Metabolism,pharmacokinetics and selected pharmacodynamic effects of codeine following a single oral administration to horses

ObjectiveTo describe the pharmacokinetics and selected pharmacodynamic variables of codeine and its metabolites in Thoroughbred horses following a single oral administration.Study designProspective experimental study.AnimalsA total of 12 Thoroughbred horses, nine geldings and three mares, aged 4–8 years.MethodsHorses were administered codeine (0.6 mg kg^–1) orally and blood was collected before administration and at various times until 120 hours post administration. Plasma and urine samples were collected and analyzed for codeine and its metabolites by liquid chromatography–mass spectrometry, and plasma pharmacokinetics were determined. Heart rate and rhythm, step counts, packed cell volume and total plasma protein were measured before and 4 hours after administration.ResultsCodeine was rapidly converted to the metabolites norcodeine, codeine-6-glucuronide (C6G), morphine, morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G). Plasma codeine concentrations were best represented using a two-compartment model. The C_max, t_max and elimination t_½ were 270.7 ± 136.0 ng mL^–1, 0.438 ± 0.156 hours and 2.00 ± 0.534 hours, respectively. M3G was the main metabolite detected (C_max 492.7 ± 35.5 ng mL^–1), followed by C6G (C_max 96.1 ± 33.8 ng mL^–1) and M6G (C_max 22.3 ± 4.96 ng mL^–1). Morphine and norcodeine were the least abundant metabolites with C_max of 3.17 ± 0.95 and 1.42 ± 0.79 ng mL^–1, respectively. No significant adverse or excitatory effects were observed.Conclusions and clinical relevanceFollowing oral administration, codeine is rapidly metabolized to morphine, M3G, M6G, C6G and norcodeine in horses. Plasma concentrations of M6G, a presumed active metabolite of morphine, were comparable to concentrations reported previously following administration of an analgesic dose of morphine to horses. Codeine was well tolerated based on pharmacodynamic variables and behavioral observations.     

The disposition and local effects of intra-articular morphine in normal ponies

Morphine (15 mg in 5 ml saline) was injected into the left, and 5 ml saline into the right, tarsocrural joint of 8 ponies. Venous blood samples were collected before and at 0.5, 1, 2, 6 and 24 h after the intra-articular morphine injection and analysed for morphine and its metabolites. Synovial fluid was sampled from both tarsocrural joints before and 24 h after injection. Synovial white blood cell and red blood cell counts, protein and hyaluronate concentrations were measured in all the samples; and the synovial fluid morphine concentration from the left tarsocrural joint was measured 24 h after the injection. The peak mean plasma morphine concentration (7.1 μg/l) was detected in samples taken 0.5 h after the intra-articular morphine injection, but neither morphine nor its metabolites were found in plasma 6 h or more post injection. Morphine was detected in the synovial fluid of each pony 24 h after the injection. The plasma morphine or morphine-6-glucuronide concentrations were lower than those likely to have any systemic effect. The synovial fluid white blood cell count and protein concentration were increased and hyaluronate concentration decreased in samples taken 24 h after the intra-articular morphine injection, compared to the pre-injection samples. No differences were found between morphine and saline injected joints. It was concluded that morphine did not irritate the joint more than saline.     

The dose–response relation for the antinociceptive effect of morphine in a fish,rainbow trout

Jones, S. G., Kamunde, C., Lemke, K., Stevens, E. D. The dose–response relation for the antinociceptive effect of morphine in a fish, rainbow trout. J. vet. Pharmacol. Therap.  35 , 563–570. There have been suggestions that analgesics be used by fish researchers. But in the absence of dose–response data for morphine, this suggestion seems imprudent. The purpose of the present study was to develop a dose–response relationship in fish using six doses of morphine. The response (movement of the fins or tail) to a noxious stimulus (electrical shock to the face region) was monitored before and after a dose of morphine intraperitoneally (i.p.). The i.p. dose of morphine ED₅₀ in rainbow trout was 6.7 ± 0.8 mg/kg (n = 12 at each dose). The plasma morphine concentration EC₅₀ was 4.1 ± 1.5 mg/L. In a second experiment, rainbow trout tested with equal amounts of morphine and naloxone (30 mg/kg) showed that the antinociceptive effect of morphine was blocked by naloxone. It has been suggested that stress‐induced analgesia has been a confounding factor in some fish studies. However, plasma cortisol levels in our study indicated that stress was not a confounding factor in the present experiments. The ED₅₀ for morphine in fish was higher than that reported for humans or other mammals. Our observation showing a dose–response relation for morphine using a noxious stimulus supports arguments for its effectiveness as an antinociceptive drug in fish.     

