In vivo and in vitro treatments against Sparicotyle chrysophrii (Monogenea: Microcotylidae) parasitizing the gills of gilthead sea bream (Sparus aurata L.)

The effect of in vitro and in vivo treatments against Sparicotyle chrysophrii, a microcotylid parasite of gilthead sea bream (Sparus aurata L.), was studied. In vitro chemical treatments were targeted to eggs, oncomiracidia and adults, and were tested both as disinfectants and therapeutics for infected animals. The compounds were: distilled water, formalin, limoseptic ®, hydrogen peroxide, chlorine, and praziquantel (PZQ). Larvae were sensitive to all the treatments, but adults were more resistant, as chlorine (60 ppm - 1 h), hydrogen peroxide (100 ppm - 30 min) and PZQ (50 ppm - 30 min) produced only 10% mortality. All adults were killed only with distilled water, limoseptic (0.1% - 5 min), formalin (300 ppm - 30 min), or hydrogen peroxide (200 ppm - 30 min). Eggs were the most resistant stage, as only 30 min in limoseptic (0.1% in distilled water) or in formalin (300 ppm) prevented hatching. PZQ was used in vivo either as a curative or preventive treatment. The highest dose tested (400 mg kg^− 1 BW; effective dose 116.3 mg kg^− 1 BW due to palatability problems leading to 45% reduction in host food intake) did not significantly decrease prevalence of infection when given for 6 consecutive days. A lower dose (200 mg kg^− 1 BW) (effective dose 158.1 mg kg^− 1 BW) was rejected to a lower degree and decreased the prevalence of infection from 90% to 40%. When a lower dose (40 mg kg^− 1 BW) was administered for longer periods (20 days), food intake was reduced slightly, but the infection did not decrease significantly. The oral intubation with PZQ (200 mg kg^− 1 BW) once a week for 4 weeks did not prevent the infection of fish by cohabitation. However, a significant reduction in the abundance of the parasite was registered. In view of the results, recommendations for fish treatment and disinfection of aquaculture facilities are discussed.     

Absence of formaldehyde residues in penaeid shrimp exposed to formalin

Cultured blue shrimp (Penaeus stylirostris) were treated with normal therapeutic amounts of formalin (50 and 150 ppm) for 6 or 24 h. Analyses for extractable formaldehyde residues were run on the tails from these shrimp immediately after death and after storage at ambient (24°C) and refrigerated temperatures. No formaldehyde residues in these shrimp were found to result from the therapeutic use of formalin. However, shrimp tail muscle was found to produce small amounts (up to 4.8 ppm) of extractable formaldehyde during postmortem decomposition.     

Effects of formalin on total haemocytes count and histopathological changes in the shrimp Litopenaeus schmitti (Pérez‐Farfante & Kensley 1997)

Formalin is used in shrimp industry to eliminate ectoparasites and as a disinfectant. However, the effect of formalin on shrimp's defense mechanism is unknown. Several biomarkers were used to assess the immune response of juveniles of the shrimp Litopenaeus schmitti exposed to different concentrations of formalin (10, 25 and 50 mg L^?1) during 24 and 48 h. Formalin concentrations of 25 and 50 mg L^?1 produced a significant reduction in the total haemocyte count at both 24 and 48 h. Peroxidase and phenoloxidase activity did not show variations. The total protein concentration in haemolymph of shrimps was not significantly different between experimental groups. Histological analysis showed hyperplasia in the gills of animals exposed at 10 mg L^?1 for 24 h; however, the exposure at 25 and 50 mg L^?1 caused necrosis in gills, hepathopancreas and the antennal gland.     

Antifungal effect of Origanum onites essential oil as an alternative to formalin in the artificial incubation of narrow‐clawed crayfish (Astacus leptodactylus Eschscholtz, 1823)

As alternative to formalin, the antifungal effect of a plant product [Origanum onites L. (Lamiaceae) oil] was investigated for use in the artificial incubation of narrow‐clawed crayfish (Astacus leptodactylus Eschscholtz) eggs. For this purpose, this study was conducted as two experiments. In experiment I, the eggs were artificially incubated for 40 days. In experiment II, juveniles were cultured to determine effects of O. onites oil on juveniles for 30 days. The experimental groups were as follows: formalin (3500 ppm for 15 min), O. onites oil (300 ppm for 15 min, 700 ppm for 2 min and 1000 ppm as a dip treatment 15 split‐second) and a control (no treatment). In the experiment I, the highest hatching rate (86%) and survival rate of stage II juveniles (80%) were observed in 1000 ppm dip group. These results were similar to that of formalin group (85% and 79%) respectively. The control group exhibited the lowest hatching rate (49%) and stage II rate (42%) compared with the 1000 ppm dip group and 3500 ppm formalin treatments. However, other concentrations (300 and 700 ppm) of O. onites showed toxic effects on the eggs and there was no hatching. In the experiment II, the survival rate and growth performance of the crayfish juveniles were similar in all groups. This study indicated that the 1000 ppm O. onites dip treatment could be a good alternative to formalin for improved egg hatchability in the artificial incubation of crayfish eggs.     

