Anthelmintic efficacy of natural saponins against Gyrodactylus kobayashii in goldfish (Carassius auratus) and their 3D-QSAR analysis.

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.

Molecular Effects of Silver Nanoparticles on Monogenean Parasites: Lessons from Caenorhabditis elegans.

The mechanisms of action of silver nanoparticles (AgNPs) in monogenean parasites of the genus Cichlidogyrus were investigated through a microarray hybridization approach using genomic information from the nematode Caenorhabditis elegans. The effects of two concentrations of AgNPs were explored, low (6 µg/L Ag) and high (36 µg/L Ag). Microarray analysis revealed that both concentrations of AgNPs activated similar biological processes, although by different mechanisms. Expression profiles included genes involved in detoxification, neurotoxicity, modulation of cell signaling, reproduction, embryonic development, and tegument organization as the main biological processes dysregulated by AgNPs. Two important processes (DNA damage and cell death) were mostly activated in parasites exposed to the lower concentration of AgNPs. To our knowledge, this is the first study providing information on the sub-cellular and molecular effects of exposure to AgNPs in metazoan parasites of fish.

Copper and cadmium administration induce toxicity and oxidative stress in the marine flatworm Macrostomum lignano.

The contamination of coastal regions with different toxicants, including heavy metal ions such as copper and cadmium jeopardize health and survival of organisms exposed to this habitat. To study the effects of high copper and cadmium concentrations in these marine environments, we used the flatworm Macrostomum lignano as a model. This platyhelminth lives in shallow coastal water and is exposed to high concentrations of all toxicants that accumulate in these sea floors. We could show that both, cadmium and copper show toxicity at higher concentrations, with copper being more toxic than cadmium. At concentrations below acute toxicity, a reduced long-term survival was observed for both metal ions. The effects of sublethal doses comprise reduced physical activities, an increase in ROS levels within the worms, and alterations of the mitochondrial biology. Moreover, cell death events were substantially increased in response to sublethal concentrations of both metal ions and stem cell activity was reduced following exposure to higher cadmium concentrations. Finally, the expression of several genes involved in xenobiotic metabolism was substantially altered by this intervention. Taken together, M. lignano has been identified as a suitable model for marine toxicological studies as it allows to quantify several relevant life-history traits as well as of physiological and behavioral read-outs.

Development of a non-lethal hydrogen peroxide treatment for surveillance of Gyrodactylus salaris on trout farms and its application to testing wild salmon populations.

This study documents the development of a non-lethal sampling method to recover gyrodactylid parasites from large numbers of fish that will underpin an improved surveillance strategy for Gyrodactylus salaris. A review of published literature identified over 80 compounds that have previously been tested against gyrodactylids or closely related parasite species. Five safe and relatively fast-acting compounds were selected for testing to determine their efficiency in removing gyrodactylids from host fish in small-scale aquaria trials using three-spined stickleback infected with Gyrodactylus gasterostei as a model host-parasite system. The most effective compound was hydrogen peroxide; short-duration exposure (3 min) achieved a parasite detection sensitivity of 80%-89%. The practicality of exposing farmed salmonids to hydrogen peroxide for G. salaris surveillance was tested in the field by conducting a parasite recovery trial using a brown trout stock endemically infected with G. truttae and G. derjavinoides and comparing this to the whole-body examination procedure currently conducted by UK authorities. Significantly more parasites were recovered after exposing fish to hydrogen peroxide and filtering the treatment solution than by direct whole-body examination of killed fish (mean: 225 vs. 138 parasites per fish). The gyrodactylid recovery rate of the two methods was 84.6% and 51.9%, respectively. A comparison of timings for the two methods indicated scope for significant time savings in adopting the chemical screening method. The study demonstrated that hydrogen peroxide bath treatment may be successfully applied to the surveillance of gyrodactylid parasites and established as a non-lethal method for sampling farmed and wild fish. This approach has the potential to reduce resources required to collect and isolate parasites for diagnostic testing and improve the sensitivity and confidence of surveillance programmes designed to demonstrate freedom from disease, thus underpinning a robust and defensible surveillance strategy for G. salaris for the UK aquatic animal disease contingency plan.

