White Sturgeon as a Potential Vector of Infectious Hematopoietic Necrosis Virus

Abstract

Cell lines from white sturgeon Acipenser transmontanus were derived from peripheral blood cells, heart, and spleen. Incubated with infectious hematopoietic necrosis virus (IHNV) for 8 d at l5°C, these cell lines produced 0.7–53.2 plaque-forming units (PFU)/cell. Waterborne exposure of larval white sturgeons (60 d posthatch) to 10⁶ PFU/mL of IHNV resulted in 10% mortality 5–6 d postinfection, with virus concentrations consistently greater than 10⁵ PFU/g. A replicate group of larval white sturgeons that were sampled at different times post-IHNV exposure had no detectable virus at 24 h, but 72% of the fish had IHNV concentrations of 10²-10⁶ PFU/g when they were examined 2–9 d postinfection. Juvenile white sturgeons (mean weight, 35 g) immersed in or injected with IHNV exhibited no mortality, and virus was only detected immediately postexposure in just 25% of the fish tested. Juvenile white sturgeons fed either virus-free rainbow trout Oncorhynchus mykiss or dead IHNV-infected rainbow trout had no viable virus in their feces. Juvenile white sturgeons fed or exposed to IHNV failed to transmit the virus to cohabiting rainbow trout fry. These results suggest that IHNV can replicate in larval white sturgeons but presumably not in juveniles or adults. Virus neutralization activity was detected in serum from adult white sturgeons (4–6 years old) cultured with rainbow trout exposed to IHNV but not in white sturgeons kept in a pathogen-free environment and fed a manufactured diet. White sturgeon serum with IHNV-neutralizing activity was used to passively immunize rainbow trout, and it provided significant (P < 0.01) protection against IHNV challenge.     

Influence of environmental temperature on experimental infection of redfin perch (Perca fluviatilis) and rainbow trout (Oncorhynchus mykiss) with epizootic haematopoietic necrosis virus, an Australian iridovirus

SUMMARY Experimental transmission of epizootic haematopoietic necrosis virus (EHNV) to adult redfin perch Perca fluviatilis and juvenile rainbow trout Oncorhynchus mykiss was undertaken at different water temperatures using intraperitoneal (IP) and bath inoculation. Redfin perch were highly susceptible to EHNV by both routes of infection. Bath inoculation with as few as 0.08 TCID₅₀. _mL^-1 was lethal. The incubation period in redfin perch was about 11 days at a water temperature of 19–21°C but was longer at colder temperatures and disease did not occur at temperatures below 12°C. The longest incubation period recorded in redfin perch was 28 days. Rainbow trout were not susceptible to infection by bath inoculation but the disease was reproduced after IP inoculation with 10^5.6 TCID₅₀ at water temperatures ranging from 8–21°C. The incubation period was 3–10 days at 19–21°C, but was up to 32 days at 8–10°C. Persistent infection with EHNV was detected by virus isolation in a clinically unaffected rainbow trout after 63 days. The implications of these findings in the understanding of the epidemiology of EHNV infection are discussed.     

Comparative susceptibility of Atlantic salmon and rainbow trout to Yersinia ruckeri: Relationship to O antigen serotype and resistance to serum killing

