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Scuticociliatosis, caused by Miamiensis avidus, is a severe parasitic disease affecting marine organisms, particularly Paralichthys olivaceus. The aim of this study was to assess the antiparasitic potential of ethanolic extracts of Carpesii Fructus (EECF), the dried fruit of Carpesium abrotanoides L., which is used in traditional Chinese medicine, in vitro. We found that 50%, 70%, and 100% EECF induced morphological changes in M. avidus, including reduced motility, cell shrinkage, and lysis. Nearly 100% cell lysis was observed in M. avidus after 2 h of treating with 100% EECF. After 24 h, the survival rates of M. avidus treated with 100%, 70%, and 50% EECF were 10%, 20%, and 30%, respectively. Additionally, the mRNA levels of immune response-related (IL-1β, IL-8, TNF-α, and CD8-α) and biotransformation-related (CYP1A, CYP1B, CYP3A4, and UGT2B19) genes increased with 70% and 100% EECF treatment and decreased with 50% EECF treatment following pretreatment with concanavalin A. The viability of hirame natural embryo (HINAE) cells was reduced by 50%, 70%, and 100% EECF (100 mg/L) and was between 67 and 80%. The IC 50 values of 50%, 70%, 90%, and 100% EECF in HINAE cells were 102.3, 42.93, 39.15, and 38.39 mg/L, respectively. These results indicated that 50% EECF was less toxic to HINAE cells than 70% or 100% EECF, while still exhibiting antiparasitic activity against M. avidus. Therefore, we demonstrated the role of EECF as a natural antiparasitic agent against M. avidus. Our findings suggest that Carpesii Fructus has potential use as an antiparasitic agent in the aquaculture industry., (Copyright © 2021 Elsevier Inc. All rights reserved.)

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.)

The nervous necrosis virus (NNV) infection is generally observed in aquafarms when the seawater temperature is higher than 24 °C and the fishes seem to be refractory to disease at suboptimal temperatures below 20 °C suggesting a role of thermoregulation in NNV pathogenesis. The present study profiled the temperature-dependent regulation of cytokines (TNF-α, IL-1β and IFN-γ), innate antiviral factors (IFN-1, Mx, ISG-15), adaptive immune factors (CD-4, CD-8, IgM), signaling regulators (SOCS-1, SOCS-3), transcription factors (STAT-1, STAT-3) and microglial and NCC/NK specific cell markers (TMEM-119 and NCCRP-1) during NNV challenge in seven-band grouper, Hyporthodus septemfasciatus. The co-habitation challenge at 17 °C with showed a sustained expression of proinflammatory cytokines and following rechallenge with a dose of 10 4 TCID 50 /100μL/fish at optimal temperature, the survivors also exhibited a stable expression of immune factors. The 100% survival following the challenge at sub-optimal (17 °C) and rechallenge at optimal (25 °C) was due to the stable and sustained activation of the immune response. However, at 25 °C, the rechallenge displayed a priming effect with hyperactivation of the immune system evident from the immune gene expression profile. The mortality pattern observed is co-related with the cytokine storm as is evident from the gene expression profile. Whereas, neither of the adaptive immune markers was suggestive of humoral immune response in the 17 °C groups. Also, the data suggest a possible role of NK cell and microglia in mediating antiviral immune response following infection in the brain at different temperatures, where, former is beneficial in restricting viral infection with higher host tolerance., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

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.

Ichthyophthiriasis, commonly known as white spot disease, occurs in both wild and cultured fish and is responsible for heavy economic losses to the aquaculture industry. In past decade, several chemical therapeutants were used to treat ichthyophthiriasis, but the effective drugs, such as malachite green, have been banned for use in food fish due to its genotoxic and carcinogenic properties. To find efficacious drugs to control Ichthyophthirius multifiliis (Ich), whole Eclipta alba plants and dried root of Arctium lappa were evaluated for their antiprotozoal activity. E. alba and A. lappa extracts significantly reduced the survival of Ich trophonts and theronts. In vitro, the E. alba and A. lappa methanol extracts killed all trophonts at 3200 mg l-1. All trophonts were killed after exposure to E. alba aqueous extract at 3200 mg l-1. The methanol extracts of E. alba and A. lappa killed 100% of I. multifiliis theronts at 400 mg l-1 and 800 mg l-1, respectively. The aqueous extract of E. alba and A. lappa killed 100% of I. mulitifiliis theronts at 1600 mg l-1 and 3200 mg l-1, respectively. E. alba and A. lappa extracts may be new and efficacious drugs for the control of ichthyophthiriasis., (Copyright© by the Polish Academy of Sciences.)

Tapeworms (Cestoda) represents a species rich (about 5000 species) group of flatworms (Neodermata) parasitizing all groups of vertebrates including humans, with about 1000 species parasitizing elasmobranchs and almost 500 occurring in teleosts as adults. They are common parasites of cultured fish, both as adults and larvae (metacestodes), but only few adult tapeworms are actually pathogenic for their fish hosts. In contrast, cestode larvae can be harmful for fish, especially plerocercoids migrating throughout their tissue and internal organs. Current knowledge of host-parasite relationships, including immune response of fish infected with tapeworms, is still insufficient to enable adequate control of cestodoses, and most data available were obtained several decades ago. Treatment of fish infected with adult tapeworms is effective, especially with praziquantel, whereas the treatment of metacestodes is problematic. Control measures include interruption of the complex life cycle and prevention of transport of uninspected fish to new region., (© 2021 John Wiley & Sons Ltd.)

Diseases caused by rhabdoviruses have had a huge impact on the productive lives of the entire human population. The main problem is the lack of drugs for the treatment of this family of viruses. Infectious hematopoietic necrosis virus (IHNV), the causative agent of IHN, is a typical rhabdovirus which has caused huge losses to the salmonid industry. Therefore, in this study, IHNV was studied as a model to evaluate the antiviral activity of 35 novel coumarin derivatives. Coumarin A9 was specifically selected for further validation studies upon comparing the half maximum inhibitory concentration (IC 50 ) of four screened candidate derivatives in epithelioma papulosum cyprinid (EPC) cells, as it exhibited an IC 50 value of 2.96 μM against IHNV. The data revealed that A9 treatment significantly suppressed the virus-induced cytopathic effect (CPE) in EPC cells. In addition, A9 showed IC 50 values of 1.68 and 2.12 μM for two other rhabdoviruses, spring viremia of carp virus and micropterus salmoides rhabdovirus, respectively. Furthermore, our results suggest that A9 exerts antiviral activity, but not by destroying the virus particles and interfering with the adsorption of IHNV. Moreover, we found that A9 had an inhibitory effect on IHNV-induced apoptosis in EPC cells, as reflected by the protection against cell swelling, formation of apoptotic bodies, and loss of cell morphology and nuclear division. There was a 19.05 % reduction in the number of apoptotic cells in the A9 treatment group compared with that in the IHNV group. In addition, enzyme activity assays proved that A9 suppressed the expression of caspase 3, 8 and 9. These results suggested that A9 inhibit viral replication, to some extent, by blocking IHNV-induced apoptosis. In an in vivo study, A9 exhibited an anti-rhabdovirus effect in virus-infected fish by substantially enhancing the survival rate. Consistent with the above results, A9 repressed IHNV gene expression in virus-sensitive tissues (brain, kidney and spleen) in the early stages of virus infection. Importantly, the data showed that horizontal transmission of IHNV was reduced by A9 in a static cohabitation challenge model, especially in fish that underwent bath treatment, suggesting that A9 might be a suitable therapeutic agent for IHNV in aquaculture. Therefore, coumarin derivatives can be developed as antiviral agents against rhabdoviruses., 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 © 2021 Elsevier Masson SAS. All rights reserved.)