Susceptibility of Caspian Roach (Rutilus rutilus caspicus) Fingerlings to Formalin

ABSTRACT

Caspian roach (Rutilus rutilus caspicus) were exposed to 0, 30, 40, 45, 50, 55, 60, 70, 100, 150 200, 250, 300, and 400 ppm formalin to determine LC₅₀. Respectively, 0.5 to 96 h LC₅₀ values were 252 to 49 ppm. The lowest observed effect concentrations (LOEC) and no observed effect concentrations (NOEC) varied between 40 to 150 and 30 to 100 ppm, respectively, across the tested time spectrum. Regression suggested that the concentrations of 88.4, 80.3, 51.2, and 40.1 ppm are maximum concentrations for 0.5, 1, 12, and 24 h bath treatment, respectively, which are not in the range suggested in the literature.     

Tolerance to Formalin by a Fluidized-Bed Biofilter and Rainbow Trout Oncorhynchus mykiss in a Recirculating Culture System

Formalin tolerances of a fluidized-bed sand biofilter and rainbow trout Oncorhynchus mykiss were tested in a semi-closed freshwater recirculating culture system. Progressively higher formalin levels were tested until fish mortality (±1.9%) occurred. A 1-h exposure to 167 ppm formalin, followed by flushing, was safe for trout (no mortality) at 15.0 C but not at 16.5 C. A 110-ppm indefinite treatment (no flushing) was safe at 17.3 C, but 120 ppm at 17.3 C and 100 ppm at 17.8 C were not. Biofilter nitrification was not impaired by 1-h formdin treatments up to 167 ppm at 16.5 C, and not, usually, by indefinite treatments up to 120 ppm at 17.3 C. However, a find indefinite treatment of 70 ppm was followed by abnormally high nitrite levels for 9 d. Formaldehyde remained detectable in the system for 11 h during indefinite treatment at 120 ppm formalin. Tests on the system's Co₂ stripper did not indicate that it removed formaldehyde.     

Lead accumulation and its effects on the branchial physiology of Prochilodus lineatus

The purpose of this work was to determine the tissue accumulation of lead (Pb) and its effects on osmoregulatory processes of the freshwater fish Prochilodus lineatus. Juvenile fish were exposed to Pb (from 1.7 to 0.7 mg of dissolved Pb L^?1) for 6, 24 and 96 h and Pb accumulation was analyzed in the gills, liver, kidneys, blood cells and muscle. The following parameters were also analyzed: hematologic (hemoglobin content, hematocrit and number of red blood cells), metabolic (blood glucose), endocrine (blood cortisol), osmo ionic (plasma osmolality and Na⁺, K⁺, Cl^? and Ca⁺² concentrations), gill enzymes (Na⁺/K⁺-ATPase and carbonic anhydrase), chloride cell (CC) density and CC location in the gills. Pb accumulated in all the analyzed tissues, with the kidneys showing the highest concentration, followed by the gills and liver. The lowest Pb concentrations were found in blood cells and muscle. Pb promoted an increase in blood glucose after 6 and 24 h exposure. Gill Na⁺/K⁺-ATPase was inhibited after 24 h of exposure, but its activity was restored at 96 h, probably due to the increase in CC in gill lamellae. Plasma Na⁺ was reduced after 6 and 96 h, while K⁺ concentrations increased at all the experimental times. Fish exposed to Pb showed reduced plasma Ca⁺² at all experimental periods. Hematologic parameters remained unchanged. Overall, this study demonstrated that Pb interferes in osmoregulatory processes of P. lineatus and the proliferation of CC in the gills is a response in order to reestablish adequate ion concentrations.     