A study was undertaken to compare the virulence and serum killing resistance properties of Atlantic salmon and rainbow trout Yersinia ruckeri isolates. Five isolates, covering heat-stable O-antigen O1, O2 and O5 serotypes, were tested for virulence towards fry and juveniles of both species by experimental bath challenge. The sensitivity of 15 diverse isolates to non-immune salmon and rainbow trout serum was also examined. All five isolates caused significant mortality in salmon fry. Serotype O1 isolate 06059 caused the highest mortality in salmon (74% and 70% in fry and juveniles, respectively). Isolate 06041, a typical ERM-causing serotype O1 UK rainbow trout strain, caused mortalities in both rainbow trout and salmon. None of the salmon isolates caused any mortalities in 150–250 g rainbow trout, and only serotype O2 isolate 06060 caused any significant mortality (10%) in rainbow trout fry. Disease progression and severity was affected by water temperature. Mortality in salmon caused by the isolates 06059 and 05094 was much higher at 16 °C (74% and 33%, respectively) than at 12 °C (30 and 4% respectively). Virulent rainbow trout isolates were generally resistant to sera from both species, whereas salmon isolates varied in their serum sensitivity. Convalescent serum from salmon and rainbow trout that had been infected by serotype O1 isolates mediated effective classical pathway complement killing of serotype O1 and O5 isolates that were resistant to normal sera. Overall, strains recovered from infected salmon possess a wider range of phenotypic properties (relative virulence, O serotype and possession of serum-resistance factors), compared to ERM-causing rainbow trout isolates.     

Susceptibility of Selected Inland Salmonids to Experimentally Induced Infections with Myxobolus cerebralis,the Causative Agent of Whirling Disease

Abstract

Laboratory exposures to the infectious stages (triactinomyxons) of Myxobolus cerebralis demonstrated a range of susceptibility to whirling disease among four species of inland salmonids. Replicate groups of each species were exposed to two concentrations of triactinomyxons, a low dose (100–200 per fish) and a high dose (1,000–2,000 per fish). Exposed fish were evaluated for clinical signs, for severity of microscopic lesions at 35 d, 2 and 5 months, and for spore concentrations in the head cartilage at 5 months. A standard strain of rainbow trout Oncorhynchus mykiss matched for age served as a susceptible species control. Rainbow trout, westslope cutthroat trout O. clarki lewisi, Yellowstone cutthroat trout O. clarki bouvieri, and bull trout Salvelinus confluentus were susceptible to M. cerebralis infections. Clinical signs, including radical swimming (“whirling”) and black tails, were observed at 7 weeks postexposure among rainbow and cutthroat trout challenged at 3 weeks of age. Clinical signs were rare among bull trout exposed at an age of 4 weeks and absent among rainbow and cutthroat trout exposed at 3 months posthatch. Most rainbow, cutthroat, and bull trout were found to be infected when examined at 5 months postexposure. The most severe microscopic lesions among infected fish at 5 months postexposure were found among rainbow trout. Cutthroat trout had less severe lesions, bull trout had mild infections, and no evidence of infection was found among Arctic grayling Thymallus arcticus. Mean spore concentrations among infected fish correlated with the severity of microscopic lesion scores. Rainbow trout had mean concentrations of spores in head cartilage reaching 10⁶, whereas more resistant species such as bull trout had 10⁴ spores; no spores were found among Arctic grayling at 5 months postexposure.     

Relation of Water Temperature to Bacterial Cold-Water Disease in Coho Salmon,Chinook Salmon,and Rainbow Trout

Abstract

The effect of water temperature on the progress of experimentally induced Cytophaga psychrophila infection was investigated in juveniles of coho salmon Oncorhynchus kisutch, chinook salmon O. tshawytscha, and rainbow trout O. mykiss (formerly Salmo gairdneri). A virulent strain of C. psychrophila was administered to fish by subcutaneous injection. Infected fish were held in tanks containing pathogen-free well water at temperatures ranging from 3 to 23°C. Mean times from infection to death of the fish were shortest at 12–15°C, which were the temperatures associated with the shortest time for doubling the population of this bacterium in vitro. Juvenile steelhead (anadromous rainbow trout) injected with viable C. psychrophila cells and held in 22°C water did not become diseased.     

