Previous Page Table of Contents Next Page


TICK-BORNE DISEASES

Immunization of cattle
imported into Mozambique

R.M. Arnold and M. Asselbergs

Tick-borne diseases are the cause of probably the most economically serious losses of ruminants in southeastern Africa, where heartwater (Cowdria ruminantium) infection, Babesia bovis and B. bigemina infections and anaplasmosis are all endemic. Tick control is becoming increasingly difficult, for economic, administrative and biological reasons, and this group of diseases seems likely to assume greater importance with the planned expansion and intensification of the livestock industry. This article, which describes the immunization of some 400 head of imported, susceptible cattle against four of the most serious tick-borne diseases, will be of particular interest to readers in Africa.

Holding paddock for the Cuban bulls at the quarantine station

Although potential vectors such as Amblyomma maculatum and A. americanum occur in Cuba, the country is known to be free of Cowdria ruminantium infection (FAO, 1978). Babesia bovis and B. bigemina infections and anaplasmosis depend for their transmission and maintenance on Boophilus microplus and B. decoloratus ticks, control programmes for which have recently gained momentum, so that the extent of infection, although documented in a general sense in papers by Rivas et al. (1974), Rodrigues et al. (1977) and Rodrigues et al. (1978), is uncertain at the individual and national herd level.

The decision by Mozambique to import over 400 adult bulls from Cuba, therefore, called for careful consideration of the disease and parasite risks involved, especially as experience of previous importations such as that reported by Valadao (1969) suggested that a mortality rate of 50 percent could be expected. In the present instance, risks were especially high because the animals were adults, and could be expected to be affected clinically more severely than would calves.

Immunization facilities and procedures

The imported animals

Four hundred Brahman and seven Holstein bulls between 18 and 36 months of age arrived by boat from Cuba in mid-December 1978. A number of selected animals (destined for the ai Centre) had been reared under tick-free conditions in Cuba but most of the bulls originated from farms or ranches where tick control was said to be good and the Cuban authorities warned that most of the bulls could be expected to be susceptible to babesiosis, anaplasmosis and heartwater.

The seven Holsteins were taken to the ai Centre, near Maputo, as were four Brahmans of special merit. The remaining 396 Brahman bulls were divided between two quarantine camps, 205 being sent to the quarantine station at Belulane, some 20 km from Maputo, and the balance of 191 being kept on the boat and shipped to Nampula in the northern part of Mozambique.

Quarantine stations

At Cefria, the Centre for Artificial Insemination near Maputo, the 11 bulls were housed in individual concrete pens, fed cut grass and concentrates and exercised by walking round the roads of the Centre. They were hand-sprayed against ticks, but precautions were not sufficient completely to preclude exposure to tickborne disease.

At Belulane, the quarantine station in Maputo Province, approximately 50 ha of a former dairy farm had been double-fenced and divided into five paddocks of 8–10 ha. Each of these paddocks held a herd of 30– 50 head. There were three crushes (races) for handling the animals and a dip (of very poor design), but ticks were present on the grazings. The construction works were not completed during the quarantine period and a good deal of improvisation was required. Feeding was partly from grazing, supplemented by hay ad libitum and approximately 3 kg per head of a balanced concentrate ration fed daily. Dipping was carried out on a five-day cycle using an organophosphorus dip (Delnav) at the recommended strength.

At Nampula, in northern Mozambique, facilities were better and were ready before the cattle arrived. There were nine herds, each of 20–25 head. Apart from limited natural grazing, each animal received 30 kg of cut elephant grass and 7 kg of concentrate feed daily. Cattle were dipped at five-day intervals, although from the beginning the station appeared free of ticks.

Daily routine

This was essentially similar at both the Belulane and Nampula stations. Cattle were brought into small corrals at first light so that they could be put through the crush for clinical examination and temperature-taking early in the morning. Since the quarantine period was during the hottest months when ambient temperatures reached 38°C during the day, it was necessary to have temperature-taking completed by 09.00 if the temperature figures were to have any clinical value. At the same time, blood sampling was undertaken and treatments were given to the animals. Animals were only brought in during the afternoon if they were seen to be sick or if they required treatment.

