Immunostimulants added to injected Aeromonas salmonicida bacterin enhance the defense mechanisms and protection in rainbow trout (Oncorhynchus mykiss).

Immunostimulants were given to rainbow trout for assaying effects on modulating non-specific defense mechanisms, specific immune response, and protection levels against pathogen challenge with Aeromonas salmonicida. Three drugs, levamisole (an approved veterinary drug in the USA), a quaternary ammonium compound (QAC), and a short-chain polypeptide (ISK) were found to affect the non-specific defense mechanism activities, which were measured by changes in circulatory neutrophil and phagocytic activity levels, and the specific immune response factors, which were measured by numbers of plaque-forming cells, and circulatory antibody levels. When given alone, the immunostimulants elevated the non-specific factors. When injected in combination with an A. salmonicida O-antigen bacterin, the non-specific factors were further elevated, and the specific response was raised over samples taken from fish given the bacterin without the immunostimulants. Challenge tests with the virulent pathogen, A. salmonicida, showed a 5-6 day delay in the onset of mortalities in the fish given the immunostimulants alone, and a 12-14 day delay when immunostimulants given were combined with the bacterin. In the groups given the QAC or ISK with the bacterin, there was a 20% and 40% survival rate, respectively.     

Effect of an immunopotentiator on Aeromonas salmonicida infection in rainbow trout (Salmo gairdneri)

The immunoactive peptide FK-565 (heptanoyl-y-D-glutamyl-(L)-mesodiaminopimelyl-(D)-alanine) was found to induce protection against intraperitoneal Aeromonas salmonicida infection in rainbow trout (Salmo gairdneri Richardson). The survival rate was as high as 60% when FK-565 was given intraperitoneally as a single dose (1mg/kg) one day before bacterial challenge. A non-specific stimulation of phagocytic cells by FK-565 at an early stage of the bacterial infection may contribute to the resistance observed. The phagocytic activity of peritoneal phagocytic cells as well as phagocytic cells of the pronephros were stimulated by FK-565 in vivo and in vitro, respectively, as compared to untreated control fish. Furthermore, decreased activity of phagocytic cells previously immunosuppressed with cyclophosphamide was rapidly restored by application of FK-565.     

Experimental Infection of Brook Trout with Infectious Hematopoietic Necrosis Virus Types 1 and 2

Abstract

Fry of brook trout Salvelinus fontinalis became infected and diseased after immersion exposure to infectious hematopoietic necrosis virus (IHNV), but a long-lasting IHNV carrier state was not induced. Duplicate groups of 100 fish were immersed for 6 h in baths containing a type 1 (Round Butte, RB) or a type 2 (Rangen, RA) IHNV isolate at a high or low dose. Brook trout mortalities induced by immersion in a bath of the RB or RA IHNV isolate at 10² plaque-forming units (pfu) per milliliter were equivalent (1 and 0%), but fish were more susceptible to infection with RA IHNV. Only the single dead fish in the RB group was infected, but 24% of the RAexposed fish were infected 1 week after exposure. At a dose of 10⁶ pfu/mL, exposure to RB IHNV resulted in a higher mortality (35%) and prevalence of infection (89% of live fish sampled at 1 week postexposure), but no infectious virus was detectable by 5 weeks after exposure. In contrast, RA IHNV exposure at a dose of 10⁴ pfu/mL resulted in only 5% mortality, and live fish killed at 1 week postexposure had a 22% prevalence of infection, but infectious virus was not detectable by week 3. Although brook trout have been previously considered to be resistant to IHNV, this study has shown that brook trout become diseased and die after exposure to a high dose of one type I IHNV isolate and can be infected after immersion exposure to even a low dose of type 1 or type 2 IHNV.     

Mobilization and activation of nonspecific cytotoxic cells (NCC) in the channel catfish (Ictalurus punctatus) infected with Ichthyophthirius multifiliis

Channel catfish demonstrate a shift in the tissue distribution of nonspecific cytotoxic cells (NCC) when infected with the protozoan parasite, Ichthyophthirius multifiliis. NCC, isolated from head kidney (HK) tissues (hemopoietic organ) or peripheral blood leukocytes, were assessed for cytotoxic activity against NC-37 (a transformed mammalian cell line). NCC activity from HK tissue of moribund I. multifiliis-infected fish was depressed compared to HK-NCC activity in uninfected or I. multifillis-immune fish. The activity of NCC, isolated from the peripheral blood of moribund I. multifiliis-infected fish was significantly greater than the NCC activity in peripheral blood from either immune or uninfected fish. Chromium-51 release assays were combined with effector and target conjugate assays to determine killing capacity (Vmax) and affinity (Km) for target cells of peripheral blood NCC from moribund I. multifiliis-infected and uninfected fish. These experiments indicated that the peripheral blood from the moribund infected fish contained an increased percentage of active NCC with increased killing capacity and target cell affinity compared to peripheral blood NCC activity of uninfected fish.     

