J.G. Grootenhuis
Wildlife Disease Research Project
National Veterinary Research Laboratory
Kenya Agricultural Research Institute
P.O. Kabete, Kenya
Most African wild Bovidae are carriers of Theileria parasites, for many of which the tick vector, host range and pathogenicity are unknown. Buffalo (Syncerus caffer) are known reservoirs of Theileria parva lawrencei and T. mutans and eland (Taurotragus oryx) of T. taurotragi, all playing an important role in the epidemiology of cattle theileriosis. In spite of the paucity of information on the involvement of wild animals in the epidemiology of cattle theileriosis, the exclusion of wildlife from cattle-grazing environments is advocated as a measure to reduce the risk of disease for cattle. Yet the buffalo, carrying T. p. lawrencei, is the only animal that is a proven risk to livestock. The exclusion of wildlife from livestock grazing areas is a drastic measure, which may cause losses in tourism and wildlife as well as ecological damage due, for example, to the loss of genetic resources. This paper discusses the role of wildlife in the epidemiology of theileriosis and immunization of cattle against T. p. lawrencei.
THEILERIA FROM BUFFALO
Theileria p. lawrencei, T. taurotragi, T. velifera and T. mutans have been isolated from buffalo and are transmissible to cattle. Of these, T. p. lawrencei is the only species highly pathogenic for cattle. The Wildlife Disease Research Project of the Kenya Agricultural Research Institute (KARI) has established breeding herds to breed and rear wild animals in captivity under tick-free conditions to study the role of wild animals in maintaining tick-borne parasites pathogenic to domestic livestock. A question frequently asked is: "Why do researchers work with wild animal hosts if the disease occurs in cattle and the parasites can be studied in domestic animals"? The answer lies in the origin and biology of these parasites.
Carrier state
The buffalo is an indigenous bovine of sub-Saharan Africa and has lived in harmony with T. parva and its vector Rhipicephalus appendiculatus since long before cattle were introduced into the region. It is assumed that when cattle were introduced into East Africa many died of theileriosis and that gradually a population of cattle evolved that was resistant or tolerant to this disease. In addition, a sub-population of the Theileria parasites transmitted from buffalo to cattle that we now refer to as T. p. parva may have become adapted to cattle. However, the major parasite pool is maintained by buffalo and when ticks from buffalo feed on cattle, the cattle are likely to become infected with T. p. lawrencei and develop Corridor disease.
In spite of the pandemic of rinderpest in Africa in the 1890s and subsequent rigorous control and culling, buffalo are widespread and commonly share habitats with domestic cattle. Surveys in Kenya, Tanzania and Uganda have shown that almost every buffalo sampled is a carrier of T. p. lawrencei (Young et al., 1978a). The carrier state of T. parva in cattle has also been well established (Maritim et al., this meeting). However, there are important differences between the carrier state in buffalo and cattle. First, it has been shown recently that not all of the antigenic types of T. parva in a stock isolated from a buffalo and used to immunize cattle were maintained in these cattle. Parasites isolated in ticks from the immunized carrier cattle were used to immunize a second group of cattle. Six out of seven of these cattle died on challenge with the original immunizing stabilate (Maritim et al., in press). Second, Theileria infection rates in ticks fed on carrier buffalo range from 10% to almost 100%, but infection rates in ticks fed on carrier cattle can be as low as 0.01% to 0.05% (Maritim et al., in press). Third, most cattle challenged with a lethal dose of T. p. lawrencei die before the piroplasm stage appears in the blood. Most cattle infected with T. p. parva die during a phase of high piroplasm parasitaemia, thus allowing ticks to pick up the parasite and transmit it to other cattle. Therefore, T. p. parva can be maintained by a cattle population, but T. p. lawrencei might not be. This parasite behaviour necessitates the study of T. p. lawrencei in buffalo.