Retention of formaldehyde in the tissues of two tropical fish species following exposure to therapeutic levels

Abstract. Formalin is an effective fish chemotherapeutant which is used to control certain ectoparasitic and bacterial infections. It is toxic to fish and its pathological and physiological effects have been reported. An experiment was conducted to elucidate the possibility of retention of formaldehyde in the muscles of two tropical food fish species, following exposure to therapeutic levels of formalin. The results indicated that Puntius gonionotus (Bleeker), retained 5·75 ppm formaldehyde following exposure to 50 ppm formalin for 24h. In contrast, Clarias batrachus (L.) retained 14·03 ppm and 18·72 ppm formaldehyde following exposure to formalin at 50 ppm and 100 ppm for 24 h respectively. However, no formaldehyde was detected after keeping the exposed fish in clean running water for 24h.     

Toxicity of Four Chemotherapeutic Agents to Rabbitfish Siganus rivulatus

As aquaculture evolves from extensive pond culture to intensive tank and cage systems, chemical dips and baths are increasingly being used to treat a concomitant increase in ectoparasitic and bacterial infestations. Some of the main disease‐causing agents are ectoparasites on the skin and gills of fish. Consequently, application of chemotherapeutics is increasing in aquaculture industries in order to control outbreaks of parasitic infestations. However, the toxic effect of commonly used chemotherapeutic agents on specific aquacultured fish species is often unknown. The present work was performed to test the effect of four commonly used chemical treatments on rabbitfish, Siganus rivulatus. The lethal concentrations for 50% of population (LC₅₀) of formalin, copper sulfate, potassium permanganate, and hydrogen peroxide for S. rivulatus juveniles treated for 1 h were assessed. Formalin and potassium permanganate tolerance values were determined by calculating 72‐h LC₅₀ values through probit analysis. The 72‐h LC₅₀ values for the formalin toxicity tests were 551.0 and 1.68 mg/L, respectively. LC₅₀ of copper sulfate and hydrogen peroxide could not be determined from the concentrations tested but were found to be >3 and >700 mg/L, respectively. Accordingly, treatment concentrations of formalin and potassium permanganate used for other fish species could be lethal to S. rivulatus, but the species appears to be quite tolerant to copper sulfate and hydrogen peroxide.     

Effects of formalin on haematological and blood chemistry in olive flounder, Paralichthys olivaceus (Temminck et Schlegel)

Erythrocytes of olive flounder, Paralichthys olivaceus (Temminck et Schlegel), were treated with serial concentrations of formalin (37% formaldehyde) to investigate in vitro haemolysis and methaemoglobin formation. In addition, the short‐term toxicity of formalin concentrations of 0, 100, 212 and 300 ppm was also studied by clinical tests in which fish were subjected to 3‐h bath exposure. There was no haemolysis of fish erythrocytes exposed to formalin concentrations ranging from 31.3 to 2000 ppm. Methaemoglobin formation, however, was induced at concentrations greater than 500 ppm. Red blood cell count, haemoglobin, haematocrit, mean corpuscular haemoglobin concentration and percentage of immature erythrocytes were also markedly elevated in all formalin‐exposed groups (P<0.05). Formalin exposure also caused significant increases in alkaline phosphatase, aspartate aminotransferase, lactate dehydrogenase, potassium, chloride, magnesium and inorganic phosphorus (P<0.05). However, total protein decreased significantly in the formalin‐exposed groups (P<0.05). No significant differences in white blood cell count, mean corpuscular volume, mean corpuscular haemoglobin, albumin, glucose, total cholesterol, high‐density lipoprotein cholesterol, free cholesterol, alanine aminotransferase, calcium, creatinine and total bilirubin were observed in the formalin‐exposed groups (P>0.05).     

Oral administration of bovine lactoferrin inhibits bacterial infection in tilapia and elevates survival after bacterial infection: an examination of its immune-modulating properties

It was previously reported that the oral administration of lactoferrin (LF) provides antimicrobial activity in animals against bacterial and viral infections and is involved in immunomodulatory properties. In this report, a hybrid strain of Oreochromis nilotica (male) × O. mossambicus (female) was fed homemade diets with or without supplementation with 10, 50, 100, and 150 mg bovine LF g^?1 feed for 60 days. After dietary treatment, the experimental fish were infected with Streptococcus agalactiae. LF supplementation resulted in a significantly higher survival rate and suppression of bacterial growth at 24–96 h in the liver, spleen, kidneys, eyes, and gills. A reduction in peroxidase activity was followed by a similar reduction in the peroxidase content of leukocytes at 24 h as analyzed by spectrophotometry. Respiratory burst (RB) activity was detected regardless of the time at which LF was administered to fish in relation to the bacterial infection. A beneficial effect of LF on RBs was also detected before bacterial challenge (with 100 mg LF/g) and 24 h after bacterial challenge with 150 mg/g. The results obtained validate LF’s beneficial effects on RBs by phagocytes in tilapia, which was only shown at 48 h after supplementation with 100 mg bovine LF g^?1 feed after the bacterial infection. Moreover, after oral administration of LF, it was detected in the mucosa of the small intestines of tilapia. In conclusion, we demonstrate a reduction in fish mortality after the oral administration of LF, and we examined its immunomodulatory properties in tilapia.     