Use of praziquantel to control platyhelminth parasites of fish.

Fish are common definitive and intermediate hosts for a variety of parasitic flatworms. In unstressed wild populations, parasitic infections often go unnoticed and are perceived to represent a lesser threat to fish health. In contrast, platyhelminth parasitism of captive fish often results in decreased weight gain and increased mortality which often necessitates chemotherapeutic treatment. The presence of platyhelminth parasites in fish tissues is not only unappealing but in some cases also represents a threat to human health. In veterinary medicine, one of the most commonly used agents with anti-flatworm activity is praziquantel; yet, no praziquantel products are labeled for use in fish in the United States. Veterinarians may use praziquantel preparations approved for other vertebrate species under the Animal Medicinal Drug Use Clarification Act (AMDUCA). However, such extra-label use should be informed by scientific evidence including efficacy and tissue residue studies. Herein, we review studies testing the efficacy of praziquantel for treatment of platyhelminthes along with an assessment of routes of administration, pharmacokinetics, and toxicity information.

Anthelmintic activity of Rosmarinus officinalis against Dactylogyrus minutus (Monogenea) infections in Cyprinus carpio.

Monogenean parasites are important ectoparasites of fish, and are responsible for severe economic impacts in the aquaculture industry. They are usually treated with chemicals, but the chemicals can have harmful side effects in the fish and may pose threats to human health. Rosemary (Rosmarinus officinalis) is a common medicinal herb, with antimicrobial and antitumor properties. Here, we examined the anthelmintic activity of rosemary extract against the monogenean (Dactylogyrus minutus) in vitro and in vivo using bath treatment and oral administration. The in vitro experiments showed that parasite survival was affected by both rosemary extract concentration and the solvent (water and ethanol). Parasites were dead at 61.8±5.6 and 7.8±1.4min when exposed to 100 and 200g aqueous rosemary extract solution/L of water respectively. It took 166.7±48.2 and 5.4±1.01min to kill the parasites when exposed to 1 and 32g ethanol rosemary extract solution/L of water respectively. Moreover, pure component of rosemary extract obtained commercially used in in vitro experiments showed that 1,8-Cineole was the most toxic component of the main components tested. Parasite intensity and prevalence in fish exposed to 50 and 100g aqueous rosemary solution/L water for 30min were significantly lower than they were in controls (p<0.05). In oral treatment experiments, diets of Cyprinus carpio were supplemented with eight different concentrations of aqueous rosemary extract. The intensity of parasites was significantly less in fish fed for 30days with feed containing 60, 80 and 100ml aqueous extract/100g feed than in control (p<0.05). Together these results indicate that rosemary is a promising candidate for prevention and control of monogenean infection.

Synthesis and anthelmintic activity of arctigenin derivatives against Dactylogyrus intermedius in goldfish.

To control the parasitic disease of Dactylogyrus intermedius, a series of new arctigenin derivatives were designed, synthesized and tested in our study. The anthelmintic activity of most of the derivatives ranged from 1 to 10mg/L. Compared to traditional drug praziquantel (EC50=2.69mg/L), ether derivatives 2g and 2h exhibited slightly higher anti-parasitic activity, with the EC50 values of 2.48 and 1.52mg/L, respectively. Furthermore, the arctigenin-imidazole hybrids 4a and 4b also removed D. intermedius effectively, with the EC50 values of 2.13 and 2.07mg/L, respectively. The structure-activity relationship analysis indicated that four carbon atoms length of linker and imidazole substitute group could significantly increase the anthelmintic activity, and reduced the toxicity. Through the scanning electron microscope observation, compounds 4a and 4b caused the D. intermedius tegumental damage such as intensive wrinkles, holes and nodular structures. Overall, the structural optimization analysis of arctigenin suggested that 4a and 4b can be used for preventing and controlling Dactylogyrus infections and considered as promising lead compounds for the development of commercial drugs.

