrafish as model. Three-month-old zebrafish were divided into nano-HW treatment-treated group and aquaculture water treated group (control group). The results revealed that the cumulative mortality rate of SVCV-infected zebrafish was reduced by 40% after treatment with nano-bubble hydrogen water, and qRT-PCR results showed that SVCV replication was significantly inhibited. Histopathological examination staining showed that SVCV infection caused tissue damage was greatly alleviated after treatment with nano-bubble hydrogen water. Futhermore, SVCV infection caused reactive oxygen species (ROS) accumulation was significantly reduced upon nano-HW treatment. The level of proinflammatory cytokines IL-1β, IL-8, and TNF-α was remarkably reduced in the nano-HW-treated group in vivo and in vitro. Taken together, our data demonstrated for the first time that nano-HW could inhibit the inflammatory response caused by viral infection in zebrafish, which suggests that nano-HW can be applied to antiviral research，and provides a novel therapeutic strategy for virus-caused inflammation related disease., (Copyright © 2022 The Authors. Publishing services by Elsevier B.V. All rights reserved.)

We used 4 years of publicly available data (2016-2019) on chemical usage at salmon sites with information on production, stocking, locations and environmental conditions to explore patterns of anti-sea lice treatments in a Canadian province. Results show that sequential chemical treatments are prevalent, emamectin benzoate (EMB) with azamethiphos being the most used combination with a decrease in ivermectin usage. Relatively high rates of usage of EMB per fish biomass may point to potential lice resistance patterns with information needed on mechanisms and local populations. Lower or no chemical usage at some sites indicate less sea lice infestations likely influenced by localized site conditions (coves), and a lessened need for medication due to the usage of cleaner fish and possibly other non-chemical methods (not documented in this report). The year/climate influenced chemical input only in sites with higher treatment levels likely due to effects on sea lice growth and reproduction. Observed differences between years are warmer surface temperature in the fall, a higher freshwater input in spring, and stronger wind conditions for 2017 and 2018 with more medication usage for these two years. The lack of significant effect of site distances calculated in zones of influence based on 24 h potential connectivity patterns highlight the need to refine the resolution of hydrodynamic processes., (© 2022. The Author(s).)

Bacteriocins are emerging as a viable alternative to antibiotics due to their ability to inhibit growth or kill antibiotic resistant pathogens. Herein, we evaluated the ability of the bacteriocin Garvicin KS (GarKS) produced by Lactococcus garvieae KS1546 isolated from cow milk to inhibit the growth of fish and foodborne bacterial pathogens. We found that GarKS inhibited the growth of five fish L. garvieae strains isolated from infected trout and eels. Among fish pathogens, GarKS inhibited the growth of Streptococcus agalactiae serotypes Ia and Ib, and Aeromonas hydrophila but did not inhibit the growth of Edwardsiella tarda . In addition, it inhibited the growth of A. salmonicida strain 6421 but not A. salmonicida strain 6422 and Yersinia ruckeri . There was no inhibition of three foodborne bacterial species, namely Salmonella enterica , Klebsiella pneumoniae , and Escherichia coli . In vitro cytotoxicity tests using different GarKS concentrations showed that the highest concentration of 33 µg/mL exhibited low cytotoxicity, while concentrations ≤3.3 µg/mL had no cytotoxicity on CHSE-214 and RTG-2 cells. In vivo tests showed that zebrafish larvae treated with 33 µg/mL and 3.3 µg/mL GarKS prior to challenge had 53% and 48% survival, respectively, while concentrations ≤0.33 µg/mL were nonprotective. Altogether, these data show that GarKS has a broad inhibitory spectrum against Gram positive and negative bacteria and that it has potential applications as a therapeutic agent for a wide range of bacterial pathogens. Thus, future studies should include clinical trials to test the efficacy of GarKS against various bacterial pathogens in farmed fish.

Present study investigate the in-vitro antibacterial and antifungal potential of Typha elephantina leaves aqueous extract (T. Eaq), ethanolic extract (T. Eeth) and methanolic extract (T. Emth) at different dosages against selected bacteria and fungi using dis diffusion method and Potato Dextrose Agar method. The study was also proceeded in- vivo against one strain of fungi (Aspergillus niger) using aqueous (T. Eaq) extract only. In-vitro study showed that Citrobacter freundii was highly sensitive while Salmonella typhimurium was the least among all. The antifungal activity was dose dependent and differs according to the fungal strain. Aspergillus niger was highly sensitive in order of aqueous extract (T. Eaq), ethanolic extract (T. Eeth) and methanolic extract (T.Emth), followed by Alterneria solani, Candida albicans and Aspergillus ustus. The in-vivo antifungal study was carried using Cyprinus carpio which were first infected with Aspergillus niger and then treated with (T. Eaq) at different doses. During in-vivo study various hematobiochemicl parameters and bio-accumulative stress of some heavy metals were assessed. Highly significant (P<0.05) remedial effects were observed at day 21st of treatment with extract at 100mg/ kg body weight. Differential accumulation was found i.e in skin the accumulation was highest followed by intestine gills and muscles tissues. Liver showed least accumulation.

