Evaluation of age-dependent susceptibility in calves infected with two doses of Mycobacterium avium subspecies paratuberculosis using pathology and tissue culture

The longstanding assumption that calves of more than 6 months of age are more resistant to Mycobacterium avium subspecies paratuberculosis (MAP) infection has recently been challenged. In order to elucidate this, a challenge experiment was performed to evaluate age- and dose-dependent susceptibility to MAP infection in dairy calves. Fifty-six calves from MAP-negative dams were randomly allocated to 10 MAP challenge groups (5 animals per group) and a negative control group (6 calves). Calves were inoculated orally on 2 consecutive days at 5 ages: 2 weeks and 3, 6, 9 or 12 months. Within each age group 5 calves received either a high – or low – dose of 5 × 10⁹ CFU or 5 × 10⁷ CFU, respectively. All calves were euthanized at 17 months of age. Macroscopic and histological lesions were assessed and bacterial culture was done on numerous tissue samples. Within all 5 age groups, calves were successfully infected with either dose of MAP. Calves inoculated at < 6 months usually had more culture-positive tissue locations and higher histological lesion scores. Furthermore, those infected with a high dose had more severe scores for histologic and macroscopic lesions as well as more culture-positive tissue locations compared to calves infected with a low dose. In conclusion, calves to 1 year of age were susceptible to MAP infection and a high infection dose produced more severe lesions than a low dose.     

Interferon, Antibody Responses and Protection Induced by an Intranasal Infectious Bovine Rhinotracheitis Vaccine

The interferon and antibody response induced by an intranasal infectious bovine rhinotracheitis vaccine was followed in 22 calves over a nine month period and the ability of these vaccinated calves to withstand challenge with virulent infectious bovine rhinotracheitis virus was assessed.Interferon was detected two to three days post-vaccination and disappeared by the tenth day. Nasal and serum antibodies appeared by day 7 and persisted for nine months.The calves challenged three days postvaccination came down with disease typical of infectious bovine rhinotracheitis, whereas calves challenged three weeks, three months or nine months postvaccination resisted infection.     

Intestinal antibody response after vaccination and infection with rotavirus of calves fed colostrum with or without rotavirus antibody

The intestinal and systemic antibody response of calves vaccinated and/or challenged with rotavirus was studied employing isotype-specific ELISAs for the detection of IgG1, IgG2, IgM and IgA antibodies to rotavirus. Monoclonal antibodies to bovine immunoglobulin isotypes of proven specificity were used as conjugated or catching antibody. Five days after oral inoculation (dpi) of a 5-day-old gnotobiotic calf with rotavirus, IgM rotavirus antibodies were excreted in faeces, followed 5 days later by IgA rotavirus antibodies. The increase in IgM rotavirus antibody titre coincided with the inability to detect further rotavirus excretion. Faeces IgM and IgA rotavirus antibody titres fell to low levels within 3 weeks post infection. IgG1 and IgG2 rotavirus antibodies were not detected in faecal samples. In serum, antibodies to rotavirus of all four isotypes were detected, starting with IgM at 5 dpi. Two SPF-calves, which were fed colostrum free of rotavirus antibodies, were vaccinated with a modified live rotavirus vaccine and challenged with virulent rotavirus 6 days later. Upon vaccination, the calves showed an antibody response similar to the response of the infected gnotobiotic calf. Intestinal IgM rotavirus antibodies were excreted before or on the day of challenge and appeared to be associated with protection against challenge infection with virulent virus and rotavirus-induced diarrhoea. In 3 control calves, which were challenged only, the antibody patterns also resembled that of the gnotobiotic calf and again the appearance of IgM rotavirus antibodies coincided with the end of the rotavirus detection period. Two other groups of 3 SPF-calves were treated similarly, but the calves were fed colostrum with rotavirus antibodies during the first 48 h of life. These calves excreted passively acquired IgG1 and IgG2 rotavirus antibodies in their faeces from 2 to 6 days after birth. After vaccination, no IgM or IgA antibody activity in serum or faeces was detectable. Upon challenge, all calves developed diarrhoea and excreted rotavirus. Seven to 10 days after challenge low levels of IgM rotavirus antibody were detected for a short period. These data indicate that the intestinal antibody response of young calves to an enteric viral infection is associated with the excretion of IgM antibodies, immediately followed by IgA antibodies. This response is absent or diminished in calves with passively acquired specific antibodies which may explain the failure to induce a protective intestinal immune response by oral vaccination with modified live rotavirus of calves fed colostrum containing rotavirus antibodies.     