Anesthesia by sprinkling method in the gills of tambaqui Colossoma macropomum does not influence intensity and morphology of monogeneans / Anestesia por meio de aspersão nas brânquias de tambaqui Colossoma maropomum não influencia a intensidade e a morfologia de monogeneas

Abstract The present study evaluates the influence of anesthesia on the parasitic fauna of monogenea fish parasites, as its intensity and viability. Two experiments were conducted: Evaluation of an anesthetic method by sprinkling eugenol directly on gills and evaluation of monogenea motility and viability; Comparison of immersion and directly sprinkling on the gills with benzocaine and eugenol followed by evaluation on parasite intensity. The results suggest that the anesthetic sprinkling didn't interfere in the parasite motility, morphology and body surface integrity analyzed by fluorescence method. The monogenean intensity in the gills was lower in fish anesthetized by immersion method compared to the sprinkling method and the control group. This method of anesthesia can be used in parasitological studies.

Resumo O presente estudo avalia a influência da anestesia sobre a fauna parasitária de monogeneas em peixes, sua intensidade e sua viabilidade. Dois experimentos foram realizados: Avaliação de um método anestésico por aspersão eugenol diretamente nas brânquias e avaliação da motilidade das monogeneas e sua viabilidade; e Comparação entre imersão e aspersão diretamente nas brânquias com benzocaína e eugenol, seguido de avaliação sobre a intensidade parasitária. Os resultados sugerem que a aspersão do anestésico não interferiu na motilidade, morfologia, superfície corporal e integridade do parasita, analisadas pelo método de fluorescência. A intensidade de monogenéticos nas brânquias foi menor nos peixes anestesiados pelo método de imersão em comparação com o método de aspersão e o grupo controle. O método de anestesia por aspersão nas brânquias pode ser utilizado em estudos parasitológicos.

Anesthesia by sprinkling method in the gills of tambaqui Colossoma macropomum does not influence intensity and morphology of monogeneans.

The present study evaluates the influence of anesthesia on the parasitic fauna of monogenea fish parasites, as its intensity and viability. Two experiments were conducted: Evaluation of an anesthetic method by sprinkling eugenol directly on gills and evaluation of monogenea motility and viability; Comparison of immersion and directly sprinkling on the gills with benzocaine and eugenol followed by evaluation on parasite intensity. The results suggest that the anesthetic sprinkling didn't interfere in the parasite motility, morphology and body surface integrity analyzed by fluorescence method. The monogenean intensity in the gills was lower in fish anesthetized by immersion method compared to the sprinkling method and the control group. This method of anesthesia can be used in parasitological studies.

Hatchability and survival of oncomiracidia of Paradiplozoon ichthyoxanthon (Monogenea: Diplozoidae) exposed to aqueous aluminium.

BACKGROUND: Monogenea is a diverse group of ectoparasites showing great potential as sentinel organisms for monitoring environmental health. Exposure to metals negatively affects infrapopulations of monogeneans and exposure to aluminium has been found to negatively impact the survival of gyrodactylids. METHODS: Samples of infected host fish, the smallmouth yellowfish Labeobarbus aeneus (Cyprinidae), were collected from the Vaal Dam, South Africa and transported back to the laboratory in dark 160 l containers. Eggs of the monogenean Paradiplozoon ichthyoxanthon infecting L. aeneus were collected and exposed to varying concentrations of aluminium along with a control group in static tanks. The eggs were checked every 24 h and hatching commenced 13-14 days after exposure. Water samples were taken from exposure tanks and acidified for analysis of Al levels with inductively-coupled plasma mass spectrometry. RESULTS: Hatching of eggs was variable between exposures, and in 30 µg Al/l and 60 µg Al/l was found to occur before eggs in control beakers, whereas, exposure to 120 µg Al/l delayed hatching and reduced hatchability. Survival of hatched oncomiracidia was concentration dependent and negatively correlated with aluminium concentrations. Lowest survival was recorded for 60 µg Al/l and 120 µg Al/l where all larvae died shortly after or during hatching. Normal development of embryos of P. ichthyoxanthon within eggs exposed to all doses of aluminium indicates that the egg shell is moderately impermeable to metals and inhibits movement of aluminium across the shell and interacting with developing embryos. CONCLUSIONS: Higher larval mortality rate in 120 µg/l exposure can be related to aluminium crossing the egg shell in the late stages and causing death of unhatched yet fully developed embryos, possibly due to changes in the permeability of the egg shell as embryos neared developmental completion. Accelerated death of oncomiracidia after hatching indicates sensitivity toward high concentrations of aluminium.