Antigenic diversity
Antigenic diversity of T. p. lawrencei from buffalo has been demonstrated in several studies (Young et al., 1977, 1978b; Conrad et al., 1987, 1989; Grootenhuis et al., 1987a). The most striking example is the lack of cross-protection between two Theileria stocks isolated in ticks fed on the same buffalo 18 months apart and used to immunize and challenge cattle. Homologous challenge gave complete cross-protection, but some of the cattle immunized with the earlier isolate were susceptible to challenge with ticks infected later (Young et al., 1977, 1978b). Conrad et al. (1987) have shown that cell lines isolated from a single buffalo consist of a mixture of four or five antigenic types identified by monoclonal antibody staining patterns. An experimentally induced carrier state established in a buffalo following a single injection of stabilate resulted in a long-term carrier state from which isolates were made over a two-and-a-half-year period. Monoclonal antibody staining patterns differed among the cell lines isolated (Grootenhuis et al., 1987a). These antigenic differences may explain the lack of cross-protection among T. p. lawrencei types and between stocks of T. p. lawrencei and T. p. parva.
Nonetheless it has been possible to immunize effectively against natural T. p. lawrencei challenge, as shown by trials in the Trans-Mara, Nanyuki and Naivasha areas of Kenya (Young, 1985; Dolan, 1985; Young et al., in preparation). In these trials T. p. lawrencei isolates from the local areas were used to avoid introduction of new antigenic types and, more importantly, to immunize against the theilerial antigens of the areas. In the Trans-Mara trials, the T. p. lawrencei components were obtained from one buffalo. The success of this immunization suggests that it may be possible to obtain very broadly immunizing stock from one animal. However, the antigenic mixture that is required for this or any other area remains unknown and needs further study.
Transformation
Theileria p. lawrencei has been shown to change its behaviour following serial passage in cattle. After three to six passages it behaves like T. p. parva, producing a high schizont parasitosis and piroplasm parasitaemia. The initial passages require large numbers of cattle, since only a small proportion of animals infected with T. p. lawrencei from the buffalo will survive and develop a piroplasm parasitaemia. Several studies have described this transformation (Barnett and Brocklesby, 1966; Young and Purnell, 1973; A.C. Maritim, J.J. Mutugi and A.S. Young, personal communication) and the phenomenon was first attributed to a change in parasite behaviour. Recent studies have indicated that this process is not reversible because buffalo infected with the "transformed" parasite develop a transient infection and no carrier state (Grootenhuis et al., 1987a), similar to experimental T. p. parva infection in buffalo (Brocklesby, 1964). In addition, there is evidence for antigenic diversity of T. parva within individual buffalo as described earlier (Conrad et al., 1987; Grootenhuis et al., 1987b). These studies suggest selection from the diverse antigenic pool of parasites carried by the buffalo as an alternative explanation of T. p. lawrencei transformation.
If selection is what takes place, then buffalo contain other antigenic types that are not passaged in cattle to produce the transformed parasite. These other types may account for the high mortality in cattle with classical Corridor disease in which piroplasms are not detected because they die so rapidly. These virulent antigenic types may be essential in an immunizing parasite mixture.
Mechanisms of disease resistance
Buffalo and cattle are related closely phylogenetically, their immunoglobulins cross-react and they share the largest proportion of cell surface markers of all Bovidae (W.C. Davis, personal communication). Their cell mediated immune response to infection with T. parva appears to be similar to the response of cattle (Baldwin et al., 1986). Yet buffalo tolerate a T. parva challenge that is lethal to cattle (Brocklesby, 1964). Understanding the mechanism of resistance to theileriosis in the buffalo might provide new avenues for exploitation in the control of cattle theileriosis.
Isolation of parasites from buffalo
The arguments presented above are a strong case for the use of buffalo and their T. parva parasites. Three methods have been used to obtain buffalo-derived parasites for use in immunization.
a) Buffalo calves may be captured from the area where cattle are to be immunized. One- to three-month-old buffalo calves adapt readily to captivity and can be handled as easily as cattle. In our experience, every calf captured has been a carrier of T. p. lawrencei. At present we keep nine buffalo from a total of four areas of Kenya.b) A carrier state may be induced in Theileria-free buffalo born and reared at the laboratory with ticks that have fed as nymphs on wild buffalo in the selected area. Adult unengorged R. appendiculatus may be collected from buffalo habitats, or if the tick population density is low, sentinel cattle may be used to collect the ticks. We have infected two parasite-free buffalo with ticks collected from two buffalo habitats in areas where cattle need to be immunized against T. p. lawrencei and where buffalo calves could not be captured easily.
c) Adult unengorged R. appendiculatus may be collected from buffalo habitats and fed on rabbits for production of immunizing stabilates.