Studies on the interaction between an irradiated bovine lungworm vaccine and a morantel sustained release bolus

The interaction of the morantel sustained release bolus with the development of immunity in calves vaccinated with two doses of gamma irradiated (40 Kr) Dictyocaulus viviparus larvae was investigated under laboratory conditions. A total of 37 helminth-naive calves were used. Eight calves were used in the first part of the study to test the efficacy of a larval vaccine prepared by using gamma rays delivered from a cobalt source. In the second part of the study, four groups of four groups of four calves each were vaccinated and of these, all the animals in two groups each received a bolus. The remaining three groups (two groups of four and one group of five calves each) remained nonvaccinated with each animal in one group receiving a bolus. All the calves were challenged with approximately 2000 lungworm larvae four months postvaccination. In order to simulate possible field conditions, two of the vaccinated groups and two of the nonvaccinated groups were given a trickle infection of 800 lungworm larvae over a four-week period, three months prior to challenge. Based on a comparison of clinical signs, pathology and lungworm burdens at necropsy, the vaccination of the calves conferred a significant degree of protection (P less than 0.001) to a subsequent challenge compared with controls. The introduction of a morantel sustained release bolus and/or a trickle infection had no effect on the high degree of protection engendered by the vaccination. Nonvaccinated calves given a trickle infection, with or without a bolus, were also highly immune to challenge.     

Protection studies with a globin-enriched protein fraction of Ostertagia ostertagi

The protective capacity of an adult stage Ostertagia ostertagi globin antigen was tested in four vaccination experiments in cattle. In a preliminary experiment, calves were vaccinated three times intraperitoneally with 250 microg globin in Freund's adjuvant and challenged with a trickled infection of 25,000 infective larvae. In three subsequent field studies, calves were vaccinated twice or three times intramuscularly with 80-100 microg globin in Quil A and challenged with a natural gastrointestinal nematode infection on pasture. Higher globin-specific antibody levels were detected in the vaccinated calves than in the control animals in all vaccine trials. In the preliminary experiment, geometric mean cumulative egg counts in the globin group were reduced by 52% and total worm burdens were reduced by 28%, compared to the controls. In the first field trial cumulative faecal egg counts were reduced by 63% in the vaccinated calves. However, the reduction in faecal egg output in these two experiments was not statistically significant and no reduction in faecal egg counts was observed in the vaccinated animals in the two last field trials. In conclusion, vaccination of calves with O. ostertagi globin resulted in highly variable protection levels after challenge infection.     

Humoral immune response of the bovine fetus to in utero vaccination with attenuated bovine coronavirus

A fetal response to in utero vaccination with attenuated bovine coronavirus (9 to 49 days before parturition) was determined in 8 calves, 5 vaccinated and 3 controls. Calves were derived by hysterotomy before parturition and were maintained in a closed gnotobiotic environment. The IgA, IgM, and IgG values and coronavirus-neutralizing antibody titers were higher in the sera and intestinal loop fluid from vaccinated calves than in those from control calves. Sections of ileum and ileal lymph nodes from 1-day-old vaccinated calves, when stained with monospecific anti-bovine IgG, IgM, and IgA had numerous positively stained plasma cells. Positive fluorescence was not detected in comparable tissues from controls. When the 8 calves were given virulent coronavirus orally at 6 days of age, vaccinated calves did not become ill, whereas control calves had diarrhea in 19 to 22 hours. All calves were killed at 10 days of age. Control calves had lesions characteristic of coronavirus infection, and intestinal epithelial cells were positive by fluorescent antibody tests. In vaccinated calves, lesions of coronavirus infection were absent, and results of fluorescent antibody tests were negative. Although in utero vaccination with a coronavirus vaccine stimulated immunity in the newborn calf, the frequency of abortions (2 of 14 cows inoculated intra-amniotically) and premature births (4 of 14) precluded practical application.     