The use of Ocimum americanum essential oil against the pathogens Aeromonas hydrophila and Gyrodactylus sp. in silver catfish (Rhamdia quelen).

UNLABELLED: The bactericidal activity (minimum inhibitory concentration (MIC)-test) of Ocimum americanum (inflorescences) essential oil (OAEO) against Aeromonas hydrophila was determined in this study. Also investigated was the potential of OAEO and the main compound found in the oil (linalool) at subinhibitory concentrations to be inhibitors of haemolysis caused by Aer. hydrophila in fish erythrocytes. An in vivo experiment was conducted to evaluate the survival of fish (Rhamdia quelen) experimentally infected with Aer. hydrophila and exposed to OAEO. A second experiment was conducted to evaluate the in vitro and in vivo activity of OAEO (mix from inflorescences and leaves) against the parasite Gyrodactylus sp. The OAEO showed weak in vitro activity against Aer. hydrophila (6400 µg ml(-1) ). Subinhibitory concentrations of OAEO (100 µg ml(-1) ) inhibited haemolysis (90%) caused by Aer. hydrophila in fish erythrocytes, however, linalool did not inhibit haemolysis activity. At the low concentrations (10 and 20 mg l(-1) ) added to the water, OAEO promoted the survival of fish experimentally infected with Aer. hydrophila. Lastly, the OAEO mix (50 mg l(-1) ) was effective against Gyrodactylus sp., significantly reducing (60%) the number of parasites in the fish. SIGNIFICANCE AND IMPACT OF THE STUDY: Phytochemicals, such as essential oils (EOs) are a great source of new molecules and have shown potential to be used in aquaculture systems. However, additional studies focused on the in vivo efficacy, mode of action and identification of the active compounds are needed. This study determined the potential of Ocimum americanum EO for use against two important fish pathogens, Aeromonas hydrophila and Gyrodactylus sp., as well as providing preliminary information about the role of the main EO compound (linalool) against Aer. hydrophila virulence.

Antihelminthic potential of quinacrine and oxyclozanide against gill parasite Microcotyle sebastis in black rockfish Sebastes schlegeli.

The aim of this study was to assess the treatment potential of quinacrine and oxyclozanide against Microcotyle sebastis (Monogenea: Polyopisthocotylea) infection in cultured black rockfish Sebastes schlegeli. The oral administration of quinacrine led to a reduction in the mean abundance of M. sebastis infection in all quinacrine-treated groups, and the groups of fish administered quinacrine at 50, 100, and 200 mg kg(-1) for 3 consecutive days showed a parasite mean abundance that was 50 to 30% lower compared to that of the control group, suggesting that quinacrine has a therapeutic potential against M. sebastis. Although oxyclozanide showed a very high in vitro killing activity, in oral administration experiments, only the groups of fish administered 200 mg kg(-1) showed less than 50% mean abundance of M. sebastis compared to the control groups, suggesting that the absorption efficiency of orally administered oxyclozanide might be low in black rockfish and/or that M. sebastis might be less sensitive to orally ingested oxyclozanide. As praziquantel has been the sole therapeutic against M. sebastis infection in Korea for a long time, a broadening of available control measures is advisable in order to reduce the possible emergence of praziquantel-resistant M. sebastis. In our study, although quinacrine and oxyclozanide showed a therapeutic potential against M. sebastis, the treatment efficacy was not high enough to replace praziquantel. Thus, after investigations on the pathological effects and pharmacodynamics, use of quinacrine or oxyclozanide in combination with praziquantel may be considered as a way to prevent praziquantel resistance in M. sebastis.