TABLE 1. Vaccination schedule for imported bulls
Vaccination/ treatmentDays after arrival of cattleExpected reaction period (days after vaccination
Heartwater and lumpy skin disease9–17 (by herds)12–22
Babesiosis and anaplasmosis45 (Belulane)12–16 (babesiosis)
Babesiosis and anaplasmosis38 (Nampula)45–50 (anaplasmosis)
Not vaccinated(Cefria Al station)-
Vitamin A/D Complex50 (approx.)-
Blackleg and anthrax90–102-
Not vaccinated(Cefria Al station)-
Isometamidium (Samorin)113 (Nampula only)-


TABLE 2. Summary of reactions to heartwater vaccination
HerdPositive reactionDoubtful reactionsUncertainNon-reactionsTotal
Belulane875043121201
Cefria Al station920011
Nampula63181090190
TOTAL1597015221402
Percentage4017385-
1 Includes four head not vaccinated.

At Belulane, each herd was dipped every fifth day and was examined in the crush leading to the dip. On the day of dipping, the other four herds were examined in small wooden crushes constructed in the dividing fence between their paddocks so that unnecessary movement of the cattle was avoided.

At Nampula, a similar routine was followed, but more crushes were available and a less rigorous routine was possible.

At the Cefria ai station, temperatures were taken early in the day before the bulls were fed or taken out for exercise. At this station, evening temperatures were also recorded and the bulls were under closer observation than elsewhere.

Vaccination schedules

The same vaccines were used at all stations, except that at the Cefria ai station the bulls were not vaccinated against Babesia. Heartwater vaccine was the blood-based vaccine prepared from the Ball 3 strain of C. ruminantium at the Veterinary Research Institute, Onderstepoort. This was maintained on dry ice or liquid nitrogen and thawed out at the crush shortly before use. This vaccine was injected intravenously (5 ml, using an ear vein in the Brahman cattle), reactions being expected between 12 and 22 days later. Reactions resulting in a rise of body temperature to 40°C or over for two days were treated on the second day with oxytetracycline at the rate of 10 mg/kg body-weight, treatment generally being continued for three days or until the temperature fell. If an animal appeared clinically infected half the first dose was given intravenously; otherwise the intramuscular route was used.

Typical group of the bulls imported from Cuba

(Photo: A. Keim)

Because of the anticipated large labour requirements involved in dealing with many reactions at one time, it was decided to treat one herd of 43 bulls at Belulane and 109 animals at Nampula by the “block treatment” method, followed by a single dose of oxytetracycline at either Day 12 after vaccination or when 25 percent of the animals in the group had reacted. This block treatment made assessment of the immunity achieved difficult, as it subsequently proved impossible to decide whether individuals had in fact reacted to the vaccine and what the immune status of such animals was.

The Babesia vaccine was a bloodbased vaccine consisting of mixed B. bovis and B. bigemina vaccine strains, prepared at the Veterinary Research Institute, Onderstepoort. The dose used was 5 ml injected subcutaneously. This vaccine was received fresh from Onderstepoort and was used within two days of receipt.

Reactions were expected between 12 and 16 days after injection. During this period, when bulls had temperatures of 40.0°C or above, blood slides were made and examined the same day. Cases of babesiosis were treated with 44-diamidino-diazoaminobenzinedi-acetamidoacetate (Berenil) at the rate of 3.5 mg/kg.