This study provides the first data related to albendazole (ABZ) and its main metabolites [albendazole sulphoxide (ABZSO), albendazole sulphone (ABZSO 2 ), and albendazole-2-amino sulphone (ABZ-2-NH 2 -SO 2 )] residue depletion in tambaqui ( Colossoma macropomum ) parasitised by acanthocephalan ( Neoechinorhynchus buttnerae ). The ABZ withdrawal period was also calculated. The fish received a daily dose of 10 mg ABZ kg -1 body weight (b.w.) via medicated feed for 34 days. Samples of target tissue (muscle plus skin in natural proportions) were collected 24, 48, 72, 120, 168, 240, and 336 h after the end of ABZ administration. The quantitation of ABZ residues and its metabolites in the target tissue was performed using a validated ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) analytical method. After treatment, ABZ in the target tissue was rapidly metabolised over time, and ABZSO was the most persistent metabolite and was shown to be at the highest levels in the target tissue. Considering the maximum residue limit (MRL) established by Codex Alimentarius in the muscle (100 μg kg -1 , species not specified), a withdrawal period of 4 days (112 °C-day) was estimated for the total residue (sum of ABZ and its metabolite residues). Considering data reported in the literature and data obtained in this study, it is suggested that the total residue be considered as marker residue to be adopted for fish in the legislative framework.

The majority of public aquaria have at least one very large tank (>370,000 liters, 100,000 gallons) as part of their collection. The large water volumes can make treatment difficult when pathogens escape quarantine or cross contamination occurs. The cost and methods of dispersion of chemotherapeutics to control infectious agents (bacterial, protozoal, or metazoan) can be prohibitive in large volumes. Currently there are few treatments for stenohaline monogeneans in large aquaria that are efficacious, safe, and affordable. This case demonstrates the safety and efficacy of a decade long use of prolonged hyposalinity that replaced the repeated use of copper and organophosphate treatments to control a Neobenedenia spp. monogenean infestation. The use of prolonged 20 ppt saline water is safe for a list of species traditionally listed as stenohaline which makes altering salinity a useful tool in treating parasites with restricted saline tolerances., (© 2021 Wiley Periodicals LLC.)

Due to the lack of relevant therapies for infectious haematopoietic necrosis virus (IHNV) infection, the viral outbreak invariably causes serious economic losses in salmonid species. In this study, we evaluated the anti-IHNV effects of 7-(6-benzimidazole) coumarin (C10) and 4-phenyl-2-thioxo-1,2,3,4-tetrahydro-5H-chromeno[4,3-d]pyrimidin-5-one (S5) in vitro and in vivo. The results revealed that C10 at 12.5 mg/L and S5 at 25 mg/L significantly inhibited IHNV replication in epithelioma papulosum cyprini (EPC) cells with a maximum inhibitory rate >90%, showing that IHNV-induced cytopathic effect (CPE) was alleviated by C10 and S5. There are two complementary effects on antiviral mechanism: 1. C10 completely inhibited IHNV infectivity when the virus was preincubated with C10 at 12.5 mg/L, determining that C10 may have a negative impact on IHNV binding to the cell; 2. C10 also up-regulated the gene expression of extracellular proto type galectin-1 (Gal1-L2) and a chimera galectin-3 (Gal3-L1) of EPC cells to inhibit IHNV adhesion. For the in vivo study, injection and immersion of the coumarins enhanced the survival rate of rainbow trout (Oncorhynchus mykiss) juveniles by 25% (at least) at 12 dpi. IHNV loads in the kidney and spleen were also obviously decreased at 96 h, and thus we considered that they had a delaying effect on IHNV replication in vivo. Meanwhile, C10 with a high stability in aquacultural water in immersion suppressed IHNV horizontal transmission by decreasing the viral loads in recipient fish. Overall, our data suggest that there is a positive effect of C10 and S5 against IHNV infection in aquaculture, and C10 had the potential to be a broad-spectrum antiviral against fish rhabdoviruses., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Parasite infections in fish require constant surveillance and strategies for efficient treatments which guarantee the fish health, their sale value and the non-propagation of pathogens in new environments. Fish treatments based on nanotechnology become of increasing interest since nanoparticles have been shown as efficient materials for optimizing administration of bioactives. In this study a chitosan derivative, alginate and praziquantel conjugated nanobioparticle of effective action for oral treatment of digenetic trematodes in highly infected Corydoras schwartzi was evaluated in terms of histological and hematological safety. The inherent absence of alterations in intestinal tissue and the reversible blood cells counting during a period up to 35 days showed the safety of the drug delivery nanobioparticles, which thus represent a promising strategy for effective applications in pathogens treatments by oral administration., (© 2021 John Wiley & Sons Ltd.)