Streptococcus zoonotic bacteria cause serious problems in aquaculture with clinical effects on humans. A structure-antibacterial activity relationships analysis of 22 isoflavones isolated from M. tricuspidata (leaves, ripe fruits, and unripe fruits) against S. iniae revealed that prenylation of the isoflavone skeleton was an important key for their antibacterial activities (minimum inhibitory concentrations: 1.95-500 μg/mL). Through principal component analysis, characteristic prenylated isoflavones such as 6,8-diprenlygenistein ( 4 ) were identified as pivotal compounds that largely determine each part's antibacterial activities. M. tiricuspidata ripe fruits (MTF), which showed the highest antibacterial activity among the parts tested, were optimized for high antibacterial activity and low cytotoxicity on fathead minnow cells using Box-Behnken design. Optimized extraction conditions were deduced to be 50%/80 °C/7.5 h for ethanol concentration/extraction temperature/time, and OE-MTF showed contents of 6,8-diprenlygenistein ( 4 ), 2.09% with a MIC of 40 µg/mL. These results suggest that OE-MTF and its active isoflavones have promising potential as eco-friendly antibacterial agents against streptococcosis in aquaculture.

Viral hemorrhagic septicemia virus (VHSV), belonging to the genus Novirhabdovirus, Rhabdoviridae family, is a causative agent of high mortality in fish and has caused significant losses to the aquaculture industry. Currently, no effective vaccines, Food and Drug Administration-approved inhibitors, or other therapeutic intervention options are available against VHSV. α-Lipoic Acid (LA), a potent antioxidant, has been proposed to have antiviral effects against different viruses. In this study, LA (CC 50  = 472.6 μmol/L) was repurposed to exhibit antiviral activity against VHSV. In fathead minnow cells, LA significantly increased the cell viability post-VHSV infection (EC 50  = 42.7 μmol/L), and exerted a dose-dependent inhibitory effect on VHSV induced-plaque, cytopathic effects, and VHSV glycoprotein expression. The time-of-addition assay suggested that the antiviral activity of LA occurred at viral replication stage. Survival assay revealed that LA could significantly upregulated the survival rate of VHSV-infected largemouth bass in both co-injection (38.095% vs. 1.887%, P < 0.01) and post-injection manner (38.813% vs. 8.696%, P < 0.01) compared with the control group. Additional comparative transcriptome and qRT-PCR analysis revealed LA treatment upregulated the expression of several antiviral genes, such as IRF7, Viperin, and ISG15. Moreover, LA treatment reduced VHSV-induced reactive oxygen species production in addition to Nrf2 and SOD1 expression. Taken together, these data demonstrated that LA suppressed VHSV replication by inducing antiviral genes expression and reducing VHSV-induced oxidative stress. These results suggest a new direction in the development of potential antiviral candidate drugs against VHSV infection., (© 2021. Wuhan Institute of Virology, CAS.)

Aeromonas hydrophila ( A. hydrophila ) can cause a number of diseases in both human and animals. A. hydrophila- related infections in aquaculture cause severe economic losses every year throughout the world. The emergence of antibiotic resistance that is due to the abuse of antibiotics has limited the application of antibiotics. Thus, novel approaches are needed to combat with treatment failure of antibiotics caused by resistant bacterial strains. Aerolysin plays a critical role in the pathogenesis of A. hydrophila and has been considered as a novel target for developing drugs based on anti-virulence strategies. Here, we reported that luteolin, a natural product with no anti- A. hydrophila activity, could reduce aerolysin-induced hemolysis by inhibiting aerolysin activity. The binding mode was simulated by molecular docking and dynamics simulation. Then the main binding sites were confirmed by fluorescence quenching assays. We found that luteolin could hindered the formation of functional heptamer of aerolysin according to the results of the oligomerization assay. Moreover, luteolin could protect A549 cells from aerolysin mediated cell death and increase the survival rate of A. hydrophila -infected channel catfish. These findings suggest a novel approach to developing drugs fighting against A. hydrophila , and luteolin can be a promising drug candidate for treatment of A. hydrophila -associated infections.

