Activity of 10 naphthoquinones, including parvaquone (993C) and menoctone, in cattle artificially infected with Theileria parva

Ten naphthoquinones, including parvaquone (993C) (Clexon; Wellcome) and menoctone, were tested for activity in cattle artificially infected with Theileria parva, the causative organism of East Coast fever (ECF). Parvaquone cured all 14 cattle treated with a single dose of 20 mg/kg intramuscularly and all five treated twice with 10 mg/kg intramuscularly. Menoctone cured seven of 10 cattle treated with a single dose of 5 mg/kg intramuscularly. Of 25 untreated control cattle, 22 died of ECF. None of the remaining eight naphthoquinones was as active as parvaquone. Three esters of active naphthoquinones, designed as 'prodrugs' of their parent compounds, showed little or no activity in infected cattle despite being highly active in vitro against T parva. These results were instrumental in the selection of parvaquone for development as the first specifically active remedy for ECF.     

Evaluation of PCR to detect Theileria parva in field-collected tick and bovine samples in Tanzania

The ability of PCR to detect infections of Theileria parva, the cause of East Coast Fever, in field-collected tick and bovine samples from Tanzania was evaluated. PCR-detected infection prevalence was high (15/20, 75%) in unfed adult Rhipicephalus appendiculatus ticks that fed as nymphs on an acutely-infected calf, but low (22/836, 2.6%) in unfed adult R. appendiculatus collected from field sites in Tanzania. Tick infection prevalence was comparable to that in previous studies that used salivary gland staining to detect T. parva infection in field-collected host-seeking ticks. Of 282 naturally-exposed zebu calves, seven had PCR-positive buffy coat samples prior to detection of Theileria spp. parasites in stained buffy coat cells or lymph node biopsies. Evidence of Theileria spp. infections was detected in stained smears of lymph node biopsies from 109 calves (38.6%) and buffy coat samples from 81 (28.7%), while buffy coat samples from 66 (23.4%) were PCR-positive for T. parva. Implications of these findings for the sensitivity and specificity of the PCR are discussed.     

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

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

Antitheilerial activity of BW720C (buparvaquone): a comparison with parvaquone

A series of hydroxynaphthoquinones, all derivatives of the antitheilerial hydroxynaphthoquinone parvaquone (993C, Clexon; Wellcome) was tested for antitheilerial activity against Theileria parva (Muguga) in vitro. BW720C (buparvaquone) was 20 times more active than parvaquone. When tested in vivo BW720C cured all 13 cattle infected with T parva and all six infected with T annulata treated at a dose rate of 2.5 mg (kg bodyweight)-1 while parvaquone at 20 mg kg-1 cured nine of 10 cattle. All 16 untreated control cattle died of theileriosis.     

Endemic stability for Theileria parva infections in Ankole calves of the Ankole ranching scheme, Uganda

A population-based study was carried out on the Ankole ranching scheme in south-west Uganda with the aim of determining the endemic status of Theileria parva infections. For this purpose, the age-related sero-prevalence of T. parva and the specific calf mortality associated with the parasite were assessed. Blood samples were collected from 931 Ankole calves of up to 12 months of age from 81 randomly selected herds. The relationship between rainfall pattern and whole-body Rhipicephalus appendiculatus counts was determined. The influence of tick control practices on East Coast fever-related calf mortality, and sero-positivity were also determined. A significant (r2 = 0.76, P = 0.000) association between R. appendiculatus counts and rainfall was observed. There was no significant (P > 0.05) association between theileriosis-related calf mortality, sero-positivity and the different tick control practices. Antibody prevalence based on the PIM ELISA was above 70% among calves of 6 months of age in 96% in all the herds. Theileria parva-related calf mortality determined by repeated herd visits and farm records ranged between 0 % and 5.4 %. It was concluded that endemic stability for theileriosis, caused by T. parva, existed in the study area, and that the risk of the occurrence of economically important outbreaks of East Coast fever in indigenous cattle was regarded as minimal under the prevailing conditions.     

Indirect fluorescent antibody test for experimental and epizootiological studies on East coast fever (Theileria parva infection in cattle). Evaluation of a cell culture schizont antigen fixed and stored in suspension

A schizont antigen for the indirect fluorescent antibody test against Theileria parva was prepared from a T parva-infected bovine lymphoblastoid cell line by fixing the cells in suspension with a mixture of acetone and formaldehyde. The antigen was stored in suspension in phosphate buffered saline for one and a half years at -60 degrees C without loss of activity; the antigen could also be lyophilised. The fluorescence of the intracellular schizonts was bright and specific with T parva positive bovine control serum and absent with negative bovine control serum and Theileria mutans positive bovine control serum. Fluorescence of the lymphoblastoid cell itself was observed with Trypanosoma brucei positive control serum and some bovine test sera: this fluorescence, which masked the intracellular schizonts, was eliminated by absorbing the sera in the supernatant of sonicated lymphocytes obtained from bovine lymph nodes. The antigen was evaluated with sera from cattle experimentally infected with T parva. In an epizootiological study on East Coast fever in the Coast Province of Kenya, there was good correlation between the serological responses of cattle to T parva schizont antigen and the distribution of Rhipicephalus appendiculatus ticks.     