Accumulating evidence supports that vitamin D 3 (VD 3 ) possesses immunomodulatory properties besides its classical actions in calcium and bone homeostasis. In this study, juvenile turbots were fed with the diets containing 0 IU/kg VD 3 or the optimum dose of 400 IU/kg VD 3 for 8 weeks. To investigate the effects of VD 3 on anti-infectious immunity in fish, 10 7  CFU Edwardsiella tarda was injected intraperitoneally to each juvenile turbot after the feeding trial. Our results showed that the mortality of infected turbots with dietary VD 3 was much lower than that in VD 3 deficient group, and the supplementation of dietary VD 3 significantly reduced the bacterial load in the spleen of infected turbots. Further analysis demonstrated that the production of reactive oxygen species (ROS) in haemocytes and lysozyme activity in serum was elevated, and the responses of T cells and B cells were modulated in VD 3 -supplemented turbots. Moreover, the inflammation was significantly exacerbated in the infected turbots fed with 0 IU/kg VD 3 compared to the fish fed with 400 IU/kg VD 3 . In addition, the head kidney macrophages (HKMs) in turbots were isolated and incubated with VD 3 in vitro, the results showed that VD 3 significantly promoted the bactericidal activity in HKMs. In conclusion, our study has shown clear evidence that VD 3 positively regulates the innate and adaptive immunity in fish, which is beneficial to the defense in fish against pathogen infection., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Aims: The streptococcal disease has been associated with serious mortality and significant global economic loss in the tilapia farming industry. The overall goal of this work was to test herbal hydrogels based on encapsulated Enterococcus faecium ABRIINW.N7 for potential probiotic anti-microbial activity against Streptococcus iniae in red hybrid tilapia., Methods and Results: Abnormal behaviour, clinical signs, postinjection survival and histopathology (kidney, liver, eye and brain) were measured. Cumulative mortality of CON + , free cells, ALG and treatments (F1-F7) was 30, 24, 22, 19, 17, 17, 16, 14, 14 and 12 out of 30 fish and the survival rates for E. faecium ABRIINW.N7 microencapsulated in an alginate-BS blend with 0·5, 1, 1·5, 2, 2·5 and 3% fenugreek were 43, 43, 47, 53, 53 and 60%, respectively. After the incorporation of fenugreek with the alginate-BS blend, there was an 8-21% increase in probiotic cell viability. Furthermore, the survival rate for the alginate-BS blend with 2·5 and 3% fenugreek (F6 and F7) was significantly (P ≤ 0·05) higher than other blends. The highest encapsulation efficiency, viability in gastrointestinal conditions and during storage time and excellent antipathogenicity against S. iniae were observed in alginate-BS +3% fenugreek formulation (F7)., Conclusions: It is recommended that probiotic strains like E. faecium ABRIINW.N7 in combination with local herbal gums, such as BS and fenugreek plus alginate, can be used as a suitable scaffold and an ideal matrix for the encapsulation of probiotics., Significance and Impact of the Study: This study proposes models connecting process parameters, matrix structure and functionality., (© 2021 The Society for Applied Microbiology.)

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 emergence of carbapenem-resistant Acinetobacter baumannii has increased the risk of nosocomial infections, which pose a huge health threat. There is an urgent need to develop alternative therapies, including broad-spectrum antimicrobial peptides. In this study, we designed, characterized, and studied the antibacterial, antibiofilm effects and possible mode of actions of a novel synthetic peptide Octopromycin, derived from the proline-rich protein 5 of Octopus minor. Octopromycin consists of 38 amino acids, (+5) net positive charge, high hydrophobic residue ratio (36%), and two α-helix secondary structures. The minimum inhibitory concentration and minimum bactericidal concentration against A. baumannii were 50 and 200 μg/mL, respectively. Time-kill kinetics and bacterial viability assays confirmed the concentration-dependent antibacterial activity of Octopromycin. Field emission scanning electron microscopy images clearly showed ultrastructural alterations in Octopromycin-treated A. baumannii cells. Propidium iodide penetrated into Octopromycin-treated A. baumannii cells, demonstrating the loss of cell membrane integrity. Octopromycin treatment increased the production of reactive oxygen species in a concentration-dependent manner, and it inhibited the biofilm formation and showed biofilm eradication activity against A. baumannii. In vitro and in vivo safety evaluation revealed that Octopromycin was nontoxic to HEK293T and Raw 264.7 cells (<400 μg/mL), as well as mice red blood cells (<300 μg/mL), and zebrafish embryos (<4 μg/mL). An in vivo study results revealed that the A. baumannii-infected fish treated with Octopromycin exhibited a significantly higher relative percent survival (37.5%) than the infected mock-treated fish with PBS (16.6%). Furthermore, a decreased bacterial load and fewer alterations in histological analysis confirmed the successful control of A. baumannii by Octopromycin in vivo. Collectively, the results indicate that the antibacterial peptide Octopromycin may achieve rapid control of A. baumannii through multi-target interactions; it presents a desirable therapeutic option for the prevention and control of the infections., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Aims: This study aimed to evaluate the in vitro cytotoxicity and efficacy of synthetic host defence peptides (HDPs), alone or in combination with florfenicol (FFC), oxytetracycline (OTC) or thiamphenicol (TAP), against different pathogenic bacteria isolated from diseased fish., Methods and Results: Solid-phase synthesis, purification and characterization of several HDPs were performed manually, using the fluorenylmethyloxycarbonyl protecting group in different resins and via high-performance liquid chromatography-mass spectrometry, respectively. The in vitro cytotoxicity and antimicrobial activity of HDPs, FFC, OTC and TAP against Nile tilapia red blood cells (RBCs) and relevant fish pathogenic bacteria (Aeromonas, Citrobacter, Edwardsiella, Streptococcus, Lactococcus and Vibrio) was determined using the haemolysis assay and broth microdilution method, respectively. The checkerboard assay was used to evaluate the synergy between the most active HDPs and other antimicrobials against the tested strains. MUC 7 12-mer, FFC, OTC and TAP were not cytotoxic to Nile tilapia RBCs, in all tested concentrations. LL-37, (p-BthTX-I) 2 and Hylin-a1 were not cytotoxic at concentrations up to 78·13, 19·53 and 9·77 μg ml -1 , respectively. HDPs demonstrated potent antimicrobial activity (minimum inhibitory concentration ≤31·25 µg ml -1 ) against Aeromonas jandaei (KR-12-a5), Citrobacter freundii (Kr-12-a5; (p-BthTX-I) 2 ; LL-37; and Hylin a1), Streptococcus agalactiae (Hylin a1; (p-BthTX-I) 2 and LL-37), Lactococcus garviae (Hylin a1), and Vibrio fluvialis (KR-12-a5). The combinations of (p-BthTX-I) 2 with TAP and LL-37 with FFC showed synergistic activity against C. freundii (fractional inhibitory concentration index of 0·25 and 0·50, respectively)., Conclusions: Synthetic HDPs have the potential as a good treatment option for bacterial diseases in aquaculture., Significance and Impact of the Study: The in vivo effectiveness of synthetic HDPs such as KR-12-a5; LL-37; (p-BthTX-I) 2 and Hylin a1 can be tested alone or in combination with conventional antimicrobials as a treatment option to reduce the use of antimicrobials in aquaculture., (© 2021 The Society for Applied Microbiology.)

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.)

