Investigation of co‐infections with pathogens associated with gill disease in Atlantic salmon during an amoebic gill disease outbreak

Gill diseases are a complex and multifactorial challenge for marine farmed Atlantic salmon. Co‐infections with putative pathogens are common on farms; however, there is a lack of knowledge in relation to the potential effect co‐infections may have on pathology. The objective of this study was to determine the prevalence and potential effects of Neoparamoeba perurans, Desmozoon lepeophtherii, Candidatus Branchiomonas cysticola, Tenacibaculum maritimum and salmon gill poxvirus (SGPV) during a longitudinal study on a marine Atlantic salmon farm. Real‐time PCR was used to determine the presence and sequential infection patterns of these pathogens on gill samples collected from stocking until harvest. A number of multilevel models were used to determine the effect of these putative pathogens on gill health (measured as gill histopathology score), while adjusting for the effect of water temperature and time since the last freshwater treatment. Results indicate that between 12 and 16 weeks post‐seawater transfer (wpst), colonization of the gills by all pathogens had commenced and by week 16 of marine production each of the pathogens had been detected. D. lepeophtherii and Candidatus B. cysticola were by far the most prevalent of the potential pathogens detected during this study. Detections of T. maritimum were found to be significantly correlated with temperature showing distinct seasonality. Salmon gill poxvirus was found to be highly sporadic and detected in the first sampling point, suggesting a carryover from the freshwater stage of production. Finally, the model results indicated no clear effect between any of the pathogens. Additionally, the models showed that the only variable which had a consistent effect on the histology score was N. perurans.     

Morphological changes in gill mitochondria-rich cells in cultured Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis> acclimated to a wide range of environmental salinity

Morphological changes in gill mitochondria-rich (MR) cells were examined in cultured Japanese eel acclimated to deionized freshwater (DFW), freshwater (FW), 30%-diluted seawater (DSW), and seawater (SW). The gill Na⁺/K⁺-ATPase activity was higher in SW-acclimated eel than in those acclimated to DFW, FW, and DSW. Immunocytochemical observations revealed that MR cells in the gill filaments were most developed in SW, whereas MR cells in the lamellae were preferentially observed in DFW, suggesting that filament and lamellar MR cells are responsible for ion secretion and absorption, respectively. In scanning electron microscopic observations, the apical membrane of lamellar MR cells appeared as a flat or slightly projecting disk with a mesh-like structure on its surface. In contrast, the apical membrane of filament MR cells showed a slightly concave surface. Whole-mount immunocytochemistry revealed that most MR cells showed cystic fibrosis transmembrane conductance regulator immunoreaction in their apical region in fish in DSW and SW, but not in those in DFW and FW, indicating that MR cells developed in DSW and SW function as an ion-secreting site. In addition to MR cells, distinct Na⁺/K⁺-ATPase immunoreaction was observed in the outermost layer of gill epithelia, suggesting that pavement cells are an additional site of ion uptake in the gills.     

Atypical bacterial gill disease: a new form of bacterial gill disease affecting intensively reared salmonids

An unusual form of bacterial gill disease (BGD) was identified which affected five species of cultured salmonids from Canada (i.e. rainbow trout, chinook salmon and Atlantic salmon), Norway (i.e. brown trout) and Chile (i.e. coho salmon). All outbreaks occurred at low water temperatures (< 10 °C) and with clinical presentations distinct from classical BGD, which is caused by Flavobacterium branchiophilum. In contrast to classical BGD, fish did not show marked respiratory distress with flaring of the opercula, the animals did not orientate at the surface of the water column near inflow water or at the margins of the tanks, and the feed response of the fish was varied. While mortality was increased, it was not precipitous as in classical BGD. Eight outbreaks were examined in greater detail using histopathology, scanning electron microscopy, bacteriology and immunohistochemistry. Large numbers of small bacterial rods were seen adhering to the lamellar epithelium of affected gills from all outbreaks. Histologically, the lamellar epithelium appeared swollen, often with evidence of single cell degeneration and exfoliation. In more severe instances, the formation of lamellar synechiae was seen, usually associated with sequestration of bacteria between fused lamellae. By contrast with typical BGD, overt epithelial hyperplasia, lamellar fusion and filamental clubbing were not common sequelae to infection; instead, the end result was shortened and somewhat stubby lamellae covered with swollen epithelial cells. The predominant bacterium recovered from affected gills was a small, Gram-negative, motile, fluorescent pigment-producing rod that shared phenotypic characteristics with Pseudomonas fluorescens. Polyclonal antisera prepared against three representative isolates indicated a weak antigenic similarity among them. Immunohistochemistry corroborated this finding, in that the antisera reacted strongly with gill sections containing the homologous bacteria, but not against morphologically similar bacteria in heterologous sections. A Gram-negative, yellow pigmented bacterium (YPB), identified as Flavobacterium psychrophilum, was also recovered, but only from the gills in the Ontario outbreaks. Antiserum prepared against this YPB indicated an antigenic similarity among isolates recovered from the Ontario outbreaks, but immunohistochemistry failed to recognize antigenically related bacteria on the gills of fish from the other outbreaks. Based on the unusual clinical presentation and the histopathological appearance of the gills, in conjunction with the absence of filamentous bacteria associated with and recovered from affected gills, the present authors have called this condition ‘atypical bacterial gill disease’ or ABGD.     