Pharmacokinetics of intra‐articular morphine in horses with lipopolysaccharide‐induced synovitis

ObjectiveTo describe the pharmacokinetics of intra-articularly (IA) administered morphine.Study designExperimental randomized, cross-over study.AnimalsEight adult healthy mixed breed horses aged 6.5 ± 2.3 (mean ± SD) years and weighing 535 ± 86 kg.MethodsUnilateral radiocarpal synovitis was induced by IA injection of 3 μg lipopolysaccharide (LPS) on two occasions (right and left radiocarpal joint, respectively) separated by a 3-week wash-out period. Treatments were administered 4 hours post-LPS-injection: Treatment IA; preservative free morphine IA (0.05 mg kg^?1) plus saline intravenous (IV) and treatment IV; saline IA plus preservative free morphine IV (0.05 mg kg^?1). Concentrations of morphine, morphine-3-glucuronide and morphine-6-glucuronide (M6G) were determined repeatedly in serum and synovial fluid (SF) by high-performance liquid chromatography mass spectrometry, at 2 and 4 hours and then at 4 hours intervals until 28 hours post-treatment.ResultsInjection of LPS elicited a marked and comparable synovitis in all LPS-injected radiocarpal joints. IA administered morphine was detectable in SF of all eight joints 24 hours post-treatment and in 6/8 joints 28 hours post-treatment. The terminal half-life of morphine in SF was estimated to be 2.6 hours. IA administration of morphine resulted in mean serum concentrations of morphine below 5 ng mL^?1 from 2 to 28 hours after treatment.Conclusions and clinical relevanceIntra-articularly administered morphine remained within the joint for at least 24 hours. At the same time only very low serum concentrations of morphine and M6G were detected. The present results suggest that IA morphine at 0.05 mg kg^?1 may be used for IA analgesia lasting at least 24 hours and give strong support to the theory that previously observed analgesic and anti-inflammatory effects of IA morphine in horses are most likely to be mediated peripherally.     

Morphine reduces spinal c-fos expression dose-dependently during experimental laparotomy in pigs: a combined pharmacokinetic and surgical study

The pharmacokinetics of intravenous morphine 2.5mg/kg (n=4) and 10mg/kg (n=4) in plasma and cerebrospinal fluid (CSF) of pigs was studied. Plasma half-life was 1.0+/-0.1h and the main metabolite was morphine-3-glucuronide, whereas morphine-6-glucuronide was negligible. CSF morphine concentration peaked after 20-30min (2.5mg/kg) and 60-120min (10mg/kg), and elimination half-life was 3.5+/-0.3h. Subsequently, the effect of morphine on surgery-induced spinal nociception in pigs subjected to unilateral laparotomy was evaluated by stereological quantification of the total number of Fos-like-immunoreactive (Fos-LI) spinal neurons of the dorsal horn. Surgery (n=4) induced 91,680+/-14,974 Fos-LI neurons ipsilaterally and morphine reduced this number to 45,771+/-8755 following the 2.5mg/kg dose (p<0.01; n=6) and 14,981+/-2327 following the 10mg/kg dose (p<0.001; n=6). These results indicate that morphine dose-dependently reduces the number of surgery-induced Fos-LI neurons in the spinal cord. As even a high dose of morphine does not reduce spinal c-fos expression to basal level, it may be appropriate to use other analgesics simultaneously with morphine during surgery.     