Possible vaccination schedules against heartwater, anaplasmosis and babesiosis

The Anaplasma vaccine was the A. centrale strain also produced at the Veterinary Research Institute, Onderstepoort. This was received together with the Babesia vaccine and injected (2 ml subcutaneously) at the same time. Reactions were expected 45–50 days after inoculation but actually appeared after 30–60 days and were considered to occur when body temperatures exceeded 39.5°C. In such cases, or where clinical signs such as anaemia, icterus or constipation suggested the disease, blood samples were taken by caudal vein puncture using vacutainers, blood slides made and haematocrit values estimated. Animals with parasitaemias in excess of one percent infected erythrocytes, or with haematocrits below 20 percent, were treated with oxytetracycline at 8 mg/ kg body-weight each day for three days.

Vaccinations against other diseases and supportive treatments were carried out at convenient times during the quarantine period, as shown in Table 1.

During the quarantine period, faecal samples from all bulls were periodically examined for gastrointestinal parasite eggs and appropriate treatment was given. All bulls were treated with anthelmintics before being discharged at the end of the quarantine period.

Animals dying from whatever cause were carefully post-mortemed, special attention being paid to any evidence of exotic disease or parasites. All bulls were subjected to tests for semen quality, physical examination of the bulls' genitalia being made by veterinarians of the Insemination Service. Records were kept of morning temperatures, clinical symptoms and treatments given. A daily diary of conditions and work done was also maintained.

Assessment of reactions

Attempts were made to assess the success of the immunization against the tick-borne diseases with a view to advising generally on the practice, on any special precautions likely to be required when the bulls were later sent to the field and as a guide on the immunization of animals in the case of future importation.

In the case of heartwater, the only available measures of probable success before the animals are exposed to field challenge are evaluation of the temperature reactions to vaccination and the response to challenge. A temperature rising over two to three days during the expected reaction period to 40.0°C or over was accepted as evidence of a reaction. Typically the temperature should continue to rise above 41.0°C, but all animals were treated before this stage was reached. Challenge was carried out on a group of 42 head at Belulane that had appeared not to react to the initial injection. For this challenge, 10 ml (a double dose) of the Ball-3 strain vaccine were administered intravenously on Day 103 approximately after immunization. The reactions were controlled in the usual way for 30 days after challenge.

TABLE 3. Temperature reactions to heartwater vaccination, by days after vaccination
Days after vaccination7891011121314151617181920
No. bulls temperature taken12114918022437837130833536736729728212730
No. with temperatures > 39.4°C636247525058447068856443103
No. reacting bulls treated101952814165231424482961
1 Excludes bulls given block treatment - on Day 12 = 47; Day 16 = 71; Day 17 = 55; Day 18 = 5; Day 19 = 32. A total of 152 bulls were given block treatment.


TABLE 4. Reactions to Babesia and Anaplasma vaccinations
Centre1No. bulls with full recordsNo. with temps during expected reaction periodNo. bulls with positive slides within (and outside) expected reaction period
> 40.0> 39.5Babesia + Anaplasma -Babesia + Anaplasma +Babesia - Anaplasma +
Belulane200545924 (10)15 (0)8 (3)
Nampula19135101291
Total3918969251799
1 Bulls were not vaccinated against these diseases at Cefria Al station.

Assessment of reactions to babesiosis were made using the indirect immunofluorescence antibody test (ifat) and using antigens kindly supplied by the Veterinary Research Institute, Onderstepoort, and standard techniques. Unfortunately no pre-vaccination sera were available for comparison. Blood samples for the test were taken from seven bulls at the Cefria ai station (which had not been vaccinated against Babesia) and from 36 bulls at Belulane 26 days after vaccination, sera being tested against either or both B. bovis and B. bigemina antigens.

No tests were carried out for antibodies against anaplasmosis.

Acclimatization

It had been planned to vaccinate the bulls four days after arrival but this proved impossible as the animals were disturbed by their sea voyage and were unaccustomed to their new surroundings and methods of management. The difficulties with handling the bulls were aggravated, especially at Belulane, by the incomplete and unsatisfactory nature of the installations. Initially some animals escaped vaccination and some, usually the same animals, also missed temperature taking. Driving the animals for daily observation or for dipping also caused many to become excited. For these reasons, some of the temperatures recorded at the outset may be of doubtful significance or are incomplete.