Salmon gill poxvirus,a recently characterized infectious agent of multifactorial gill disease in freshwater‐ and seawater‐reared Atlantic salmon

Gill diseases cause considerable losses in Norwegian salmon farming. In 2015, we characterized salmon gill poxvirus (SGPV) and associated gill disease. Using newly developed diagnostic tools, we show here that SGPV infection is more widely distributed than previously assumed. We present seven cases of complex gill disease in Atlantic salmon farmed in seawater and freshwater from different parts of Norway. Apoptosis, the hallmark of acute SGPV infection, was not easily observed in these cases, and qPCR analysis was critical for identification of the presence of SGPV. Several other agents including Costia‐like parasites, gill amoebas, Saprolegnia spp. and bacteria were observed. The studied populations experienced significant mortalities, which increased to extreme levels when severe SGPV infections coincided with smoltification. SGPV infection appears to affect the smoltification process directly by affecting the gills and chloride cells in particular. SGPV may be considered a primary pathogen as it was often found prior to identification of complex gill disease. It is hypothesized that SGPV‐induced gill damage may impair innate immunity and allow invasion of secondary invaders. The distinct possibility that SGPV has been widely overlooked as a primary pathogen calls for extended use of SGPV qPCR in Atlantic salmon gill health management.     

Acid water and aluminium exposure: gill lesions and aluminium accumulation in farmed brown trout, Salmo trutta L.

Abstract. Brown trout, Salmo trittta L., from two fish farms, situated within acid-susceptible areas in Sweden, were studied with respect to aluminium accumulation and gill pathology. The total aluminium concentrations, after liming of the water in both farms, were between 200 and 300μg/1 at sampling. Brown trout from a third farm in a non-acidified area (total aluminium concentration of water 35μg/1 were included as a control. The aluminium concentrations in gills from farmed brown trout in acidified areas were approximately 20μg/g and 90μg/g wet weight respectively compared with approximately 2μg/g wet weight in control brown trout gills. Light- and electron microscopic examination disclosed two major types of gill lesions characterized by chloride cell hyperplasia in the secondary lamellar epithelium and enlargement of the intercellular spaces in the secondary lamellar epithelium. The difference in the aluminium speciation and accumulation may explain the difference in the gill pathology of farmed fish from acidified areas. The chloride cell hyperplasia together with a relatively low aluminium accumulation is considered to reflect a long-term exposure effect.     

Efficacy of bithionol as an oral treatment for amoebic gill disease in Atlantic salmon Salmo salar (L.)

This study examined the efficacy of bithionol as an oral treatment for Atlantic salmon Salmo salar affected by amoebic gill disease (AGD). The current commercial management strategy of AGD is a costly 3 h freshwater bath. It is labour intensive and the number of baths needed appears to be increasing; hence, there is an effort to identify alternative treatments. Efficacy was examined by feeding AGD-affected Atlantic salmon twice daily to satiation with bithionol, an antiprotozoal, at 25 mg kg^− 1 feed. Three seawater (35‰, 17 °C) re-circulation systems were used each consisting of three tanks containing 32 Atlantic salmon smolts with an average (± SEM) mass of 90.4 g (± 5.2). Three feeds were examined in the trial including bithionol, plain commercial control and oil coated commercial control. Feeding commenced 2 weeks prior to exposure to Neoparamoeba spp. at 300 cells L^− 1 and continued for 28 days post-exposure. Efficacy was determined by examining gross gill score and identifying percent lesioned gill filaments twice weekly for 4 weeks post-exposure. Bithionol when fed as a two-week prophylactic treatment at 25 mg kg^− 1 feed delayed the onset of AGD pathology and reduced the percent lesioned gill filaments by 53% and halved the gill score from 2 to 1 when compared with both the plain and oil controls during an experimental challenge. There were no palatability problems observed with mean feed intake of bithionol over the trial duration with fish consuming higher levels of the bithionol diet compared to both the oil and plain controls. This study demonstrated that bithionol at 25 mg kg^− 1 feed, when fed as a two-week prophylactic treatment for Neoparamoeba spp. exposure, delayed and reduced the intensity of AGD pathology and warrants further investigation as an alternative to the current freshwater bath treatment for AGD-affected Atlantic salmon.     

