Investigation of the effects of transport and lairage on hydration state and resting behaviour of calves for export.

Plasma sodium concentrations, packed cell volume, total plasma protein concentration and dermal skinfold thickness were used to assess the state of hydration of 113 transported calves before and after lairage and in 30 control calves on the farm of origin. Skin thickness increased significantly (P less than 0.05) and total protein increased slightly during transport and decreased during lairage. These changes suggest that transport may cause dehydration and lairage may help in recovery. Plasma potassium concentration decreased during transport, but the effect was inversely related to the distance travelled, and the concentration increased during lairage. These changes are consistent with recovery from initially high cortisol levels at loading. The resting behaviour of 150 transported calves was recorded hourly for six hours. More time was spent resting and sleeping by the transported calves than has been reported for non-transported calves and more still by small transported calves, suggesting that transport is exhausting, that lairage helps recovery and that small calves are more adversely affected. During lairage the numbers of calves asleep decreased to values reported for normal calves, suggesting that 10 hours lairage was adequate. However, small calves did not return to their normal rest patterns within the observation period.     

Effects of food withdrawal and transport on 5- to 10-day-old calves

The effects of food withdrawal for 30 hours and transport for up to 12 hours on 5- to 10-day-old calves were determined by monitoring plasma concentrations of glucose, beta-hydroxybutyrate, urea, lactate and creatine phosphokinase. In addition gamma-glutamyl transferase activity, packed cell volume, total plasma protein concentration, bodyweight and rectal temperature were recorded. Food withdrawal for 30 hours caused hypoglycemia but calves maintained normothermia and although their beta-hydroxybutyrate level increased indicating lipid mobilisation, the changes in blood urea level were minimal indicating low amino acid catabolism. When calves were transported for 12 hours in addition to having food withdrawn for 30 hours, they remained normoglycemic for six hours longer than non-transported calves, probably due to an effect of muscular activity during bracing against truck movements. Stocking calves at a lower density during transport apparently had a beneficial effect, as calves were able to lie down throughout the journey and therefore produced a similar metabolic profile to resting, non-transported animals. Transport and food withdrawal had no obvious effects on calf hydration. The results of this study suggest that food withdrawal for up to 30 hours and transport for up to 12 hours had no detrimental effects on the metabolism of healthy and clinically normal calves. With correct feeding regimes and transport protocols, welfare compromise in young, healthy calves being transported for up to 12 hours can be minimised when they are slaughtered within 30 hours of the start of transport.     

A study of the effect of acute systemic anaphylaxis on blood coagulation and TAME esterase activity in calves.

Sensitized and control calves were challenged with an intravenous dose of 0.2 mL/kg horse serum. Changes in the blood pressure, blood cellular components, plasma kaolin activated p-tosyl-1-arginine methyl ester esterase activity, plasma antithrombin III levels and activated partial thromboplastin time were monitored. Anaphylaxis induced a severe drop in carotid arterial pressure and respiratory distress. There was a decrease in the total white blood cell count from a mean of 10,000 to a low of 1,900 per microL, a decrease in the percentage neutrophils and an increase in the percentage of lymphocytes from 57.4% to 94.8%. A drop was observed in the mean platelet count from 463 x 10(3)/microL. The time required for kaolin to produce maximum p-tosyl-1-arginine methyl ester esterase activation increased from two minutes in the controls to three minutes in calves undergoing anaphylaxis and was observed three to 90 minutes after the administration of horse serum. Antithrombin III levels in the plasma appeared to drop during anaphylaxis but were not significantly depressed (p greater than or equal to 0.5). A statistically significant (p < 0.5) drop in the rate of blood coagulation was observed by the activated partial thromboplastin time assay at 15 to 30 minutes after horse serum injection.     

Selenium effects on glutathione peroxidase and the immune response of stressed calves challenged with Pasteurella hemolytica