The handling of the bulls continued to be difficult for up to six weeks at both main centres, although problems occurred to a lesser degree at Nampula. However, when a few Amblyomma hebraeum, Rhipicephalus spp. and Boophilus decoloratus ticks had been detected on the bulls, it was decided on Day 9, notwithstanding these disadvantages, to go ahead and start immunization. Ticks continued to be found on the bulls during the first five weeks of the quarantine period at Belulane in spite of a five-day dipping schedule, although in decreasing numbers. Ticks were not found on bulls at Nampula or the Cefria ai station.

Results of vaccinations

The reactions to the injection of the C. ruminantium vaccine are summarized in Table 2.

A clear-cut temperature reaction occurred in the “positive” reactors although it was difficult to interpret the temperature curve in the case of “doubtful” reactions, because of the influence of treatment or other factors. In retrospect, it is considered that most of these were “positive” reactors. The large “uncertain” group includes 147 bulls given “block treatment”, which prevents the full development of almost all reactions and makes interpretation impossible. The possible reactions of these bulls cannot be interpreted satisfactorily and therefore must be excluded from any analysis, thus leaving only 250 bulls for analysis. Of these, 159 (64 percent) gave clear positive temperature responses, 70 (28 percent) had “doubtful” responses, while 21 (8 percent) did not react.

The period elapsing between immunization and the temperature reaction is given in Table 3.

In an attempt to assess the protection given by the vaccine at Belulane, 21 animals that had not reacted to the first injection, plus 21 from the “uncertain” reactors that had been “block” treated, were re-vaccinated with a double dose of heartwater vaccine. During the 30 days' observation period, temperature reactions above 39.5°C were seen in only four animals. Two of these were attributed to ephemeral fever (which was occurring on the station at the time) and in the other two the cause of fever, which subsided without treatment, was undiagnosed. Blood slides showed no parasites and it is probable that these periods of above-normal temperatures represented mild reactions to the vaccine.

Reactions to the Babesia and Anaplasma vaccinations are summarized in Table 4.

The reactions to inoculations, related to days elapsed after vaccination, are given in Table 5, while the results of the ifat test for Babesia antibodies, related to reactions to vaccination, are given in Table 6.

TABLE 5. Reactions to Babesia and Anaplasma inoculations related to days after injection (both centres)
Babesia Days567891011121314151617181920>20       
Temp > 40.0°C8152019102562281091400       
Slides positive123661120201100010       
Anaplasma Days3030313233343536373839404142434445464748495051>51
Temp > 40.0°C0110911138991081010115365575421
Slides positive312255343331283611731052639
PVC1 <30101011202222151111100111
1 PVC = Packed volume cells.


TABLE 6. Results of IFAT test for Babesia antibodies, related to reactions to vaccination
Reaction to IFAT testNon-vaccinated animals at CefriaBulls reacting (temp. or slides) to vaccinationBulls not reacting to vaccinationTotals tested positive either B. bovis or B. bigemina
B. bovisB. bigemina
++05621
+-00321
-+00021
--170021
+NT05510
-NT00010
NT-01112
NT+02812
NOTE: + = strong or weak reaction to IFAT test;
- = negative reaction to IFAT test; NT = not tested.
1 All seven were negative to both antigens.,

Morbidity and mortality resulting from tick-borne disease

During the quarantine period, four animals died of drowning in the dip at Belulane, one was slaughtered after breaking a leg and one (which had escaped vaccination) died of naturally acquired heartwater. At the Cefria ai station, one Brahman bull, apparently successfully vaccinated, died 54 days later of heartwater. At Nampula, one bull died from a severe and uncontrolled reaction to heartwater vaccination, one from an encephalitis of undetermined origin and three from accidents resulting from sporadic diseases.