Regulatory Effects of Water Temperature on Loma salmonae (Microspora) Development in Rainbow Trout

Abstract

Water temperature, a pivotal factor influencing interactions between teleosts and pathogens, was examined to determine its effects on the kinetics of xenoma formation and dissolution subsequent to experimental exposure of rainbow trout Oncorhynchus mykiss to the microsporidian gill pathogen Loma salmonae. The permissive water temperature range in which xenomas developed was between 9° and 20°C. Parasite development was arrested at temperatures outside this range, as indicated by the absence of visible xenomas among exposed fish. In addition, when these trout were subsequently moved to temperatures within the permissive range, xenomas failed to develop. Water temperature, within the permissive range, had no significant effect on either the number of xenomas that formed or the proportion of fish that developed xenomas following gastric intubation with a standard dose of spores. The relationship between water temperature and xenoma onset-time was best described (R ² = 88.3%) by polynomial regression analysis: onset = 320 ? 33.4T + 0.9547T ², where T is temperature (°C). Xenoma onset rate was also described through a modified degree-days model, yielding a predictive equation appropriate for use under conditions of fluctuating temperature. The thermal units, expressed as days × (°C above 7°C) necessary for xenoma onset were 298.6 on average. Xenoma dissolution rates, from the time of onset, also appeared to have a trend; more rapid dissolution occurred as temperatures increased. However, this trend correlated minimally with regression models.     

Identification and characterization of serum complement activity in the American crocodile (Crocodylus acutus)

Incubation of unsensitized sheep red blood cells with serum from the American crocodile (Crocodylus acutus) resulted in a concentration-dependent hemolysis. The hemolytic activity was heat-sensitive, and inhibited by EDTA in a concentration-dependent manner. The EDTA-inhibited SRBC hemolysis could be restored by the addition of excess Ca²⁺ or Mg²⁺, but not Ba²⁺ or Cu²⁺, revealing the specificity of this activity for these two divalent cations. The hemolytic activity of crocodile serum was titer-dependent, with 329 μL producing 50% of maximal SRBC hemolysis. The complement activity was also temperature-dependent, with decreased activity at lower temperatures (5–15 °C) and maximal activity occurred at 30–40 °C. The hemolysis occurred relatively slowly, with near zero activity after 10 min, 40% of activity observed within 15 min of exposure to SRBCs, and maximal activity at 30 min.     

Effect of temperature on the pharmacokinetics of benzocaine in rainbow trout (Oncorhynchus mykiss) after bath exposures

The pharmacokinetics of benzocaine during bath exposures at 1 mg/L were determined in rainbow trout acclimated at 6 °C, 12 °C or 18 °C for at least 1 month. Individual fish were exposed to benzocaine in a recirculating system for 4 h and pharmacokinetic parameters were estimated in a unique manner from the concentration of benzocaine in the bath water vs. time curve. Elimination from plasma was also determined after the 4 h exposure. The uptake clearance and metabolic clearance increased with increased acclimatization temperatures (uptake clearance 581 ± 179 mL/min/kg at 6 °C and 1154 ± 447 mL/min/kg at 18 °C; metabolic clearance 15.2 ± 4.1 mL/min/kg at 6 °C and 22.3 ± 4.2 mL/min/kg at 18 °C). The apparent volume of distribution had a trend for increasing with temperature that was not significant at the 5% level (2369 ± 678 mL/kg at 6 °C to 3260 ± 1182 mL/kg at 18 °C). The elimination half-life of benzocaine in plasma was variable and did not differ significantly with temperature (60.8 ± 30.3 min at 6 °C to 35.9 ± 13.0 min at 12 °C). Elimination of benzocaine from rainbow trout is relatively rapid and even more rapid at higher acclimatization temperatures based on calculated metabolic clearances and measured plasma concentrations, but was not evident by measurement of terminal plasma half-lifes.     

