Trypanotolerance – a review

M. Murray, W.L Morrison, P.K. Murray, D.3. Clifford and J.C.M. Trail

It is a wellaccepted epidemiological fact that many West African breeds of cattle are able to survive without the aid of chemotherapy in areas of tsetse fly challenge where the humped zebu cannot (Stewart, 1951). In the same way, it is recognized that some breeds of sheep and goats (E.W. AIlonby, personal communication), many species of wild life (Ashcroft, Burtt and Fairbairn, 1959) and certain inbred strains of mice (Morrison et al., 1978), also exhibit increased resistance to trypanosomiasis. This phenomenon is known as trypanotolerance although in immunological terms this is a mis­nomer since trypanotolerant animals do become infected with trypanosomes with adverse effects and they do re­spond immunologically. Thus, while the term trypanotolerance is now widely accepted, it should be under­stood to mean no more than reduced susceptibility.

The lack of vaccine and the limi­tations of the present methods of control, namely chemotherapy and tsetse control, have stimulated the desire to develop additional approaches that might allow more efficient land utilization in the vast areas of Africa dominated by the tsetse fly. Thus, there is now considerable interest in the potential use of trypanotolerant livestock. Particular attention has been focussed on the N'Dama breed of cattle because it has a relatively fixed phenotype and can be improved in productivity. Furthermore, pub­lished information has consistently confirmed the greater resistance to trypanosomiasis of the N'Dama over the zebu (Chandler, 1952, 1958; Desowitz, 1959; Stephen, 1966; Roberts and Gray, 1972). However, precise comparative information cannot always be obtained from these publications because in some cases the animals under study had previously been ex­posed to trypanosomiasis and in others their disease history was not known. In addition group numbers were fre­quently small, the groups were of widely different ages and of mixed sex. Moreover, many of these inves­tigations were conducted on experi­mental stations where the general husbandry and feeding were of a high standard; as nutrition has a profound effect on the outcome of the disease (Mortelmans and Kageruka, 1976), it is likely that cattle kept on experi­mental stations will be more resistant to infection than cattle in natural field conditions.

Figure 1. Zebu. The preinoculation packed red blood cell volume of the zebu in the described experiments was 35 +4 (standard deviation)

Figure 2. N'Dama. The preinoculation packed red cell volume of the N'Dama in the described experiments was 35 +4 (standard deviation).

Thus, most of the basic questions about trypanotolerance remain to be answered precisely, questions such as, to what extent is inheritance important and how is it influenced by environ­mental factors; what are the mecha­nisms underlying increased resistance; and how productive are these breeds under various levels of challenge and in different management and ecological situations. The present article at­tempts to evaluate the current state of knowledge on these questions, lean­ing heavily on the authors' own ex­perience with cattle and inbred strains of mice.

Trypanotolerance in N'Dama cattle.

Over the past few years, a series of experiments on N'Dama and zebu cattle in The Gambia, West Africa, have been carried out. The objectives of this work were, first, to confirm the existence of trypanotolerance in N'Dama and zebu cattle that had no Previous experience of trypanosomiasis under different types of challenge, second, to compare the progress of the disease and its immunology in the hope of defining the underlying mecha­nism of trypanotolerance and, lastly, to evaluate the impact on productivity of different levels of tsetse challenge in N'Dama cattle living under natural field conditions. Preliminary results of these studies have been published by Murray et al. (1977 b, c, d and e). A complete report is being prepared by Dr P.K. Murray.

Perhaps the most important aspect of this experimental work is the back­ground of the cattle used. The zebu cattle (Figure 1) were from northern Senegal, purchased from a ranch well beyond the northern limits of the tsetse fly belt. N'Dama cattle (Figure 2) were obtained from the Govern­ment Agricultural Experimental Sta­tion at Yundum, The Gambia, a lo­cation considered tsetsefree, although surrounded by areas infested with Glossina palpalis and G. morsitans submorsitans. All animals studied were clinically, parasitologically and serologically negative for trypanosomiasis. There are divisions of opinion on what constitutes an N'Dama and, unfor­tunately, there is no genetic definition. The N'Dama used in this work con­formed closely to the accepted phenotype in this area of West Africa. Furthermore, blood analysis by starchgel electrophoresis showed a haemo­globin frequency (0.89) similar to that recorded for N'Dama in other parts of Africa.

Figure 3. Experimental design. B = time of Trypanosoma brucei challenge; C = time of T. congolense challenge; T—T =period when the cattle were exposed to Glossina palpalis.

Figure 4. Zebu suffering from the effects of inoculation with Trypanosoma congotense.

The design of the first experiment carried out is shown in Figure 3. All animals used were aged three to four years. The 60 zebu were male, where­as of the 61 N'Dama, 34 were female and 27 were male; 8 of these zebu and 9 N'Dama were kept as unchal­lenged controls. In order to simulate field conditions of nutrition and exer­cise, the cattle were maintained under such conditions in an area initially thought to be tsetsefree; tsetse traps were set in order to keep the area under surveillance. During the day the cattle grazed extensively, walking up to approximately 27 km and they were tethered at night. No supple­mentary feed was given.

With the onset of the rainy season in August 1976, the herd came under challenge from G. palpalis and this lasted until November. A daily mean of 0.17 flies of both sexes was caught per Malaise trap (W.F. Snow, personal communication). It was estimated that this tsetse challenge commenced 15S days after the needle challenge with Trypanosoma brucei and 59 days after inoculation with T. congolense. During this time several animals be­came infected with T. vivax.