Neobenedenia girellae is a pathogenic ectoparasite of many marine fishes, and it causes major epidemics in marine aquaculture. In this study, the efficacy of ethanol extracts of huangqi Astragalus membranaceus (known as milkvetch in North America), guanzhong Dryopteris setosa (known as beaded wood fern in North America), gancao Glycyrrhiza uralensis (known as Chinese licorice in North America), danshen Salvia miltiorrhiza (known as red sage in North America), and pomegranate Punica granatum, as well as seven phytochemicals (10-gingerol, curcumin, cynatratoside-C, emodin, kuwanon-G, kuwanon-O, and sophoraflavanone-G), against adult N. girellae was investigated. In vitro results indicated that pomegranate extract killed all adult N. girellae at a 62.5-mg/L concentration with an 8-h exposure, but gancao extract did not cause 100% mortality until a 1,000-mg/L concentration was used. Additionally, all adult N. girellae died after an 8-h exposure to cynatratoside-C, kuwanon-G, kuwanon-O, or sophoraflavanone-G at a concentration of 125 mg/L. Curcumin, emodin, and 10-gingerol at a concentration of 1,000 mg/L did not kill all parasites after an 8-h exposure. These findings demonstrate that plant extracts and active phytochemicals are potential sources of botanical drugs for controlling N. girellae infection in aquaculture., (© 2021 American Fisheries Society.)

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.

Research on immunotherapeutic agents has become a focus for the treatment of fish diseases. The ability of algae to produce secondary metabolites of potential interest as immunotherapeutics has been documented. The present research intended to assess antiviral and antibacterial activities of macro- and microalgae extracts against viral and bacterial pathogens and explore their immunomodulatory potential using zebrafish (Danio rerio) larvae as a model organism. The cytotoxicity and antiviral activity of eight methanolic and ethanolic extracts from two macroalgae (Fucus vesiculosus, Ulva rigida) and two microalgae (Nannochloropsis gaditana, Chlorella sp.) were analyzed in established fish cell lines. Six extracts were selected to evaluate antibacterial activity by disk diffusion and growth inhibition assays. The three most promising extracts were characterized in terms of fatty acid composition, incorporated at 1% into a plant-based diet, and evaluated their effect on zebrafish immune response and intestinal morphology in a short-term feeding trial. All extracts exhibited in vitro antiviral activity against viral hemorrhagic septicemia and/or infectious pancreatic necrosis viruses. Methanolic extracts from F. vesiculosus and U. rigida were richer in saturated fatty acids and exhibited in vitro antibacterial action against several bacteria. Most promising results were obtained in vivo with F. vesiculosus methanol extract, which exerted an anti-inflammatory action when incorporated alone into diets and induced pro-inflammatory cytokine expression, when combined with the other extracts. Moreover, dietary inclusion of the extracts improved intestinal morphology. In summary, the results obtained in this study support the potential of algae as natural sources of bioactive compounds for the aquaculture industry., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Cyprinid herpesvirus 3 (CyHV-3) causes high mortality in carp. Emodin has been shown of the effects of antioxidant, anti-inflammatory and antiviral. In present study, we investigated the preventive effects and mechanism of emodin on CyHV-3 infection. The ornamental koi carp (Cyprinus carpio haematopterus) were intraperitoneally injected with emodin (10 mg/kg, 20 mg/kg, or 40 mg/kg). 72 h later, an intraperitoneal injection of CyHV-3 was administered, and collected the samples one week later to detect the antioxidant parameters, antioxidant genes, inflammatory genes and to perform histopathology assays. The results showed that emodin significantly suppressed CyHV-3 replication (P < 0.05), improved the koi survival rate and slowed the damage caused by CyHV-3. Emodin treatment increased the antioxidant activity and decreased the lipid peroxidation level of the koi. Compared to the CyHV-3 group, emodin treatment resulted in the same antioxidant parameters after CyHV-3 infection. Emodin treatment activated the Nuclear factorery throid 2-related factor 2/Kelch-like ECH-associated protein 1-antioxidatant response element (Nrf2/Keap1-ARE) pathway and upregulated the expression of heme oxygenase 1 (HO-1), superoxide dismutase (SOD), and catalase (CAT) in the hepatopancreas after CyHV-3 infection. Emodin activated the nuclear factor kappa-B (NF-κB) pathway and decreased the expression of interleukin-6 (IL-6), interleukin-8 (IL-8), and tumour necrosis factor-α (TNF-α) in the koi induced by CyHV-3. In conclusion, emodin treatment can suppress CyHV-3 replication and reduce the mortality of koi caused by CyHV-3. Emodin improves antioxidant function, relieves oxidative stress and inflammation cytokines via Nrf2/Keap1-ARE and NF-κB pathways, and protects against the adverse effects induced by CyHV-3., (Copyright © 2021. Published by Elsevier Inc.)

Diet-induced fatty liver is a considerable threaten to fish aquaculture due to the popularity of the high-fat diet (HFD) feeding. Our study aims to investigate the effects of flavanones from Sedum sarmentosum Bunge (FSSB) on the liver function to identify a potential treatment for HFD-induced fatty liver disease. Physiological and pathological indicators were tested in the liver of Nile tilapia (Oreochromis niloticus) and results showed parameters including lipid metabolites, redox parameters, and inflammatory factors could be adequately restored to normal level by addition of 150 mg/kg FSSB to HFD. Proteomics analysis was performed in liver tissues from tilapia with normal diet (ND), HFD, and HFD+FSSB. Totally, 51 upregulated proteins and 77 downregulated proteins were identified in HFD groups and 67 proteins of them were restored after treated with FSSB. Bioinformatics analysis showed that differentially expressed proteins (DEPs) in HFD+FSSB150 group compared with HFD group are mainly enriched in acety-CoA metabolic process, adenosine-triphosphate (ATP) biosynthetic process, lipid metabolic process, and phospholipid metabolic process. The dysregulated proteins were involved in peroxidosome proliferators-activated receptor (PPAR) signaling pathway, fat digestion and absorption, and immune system. The quantitative real-time PCR (qRT-PCR) assay further revealed that the expression of GST, PPARα, PPARγ, and multiple-inflammatory cytokines could be also reversed in HFD group under the treatment of 150 mg/kg FSSB. Our findings demonstrated FSSB is efficient for the treatment of fatty liver disease through regulation of lipid metabolism and antioxidation in Nile tilapia, providing a new treatment of non-alcoholic fatty liver disease (NAFLD) in fish aquaculture., (© 2021. The Author(s), under exclusive licence to Springer Nature B.V.)