Fighting fish parasites with photodynamically active chlorophyllin.

Water-soluble chlorophyll (chlorophyllin) was used in a phototoxic reaction against a number of fish ectoparasites such as Ichtyobodo, Dactylogyrus, Trichodina, and Argulus. Chlorophyllin is applied to the water at concentrations of several micrograms per milliliter for a predefined incubation time, and afterwards, the parasites are exposed to simulated solar radiation. Application in the dark caused only little damage to the parasites; likewise, light exposure without the addition of the photosensitizer was ineffective. In Ichthyobodo, 2 µg/mL proved sufficient with subsequent simulated solar radiation to almost quantitatively kill the parasites, while in Dactylogyrus, a concentration of about 6 µg/mL was necessary. The LD50 value for this parasite was 1.02 µg/mL. Trichodina could be almost completely eliminated at 2 µg/mL. Only in the parasitic crustacean Argulus, no killing could be achieved by a photodynamic reaction using chlorophyllin. Chlorophyllin is non-toxic, biodegradable, and can be produced at low cost. Therefore, we propose that chlorophyllin (or other photodynamic substances) are a possible effective countermeasure against several ectoparasites in ponds and aquaculture since chemical remedies are either forbidden and/or ineffective.

Killing of Gyrodactylus salaris by heat and chemical disinfection.

BACKGROUND: Gyrodactylus salaris is a monogenean, which has collapsed tens of wild Atlantic salmon populations. One of the means of preventing the spread of the parasite is the disinfection of the fishing equipment, which is used in the rivers having susceptible salmon populations. Little is known about the dosage of disinfectants against G. salaris. There are not standards for the testing of disinfectants against multicellular parasites. The present investigation developed a method to test disinfectants and examined the effectiveness of heated water and a commercially available disinfectant (Virkon S) in killing G. salaris. Individual G. salaris worms were followed under the microscope during treatment with heated water or Virkon S disinfectant blend. The logarithm of the time needed to kill the parasite was used as a dependent variable in linear regression. The upper 99.98 % prediction line for the dependent variable was used to obtain a value resembling the time needed for a 4 log reduction of the microbial pathogen, which is commonly used as a criterion for disinfectants. Also 6 log reduction was applied. RESULTS: Exposure to a relatively low temperature was found to kill the parasite. Even 5-50 min treatment (=10-100 times the 99.98 % upper prediction value) with heated water at 40 °C might be used. This would enable the utilisation of hot tap water in the disinfection of fishing gear. The present practice of 1 % Virkon S for 15 min was also found to kill the parasite. CONCLUSIONS: The follow-up of single parasites of a test population and the use of the calculated upper predictive line in the regression analysis offers a method to analyse the effects of disinfectants on parasites like G. salaris. The results of our tests give possibilities for using disinfection methods, which may be more acceptable by the fishermen than the present ones.

Anthelmintic efficacy of cinnamaldehyde and cinnamic acid from cortex cinnamon essential oil against Dactylogyrus intermedius.

Utilization of chemical pesticide to control monogenean diseases is often restricted in many countries due to the development of pesticide resistance and concerns of chemical residues and environmental contamination. Thus, the use of antiparasitic agents from plants has been explored as a possible way for controlling monogenean infections. Extracts from Cinnamomum cassia were investigated under in vivo conditions against Dactylogyrus intermedius in goldfish. The two bioactive compounds, cinnamaldehyde and cinnamic acid, were identified using nuclear magnetic resonance and electrospray ionization mass spectrometry. The 48 h median effective concentrations (EC(50)) for these compounds against D. intermedius were 0·57 and 6·32 mg L(-1), respectively. The LD(50) of cinnamaldehyde and cinnamic acid were 13·34 and 59·66 mg L(-1) to goldfish in 48 h acute toxicity tests, respectively. These data confirm that cinnamaldehyde is effective against D. intermedius, and the cinnamaldehyde exhibits potential for the development of a candidate antiparasitic agent.

Ergot Alkaloids (Re)generate New Leads as Antiparasitics.