All 52 inoculated zebu and 52 N'Dama became infected and ill — as judged by deterioration of body con­dition and the development of anaemia (Figure 4). During the course of the experiment, 12 infected zebu and 15 infected N'Dama were killed for histopathological examination. In the remainder, clear differences in suscep­tibility emerged between the two breeds. Of the 37 N'Dama allowed to survive, none died, whereas 30 of the 40 zebu died of ?trypanosomiasis; 9 died after needle challenge (3 infected with T. congolense), while the other 21 died during or after the fly challenge. These provided the additional histopathological material.

This result was reflected in clear differences in the degree of anaemia that developed in the two breeds. Whereas in both N'Dama and zebu, the onset of the anaemia occurred within a few days of inoculation and was associated with appearance of parasites in the blood, the rate of de­velopment and severity of the anaemia was significantly greater in the zebu. This was true for both T. congolense and for T. brucei, although the severity of the anaemia and the extent of the difference between breeds was much greater in T. congolenseinfected ani­mals. Nevertheless, it must be pointed out that the isolate of T. brucei em­ployed was pathogenic for both N'Dama and zebu and 3 zebu died as a direct result. It was also obvious that N'Dama and zebu previously infected with T. brucei were equally susceptible to inoculation with T. con­golense, although as before the anae­mia became more severe in the zebu (Figure 5).

Figure 5. Average parasitaemia score re­corded in N'Dama and zebu inoculated with 10^s Trypanosoma brucei followed by 1(P T. congolense 96 days later. Details of this scoring system have been described by Murray et al. (1977a). The level of the first peak of parasitaemia plus one standard deviation is shown. The percentage fall in packed red cell volume (pcv) is demonstrated and zebu mor­talities are given. The hatched areas of the histogram represent the N'Dama while the open areas represent zebu. Elsewhere N'Dama = •; Zebu = o.

Figure 6. Average parasitaemia score re­corded in N'Dama and zebu inoculated with 10⁸ Trypanosoma brucei. Percentage fall in packed red cell volume (pcv) is demon­strated and zebu mortalities are given. The symbols are as in Figure 5.

During August when there was some indication that animals were recover­ing from the needle challenge, as judged by levels of anaemia, G. pal­palis moved into the area and all zebu infected with T. brucei except one were reinfected and severe anae­mia developed (Figures 6 and 7); a similar situation occurred with the zebu infected with T. congolense (Figures 5 and 8). With N'Dama, how­ever, only a few animals became rein­fected and even then only transiently. The tsetse challenge appeared to have little effect on the course of the disease.

On examining the parasitaemia of infected N'Dama and zebu, it was immediately obvious that, while the prepatent period was similar, the level of parasitaemia which developed in the N'Dama was consistently and sig­nificantly lower than in the zebu. Furthermore, all of the 37 N'Dama allowed to survive and all 10 of the zebu survivors had the apparent ability to eliminate trypanosomes, or "self cure". There was some indication that the duration of parasitaemia was shorter in the N'Dama, but this observation was complicated by the tsetse challenge when most of the zebu, but only a few N'Dama, became reinfected. Nevertheless, from several months after needle challenge onward in most animals and for 90 days prior to termination of the experiment all surviving animals were negative for detectable parasites in the blood and tissues.

Following the disappearance of the parasites from the circulation, 25 of the 37 N'Dama and 3 of the 10 sur­viving zebu made a slow but complete clinical recovery, as judged by their physical improvement and a return to normal haematological values. How­ever, the remainder did not and, despite the absence of detectable para­sites, continued to be anaemic with packed red cell volumes (pcv) ap­proximately 30 to 40 percent below normal. The authors believe this neg­ative anaemic aspect of the trypanosomiasis syndrome to be widespread in the field (Murray, 1979). This outcome should be borne in mind when evaluating Figures 5, 6 and 7, where, by presenting the group mean pcv, a slightly misleading trend is ap­parent toward the end of the experiment.

Figure 7. Average parasitaemia score re�corded in N'Dama and zebu inoculated with I0³ Trypanosoma brucei. The level of the first peak of parasitaemia plus one standard deviation is shown. Percentage fall in packed cell volume (pcv) is demonstrated and zebu mortalities are given. The symbols are as in Figure 5.

Figure 8. Average parasitaemia score re�corded in N'Dama and zebu inoculated with 10⁷ Trypanosoma congolense. The level of the first peak of parasitaemia plus one standard deviation is shown. Percentage fall in packed red cell volume (pcv) is demonstrated and zebu mortalities are given. The symbols are as in Figure 5.

Another important finding to emerge from this study was that weight of challenge, as judged by the number of bloodstream forms of T. brucei inoculated, had a significant effect on the sequential development of the disease. Both the N'Dama and the zebu that received the heaviest chal­lenge became more rapidly and severely ill and more anaemic than those receiving the lowest dose. The parasitaemic profile was also influenced in that the prepatent period and the level of parasitaemia were lower in the animals inoculated with the lowest dose. In the same way, at least in the N'Dama, it appeared that the dose also affected the period of parasitae­mia, the duration being shorter in the group that received the lowest dose; in the zebu, however, this conclusion could not be made because most of them became reinfected during the period of G. palpalis challenge. In confirmation of the trypanotolerant nature of the N'Dama breed, but reflecting the quantitative rather than the absolute nature of trypanonotolerance, it was found that the N'Dama receiving the highest challenge, namely, 10⁸ T. brucei, developed a disease picture of the same order of magnitude as zebu given the lowest dose of 10³ T. brucei. This observation was made only in the initial few months following inoculation and was then complicated by reinfection of the zebu during the period of tsetse fly challenge.

In a further experiment, a group of ten female three-year-old zebu and nine female three-year-old N'Dama were subjected to what was considered a heavy G. m. submorsitans challenge. A daily mean of between 20 and 40 flies was caught per Malaise trap (W.F. Snow, personal communication). These cattle had no previous experi­ence of infection. The results were similar to the findings following needle and G. palpalis challenge. The zebu developed significantly higher levels of parasitaemia, more severe anaemia and all 10 had died of trypanosomiasis within 242 days of being moved into the challenge area. At this time the average pcv of the N'Dama was still above 30 percent; the only N'Dama death was caused by anthrax.