Effect of parainfluenza-3 virus challenge on cell-mediated immune function in parainfluenza-3 vaccinated and non-vaccinated calves

A group of four conventional, colostrum-fed calves was vaccinated with live parainfluenza type 3 (PI-3) virus vaccine at 1 and 5 weeks of age. A group of four control calves was treated with cell culture medium at the same time. Two weeks after the second vaccination, both groups of calves were challenged with PI-3 virus by a combined respiratory route. Blood and nasal mucus samples were collected at intervals, and alveolar macrophages were recovered before and after challenge by bronchoalveolar lavage. The results demonstrated that clearance of virus, as indicated by presence of virus antigen was more rapid in previously vaccinated calves. Several alveolar macrophage functions were markedly reduced in all calves 5 to 7 days following virus challenge, although microbicidal activity was unaffected, compared to the controls. The production of neutrophil chemotactic factors by alveolar macrophages occurred more rapidly after virus challenge in the previously vaccinated calves and this correlated with a more rapid neutrophil influx into the lungs in these animals.     

Pathological changes in the small intestine of neonatal calves naturally infected with reo-like virus (rotavirus).

Nine calves, of which six had been challenged with an enteropathogenic strain of Escherichia coli, were found to be naturally infected with rotavirus. Rotavirus was recovered from the faeces of six calves and rotavirus antigen was detected in the intestinal mucosa of all calves. Stunting and fusion of villi were seen principally in the proximal and middle small intestine, where rotavirus antigen was detected by immunofluorescence. Typical lesions of enteric colibacillosis were found in the distal ileum of the challenged calves, associated with adhesion of the challenge strain of E coli to the mucosa. All samples were removed from the intestine under general anaesthesia and denudation of villi was not observed. However, following exsanguination and resampling at the same site in the small intestine of one calf, denudation was a constant feature.     

Immunological relationship between strains of Theileria annulata Dschunkowsky and Luhs 1904

The immunological relationship between the Ludhiana and Hissar strains, and between the Uruli-Kanchan, Bangalore and Jaipur strains of Theileria annulata was studied. Calves were immunised against the Ludhiana strain by the infection-treatment method, against the Uruli-Kanchan strain by the infection-treatment method or by untreated low-grade infection and against the Jaipur strain by untreated low-grade infection. After 45 days, groups of immune calves and susceptible controls were challenged with 10-tick equivalent stabilates. The ensuing host responses, body temperature, swelling of the regional lymph node, appearance of schizonts in the regional lymph node and piroplasms in blood and mortality, were studied. All five strains proved very virulent and produced severe disease in susceptible calves. Immunisation with the Ludhiana, Uruli-Kanchan and Jaipur strains conferred absolute protection against severe homologous challenges. Immunisation with the Ludhiana and Uruli-Kanchan strains conferred substantial protection against heterologous challenges with the Hissar strain, and the Bangalore and Jaipur strains, respectively. Immunisation with the Uruli-Kanchan strain gave less protection against heterologous challenge with the Jaipur strain and vice versa as some of the challenged calves showed slight swelling of the regional lymph node, two had fever, all exhibited a low-grade parasitaemia and four developed anaemia.     

β-Glucan plus ascorbic acid in neonatal calves modulates immune functions with and without Salmonella enterica serovar Dublin

To determine if β-glucan plus ascorbic acid affects adherence and pathogenicity of Salmonella Dublin and innate immune response in neonatal calves, 20 calves were fed control or supplemented diets (β-glucan, 0.9 g/d, plus ascorbic acid, 500 mg/d) until d 23. On d 21, 5 calves per treatment received 2.4 × 10(8)CFU of S. Dublin orally. S. Dublin spread through intestinal tissues into mesenteric lymph nodes (MLN), spleen, and lung tissues within 48 h. All supplemented calves had less mRNA expression of IL-1 receptor antagonist in liver. Leukocyte cell surface markers changed in lung cells, but not in blood, MLN, or spleen. CD14 in lungs was greatest for calves receiving supplement and challenge, but CD18 in lungs was greater for challenged than control calves. Lung DEC205 was greatest for challenged calves with and without supplement compared to controls, but more lung cells expressed CD14 for all treated groups compared to controls. These data show that S. Dublin briefly inhabited the intestinal tract, moving quickly to spleen, MLN, and lung tissues. Lung tissue was modulated by S. Dublin, but supplement alone increased CD14 expressing cells. The supplement appears not to attenuate invasiness but modified some lung cell populations by 48h.     