Treatment of Ichthyophthiriasis in Rainbow Trout and Common Carp with Common and Alternative Therapeutics

Abstract

The goal of this laboratory study was to provide better knowledge about the treatment of ichthyophthiriasis (causative agent: Ichthyophthirius multifiliis, a ciliate bacteria) in rainbow trout Oncorhynchus mykiss and common carp Cyprinus carpio. The following questions were investigated: (1) the effectiveness of different chemicals (formalin, sodium chloride, hydrogen peroxide, Perotan, Virkon, Aquahumin, Baycox, and Ivomec) and at different concentrations and durations of application, (2) the number of treatments and the time intervals between treatments that were necessary to remove the parasite, and (3) how treatment effectiveness differed between the two species. The most effective treatment was a 37% stock solution of formalin at 110 μL/L of bath water for 1 h in rainbow trout and for 2 h in common carp. Aquahumin (150 μL/L for 2 h) was effective in slightly or moderately infected rainbow trout and at low water temperatures, but it was not effective for common carp. All other tested chemicals were ineffective. With formalin and Aquahumin, five treatments were necessary to remove I. multifiliis infestation. At 10 ± 1°C, the parasites were eradicated when the treatment was performed at 48-h intervals. At 18 ± 1°C the infestation was eliminated when treatment was performed at 24-h intervals but not at 48-h intervals. At 25 ± 1°C, treatment at 24-h intervals was ineffective; however, shorter intervals between treatments might improve treatment efficacy at this temperature. In contrast, the number of treatment repetitions played a minor role, and parasites were eliminated with five treatments in all experiments when the type of chemical and treatment interval were optimal.     

Assessment of serum protein fractions in rainbow trout using automated electrophoresis and densitometry

Background: Although rainbow trout (Oncorhynchus mykiss, Walbaum) are one of the most‐studied fish, electrophoretic techniques and classification of serum protein fractions have not been standardized, such that clinically useful values are lacking. Objective: The aim of the present study was to evaluate preliminarily the serum protein fractions of rainbow trout using automated cellulose acetate electrophoresis and densitometry. Methods: Serum samples from 25 rainbow trout (Oncorhynchus mykiss, Walbaum) were electrophoresed on cellulose acetate plates and quantified using densitometry. Results: A maximum of 6 fractions were identified and numbered, in order of decreasing mobility, as I, II, III, IV, V, and VI. In 3 of 25 (12%) samples, 6 fractions were identified; in 18 (72%) samples, 5 fractions were identified; and in 4 (16%) samples, 4 fractions were identified. Fractions I, V, and VI were always clearly identifiable, whereas fractions II and IV were frequently fused and indistinguishable from fraction III. The pattern with 5 fractions was the most probable type (χ², P<.01). The mean (±SEM) protein concentrations of the 6 fractions were I, 0.8±0.1 g/dL; II, 0.3±0.0 g/dL; III, 1.6±0.1 g/dL; IV, 0.3±0.1 g/dL; V, 0.6±0.0 g/dL; and VI, 0.2±0.0 g/dL. Based on comparison of serum and plasma electrophoretic patterns from 8 fish, fibrinogen was found in fraction V. Conclusion: Automated cellulose acetate electrophoresis and densitometry appear to be a practical method for estimation of serum protein fractions in rainbow trout.     

Comparison of the susceptibility of brown trout (Salmo trutta) and four rainbow trout (Oncorhynchus mykiss) strains to the myxozoan Tetracapsuloides bryosalmonae, the causative agent of proliferative kidney disease (PKD)

Differences in susceptibility to the myxozoan parasite Tetracapsuloides bryosalmonae, the causative agent of proliferative kidney disease (PKD), between four strains of rainbow trout (Oncorhynchus mykiss) and brown trout (Salmo trutta) were evaluated. Fish were exposed to water enzootic for the parasite in the field for 5 days and were subsequently transferred to the laboratory. Relative parasite load was determined after 2, 3 and 4 weeks post-exposure (wpe) by quantitative real-time PCR (qPCR) of kidney samples and number of parasite stages was determined in immunohistochemical stained sections of kidney, liver and spleen tissues. According to qPCR results, the highest amount of parasite DNA per equal amount of host tissue at all time points was measured in brown trout. Two of the rainbow trout strains showed lower relative parasite load than all other groups at the beginning of the experiment, but the parasite multiplied faster in these strains resulting in an equal level of relative parasite load for all rainbow trout strains at 4 wpe. A weak negative correlation of fish size and parasite load was detected. Only in samples of a few fish, single stages of T. bryosalmonae were found in sections stained by immunohistochemistry impeding quantitative evaluation of parasite numbers by this method. The results indicate a differential resistance to T. bryosalmonae between the rainbow trout strains investigated and between rainbow trout and brown trout.     