Target identification is important for drug discovery. Unfortunately, no drug targets have been found in Ichthyophthirius multifiliis until now and further limited development of the novel drug for Ichthyophthiriasis. In this study, an iTRAQ-based quantitative proteomic analysis was used to find the target of malachite green (MG), exhibiting greater efficacy than the existing drugs, against I. multifiliis trophonts in situ. We also verified the proteomic results by RT-qPCR, TEM and cell apoptosis assay. Our results showed that major variations in protein abundance were found among many of the ribosome proteins, indicating ribosome might be a candidate target. Furthermore, GO and KEGG pathway analyses of differentially expressed proteins (DEPs) revealed that ribosome and PI3K-Akt signalling pathway were remarkably enriched. Taken together, the above DEPs were also verified by RT-qPCR and morphological observations. This study provides insights into the key proteins enriched in PI3K-Akt signal pathway and ribosome pathway as potential targets of MG killing I. multifiliis, which could be served as targets for other less toxic drugs and be tested as potential treatments for I. multifiliis., (© 2021 John Wiley & Sons Ltd.)

We previously identified a novel acidic polysaccharide, silkrose-AY, from the Japanese oak silkmoth (Antheraea yamamai), which can activate an innate immune response in mouse macrophage cells. However, innate immune responses stimulated by silkrose-AY in teleosts remain unclear. Here, we show the influence of dietary silkrose-AY in medaka (Oryzias latipes), a teleost model, in response to Edwardsiella tarda infection. Dietary silkrose-AY significantly improved the survival of fish and decreased the number of bacteria in their kidneys after the fish were artificially infected with E. tarda by immersion. We also performed a microarray analysis of the intestine, which serves as a primary barrier against microbial infection, to understand the profiles of differentially expressed genes (DEGs) evoked by silkrose-AY. The dietary silkrose-AY group showed differential expression of 2930 genes when compared with the control group prior to E. tarda infection. Gene ontology and pathway analysis of the DEGs highlighted several putative genes involved in pathogen attachment/recognition, the complement and coagulation cascade, antimicrobial peptides/enzymes, opsonization/phagocytosis, and epithelial junctional modification. Our findings thus provide fundamental information to help understand the molecular mechanism of bacterial protection offered by insect-derived immunostimulatory polysaccharides in teleosts., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

This retrospective descriptive study estimates cage-level mortality distributions after six immediate delousing methods: thermal, mechanical, hydrogen peroxide, medicinal, freshwater and combination of medicinal treatments. We investigated mortality patterns associated with 4 644 delousing treatment of 1 837 cohorts of farmed Atlantic salmon (Salmo salar) stocked in sea along the Norwegian coast between 2014 and 2017. The mortality is expressed as mortality rates. We found distributions of delta mortality rate within 1, 7 and 14 days after all six delousing treatments, using mortality rate within 7 days before treatments as baseline. The results show that we can expect increased mortality rates after all six delousing methods. The median delta mortality rates after thermal and mechanical delousing are 5.4 and 6.3 times higher than medicinal treatment, respectively, for the 2017 year-class. There is a reduction in the delta median mortality for thermal and freshwater delousing from 2015 to 2019. There is a wide variability in the mortality rates, in particular for thermal delousing. Our results suggest that the variability in delta mortality for thermal delousing has been reduced from the 2014 to 2017 year-class, indicating an improvement of the technique. However, a significant increase in the number of thermal treatments from 14 in 2015 to 738 in 2018 probably contributes to the overall increased mortality in Norwegian salmon farming., (© 2021 The Authors. Journal of Fish Diseases published by John Wiley & Sons Ltd.)

Background: Use of enrofloxacin in trout farms is reported, especially for the treatment of yersiniosis, albeit various dosing regimens have been used. Therefore, optimal doses should be investigated., Methods: Five groups of 15 fish were challenged with Y. ruckeri. Two days later, three groups received feed containing enrofloxacin (ENR) at 1, 2.5 and 5 mg/kg fish respectively, during 7 days; one group received a single intraperitoneal injection of ENR at 10 mg/kg; and one group was left untreated. On day 15, surviving fish were euthanized., Results: All fish survived in the group treated by injection, compared to 53%, 60% and 40% of the fish treated with 1, 2.5 and 5 mg/kg oral ENR, respectively, and 53% in the infected untreated group., Conclusion: A single intraperitoneal injection of ENR at 10 mg/kg seems more relevant than repeated oral administrations. The ENR oral doses used in trout farms should be revised., (© 2021 British Veterinary Association.)

This study investigated the effects of Greek juniper extract on immune responses of rainbow trout. In this experiment, 4 doses [0 (Control), 1 (J1), 4 (J4) and 8 (J8) mg/kg] of the extract were administered orally using an oral gavage twice a day for 14 days. Immune responses were measured on 7th and 14th days. On 14th day, Yersinia ruckeri was injected intraperitoneally to all fish of all groups. On 14th day, ORP in fish of J1 group increased significantly. Lysozyme activity (LA) was increased in J8 group on 7th day (p < .05). On 14th day, a significant decrease was determined in J1 and J4 treatments in LA. Myeloperoxidase activity was significantly decreased in all groups irrespective of sampling times (p < .05). Interleukin (IL)-1β was significantly elevated in fish of J8 group on 7th day. IL-8 increased in fish of J8 and J4 groups on 7th day of the study. IL-12 gene expression was significantly up-regulated in J8 fish group on 7th day, and in J4 fish group on 14th day. Survival rate was higher in J8 treatment compared to the control and other treatments (p < .05). The results suggest that Juniperus excelsa provides protection against Y. ruckeri in rainbow trout., (© 2020 John Wiley & Sons Ltd.)

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.