Pathology of experimental amoebic gill disease in Atlantic salmon, Salmo salar L., and the effect of pre-maintenance of fish in sea water on the infection

Atlantic salmon were exposed to amoebic gill disease (AGD) immediately following their acclimatization to sea water (group 1), or following a 2 week period of maintenance in sea water (group 2). Three fish from each group were sampled on days 0, 1, 2, 4, 7, 14 and 28 post-infection. Characteristic gill lesions began to occur between days 2 and 4, and dramatically increased by day 7. The number of gill lesions on fish from group 2 was significantly higher than on fish from group 1 on days 7 and 14 ( P <0.001), but the two groups did not differ in any other parameter. Histologically, Paramoeba sp., the aetiological agent of AGD, could be seen on the gills of fish as soon as 1 day post-exposure, attached to healthy-appearing gills. Gill pathology in the form of hyperplasia and lamellar fusion followed shortly. AGD infection was accompanied by a significant increase in the number of gill mucous cells ( P =0.002). Different methods for the diagnosis of AGD are discussed.     

Detection of Neoparamoeba perurans by duplex quantitative Taqman real-time PCR in formalin-fixed, paraffin-embedded Atlantic salmonid gill tissues

The development and the application of a quantitative duplex real‐time PCR for the detection of Neoparamoeba perurans and the elongation factor α 1 gene (ELF) of Atlantic salmon, Salmo salar L., and rainbow trout, Oncorhynchus mykiss (Walbaum), are described. A set of primers and probe was designed to amplify a 139‐bp fragment specific to the N. perurans 18S rRNA gene. The test was shown to be very sensitive, being able to detect as little as 13.4 DNA copies per μL corresponding to 0.15 fg of template DNA. In addition, the reaction that detected N. perurans was found to have a high degree of repeatability and reproducibility, to have a linear dynamic range (R² = 0.999) extending over 5 log₁₀ dilutions and to have a high efficiency (104%). The assay was applied to DNA samples extracted from 48 formalin‐fixed, paraffin‐embedded (FFPE) salmon gill tissues showing varying degrees of gill histopathology and amoebic gill disease (AGD)‐type histopathology ranging from absent to severe (each scored 0–3). Neoparamoeba perurans DNA was detected in all the blocks where AGD‐type histopathology was diagnosed microscopically and in 43.6% of the blocks showing signs of gill pathology. The association between parasitic load and gill histopathology and AGD‐type histopathology severity was also investigated. This study also describes the development and the application of a second real‐time PCR for the generic detection of Neoparamoeba spp., Page, 1987. A set of primers and probe conserved among the Neoparamoeba spp. was designed to amplify a 150‐bp fragment within the 18S rRNA gene. Applied to N. perurans‐negative gill tissues, the method was used to exclude the presence of other Neoparamoeba spp. in those blocks where gill pathology was observed microscopically.     

Toxic effects of indoxacarb on gill and kidney histopathology and biochemical indicators in common carp (Cyprinus carpio)

