The effects of levamisole on oxidative stress induced by copper intoxication in broilers

Abstract

AIM: To determine the effects of Cu and levamisole on concentrations of Cu and Fe in plasma and liver, and the effects of levamisole on lipid peroxidation induced by Cu intoxication in broiler chickens.

METHODS: In a 2×4 factorial study, 80 one-day-old Ross PM3 broiler chicks were fed diets for 21 days containing either 8 mg/kg Cu (Low Cu) or 250 mg/kg Cu (High Cu) and were treated with 0 (L0), 4 (L4), 8 (L8) or 16 (L16) mg/kg bodyweight levamisole per day from Day 7 of the study, on three consecutive days in their drinking water. This treatment was repeated three times, at 3-day intervals. On Day 21, blood samples were collected from each bird for analysis of concentrations of Cu, Fe and malondialdehyde, and activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), superoxide dismutase, catalase and glutathione peroxidase (GSH-Px). The birds were killed and liver samples collected for analysis of Cu and Fe.

RESULTS: Mean concentrations of Cu and Fe in plasma, and Cu in liver, were increased overall in the High Cu groups compared with the Low Cu groups (p<0.001). Compared with the L0 treatment group on the High Cu diet, treatments L4, L8 and L16 decreased concentrations of Cu in plasma, and L8 and L16 increased concentration of Cu in liver (p<0.05). Mean activities of AST and ALT were increased in untreated birds (L0) fed the High compared with Low Cu diets (p<0.01). In birds receiving the High Cu diet, treatments L4 and L8 decreased activities of AST, and L4 and L16 decreased activity of ALT, compared with L0 (p<0.05). The High Cu diet induced an oxidative stress characterised by increased mean concentrations of malondialdehyde and decreased activities of superoxide dismutase, catalase and GSH-Px (p<0.001). Concentration of malondialdehyde, and activities of superoxide dismutase and catalase were not changed following levamisole treatment in birds on the High Cu diet, and activity of GSH-Px was decreased by the L4 and L8 treatments compared with the L0 group.

CONCLUSIONS AND CLINICAL RELEVANCE: The results of the study suggest that treatment with levamisole might alleviate the harmful effects of Cu on the liver, as demonstrated by decreased activities of AST and ALT induced by a diet containing 250 mg/kg Cu.     

Fatal levamisole toxicosis of captive kiwi (Apteryx mantelli)

CASE HISTORY: Nine of 24 captive kiwi treated with oral levamisole at a dose between 25–43 mg/kg showed signs of respiratory distress. Six died within 4 h of treatment and the remaining three made a full recovery within 24 h.

CLINICAL AND PATHOLOGICAL FINDINGS: Within 3–4 h of treatment, the affected birds had an elevated respiratory rate, mucoid nasal discharge and rapidly became comatose. Post mortem examination revealed accumulation of thick mucus in the oral cavity and trachea. There was severe pulmonary congestion and oedema and early bronchopneumonia in the lungs of five of the birds. In two birds, there was acute hepatic degeneration and necrosis and one bird had acute pancreatic degeneration and necrosis.

DIAGNOSIS: Acute levamisole toxicity.

CLINICAL RELEVANCE: Kiwi were acutely sensitive to levamisole toxicity at doses that are well within the safe range for domestic poultry. Levamisole should not be used as an anthelmintic in kiwi.     

Subarachnoid butorphanol augments lidocaine sensory anaesthesia in calves

To determine the efficacy and safety of subarachnoid butorphanol combined with lidocaine, six calves were studied. Each calf underwent two treatments, at least one week apart, via subarachnoid injection: (1) butorphanol (0.03 mg/kg) plus 2% lidocaine (4 mg/kg) and (2) 2% lidocaine (4 mg/kg) alone. Subarachnoid injections were performed at the lumbosacral space. Analgesia, motor block, sedation, heart rate, arterial blood pressure, respiratory rate, arterial oxygen saturation as measured by pulse oximetry, and rectal temperature were compared before and after subarachnoid administration of drugs. Subarachnoid administration of the butorphanol-lidocaine combination induced bilateral prolonged analgesia extending from the coccygeal to the T11-T13 dermatomes in the calves, with minimal sedation and severe ataxia. Cardiovascular effects were significant in both treatments: heart rate was increased, and there was a minimal decrease in arterial pressure. It was concluded that adding a small dose of butorphanol to subarachnoid lidocaine in calves is effective and safe.     