Based on clinical and postmortem findings in the above experiments, death from trypanosomiasis was the result of acute congestive heart failure brought about by a combination of anaemia, circulatory disturbance as­sociated with increased vascular per­meability, and myocardial damage. The authors believe that the fact that cattle in this study had to forage for their feed was a contributory factor. Tired, anaemic animals are probably unable to trek the distances necessary to satisfy their nutritional requirements but in their efforts to achieve this they develop cardiac decompensation. It was noticeable that when sick animals were put under intensive care and did not have to forage their clinical con­dition often improved.

While many aspects of these ex­periments still await evaluation, what has been established so far is that N'Dama, with no previous experience of infection, were less susceptible than zebu to a variety of challenge situations, including inoculation with T. brucei and/or T. congolense and to fly chal­lenge with G. palpalis and G. m. submorsitans; the more virulent the or­ganism the more significant were the differences between the breeds. How­ever, it was found that the trypanotolerant status of the N'Dama was not absolute and was affected by weight of challenge.

Trypanotolerance in other breeds of cattle.

There are several other small breeds of cattle in West Africa that are considered to be trypano­tolerant (Pagot, 1974). For example, Roberts and Gray (1973) found that Muturu were less susceptible than zebu but more susceptible than N'Dama; however, Desowitz (1959) found that two Muturu from a herd that had not been exposed to trypanosomiasis for 50 years were highly susceptible and succumbed three weeks after infection. However, in an outbred species such as the bovine it is difficult to draw firm conclusions when such small numbers of animals are involved; clearly more experimental data are required for these breeds.

It is generally considered that the zebu is the most susceptible of the African breeds. However, there is considerable epidemiological evidence that, in some areas, zebu have de­veloped a degree of tolerance; for example, Cunningham (1966) has de­scribed how thousands of zebu cattle survive around the shores of Lake Victoria even though they are contin­uously exposed to tsetse. He reported a 30 percent prevalence of parasites and the presence of neutralizing anti­bodies in 90 percent of such animals. By definition, these animals must be considered trypanotolerant. It is im­portant that further experimental and epidemiological studies be carried out to evaluate the extent of this situation.

Trypanotolerance in sheep and goats.

As with cattle, it is a well recognized but poorly documented fact that certain breeds of sheep and goats survive in endemic tsetsefly areas without the aid of chemotherapy and must be considered trypanotol­erant. There is a paucity of published data on the susceptibility of different breeds to trypanosomiasis under vari­ous challenge and ecological regimes. Work in Kenya has confirmed that differences in susceptibility do exist between different breeds of sheep and goats to both needle and fly challenge (E.W. Allonby, personal communica­tion); local breeds are much more resistant than imported ones. It is essential that the extent and basis of this difference be further investigated.

Trypanotolerance in wildlife.

Wild­life have an established reputation for being trypanotolerant or even com­pletely resistant to trypanosomiasis. There is, however, little published information on infection and the clini­cal course of the disease in experi­mentally infected wildlife. Surveys involving the demonstration of the presence or absence of trypanosomes in one blood sample on one occasion yield little useful information on sus­ceptibility to the disease of trypano­somiasis. Sequential studies following experimental infections are required where the clinical, parasitological, immunological and pathological parame­ters are assessed.

In one of the few studies of this type, Ashcroft, Burtt and Fairbairn (1959) examined the susceptibility of various wildlife species to needle chal­lenge with T. rhodesiense and T. brucei. They found that the animals examined could be considered to lie in two main categories: first, those species such as Thomson's gazelle, dikdik, Blue Forest duiker, jackal, bateared fox, Ant bear, hyrax, serval and monkey, which usually died of the infection: the second category included less sus­ceptible or resistant animals. These could be divided into species that became infected and had parasitaemias of considerable duration, such as the common duiker, eland, Bohar reedbuck, spotted hyena, oribi, bushbuck and impala; species usually infectible but with scanty parasitaemias, such as the warthog, bushpig and porcupine; and the baboon, which was refractory to infection. When the species of game that tsetse normally feed on was evaluated, what was of considerable interest was the observa­tion that the species most susceptible to needle infection were not popular with the tsetse, as judged by blood meals, whereas most of the species of the second group, i.e., the lesssus­ceptible group, were fed on more regularly, with wild pigs being most popular. This observation would sug­gest that species in the second group may have evolved by the survival and selection of the more resistant members within each of these species. In a similar but more limited study, Carmichael (1934) also found a range of susceptibility to T. brucei between different species of wildlife. Both Carmichael (1934) and Ashcroft, Burtt and Fairbairn (1959) attempted to infect a small number of game ani­mals with T. congolense. They found that in most cases the animals tested were not infectible or developed only transient infections and then recovered.

It would appear essential to the authors that such studies be extended. Not only is it important to evaluate the epidemiological role of game ani­mals in African trypanosomiasis but also the fact that certain species are less susceptible or even refractile to trypanosome infection makes them, important subjects for studies into the basic mechanisms of trypanotolerance. It may be, for example, that the resistant wildlife host can "see" anti­gens in the trypanosome that make the trypanosome more vulnerable; altern­atively, these species may have certain blood proteins that are active in a non­specific way against the trypanosome.

Productivity of trypanotolerant livestock.