The aim of this study was to determine the biochemical and histopathological effects of indoxacarb on an economic fish species Cyprinus carpio. The fish were divided into four triplicated groups exposed to either of 0% (control), 5% (0.75 mg/L), 10% (1.5 mg/L) or 20% (3 mg/L) of the pesticide LC₅₀. Blood sampling was performed after 7, 14 and 21 days exposure. There were no changes in blood calcium levels at any times. In the indoxacarb‐treated fish, chloride levels decreased at the first and increased at the second sampling compared to the control. In 3 mg/L treatment, phosphorus increased significantly after 7 days compared to the control. Plasma glucose levels decreased significantly at the 7th and 14th days sampling; however, it increased at the 21st day. After 7 days, creatinine level in 3 mg/L treatment was significantly lower than the control; however, the creatinine levels of indoxacarb treatments were higher than the control at the 21st day. The indoxacarb‐treated fish had higher plasma urea levels compared to the control at the 7th day. At the 21st day, plasma urea level at 3 mg/L was significantly lower than the control treatment. Edema, Lamellar curling, hyperplasia, lamellar fusion and hyperaemia were observed in the indoxacarb‐treated fish gill. Tubular necrosis, hematopoietic necrosis, melanomacrophage aggregates, Bowman's capsule edema, glomerulus degeneration and hyperaemia were observed in the indoxacarb‐treated fish kidney. Generally, sublethal concentrations of indoxacarb cause stress, hydromineral imbalance, metabolism alteration and gill and kidney damages in common carp.     

Pathology of bacterial gill disease: ultrastructure of branchial lesions

Abstract. The range of branchial lesions associated with bacterial gill disease (BGD) in rainbow trout, Oncorhynchus mykiss (Richardson), was investigated through the ultrastructural examination of 23 separate outbreaks of the disease condition within commercially reared stocks. Diseased branchial tissues had a large and diffusely distributed monomorphic population of filamentous bacteria which were strictly epicellular. Although bacterial colonization was restricted to the branchial cavity, it was neither site nor cell specific: epithelial and chloride cells of the lamellae, filaments and lining tissues of the branchial cavity were all uniformly affected. The bacteria possessed an extensive glycocalyx which appeared to facilitate adhesion to the apices of the microridged sub-unit modifications of the cell surface. Sites of colonization were accompanied by a diffuse pattern of cellular degeneration and necrosis that was generally restricted to the outer layers of epithelium. The polarity and nature of these changes suggest that the mechanism of interaction between the bacteria and host cells includes progressive hydropic change as a sequel to primary membrane damage and consequent increased cell permeability. These cellular changes were accompanied by the range of stereotypical responses of the gill to damage frequently reported for BGD including lamellar fusion, epithelial hyperplasia, and squamous and mucous cell metaplasia, in addition to lamellar spongiosis.     

Pathology of bacterial gill disease: sequential development of lesions during natural outbreaks of disease

Abstract. This study examines the sequential pattern of bacterial colonization and the sequential development of branchial lesions during typical outbreaks of bacterial gill disease (BGD) among four groups of commercially reared rainbow trout fingerlings, Oncorhynchus mykiss (Richardson). During a 5-month monitoring regime, prior to the onset of natural outbreaks of BGD, gill morphology of examined fish remained unaltered. Bacterial colonization of the gills was immediately preceded by several gill changes which were widely present in fish from all groups under study. These changes, which were detectable at the ultrastructural level only, included cytoplasmic blistering and degeneration of the microridges of the superficial filament epithelium, in addition to slight topographical irregularity of the filament tips suggestive of mild hyperplasia. Bacterial colonization began at these altered filament tips before spreading to more proximal regions of the filament and adjacent lamellar surfaces. Explosive increases in proportional morbidity and mortality coincided with the development of the following gill lesions: extensive bacterial colonization of lamellar surfaces, lamellar epithelial hydropic degeneration and necrosis, and lamellar oedema. In contrast, gill lesions such as lamellar fusion, epithelial hyperplasia and various metaplastic responses were detected later either as subacute (3–5 days) or chronic (7–14 days) changes. The role of epithelial necrosis as an important event during BGD is discussed as a possible mechanism leading to the development of sub-acute and chronic gill lesions.     

Effect of hydrogen peroxide as treatment for amoebic gill disease in Atlantic salmon (Salmo salar L.) in different temperatures

Amoebic gill disease (AGD) is a pathogenic disease in salmonids caused by Neoparamoeba perurans. Treatment of AGD infection has been through freshwater bathing of the fish. However, as the availability of fresh water is often limited, hydrogen peroxide has been introduced as an alternative treatment. This study investigated the effect of hydrogen peroxide as treatment for AGD‐infected salmon (Salmo salar L.,) at different seawater temperatures and hydrogen peroxide dosages. In total, 600 fish were challenged with N. perurans and the severity of the AGD infection was measured using a gill score scale. After challenge and disease development, the fish were distributed into 12 tanks. The treatment was performed at different seawater temperatures (8°C, 12°C, 17°C) using different hydrogen peroxide doses. Each temperature included an untreated control group. Linear models were used to analyse gill score. A significant effect of treatment was found (?0.68 ± 0.05) regardless of dose and temperature, suggesting that hydrogen peroxide was effective in treating AGD. When the model included dose, a negative linear relationship between dose and gill score was found. The study proved that treatment of AGD with hydrogen peroxide was successful, as gills partially recovered following treatment and further disease development was delayed.     