Saikosaponin a (SSa), a triterpene saponin extracted from Bupleurumfalcatum L., has been reported to have anti-inflammatory activity. In the present study, we investigated the effects of SSa on lead-induced kidney injury in common carp. Kidney histological changes were detected by H&E staining. The levels of TNF-α, IL-1β, MPO, MDA, GSH, and SOD activity were also measured. Furthermore, the NF-κB and Nrf2 signaling pathways were tested by western blot analysis. The results showed that lead-induced kidney histological change was attenuated by SSa. Lead-induced TNF-α, IL-1β, MPO, and MDA production were also suppressed by SSa. Meanwhile, lead could decrease GSH level and SOD activity and the decreases were inhibited by SSa. Furthermore, we found SSa significantly inhibited lead-induced NF-κB translocation. In addition, the expression of Nrf2 and HO-1 were increased by the treatment of SSa and Keap1 expression was decreased by SSa. In conclusion, this study indicated that SSa inhibited lead-induced kidney injury in carp through suppressing inflammatory and oxidative responses, and the mechanism may be involved in the inhibition of NF-κB and activation of Nrf2 signaling pathway., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Blunt snout bream (Megalobrama amblycephala) were randomly assigned into three diets: normal-carbohydrate diet (NCD, 30% carbohydrate, w/w), high-carbohydrate diet (HCD, 43% carbohydrate), and HCB (HCD supplemented with 50 mg/kg berberine (BBR)). After 10 weeks' feeding trial, the results showed that higher levels of plasma glucose, triglyceride, and total cholesterol were observed in HCD-fed fish than in NCD-fed fish, while HCB feeding significantly ameliorated this effect. Moreover, HCB feeding remarkably reversed HCD-induced hepatic glycogen and lipid contents. In insulin signaling, BBR inclusion restored HCD-induced suppression of insulin receptor substrate mRNA expression and elevation of forkhead transcription factor 1 mRNA expression. In glucose metabolism, upregulated glucose transporter 2 and glycogen synthase mRNA expressions in the HCD group were observed compared to the NCD group. However, BBR adding reduced the mRNA expressions of glycogen synthase, phosphoenolpyruvate carboxykinase, and glucose-6-phosphatase and increased the transcriptional levels of glucose transporter 2 and pyruvate kinase. In lipid metabolism, BBR supplementation could reverse downregulated hepatic carnitine palmitoyl transferase I mRNA expression and upregulated hepatic acetyl-CoA carboxylase and fatty acid synthetase mRNA expressions in the HCD group. Taken together, it demonstrates that BBR could improve glucose metabolism of this species via enhancing liver's glycolysis and insulin signaling, while inhibiting liver's glycogen synthesis and gluconeogenesis. It also indicates that BBR could reduce the metabolic burden of the liver by inhibiting fat synthesis and promoting lipid decomposition, and then enhance fat uptake in peripheral tissues.

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.)

Gyrodactylus spp. are common monogenean ectoparasites that may lead to significant fish mortality. To find effective anthelmintic agents with lower toxicity, a series of natural saponins were obtained and evaluated for their anthelmintic activity against Gyrodactylus kobayashii and acute toxicity to goldfish (Carassius auratus). Among all tested compounds, six compounds (1, 2, 3, 8, 10, and 13) shown higher anthelmintic activity and safety than widely used formaldehyde-based parasiticides, especially compound 1 having 100% anthelmintic efficacy against G. kobayashii at 0.3 mg/L and a therapeutic index of 16.6. Also, the three-dimensional quantitative structure-activity relationship (3D-QSAR) studies of these saponins have been performed to explore the structural features reasonable for the anthelmintic activity against G. kobayashii. These models demonstrated that the hydroxyl group at C-17 position and the sugar moieties at C-3 position, especially the hydroxyl groups of the sugar moieties, were critical to the anthelmintic activity. The QSAR studies could provide useful information for further rational design and optimization of novel saponins for the control of gyrodactylosis.

Surveillance of antibiotic resistance is of paramount importance for animal welfare and production. Despite aquaculture being a main source of animal protein, studies on antibiotic susceptibility in fish pathogens are scarce. Renibacterium salmoninarum, the aetiological agent of bacterial kidney disease (BKD), is one of the most common bacterial pathogens affecting salmon farming. In this work, we present an analysis of susceptibility patterns using determinations of minimum inhibitory concentration (MIC) for 65 field isolates, which were collected over seven years (2013-2019) from Atlantic salmon (Salmo salar) and coho salmon (Oncorhynchus kisutch) farms across southern Chile. The MIC protocol described by the Clinical Laboratory Standards Institute (CLSI) was used, but with microdilution instead of macrodilution and eight instead of four days of incubation. Two laboratories independently conducted analyses to provide data on the epidemiological cut-off values for R. salmoninarum to florfenicol, oxytetracycline and erythromycin. By using two calculation methods, our results provide evidence for an evolving subpopulation of non-wild-type isolates for the macrolide erythromycin, which is consistent with the respective treatment frequencies prescribed against BKD. Contrasting with what was expected, R. salmoninarum isolates were most susceptible to florfenicol and oxytetracycline, both of which are widely used antibiotics currently used in the Chilean salmon industry. The presented findings can serve as a reference for national or international antibiotic surveillance programmes, for both MIC interpretation and to identify emerging resistance to the conventional drugs used in BKD management. Finally, our results indicate that an 8-day incubation period for establishing MIC values of R. salmoninarum should be considered in a future revision of the CLSI guidelines., (© 2020 John Wiley & Sons Ltd.)

Soybean meal-induced enteropathy (SBMIE) is prevalent in aquaculture. The aim of this study is to evaluate the role of daidzein on SBMIE of juvenile turbot (Scophthalmus maximus L.) by feeding with fish meal diet (FM), soybean meal diet (SBM, 40% fish meal protein in FM replaced by soybean meal protein) and daidzein diet (DAID, 40 mg/kg daidzein supplemented to SBM) for 12 weeks. We found that daidzein supplementation elevated the gene expression of anti-inflammatory cytokine TGF-β, decreased gene expression of pro-inflammatory cytokines TNF-α and signal molecules p38, JNK and NF-κB. SBM up-regulated the genes expression related to oxidative stress and apoptosis, but dietary daidzein restored it to the similar level with that in FM group. Moreover, dietary daidzein up-regulated gene expression of tight junction protein, and modified the intestinal microbial profiles with boosted relative abundance of phylum Proteobacteria and Deinococcus-Thermus, genera Sphingomonas and Thermus, species Lactococcus lactis, and decreased abundance of some potential pathogenic bacteria. In conclusion, dietary daidzein could ameliorate SBM-induced intestinal inflammatory response, oxidative stress, mucosal barrier injury and microbiota community disorder of turbot. Moreover, p38, JNK and NF-κB signaling might be involved in the anti-inflammatory process of daidzein, and daidzein itself might act as an antioxidant to resist SBM-induced oxidative damage.