Flavobacterium psychrophilum, the etiological agent of bacterial coldwater disease (BCWD) and rainbow trout fry syndrome, causes great economic losses in salmonid aquaculture worldwide. Recent molecular studies have uncovered important epidemiological and ecological aspects of this pathogen; however, such data are lacking for F. psychrophilum populations affecting aquaculture in China. Herein, F. psychrophilum phenotype, genotype, and virulence were characterized for isolates recovered from epizootics in multiple salmonid aquaculture facilities across China. Thirty-one F. psychrophilum isolates, originating from four provinces and three host fish species, were predominantly homogeneous biochemically but represented 5 sequence types (STs) according to multilocus sequence typing (MLST) that belonged to clonal complex CC-ST10 or 3 newly recognized singleton STs. PCR-based serotyping classified 19 and 12 F. psychrophilum isolates into molecular serotypes 1 and 0, respectively, showing an obvious relationship with host species. Antimicrobial susceptibility analysis via broth microdilution revealed reduced susceptibility to enrofloxacin, flumequine, and oxolinic acid, moderate susceptibility to gentamicin, erythromycin, and florfenicol, and variable susceptibility to ampicillin and oxytetracycline. In vivo challenge experiments confirmed the ability of two representative Chinese F. psychrophilum isolates to induce typical signs of BCWD and mortality in 1-year-old rainbow trout (Oncorhynchus mykiss). Findings collectively demonstrate (i) that BCWD outbreaks in China studied thus far are caused by F. psychrophilum lineages that are common on other continents (e.g., CC-ST10) and others that have not been reported elsewhere (e.g., ST355, ST356, ST357), (ii) that F. psychrophilum molecular serotypes distinguish isolates from different host fish species, even within STs, and (iii) reduced F. psychrophilum antimicrobial susceptibility against compounds used for BCWD control in China. IMPORTANCE Flavobacterium psychrophilum causes substantial economic losses in salmonid aquaculture worldwide. Although this bacterium is also believed to be a disease source in China, published reports of its presence do not yet exist. Herein, F. psychrophilum was linked to multiple disease outbreaks in several salmonid aquaculture facilities within four Chinese provinces, and polyphasic characterization revealed that most isolates were genetically distinct from strains recovered on other continents. Analyses further revealed the predominating molecular serotypes, antimicrobial susceptibility profiles, and pathogenic potential of two representative recovered isolates. Collectively, the results presented here provide important data on the epidemiology and disease ecology of F. psychrophilum in China and pave the way for targeted prevention and control methods to be pursued in the future.

The current treatments applied in aquaculture to limit disease dissemination are mostly based on the use of antibiotics, either as prophylactic or therapeutic agents, with vaccines being available for a limited number of fish species and pathogens. Antimicrobial peptides are considered as promising novel substances to be used in aquaculture, due to their antimicrobial and immunomodulatory activities. Hepcidin, the major iron metabolism regulator, is found as a single gene in most mammals, but in certain fish species, including the European sea bass ( Dicentrarchus labrax ), two different hepcidin types are found, with specialized roles: the single type 1 hepcidin is involved in iron homeostasis trough the regulation of ferroportin, the only known iron exporter; and the various type 2 hepcidins present antimicrobial activity against a number of different pathogens. In this study, we tested the administration of sea bass derived hepcidins in models of infection and iron overload. Administration with hamp2 substantially reduced fish mortalities and bacterial loads, presenting itself as a viable alternative to the use of antibiotics. On the other hand, hamp1 seems to attenuate the effects of iron overload. Further studies are necessary to test the potential protective effects of hamp2 against other pathogens, as well as to understand how hamp2 stimulate the inflammatory responses, leading to an increased fish survival upon infection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Barroso, Carvalho, Nunes, Gonçalves, Rodrigues and Neves.)

Antimicrobial resistance (AMR) is a growing threat to human and animal health. However, in aquatic animals-the fastest growing food animal sector globally-AMR trends are seldom documented, particularly in Asia, which contributes two-thirds of global food fish production. Here, we present a systematic review and meta-analysis of 749 point prevalence surveys reporting antibiotic-resistant bacteria from aquatic food animals in Asia, extracted from 343 articles published in 2000-2019. We find concerning levels of resistance to medically important antimicrobials in foodborne pathogens. In aquaculture, the percentage of antimicrobial compounds per survey with resistance exceeding 50% (P50) plateaued at 33% [95% confidence interval (CI) 28 to 37%] between 2000 and 2018. In fisheries, P50 decreased from 52% [95% CI 39 to 65%] to 22% [95% CI 14 to 30%]. We map AMR at 10-kilometer resolution, finding resistance hotspots along Asia's major river systems and coastal waters of China and India. Regions benefitting most from future surveillance efforts are eastern China and India. Scaling up surveillance to strengthen epidemiological evidence on AMR and inform aquaculture and fisheries interventions is needed to mitigate the impact of AMR globally., (© 2021. The Author(s).)