Early Life Stage Survival and Susceptibility of Brook Trout,Coho Salmon,Rainbow Trout,and Their Reciprocal Hybrids to Infectious Hematopoietic Necrosis Virus

Abstract

Triploid (heat-shocked) and diploid groups of rainbow trout Oncorhynchus mykiss, brook trout Salvelinus fontinalis, coho salmon Oncorhynchus kisutch, and reciprocal hybrids were produced, monitored for early life stage survival, and evaluated for susceptibility to infectious hematopoietic necrosis virus (IHNV). The female rainbow trout × male brook trout triploid hybrids had significantly greater (P < 0.01) survival than the diploid hybrids of this cross. The heat-shocked hybrid group of the female rainbow trout × male coho salmon also exhibited significantly greater survival to the eyed egg stage of development than the untreated group of this hybrid. Studies of the susceptibility of treatment groups to a 1990 IHNV isolate from the Hagerman Valley were conducted by using a standardized immersion exposure procedure at one or two different mean body weights. The diploid brook trout and coho salmon and two triploid hybrids (female rainbow trout × male brook trout or male coho salmon) were significantly less (P < 0.05) susceptible to IHNV than the pure-species diploid and triploid rainbow trout groups.     

Haemolytic activity of sheep complement for two assay systems

Sheep complement (C) is haemolytic for sheep erythrocytes sensitized with rabbit antibody (sheep E-rabbit A) provided serum is used as soon as possible after collection. If left at 4 °C to separate from the clot, serum C activity for sheep E-rabbit A is markedly reduced. Heparinized plasma retains its haemolytic titre for at least 24 h at 4 °C. Plasma from Mg²⁺-ethyleneglycoltetraacetic acid (EGTA) blood is non-haemolytic, but addition of Ca²⁺ partially restores the titre. A high concentration of rabbit A is necessary to sensitize sheep E.Sheep C is haemolytic for human erythrocytes sensitized with sheep antibody (human E-sheep A) in the presence of Mg²⁺-EGTA. This C activity is stable at 4 °C for 24 h in serum, Mg²⁺-EGTA plasma and heparinized plasma. Haemolytic activity of serum heated at 50 °C for 30 min was restored by a factor B containing CM-cellulose fraction of foetal lamb serum in the presence of Mg²⁺-EGTA for human E-sheep A but not sheep E-rabbit A.These findings show that sheep C haemolysis of sheep E-rabbit A requires a Ca²⁺-Mg²⁺-dependent pathway that is labile in vitro for 24 h at 4 °C.     

Pharmacokinetics of sulphadimidine in carp (Cyprinus carpio L.) and rainbow trout (Salmo gairdneri Richardson) acclimated at two different temperature levels

Summary

The influence of temperature (10° C and 20° C) on pharmacokinetics and metabolism of sulphadimidine (SDM) in carp and trout was studied.

At 20° C a significantly lower level of distribution (Vd_area ) and a significantly shorter elimination half‐life (T _(½>) β) was achieved in both species compared to the 10° C level. In carp the body clearance parameter (Cl_B (SDM) was significantly higher at 20° C compared to the value at 10° C, whereas for trout this parameter was in the same order of magnitude for both temperatures.

N₄‐acetylsulphadimidine (N₄‐SDM) was the main metabolite of SDM in both species at the two temperature levels. The relative N₄‐SDM plasma percentage in carp was significantly higher at 20° C than at 10° C, whereas there was in trout no significant difference.