An outbreak of East Coast Fever on the Comoros: A consequence of the import of immunised cattle from Tanzania?

In 2003 and 2004, a severe epidemic decimated the cattle population on Grand Comore, the largest island of the Union of Comoros. Fatalities started soon after the import of cattle from Tanzania. Theileria parva and its vector, Rhipicephalus appendiculatus, could be identified as the main culprits of the epidemic. Characterisation by multilocus genotyping revealed that the T. parva parasites isolated on the Comoros were identical to the components of the Muguga cocktail vaccine used in Tanzania to immunise cattle. Therefore, it is believed that East Coast Fever reached the Comoros while some of the imported livestock got infected in Tanzania by ticks of which the immature stadia fed on Muguga cocktail vaccinated animals. Since the Comorian government neither has the financial means nor the competent staff to pursue an adequate epidemiosurveillance, the danger exists that without external assistance and in a context of continuing globalisation more transboundary diseases will affect the Comorian livestock sector in the future.     

The effect of East Coast fever immunisation and different acaricidal treatments on the productivity of beef cattle

A trial was performed on a farm in the Coast Province of Kenya to study the effects of East Coast fever immunisation and different acaricidal treatments on the productivity of immunised and unimmunised beef cattle. Eighty cattle were immunised against Theileria parva parva (Marikebuni) by the infection and treatment method and a similar group was left as an unimmunised control. Immunisation had no deleterious effect on the cattle. After immunisation, the immunised and control groups were each subdivided into four groups of 20 and each subgroup was managed under a different tick control regimen. The tick control regimen were, acaricidal spraying twice a week or once every three weeks, the application of acaricide-impregnated ear-tags, and no tick control. During a nine-month exposure period there were 18 cases of East Coast fever among the 80 immunised cattle, three which were severe and the others mild. Among the 80 unimmunised cattle there were 57 cases of East Coast fever, 50 of which were severe. The highest morbidity and mortality occurred in the groups under limited tick control or without tick control. Overall weight gain in the immunised cattle, irrespective of the tick control regimen, was better than the weight gain in the unimmunised groups. Within the immunised groups, the weight gain of the cattle sprayed twice weekly was comparable to the weight gain of the animals with acaricidal ear-tags and was significantly higher than the weight gains in the groups sprayed once every three weeks or with tick control. Preliminary cost/benefit analysis showed that it was uneconomical to maintain unimmunised cattle under limited or no tick control.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immunization against East Coast fever: the use of selected stocks of Theileria parva for immunization of cattle exposed to field challenge

Two antigenically different stocks of Theileria parva parva (Kilifi and Marikebuni), previously characterized as belonging to groups A and C respectively on monoclonal antibody (MAb) profiles, were selected for immunization of different breeds of cattle against East Coast fever (ECF) by the infection and treatment method. A total of 52 immunized cattle and 33 susceptible controls of different group sizes were exposed to field challenge by ticks for periods of 42-90 days at three field sites where ECF is endemic on the Kenyan coast. All immunized cattle survived ECF challenge, but 87% of the controls died of the disease. The cattle exposed at one site had been immunized 1 year earlier and maintained tick-free in the intervening period. The level of immunity in these cattle was similar to that of cattle which had been immunized 1 or 2 months prior to exposure. Thus, immunity had not waned over the 1-year period. A study at another site showed that acaricidal treatment of immunized cattle could be safely extended from twice a week to once every three weeks, whereas in susceptible cattle even twice weekly spraying did not control ECF. The isolates made from infected controls during the trials indicated the presence of three T. p. parva stocks as defined by MAb profiles. Of the two stocks used for immunization, T. p. parva Marikebuni induced broader protection. In view of the apparent limited antigenic diversity of T. p. parva strains within the Coast Province it is suggested that the Marikebuni stock might represent a key stock for vaccination in this area.     