Influence of recombinant bovine interferon gamma and dexamethasone on pneumonia attributable to Haemophilus somnus in calves

The influence of recombinant bovine interferon gamma (rBoIFN-gamma) treatment on resistance of clinically normal and dexamethasone-treated calves to Haemophilus somnus infection was evaluated. Four groups of 6 calves each were treated with saline solution (controls), dexamethasone (0.04 mg/kg of body weight/for 3 days), rBoIFN-gamma (2 micrograms/kg for 2 days), or dexamethasone and rBoIFN-gamma (aforementioned dosages). All treatments were started 24 hours before intrabronchial challenge exposure with 5 x 10(9) colony-forming units of H somnus. Rectal temperature and WBC count were monitored daily. Two of the dexamethasone-treated calves died of pneumonia 4 days after challenge exposure and were necropsied. All other calves were euthanatized and necropsied 7 days after challenge exposure. All calves had pneumonia of variable intensity. Dexamethasone-treated calves had increased volume of pneumonic lung (P less than 0.05) and increased severity of pneumonia, compared with control calves. Recombinant bovine interferon gamma treatment resulted in reduction in pneumonic lung volume and severity of pneumonia in dexamethasone-treated calves (P less than 0.05), although it did not influence severity of pneumonia in nondexamethasone-treated calves.     

Immunity of ivermectin treated cattle to challenge from helminth parasites in the following season

Two groups of yearling cattle which had been treated with ivermectin either three and eight, or three, eight and 13 weeks after turn out to trichostrongyle contaminated pasture in their first grazing season were exposed in the following season to natural challenge with helminth parasites. To assess their immunity to this challenge each group shared a pasture with parasite naive first season calves. No anthelmintic treatments were administered at any time during the year. Throughout the grazing period the yearlings showed normal respiratory rates, negative faecal lungworm larval counts, and, relative to the calves, low faecal trichostrongyle egg counts. All the first season calves developed patent lungworm infections and on one occasion the mean respiratory rates of each group of calves were significantly greater than those of the yearling cattle. At the end of the grazing period, from early May until late September or October 1986, the cattle were removed from pasture and together with parasite naive controls challenged with either 10 or 22 third stage larvae of Dictyocaulus viviparus/kg bodyweight and necropsied between 18 and 23 days later. Although the experimental challenge resulted in relatively heavy lungworm infection of the naive controls, none of the yearlings and only three of the 11 calves which had been at pasture were found to be infected. However, large numbers of arrested fourth stage larvae of Ostertagia ostertagi were present in all the naturally infected yearlings and calves.     

Intrauterine infection with bovine virus diarrhoea virus on alpine communal pastures in Switzerland

This study involved 13 calves, one to 50 days of age, born to first calf heifers that had been pastured on one of seven alpine communal pastures in the canton of St. Gallen during the summer of 1995. Of a total of 993 cattle pastured, 61 were pregnant heifers that were negative for bovine virus diarrhoea (BVD) antigen and for BVD antibodies at the start of pasturing. Seroconversion occurred in 26 of these pregnant heifers during the pasture period. Blood samples and skin biopsy specimens of calves born to 13 of these were examined for BVD antigen by antigen-ELISA and by an immunohistochemical technique, respectively. Blood samples were positive for BVD antigen in four calves, questionable in one calf, negative in seven and missing in one prematurely born calf. The four calves that were positive for BVD antigen in the blood were also positive in skin biopsies. Of the seven calves with a negative or missing blood test, six had positive and two had negative skin samples. Based on the combined results of blood and skin testing, 11 of 13 calves were positive for BVD antigen. Of the 11 infected calves, six were normal at birth, four were smaller than normal and one was premature and weak and was euthanized on humane grounds. Of the four small calves, two developed diarrhoea and died within the first month of life. The two calves that were negative for BVD antigen were clinically normal. The results of this study not only demonstrate the occurrence of in-utero infection with BVD virus, but also stress the importance of alpine communal pasturing in the spread of BVD virus. Because the prevention of infection with BVD virus on communal pastures does not seem feasible, it is recommended that all calves born to cows from such pastures be tested for BVD antigen.     