Following vaccination, the bulls were sent in batches for distribution over the major livestock areas of the country, starting four and a half months after their arrival. Attempts were made by means of verbal inquiries and departmental circulars to obtain information on the fate of the bulls, particularly in respect of tickborne disease. Although not all circulars were answered it was believed that all serious cases were reported.

A year after the bulls had been dispersed, two had been reported as having been successfully treated for B. bovis infections and three for anaplasmosis. No other cases of tickborne disease had been reported, although trypanosomiasis affected several bulls in Zambezia Province. Seven bulls had been recorded as dying from poisoning or accidents.

Conclusions

Judging by the negative reports of morbidity and mortality from the field in the 12 months after immunization, the bulls can be considered to have been protected against the four tick-borne diseases concerned. This view is supported by the results of the laboratory and challenge tests carried out before the bulls were dispersed.

Although two bulls died of heartwater during the quarantine period, the lack of response in the group of 42 bulls that were challenged and the complete lack of reports of the disease in the field for 12 months after immunization suggest that protection against C. ruminantium infection was extremely good.

Above: Collection of capillary blood from tail for preparation of thick and thin blood smears
Above left: Daily body temperature being measured early in the morning at the quarantine station
Far left: One of the dips used in the regular five-day dipping programme
Left: Intravenous inoculation of C. ruminantium (heartwater) vaccine

(Photos: A. Keim)

In the case of babesiosis, it appears from analysis of Tables 4 and 6 that the bulls were mostly already immune to both Babesia bovis and B. bigemina infections as a result of natural exposure before they left Cuba: only 10.9 percent of the bulls responded to the inoculation of the mixed Babesia vaccine by the manifestation of parasitaemias. All animals tested appeared to be immune at the end of the quarantine, but two breakdowns were later reported from the field. Exceptions are the animals taken to the Cefria ai station, which showed no evidence of previous Babesia infection, had not been vaccinated and all of which appeared susceptible (negative for antibody) in the ifat test. These animals, however, had all been reared indoors in Cuba under conditions of special tick control. In the case of anaplasmosis, the position is similar. Only 25.3 percent of the bulls appeared to respond to vaccination by significant parasitaemias.

It is probable that 90 percent of the bulls had experienced both babesiosis and anaplasmosis in Cuba and had maintained a level of humoral or cellular immunity. Since the strains of B. bovis (B. argentina) from South America and Mozambique are immunologically identical, it appears in retrospect that vaccination against babesiosis and anaplasmosis was only necessary in the case of a few selected individuals. This points to the value of testing for Babesia and Anaplasma antibodies of animals in future importations before proceeding with widespread vaccination involving an extended quarantine period.

The process of immunization against these four diseases, using present vaccines, is lengthy and tedious. Animals must be monitored for body temperature and general health and haematological studies carried out for almost four months. Alternative regimes to shorten this period are worthy of consideration. In some cases, testing for antibodies against babesiosis and anaplasmosis might indicate that immunization against these diseases was not justified. The practical feasibility and safety of alternative schedules to vaccination (set out in the figure) might be worth investigating. In the case of each of these schedules, it is important to bear in mind that good tick control must be maintained throughout and that vaccination should not be started until the animals have acclimatized and settled down in their new environment.

The first scheme calls for no special comment, except that if animals react severely to the Babesia vaccine, the additional stress of the reactions to the Anaplasma vaccine, which follows immediately after, might be excessive. In the second scheme, it would be impossible to assess the success of the vaccination against heartwater since the “blocking” method of controlling reactions is used. Vigilance over the animals would need to be especially strict for the 60 days of the observation period and, subsequently, after they had been sent to the field.

Other possibilities for reducing the present laborious procedure include the use of more attenuated and safer vaccines such as those now available in Australia against babesiosis. These have been produced and used in South America (FAO, 1975) and can be expected to protect against African strains of the organisms. However, vaccines with greater immunogenicity and safety are still required before vaccination against heartwater and anaplasmosis can be undertaken without the need for monitoring the animals after vaccination.