Effect of a mixed herb-enriched diet on the innate immune response and disease resistance of Paralichthys olivaceus against Philasterides dicentrarchi infection

We investigated the effect of a mixed herb-enriched diet obtained from pomegranate Punica granatum, Dalmatian chrysanthemum Chrysanthemum cinerariaefolium, and mastic-leaved prickly-ash Zanthoxylum schinifolium on innate immune mechanisms (e.g., phagocytosis activity, respiratory burst activity, alternative complement activity, lysozyme activity, and disease resistance) of olive flounder Paralichthys olivaceus against the scuticociliate Philasterides dicentrarchi. All experimental groups were challenged with P. dicentrarchi (1 x 10(5) ciliates/mL) through intraperitoneal administration of the pathogen (50 microL) on day 1. On day 7, the infected groups were fed 0, 5, 50, and 100 mg/kg of the enriched diets. The innate immune parameters, cumulative mortality, and relative percent survival (RPS) were assessed at weeks 1, 2, 3, and 4. Administration of 50 or 100 mg/kg of the herbal-enriched diet enhanced immunity throughout the experimental period. However, at the 5-mg/kg dosage, the enriched diet did not enhance the innate immune estimates at any time. At doses of 50 and 100 mg/kg, administration of the diet preceding the challenge with P. dicentrarchi decreased the percentage cumulative mortality in the experimental groups and thereby increased RPS values. This study reports that administration of 50 or 100 mg/kg mixed herbal-enriched diet could positively influence the innate immune response to P. dicentrarchi and enhance the health status of olive flounder with respect to this microbe.     

Phagocytic activity of macrophages of rainbow trout against Vibrio anguillarum and the opsonising effect of antibody and complement

The phagocytosis of Vibrio anguillarum by peritoneal macrophages from normal rainbow trout was enhanced by antibody and complement. Treatment of either macrophages or the bacteria by antibody also enhanced opsonisation. Five weeks after immunisation with V anguillarum, the phagocytic activity of macrophages from rainbow trout was increased significantly compared with the activity of those from unvaccinated fish. Although agglutinin titres did not increase until three weeks after immunisation, seven out of 10 fish challenged one week after immunisation survived, indicating that immunised fish had developed resistance to vibrio infection before significant levels of antibody or phagocytic activity became detectable.     

Experimental induction of the carrier state in yearling brook trout: a model challenge protocol for IPNV immunization

Brook trout fry (Salvelinus fontinalis) were not protected from infectious pancreatic necrosis virus (IPNV) challenge by immersion vaccination with inactivated, purified virus at concentrations of 10(7) to 10(9) pfu/ml. Mortalities in vaccinated groups were higher than for the unvaccinated control group and appeared to be dose-dependent. A challenge protocol for adult brook trout was developed for future vaccine trials. A single intraperitoneal injection of virulent, purified virus was sufficient to make long-lasting carriers of 16 month-old trout. Fish underwent a transient viremia, identified by virus isolation from plasma and leucocytes. Feces were the most reliable samples for identification of IPNV carriers by non-sacrificial testing. Many fish in the remaining infected group were still carriers 12 and 27 weeks post-infection.     

Supernatants from leucocytes treated with melanin-concentrating hormone (MCH) and alpha-melanocyte stimulating hormone (alpha-MSH) have a stimulatory effect on rainbow trout (Oncorhynchus mykiss) phagocytes in vitro