Characterization of the pharmacokinetics,behavioral effects and effects on thermal nociception of morphine 6-glucuronide and morphine 3-glucuronide in horses

ObjectiveTo describe the pharmacokinetics, behavioral and physiologic effects and effects on thermal thresholds of morphine, morphine 6-glucuronide (M6G) and morphine 3-glucuronide (M3G) following administration to horses.Study designRandomized balanced crossover study.AnimalsA total of seven University-owned horses, five mares and two geldings, aged 3–6 years.MethodsHorses were treated with a single intravenous dosage of saline, morphine (0.2 mg kg^–1), M6G (0.01 mg kg^–1) and M3G (0.03 mg kg^–1). Blood was collected prior to (baseline) and at several times post administration. Drug and metabolite concentrations were determined by liquid chromatography–mass spectrometry, and plasma pharmacokinetics were calculated. Behavioral observations and physiologic variables (heart rate, step counts, packed cell volume, total plasma protein and gastrointestinal sounds) were determined at baseline and for up to 6 hours. The effects on thermal nociception were determined and thermal excursion was calculated.ResultsThe volumes of distribution were 4.75–10.5, 0.244–0.295 and 0.215–0.356 L kg^–1 for morphine, M6G and M3G, respectively. Systemic clearances were 26.8–39.6, 3.16–3.88 and 1.46–2.13 mL minute^?1 kg^?1 for morphine, M6G and M3G, respectively. Morphine administration resulted in signs of excitation as evidenced by an increase in step counts and subjective behavioral observations, whereas M6G and M3G, based on the same criteria, appeared to cause sedative-like effects. Significant effects on thermal nociception were observed until 4 hours post morphine administration, 1 hour post M6G administration and at various times post M3G administration.Conclusions and clinical relevanceResults of this study provide additional information regarding the use of morphine in horses. Less locomotor excitation and gastrointestinal adverse effects, compared with morphine, coupled with favorable effects on thermal nociception are encouraging for further study of the pharmacodynamics of both M6G and M3G in horses.     

Pharmacokinetics and selected pharmacodynamics of morphine and its active metabolites in horses after intravenous administration of four doses

The objective of the current study was to describe and characterize the pharmacokinetics and selected pharmacodynamic effects of morphine and its two major metabolites in horses following several doses of morphine. A total of ten horses were administered a single intravenous dose of morphine: 0.05, 0.1, 0.2, or 0.5 mg/kg, or saline control. Blood samples were collected up to 72 hr, analyzed for morphine, and metabolites by LC/MS/MS, and pharmacokinetic parameters were determined. Step count, heart rate and rhythm, gastrointestinal borborygmi, fecal output, packed cell volume, and total protein were also assessed. Morphine‐3 glucuronide (M3G) was the predominant metabolite detected, with concentrations exceeding those of morphine‐6 glucuronide (M6G) at all time points. Maximal concentrations of M3G and M6G ranged from 55.1 to 504 and 6.2 to 28.4 ng/ml, respectively, across dose groups. The initial assessment of morphine pharmacokinetics was done using noncompartmental analysis (NCA). The volume of distribution at steady‐state and systemic clearance ranged from 9.40 to 16.9 L/kg and 23.3 to 32.4 ml min^?1 kg^?1, respectively. Adverse effects included signs of decreased gastrointestinal motility and increased central nervous excitation. There was a correlation between increasing doses of morphine, increases in M3G concentrations, and adverse effects. Findings from this study support direct administration of purified M3G and M6G to horses to better characterize the pharmacokinetics of morphine and its metabolites and to assess pharmacodynamic activity of these metabolites.     