Bacterial infection presents severe challenge to tilapia farming, which is largely influenced by water temperature. However, how water temperature determines tilapias' survival to infection is not well understood. Here, we address this issue from the perspective of metabolic state. Tilapias were more susceptible to Aeromonas sobria infection at 33°C than at 18°C, which is associated with differential metabolism of the fish. Compared to the metabolome of tilapia at 18°C, the metabolome at 33°C was characterized with increased an tricarboxylic acid cycle and a reduced level of myo -inositol which represent the most impactful pathway and crucial biomarker, respectively. These alterations were accompanied with the elevated transcriptional level of 10 innate immune genes with infection time, where il-1b , il-6 , il-8 , and il-10 exhibited a higher expression at 33°C than at 18°C and was attenuated by exogenous myo -inositol in both groups. Interestingly, exogenous myo -inositol inactivated the elevated TCA cycle via inhibiting the enzymatic activity of succinate dehydrogenase and malate dehydrogenase. Thus, tilapias showed a higher survival ability at 33°C. Our study reveals a previously unknown relationship among water temperature, metabolic state, and innate immunity and establishes a novel approach to eliminate bacterial pathogens in tilapia at higher water temperature., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Yang, Jiang, Peng and Li.)

In this study, it was determinated the essential oil of cultivated apple mint, Mentha suaveolens Ehrh. composition and in vitro antibacterial activity of against 11 fish pathogen bacteria including Gram-positive (Staphylococcus warneri, Staphylococcus sp., Lactococcus garvieae, Vagococus salmoninarum) and Gram-negative (Aeromonas hydrophila, Aeromonas sobria Aeromonas cavieae, Vibrio anguillarum, Pseudomonas aeroginosa, Yersinia ruckeri, Edwardsiella tarda) by using agar diffusion assay. The main component of M. suaveolence oil was obtained as piperitenone oxide. The essential oil exhibited strong inhibitory activity such as inhibition zone sizes: 30-50mm at 250-1000 µL mL-1 concentrations against V. anguillarum; 16-20mm at 31.25-125 µL mL-1 concentrations against P. aeroginosa; 15-18mm at 500-1000 µL mL-1 concentrations against A. sobria. However, it was found to be moderately effective against E. tarda (8-15 mm), Y. ruckeri (9-12mm), S. warneri (9-10mm), V. salmoninarum (9mm) and Staphylococcus sp. (8-9mm). The essential oil showed weak inhibitory activity against A. cavieae (5-8), A. hydrophila (6-7mm), L. garvieae (5-7mm). Thus, effect of essential oil of M. suaveolens on immune response and disease resistance against Vibrio anguillarum, A. sobria and P. aeroginosa should be investigated in vivo in cultured fish species in future studies.

The myxozoan Tetracapsuloides bryosalmonae is a widely spread endoparasite that causes proliferative kidney disease (PKD) in salmonid fish. We developed an in silico pipeline to separate transcripts of T. bryosalmonae from the kidney tissue of its natural vertebrate host, brown trout (Salmo trutta). After stringent filtering, we constructed a partial transcriptome assembly T. bryosalmonae, comprising 3427 transcripts. Based on homology-restricted searches of the assembled parasite transcriptome and Atlantic salmon (Salmo salar) proteome, we identified four protein targets (Endoglycoceramidase, Legumain-like protease, Carbonic anhydrase 2, Pancreatic lipase-related protein 2) for the development of anti-parasitic drugs against T. bryosalmonae. Earlier work of these proteins on parasitic protists and helminths suggests that the identified anti-parasitic drug targets represent promising chemotherapeutic candidates also against T. bryosalmonae, and strengthen the view that the known inhibitors can be effective in evolutionarily distant organisms. In addition, we identified differentially expressed T. bryosalmonae genes between moderately and severely infected fish, indicating an increased abundance of T. bryosalmonae sporogonic stages in fish with low parasite load. In conclusion, this study paves the way for future genomic research in T. bryosalmonae and represents an important step towards the development of effective drugs against PKD.