Pemoline and tetramisole 'positives' in English racehorses following levamisole administration

Pemoline is a central nervous system stimulant that has been used to treat attention-deficit hyperactivity disorder and narcolepsy in humans; its identification in horses could be considered evidence of attempts to influence performance. Two recent pemoline 'positives' in English racehorses led us to review the chemical relationships between tetramisole, levamisole, aminorex and pemoline. Pemoline is a simple oxidation product of aminorex, which has been shown in the United States and elsewhere to be an equine metabolite of levamisole. Based on the clear structural relationships between aminorex and pemoline, we conclude that levamisole can metabolise to pemoline in horses and that pemoline identifications in horses post levamisole administration are likely to be associated with levamisole administration. Levamisole should not be administered to horses about to compete because of its ability to metabolise to two central nervous system stimulants, aminorex and pemoline.     

Efficacies of fenbendazole and levamisole in the treatment of commercial turkeys for Ascaridia dissimilis infections

A field trial was conducted on a farm in Michigan with commercial turkeys naturally infected with Ascaridia dissimilis. The birds were treated at approximately 8 and 14 wk of age on a per-barn basis with fenbendazole (Safeguard) in the feed at 16 ppm for 6 d (1 barn), or with levamisole (Prohibit) in the water at 16 mg/kg of BW (2 barns). Six birds were obtained at random and sacrificed in each barn for worm burden determinations before treatments and again soon after treatments. Nematicidal efficacies for fenbendazole and levamisole were shown to be 99.3 to 99.9% and 54.6 to 85.8%, respectively. Despite the far greater reductions in worm burdens by fenbendazole as compared with levamisole, the burdens at the time of retreatment had equalized in all barns, illustrating the role of premunition regarding turkey ascaridiasis, and highlighting the overriding importance of reducing the challenge to provide long-term control of this parasitism.     

Dynamics of viral spread in bluetongue virus infected calves

The kinetics of viremia and sites of viral replication in bluetongue virus (BTV) infected calves were characterized by virus isolation, serology and immunofluorescence staining procedures. In addition, the role of the regional lymph node and lymphatics draining inoculated skin in the pathogenesis of BTV infection was determined by analyzing efferent lymph collected from indwelling cannulas. Viremia persisted for 35 to 42 days after inoculation (DAI) and virus co-circulated with neutralizing antibodies for 23 to 26 days. Virus was first isolated from peripheral blood mononuclear (PBM) cells at 3 DAI, after stimulation of PBM cells with interleukin 2 and mitogen. BTV was frequently isolated from erythrocytes, platelets and stimulated PBM cells but never from granulocytes and rarely from plasma during viremia. Virus was consistently isolated from erythrocytes late in the course of veremia. Interruption of efferent lymph flow by cannulation delayed the onset of viremia to 7 DAI. BTV was infrequently isolated from lymph cells, and few fluorescence positive cells were observed after lymph and PBM cells were labelled with a BTV-specific monoclonal antibody. Virus was isolated from spleen by 4 DAI and most tissues by 6 DAI, whereas virus was isolated from bone marrow only at 10 DAI. Virus was not isolated from any tissue after termination of viremia. It is concluded that primary viral replication occurred in the local lymph node and BTV then was transported in low titer to secondary sites of replication via infected lymph and PBM cells. We speculate that virus replication in spleen resulted in release of virus into the circulation and non-selective infection of blood cells which disseminated BTV to other tissues. Virus association with erythrocytes likely was responsible for prolonged viremia, although infected erythrocytes eventually were cleared from the circulation and persistent BTV infection of calves did not occur.     

我国发生Leachii支原体引起的犊牛多发性关节炎

2009年2月～6月我国某奶牛场发生严重的犊牛多发性关节炎,发病犊牛的症状与最早发生于澳大利亚的犊牛Leachii支原体关节炎非常相似。为确定病原,我们无菌采集2份具有典型症状犊牛的关节液样品进行实验室诊断,2份样品中均检测和分离出支原体。将2个分离菌株的16S rRNA基因和LppA基因与参考支原体菌株进行核苷酸序列比对,发现这2株支原体的16S rRNA基因和LppA基因均与Leachii支原体具有最高的核苷酸序列同源性,分别为99.9%和99.6%。结果显示,本研究分离的2株支原体为Leachii支原体,分别命名为GN407和GN408。综合分析发病犊牛的临床病理学观察和关节液样品的实验室诊断结果,我们确定Leachii支原体为该奶牛场犊牛多发性关节炎的病因。     

Adverse reactions to fentanyl transdermal patches in calves: a preliminary clinical and pharmacokinetic study

Objective

To describe adverse reactions and measure plasma fentanyl concentrations in calves following administration of a fentanyl transdermal patch (FTP).