Melanin-concentrating hormone (MCH) and alpha-melanocyte stimulating hormone (alpha-MSH) are widespread vertebrate neuropeptides. In teleost fish the peptides are involved in the hormonal control of skin pigmentation, but they have also been shown to modulate corticosteroid secretion in both fish and mammals. alpha-MSH has additional potent anti-inflammatory actions in mammals and both peptides stimulate leucocyte phagocytosis in rainbow trout in vitro. The effects of these peptides on phagocytosis and the release of immunomodulatory factors by rainbow trout head kidney leucocytes were investigated in vitro. Neither MCH nor alpha-MSH had any effect on the adherence of phagocytes to glass slides or the activity of isolated phagocytes. When added to mixed leucocyte suspensions, however, MCH (50 and 100nM) and alpha-MSH (1 and 10nM) significantly increased the percentage of cells undergoing phagocytosis and MCH (50nM), but not alpha-MSH, stimulated the phagocytic index. In subsequent experiments, isolated phagocytes were exposed to supernatants derived from mixed leucocyte suspensions exposed to MCH (50 and 100nM) and alpha-MSH (1 and 10nM). Supernatants from leucocytes exposed to all doses of the peptides significantly increased the percentage phagocytosis and those from cells stimulated with MCH (100nM) and alpha-MSH (1 and 10nM) increased the phagocytic index of the phagocytes. The results suggest that cells other than phagocytes are required for MCH and alpha-MSH to exert their stimulatory effects on trout phagocytic cells through the release of one or more macrophage-activating factors.     

Interaction between Aeromonas salmonicida and peritoneal macrophages of brook trout (Salvelinus fontinalis)

The phagocytic and bactericidal properties of peritoneal macrophages obtained from brook trout (Salvelinus fontinalis), injected with either glycogen or a modified Freund's complete adjuvant (MFCA), were evaluated against an avirulent and a virulent strain of Aeromonas salmonicida. Avirulent bacteria were effectively phagocytized and killed by macrophages obtained from fish injected with both irritants. With glycogen-elicited macrophages, no enhancement of killing was observed following opsonization of avirulent bacteria with specific antibodies. A killing index (K.I.) of 38 was obtained, compared to a K.I. of 39 for unopsonized bacteria. When avirulent bacteria were opsonized with complement, the K.I. was increased to 67. Virulent bacteria were less susceptible to the phagocytic and the bactericidal activities of glycogen-elicited macrophages, even after opsonization with antibodies and/or complement, K.I. of 9 to 15. In contrast, MFCA-elicited macrophages showed increased phagocytic and bactericidal activities against both strains. The K.I. of unopsonized virulent bacteria was increased to 47 and 46 compared to K.I. of 4 and 7 obtained with glycogen-elicited macrophages.     

壳寡糖对虹鳟生长性能、血清生化指标及非特异性免疫功能的影响

本文旨在研究壳寡糖对虹鳟生长性能、血清生化指标及非特异性免疫功能的影响。选用体重约为 ６０ｇ的虹鳟 ４８０尾,随机分为 ４组（每组 ４个重复,每个重复 ３０尾）,分别饲喂在基础饲料中添加 ０、１００、２００和 ４００ｍｇ／ｋｇ壳寡糖的试验饲料。试验期为 ４９ｄ。结果表明：１）与对照组相比,１００、２００和 ４００ｍｇ／ｋｇ壳寡糖组的特定生长率分别提高了 ５．６０％（Ｐ＜０．０５）、１５．８９％（Ｐ＜０．０５）和１８．６９％（Ｐ＜０．０５）,增重率分别提高了７．５３％（Ｐ＜０．０５）、２１．２１％（Ｐ＜０．０５）和 ２６．０２％（Ｐ＜０．０５）,蛋白质效率分别提高了 ０．６０％（Ｐ＞０．０５）、７．０２％（Ｐ＞０．０５）和９．７９％（Ｐ＜０．０５）,饲料系数分别降低了 ０．６０％（Ｐ＞０．０５）、６．５５％（Ｐ＞０．０５）和 ８．９３％（Ｐ＜０．０５）;２）饲料中添加壳寡糖提高了虹鳟血清总蛋白、白蛋白、球蛋白含量（Ｐ＞０．０５）,降低了血清甘油三酯、胆固醇、葡萄糖含量以及谷丙转氨酶和谷草转氨酶活性（Ｐ＞０．０５）;３）饲料中添加２００和 ４００ｍｇ／ｋｇ壳寡糖可显著或极显著提高虹鳟的白细胞吞噬百分率（Ｐ＜０．０５）、吞噬指数（Ｐ＜０．０５）,血清杀菌百分率（Ｐ＜０．０１）以及血清、肝脏和鳃中溶菌酶活性（Ｐ＜０．０５）;４）虹鳟全鱼水分、粗蛋白质、粗脂肪、灰分含量在各组间差异不显著（Ｐ＞０．０５）。由此得出,在本试验条件下,饲料中添加壳寡糖可提高虹鳟的生长性能和非特异性免疫功能,建议添加量为２００ｍｇ／ｋｇ。     

