The effect of buparvaquone treatment on the levels of some antioxidant vitamins, lipid peroxidation and glutathione peroxidase in cattle with theileriosis.

Plasma levels of vitamins A, E, beta carotene, both plasma and erythrocyte glutathione peroxidase (GSHPx), lipid peroxidation (LPO) and reduced glutathione (GSH) were investigated in cattle naturally infected with Theileria annulata and treated with buparvaquone. There were two groups each containing 30 cattle. Naturally infected cattle were used in the second group. Buparvaquone (2.5 mg/kg body weight) was administered to animals in the second group. Blood samples were taken from control animals, and immediately before treatment, and from animals 10 days after the injection of buparvaquone. Detection of the infected animals was carried out by blood smears. Plasma vitamins A, E, beta carotene, both plasma and erythrocyte GSHPx, LPO and GSH levels were determined. The levels of LPO in plasma and erythrocyte samples were significantly (P < 0.05, P < 0.01) higher after treatment than in either control animals or before treatment. Plasma levels of antioxidant vitamins, vitamin E and beta carotene were significantly (P < 0.05, P < 0.01) lower after treatment than in either control animals or before treatment, while the vitamin E level was found to be higher before treatment than in either the control group or animals after treatment (P < 0.05, P < 0.01). The levels of vitamin A in plasma and the activity of GSHPx and GSH in both plasma and erythrocytes in control animals after and before treatment did not differ significantly. In conclusion, we observed that there was a decreased plasma level of vitamin E and beta carotene and an increased level of LPO in cattle treated with buparvaquone. Buparvaquone might function in the treatment of Theileria annulata by forming free radicals.     

Further evaluation of the use of buparvaquone in the infection and treatment method of immunizing cattle against Theileria parva derived from African buffalo (Syncerus caffer).

Three experiments were undertaken to determine the efficacy of different doses of buparvaquone in the infection and treatment immunization of cattle against Theileria parva derived from African buffalo (Syncerus caffer). Two of these experiments also compared buparvaquone with standard doses of long- and short-acting formulations of oxytetracycline. In addition, different dilutions of stabilates were used in the experiments. In the first experiment, a 10(-1.0) dilution of stabilate was used to infect groups of cattle treated with buparvaquone at doses of between 5 and 0.625 mg kg-1 body weight (bwt) on Day 0 after infection. All control cattle developed severe theileriosis and none of the treatment regimes (including those utilizing long-acting oxytetracycline) prevented the development of theileriosis. Treatment with buparvaquone at 2.5 mg kg-1 bwt or oxytetracycline gave the most satisfactory results. In the second experiment when the sporozoite dose was reduced to 10(-2.0) dilution, buparvaquone treatment at 5 and 2.5 mg kg-1 bwt and short- and long-acting formulations of oxytetracycline reduced reactions greatly. While all the oxytetracycline treated animals produced a serological response and were immune to a 50-fold higher challenge with the immunizing stabilate, several animals in the buparvaquone groups did not show a serological response and were not immune to challenge. In the third experiment, groups of cattle were infected with 10(-1.2), 10(-1.4) and 10(-1.6) dilutions of stabilate and were treated with 2.5 mg kg-1 bwt of buparvaquone. No animals developed severe theileriosis and all seroconverted. On homologous challenge, however, two out of 14 cattle showed severe reactions. It was concluded that further work on immunization using buparvaquone treatment at 2.5 mg kg-1 bwt and 10(-1.6) dilution of the stabilate would have to be carried out before such a system could be used in the field.     

Chemotherapeutic value of parvaquone and buparvaquone against Theileria annulata infection of cattle

Parvaquone (BW993C), 2-cyclohexyl-3-hydroxy-1,4-naphthoquinone, and buparvaquone (BW720C) 2-(trans-4-t-butylcyclohexyl-methyl)-3-hydroxy-1,4-naphthoquinone, were evaluated to determine their therapeutic efficacy in the treatment of theileriosis caused by Theileria annulata infection in cattle in Iran. One hundred and fifty-nine pure and crossbred Bos taurus cattle, experimentally or naturally infected with T annulata, were treated. Parvaquone was injected into 86 animals with up to three doses of 20 mg kg-1 or 10 mg kg-1 at intervals of 48 hours between doses. Buparvaquone was injected into 73 animals. Up to three doses of 2.5 mg kg-1 were injected with an interval of 48 hours between doses. The recovery rate of animals treated with parvaquone was 60.7 per cent and with buparvaquone it was 88.7 per cent. No significant side effects of relapse of disease were observed following the use of either compound. It is concluded that buparvaquone at a dose of 2.5 mg kg-1 has a satisfactory therapeutic index and is a more effective treatment of T annulata infection than parvaquone. The prophylactic use of schizont tissue culture vaccine and chemotherapy with buparvaquone could be the most promising means of controlling theileriosis in Iran.     