There is a lack of scientific data on the productivity of "trypano­tolerant" livestock living in the field under natural tsetsefly challenge. As a result, a variety of opinions exists as to just how tolerant and how pro­ductive these animals really are under various ecological and management regimes and levels of challenge. The productivity of trypanotolerant live­stock is especially called into question on the basis of their small size in comparison with more susceptible breeds. At one extreme is the view that N'Dama cattle are genetically resistant and do not suffer from trypanosomiasis and should be intro­duced widely into high tsetsefly chal­lenge areas throughout Africa (Pagot, 1974). Other workers feel that further information is necessary before taking such an ambitious step (Stewart, 1951; Chandler, 1952; Roberts and Gray, 1973). At the other end of the scale, Stephen (1966) concluded that the propagation of these breeds, because of their small size, is not to be rec­ommended as a satisfactory means of protein production.

Leastsquares' means and constants for production indices of trypanotolerant cattle breeds under different management systems and tsetse challenges

In a recent major survey of the status of the trypanotolerant livestock of West and Central Africa (ILCA, 1979), indices of productivity of try­panotolerant cattle were examined, using all the basic production data that could be found in the region. In 30 herds in the 18 countries studied, sufficient information was available on the main production traits to produce indices. The traits evaluated were reproductive performance, cow and calf viability, milk production, growth and cow body weight. These were used to compute the index of the total weight of calf and liveweight equivalent of milk produced, first per cow per year and finally per 100 kg of cow maintained per year. This final index related these important production traits back to the actual weight of breeding cow that had to be supported, this being closely con­nected with maintenance costs. The traits and production indices were derived for two basic management systems, village and ranch or station and for four levels of tsetse challenge rather arbitrarily designated zero, low, medium and high. The table indicates the effects of breed groups, manage­ment system and tsetse challenge on the two productivity indices.

A tremendous range of productivity levels was spanned by both the N'Dama and West African Shorthorn relative to the different production systems and level of tsetse challenge involved. In both breeds, the range extended from about 15 kg of one-year-old calf and liveweight equivalent of milk pro­duced per 100 kg of cow maintained per year under village conditions in a high tsetsechallenge area, to about 50 kg under improved ranch or station conditions in a low tsetsechallenge area.

The table indicates no significant difference between N'Dama and West African Shorthorn for the major index of "productivity per 100 kg of cow maintained", the actual values being 28.4 kg per annum for N'Dama and 27.6 kg for West African Shorthorn. The only significant differences in individual traits leading to this index were of one-year-old calf and weight of mature cow, the N'Dama group being very much heavier in each case. The higher calf weight led to a higher index per cow for the N'Dama, but the higher maturecow weight resulted in similar indices per 100 kg of cow maintained. The effect of management system was a 38 percent lower pro­ductivity index per cow and 30 percent lower productivity index per 100 kg of cow maintained from the village compared with the ranch or station. The performance attributable to zero tsetse challenge was masked by the ef­fect of very intensive feeding and man­agement, thus only low, medium and high can be directly compared. Pro­ductivity indices per cow were 30 per­cent and 56 percent less for medium and high challenge respectively com­pared with low, while indices per 100 kg of cow maintained were 26 percent and 41 percent less for medium and high respectively compared with low.

Estimates of productivity for 16 zebu and Sanga herds under ranch/station conditions in tsetsefree areas of Africa covering Botswana, Kenya, Mali, Ni­geria, Senegal and Uganda, have been built up from the available literature. These averaged 133.4 kg of one-year-old calf and liveweight equivalent of milk produced per cow maintained per year and 37.7 kg per 100 kg of cow maintained per year. Compared with the estimates of 79.7 kg and 36.1 kg for the 30 trypanotolerant groups under ranch/station conditions in light tsetsechallenge areas, these represent a superiority of 67 percent per cow. maintained per annum, but only 4 per­cent per 100 kg of cow maintained per annum for the zebu and Sanga over the trypanotolerant breeds. This strongly suggests that the productivity of trypanotolerant cattle relative to other indigenous types may be much higher than previously assumed.

A preliminary survey of the impact of trypanosomiasis has been carried out on the N'Dama on The Gambia¹, which live under different levels of tsetse challenge (Murray et al., 1977b; Clifford and McIntyre, 1977). It was found that in heavy G. m. submorsitans areas anaemia was widespread and up to 50 percent of the herd could be infected. In areas of lighter G. palpalis challenge, the prevalence of trypanosomes was less as was the extent of anaemia. In the heavy chal­lenge areas, while some of the trypanosomeinfected N'Dama died, most N'Dama survived but they often did so in a poor productive state with wasting, stunting (Figure 9), abortion, high calf mortality and with a persistent lowgrade anaemia being manifest. Thus, there is little doubt that trypano­somiasis must be considered a dis­ease of importance in N'Dama in terms of morbidity if not mortality. However, it should be emphasized that many other animals in the same herds were in an excellent productive state, suggesting that a wide range of sus­ceptibility exists within the N'Dama breed. It must be remembered that these results were obtained in areas where zebu could not survive; of 31 zebu introduced in the G. m. submor­sitans area in June 1977, only one survived until December 1978, and this animal was in poor condition. In the same way, of the 31 zebu studied in the G. palpalis area 21 died; the N'Dama in this area were hardly affected.

These N'Dama herds, numbering around 2 000 head, are now double eartagged and have been investigated from a disease and productivity point of view over several years. Detailed quantitative data are at present being evaluated by one of the authors (D.J. Clifford).

Basic mechanisms of trypanotol erance.

While the basic mechanism of trypanotolerance is still to be precisely defined, there is at least circumstantial evidence that the mech­anism is related to a hostresponse factor and that it is a heritable trait.