Adequate installations and handling facilities and a good system of management are essential in achieving successful immunization both because of the significance of body temperature response in assessing reactions to vaccination and because adequate nutrition is important in ensuring a satisfactory response and a short recovery period in vaccination against anaplasmosis (FAO, 1975).

Postscript

As a postscript to this account of the mass immunization of some 400 bulls imported from Cuba, the results obtained by one of the authors (Dr Asselbergs) two years later, in immunizing 220 Friesian heifers imported from South Africa against heartwater using the methods described in this article, are of interest.

Because of the recognized risks involved in vaccinating pregnant cattle, only non-pregnant heifers or heifers less than four months pregnant were vaccinated initially. These numbered 140 and 55 showed clear-cut reactions to the first vaccination. Three died of rickettsiosis and one, which died from other causes, was found on autopsy to have Cowdria bodies in brain smears. Of those which had not reacted clearly to the first vaccination, 74 were re-vaccinated twice at five-day intervals (Van de Merwe, 1979). Of these, one died of heartwater overnight, having been normal on the previous day's check. One was seriously ill with symptoms typical of heartwater (although it never showed fever), but recovered in two days. Seventeen animals that had shown no reaction before now showed clearly defined reactions, while six that had shown low-grade fevers on first vaccination again showed poor responses. Twenty animals that reacted first, with early or low fever, were now negative and, it was supposed, had been immunized by the first vaccination. It was assumed that the remaining 29, which did not react to either of the vaccinations, were either inherently resistant or had acquired immunity before importation. After immunization, the heifers were transferred from the quarantine farm (where no ticks had been seen during the quarantine period) to a dairy farm in the area where, in the herd of 150 cattle, seven had died and seven had been acutely ill with heartwater during the previous two months. This figure included one of the Cuban bulls from the Cefria herd that had been immunized two years before. It is planned to control ticks here only when they appear as a problem per se in order to maintain the immunity.

The vaccine used had been produced at INIVE, Maputo, using the Ball 3 strain and techniques developed at the Veterinary Research Station, Onderstepoort. Unfortunately supplies of Babesia and Anaplasma vaccine (which are not produced in Mozambique) were not available, but serological tests indicated that the majority of the animals possessed some antibodies to both Babesia bovis and Anaplasma, although one died of B. bovis infection and several were severely affected with anaplasmosis after their arrival.

References

FAO. 1975. Control of piroplasmosis and anaplasmosis in cattle - a practical manual, by P.J. McCosker. Rome.

FAO. 1978. FAO animal health yearbook. Rome.

Rivas, A., Rodrigues, O.N., Espaine, L. & Rodrigues, P. 1974. Aplicación simultánea de las reacciones de fijación de complemento y la aglutinación lenta en tubos para serodiagnóstico de la anaplasmosis en los bovinos. Rev. Cubano Ciencias Vet., 5: 1–9.

Rodrigues, O.N., Rodrigues, P., Rivas, A., Espaine, L. & Polarco, R. 1977. Evaluación y comparación de las pruebas de fijación del complemento para el diagnóstico de la babesiosis bovina. Rev. Cubano Ciencias Vet., 8: 1–7.

Rodrigues, O.N., Espaine, L., Rodrigues, P. & Rivas, A. 1978. Nuevos aspectos en la investigación serológica de la babesiosis y anaplasmosis bovinas, mediante microtécnicas de la fijación del complemento y aglutinación capilar. Rev. Cubano Ciencias Vet., 9: 87–94.

Valadao, F.G. 1969. Occurrence of hyperacute heartwater in Mozambique and problems of premunition against this disease. Annls Serv. Vet. Moçambique, 12–14: 85–90.

Van de Merwe, L. 1979. J. South African Vet. Ass., 50: 4.


Previous Page Top of Page Next Page