The present study was conducted to determine whether a marginal Se deficiency affects health, blood characteristics and the immune response of calves subjected to stresses associated with weaning, shipping (332 km) and Pasteurella hemolytica inoculation. Treatments were 1) -Se, 2) -Se/P. hemolytica, 3) +Se (.1 mg Se/kg feed) and 4) +Se/P. hemolytica. Previous Se intake was controlled; dams of -Se calves were fed diets marginally deficient in Se (.03 to .05 mg/kg), whereas dams of +Se calves received a s.c. injection of 30 mg Se (as sodium selenite) every 60 d. Calves were inoculated with P. hemolytica intratracheally on d 3 following weaning and transport. Inoculation with P. hemolytica increased (P less than .05) body temperatures, platelet counts, serum IgM concentrations and serum antibody titers and decreased serum albumin concentrations at 4 to 7 d postinoculation. Weight gains for the 21-d study were not affected by Se status, although whole blood and plasma glutathione peroxidase (GSH-Px) were higher (P less than .05) for +Se calves. Plasma GSH-Px increased (P less than .01) in calves showing signs of morbidity. Increases in plasma GSH-Px were correlated positively with body temperature. Serum IgM concentrations were higher (P less than .05) in +Se calves on d 17, but Se-supplemented calves had lower (P less than .05) anti-P. hemolytica titers on d 17 than -Se calves. Selenium status did not affect body temperatures, plasma creatine phosphokinase or serum IgG and albumin concentrations. These results indicate that Se status can affect IgM concentrations following stress.     

Blood bactericidal assay (Pasteurella haemolytica) comparison of morbidity in marketed feeder calves

An in vitro bactericidal assay that used bovine heparinized blood was investigated for its usefulness in detecting differences in the bactericidal immunity of calves against Pasteurella haemolytica serotype 1 (Ph1). Greater than 90% of killing occurred within 30 minutes. The substitution of fetal calf serum for autologous calf plasma caused loss of bactericidal activity of the blood. Decomplemented calf serum also was low in bactericidal activity. The blood bactericidal assay appears to be opsonin antibody-dependent and complement-dependent. The coefficient of variation (CV) that can be expected with this assay was established by use of a group of 8 calves; within-day CV maximum was 0.9, and between-day CV maximum was 2.1. The blood bactericidal assay was used to evaluate 30 calves under typical market stress from 4 farms in eastern Tennessee. All calves had decreased bactericidal activity, as they moved into a feedyard in Texas. The bactericidal activity was reduced among sick calves, based on the severity of clinical signs. Morbidity was highest during the first 14 days in the feedlot. During this period, healthy calves had a decreased bactericidal index (BI) of 4 points, and calves with clinical signs of bovine respiratory tract disease for 3 days had a decreased BI of 8 points. The average reduction in the BI of calves with clinical signs of bovine respiratory tract disease for 6 or more days was 14 points.     

The influence of farmers' behavior on calves' reactions to transport and quality of veal meat

The relationships between farmers' behavior toward veal calves, calves' responses to handling and transport, and veal meat quality were assessed. Two groups of 10 veal units were selected based on previous observed farmers' behavior toward the calves: one group consisted of farmers who had shown predominantly "positive" behavior toward the calves, and the other group of farmers had shown predominantly "negative" behavior. Calves were observed for their reactions to people at the unit, and 20 calves per veal unit were transported either directly to the slaughterhouse or subjected to additional transport consisting of a supplementary 20-min transport with additional unloading and loading. The effort needed to load the calves onto the truck and their behavior during loading was observed. During loading and unloading, and during lairage at the slaughterhouse, potentially traumatic incidents (falling down, hits against structures, slips) were recorded, and heart rate and cortisol measurements were taken. Carcasses were evaluated on their weight, color, conformation, pH, and bruise level. A meat sample was taken from the longissimus thoracis muscle for physical, chemical, and sensory analysis. Calves originating from "positive behavior" units showed fewer fear responses to people at the veal unit, needed less effort to be loaded to the truck, had lower heart rates during loading and unloading, and had fewer incidents at the slaughterhouse than calves from "negative behavior" units (P < 0.05). Carcasses from calves from "positive behavior" units were paler, and analyses of the meat sample revealed lower pH, moisture level, and redness compared to carcasses from calves from "negative behavior" units (P < 0.05). Additional transport led to a lower cortisol level after transport and to higher carcass pH values at slaughter compared to direct transport (P < 0.05) but did not affect meat quality. We concluded that farmers' positive behavior toward veal calves during rearing is likely to reduce the emotional responses of calves to handling and transport and to lead to fewer incidents, compared to negative behavior. This reduction of calves' emotional responses seems to be the reason for improved veal meat color.     