The trypanotolerant nature of the N'Dama and the capacity of certain strains of mice to survive a trypanosome infection longer than others (Morrison et al, 1978b) would appear to be related to their ability to limit the level of peaks of parasitaemia and subsequently to control, reduce or even eliminate the parasite. The find­ing of a similar prepatent period between N'Dama and zebu, and be­tween strains of mice of high and low susceptibility suggested that the initial replication rates in all groups were similar. Furthermore, dose titration studies showed that there was no difference in the infectivity of T. congolense for mice of high and low susceptibility (Morrison et al., 1978b). These findings indicated that the dif­ferent levels of parasitaemia found between breeds of cattle and strains of mice might reflect differences in the nature or quality of the immune response to the trypanosome. This hypothesis requires experimental verification.

Evidence that a more effective im­mune response might be responsible for the differences in susceptibility be­tween N'Dama and zebu cattle comes from the work of Desowitz (1959). He found that N'Dama with previous experience of trypanosomiasis were able to eliminate trypanosomes more rapidly than their zebu counterparts following a renewed challenge. Em­ploying an in vitro test that involved the use of sera from the challenged animals to inhibit trypanosome res­piration, it was found that the activity of N'Dama sera was superior to that of zebu sera. Desowitz (1959) con­cluded that the trypanotolerant nature of the N'Dama lay in its capacity to mount a better secondary immune response. Unfortunately, in these stud­ies the trypanosomal antigenic history of the N'Dama and zebu used was not known precisely. Thus, while the results achieved are indicative of a moreeffective immune response in the N'Dama they require confirmation. Similarly, using a serum neutralization test, Chandler (1958) stated, without supplying details, that the immune response of zebu to the trypanosome was inferior to that of the N'Dama.

A range of susceptibility to T. con­golense (Morrison et ah, 1978) and to T. brucei and T. vivax (Morrison and Murray, 1979) has been shown to occur in different inbred strains of mice. Following T. congolense infection, the C57BI was the least susceptible and the A/J was the most susceptible of the strains examined. When the spleen lymphocyte populations were studied in these strains of mice infected with T. congolense, it was found that there was a marked increase in splenic Blymphocytes and null cells (Mor­rison et al., 1978). This, allied to the findings of an increase in background plaqueforming cells to sheep erythrocytes, indicated that trypanosome infection resulted in a nonspecific polyclonal activation of lymphocytes, affecting primarily Blymphocytes., Tn the strains of mice that survived longest, the C57BI and AKR, the increase in Bcells and null cells was less than in the highly susceptible strains. Furthermore, the immunosuppression observed in mice infected with T. congolense occurs earlier in the highly susceptible strains. It might be, therefore, that susceptibility is related to sensitivity to polyclonal activation or to immunosuppression induced by the trypanosome infection. Alternatively, differences in the cellular response and degree of immunosup­pression might merely reflect differ­ences in levels of parasitaemia observed in the various strains of mice (Mor­rison et al, 1978).

Further support for the hypothesis that trypanotolerance has an immunological basis comes from the effect of immunostimulants on the susceptibility of mice to trypanosomiasis (Morrison and Murray, 1979). The authors found that the administration of Bordetella pertussis, Corynebacterium parvum or Bacillus Calmette-Guérin (BCG) prior to or on the day of challenge with T. congolense significantly delayed or re­duced parasitaemias and increased sur­vival time. In this way, it was possible to change the survival time and parasitaemia levels of the highly sus­ceptible AjJ strain to values more akin to the less susceptible C57BI. This strategy might have practical significance in rearing domestic live­stock.

Further work is now required to define both qualitatively and quanti­tatively the host's immune response to the trypanosome and to compare this between breeds and strains of animals with different susceptibilities. In this way the important effector mechanisms might be clearly defined and, moreover, it might be possible to potentiate them with the object of increasing host resistance to trypanosomiasis.

Figure 9. 'The effect of trypanosomiasis on a naturallyinfected N'Dama yearling. The animal is stunted and has a characteristic "nagana" pose.

It is also essential that possible physiological factors be considered in the construction of the overall picture of the underlying mechanisms of trypanotolerance. As the N'Dama tend to develop lesssevere anaemia than zebu, the authors considered that this might be related to the capacity to mount a more effective erythropoietic response. Thus, a series of in vivo pathological studies were carried out in N'Dama and zebu cattle infected with T. congolense and T. brucei; these studies involved the use of ⁵¹Crlabelled red cells, ¹²⁵Ilabelled al­bumin and ⁵⁹Felabelled transferrin (Dargie et al., 1979; Dargie et al., 1979). The findings, however, showed that the anaemia and its underlying processes were broadly related to the number of parasites in the blood and that the superior resistance of the N'Dama lay in their capacity to control parasitaemia rather than their ability to mount a more efficient erythropoietic response.

Nevertheless, it would seem to us that, when N'Dama and zebu are kept under the same conditions and have anaemia of similar severity, the N'Dama always appear to be clinically and physically superior.

Under normal field conditions where nutrition is poor, cattle often have to forage up to about 27 km in a day; perhaps the N'Dama's ability to forage and to digest what it gets is superior to the zebu. Several other physio­logical mechanisms are also worthy of consideration. The authors have observed remarkable water conserva­tion and heat tolerance in N'Dama; rectal temperatures can range from 34.4°C at dawn to 41.1°C by late afternoon. Furthermore, Pagot (1974) has pointed out that N'Dama can withstand higher levels of humidity than zebu. Zebu appear to have evolved a capacity for conserving water and in East Africa it has been shown that the water requirements of zebu steers is half that of Hereford steers and is as specialized as several species of game animals (EAVRO, 1967). Zebu were found to be better able to conserve evaporative and faecal water than Hereford. Zebu deprived of water stopped eating and metabolised fat consequently reducing urinary and faecal water losses; zebu cattle form faeces as dry as 190 g water/100 g dry matter whereas Hereford are unable to form faeces containing less than 300 g water/100 g dry matter. As a, result, zebu were able to live com­fortably without water for two months at an environmental temperature of 22°C or until their fat supplies were depleted, a fact confirmed by field observations on Turkana cattle living under drought conditions. This ca­pacity for conserving water was in­herited as a dominant trait in zebu-Hereford F₁crosses (EAVRO, 1967). In possible contrast to N'Dama, zebu would appear to regulate their body temperature within a range of 2°C and neither a periodic heat load nor dehydration had any effect on the range over which zebu regulate (EAVRO, 1967). The greater variation in body temperature in the N'Dama compared with zebu might result in better conservation by N'Dama of water which would otherwise be lost by evaporation. The authors believe that these observations require further investigations to evaluate the possible role played by adaptation of physiolog­ical mechanisms in trypanotolerance.