Praziquantel (PZQ) is a key therapy for treatment of parasitic flatworm infections of humans and livestock, but the mechanism of action of this drug is unresolved. Resolving PZQ-engaged targets and effectors is important for identifying new druggable pathways that may yield novel antiparasitic agents. Here we use functional, genetic and pharmacological approaches to reveal that serotonergic signals antagonize PZQ action in vivo. Exogenous 5-hydroxytryptamine (5-HT) rescued PZQ-evoked polarity and mobility defects in free-living planarian flatworms. In contrast, knockdown of a prevalently expressed planarian 5-HT receptor potentiated or phenocopied PZQ action in different functional assays. Subsequent screening of serotonergic ligands revealed that several ergot alkaloids possessed broad efficacy at modulating regenerative outcomes and the mobility of both free living and parasitic flatworms. Ergot alkaloids that phenocopied PZQ in regenerative assays to cause bipolar regeneration exhibited structural modifications consistent with serotonergic blockade. These data suggest that serotonergic activation blocks PZQ action in vivo, while serotonergic antagonists phenocopy PZQ action. Importantly these studies identify the ergot alkaloid scaffold as a promising structural framework for designing potent agents targeting parasitic bioaminergic G protein coupled receptors.

Anthelmintic Effect of Biocompatible Zinc Oxide Nanoparticles (ZnO NPs) on Gigantocotyle explanatum, a Neglected Parasite of Indian Water Buffalo.

Helminth parasites of veterinary importance cause huge revenue losses to agrarian economy worldwide. With the emergence of drug resistance against the current formulations, there is a need to focus on the alternative approaches in order to control this menace. In the present study, biocompatible zinc oxide nanoparticles (ZnO NPs) were used to see their in vitro effect on the biliary amphistomes, Gigantocotyle explanatum, infecting Bubalus bubalis because these nanoparticles are involved in generation of free radicals that induce oxidative stress, resulting in disruption of cellular machinery. The ZnO NPs were synthesized by using egg albumin as a biotemplate and subsequently characterized by Scanning Electron Microscopy (SEM), Transmission Electron Microscopy (TEM), X-ray Diffraction and Spectrophotometrical, which showed that ZnO NPs were highly purified wurtzite type polycrystals, with a mean size of 16.7 nm. When the parasites were treated with lower concentrations (0.004% and 0.008%) of the ZnO NPs, the worms mounted a protective response by stimulating the antioxidant system but the treatment of G. explanatum with 0.012% ZnO NPs produced significant inhibition of the antioxidant enzymes like superoxide dismutase (SOD) (p< 0.05) and glutathione S- transferase (GST) (p<0.01), while the level of malondialdehyde (MDA), a lipid peroxidation marker, was significantly (p< 0.01) elevated. SEM and histopathology revealed pronounced tegumental damage showing the disruption of surface papillae and the annulations, particularly in the posterior region near acetabulum. The under expression of a number of polypeptides, loss of worm motility in a time dependent manner, further reflect strong anthelmintic potential of ZnO NPs. It can be concluded that the anthelmintic effect might be due to the production of reactive oxygen species that target a variety of macromolecules such as nucleic acid, protein and lipids which are involved in different cellular processes.

In vitro immunocompetence of two compounds isolated from Polygala tenuifolia and development of resistance against grass carp reovirus (GCRV) and Dactylogyrus intermedius in respective host.