Pharmacokinetics of morphine and plasma concentrations of morphine-6-glucuronide following morphine administration to dogs

The purpose of this study was to evaluate the pharmacokinetics of morphine and morphine-6-glucuronide (M-6-G) following morphine administered intravenously and orally to dogs in a randomized crossover design. Six healthy 3–4-year-old Beagle dogs were administered morphine sulfate (0.5 mg/kg) as an i.v. bolus and extended release tablets were administered orally as whole tablets (1.6 ± 0.1 mg/kg) in a randomized crossover design. Plasma concentrations of morphine and M-6-G were determined using high-pressure liquid chromatography and electrochemical coulometric detection. Following i.v. administration all dogs exhibited dysphoria and sedation, and four or six dogs vomited. Mean ± SE values for half-life, apparent volume of distribution, and clearance after i.v. administration were 1.16 ± 0.15 h, 4.55 ± 0.17 L/kg, and 62.46 ± 10.44 mL/min/kg, respectively. One dog vomited following oral administration and was excluded from the oral analysis. Oral bioavailability was 5% as determined from naïve-averaged analysis. The M-6-G was not detected in any plasma samples following oral or i.v. administration of morphine at a 25 ng/mL the limit of quantification. Computer simulations concluded morphine sulfate administered 0.5 mg/kg intravenously every 2 h would maintain morphine plasma concentrations consistent with analgesic plasma concentrations in humans. Oral morphine is poorly and erratically absorbed in dogs.     

Equine uridine diphospho-glucuronosyltransferase 1A1, 2A1, 2B4, 2B31: cDNA cloning,expression and initial characterization of morphine metabolism

ObjectiveUridine diphospho-glucuronosyltransferases (UGTs) are membrane-bound enzymes that catalyze the conjugation of glucuronic acid onto a diverse set of xenobiotics. Horses efficiently and extensively glucuronidate a number of xenobiotics, including opioids, making UGTs an important group of drug-metabolizing enzymes for the clearance of drugs. Recombinant enzymes have allowed researchers to characterize the metabolism of a variety of drugs. The primary objective was to clone, express and characterize equine UGTs using drugs characterized as UGT substrates in other species. A secondary objective was to characterize the in vitro metabolism of morphine in horses.Study designIn vitro drug metabolism study using liver microsomes and recombinant enzyme systems.AnimalsLiver microsomes and RNA from tissue collected from two Thoroughbred mares euthanized for other reasons.MethodsBased on homology to the human UGT2B7, four equine UGT variants were expressed: UGT1A1, UGT2A1, UGT2B31 and UGT2B4. cDNA sequences were cloned and resulting protein expressed in a baculovirus expression system. Functionality of the enzymes was assessed using 4-methylumbelliferone, testosterone, diclofenac and ketoprofen. Recombinant enzyme, control cells, equine liver microsomes and human UGT2B7 supersomes were then incubated with morphine. Concentrations of metabolites were measured using liquid chromatography–tandem mass spectrometry and enzyme kinetics determined.Results4-Methylumbelliferone was glucuronidated by all expressed equine UGTs. Testosterone glucuronide was not produced by any of the expressed enzymes, and diclofenac glucuronide and ketoprofen glucuronide were produced by UG2A1 and UGT1A1, respectively. UGT2B31 metabolized morphine to morphine-3-glucuronide and low concentrations of morphine-6-glucuronide.Conclusions and clinical relevanceThis is the first successful expression of functional recombinant equine UGTs. UGT2B31 contributes to the glucuronidation of morphine; however, it is probably not the main metabolizing enzyme. These results warrant further investigation of equine UGTs, including expression of additional enzymes and further characterization of UGT2B31 as a contributor to morphine metabolism.     

Pharmacokinetics and pharmacodynamics of morphine in llamas

OBJECTIVE: To assess the pharmacokinetics and pharmacodynamics of morphine in llamas. ANIMALS: 6 healthy adult llamas. PROCEDURES: Llamas received morphine sulfate in a randomized crossover design. In phase 1, they received IV or IM administration of morphine at 0.05 or 0.5 mg/kg, respectively; in phase 2, they received IV administration of morphine at 0.05, 0.25, or 0.5 mg/kg. Plasma morphine and morphine-6-glucuronide concentrations were determined by validated methods. Body temperature, heart rate, respiratory rate, sedation, and analgesia were assessed and compared with plasma concentrations by regression analysis. RESULTS: Total body clearance was similar between IV administration of morphine sulfate at 0.25 and 0.5 mg/kg (mean +/- SD, 25.3 +/- 6.9 mL/min/kg and 27.3 +/- 5.9 mL/min/kg, respectively), and linearity was demonstrated between these doses. Bioavailability of morphine following IM administration at 0.5 mg/kg was 120 +/- 30%. Body temperature and sedation increased as the dose of morphine administered increased. Heart rate was unaffected by varying doses. Respiratory rate decreased as dose increased. Analgesia was difficult to assess as a result of high individual variability. Intravenous administration of morphine at 0.25 mg/kg provided the most consistent increase in tolerance to electric stimulation. Pharmacodynamic modeling revealed a sigmoidal relationship between plasma concentration and sedation score. CONCLUSIONS AND CLINICAL RELEVANCE: Morphine was characterized by a large apparent volume of distribution and high systemic clearance in llamas. A prolonged half-life was observed with IM injection. Intravenous administration of morphine sulfate at 0.25 mg/kg every 4 hours is suggested for further study.     