Another aspect that might play a potential role in trypanotolerance is skin physiology, including colour and smell, in relation to attractiveness to the tsetse; skin structure might also be important. In addition the role of nonspecific factors in host sus­ceptibility to trypanosomiasis should be compared between breeds; these factors might include the extent and activity of the mononuclear phagocytic system as well as complement, properdin and conglutinin reactivity. One interesting finding is that the N'Dama breed has a significantly higher level of white blood cells, particularly eosinophils, than the zebu. This ob­servation was made by Oduye and Okunaiya (1971) and the authors sub­sequently have confirmed their findings.

The genetics of trypanotolerance.

There is now a considerable body of evidence to show that trypanotolerance has a genetic basis. Thus, studies on cattle that have had no previous ex­perience of trypanosomiasis have clear­ly established that N'Dama are sig­nificantly more resistant than zebu (Stephen, 1966; Roberts and Gray, 1973). It is likely that trypano­tolerance has evolved in tsetsefly infested areas by natural selection of the more resistant animals within a breed. In this respect (in the authors' experience) a range of susceptibility is found both within groups of N'Dama and zebu cattle. Under such circum­stances it is likely that the factors governing the susceptibility have a complex genetic basis.

There have been few breeding studies with trypanotolerant breeds of cattle. Stewart (1951) reported crossbreeding studies, without supplying details, in­volving the West African Shorthorn, a trypanotolerant genetic mix, and zebu, in which trypanosomiasis re­sistance was retained in the crossbred offspring. However, Chandler (1958) found that the resistance of N'Damazebu crosses to trypanosome challenge was about half way between the two parent breeds. In an extended cross­breeding trial in the Ivory Coast involving large numbers of N'Dama and Jersey, it was found that the F₁ cross produced an excellent animal as regards growth and milk production (Letenneur, 1978). It was stated that such crosses retained their tolerance although no data were supplied on the level of fly challenge or on the preva­lence of trypanosomes. Crossbreds with greater than 50 percent Jersey background appeared to be less hardy and gave equivocal results. The fore­going reports indicate that the trypano­tolerant trait is at least partly dom­inant. However, in using such outbred populations, differences in results on crossbreeding must be expected. Indeed, it is likely that the degree of trypanotolerance observed in first gen­eration crosses will vary widely de­pending on the individual parental combination. In this respect, any future investigation of the inheritance of trypanotolerance will require the use of large groups of both parental breeds if reliable results are to be obtained. Thus, only by using suffi­cient numbers of animals will it be possible to determine the feasibility of selection for maximum trypano­tolerance along with the retention of the required characteristics of the nontrypanotolerant parent. The main problem in undertaking such a study at present is the lack of genetic markers that would allow monitoring of sus­ceptibility without having to infect all of the animals involved.

It has been proposed that, as N'Dama cattle show almost 100 percent gene frequency for Haemoglobin (Hb) A, while zebu are a mixture of A and B, animals could be selected by Hb type. However, the fact that certain exotic breeds such as the Friesian aie predominantly HbA (Bangham and Blumberg, 1958) and are highly sus­ceptible to trypanosomiasis, makes it unlikely that Hb type will be of value as a marker. With the Tecent upsurge of interest in the major histocompatibility complex (mhc) in cattle and the identification of a series of gene prod­ucts, the possible association of trypano­tolerance with particular mhc prod­ucts might prove a more profitable avenue of research.

The lack of suitable herds for study, the absence of genetic markers for resistance and the genetic heterogeneity of bovine populations, at present pre­clude a critical analysis of the genetics of trypanotolerance in the bovine. Thus, the authors have carried out a series of experiments to compare the susceptibility of different inbred strains of mice to trypanosomiasis and to in­vestigate the underlying genetics (Mor­rison et al., 1978; Morrison and Murray, 1979).

As stated earlier, it was found that strains of mice differed markedly in their susceptibility to African trypano­somiasis. Breeding studies indicated that reduced susceptibility (as judged .by survival times) was inherited as a dominant trait, in that F₁ hybrids be­tween the highly susceptible A/J strain and the moreresistant C57B1J6 showed similar survival times to the C57BI/6 parents (Figure 10). When these F₁ hybrids were then back-crossed onto the parent strains, the extent of het­erogeneity in survival indicated that susceptibility was under polygenic con­trol (Morrison and Murray, 1979).

In recent years, the susceptibility to a number of experimental infections in mice and the prevalence of certain diseases in man has been shown to be at least partially linked to H-2 hap-lotype (Lilly and Pincus, 1973; Mc Devitt, Oldstone and Pincus, 1974) and with particular HLA antigens (Vladutiu and Rose, 1974) respectively. It has been suggested that H-2 may exert its influence through immune response (Ir) genes present in the I region of the H-2 complex. It is thought that immune response genes may also be associated with the HLA complex in man. However, the authors' studies on the comparative susceptibility of congenic resistant mice, i.e., mice with a genetic background differing only at the H-2 locus, have failed to demon­strate a major relationship between H-2 haplotypes and susceptibility (Morrison and Murray, 1979). So far, the authors have carried out these experiments only on mice of the C57BI genetic background and it may be that the genes responsible for re­duced susceptibility in this strain over­ride any influence exerted by the H-2 haplotype.