This study was conducted to explore the beneficial role of taurine against chronic high carbohydrate diet-induced oxidative stress, endoplasmic reticulum (ER) stress and inflammation, and to understand the underlying molecular mechanisms in turbot. Two 10-week feeding trials were simultaneously conducted. For the one, six experimental diets with graded levels of taurine supplementation (0, 0.4%, 0.8%, 1.2%, 1.6% and, 2.0%, respectively) and 15% of carbohydrate were used. For the other one, three graded levels of dietary taurine supplementation (0.4%, 1.2% and 2.0%, respectively) with 21% of carbohydrate were used. The results showed that higher expression level of inflammation cytokines and ER stress related genes were detected in higher dietary carbohydrate group. In both feeding trials, 1.2% of dietary taurine supplementation improved anti-oxidative status by decreasing the content of malondialdehyde, increasing the catalase activity and total anti-oxidative capacities. In feeding trial 1, appropriate taurine supplementation lowered contents of tumour necrosis factor-a, interleukin-6, aspartate aminotransferase and alkaline phosphatase in plasma, and decreased the expressions of pro-inflammatory cytokines, such as interleukin-8 (il-8) and interferon-γ (ifn-γ). Furthermore, dietary taurine reduced ER stress by decreasing the mRNA levels of activating transcription factor 6, protein kinase R-like endoplasmic reticulum kinase and G protein-coupled receptor 78. The optimal dietary taurine content was estimated as 1.40% based on the analysis of specific growth rate. In feeding trial 2, dietary taurine supplementation attenuated liver inflammation partly referring to significantly down-regulated mRNA levels of nuclear transcription factor-κB p65, ifn-γ, interleukin1β and up-regulate the transcript of ribosomal protein S6 kinase 1. Dietary taurine supplementation in feeding trial 2 significantly increased the Nrf2-related factor 2 protein level and decreased the NFκB p65 protein level only at 21% of dietary carbohydrate level. Taurine can alleviate the oxidative damage and inflammation caused by 21% of dietary carbohydrate to a certain degree. Overall, the present study confirmed that dietary taurine supplementation improved growth performance and anti-oxidative response, and reduced liver inflammatory and ER stress processes induced by high dietary carbohydrate in turbot., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Cyprinid herpesvirus 2 (CyHV-2), which infects silver crucian carp including goldfish (Carassius auratus auratus) and Crucian carp (Carassius auratus gibelio) with high mortality, is an emerging viral pathogen worldwide. Previous studies showed that berberine (BBR), a bioactive plant-derived alkaloid, demonstrated potential antiviral actions against many different viruses. Here, we assessed the effect of berberine hydrochloride (BBH) on the replication of CyHV-2 in vitro and in vivo. Cytotoxicity assay indicated that 5-25 μg/mL BBH was non-toxic to the RyuF-2 cells. In viral inhibition assays, real time PCR was employed to titrate the genomic copy number of progeny virus, real time RT-PCR was applied to monitor the transcriptional levels of viral genes, and Western blot analysis was performed to detect the synthetic levels of viral proteins. The results demonstrated that BBH systematically impedes the viral gene transcription and suppressed the replication of CyHV-2 in RyuF-2 cells. In animal challenge test, BBH was confirmed to protect Crucian carps from CyHV-2 infection in a dose-dependent manner, which was supported by suppressed viral replication levels, reduced viral pathogenesis and higher survival rates. Furthermore, pharmacokinetics data of BBH in Crucian carp revealed its rapid absorption (T max of 1.5 h), suitable plasma half-life (t 1/2 z/h of 7-12 h depending on oral dosage), and dose-dependent drug exposure properties following oral administration (revealed by AUC 0-t values). These findings shed light on repurposing BBH to treat CyHV-2 infections in silver crucian carp., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Bio-nanotechnology employing bio-sourced nanomaterial is an emerging avenue serving the field of fish medicine. Marine-sourced chitosan nanoparticles (CSNPs) is a well-known antimicrobial and immunomodulatory reagent with low or no harm side effects on fish or their human consumers. In this study, in vitro skin mucus and serum antibacterial activity assays along with intestinal histology, histochemical, and gene expression analyses were performed to evaluate the impact of dietary CSNPs (5 g kg -1 dry feed) on rainbow trout resistance against 'enteric redmouth' disease. Two treatment conditions were included; short-term prophylactic-regimen for 21 days before the bacterial challenge, and long-term therapeutic-regimen for 21 days before the challenge and extended for 28 days after the challenge. Our results revealed higher antibacterial defense ability and positive intestinal histochemical and molecular traits of rainbow trout after dietary CSNPs. The prophylactic-regimen improved trout health while the therapeutic regimen improved their disease resistance and lowered their morbidity. Therefore, it is anticipated that CSNPs is an effective antibacterial and immunomodulatory fish feed supplement against the infectious threats. However, the CSNPs seem to be more effective in the therapeutic application rather than being used for short-term prophylactic applications.

The channel catfish virus (CCV) can cause lethal hemorrhagic infection in channel catfish, resulting in significant economic losses in the fish industry. Effective drugs for the virus are still lacking. Acyclovir is known as a potent antiviral agent against human herpes viruses and some animal DNA viruses. The present study was undertaken to explore the antiviral response and mechanism of acyclovir against CCV in channel catfish ovary (CCO) cells. Acyclovir was able to significantly inhibit the expression of viral genes related to CCV viral DNA synthesis and suppress viral replication at a safe concentration. Furthermore, acyclovir blocked the cytopathic effects and apoptosis induced by CCV, thereby maintaining the normal cellular morphological structure, as shown by the protection of CCO cells from the formation of apoptotic bodies or nuclear fragmentation. Moreover, reverse transcript quantitative polymerase chain reaction (RT-qPCR) demonstrated that acyclovir suppressed the expression of caspase 3, caspase 8 and caspase 9, while there was no significant impact on the expression of the apoptosis-inhibiting gene bcl-2 in CCV-infected cells. In addition, acyclovir did not promote the expression of immune-related genes such as MyD88, Mx1, IRF3, IRF7, IFN-I, NF-kB and IL-1β, suggesting that the antiviral activity of acyclovir to CCV infection is not achieved by facilitating the expression of immune-related genes in CCO cells. Taken together, the results from this study suggest that acyclovir could effectively regulate CCV-induced infection, and thus is a promising therapeutic agent against CCV. Our results will aid our understanding of the pharmacological mechanisms of antiviral agents., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Phenylpropanoids, common natural compounds, possess many different biological activities such as antioxidant, anti-inflammatory and antiviral. Spring viraemia of carp virus (SVCV) can cause a high mortality in common carp (Cyprinus carpio). However, there are currently no licenced drugs that effectively cure this disease. In this study, we designed and synthesized a phenylpropanoid derivative 4-(4-methoxyphenyl)-3,4-dihydro-2H-chromeno[4,3-d]pyrimidine-2,5(1 H)-dione (E2), and explored the antiviral effect against SVCV in vitro and in vivo. Up to 25 mg/L of E2 significantly inhibited the expression levels of SVCV protein genes in the epithelioma papulosum cyprini (EPC) cell line by a maximum inhibitory rate of >90%. As expected, E2 remarkably declined the apoptotic of SVCV-infected cells and suppressed potential enhancement of the mitochondrial membrane potential (ΔΨm), these data implied that E2 could protect mitochondria from structural damage in response to SVCV. Meanwhile, E2 was added to EPC cells under four different conditions: time-of-addition, time-of-removal, pre-treatment of viruses and pre-treatment of cells indicated that E2 may block the post-entry transport process of the virus. Additionally, the up-regulation of six interferon (IFN)-related genes also demonstrated that E2 indirectly activated IFNs for the clearance of SVCV in common carp. Drug cure effect showed that treatment with E2 at 0.5 d post infection (dpi) is more effective than at 0, 1 or 2 dpi. Most importantly, intraperitoneal therapy of E2 markedly improved common carp survival rate and reduced virus copies in body. Therefore, the E2 has potential to be developed into a novel anti-SVCV agent., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background and Objective: Sessiline ciliates live as eco commensals (low numbers) and parasites (high numbers) on different hosts, like mollusks copepods, mysids and fish. Riboscyphidia ecto-protozoan is moderately pathogenic but high numbers of it on the gills can physically prevent gas exchange. The present study aimed to describe the epizoic ciliates Riboscyphidia found on the Red Sea cultured Asian sea bass and obtain more information on the Epidemiology of the parasite with special references to control and histopathological examination of naturally infected sea bass., Materials and Methods: The occurrence of epizoic ciliates on the adult Asian Sea bass. About 100 Asian sea bass were collected by the fishing net at a private marine fish farm at Ismailia governorate and transferred to the hydrobiology laboratory at National Research Centre. A parasitological and histopathological study of epizoic sessile ciliate species was done. ANOVA test was used for Statistical analysis., Results: Riboscyphidia sp. was found and isolated after parasitological examination of investigated adult's Asian sea bass. The prevalence of Riboscyphidiosis was 64%. Sessile ciliates were found on gills, skin and fins. The clinical signs of Riboscyphidiosis were respiratory distress, flashing and off food. Histopathological alterations in naturally infested Asian sea bass were investigated., Conclusion: The treatment of choice of Riboscyphidiosis was prolonged immersion by Copper citrate with a dose of 0.56 mg mL-1 for 7 days.