Non-lethal loop-mediated isothermal amplification assay as a point-of-care diagnostics tool for Neoparamoeba perurans,the causative agent of amoebic gill disease

Neoparamoeba perurans is the causative agent of amoebic gill disease (AGD). Two loop-mediated isothermal amplification (LAMP) assays targeting the parasite 18S rRNA and the Atlantic salmon EF1α, used as internal control, were designed. The N. perurans LAMP assay did not amplify close relatives N. pemaquidensis and N. branchiphila, or the host DNA. This assay detected 10⁶ copies of the parasite 18S rRNA gene under 13 min and 10³ copies under 35 min. Five “fast-and-dirty” DNA extraction methods were compared with a reference method and further validated by TaqMan™ qPCR. Of those, the QuickExtract buffer was selected for field tests. Seventy-one non-lethal gill swabs were analysed from AGD-clinically infected Atlantic salmon. The pathogen was detected under 23 min in fish of gill score >2 and under 39 min for lower gill scores. About 1.6% of the tests were invalid (no amplification of the internal control). 100% of positives were obtained from swabs taken from fish showing gill score ˃3, but only ~50% of positives for lower gill scores. The present LAMP assay could be implemented as a point-of-care test for the on-site identification of N. perurans; however, further work is required to improve its performance for lower scores.     

Environmental risk factors associated with amoebic gill disease in cultured salmon, Salmo salar L., smolts in Ireland

A 2-year study was carried out on amoebic gill disease (AGD) involving monthly samples of 1+ Atlantic salmon, Salmo salar L., smolts, histological assessment of the gills and analysis of environmental data. Gill pathology was seen before amoebae could be detected microscopically. These changes in gill integrity were associated with marine environmental conditions, particularly elevated ammonium, nitrite and chlorophyll levels. The results suggest that the environmental changes predispose salmon to colonization by amoebae and ciliates. High densities of histophagous scuticociliates were observed in the gills during periods of advanced gill pathology. A number of different amoebae were observed in close association with gill pathology. Neoparamoeba was not seen in high densities, nor was it associated with gill pathology, indicating that Neoparamoeba may not be the primary agent of the AGD in Irish salmonid culture.     

Gill pathology in Scottish farmed Atlantic salmon,Salmo salar L., associated with the microsporidian Desmozoon lepeophtherii Freeman et Sommerville, 2009

Gill disorders have emerged in recent years as a significant problem in the production of marine‐stage Atlantic salmon Salmo salar L. The multi‐aetiological condition ‘proliferative gill inflammation’ (PGI) has been reported to cause heavy losses in western Norway, yet reports of Scottish cases of the disease have remained anecdotal. In the present study, histopathological material from a marine production site in the Scottish Highlands experiencing mortalities due to a seasonal gill disease with proliferative‐type pathology was examined using light microscopy, special staining techniques and transmission electron microscopy (TEM). The microsporidian Desmozoon lepeophtherii Freeman et Sommerville, 2009 (syn. Paranucleospora theridion) was identified by staining using a Gram Twort method and TEM associated with distinctive proliferative and necrotic pathology confined to the interlamellar Malpighian cell areas of the primary filaments. Epitheliocystis was not a feature of the gill pathology observed. It is believed this is the first report of D. lepeophtherii being identified associated with pathology in a Scottish gill disease case, and supports anecdotal reports that a disease at least partly synonymous with PGI as described by Norwegian researchers is present in Scottish aquaculture.     