Despite the lack of precise data there would appear to be overwhelming evidence that susceptibility of cattle to African trypanosomiasis is under ge­netic control. Nevertheless, there is a considerable body of evidence to support the fact that the innate sus­ceptibility of cattle is decreased by repeated exposure to the same pop­ulation of trypanosomes in a given area. Thus Desowitz (1959) demon­strated the ability of N'Dama and, to a lesser extent, zebu that had been previously exposed, to mount what could be described as a secondary immune response with the elimination of the parasite. He believed that the course of the disease was dependent not only on the breed of animal but also on the nature of the individual's past contact with the trypanosome. The fact that cattle previously exposed to trypanosomiasis are more resistant was described many years ago by Bevan (1928) and more recently by Wilson et al. (1976) who showed that zebu cattle kept under an infection-and-treatment regime did become more resistant.

Figure 10. Percentage deaths in groups of A/J, C57B1/6J, F₁ hybrids and backcrosses after infection with Trypanosoma congolense. Morrison and Murray, 1979

In addition to previous exposure to infection, it is established that other factors, such as weight of challenge, influence susceptibility. Studies have shown that if the dose of inoculum is heavy enough N'Dama can become very ill and may even die (Murray et al, 1977 a, e).

In addition, it is essential that the effect on susceptibility of such factors as nutrition, stress, exercise, age of first exposure, effect of colostrum, sex, pregnancy and parturition be fully investigated. The effect of intercurrent disease must also be considered as must the relative susceptibility of trypanotolerant breeds to other in­fections; it has been reported, for example, that N'Dama are more re­sistant to streptothricosis (Oduye and Okunaiya, 1971). In addition, the impact of transferring trypanotolerant stock to a distant location must be critically evaluated in order that the conflicting opinions that exist on move­ment may be resolved; this is obviously one of the most important questions to be answered about trypanotolerance although the successful establishment of trypanotolerant breeds in Zaire and elsewhere would suggest that these breeds do adapt (Pagot, 1974; Mortelmans and Kageruka, 1976).

Conclusions.

There is now a con­siderable body of evidence, both epidemiological and experimental, to con­firm the existence of trypanotolerance. It would also appear that trypano­tolerance has a genetic basis although this may be supplemented by repeated exposure to the same population of trypanosomes in a given area. How­ever, trypanotolerance is not absolute and breaks down if the weight of challenge is heavy enough. Further work on the genetics of susceptibility in cattle, sheep and goats and precise information on the effect of environ­mental factors are necessary.

The productivity of trypanotolerant livestock relative to other indigenous types may be much higher than pre­viously assumed; they could well be an economically viable proposition in their own right and be introduced into areas where other livestock cannot exist. Furthermore, it is likely that strategies involving immunotherapy, if and when available, and drugs will be more effective and economically viable if carried out on trypanotolerant stock. Finally, there is little doubt that comparative studies of trypano­tolerant and susceptible animals offer one of the best approaches for the understanding of the important immunological mechanisms and physio­logical factors involved in hostparasite interactions operative in African try­panosomiasis. ■

References

Aschroft, M.T., Burtt, E. & Fairbairn, H. 1959. The experimental infection of some African wild animals with Trypanosoma rhodesiense, T. brucei and T. congolense. Ann. trop. Med. Parasit., 53: 147-160.

Bangham, A.D. & Blumberg, B.S. 1958. Distribution of electrophoretically different haemoglobins among some cattle breeds of Europe and Africa. Nature, 181: 1551-1552.

Bevan, L.E.W. 1928. Method of inocu­lating cattle against trypanosomiasis. Trans. R. Soc. Trop. Med. Hyg., 22: 147-156.

Carmichael, J. 1934. Trypanosomes patho­genic to domestic stock and their effect in certain species of wild fauna in Uganda. Ann. trop. Med. Parasit., 28: 41-45.

Chandler, R.L. 1952. Comparative toler­ance of West African N'Dama cattle to trypanosomiasis. Ann. trop. Med. Parasit., 46: 127-134.

Chandler, R.L. 1958. Studies on the tol­erance of N'Dama cattle to trypanoso­miasis. J. comp. Path., 68: 253-260.

Clifford, D.J. & McIntyre, W.I.M. 1977. An epidemiological study of trypanoso­miasis in N'Dama cattle in The Gambia. In International Scientific Council for Trypanosomiasis Research and Control, 15th meeting. The Gambia, OAU/STRC. Publication No. 110, p. 152-154.

Cunningham, M.P. 1966. Immunity in bovine trypanosomiasis. E. Afr. med. J., 43:394-397.

Dargie, J.D., Murray, P.K., Murray, M. & McIntyre, W.I.M. 1979. The blood volumes; and erythrokinetics of N'Dama and Zebu cattle experimentally infected with Trypanosoma brucei. Res. vet. Sci. (In press)

Dargie, J.D., Murray, P.K., Murray, M., Grimshaw, W.T.R. & McIntyre, W.I.M. 1979. Bovine trypanosomiasis: the red cell kinetics of N'Dama and Zebu cattle infected with Trypanosoma congolense. Parasitology. (In press)

Desowitz, R.S. 1959. Studies on immunity and hostparasite relationship. I. The immunological response of resistant and susceptible breeds of cattle to trypanosomal challenge. Ann. trop. Med. Parasit., 53: 293-313.

EAVRO. 1967. Annual report. Physiology Section, Animal Production Division, East African Veterinary Research Organization, p. 56-65.

ILCA. 1979. Trypanotolerant livestock in West and Central Africa. Vol. 1: General study. Addis Ababa, Ethiopia, Interna­tional Livestock Centre for Africa.