The calf training for loading onto vehicle at weaning

The aim of this study is to examine whether calf training for loading at the weaning period improves later loading. Five calves were allocated to the trained group and the control group, respectively. Calves in the trained group were loaded onto a livestock trailer for 5 successive days at weaning. Trainers led or hauled the calves by rope only. When loading was completed, trainers fed calves with sugar cubes as rewards. Calves in the control group were weaned without any treatment. Five weeks after weaning, tests were carried out under similar conditions as the trained group. Loading efficiency, physical effort on the handler and handling stress on calves between groups were compared. Trained calves were loaded significantly faster than control calves. Trained calves balked less during loading than control calves. Heart rates of handlers after loading were significantly lower in the trained group than in the control group; however, salivary amylase activity and cortisol concentration was not different between groups. Physical effort and stress on handlers would be almost the same in both groups. Heart rate, plasma cortisol, NEFA and CPK of calves were significantly increased only in the control group after loading. These results show calf training improves loading efficiency and reduces stress on calves.     

Effects of pasteurization of colostrum on subsequent serum lactoferrin concentration and neutrophil superoxide production in calves

OBJECTIVE: To determine the effects of pasteurization of colostrum on serum lactoferrin concentration and neutrophil oxidative function by comparing values from calves given pasteurized (76 C, 15 minutes) colostrum versus calves given fresh frozen colostrum. ANIMALS: 8 Holstein bull calves were used to study the effects of pasteurization of colostrum on the absorption of lactoferrin and neutrophil oxidative burst. Three additional calves were used to study the effect of exogenous lactoferrin on neutrophil oxidative burst. METHODS: Calves were fed fresh frozen or heat pasteurized colostrum (76 C for 15 minutes) via esophageal feeder within 4 hours of birth. Neutrophils were isolated from whole blood samples. Neutrophil oxidative burst was induced by phorbol ester (300 ng/ml) stimulation of cells (1 X 10(6) cells) at 37 C. Serum lactoferrin concentrations were compared, using immunoblot analysis. Serum IgG concentrations were determined by radial immunoassay. Comparisons were made between the use of the 2 types of colostrum in calves by measuring subsequent serum IgG and lactoferrin concentrations and neutrophil superoxide production. RESULTS: Serum IgG and lactoferrin concentrations increased more in calves receiving fresh frozen colostrum. Neutrophil superoxide production was higher in neutrophils prepared from calves receiving fresh frozen colostrum. Colostral lactoferrin addition to neutrophil incubations resulted in increased oxidative burst. CONCLUSIONS AND CLINICAL RELEVANCE: Compared with calves given fresh frozen colostrum, calves given pasteurized colostrum had decreased serum IgG and lactoferrin concentrations and neutrophil superoxide production 24 hours after administration. These results suggest that pasteurizing bovine colostrum at 76 C for 15 minutes has substantial effects on passive transfer of proteins and neutrophil function.     

Sternal recumbency or suspension by the hind legs immediately after delivery improves respiratory and metabolic adaptation to extra uterine life in newborn calves delivered by caesarean section

The aim of this study was to evaluate the effect of body positioning immediately after delivery on respiratory and metabolic adaptation to extra-uterine life in newborn calves. One hundred and one Belgian White and Blue calves were delivered at term by an elective caesarean section and were assigned into three categories according to the body position imposed immediately after umbilical cord rupture: 71 calves were placed in lateral recumbency; 16 calves were placed in sternal recumbency and 14 calves were suspended by the hind legs for less than 90 seconds (75 +/- 5 s). Following this initial body position, the calves were allowed to move without restraint. They were examined at birth, 5, 15, 30, 45 and 60 minutes, and 2, 3, 6, 12 and 24 hours after birth by the following measurements: physical examination, heart rate, arterial blood gas analysis, pulmonary function tests using the esophageal balloon catheter technique, arterial and venous blood acid-base balance analysis, rectal temperature, jugular venous blood sampling for the determination of blood glucose, plasma lactate and serum cortisol concentrations, haematologic variables and passive immune transfer variables. Body positioning immediately after delivery clearly influenced respiratory and metabolic adaptation to extra-uterine life in term calves delivered by an elective caesarean section. Systematic sternal recumbency and suspension by the hind legs for less than 90 seconds immediately after umbilical cord rupture had a positive functional impact on postnatal pulmonary mechanics and gas exchange and on postnatal correction of mixed acidosis present at birth, contributing in turn to an enhanced passive immune transfer. These two body positions should be encouraged to improve adaptation at birth in healthy term calves delivered by an elective caesarean section. Evaluation of possible side-effects is required before application in severely asphyxiated calves.     