In neither species was the peak plasma concentration of N_4‐SDM (C_maxN4‐SDM)) significantly different at two temperatures.

The corresponding peak time of this metabolite (T_{max (N4‐SDM)}) was significantly shorter at 20° C compared to 10° C in both carp and trout.

In carp at both temperatures, acetylation occurs to a greater extent than hydroxylation. Only the 6‐hydroxymethyl‐metabolite (SCH₂OH) was detected in carp, at a significant different level at the two temperatures. Concentrations of hydroxy metabolites in trout were at the detection level of the HPLC‐method (0.02‐μg/ml). The glucuronide metabolite (SOH‐gluc.) was not detected in either species at the two temperatures.     

Early Kinetics of Infectious Hematopoietic Necrosis Virus (IHNV) Infection in Rainbow Trout

Abstract

A series of experiments was carried out with infectious hematopoietic necrosis virus (IHNV; 193-110 isolate) in rainbow trout Oncorhynchus mykiss (weight, ～1.2 g) to determine the duration of the patent period and the timing of onset of the infectious periods. We first attempted to transmit IHNV to recipient fish from infected rainbow trout 2–3 d after they had been exposed. No infection transfer occurred despite high titers (10^4.79 to 10^4.91 plaque-forming units 5–8 d postexposure (dpe). To determine the number of secondary cases produced by one infectious individual, we exposed approximately 50 rainbow trout (weight, ～1.5 g) in each of seven replicate tanks to a donor fish that had been infected with virus by bath exposure 3 d earlier. The prevalence of infection in recipient fish rose from 0.84% at 2 dpe to 7.9% at 6 dpe. Maximum incidence (22 cases) occurred between 2 and 4 dpe. No disease-specific mortalities occurred in recipient fish during the experiment. The titer of virus in both recipient and donor fish increased from 2 to 4 dpe. There was a positive correlation between the level of infection among donors and prevalence values among recipient fish (r ² = 0.60). The level of challenge by one infectious fish under the conditions provided was enough for infection transfer from sick cohabitant to susceptible fish but was not enough for initiation of a full-scale epizootic among recipients.     

Increasing stocking density causes inhibition of metabolic–antioxidant enzymes and elevates mRNA levels of heat shock protein 70 in rainbow trout

The objective of this study was to determine the effects of stocking density on the activity of glucose 6-phosphate dehydrogenase (G6PD), 6-phosphogluconate dehydrogenase (6PGD) and glutathione reductase (GR) enzymes in liver, muscle, gill and kidney tissues of rainbow trout. Fish were reared at different stocking densities (15 kg/m³, 20 kg/m³, 25 kg/m³ and 30 kg/m³). After adaptation period of 30 days, the experiment was carried out for two months. Stocking density of the control group was 15 kg/m³. Increasing stocking density caused inhibition in the activities of the enzymes except for kidney G6PD and 6PGD. Activity of both pentose phosphate pathway enzymes unexpectedly increased only in kidney whereas inhibition was observed in other tissues. Since the most powerful and gradual attenuation was observed in muscle tissue for all enzymes, we performed quantitative Real Time PCR to examine the expression of heat shock protein (Hsp70) gene in muscle in order to understand whether the decrease in enzyme activities is associated with stress. The mRNA expression data showed that Hsp70 expression levels significantly elevated at 25 kg/m³ and 30 kg/m³ stocking densities. Overall results indicate that increasing stocking density blocks the activity of metabolic and antioxidant enzymes and causes considerable stress in rainbow trout. The most susceptible tissue to increasing stocking density was observed to be the muscle.     