Methods a and b have the advantage that a tick feed can always be repeated. Although the antigenic composition of the parasite population from individual buffalo may change over time, experience to date indicates that good immunizing stabilates can be obtained repeatedly from the same buffalo. Tick collections from buffalo habitats are practical only as a "one off" exercise and only if there are high densities of both buffalo and ticks. Another problem encountered was that highly infected salivary glands were uninfective for cattle. These ticks could have been infected with a Theileria species from other wild Bovidae inhabiting the area and from a species that could not be transmitted to cattle (T.T. Dolan and D.A. Stagg, personal communication).
THEILERIA FROM ELAND
Theileria parasites initially thought to be mild strains of T. p. parva have been isolated in R. appendiculatus ticks and in cell culture from cattle in Kenya, Tanzania and Zimbabwe (Burridge et al., 1974; Uilenberg et al., 1977; Koch et al., 1988). These parasites cross-reacted serologically with T. taurotragi, a Theileria species isolated from eland (Taurotragus oryx) (Grootenhuis et al., 1981). Although T. taurotragi was named for its initial detection in eland (Martin and Brocklesby, 1960; Grootenhuis et al., 1979), it was found later to have a potentially wide host range (Stagg et al., 1983). Experimental infections have been successful in a captured buffalo calf while in vitro infection of lymphocytes has been successful in several other species of Bovidae.
Theileria taurotragi from eland causes a mild transient infection in cattle which is not readily detectable. Using an indirect fluorescent antibody (IFA) test, antibodies can be detected in both cattle and eland. Fatal T. taurotragi infection has been reported in eland, but infections in cattle are usually subclinical (Grootenhuis et al., 1979). The piroplasm and schizont stages in cattle are usually indistinguishable from those of T. parva. One isolate from cattle caused a very mild and transient infection in eland. This group of parasites was called T. taurotragi because of these results (Grootenhuis et al., 1981).
Although the reasons for placing these T. taurotragi-like isolates in the same group appear to be valid, there are behavioural differences between the cattle and the eland isolates. Eland isolates cause a very mild transient infection in cattle where schizont and piroplasm stages are difficult to detect. Conversely, it is difficult to infect eland with the cattle isolates. It appears that the T. taurotragi originating from wildlife may become adapted to cattle and that populations of these parasites are being maintained within cattle.
Serological tests to distinguish T. taurotragi from T. parva are unsatisfactory. Good positive control sera are not available and the IFA test is most reliable using piroplasm antigen. However, this stage generally does not occur at a sufficiently high parasitaemia to make suitable antigens. It would be preferable to develop a test making use of schizont antigen, which can be propagated in culture. Once better methods for the identification of this parasite are developed, the distribution of this Theileria species can be established. It is important to be able to recognize this parasite in the course of immunization trials, because vaccination against T. parva cannot prevent infection with T. taurotragi. Because of misdiagnosis, expensive drugs may be wasted when treatment is unnecessary. It is also important to be able to distinguish T. taurotragi from T. parva in ticks because these species are morphologically identical and therefore cause confusion in this epidemiological parameter.
THEILERIA FROM OTHER WILD BOVIDAE
Attempts have been made to transmit Theileria species detected in impala (Aepyceros melampus), wildebeest (Connochaetes taurinus) and waterbuck (Kobus defassa). Transmission from impala and wildebeest to cattle was unsuccessful, although the parasites could be transmitted between animals of the same species (Purnell et al., 1973; Grootenhuis et al., 1975). This is likely to be the situation with many of the wild Bovidae. The waterbuck can be infected with T. p. lawrencei (Stagg, Young and Grootenhuis, unpublished results), but its importance as a reservoir of this parasite remains to be established. The waterbuck also carries its own Theileria and Babesia piroplasms (Fawcett et al., 1987).
The genus Cytauxzoon, in which T. taurotragi was placed originally, appears to be pathogenic only for certain wild Bovidae and was not considered in this presentation.
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