Letenneur, L. 1978. Crossbreeding N'Da­ma and Jersey cattle in Ivory Coast. Wld Anim. Rev., 27: 36-42.

Lilly, F. & Pincus, T. 1973. Genetic control of murin virale leukaemogenesis. Advanc. Cancer Res., 17: 231-277.

McDevitt, H.O., Oldstone, M.B.A. & Pincus, T. 1974. Histocompatibility linked genetic control of specific immune responses to viral infection. Transplant. Rev., 19: 209-225.

Morrison, W.I., Roelants, G.E., Mayor-Withey, K.S. & Murray, M. 1978. Susceptibility of inbred strains of mice to Trypanosoma congolense. Correlation with changes in spleen lymphocyte popu­lations. Clin. Exp. Immunol., 32: 25-40.

Morrison, W.I., Roelants, G.E., Pearson, T.W. & Murray, M. 1978b. Genetic control of susceptibility to Trypanosoma congolense infection in inbred strains of mice. Advances in Experimental Medicine and Biology. Vol. 69. New York and London, Plenum Press. (In press)

Morrison, W.I. & Murray, M. 1979. Inheritance of susceptibility Trypanosoma congolense infection in inbred strains of mice. Exp. Parasit. (In press)

Mortelmans, J. & Kageruka, P. 1976. Trypanotolerant cattle breeds in Zaire. Wld Anim. Rev., 19: 14-17.

Murray, C, Murray, M., Murray, P.K., Morrison, W.I., Pyne, C. & McIntyre, W.I.M. 1977. Diagnosis of African try­panosomiasis in cattle. Improved parasitological and serological techniques. In International Scientific Council for Try­panosomiasis Research and Control. 15th meeting. The Gambia, OAU/STRC. Pub­lication No. 110, p. 247-254.

Murray, M., McIntyre, W.I.M., Murray, P.K., Urquhart, G.M., Jennings, F.W., Greig, W.A., Clifford, D.J., N'Dow, W.S.M., Touray, B., Sanyang, B.J. & Bray, R.S. 1977a. Cattle diseases and trypanosomiasis in The Gambia. I. Clini­cal studies. In International Scientific Council for Trypanosomiasis Research and Control. 15th meeting. The Gambia, OAU/STRC. Publication No. 110, p. 83-91.

Murray, M., McIntyre, W.I.M., Murray, P.K., Urquhart, G.M., Jennings, F.W., Greig, W.A., Clifford, D.J., N'Dow,

W.S.M., Touray, B., Sanyang, B.J. & Bray, R.S. 1977b. Cattle diseases and trypanosomiasis in The Gambia. II. Pathology studies. In International Scientific Council for Trypanosomiasis Research am Control. 15th meeting. The Gambia OUA/STRC. Publication No. 110 p. 92-98.

Murray, P.K., Murray, M., Wallace, M. Morrison, W.I. & McIntyre, W.I.M 1977a. Trypanosomiasis in N'Dama and Zebu cattle. I. An experimental inves tigation of susceptibility to Trypanosomt brucei, T. congolense and mixed infections In International Scientific Council for Trypanosomiasis Research and Control 15th meeting. The Gambia, OAU/STRC Publication No. 110, p. 470-481.

Murray, P.K., Murray, M., Wallace, M. Morrison, W.I. & McIntyre, W.I.M 1977b. Trypanosomiasis in N'Dama and Zebu cattle. II. The influence of weight of infection on the severity of the disease In International Scientific Council for Trypanosomiasis Research and Control 15th meeting. The Gambia, OAU/STRC Publication No. 110, p. 482-487.

Oduye, O.O. & Okunaiya, O.A. 1971 Haematological studies on the White Fulani and N'Dama breeds of cattle. Bull. epiz. Dis. Afri., 19: 213-218.

Pagot, J. 1974. Les races trypanotolérantes. In Control programmes for trypa­nosomes and their vectors. Rev. Elev. Med. Vet. Pays. Trop. Actes du Colloque. Paris, p. 235-248.

Roberts, C.J. &Gray, A.R. 1973. Studies on trypanosomeresistant cattle. II. The effect of trypanosomiasis on N'Dama, Muturu and Zebu cattle. Trop. Anim. Health Prod., 5: 220-233.

Stephen, L.E. 1966. Observations on the resistance of West African N'Dama and Zebu cattle to trypanosomiasis following challenge by wild Glossina morsitans from an early age. Ann. trop. Med. Parasit., 60: 230-246.

Stewart, J.L. 1951. The West African Shorthorn cattle. Their value to Africa as trypanosomiasisresistant animals. Vet. Rec, 63: 454-457.

Vladutiu, A.O. & Rose, M.R. 1974. hl-a Antigens: association with disease. Immunogenetics, 1: 305-328.

Wilson, A.J., Paris, J., Luckins, A.G., Dar, F.K. & Gray, A.G. 1976. Obser­vations on a herd of beef cattle maintained in a tsetse area. II. Assessment of the development of immunity in association with trypanocidal drug treatment. Trop, Anim. Health Prod., 8: 1-12.

The first N'Dama cattle were brought to southwestern Zaire in the early 1920s

M. Murray and W.I. Morrison are with the International Laboratory for Research on Animal Diseases (ILRAD), PO Box 30703, Nairobi, Kenya; P.K. Murray is with Merck Sharp and Dohme Research Laboratories, Rahway, New Jersey, USA; D.J. Clifford is presently at the Department of Veterinary Services, Abuko, The Gambia (attached to th^e Glasgow University Veterinary School, Scotland); J.C.M. Trail is at the International Livestock Centre for Africa (ILCA), PO Box ,46847, Nairobi, Kenya.

¹ Not all of these herds are pure N'Dama and some are judged to have a component of West African Shorthorn.