An extended outbreak of congenital chondrodysplasia in calves in South East Australia

OBJECTIVE: To report an outbreak of congenital chondrodystrophy in calves in South East Australia. METHODS: District veterinarians investigated reported cases of calf deformities. Owners of affected herds were interviewed using a standard questionnaire to identify potential risk factors. Dams of several affected calves were serologically tested for Akabane virus, Aino virus, pestivirus and bluetongue, and affected calves were tested for pestivirus antigen and serum immunoglobulin concentrations. Gross and histopathological examinations of numerous calves were performed, concentrating on the musculoskeletal system. RESULTS: A case definition of distinctive skeletal deformities was established, and 89 property owners reported calves with chondrodystrophy in Spring 2003, 2004 or 2005. Some 14 property owners reported affected calves in more than one year. Prevalence and severity of deformity varied greatly between and within properties. None of breed, sex, age of dam, lineage, pasture type, supplementary feeding, fertiliser use or toxic plants was consistently associated with the disease. All dams experienced hot, dry conditions during the first trimester of pregnancy and were exposed to adverse conditions thereafter. Consistently dams were reported to have been grazing undulating to hilly terrain during early pregnancy. All serological tests were negative for Akabane virus, Aino virus, pestivirus and bluetongue. Histopathology of affected skeletal samples showed chondrodysplasia. CONCLUSION: The outbreak had similarities with previous outbreaks reported in the region. No specific aetiology could be determined. There is some evidence that the cause of the deformities could be a manganese deficiency during foetal development. Ongoing work to test this hypothesis is therefore warranted.     

Pharmacokinetic study of an injectable long-acting parenteral formulation of doxycycline hyclate in calves

Doxycycline hyclate (DOX-h) can be regarded as a time-dependant antibacterial. Hence, a parenteral long-acting formulation may be regarded as more pharmacologically sound. A poloxamer-based matrix was used to produce a long-acting injectable preparation (DOX-h-LA) and its serum concentrations vs. time profile investigated after its s.c. injection to calves. Serum concentrations profiles for such a prepartion were compared to the corresponding profiles obtained with an aqueous formulation of DOX-h injected either i.m. or i.v. in 10 calves in a crossover study at dose of 10mg/kg, with washout periods. DOX-h-LA showed the greatest values for bioavailability (602%); maximum serum concentration (C(max)) value was 1.99microg/mL with a time to reach C(max) (T(max)) of 25h and an elimination half-life of 40.81h. Considering minimum effective serum concentration of 0.5microg/mL a dose-interval of 80h can be achieved for DOX-h-LA, and only 9.7h and 17h after the i.v. or i.m. administration of DOX-h, respectively.     

High infection rate of Trichophyton verrucosum in calves from Central Italy

Recent epidemiological features of the zoonotic dermatophyte Trichophyton verrucosum were investigated in 294 calves from 20 farms located in Central Italy. By the combination of microscopic examinations and fungal cultures, the total prevalence rate of T. verrucosum infection was found to be high, with 87.7% of samples yielding positive results and 100% of farms being infected. Farm to farm prevalence of infection varied from 25% to 100% of the examined calves. Prevalence rates were higher in females than males (91.6% versus 84%), in >3-< or =6 than >6-< or =9 month old calves (89.8% versus 85.7%), in purebred than in crossbred (88.5% versus 81.8%), and in symptomatic than asymptomatic ones (95.7% versus 80.4%). T. verrucosum was as prevalent in semi-intensive as in intensive breeding system (88.2% versus 87.6%) but more prevalent in calves raised for meat rather than milk production (91.1% versus 84.9%), and reared under conventional than organic production management system (89.3% versus 82.6%). Prevalence rates were also higher when the cattle present in the farm were of mixed origin, rather than when the whole stock was purchased or was born in the farm (92% versus 88.2% and 85.7%), when other cattle in the farm were showing skin signs (90% versus 84.2%), and at least one case of human ringworm was recorded in the farm (90.9% versus 86.1%). No statistically significant association was found. Therefore, T. verrucosum infection may be more widespread in calves from Central Italy than it was previously thought to be. Calves of any sex, breed, age, and of any of the farm condition considered, are highly exposed to the risk of infection. As calves may play a considerable role as reservoirs of the infection, our results highlight the need to determine the incidence and the importance of T. verrucosum ringworm in personnel involved with cattle care in Italy and elsewhere.     