Differences between Plasma and Serum Samples for the Evaluation of Blood Chemistry Values in Rainbow Trout,Channel Catfish,Hybrid Tilapias,and Hybrid Striped Bass

Abstract

Blood chemistry analytes are determined for fish from either serum or plasma samples. Serum and plasma are similar in that they both represent the fluid component of blood; however, plasma contains clotting factors that are not present in serum. This study was conducted to determine whether the type of sample—plasma or serum—had an effect on measured blood analytes in rainbow trout Oncorhynchus mykiss, channel catfish Ictalurus punctatus, hybrid tilapia Oreochromis spp., and hybrid striped bass (striped bass Morone saxatilis × white bass M. chrysops). Paired plasma and serum samples were analyzed for the following standard biochemical analytes: total protein, albumin, globulin, creatinine, total bilirubin, alkaline phosphatase, aspartate aminotransferase, sodium, potassium, chloride, calcium magnesium, phosphorus, glucose, and cholesterol. For all four taxa, values for potassium were lower in the serum and magnesium and phosphorus values were higher in the serum. Glucose values were lower in the serum from rainbow trout, hybrid striped bass, and channel catfish; whereas cholesterol values were higher in the serum of rainbow trout, channel catfish, and hybrid tilapias. The differences observed between serum and plasma were distinct from changes occurring with hemolysis and, therefore, do not represent release of erythrocyte constituents. The differences most likely represent metabolic utilization of blood constituents while the blood was clotting. This work indicates that plasma should be used preferentially to serum for biochemical analysis because analyte levels determined from serum may not accurately reflect those found in circulating blood.     

Serological Evidence of Infectious Hematopoietic Necrosis in Rainbow Trout from a French Outbreak of Disease

Abstract

Following the detection of infectious hematopoietic necrosis virus (IHNV) in France in April 1987, a serological survey was conducted of the rainbow trout Oncorhynchus mykiss (formerly Salmo gairdneri) from an infected cultured stock previously known to be contaminated with viral hemorrhagic septicemia virus (VHSV) for 3 years. The work lasted from April to December 1987, at which time all the remaining fish were slaughtered. Serum samples were assayed by a plaque-reduction test and a simplified neutralization test that is more suitable for processing large numbers of serum samples. Such investigations revealed that IHNV neutralization by trout antibodies depended on trout complement, as did neutralization of VHSV. Incubation for 16 h at 4°C increased the sensitivity of the test compared to incubation for 1 h at 20°C. During the course of clinical IHN from April to June, young fish did not display any neutralizing activity, but in September, 29 of 50 of them exhibited significant anti-IHN neutralizing antibody titers ranging from 21 to over 160, and 18 of 46 of these same fingerlings did so in December. Similarly, fish that had undergone VHS infection in August began to develop anti-VHSV antibodies in December (5 of 50), demonstrating that one fish can harbor neutralizing antibodies to both IHNV and VHSV, and that these antibodies had required 14 weeks to appear under fish culture conditions at 10°C. As could be expected from seroneutralization tests, neutralizing antibodies to IHNV did not result in protection against VHS. Sera from 13 of 20 adult fish sampled in mid-June revealed neutralizing antibodies to IHNV, suggesting that they harbored the virus prior to the clinical infection that affected their progeny. Only two of the fish showed low anti-VHSV antibody titers. Similarly, neutralizing antibodies to IHNV were detected in 53 of 73 other adult fish sampled in late October, 10 months after they had spawned and 7 months after mortality had occurred among their progeny. Given the prevalence, level, and persistence of neutralizing antibody titers, the seroneutralization test would be worth investigating more thoroughly to define the conditions that could make it a reliable tool for checking the virus status of trout carriers.     

The anaesthetic potency of benzocaine-hydrochloride in three freshwater fish species

Anaesthesia was induced in the common carp, Cyprinus carpio, tiiapia, Oreochromis mossambicus and rainbow trout, Salmo gairdneri, at concentrations of 25; 50; 75 and 100 mg/1 of benzocaine-hydrochloride as well as neutralized benzocaine-hydrochloride at water temperatures of 15; 20 and 25 °C. The results obtained indicated intra-and interspecific differences in the susceptibility of fish to anaesthesia due to metabolic, chemoreceptive and temperature tolerance differences in and amongst the three species.