Changes of Non-Specific Immune Parameters in Rainbow Trout,Oncorhynchus mykiss,After Exposure to Antimicrobial Agents Used in Aquaculture

Changes of plasma lysozyme and serum bactericidal activity, ceruloplasmin, total plasma protein, serum glucose, hematocrit and leucocrit levels in rainbow trout (Oncorhynchus mykiss) were studied after exposure to formalin and chloramine-T, which are used in aquaculture as anti-ectoparasitic and antibacterial agents. Fish were exposed to formalin at a concentration of 250 ppm for 60 min and exposed to chloramine-T for 5 ppm for 3 h. The parameters were analyzed for rainbow trout exposed to formalin and chloramine-T soon after treatments and rainbow trout exposed to freshwater after 24 h (return) following the treatments. Plasma lysozyme activity decreased after exposure to formalin; however, it was not affected by chloramine-T exposure. Serum bactericidal activity reduced after both formalin and chloramine-T exposure and did not change after returning to freshwater. Ceruloplasmin and total plasma protein were not affected by both treatments. Serum glucose levels of exposed fish elevated and remained at higher levels during recovery period when compared to control values. Hematocrit values of fish exposed to formalin increased; however, no significant change was observed in fish exposed to chloramine-T. Leucocrit levels of fish treated with both formalin and chloramine-T were higher than that of controls.     

Investigations on the effect of formalin and iodophor on embryo and larvae development in pikeperch,Sander lucioperca

ABSTRACT

An experiment was conducted to evaluate the effects of incubating pikeperch, Sander lucioperca, eggs in formalin and iodophor solutions for 15 min on embryo survival, the hatching rate, as well as on the rate of misshaped larvae, in order to develop methods for egg surface disinfection. Embryos in the morula stage, in the epiboly stage, and at the beginning of heart beat and blood circulation tolerated formalin concentrations up to 1,500 ppm for 15 min. However, they were very susceptible to iodophor treatment, as >0.1% iodophor solution (=13 ppm active iodine) significantly decreased the percentage of ready-to-hatch embryos and the percentage of hatched larvae. These data of this study recommend the use of formalin at a concentration of up to 1,500 ppm to disinfect pikeperch eggs.     

Instantaneous effect of dibromomethane on metamorphosis of larvae of the sea urchins Strongylocentrotus nudus and Strongylocentrotus intermedius

A volatile chemical, dibromomethane (DBM), produced from red coralline algae is known as a chemical inducer of larval metamorphosis of the sea urchin Strongylocentrotus nudus. We performed experiments exposing DBM to the larvae of S. nudus and Strongylocentrotus intermedius through a hydrophobic membrane. Metamorphic rates resulting from different diluted DBM solutions and exposure times were ascertained. The highest metamorphic rate, more than 80% in both species, was found after 1 h exposure to 1/2 diluted DBM. With this dilution, more than 80% of S. nudus and S. intermedius larvae metamorphosed 1 h after start of the experiment after only 10 and 5 min exposure, respectively, which corresponded to the low concentrations of 52–61 ppm and 34–43 ppm DBM by GCMS analysis, respectively. These findings suggest that DBM has an instantaneous effect on high success of metamorphosis of larvae of S. nudus and S. intermedius.     

Effects of formalin on water quality and parasitic monogeneans on silver perch (Bidyanus bidyanus Mitchell) in earthen ponds