Changes in peripheral leukocytes enzymes activity and plasma metabolite concentrations in growing Holstein calves

Plasma metabolite and immunoreactive insulin concentrations and activities of enzymes related to energy metabolism in peripheral leukocytes were measured in growing Holstein calves. A ratio of girth of abdomen divided by girth of thorax (A/T ratio) of calves was significantly elevated after weaning, and the A/T ratio maybe a good indicator to evaluate rumen development. Plasma glucose and free fatty acid concentrations were changed in calves accompanying change in feeding. Activities of lactate dehydrogenase with pyruvate as substrate (LDH-P) and hexokinase (HK) in cytosolic fractions of peripheral leukocytes decreased significantly after weaning the calves reflecting the change of energy source from milk replacer with high percentages of fat and glucose and lactose as absorbable carbohydrate to pelleted feed containing starch as less absorbable carbohydrate and roughage. Some peripheral leukocyte enzymes such as LDH and HK may be good indicators to evaluate changes in energy metabolism of growing calves.     

Clinical assessment of epidural analgesia induced by xylazine-lidocaine combination accompanied by xylazine sedation in calves

: The aim of the present study was to investigate whether epidural administration of a xylazine-lidocaine combination accompanied by xylazine sedation would provide satisfactory analgesia for some surgical procedures on 10 calves admitted to the Department of Veterinary Surgery, University of Kafkas with perineal urolithiasis (n:2), rectovaginal fistula (n:1), atresia ani (n:2), omphalophlebitis (n:2), omphaloarteritis (n:1) and umbilical hernia (n:2).Following intramuscular injection of xylazine at a dose of 0.05 mg/kg for sedation, xylazine-lidocaine combination (0.2 mg/kg lidocaine + 0.02 mg/kg xylazine + 5 ml 0.9% NaCl) was administrated into the lumbosacral (L6-S1), sacrococcygeal (S5-Co1) or intercoccygeal (Co1-Co2) space. Heart rate, respiratory rate and rectal temperature were recorded prior to and during analgesia at 5, 10, 15, 30 and 60 minutes. Furthermore, depth and duration of analgesia were evaluated during surgical intervention.The study revealed that the combination of epidural xylazine-lidocaine with xylazine sedation was highly satisfactory for surgery of the lower urinary tract and the perineal region, but it was less so for surgery of the umbilical area.     

Weaning and post-weaning performance by fall-born beef calves weaned on different dates in the spring from Neotyphodium coenophialum-infected tall fescue pastures

Fall-born calves grazing Neotyphodium coenophialum-infected tall fescue [E+; Lolium arundinaceum (Schreb.) Darbysh.] pastures should benefit from early weaning because of reduced exposure to fungal toxins. However, fall-born calves that grazed E+ and were weaned in mid-April had reduced post-weaning performance compared with calves managed similarly but weaned in early June. Gelbvieh × Angus calves (n = 238) were used in a 3-year study to determine the optimal time to wean fall-born calves grazing E+. Cow/calf pairs were allocated randomly to one of four weaning date treatments: 1) March 16 (177 ± 4.7 days of age; MarW), 2) April 13 (204 ± 4.7 days of age; AprW), 3) May 11 (236 ± 4.7 days of age; MayW), and 4) June 8 (264 ± 4.8 days of age; JuneW). On their assigned weaning date, calves were weighed, vaccinated, blood samples were collected, and calves were moved to 3.2-ha pastures adjacent to their dams for 14 days. After the weaning period, calves were weighed, blood samples were collected, and then calves were moved to pastures containing non-toxic forages. Birth weight and calf weights on MarW, AprW, and MayW weaning dates did not differ (P ≥ 0.21) across weaning date treatments. However, calf weaning weights on their respective weaning dates, weight on JuneW and on June 22 (14 days following the JuneW), daily gain between birth and June 22, and weight change between MarW and JuneW increased linearly (P < 0.05) across weaning dates. Linear decreases (P < 0.05) across weaning dates were noted for antibody titers to bovine virus diarrhea (BVD) and infectious bovine rhinotracheitis (IBR), serum Cu, and platelets at the time of actual weaning. Linear increases (P < 0.05) across weaning dates were noted for: i) total antioxidant potential and eosinophils at weaning; ii) changes in antibody titers to BVD and IBR, total antioxidant potential, and serum Zn during the 14-day post-weaning period; and iii) and changes in BVD and BRSV titers and serum Cu between actual weaning date and June 22. Heifer weight at breeding increased (P < 0.05) and steer weight at shipping, hot carcass weight and backfat thickness tended (P ≤ 0.10) to increase linearly across weaning dates. Final feedlot weight and gain did not differ (P ≥ 0.18) among weaning dates. Therefore, delaying weaning of fall-born calves grazing E+ pastures until early June may be beneficial for calf weight and immune function at weaning, and heifer weight at breeding, but those benefits in steers may be mitigated through the feedlot period.     