Influence of naturally acquired feline leukemia virus (FeLV) infection on the phagocytic and respiratory burst activity of neutrophils and monocytes of peripheral blood

The purpose of this study was cytometric evaluation of phagocytic and oxidative burst activity of neutrophils and monocytes in cats naturally infected with FeLV. To conduct the study, the peripheral blood was obtained from 33 cats naturally infected with FeLV. The control group consisted of 30 FeLV-, FIV-, clinically healthy cats. The percentage of phagocytizing neutrophils of peripheral blood was lower in FeLV+ than in FeLV- cats. The percentage of neutrophils and monocytes in which an oxidative burst occurred was lower in FeLV+ than in FeLV-animals. Also an oxidative product formation in neutrophils after E. coli and PMA stimulation was lower in FeLV+ than in FeLV-animals. Obtained results allow to conclude that diminished phagocytic and oxidative burst activity of peripheral blood leukocytes may cause impairment of innate immunity in cats infected with FeLV.     

Characterisation of the green turtle’s leukocyte subpopulations by flow cytometry and evaluation of their phagocytic activity

Phagocytosis is a fundamental aspect of innate immunity that is conserved across many species making it a potentially useful health-assessment tool for wildlife. In non-mammalian vertebrates, heterophils, monocytes, macrophages, melanomacrophages, and thrombocytes all have phagocytic properties. Recently, B lymphocytes from fish, amphibians, and aquatic turtles have also showed phagocytic capacity. Phagocytes can be studied by flow cytometry; however, the use of this tool is complicated in reptiles partly because nucleated erythrocytes complicate the procedure. We separated green turtle leukocytes by density gradient centrifugation and identified subpopulations by flow cytometry and confocal microscopy. Additionally, we assessed their ability to phagocytize Fluorspheres and Ovoalbumin-Alexa. We found that heterophils and lymphocytes but not monocytes could be easily identified by flow cytometry. While heterophils from adults and juvenile turtles were equally able to phagocytize fluorspheres, adults had significantly more phagocytic ability for OVA-Alexa. Lymphocytes had a mild phagocytic activity with fluorospheres (27–38 %; 39–45 %) and OVA-Alexa (35–46 %; 14–22 %) in juvenile and adult green turtles, respectively. Confocal microscopy confirmed phagocytosis of fluorospheres in both heterophils and lymphocytes. This provides the first evidence that green turtle lymphocytes have phagocytic activity and that this assay could potentially be useful to measure one aspect of innate immunity in this species.     

蛋氨酸对幼建鲤疾病抵抗能力及免疫应答的影响

本试验旨在研究蛋氨酸对幼建鲤疾病抵抗能力及免疫应答的影响.选择体重为(12.34±0.02)g健康建鲤864尾,平均分成6组,每组144尾(每组设3个重复,每个重复48尾),分别饲喂蛋氨酸水平为0.39%、0.70%、1.00%、1.30%、1.60%和1.90%的饲料,饲养60 d后用嗜水气单胞菌攻毒17 d,考察蛋氨酸对幼建鲤免疫功能的影响.结果表明:适宜蛋氨酸水平极显著提高了攻毒前头肾体指数、后肾体指数、脾体指数、血液红、白细胞数量和攻毒后红细胞数量(P<0.01);显著提高了攻毒前白细胞吞噬率和攻毒后成活率、白细胞数量、白细胞吞噬率、血清酸性磷酸酶活力、溶菌酶含量、补体C3含量、凝集素水平和抗嗜水气单胞菌抗体效价(P<0.05).由此得出,蛋氨酸通过增强白细胞吞噬能力和提高特异性抗体效价从而提高幼建鲤的非特异性和特异性免疫力,增强疾病抵抗力.     