The hematocrit values and hemoglobin content of blood and the level of total protein, free fatty acids, glucose, lactate, Ca, Mg, Na, K, Pa, Fe, Fe-binding capacity, Cu and Zn in the blood plasma of newborn calves and their mothers immediately after birth

Blood samples were collected from 22 calves in three weight classes (A: 29.3 +/- 0.5 kg, B: 36.0 +/- 1.3 kg, C: 42.7 +/- 3.7 kg), 1-3 minutes after parturition and prior to uptake of foremilk as well as 24-26 hours after parturition. Other blood samples were collected from 45 calves, 5-60 minutes after parturition, and from their mothers, 3-5 or 5-60 minutes from calving. Haematocrit and haemoglobin in the blood of the calves, immediately after parturition, were higher the values recorded from adult cattle. Major differences were found to exist between individual calves regarding total protein of blood plasma. Protein levels in 14 calves were below 50 g/l, within 24-26 hours from parturition. Free fatty acids in blood plasma of calves were lower than those in cattle, 1-3 minutes from parturition, and were higher, 5-60 minutes from parturition. Glucose levels in Group C were higher than those in A and B, 1-3 minutes postpartum. Lactate in Group C was higher than that in B. An account is also given of blood plasma levels of Ca, Pa, Mg, Na, K, Cu, and Zn.     

Granulocyte plasma membrane damage by leukotoxic supernatant from Pasteurella haemolytica A1 and protection by immune serum.

Bovine respiratory disease caused by Pasteurella haemolytica may be partially mediated by a leukotoxin secreted by the microorganism. We examined the effect of leukotoxic Pasteurella supernatants on leakage of the cytosol enzyme lactate dehydrogenase and the lysosomal enzyme arylsulfatase from bovine granulocytes. Lactate dehydrogenase release (94%) was much higher than arylsulfatase release (38%) over 30 minutes of incubation. The Pasteurella supernatants inhibited superoxide production by stimulated granulocytes at concentrations which also caused substantial cell death as measured by failure to exclude trypan blue. Both toxic effects were prevented by serum from aerosol-immunized calves, and protection appeared to be antibody-specific by comparison with fetal bovine serum or with serum absorbed against intact P. haemolytica. These findings suggest that the leukotoxin may selectively disrupt the granulocyte plasma membrane, and that antibody directed against a surface component of the microorganism is also capable of protecting against the leukotoxin effect.     

Suppressed lymphocyte blastogenic responses and enhanced in vitro growth of infectious bovine rhinotracheitis virus in stressed feeder calves

Hereford steers were stressed on a large-animal treadmill operating at speeds of 1.8 to 2.2 m/s. Blood samples were collected from indwelling jugular catheters before, during, and after exercise. Peripheral blood lymphocytes from stressed calves at 5 and 30 minutes after exercise had less (P less than 0.01) mitogen-induced blastogenic responses when compared to pre- or 60-minute postexercise values. Serum from stressed calves incorporated into lymphocyte cultures from nonstressed steers resulted in less (P less than 0.01) lymphocyte blastogenic responses. Infectious bovine rhinotracheitis viral growth in bovine kidney cell cultures was enhanced 4-fold when cultured with serum from stressed calves. These data indicate that acute physical exertion may cause physiologic alterations in calves that modulate cellular immunity and viral replication.     

Relationship of serum and plasma copper and ceruloplasmin concentrations of cattle and the effects of whole blood sample storage

The present study was conducted to determine whether plasma and serum copper and ceruloplasmin concentrations in cattle are different and whether transport of samples with storage on ice before centrifugation affects the measurements. Mean copper and ceruloplasmin values were higher in plasma than in serum. Linear regressions were plasma copper (microgram/ml) = 1.200 serum copper -0.032 (r2 = 0.99), serum ceruloplasmin (mg/dl) = 14.0 serum copper + 2.34 (r2 = 0.48), and plasma ceruloplasmin (mg/dl) = 18.2 plasma copper + 2.1 (r2 = 0.43). The percentage of copper associated with ceruloplasmin was less in serum (55%) than in plasma (66%). Storage of blood samples on ice for 3 days decreased serum copper value by 3.5%. Linear regressions to correct for storage effects were corrected serum copper = 1.11 stored serum copper -0.04 (r2 = 0.94) and corrected plasma copper = 1.22 stored plasma copper -0.17 (r2 = 0.86). A cuproprotein may be involved in the blood clotting process, and some ceruloplasmin and its copper are apparently trapped in the fibrin clot, causing less copper in serum, compared with that in plasma. The difference between plasma and serum copper concentrations of calves was slightly increased by dietary copper supplementation.     