Chemotherapy of East Coast fever: parvaquone treatment of clinical disease induced by isolates of Theileria parva

Two experiments were carried out in which parvaquone was used to treat experimentally-induced acute clinical East Coast fever infections. In the first experiment, infections with Theileria parva parva (Kiambu 5) were induced by applying infected Rhipicephalus appendiculatus ticks or by inoculation of triturated infected-tick stabilate. The character of the disease was similar with both methods of infection and following a single treatment with parvaquone at 20 mg kg-1, 5 of 7 cattle in each group recovered. All untreated control cattle died. In the second experiment, 5 stabilate isolates from different locations within East Africa, and representative of the challenge likely to be met in the field, were used. Treatment was administered in 2 X 10 mg kg-1 doses 48 h apart. The isolates used were T. p. parva (Mbita), T. p. parva (Pugu), T. p. parva (Entebbe), T. p. lawrencei (Mara) and T. p. lawrencei/(Manyara); following treatment 3/7, 6/6, 6/7, 5/7 and 6/7 animals recovered, respectively. All untreated control cattle died. There was evidence of a difference in susceptibility of isolates to treatment, and some animals showed prolonged disease episodes. The nature of the response to treatment and the problems in treating a lympho-destructive disease are discussed.     

Histopathologic and electron microscopic studies of cutaneous lesions in calves with experimentally induced East Coast fever (theileriosis).

Skin lesions of bovine East Coast fever were examined by light and electron microscopy at 120 hours after attachment of Rhipicephalus appendiculatus ticks infected with Theileria parva. Lesions included epidermal ulcer, hemorrhage, edema, necrosis, and inflammatory cell infiltration by mostly polymorphonuclear and mononuclear (lymphoid) cells. The trophozoite state of T parva was observed in a parasitophorous vacuole and feeding on neutrophil granules. Schizonts, some in budding process, and merozoites were extracellular and among many mononuclear phagocytes (lymphoid cells). Some merozoites were in mononuclear phagocytes and a granulocyte. Some of the cells had already transformed into lymphoblasts with pseudopodia.     

Immunization of cattle with a Theileria parva bovis stock from Zimbabwe protects against challenge with virulent T.p. parva and T.p. lawrencei stocks from Kenya

Following inoculation of 34 Bos indicus (Boran) cattle with a Theileria parva bovis (Boleni) stock from Zimbabwe, 18 animals underwent mild theilerial reactions, 12 underwent moderate reactions, three suffered severe reactions and one died. When these animals were subsequently challenged with different virulent stocks of either T.p. parva (Muguga, Marikebuni or Mariakani) or T.p. lawrencei (Ngong 1 or Nanyuki) from Kenya, all except two animals resisted challenge. The two reactors were part of the group challenged with the T.p. parva (Mariakani) stock. All 12 susceptible control animals underwent severe reactions and 11 died. The results of these experiments suggest that T.p. bovis (Boleni) may be used in some situations to immunize cattle against East Coast fever without the need to provide concomitant chemotherapy as in the infection and treatment method of immunization.     

A climate-based model for the development of the ixodid tick, Rhipicephalus appendiculatus in East Coast fever zones

East Coast fever (Theileria parva infection) is an important parasitic disease of cattle in East and Central Africa. Past experimental studies have provided a great deal of information on the dynamics of the life cycle of the tick vector, Rhipicephalus appendiculatus. This paper describes a mathematical model based on field observations to explain the close relationship between the tick life cycle and climate. The model provides a basis for the future study of different ECF control programmes using computer experiments.     

Induction of immunity against infection with Theileria parva (East Coast fever) in cattle using plasma membranes from parasitized lymphoblasts

Protection against challenge with Theileria parva was conferred on three of four calves given three or four inocula of plasma membranes prepared from 6 to 12 X 10(8) autologous parasitized lymphoblasts from cultured cell lines. In contrast, calves remained susceptible to infection following immunization with membranes prepared from allogeneic parasitized lymphoblasts. Similarly, calves vaccinated with either gamma-irradiated autologous or allogeneic infected cells also died of East Coast fever after challenge. The results raise the possibility of vaccination against T. parva using subcellular material from infected lymphoblasts.     

Clinical efficacy and plasma concentrations of two formulations of buparvaquone in cattle infected with East Coast fever (Theileria parva infection)

East Coast fever, caused by the protozoan parasite Theileria parva, kills about 600,000 cattle annually in Africa. The hydroxynaphthoquinone compound buparvaquone (BPQ) is curative. Sixteen calves were infected with T. parva. On manifestation of disease symptoms, eight were injected with the original (pioneer) BPQ product and eight with a test product containing BPQ. All 16 calves were cured by one injection of 2.5 mg BPQ/kg bodyweight. The concentration of BPQ in blood plasma was monitored by HPLC. The mean observed C(max) of BPQ was 0.229 and 0.253 microg/mL of plasma, the mean observed time to reach this concentration (T(max)) was 2.62 and 2.12 h and the AUC (area under curve) was 4.785 and 4.156 microg h/mL, respectively, for the pioneer and test product. Considerable variations occurred in the plasma concentration of BPQ within each group. They showed no relationship with either clinical or parasitological parameters following treatment.     