Myxobolus cerebralis infection in rainbow trout (Oncorhynchus mykiss) and brown trout (Salmo trutta) exposed under natural stream conditions.

From early April into mid-June 1977, sequential groups of juvenile rainbow trout (Oncorhynchus mykiss) and brown trout (Salmo trutta) were each exposed for 10 days to the parasite Myxobolus cerebralis by immersion in a stream inhabited by infected wild trout. Following incubation in a M. cerebralis-free facility, trout were subsequently killed, and heads and gill arches were examined by routine histologic methods. A grading scale to quantify lesion severity was developed and applied. Percentage infected, lesion severity scores, effects of water temperature and flow rates on percentage infected and lesion severity scores, and resulting pathology were determined for each species at each exposure period. The percentage of rainbow trout infected with M. cerebralis was significantly higher than the percentage of brown trout infected for each exposure period. The percentages of rainbow trout infected in exposure periods later in the calendar year were significantly higher than those in earlier periods. The percentages of brown trout infected were not significantly different among exposure periods. Overall average lesion severity scores were significantly higher in rainbow than in brown trout. Lesion severity scores in rainbow trout increased over time (a positive correlation with exposure period). Lesion severity scores were not significantly different for brown trout among exposure periods. A significant correlation existed between water temperature and percentage of rainbow trout infected; a significant correlation also existed between water temperature and lesion severity scores in rainbow trout. Similar correlations did not exist for percentage of brown trout infected or accompanying lesion severity scores. In rainbow trout, ventral calvarium was the most common site of M. cerebralis replication, followed by gill arches. In brown trout, lesions were virtually confined to gill arches. Early lesions consisted of foci of cartilage necrosis with small numbers of M. cerebralis developmental stages. More advanced lesions consisted of multifocal areas of cartilage necrosis with numerous M. cerebralis developmental stages and/or mature myxospores bordered and/or infiltrated by mono- and multinuclear leukocytes. Lesions in brown trout were smaller and had fewer associated leukocytes and M. cerebralis developmental stages and/or mature myxospores. Higher infection rates, lesion severity scores, and differences in lesion location in rainbow versus brown trout explain in part why numbers of rainbow but not brown trout have fallen in western rivers inhabited with M. cerebralis-infected trout.     

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.     

Comparative pharmacokinetics of oxytetracycline in rainbow trout (Salmo gairdneri) and African catfish (Clarias gariepinus)

A comparative pharmacokinetic study was conducted in rainbow trout (Salmo gairdneri) and African catfish (Clarias gariepinus) following intravenous (i.v.) and intramuscular (i.m.) administration of oxytetracycline (OTC) at a dose rate of 60 mg/kg body weight. Trout and catfish were kept in aerated tap water in tanks at constant temperatures of 12 degrees C and 25 degrees C, respectively. The two- and three-compartment open models adequately described plasma drug disposition in African catfish and rainbow trout respectively, following i.v. OTC administration. Compared to catfish (COP = 86 +/- 10 micrograms/ml) an eightfold higher extrapolated zero time concentration was obtained in trout (COP = 753 +/- 290 micrograms/ml). A significant difference was observed with respect to the relatively large apparent distribution volumes (Vd(area] after i.v. OTC administration (trout, mean value: 2.1 l/kg; catfish, mean value: 1.3 l/kg). The mean final elimination half-lives of both fish species were greater than previously reported in mammals (trout, 89.5 h; catfish, 80.3 h). A mean maximum plasma concentration (Cmax = 56.9 micrograms/ml) was obtained in trout at 4 h after i.m. administration of OTC. In catfish a lower Cmax of 43.4 micrograms/ml was determined at about 7 h. No significant difference was observed with respect to bioavailability following i.m. administration of OTC (trout, 85%; catfish, 86%).     