Activity of buparvaquone against Theileria cervi in white-tailed deer

Buparvaquone, a naphthoquinone with known efficacy against Theileria parva parva in cattle, was tested for activity against Theileria cervi piroplasms in both an in vitro culture system and in vivo in experimentally infected white-tailed deer. The in vitro data showed a significant decrease in the incorporation of 3H-hypoxanthine by infected red blood cells treated with buparvaquone when compared to that seen with imidocarb and chloroquine treatment. In both intact and splenectomized deer treated with buparvaquone (2.5 mg kg-1) a gradual decrease in piroplasm parasitaemia was observed following treatment. However, in the splenectomized deer, parasitaemia levels returned to near pretreatment values after approximately 2 weeks.     

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.     

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.     

Treatment of theileriosis with parvaquone in Zambia

Parvaquone was used to treat 126 cattle with theileriosis. Theileria species schizonts were present in their lymph node biopsy smears and the majority of the animals had clinical signs of theileriosis. One hundred and fifteen treated and one untreated cattle survived the infection while 11 treated and 12 untreated animals died of the disease. Despite serological evidence of a parasite challenge during the subsequent rainy season, recovered cattle did not develop clinical signs of theileriosis but untreated cattle in the area continued to die from the disease. An intermittent low piroplasm parasitaemia (less than 1 per cent) was observed in recovered cattle for up to 14 months after detailed monitoring of cattle in the trial; this could be evidence for a carrier status for the Theileria species or strains involved.     

Treatment of East Coast fever: a comparison of parvaquone and buparvaquone

Two trials were conducted to compare the efficacy of parvaquone and buparvaquone for the treatment of naturally acquired East Coast fever (ECF, Theileria parva infection) which, if untreated, is almost invariably fatal in European breeds of cattle. In the first trial 28 naive cattle were exposed in a paddock infested with ticks carrying a virulent form of the disease. Twelve were treated with each drug when they developed clinical ECF. All 24 cattle were cured. In the second study, 100 cases of ECF occurring naturally on farms in Kenya were treated, 50 with each drug. Parvaquone cured 44 (88%) buparvaquone cured 45 (90%). Intercurrent infections, predominantly anaplasmosis and bacterial pneumonia or scour, were treated specifically. It is concluded that parvaquone and buparvaquone are similarly effective in curing ECF and cure rates are maximised by accurate diagnosis and prompt treatment of both ECF and intercurrent infections.     

Infectivity and cross-immunity studies of Theileria lestoquardi and Theileria annulata in sheep and cattle: I. In vivo responses.

In a series of experiments, sporozoite stabilates of a Theileria lestoquardi (Lahr) and a T. annulata (Ankara) stock prepared from Hyalomma anatolicum anatolicum ticks, were used to examine the infectivity of both parasite species for sheep and cattle and to study the development of cross-immunity between these parasite species. In the first experiment sheep and cattle were inoculated with T. lestoquardi sporozoites. Surviving animals and naive sheep and cattle were, in the second experiment, inoculated with T. annulata. In the third experiment, naive sheep and sheep previously infected with T. annulata, were inoculated with T. lestoquardi. The following responses to inoculations were monitored: clinical and haematological signs of infection, appearance of parasitic stages of the parasites in lymph node biopsies and in peripheral blood and serological response to T. lestoquardi and T. annulata schizont antigens. While T. lestoquardi readily infected sheep and caused severe disease, it did not infect cattle. On the other hand, T. annulata infected both cattle and sheep. However, whereas cattle became severely affected, infected sheep showed mild clinical symptoms only and piroplasms did not develop. Despite their different behaviour in the host species examined, cross-immunity studies suggested that the parasite species are very closely related. Experiments in sheep indicated that T. lestoquardi infection protected against subsequent T. annulata infection. On the other hand, recovery from T. annulata infection did not prevent infection by sporozoites of T. lestoquardi, resulting in the establishment of schizonts and their subsequent development into piroplasms, although it protected against the major clinical effects of T. lestoquardi infection.     