Infestations of parasitic monogenean trematodes (Lepidotrema bidyana and Gyrodactylus sp.) on freshwater silver perch (Bidyanus bidyanus Mitchell) in earthen ponds were treated with formalin (37% formaldehyde). Concentrations of 30 and 40 mg L^?1 formalin were effective, but fish in ponds treated with 20 or 25 mg L^?1 remained infested. At temperatures of 24.1–26.9°C, concentrations of 30 or 40 mg L^?1 formalin caused dissolved oxygen (DO) to decline from 10.1–11.9 to 3.0–3.3 and 1.2–1.7 mg L^?1, respectively, within 36–42 h of treatment. In addition, pH declined from 7.2–8.4 to 6.3–6.7, within 36 h and turbidity decreased over 48 h. In the ponds where DO was 1.2–1.7 mg L^?1, silver perch showed signs of severe stress, but continuous aeration (10 hp ha^?1) for 3 days and inflow of well‐oxygenated water for 6–8 h prevented mortalities. At temperatures of 13.2–15.7°C, concentrations of 30 or 40 mg L^?1 formalin caused DO to decline from 9.0–10.0 to 6.0–8.1 mg L^?1 and pH from 7.0–7.3 to 5.9–6.6 within 72 h. Total ammonia‐nitrogen increased over 72 h in ponds treated with 30 or 40 mg L^?1 formalin. Fish became re‐infested with L. bidyana in all ponds within 30 days of treatment. A concentration of 30 mg L^?1 formalin is recommended as a treatment for monogeneans on silver perch in ponds, but aeration is necessary to maintain adequate water quality at higher temperatures.     

Studies on the physiology of somatolactin secretion in red drum and Atlantic croaker

The effects of exposure to water of different salinities and calcium concentrations, and to various backgrounds and illuminations on somatolactin (SL) levels in juvenile red drum (Sciaenops ocellatus) were investigated using a recently developed red drum SL radioimmunoassay. Plasma SL concentrations were also monitored in wild-caught Atlantic croaker (Micropogonias undulatus) during gonadal recrudescence. No correlations were obtained between plasma SL concentrations and different salinities, external calcium concentrations or reproductive condition in these sciaenid fishes. Plasma SL concentrations remained low (<1 ng ml^-1) in red drum 1h, 1 day and 1 week after exposure to full strength seawater (salinity 37%, calcium 1290 ppm), half strength seawater (salinity 18%, calcium 744 ppm), fresh water (salinity 1%, calcium 260 ppm) or soft water (salinity 0%, calcium 0 ppm). Circulating levels of SL did not change significantly in wild-caught croaker at the onset of vitellogenesis. However, by the end of ovarian recrudescence (late-yolk globule stage), plasma SL levels were significantly lower than those observed in females with immature (perinucleolar) oocytes. In contrast, plasma SL levels showed marked differences in red drum exposed to various backgrounds and illuminations. Plasma SL was lower in red drum kept in a light background (<1 ng ml^-1) than in those kept in a black background or in the dark (1.4-6.9 ng ml^-1). The highest plasma SL concentration (4–30 ng ml^-1) was obtained in red drum kept in a black background without illumination. These results suggest that SL is involved in the adaptation of the red drum to various backgrounds and illumination levels. SL may not have an important role during the reproductive cycle and adaptation to water of different salinities and calcium concentrations in sciaenid fishes.     

Gill histopathological characteristics of Caspian roach (Rutilus rutilus caspicus) fingerlings treated with potassium permanganate and formalin

The aim of this study was to determine the gill histopathological characteristics of Caspian roach (Rutilus rutilus caspicus) fingerlings exposed to different concentrations of potassium permanganate (PM) and formalin (FO). The fish were exposed to different PM and FO concentrations over different periods: 1 ppm PM for 24 h (1‐P24), 2 ppm PM for 24 h (2‐P24), 5 ppm PM for 0.5 h (5‐P0.5), 10 ppm PM for 0.5 h (10‐P0.5), 10 ppm FO for 24 h (10‐F24), 20 ppm FO for 24 h (20‐F24), 40 ppm FO for 0.5 h (40‐F0.5) and 80 ppm FO for 0.5 h (80‐F0.5). After exposure, gill samples were collected for histological examination. The most frequent gill histopathological symptoms were hyperplasia and epithelial lifting. The order of the gill injury index (I_g.i) among the treatments was as follow: 1‐P24, 2‐P24 < 5‐P0.5, 40‐F0.5 < 10‐F24 < 10‐P0.5, 20‐F24, 80‐F0.5. Comparing the two agents, it is concluded that there is no significant difference in the gill damages between the agents during the short‐bath treatments. However, PM causes less pathological symptoms than FO during the long‐bath treatments. Comparing the bath durations, the agents showed different behaviours. PM‐treated fish showed significantly lower gill damage during the long‐bath compared to the short‐bath treatment; whereas, pathological symptoms of FO‐treated fish was dose‐dependent for each exposure period, separately. Overall, exposure of Caspian roach to PM, at the concentrations recommended for long bath, resulted in milder gill damages compared to FO.     