Background: The only remedy for up surging problem of antibiotic resistance is the discovery of antibacterial agents of natural origin., Objective: The present study was aimed at finding antibacterial potential of crude and solvent extracts of mature leaves of Plumeria pudica., Methods: Antibacterial activity of three different solvent extracts were evaluated in four human and four fish pathogenic bacteria by measuring the zone of inhibition and determining Minimum Inhibitory Concentration and Minimum Bactericidal Concentration values. Standard antibiotics were used as positive control. Preliminary phytochemical screening of most effective extract i.e., ethyl acetate extract, Fourier Transform Infra Red analysis and GC-MS analysis of the Thin Layer Chromatographic (TLC) fraction of ethyl acetate extract were done meticulously. All experiments were done thrice and analyzed statistically., Results: Crude leaf extracts and solvent extracts caused good inhibition of bacterial growth in all selected bacteria. Ethyl acetate extract showed highest inhibition zones in all tested strains with maximum inhibition (19.50±0.29 mm) in Escherichia coli (MTCC 739). MBC/MIC of the extracts indicated that all three solvent extracts were bactericidal. Preliminary phytochemical tests revealed the presence of tannins, steroids and alkaloids and FT-IR analysis revealed presence of many functional groups namely alcoholic, amide, amine salt and aldehyde groups. From the GC-MS analysis of TLC fraction of ethyl acetate extract, five different bioactive compounds e.g., 2,4-ditert -butylphenyl 5-hydroxypentanoate, Oxalic acid; allyl nonyl ester, 7,9-Ditert-butyl-1-oxaspiro(4,5)deca-6,9-diene- 2,8-dione, Dibutyl phthalate and 2,3,5,8-tetramethyl-decane were identified., Conclusion: Leaf extracts of P. pudica contain bioactive compounds that can be used as broad spectrum bactericidal agent., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Background and Objective: Leek (Allium ampeloprasum) is one of the most commonly used herbal foods all over the world. This study was conducted to evaluate the protective effect of leek extract on catfish experimentally challenged with Aeromonas hydrophila, a problematic bacterial pathogen that affects various freshwater fish species., Materials and Methods: Aeromonas hydrophila was isolated and identified from catfish showing clinical signs of septicemia. The in vitro activity of leek extract to control the growth of Aeromonas hydrophila was investigated. In the in vivo experiment, about 240 adult catfish (Clarias gariepinus) were fed three different leek extract concentrations (10, 25 and 50 mg kg-1 body weight) for 1 month. Later on, a challenge study was conducted using an identified A. hydrophila strain. Morbidity and mortality were recorded throughout one week post-challenge. Furthermore, the effect of leek extract on some immune-related genes was investigated., Results: Under the in vitro testing, a significant increase (10 and 13 mm) in the inhibition zone was recorded in wells treated with 25 and 50 mg L-1 leak extract, respectively. A significant reduction in fish mortalities was reported in all leek extract treated groups compared to the control group which was given water. TLR1 gene expression was upregulated in fish treated with leek extract while TNFα gene expression was down-regulated., Conclusion: Overall, results suggested that the leek extract has immunostimulating effects that can help control bacterial infections in catfish and probably other fish species.

Selenium, as an essential trace element, interferes through selenoproteins in many physiological processes of plants and mammals. Its antiviral activity has recently attracted much attention because selenium improves the antiviral capacity of animal cells against a few viruses relevant to human diseases. In this study, the red elemental selenium was purified from the fermentative culture of Herbaspirillum camelliae WT00C and then used to culture epithelioma papulosum cyprinid (EPC) cells or feed crucian carp and zebrafish. Finally, its antiviral effects were investigated at the cell level and living fishes after spring viraemia of carp virus infection. At the cell level, 5, 10 and 20 μg ml -1 red elemental selenium significantly induced the expression of interferon (IFN) and ISG15 genes in EPC cells. The viral TCID 50 (50% tissue culture infective dose) values in the EPC cells incubated with 5, 10 and 20 μg ml -1 red elemental selenium were significantly less than those of the control. More expression of IFN and ISG15 genes and less TCID 50 values indicate that red elemental selenium indeed improves the antiviral capability of EPC cells. In the crucian carp fed with the food containing 5 and 10 μg g -1 red elemental selenium, IFN expressions showed 13- and 39-fold increases at the 16th day of post-injection, and its expression was dependent on selenium concentrations. Meanwhile, no fish death occurred in all the experimental groups. In the zebrafish fed with the red worm containing 5 μg g -1 red elemental selenium, IFN and Mx expressions and survival rate were significantly higher than those of the control. The results of this study show that red elemental selenium indeed improves the antiviral activity of fish. The antiviral effects of selenium mainly come from its immune regulation through its incorporation into selenoproteins. The optimum level of selenium contributes to improving fish immunity, whereas excess selenium causes excessive immune and inflammatory responses., (© 2020 Fisheries Society of the British Isles.)