Protection from parasite-induced weight loss by the vaccination of calves with excretory-secretory products of larval Oesophagostomum radiatum

Excretory-secretory products (ESP) were collected from in vitro maintained Oesophagostomum radiatum larvae during the period in which the larvae molt from the third to fourth larval stage. The ESP were used to immunize 13 uninfected calves which were subsequently challenged with 1.7 X 10(4) infective O. radiatum larvae. Worm recoveries from immunized calves were reduced 23% compared to 12 unimmunized controls, while the number of intestinal nodules was 72% greater compared to unimmunized controls; however, neither difference was statistically significant. Immunized calves had enhanced serum IgG and IgA anti-ESP antibody responses upon challenge. No differences in serum IgG2 or IgM antibody or cellular immune responsiveness, as determined by in vitro antigen-induced lymphocyte proliferation, were seen. Vaccination with ESP did significantly protect calves from the weight loss seen in non-immunized calves. Unimmunized calves ceased gaining weight approximately 5 weeks after challenge and by 10 weeks after challenge had lost an average of 4 lbs. per calf. Over the same time interval (i.e., 5-10 weeks after challenge), the immunized calves gained an average of 23 lbs. per calf. These results strongly suggest that vaccination with ESP conferred an advantage to calves that is not correlated solely with the number of worms developing from challenge infection.     

Theileria parva: the immune status of calves born of dams immunised against East Coast fever

Eight Friesian cross cows three months pregnant to a single Friesian bull were immunised against East Coast fever by infection with Theileria parva (Muguga) sporozoite stabilate and treatment with pyrrolidino-methyl tetracycline. They were challenged with the homologous stock four times before calving and a fifth time after calving, and resisted all five challenges which killed all of the five groups of five susceptible controls. Calves born to these hyperimmunised dams were fully susceptible on challenge with the same stabilate, as were susceptible cows from the same farm and their calves. In both instances the calves died three to seven days earlier than the cows which were approximately 10 times heavier. These results show that one- to two-month-old taurine calves from artificially immunised dams are not protected from experimental T parva sporozoite challenge and that there is no inherent calfhood resistance to East Coast fever.     

Immune response to and faecal shedding of Mycobacterium avium ssp. paratuberculosis in young dairy calves, and the association between test results in the calves and the infection status of their dams

The objectives of this study were to: (i) evaluate the results of an intradermal skin test, a modified IFN-gamma test, and a commercial ELISA in commercially raised dairy calves at 2, 4, 6 and 8 months of age relative to faecal shedding of Mycobacterium avium ssp. paratuberculosis (MAP); (ii) determine the proportion of 8-month-old calves shedding MAP in faeces as detected by culture and One Tube Nested Polymerase Chain Reaction (OTSN-PCR) and (iii) explore the association between results of tests described above in the calves and the Paratuberculosis (PTB) status of their dams as determined by faecal culture and/or serology. The study calves belonged to two dairy herds with different risk of exposure to MAP (high and low) and were enrolled based on their dam's ELISA results prior to calving. Approximately 3% of the calves were shedding MAP in faeces at 8 months of age. No agreement was observed among the evaluated immunity-based tests or between the immunity-based tests and the detection of MAP in faeces. Although no association was observed between the infection status of the dam and the results from the IFN-gamma and skin tests on the calves, there is an indication that calves born from dams that were faecal shedders might be at a higher risk of testing positive to the IFN-gamma test at 8 months of age. The disagreement among all tests evaluated in this calf cohort suggests that the detection of MAP infection in young stock requires the use of combined multiple tests. The early detection of PTB in calves is a challenge that requires further exploration of new methods to confirm infection status. These new testing methods should be both affordable and compatible with regular husbandry practices.     

Vaccine potential of recombinant antigens of Theileria annulata and Hyalomma anatolicum anatolicum against vector and parasite

In an attempt to develop vaccine against Hyalomma anatolicum anatolicum and Theileria annulata, three antigens were expressed in prokaryotic expression system and protective potentiality of the antigens was evaluated in cross bred calves. Two groups (grs. 1 and 4) of male cross-bred (Bos indicus × Bos taurus) calves were immunized with rHaa86, a Bm86 ortholog of H. a. anatolicum, while one group of calves (gr. 2) were immunized with cocktails of two antigens viz., surface antigens of T. annulata (rSPAG1, rTaSP). One group each was kept as negative controls (grs. 3 and 5). The animals of groups 1, 2 and 3 were challenged with T. annulata infected H. a. anatolicum adults while the animals of groups 1, 3, 4 and 5 were challenged with uninfected adult ticks. A significantly high (p<0.05) antibody responses to all the three antigens were detected in immunized calves, but the immune response was comparatively higher with rHaa86 followed by rTaSP and rSPAG1. Upon challenge with T. annulata infected ticks, animals of all groups showed symptoms of the disease but there was 50% survival of calves of group 1 while all non immunized control calves (group 3) and rSPAG1+rTaSP immunized calves died. The rHaa86 antigen was found efficacious to protect calves against more than 71.4-75.5% of the challenge infestation. The experiment has given a significant clue towards the development of rHaa86 based vaccine against both H. a. anatolicum and T. annulata.     