Cytokine responses to EHV-1 infection in immune and non-immune ponies

Protecting equids against equine herpesvirus-1 (EHV-1) infection remains an elusive goal. Repeated infection with EHV-1 leads to protective immunity against clinical respiratory disease, and a study was conducted to measure the regulatory cytokine response (IFN-gamma and IL-4) in repeatedly infected immune ponies compared to non-immune ponies. Two groups of four ponies were established. Group 1 ponies had previously been infected on two occasions, and most recently 7 months before this study. Group 2 ponies had no history no vaccination or challenge infection prior to this study. Both groups were subjected to an intranasal challenge infection with EHV-1, and blood samples were collected pre-infection, and at 7 and 21 days post-infection for preparation of PBMCs. At each time point, the in vitro responses of PBMCs to stimulation with EHV-1 were measured, including IFN-gamma and IL-4 mRNA production, and lymphoproliferation. Group 1 ponies showed no signs of clinical disease or viral shedding after challenge infection. Group 2 ponies experienced a biphasic pyrexia, mucopurulent nasal discharge, and nasal shedding of virus after infection. Group 1 ponies had an immune response characterized both before and subsequent to challenge infection by an IFN-gamma response to EHV-1 in the absence of an IL-4 response, and demonstrated increased EHV-1-specific lymphoproliferation post-infection. Group 2 ponies had limited cytokine or lymphoproliferative responses to EHV-1 pre-challenge, and demonstrated increases in both IFN-gamma and IL-4 responses post-challenge, but without any lymphoproliferative response. Protective immunity to EHV-1 infection was therefore characterized by a polarized IFN-gamma dependent immunoregulatory cytokine response.     

Characterization of two key molecules of teleost innate immunity from rainbow trout (Oncorhynchus mykiss): MyD88 and SAA

Toll-like receptors (TLR) are relevant for piscine innate immunity. TLR activation recruits several downstream factors regulating the expression of immunorelevant genes. We have characterized two key factors of innate immunity from rainbow trout: MyD88 as an adaptor protein interacting directly with TLRs, and serum amyloid A as an effector molecule induced by the activated Toll-like receptor signaling cascade.Both factors share a remarkable high degree of structural conservation with their mammalian orthologs suggesting that innate immune defense mechanisms may also functionally be conserved between fish and mammals.     

Journal of Aquatic Animal Health: Guide for Authors 2014

Abstract

Brook trout Salvelinus fontinalis were treated with single 60-min static baths of 250 mg formalin/L, 3% NaCl, and 15 mg Chloramine-T/L to evaluate the efficacy of these compounds against external infections of Aeromonas salmonicida. Prevalence of A. salmonicida was significantly lower in brook trout treated with Chloramine-T than among those treated with formalin or salt. Further laboratory tests substantiated the therapeutic value of a single treatment of ChloramineT (15 mg/L) against A. salmonicida. In two experiments, viable counts of A. salmonicida in mucus did not vary among replicate groups of treated brook trout, but the counts for treated fish were significantly (P < 0.05) lower than those for untreated controls. In vitro tube dilution assays indicated that mean minimum inhibitory concentrations of Chloramine-T for 10 isolates of A. salmonicida were 9.0 mg/L for 1 h and 2.25 mg/L, for 24 h. In field trials at the White River National Fish Hatchery (Bethel, Vermont), the pathogen was detected principally as an external infection of juvenile Atlantic salmon Salmo solar maintained in two culture ponds. In one pond, the bacterium accounted for 100% of the total distribution of tnicroflora isolated from mucus. Seven days after treatment with Chloramine-T, A. sahnonicida accounted for 11% of the total bacterial counts identified from these fish. In the second pond, A. salmonicida composed 3% of the counts of bacteria isolated from the mucus of fish before treatment but was not isolated after treatment.     

Functions of neutrophils in sheep experimentally infected with Ehrlichia phagocytophila

Ehrlichia phagocytophila infection in sheep is characterized by persistent neutropaenia, indicative of decreased phagocytic capacity. This predisposes infected animals to other infections. A whole blood flow cytometrical method was used to document the degree and extent of reduced phagocytic and respiratory burst activity in phagocytes during an experimental infection with E. phagocytophila, and monitored until 56 days post-infection. Six sheep at 5 months of age were inoculated with an intravenous injection of infected blood. Six age-matched sheep were used as controls. A period of reduced respiratory burst lasting up to Day 17 post-infection was recorded. The population of cells showing phagocytic activity without respiratory burst was larger in the infected animals compared to controls up to Day 45 post-infection.     