Combined administration of ghrelin and GHRH synergistically stimulates GH release in Holstein preweaning calves

Ghrelin is a gut peptide which participates in growth regulation through its somatotropic, lipogenic and orexigenic effects. Synergism of ghrelin and growth hormone-releasing hormone (GHRH) on growth hormone (GH) secretion has been reported in humans and rats, but not in domestic animals in vivo. In this study, effects of a combination of ghrelin and GHRH on plasma GH and other metabolic parameters, and changes in plasma active and total ghrelin levels were studied in Holstein bull calves before and after weaning. Six calves were intravenously injected with vehicle (0.1% BSA-saline), ghrelin (1 microg/kg BW), GHRH (0.25 microg/kg BW) or a combination of ghrelin plus GHRH at the age of 5 weeks and 10 weeks (weaning at 6 weeks of age). Ghrelin stimulated GH release with similar potency as GHRH and their combined administration synergistically stimulated GH release in preweaning calves. After weaning, GH responses to ghrelin and GHRH became greater compared with the values of preweaning calves, but a synergistic effect of ghrelin and GHRH was not observed. The GH areas under the concentration curves for 2h post-injection were greater in weaned than in preweaning calves (P<0.05) if ghrelin or GHRH were injected alone, but were similar if ghrelin and GHRH were injected together. Basal plasma active and total ghrelin levels did not change around weaning, but transiently increased after ghrelin injection. Basal plasma insulin, glucose and non-esterified fatty acid levels were reduced after weaning, but no changes by treatments were observed. In conclusion, ghrelin and GHRH synergistically stimulated GH release in preweaning calves, but this effect was lost after weaning.     

Effects of natural zeolite clinoptilolite on passive immunity and diarrhea in newborn Holstein calves

In the present study, the effects of natural zeolite clinoptilolite on absorption of immunoglobulins from colostrum and incidence of enteric diseases were evaluated. In a completely randomised design, thirty Holstein calves were fed pooled colostrum and then milk containing zero (control), 0.5 (T1), 1.0 (T2), 1.5 (T3) and 2.0 (T4) g clinoptilolite per kg body weight per day through day 45. Blood was collected after birth and at 24 h of age and plasma IgG and IgM concentrations were determined. Fecal consistency score and severity of diarrhea were recorded for each calf twice daily. Calves receiving T3 and T4 had lower (P < 0.05) plasma IgG concentration than control and other treatments. Calves on T2 had higher (P < 0.05) plasma IgG concentration than T3 and T4, but not T1 and control. Inclusion of clinoptilolite to colostrum did not affect (P > 0.05) IgM absorption from the intestine of newborn calves. Fecal consistency scores were lower (P < 0.05) for calves on T1 and T2 and higher for calves on T3 and T4 than calves on control. Percent calf days with diarrhea followed the same trend. In overall, seven calves died, those being one each on control and T1, two on T3 and three on T4. Based upon these results, addition of 1.0 g clinoptilolite per kg body weight per day to colostrum and milk could reduce diarrhea, but its effect on passive immunity was negligible. Over 1.0 g/kg body weight per day, clinoptilolite had adverse effect on passive immunity and diarrhea.     

Pharmacokinetics of levamisole in the red‐eared slider turtles (Trachemys scripta elegans)

The pharmacokinetics and bioavailability of levamisole were determined in red‐eared slider turtles after single intravenous (IV), intramuscular (IM), and subcutaneous (SC) administration. Nine turtles received levamisole (10 mg/kg) by each route in a three‐way crossover design with a washout period of 30 days. Blood samples were collected at time 0 (pretreatment), and at 0.25, 0.5, 1, 1.5, 3, 6, 9, 12, 18, 24, 36, and 48 hr after drug administration. Plasma levamisole concentrations were determined by a high‐performance liquid chromatography assay. Data were analyzed by noncompartmental methods. The mean elimination half‐life was 5.00, 7.88, and 9.43 hr for IV, IM, and SC routes, respectively. The total clearance and volume of distribution at steady state for the IV route were 0.14 L hr^?1 kg^?1 and 0.81 L/kg, respectively. For the IM and SC routes, the peak plasma concentration was 9.63 and 10.51 μg/ml, respectively, with 0.5 hr of T_max. The bioavailability was 93.03 and 115.25% for the IM and SC routes, respectively. The IM and SC route of levamisole, which showed the high bioavailability and long t_1/2?z, can be recommended as an effective way for treating nematodes in turtles.     