Marine sponges are sessile invertebrates that can be found in temperate, polar and tropical regions. They are known to be major contributors of bioactive compounds, which are discovered in and extracted from the marine environment. The compounds extracted from these sponges are known to exhibit various bioactivities, such as antimicrobial, antitumor and general cytotoxicity. For example, various compounds isolated from Theonella swinhoei have showcased various bioactivities, such as those that are antibacterial, antiviral and antifungal. In this review, we discuss bioactive compounds that have been identified from marine sponges that showcase the ability to act as antibacterial, antiviral, anti-malarial and antifungal agents against human pathogens and fish pathogens in the aquaculture industry. Moreover, the application of such compounds as antimicrobial agents in other veterinary commodities, such as poultry, cattle farming and domesticated cats, is discussed, along with a brief discussion regarding the mode of action of these compounds on the targeted sites in various pathogens. The bioactivity of the compounds discussed in this review is focused mainly on compounds that have been identified between 2000 and 2020 and includes the novel compounds discovered from 2018 to 2021.

Background : Turmeric is known as a natural remedy to improve the immunity of organisms. This study aims to understand the effectiveness of turmeric-enriched pellets to improve the immunity of Clarias batrachus to Aeromonas hydrophila . Methods: The study was conducted from May to August 2020. C. batrachus fingerlings, 7-8 cm total length (TL) and 4-5 g (BW) at baseline, were kept in 30 L aquaria (10 fishes/aquarium; three replicated/treatment). Commercial pellets were mixed with turmeric powder. There were five treatment groups: P0 (control, no turmeric); P1 (0.5 g turmeric per Kg of pellets); P2 (0.7 g/Kg); P3 (0.9 g/Kg); Pp (positive control). Thirty days after being feed with turmeric-enriched pellets, all groups of fish were infected with 0.1 ml (10 8 ) of A. hydrophila suspension, intramuscularly. The P0 group did not receive injection, while Pp group were not fed with turmeric-enriched pellets but were infected with the bacteria. Fourteen days after infection, clinical signs and hematology of the fish were studied. Results: Pp fish showed heavy clinical signs of A. hydrophila , such as loss of balance, pigmentation, hemorrhages and ulcers. P0 fish did not show any symptoms, while the treated fish reveled some clinical signs of A. hydrophila to a lesser extent than Pp, indicating that the fish is able to face the A. hydrophila attack. Hematology for Pp fish revealed high white blood cells, indicating that the fish were infected. The blood condition of the P0 fish, as well as those of the turmeric-treated fish were normal. In general, the P3 fish showed the least clinical signs of A. hydrophila and normal blood condition, indicating that P3 treatment is best. Conclusion: The best turmeric dosage to improve the immunity of C. batrachus toward A . hydrophila infection is 0.9 g/Kg pellets., Competing Interests: No competing interests were disclosed., (Copyright: © 2021 Riauwaty M et al.)

Background : Turmeric is known as a natural remedy to improve the immunity of organisms. This study aims to understand the effectiveness of turmeric-enriched pellets to improve the immunity of Clarias batrachus to Aeromonas hydrophila . Methods: The study was conducted from May to August 2020. C. batrachus fingerlings, 7-8 cm total length (TL) and 4-5 g (BW) at baseline, were kept in 30 L aquaria (10 fishes/aquarium; three replicated/treatment). Commercial pellets were mixed with turmeric powder. There were five treatment groups: P0 (control, no turmeric); P1 (0.5 g turmeric per Kg of pellets); P2 (0.7 g/Kg); P3 (0.9 g/Kg); Pp (positive control). Thirty days after being feed with turmeric-enriched pellets, all groups of fish were infected with 0.1 ml (10 8 ) of A. hydrophila suspension, intramuscularly. The P0 group did not receive injection, while Pp group were not fed with turmeric-enriched pellets but were infected with the bacteria. Fourteen days after infection, clinical signs and hematology of the fish were studied. Results: Pp fish showed heavy clinical signs of A. hydrophila , such as loss of balance, pigmentation, hemorrhages and ulcers. P0 fish did not show any symptoms, while the treated fish reveled some clinical signs of A. hydrophila to a lesser extent than Pp, indicating that the fish is able to face the A. hydrophila attack. Hematology for Pp fish revealed high white blood cells, indicating that the fish were infected. The blood condition of the P0 fish, as well as those of the turmeric-treated fish were normal. In general, the P3 fish showed the least clinical signs of A. hydrophila and normal blood condition, indicating that P3 treatment is best. Conclusion: The best turmeric dosage to improve the immunity of C. batrachus toward A . hydrophila infection is 0.9 g/Kg pellets., Competing Interests: No competing interests were disclosed., (Copyright: © 2021 Riauwaty M et al.)