Enhancement of phagocytic and chemokinetic activities of rainbow trout head kidney cells by C-reactive protein

OBJECTIVE: To investigate immunostimulating activity of purified rainbow trout (Oncorhynchus mykiss) C-reactive protein (CRP) on trout phagocytic cells. ANIMALS: 20 rainbow trout and 2 rabbits. PROCEDURE: The effect of CRP on phagocytic activity of head kidney (HK) cells was examined by use of a phagocytosis assay with plastic particles. The enhancing effect of CRP on migration activity of HK cells was examined by use of the blind well assay. RESULTS: Glass-adherent cells from clinically normal trout had increased dose-dependent phagocytic activity against plastic particles when cells were incubated in the presence of CRP. Pretreatment of particles with CRP also enhanced phagocytic activity of the cells, indicating an opsonic effect of CRP. Rabbit anti-trout CRP serum suppressed the enhancing activity of CRP. The HK cells had significant dose-dependent chemokinetic activity against CRP that was not inhibited by anti-CRP serum, indicating that a CRP-antibody complex also could be chemokinetic. CONCLUSIONS: Rainbow trout CRP has immunostimulating activity for HK cells, resulting in enhanced phagocytic and chemokinetic activities.     

The effects of five different glycans on innate immune responses by phagocytes of hybrid tilapia and Japanese eels Anguilla japonica

The aim of this study was to evaluate the immune responses in hybrid tilapia (Nile tilapia Oreochromis niloticus x Mozambique tilapia O. mossambicus) and Japanese eels Anguilla japonica after treatment with five glycans: barley, krestin, MacroGard, scleroglucan, and zymosan. The effects of the glycans on the innate immune responses of the fish were investigated using the phagocytic index (PI), lysozyme activity, complement opsonization, and activation assay. The results of the lysozyme assay demonstrated that the lysozyme activities increased after treatment with glycans. Moreover, based on the PI, treatment with each of the five glycans resulted in increased phagocytic activities in anterior kidney and peripheral blood phagocytes in both tilapia and Japanese eels. The opsonic effect of complement on phagocytosis in tilapia and Japanese eels were investigated using baker's yeast, which served as the activator in the classical complement pathway (CCP) and in the alternative complement pathway (ACP). Tilapia and Japanese eel sera that were treated with glycans greatly enhanced phagocytosis. The classical pathway--hemolytic complement titer (CH50) of Japanese eels treated with glycans was slightly increased in vitro and in vivo. While glycan treatment enhanced the CCP of both species in vitro and in vivo, the alternative pathway-hemolytic complement titer (ACH50) was only increased in vitro and in vivo in glycan-treated tilapia. Thus, it follows that the ACP must have been activated in tilapia treated with glycans. However, in Japanese eels, the ACH50 of the ACP activation assay was undetected in vitro or in vivo due to possible unknown factors in the Japanese eel serum that caused lysis of the rabbit red blood cells. Our study investigated the effects of glycans used to enhance phagocytosis and activate both of the complement pathways involved in stimulating the innate immune responses of Japanese eels and tilapia.     

Antiparasitic and immunomodulatory effect of innovative treatments against Myxobolus sp. infection in Diplodus puntazzo

The potential antiparasitic and immunomodulatory effect of three treatments against myxosporean parasites on the innate immune system of sharpsnout sea bream (Diplodus puntazzo) was investigated. Fish naturally infected with Myxobolus sp. (Bivalvulida/Platysporina), a histozoic parasite mainly affecting the renal interstitial tissue, were treated by oral administration of a combination of salinomycin with amprolium, Origanum essential oil or fumagillin in a small-scale field trial. Various leucocyte functions influenced by myxosporean infection were examined in order to determine treatment effects on leucocyte immunocompetence of treated fish. One month post treatment all drugs caused a significant decrease in prevalence and intensity of infection in comparison to untreated, infected fish. The effect was most prominent in salinomycin with amprolium treated fish, which 1-month post treatment contained either no cysts at all or a few spores free in melanomacrophage centres revealing almost total elimination of the parasite and the antiparasitic action of the treatment. There was no histopathological evidence of drug toxicity. Antiparasitic action was accompanied by a significant enhancement of phagocytic activity demonstrated by ingestion of large numbers of latex beads and the secretion of high levels of reactive nitrogen intermediates by phagocytes in vitro. Complete restoration of the diminished mitogenic responses and serum lysozyme secretion was also detected in salinomycin with amprolium-treated fish compared to untreated, infected fish. These data suggest that salilomycin with amprolium may be a promising treatment for myxosporean infections in intensively cultured warm-water fish, exhibiting action partially via the enhancement of host, innate immune functions and leading to parasite elimination.