Changes in ghrelin levels of plasma and proventriculus and ghrelin mRNA of proventriculus in fasted and refed layer chicks

This is a test-report of ghrelin levels in plasma and proventriculus, the glandular portion of the avian stomach, by using a specific radioimmunoassay for acylated ghrelin, as well as the expression of the ghrelin gene in the proventriculus after a 12-h fasting period followed by a 6-h feeding period with 6-day-old layer chicks. After fasting, the plasma ghrelin levels increased from 21.3 ± 4.5 to 32.9 ± 5.0 fmol/ml, but once refed it returned to the control value. After fasting, the ghrelin mRNA and the peptide levels in the proventriculus increased, and ghrelin mRNA levels remained high but once refed the ghrelin content returned to the control level. Furthermore, in order to examine the effect of increased circulating ghrelin on food intake, a bolus intravenous injection of 500 pmol of chicken ghrelin was given to 8-day-old chicks. The ghrelin injection did not cause any significant changes in food intake. These results indicate that the levels of ghrelin and its mRNA with layer chicks are altered according to the feeding state and this in a similar manner as has been observed in mammals. Unlike in mammals, an increase in circulating ghrelin does not cause the promotion of food intake in chicks.     

In vitro kinetics of hepatic albendazole sulfoxidation in channel catfish (Ictalurus punctatus), tilapia (Oreochromis sp.), rainbow trout (Oncorhynchus mykiss) and induction of EROD activity in ABZ-dosed channel catfish

Liver microsomes from market-size ( n  = 6) rainbow trout, channel catfish and tilapia were used to investigate in vitro biotransformation kinetics of albendazole (ABZ). ABZ was transformed to a single metabolite, ABZ sulfoxide (ABZ-SO). Catfish displayed the highest maximal velocity ( V _max = 264.0 ± 58.6 pmols ABZ-SO/min/mg protein) followed by tilapia (112.3 ± 8.2) and rainbow trout (73.3 ± 10.3). V _max in catfish was significantly different ( P  < 0.05) from the other two species. Michaelis–Menten constant ( K _m) values (μ m ) varied significantly among the species: rainbow trout (3.9 ± 0.5), tilapia (9.2 ± 1.7) and catfish (22.0 ± 3.2). However, V _max/ K _m ratios showed no difference among the three species, making them equally efficient performing this phase I biotransformation reaction. In a second series of experiments, channel catfish ( n  = 6 per treatment) were dosed in vivo with gel-food containing ABZ (10 mg/kg, p.o.). Fish were killed at 24, 48, 72 and 120 h after dosage. Control fish were fed ABZ-free feed. Induction of ethoxyresorufin-o-deethylase activity was significant ( P  < 0.05) in all ABZ-dosed treatments as compared with controls.     

The Effects of Whirling Disease on Growth and Survival of Snake River Cutthroat and Colorado River Rainbow Trout Fingerlings