Protection against bovine respiratory syncytial virus challenge following a single dose of vaccine in young calves with maternal antibody

Twenty-one young calves with maternally derived antibody to bovine respiratory syncytial virus (BRSV) were divided into three groups of seven, each group balanced for BRSV antibody titre. The calves had no evidence of previous exposure to BRSV. The calves in one group were given a single dose of a monovalent modified live BRSV vaccine; the calves in the second group were given a single dose of an inactivated combined BRSV, parainfluenza virus type 3, Mannheimia haemolytica vaccine and the calves in the third group were left as unvaccinated controls. Three weeks after the single doses of vaccine, all the calves were challenged with BRSV. The clinical signs of disease were mild, and virus excretion was limited to two calves in the group given the inactivated vaccine, compared with six in the negative controls (P = 0.05) and five in the group given the live vaccine. The mean virus excretion titres after the challenge were not significantly different between the groups. There was little seroconversion before the challenge, but six of the seven calves in the group given the inactivated vaccine showed significant seroconversion within two weeks after the challenge, compared with only one calf in each of the other two groups (P = 0.015).     

Immune response to Neospora caninum native antigens formulated with immune stimulating complexes in calves

The aim of this study was to compare the immune responses to live Neospora caninum tachyzoites and N. caninum native antigens formulated with immune stimulating complexes matrix (ISCOM-matrix) in calves. Fifteen calves were used in this study: 3 were intravenously inoculated with 1 × 10(8) live tachyzoites (Group A), 3 were inoculated twice with N. caninum native antigens formulated with ISCOMs (Group B); 3 with N. caninum native antigens in phosphate-buffered saline (PBS) (Group C); 3 received ISCOM-matrix (ISCOMs without antigen) (Group D) and 3 were negative controls receiving PBS (Group E). The last four groups were inoculated subcutaneously. The specific total IgG and its subtypes were analyzed by an indirect enzyme-linked immunosorbent assays (ELISAs) and by Western blot. IFN-γ levels in plasma was quantified using a commercial kit. All calves were challenged intravenously with 1 × 10(8) live tachyzoites at week 11 after receiving the first dose. Parasitemia was assessed in plasma samples by semi-nested PCR. Neospora-specific antibodies were detected in animals from Groups A and B in the week 2 after inoculation. The ELISA OD values were higher in Group B compared with Group A from weeks 6 to 11 (P<0.05). Analysis of the subisotype specific antibodies in experimentally infected calves revealed a predominant IgG(2) response; however, a predominant IgG(1) response was observed in animals inoculated with N. caninum native antigens formulated with ISCOM-matrix. Control calves remained seronegative until challenge infection. The pattern of bands by Western blot was similar when testing sera from animals in Groups A and B. The levels of IFN-γ production after respective immunization schedules were similar between Groups A and B. Neospora-DNA was detected in plasma samples shortly after intravenous challenge in calves from all groups including those receiving the experimental vaccine formulation. The duration of the parasitemia was similar in all groups.     

The immunization of calves against heartwater: subsequent immunity both in the absence and presence of natural tick challenge

Cattle, vaccinated as calves with Cowdria ruminantium-infected tick stabilate, were challenged 6, 12 and 24 months later. In the absence of tick challenge, vaccination of calves induced a partial immunity against subsequent challenge at 12 and 24 months. In animals exposed to ticks, the resistance was no better than that of control, unvaccinated cattle. When they were challenged at 6 months of age there was no difference between vaccinated and unvaccinated calves, either in the absence or presence of tick challenge, and all the animals manifested a high degree of natural resistance. This study therefore suggests that the value of vaccinating Afrikander-cross calves in heartwater endemic areas should be further investigated. The indirect fluorescent antibody (IFA) test proved to be a valuable means of monitoring the serological response of vaccinated animals and detecting the sero-conversion of animals exposed to tick infection. On one hand, there was good correlation between the febrile reaction and the results of the IFA test on the sera of vaccinated and control cattle challenged with the heartwater agent, in that all sero-positive animals were resistant to challenge. On the other hand, though, a considerable percentage of the animals that were serologically negative were also resistant to challenge.