Effect of chronic trypanosomiasis on immunization against East Coast fever

Two experiments were carried out in which uninfected cattle, or cattle chronically infected with Trypanosoma congolense, were immunized by the infection and treatment method against East Coast fever (ECF; Theileria parva infection). Chronic trypanosomiasis did not prevent cattle mounting an effective immunological response to ECF immunization and resisting subsequent lethal challenge. There appeared to be no difference in the level or quality of immunity between uninfected cattle and trypanosome-infected cattle. Thus, T. congolense infection on its own does not appear to provide a constraint to ECF immunization in the field.     

Epidemiological analysis of tick-borne diseases in Zambia

Tick-borne diseases are a constraint to livestock production in many developing countries as they cause high morbidity and mortality, which results in decreased production of meat, milk and other livestock by-products. The most important tick-borne diseases of livestock in sub-Saharan Africa are East Coast fever (caused by Theileria parva), babesiosis (caused by Babesia bigemina and B. bovis), anaplasmosis (caused by Anaplasma marginale) and heartwater (caused by Ehrlichia ruminantium). Despite their economic importance, information on the epidemiology of these diseases in many countries, including Zambia, is often inadequate, making rational disease control strategies difficult to implement. In this study 18S and 16S rRNA gene PCR assays were used for a comprehensive epidemiological analysis of tick-borne disease of cattle in three provinces of Zambia (Lusaka, Central and Eastern). All the disease pathogens under study (T. parva, T. mutans, T. taurotragi, B. bovis, B. bigemina, Anaplasma spp and E. ruminantium) were prevalent in each of the provinces surveyed. However, variation was observed in prevalence between regions and seasons. There was no association between live vaccination against East Coast fever and being PCR positive for T. parva. A number of risk factors were shown to be associated with (a) the occurrence of tick-borne pathogens in cattle and (b) cattle tick burdens in the wet season. A negative association was observed between the number of co-infecting pathogens and the erythrocyte packed cell volume (PCV) of carrier cattle.     

Transcriptional profiling of inflammatory cytokine genes in African buffaloes (Syncerus caffer) infected with Theileria parva

Theileria parva (T. parva) causes East Coast fever (ECF), which is of huge economic importance to Eastern and Southern African countries. In a previous bovine model, inflammatory cytokines were closely associated with disease progression in animals experimentally infected with T. parva. The African Cape buffalo (Syncerus caffer), the natural reservoir for T. parva, is completely resistant to ECF despite a persistently high parasitaemia following infection with T. parva. Characterizing basic immunological interactions in the host is critical to understanding the mechanism underlying disease resistance in the African Cape buffalo. In this study, the expression level of several cytokines was analyzed in T. parva-infected buffaloes. There were no significant differences in the expression profiles of inflammatory cytokines between the infected and uninfected animals despite a remarkably high parasitaemia in the former. However, the expression level of IL-10 was significantly upregulated in the infected animals. These results indicate a correlation between diminished inflammatory cytokines response and disease resistance in the buffalo.     

Skin nodules in East Coast fever.

A skin eruption developed in five calves with advanced experimental East Coast fever (ECF) (Theileria parva infection). In three cases the eruption was due to intradermal nodules containing numerous theilerial schizonts, in one animal the nodules contained abscesses filled with Demodex mites, and in the fifth calf both types of nodules were present. No nodules were seen in 15 other experimental cases of ECF, which generally died or were killed in an earlier stage of the disease. It is postulated that demodicosis may become apparent in healthy carriers of the mite in the course of ECF, because of immuno-depression caused by the latter disease.     

Comparison of indirect fluorescent antibody test and enzyme linked immunosorbent assay in the detection of exposure of cattle to Theileria parva in Kenya.

Appraisal of the indirect fluorescent antibody test (IFAT) and antigen enzyme linked immunosorbent assay (ELISA) serological tests as carried out to detect cattle exposed to Theileria parva at the National Veterinary Research Centre, Muguga (NVRC), Kenya is reported. Using sera from T. parva naive cattle and cattle experimentally exposed to T. parva, the two tests were appraised in terms of their sensitivity and specificity. IFAT and ELISA had the same sensitivity of 90% while ELISA had a higher specificity (90%) than IFAT (80%). A comparison was also made of the capability of the two tests to detect exposure of dairy cattle to T. parva prior to immunization against East Coast fever (ECF). The positive outcome from the IFAT was significantly higher (chi 2 = 30.36; P < 0.001) than that from the ELISA. The agreement between the two tests was low (Kappa = 0.21). The two tests indicated a higher risk of ECF in the study area than was expected. Indications are that the ELISA has been effectively adopted at NVRC.