Abstract

Two sizes of fingerling Snake River cutthroat trout Oncorhynchus clarkii behnkei and Colorado River rainbow trout O. mykiss were raised at hatcheries testing negative for Myxobolus cerebralis and stocked into the Dolores and Cache la Poudre rivers from 1999 to 2001. Populations were resampled over a 2-year period to determine which species and size combination had the highest growth and survival rates. Fish were tested for M. cerebralis via polymerase chain reaction and pepsin?trypsin digest analyses. Growth and survival rates between the species and size groups were not significantly different in either river. In the Dolores River, annual survival for both species and sizes of fish combined ranged from 0.063 to 0.12. In the Cache la Poudre River, survival for both sizes of rainbow trout was 0.004; survival for cutthroat trout ranged from 0.182 to 0.53. Larger fish had higher growth rates than smaller fish, and cutthroat trout had higher rates than similar sizes of rainbow trout. In both rivers, a higher percentage of the rainbow trout sample was infected than in the cutthroat trout sample. Rainbow trout also had a higher mean number of spores per head than cutthroat trout, and small rainbow trout had higher spore counts than large rainbow trout. Survival rates for cutthroat trout in the Cache la Poudre River were the highest of any of the groups, suggesting a difference that is biologically significant. Raising fingerlings to sizes greater than 100 mm can improve poststocking survival. If rainbow trout are stocked into contaminated waters, raising fingerlings to a larger size does not appear to improve growth or survival rates. Stocking rainbow trout in the spring could maximize growth rates but will expose fish to greater triactinomyxon densities, resulting in higher intensities of infection.     

A Comparative Investigation of Arcobacter cryaerophilus Infection among Albino Crosses and High- and Low-Body-Weight Rainbow Trout

Abstract

Arcobacter cryaerophilus was isolated from naturally infected rainbow trout Oncorhynchus mykiss, and its pathogenicity was tested by intramuscular injection into healthy 1-year-old high-body-weight (HBW) and low-body-weight (LBW) normally pigmented rainbow trout and albino crosses. Experimental infections caused deaths with gross clinical abnormalities such as exophthalmia, liver damage, bloody hemorrhagic kidney and heart, and swollen intestines. No significant differences in deaths were observed among the three infected fish groups. Hematocrit levels in blood of the experimentally infected HBW rainbow trout were significantly less than in healthy fish. No significant decreases were observed in the serum total protein of both the experimentally infected albino crosses and the high weight groups. Albumin and creatinine concentrations in serum were not significantly different among the three treatments.     

Distribution of Myxobolus cerebralis in Salmonid Fishes in Montana

Abstract

Over an approximately 2-year period, 20,974 fish (trout and other salmonid species) from 230 separate waters (creeks, rivers, lakes, reservoirs, ponds, hatcheries, and irrigation ditches) within 21 of the 22 major drainages in Montana were examined for Myxobolus cerebralis. Nine of the major river drainages have waters containing infected fish: Beaverhead, Big Hole, Blackfoot, Clark Fork above the Bitterroot River, Flathead below the south fork of the Flathead River, Jefferson, Madison, Missouri above the Marias River, and Yellowstone above the Bighorn River. The Beaverhead, Clark Fork above the Bitterroot River, Jefferson, Madison, and Missouri above the Marias River have the greatest number of waters containing fish infected with M. cerebralis. Comparisons of infection levels (number of pooled samples that contain fish infected with M. cerebralis) between species among these drainages show significantly lower levels of infection in brown trout Salmo trutta and rainbow trout Oncorhynchus mykiss in the Missouri above the Marias River drainage and significantly higher levels of infection of rainbow trout in the Jefferson. Comparisons of differences in infection levels between drainages among species show that, in the Beaverhead, Clark Fork above the Bitterroot River, and Madison, infection levels in brown trout are significantly higher than in rainbow trout. This is partially attributed to losses of juvenile rainbow trout because of M. cerebralis infection, leading to biased samples. Histopathologic studies showed lesions were consistently less severe in brown trout than other species and occurred in a different location (gill arches versus ventral calvarium). In six of the nine affected drainages (Beaverhead, Blackfoot, Clark Fork above the Bitterroot River, Flathead below the South Fork, Jefferson, and Madison), infected fish were found at or near the time that intensive sampling was initiated in the spring of 1995. In the three remaining affected drainages (Missouri above the Marias River, Yellowstone above the Bighorn River, and the Big Hole), infected fish were not identified until at least 15 months after the initiation of widespread testing. This indicates that in the first six drainages listed above, the infection was well established prior to 1995 but spread into the last three drainages in the ensuing months. Methods of transmission and the sources of infection are unknown, although the absence of infected fish in state, private, and federal hatcheries in Montana indicates hatchery fish from these sources are not likely to be responsible.     

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.