Efficacy and toxicity of oxidative disinfectants for the removal of gill amoebae from the gills of amoebic gill disease affected Atlantic salmon (Salmo salar L.) in freshwater

Amoebic gill disease (AGD) of Atlantic salmon is treated commercially by bathing affected fish in freshwater. Recently, the efficacy of freshwater bathing has been questioned, and the aim of this study was to examine the potential for improving bathing efficacy using additives to the freshwater bath. AGD‐affected Atlantic salmon were bathed in 350 L tanks containing oxygenated freshwater to which chlorine dioxide (0–50 mg L^?1), chloramine‐T (0–50 mg L^?1) or hydrogen peroxide (0–100 μL L^?1) was added. Before and following a 3‐h exposure to the freshwater and chemical additive, the gills were removed from a sub‐sample of fish and the number of live amoebae on the gills were counted and smears made for confirmation of the presence of Neoparamoeba pemaquidensis, the causative agent of AGD. Following a further 3‐h exposure, a sub‐sample of fish was bled from the caudal vein and the gills were removed for histological examination. Chlorine dioxide and chloramine‐T at 25–50 and 10–50 mg L^?1, respectively, reduced the number of amoebae on the gills by approximately 50% compared with pre‐exposure numbers. The results from hydrogen peroxide treatment were equivocal and the toxicity of hydrogen peroxide was high. The toxicity of chlorine dioxide varied with freshwater hardness and/or suspended solid load, whereas chloramine‐T toxicity was low, with mortalities attributable only to elevated temperatures at the highest concentration tested. In conclusion, chlorine dioxide and chloramine‐T show promise as potential freshwater additives for the improved removal of N. pemaquidensis and possibly, other amoebae from the gills of commercially farmed Atlantic salmon.     

Efficacy of formalin in reducing the levels of peritrichous ciliates on cultured marine shrimp

Abstract. Tests were performed to measure the efficacy of formalin in reducing peritrichous ciliates on the surface of cultured marine shrimp. The tests were conducted at concentrations above and below those being used in shrimp culture and were designed to define the concentration providing the most effective rapid reduction of ciliates. Penaeus stylirostris Stimpson,'preconditioned' in a single large tank, were divided equally (after diagnosis of significant ciliate epifaunation) into ten 1000 litre fibreglass tanks; each received 235 shrimp of 10–15 g average weight. These tanks were divided into five treatments: 0, 12·5, 25, 50 and 100 ppm; two replicates per treatment. Treatments were conducted under static conditions for 4 h. Ciliate levels were measured on day 0 (pretreatment) and days 1, 2 and 7 post-treatment. Formalin at 25, 50 and 100 ppm was demonstrated to be effective in reducing ciliates, while 12·5 ppm did not differ from the control (0 ppm). In addition, the higher the level of formalin used (within the effective range), the more pronounced was the initial ciliate reduction and the longer was the protection from further ciliate epifaunation.     

In vitro and in vivo efficacy of drugs against the protozoan parasite Azumiobodo hoyamushi that causes soft tunic syndrome in the edible ascidian Halocynthia roretzi (Drasche)

It was discovered recently that infection by a protozoan parasite, Azumiobodo hoyamushi, is the most probable cause for soft tunic syndrome in an edible ascidian, Halocynthia roretzi (Drasche). In an attempt to develop measures to eradicate the causative parasite, various drugs were tested for efficacy in vitro and in vivo. Of the 20 antiprotozoal drugs having different action mechanisms, five were found potent (24‐h EC₅₀ < 10 mg L^?1) in their parasite‐killing effects: formalin, H₂O₂, bithionol, ClO₂ and bronopol. Moderately potent drugs (10 < 24‐h EC₅₀ < 100 mg L^?1) were quinine, fumagillin, amphotericin B, ketoconazole, povidone‐iodine, chloramine‐T and benzalkonium chloride. Seven compounds, metronidazole, albendazole, paromomycin, nalidixic acid, sulfamonomethoxine, KMnO₄, potassium monopersulphate and citric acid, exhibited EC₅₀ > 100 mg L^?1. When ascidians were artificially infected with A. hoyamushi, treated using 40 mg L^?1 formalin, bronopol, ClO₂, or H₂O₂ for 1 h and then monitored for 24 h, very low mortality was observed. However, the number of surviving parasite cells in the ascidian tunic tissues was significantly reduced by treating with 40 mg L^?1 formalin or ClO₂ for 1 h. The data suggest that we might be able to develop a disinfection measure using a treatment regimen involving commonly available drugs.