Pharmaco‐parasitological evaluation of the ricobendazole plus levamisole nematodicidal combination in cattle

The goals of the current study were to evaluate the potential pharmacokinetic (PK) interactions and the clinical efficacy occurring after the subcutaneous (s.c.) administration of ricobendazole (RBZ) and levamisole (LEV) given both separately and co‐administered to calves naturally infected with susceptible gastrointestinal nematodes. The clinical efficacy was shown in two seasons, winter and spring, with predominance of different nematode populations. Groups of 15 calves were treated with RBZ alone, LEV alone and RBZ + LEV combination, and an untreated group was kept as a Control. RBZ and LEV plasma concentrations were quantified by HPLC. The clinical efficacy was determined by the faecal egg count reduction test. RBZ and LEV have similar plasma persistence, being detected in plasma over 24 hr post‐treatment. No PK interactions were observed after the combined treatment, with similar PK parameters (p > .05) obtained for the single‐drug and the combination‐based strategy. In winter, the observed clinical efficacies were 96%, 99% and 100% for groups treated with RBZ, LEV and RBZ + LEV, respectively; however, in spring, the efficacies were 95%, 93% and 96% for the same groups. Remarkably, the combination was the only treatment that achieved 100% clinical efficacy against both Haemonchus spp and Ostertagia spp in winter; but the increased presence of Ostertagia spp. in spring (28% in untreated group) determined a tendency to reduced efficacies compared to winter time (only 10% of Ostertagia spp. in untreated group), even for the combined treatment. Overall, in a scenario where the nematode population is susceptible, the RBZ + LEV treatment may be a valid combination in cattle to delay the development of resistance, especially in winter when this combination achieved 100% of efficacy. Thus, selection of anthelmintic resistance will never occur. In fact, this is one of the greatest challenges for the whole cattle production system: to be one step ahead of anthelmintic resistance.     

Unusual congenital deformities of the lower limb in two calves

Congenital deformities of the lower limb were observed in 2 calves from different properties. One was a newborn Holstein-Friesian bull calf with polydactyly and polypodia of the right fore-limb.The other was an 8-month-old Friesian/Jersey crossbred heifer calf, which had a mirror image duplication of the plantar/palmar half of the distal portion of the digits. The abnormalities in the second calf have not previously been described in cattle. This paper presents details of the clinical, radiological and necropsy findings in these 2 calves.     

Prevalence of Giardia and Cryptosporidium in dairy calves in three areas of Norway

A study was undertaken to determine the prevalences of Cryptosporidium and Giardia in dairy calves less than 6 months of age in Norway. Faecal samples were collected from a total of 1386 calves, between 3 and 183 days of age, in 136 dairy farms from three different areas of Norway. Faecal samples were examined for Cryptosporidium oocysts and Giardia cysts after concentration and immunofluorescent staining.

Giardia was found in 93% (127 out of 136) of the farms and in 49% (679 out of 1386) of the calves. Cryptosporidium was found in 53% (72 out of 136) of the farms and in 12% (167 out of 1386) of the calves. The level of Giardia and/or Cryptosporidium was low in the majority of the infected calves.

Infection peaked in the age group 2–3 months for both Cryptosporidium and Giardia. The prevalences of both parasites were higher in samples taken during winter than in samples taken during summer, and statistically significant differences were found when prevalences in different age groups of calves were compared between the three areas. A significantly lower prevalence of Cryptosporidium was found in calves housed in shared pens that were thoroughly washed more than three times a year than in calves from pens washed less often. For Giardia there was a trend for decreasing intensity of infection with increasing age in the sampled calves. For Cryptosporidium there was a trend for increasing herd prevalence with increasing number of calves in the herd, but this trend was not statistically significant. Other parameters which were investigated such as housing, feeding or management routines were not associated with prevalence or intensity of infection with either parasite.