Use of metamizole as an additional analgesic during umbilical surgery in calves

ObjectiveTo investigate the effect of metamizole on physiologic variables in calves undergoing surgical extirpation of the navel during anaesthesia using xylazine, ketamine and isoflurane.Study designDouble-blind, randomized trial.AnimalsA total of 26 calves.MethodsCalves with uncomplicated umbilical hernias and otherwise clinically healthy were randomly allocated to one of two groups: the control group (CG) and metamizole group (MG). All calves were administered meloxicam (0.5 mg kg^–1) intravenously (IV) 150 minutes before skin incision (SI). Animals were premedicated with xylazine (0.2 mg kg^–1) intramuscularly 50 minutes before SI. Anaesthesia was induced with ketamine (2 mg kg^–1) IV 30 minutes before SI and maintained with isoflurane in oxygen. MG calves were given metamizole (40 mg kg^–1) IV 60 minutes before SI. CG calves were administered an equivalent volume of saline. Heart rate (HR) and mean arterial blood pressure (MAP) were recorded from 5 minutes before SI until the end of anaesthesia (60 minutes after SI). Blood samples for determination of the plasma cortisol concentration (PCC) were drawn 60 minutes before SI and at 5, 30, 60, 150, and 510 minutes after SI.ResultsIn both groups, PCC increased during surgery and decreased after surgery. PCC was consistently lower in MG than in CG and was significantly (p = 0.0026) lower at 150 minutes after SI in the MG. Overall, the mean PCC in MG was 10.9 nmol L^–1 lower than that in CG (p = 0.01). In both groups, HR decreased during anaesthesia, whereas MAP increased, albeit with no statistically significant (p > 0.05) differences between groups.Conclusions and clinical relevanceOur study results suggest that a single preoperative dose of metamizole may have a positive impact on intra- and immediate postoperative analgesia by reducing PCC when used as an indicator of nociception.     

Changes in leukocyte populations in pulmonary lavage fluids of calves after inhalation of Pasteurella haemolytica

The distribution of leukocytes in bovine bronchoalveolar lavage fluids was determined in 15 calves at various times after aerosol exposure to Pasteurella haemolytica. For comparison, 10 calves were exposed to aerosols of phosphate-buffered saline solution; 15 calves, to Staphylococcus epidermidis; and 10 calves, to Salmonella typhimurium endotoxin. At 10 minutes after inhalation exposure for each group, the predominant cell type was the macrophage. Macrophages remained the predominant cell type throughout each lavage interval for calves exposed to phosphate-buffered saline solution and Staph epidermidis. For calves exposed to P haemolytica, there was a decrease in the percentage of macrophages detectable by 30 minutes after exposure, with a corresponding increase in the percentage of neutrophils. Sixty minutes after the inhalation exposure to P haemolytica, the percentages of macrophages and neutrophils in the lavage fluid were equal. By 240 minutes after exposure to P haemolytica, greater than 90% of the cells in the lavage fluids was neutrophils. The increase in the percentage of neutrophils in lavage fluids from calves exposed to S typhimurium endotoxin was similar to that seen for the calves exposed to P haemolytica.     

A study of the influence of the method of oral administration of ampicillin upon plasma drug levels in calves.

Ampicillin was given orally to five Holstein calves using the following four different methods of administration: via stomach tube, mixed and fed in the calf starter ration, dissolved in milk and pail fed and administered orally as 400 mg commercial calf tablets. In addition, ampicillin was dissolved in milk and pail fed 20 to 30 minutes following intramuscular atropine. Ampicillin was only detected in the plasma of calves which had received the drug, pail fed in milk. Atropine administration slowed the appearance of the drug in the plasma but did not decrease the efficiency of absorption.     