The present study was undertaken to isolate some compounds from methanol extract of Polygala tenuifolia and evaluate their immunostimulatory properties and antiviral activity using grass carp Ctenopharyngodon idella kidney (CIK) cells and GCRV. By applying insecticidal bioassay-guided, chromatography techniques and successive recrystallization, two purified compounds were obtained. The changes of expression of selected immune genes (Mx1, IL-1ß, TNFα, MyD88 and IgM) in C. idella kidney cell lines were evaluated after exposure to these isolated compounds. The results showed that compound 1 and 2 up-regulated to varying degrees of Mx1, IL-1ß, TNFα, and MyD88 in C. idella kidney cells. WST-8 kit assay verified the two compounds has no toxic effects on CIK cell, and furthermore, have in vitro antivirus activity. Especially, that there is keeping 79% cell viability when exposure to compound 2 (100 mg L(-1)). According to in vivo insecticidal assays against Dactylogyrus intermedius, compound 2 exhibited higher efficacy than compound 1, which was found to be 87.2% effective at the concentrations of 5 mg L(-1) and safe to goldfish (Carassius auratus). Besides, the purified compounds were identified by spectral data as: (1) 1,5-Anhydro-D-glucitol and (2) 3,4,5-trimethoxy cinnamic acid. Overall, the results indicate that bath administration of these compounds modulates the immune related genes in C. idella kidney cells and to some extent, eliminate the virus and parasitic infections.

Screening Medicinal Plants for Use against Dactylogyrus intermedius (Monogenea) Infection in Goldfish.

Abstract Methanol extracts of 24 traditional medicinal plants with potential anthelmintic activity against Dactylogyrus intermedius (Monogenea) in Goldfish Carassius auratus were investigated. Abrus cantoniensis, Citrus medica, Dioscorea collettii, and Polygonum multiflorum exhibited 100% activity and were selected for further evaluation by applying five solvents (petroleum ether, chloroform, ethyl acetate, methanol, and water) for the extraction of the samples, followed by an in vivo bioassay. Among the plants tested, water, methanol, and ethyl-acetate extracts of P. multiflorum showed the highest efficacies; EC50 values (median concentration that results in 50% of its maximal effect) were 1.9, 5.4, and 9.1 mg/L, respectively, and extracts showed 100% efficacy against Dactylogyrus intermedius at 100, 12.5, and 25 mg/L. This was followed by ethyl-acetate, chloroform, and methanol extracts of Dioscorea collettii, which demonstrated 100% efficacy at 80, 80, and 120 mg/L and had EC50 values of 19.7, 27.1, and 37.8 mg/L, respectively, after 48 h of exposure. Chloroform and ethyl-acetate extracts of C. medica, which exhibited 100% efficacy against Dactylogyrus intermedius at 100 and 125 mg/L, revealed similar activity and had EC50 values of 58.7 and 51.3 mg/L, respectively. The ethyl-acetate and methanol extracts of A. cantoniensis exhibited the lowest activity and had EC50 values of 279.4 and 64.3 mg/L. Acute toxicities of these active extracts were investigated on Goldfish for 48 h. The findings indicated that extracts of the four plants can be developed as a preferred natural antiparasitic for the control of D. intermedius. Received June 15, 2013; accepted February 11, 2014.

Efficacy of garlic (Allium sativum) extract applied as a therapeutic immersion treatment for Neobenedenia sp. management in aquaculture.

Garlic, Allium sativum L., extract administered as a therapeutic bath was shown to have antiparasitic properties towards Neobenedenia sp. (MacCallum) (Platyhelminthes: Monogenea) infecting farmed barramundi, Lates calcarifer (Bloch). The effect of garlic extract (active component allicin) immersion on Neobenedenia sp. egg development, hatching success, oncomiracidia (larvae) longevity, infection success and juvenile Neobenedenia survival was examined and compared with freshwater and formalin immersion. Garlic extract was found to significantly impede hatching success (5% ± 5%) and oncomiracidia longevity (<2 h) at allicin concentrations of 15.2 µL L(-1) , while eggs in the seawater control had >95% hatching success and mean oncomiracidia longevity of 37 ± 3 h. At much lower allicin concentrations (0.76 and 1.52 µL L(-1)), garlic extract also significantly reduced Neobenedenia infection success of L. calcarifer to 25% ± 4% and 11% ± 4%, respectively, compared with 55% ± 7% in the seawater control. Juvenile Neobenedenia attached to host fish proved to be highly resistant to allicin with 96% surviving 1-h immersion in 10 mL L(-1) (15.2 µL L(-1) allicin) of garlic extract. Allicin-containing garlic extracts show potential for development as a therapy to manage monogenean infections in intensive aquaculture with the greatest impact at the egg and larval stages.