Evaluation of sodium percarbonate as a bath treatment for amoebic gill disease in Atlantic salmon

Amoebic gill disease (AGD), caused by Neoparamoeba perurans, is a major health challenge for Atlantic salmon aquaculture globally. While freshwater bathing for 2 hr is effective in reducing infection severity, there is need for more rapid and lower cost alternatives. To this end, a combination of sodium percarbonate (SPC) in freshwater was examined for its treatment efficacy. Initial in vitro studies showed a reduction in amoeba viability when exposed for 30 min to freshwater containing >500 mg/L SPC. Subsequently, AGD‐affected salmon were bathed for 30 min in 16°C freshwater containing 100, 500 or 1,000 mg/L SPC, or for 2 hr in 16°C freshwater to mimic industry practice. Treatment at the highest SPC concentration caused extensive gill damage and substantial mortality. Neither occurred to a significant extent at lower SPC concentrations. Gill pathology of surviving fish 10 days post‐treatment (dpt) was comparable to or more severe than pre‐treatment, and significantly (p < .001) more severe than in 2 hr freshwater bathed fish. N. perurans DNA was confirmed by qPCR in all treatment groups at 10 dpt. The data indicate that a 30‐min exposure to SPC in freshwater is not a suitable alternative to existing freshwater treatment of AGD.     

The induction of laboratory-based amoebic gill disease revisited

Previous work in our laboratory defined a method of inducing laboratory‐based amoebic gill disease (AGD) in Atlantic salmon, Salmo salar L. Gills of AGD‐affected fish were scraped and the debris placed into fish‐holding systems, eliciting AGD in naïve Atlantic salmon. While this method is consistently successful in inducing AGD, variability in the kinetics and severity of infections has been observed. It is believed that the infections are influenced by inherently variable viability of post‐harvest amoeba trophozoites. Here, a new method of experimental induction of AGD is presented that redefines the infection model including the minimum infective dose. Amoebae were partially purified from the gills of AGD‐affected Atlantic salmon. Trophozoites were characterized by light microscopy and immunocytochemistry and designated Neoparamoeba sp., possibly Neoparamoeba pemaquidensis. Cells were placed into experimental infection systems ranging in concentration from 0 to 500 cells L^?1. AGD was detected by gross and histological examination in fish held in all systems inoculated with amoebae. The number of gross and histological AGD lesions per gill was proportional to the inoculating concentration of amoebae indicating that the severity of disease is a function of amoeba density in the water column. The implications of these observations are discussed in the context of the existing AGD literature base as well as Atlantic salmon farming in south‐eastern Tasmania.     

Environment-induced gill disease of cultured rainbow trout (Salmo gairdneri)

Rainbow trout from eight North German freshwater culture facilities were investigated for chronic health damage resulting from captivity. The fishes were raised under a wide variety of conditions in net cages, circulation tanks, raceways, and earthen ponds.The most obvious non-contagious health problem was gill injury. Hyperplasia of the gill filaments, leading to lamellar fusion or the development of apical caps, as well as necrotic or advanced degenerative processes were observed. In addition, the gill filaments were frequently shortened, significantly reducing the functional surface of the organ.The lesions occurred in almost the same form in all of the culture units. It is likely that they are part of a syndrome induced by the environmental conditions typical of freshwater aquaculture facilities. These are characterized by an accumulation of nitrogenous excretion and decomposition products, extreme diurnal fluctuations in the concentration of dissolved gases, and intensive feeding involving the sudden addition of large amounts of artificial products. Under such conditions even small quantities of un-ionized ammonia can be toxic for epithelial tissues and disturb the normal elimination of protein metabolites across the gills. The pathological processes may be accelerated by secondary bacterial infections.     

Effects of monensin dose and treatment time on xenoma reduction in microsporidial gill disease in rainbow trout, Oncorhynchus mykiss (Walbaum)

The objectives of the study were (1) to determine an acceptable dose of dietary monensin against microsporidial gill disease (MGD) of salmon caused by Loma salmonae and (2) to determine the life stage of L. salmonae at which the action of monensin was most effective at reducing clinical disease. Rainbow trout (RBT) were fed monensin diets at concentrations of 0, 10, 100, 1000 and 5000 ppm starting 1 week before per os exposure to L. salmonae infected gill material. Lethal sampling occurred at weeks 6, 7, 8 and 9 post‐exposure (PE). The acceptable dose of dietary monensin was determined to be 1000 ppm because it was the lowest concentration that produced a significant and constant reduction in xenoma formation. RBT were fed monensin at 1000 ppm beginning at 0, 1, 2 and 3 weeks before exposure and at 1, 2 and 3 weeks after exposure and continuing until week 9 PE. Fish in which treatment was initiated at the time of exposure or 1 week before, showed the greatest reduction in xenoma production when compared with the untreated RBT.