Sublingual administration of detomidine to calves prior to disbudding: a comparison with the intravenous route

ObjectiveTo study the effects of oromucosal detomidine gel administered sublingually to calves prior to disbudding, and to compare its efficacy with intravenously (IV) administered detomidine.Study designRandomised, prospective clinical study.AnimalsTwenty dairy calves aged 12.4 ± 4.4days (mean ± SD), weight 50.5 ± 9.0 kg.MethodsDetomidine at 80 μg kg^?1 was administered to ten calves sublingually (GEL) and at 30 μg kg^?1 to ten control calves IV (V. jugularis). Meloxicam (0.5 mg kg^?1) and local anaesthetic (lidocaine 3 mg kg^?1) were administered before heat cauterization of horn buds. Heart rate (HR), body temperature and clinical sedation were monitored over 240 minutes. Blood was collected from the V. cephalica during the same period for drug concentration analysis. Pharmacokinetic variables were calculated from the plasma detomidine concentration-time data using non-compartmental methods. Statistical analyses compared routes of administration by Student’s t-test and linear mixed models as relevant.ResultsThe maximum plasma detomidine concentration after GEL was 2.1 ± 1.2 ng mL^?1 (mean ±SD) and the time of maximum concentration was 66.0 ± 36.9 minutes. The bioavailability of detomidine was approximately 34% with GEL. Similar sedation scores were reached in both groups after administration of detomidine, but maximal sedation was reached earlier in the IV group (10 minutes) than in the GEL group (40 minutes). HR was lower after IV than GEL from 5 to 10 minutes after administration. All animals were adequately sedated, and we were able to administer local anaesthetic without resistance to all of the calves before disbudding.Conclusions and clinical relevanceOromucosally administered detomidine is an effective sedative agent for calves prior to disbudding.     

A comparison of the welfare and meat quality of veal calves slaughtered on the farm with those subjected to transportation and lairage

Comparisons were made of the behaviour, physiology and meat quality of 24-week-old male veal calves (mean live weight 234 kg) that were slaughtered either at the veal unit (the control group) or following 3 h transportation at space allowances of either 0.7 or 0.95 m² calf⁻¹ and 1.5 h of lairage. Compared with nontransported calves, transported calves had a greater heart rate during the journey and during lairage; greater plasma cortisol concentration immediately after the journey, but not after 1.5 h of lairage; and greater plasma creatine kinase activity immediately after the journey and after 1.5 h of lairage. The transported calves did not lie down during the journey. There was no effect of space allowance on either the heart rate or frequency of potentially traumatic events during the journey or the plasma cortisol concentration and plasma creatine kinase activity sampled after the journey. There was no effect of transport on either carcase bruising or muscle pH 24 h after slaughter. Although compared with ‘on-farm’ slaughter, the transport and handling associated with the journey to the slaughterhouse were stressful to the calves; there were no adverse effects of transport on either carcase bruising or meat quality.     

The route of liquids administered to calves by esophageal feeder.

An esophageal feeder and a rubber nasoesophageal tube were used to administer fluids to calves. Radio-opaque fluids were given and their destination determined by fluoroscopy and radiography. Fluids containing glucose and xylose were also given and plasma glucose and xylose concentrations measured. In at least 93% of calves, the radio-opaque fluids entered the reticulum, indicating that the reticular groove did not close. Oral administration of sodium bicarbonate, copper sulfate and guanidine HCl did not influence groove closure in calves that received fluids through an esophageal feeder. As administration of the fluids continued, overflow to the abomasum occurred after about 400 mL had been given. When 2.0 L of glucose and electrolyte solution was given by esophageal feeder, plasma glucose levels rose significantly (p less than 0.01), showing that absorption had occurred. Plasma xylose levels rose in seven out of eight calves 30 minutes after a second 2.0 L dose (containing xylose) had been administered. Thus, even though esophageal feeders do not cause reticular groove closure, they can be used to administer fluids for enteric absorption, provided large quantities are given.     

III. Comparison of Two Complement-Fixation Methods

The complement-fixation test used at Onderstepoort was compared with the method used at A.D.R.I. on infected calf and sheep sera. In the first method, the tests are incubated at 37°C for 90 minutes and the test sera are inactivated at 53°C; whereas in the A.D.R.I. method, the test sera are inactivated at 60°C for 30 minutes, incubation is at 9°C for 18 hours, and guinea-pig complement is supplemented with 5 per cent fresh, non-inactivated, normal calf serum. Serial serum samples from one of six experimentally infected calves were negative in the Onderstepoort test, three calves gave only trace reactions and two showed maximum titres of 1:10 whereas all six had maximum serum titres of 1:10 to 1:80 in the A.D.R.I. test. A good correlation was obtained, however, between the results of the two methods with the sera of experimentally inoculated sheep although titres 3 to 8 times higher were obtained with the A.D.R.I.'s test. Post inoculation bleedings from each sheep reacted in both tests.