Isoenzyme studies on Theileria (Protozoa,Sporozoa). Enzyme activity associated with the erythrocytic stage

Summary

Bovine blood containing piroplasms of Theileria parva, as well as non‐injected blood, was lysed and subjected to iso‐electric focussing.

Staining for 13 different enzymes revealed parasite‐associated bands of glucose phosphate isomerase (GPI) activity, not of any of the other enzymes. There were no variations between individual donor animals in the host cell GPI bands and these bands did not interfere with the recognition of the parasite‐associated bands, so that purification of the piroplasms was unnecessary. Blood from cattle infected with T. mutans also gave parasite‐associated bands of GPI, but no such bands were seen in zymograms of blood from cattle infected with a Theileria sp. from Japan. Depending on the level of parasitaemia, up to four parasite‐associated bands were found in one strain of T. parva and up to three in two other strains. Among the disadvantages of using piroplasm material for the study of isoenzymes of T. parva is the fact that animals often die before their parasitaemia is sufficiently high, and that some strains never give rise to a high parasitaemia.     

A molecular survey of bovine Theileria parasites among apparently healthy cattle and with a note on the distribution of ticks in eastern Turkey

A survey of Theileria parasites in cattle in eastern Turkey was carried out using specific polymerase chain reaction. A total of 252 blood samples were collected from clinically healthy cattle between June and July 2004. Of 252 blood samples examined, 41 (16%) were positive for piroplasms by microscopy, whereas 114 (45%) were positive for the presence of at least one species of Theileria by PCR. The percentages of positive animals for Theileria annulata and benign Theileria species (Theileria sergenti/buffeli/orientalis) were 39% (99/252) and 7% (18/252), respectively. By allele-specific PCR examination of 18 field isolates which were positive for benign Theileria parasites, 8 samples were only amplified by B-type specific primers and 10 samples were amplified by both of the B and C-type specific primers, indicating a mixed infection with B and C-type of the parasite. None of the field isolates was amplified by I-type specific primers. Three samples were co-infected with T. annulata and benign Theileria parasites. Two of them which were infected with B-type parasite were also infected with T. annulata, the other sample which was infected both of B and C-type parasites was also infected with T. annulata. A total of 724 ixodid ticks were collected from the cattle. Hyalomma anatolicum anatolicum was the dominant species with 32% (230/724) in the region. H. a. excavatum, Boophylus annulatus and Rhipicephalus bursa represented 25% (183/724), 19% (140/724) and 15% (112/724) of the total number of ticks, respectively. R. sanguineus was the minor species and represented 8% (59/724) of the tick population.     

Occurrence of Theileria parva infection in cattle on a farm in the Ladysmith district, KwaZulu-Natal, South Africa

Theileria parva causes widespread morbidity and mortality in cattle in endemic regions. An outbreak of theileriosis occurred on a farm near Ladysmith in KwaZulu-Natal, South Africa, which is not a declared Corridor disease-infected area. A survey of Red Brangus cattle from all age groups and areas of the farm was performed. Transmission of the parasite from infected animals on the farm to susceptible animals by tick transmission and tick-stabilate injection, was attempted. The survey indicated high numbers of animals with antibody titres to T. parva but only 6 infected animals, based on real-time PCR and RLB analysis. The transmission experiments failed to transmit the parasite. The study shows the difficulty in elucidating a source of infection and determining the dynamics of new infections in a herd where multiple possible sources are present and treatment with tetracyclines has taken place.     

Elimination ofTheileria buffeli infections from cattle by concurrent treatment with buparvaquone and primaquine phosphate

Attempts (some successful) were made to eliminate Theileria buffeli infections from 23 naturally or experimentally infected, splenectomised calves. The anti-theilerial hydroxynaphthoquinone derivative buparvaquone was used either alone or in combination with primaquine phosphate. After treatment the calves were monitored for infection for up to 26 weeks. Blood films were examined for piroplasms and serum antibody levels were measured using an immunofluorescence technique. Buparvaquone alone failed to eliminate infections. Infections were eliminated from 11 of 16 calves treated twice with buparvaquone and three or six times with primaquine phosphate. Theilerial parasites were not subsequently seen in these 11 calves nor were antibodies detected beyond the eighth week after treatment.     

Treatment of experimentally induced cytauxzoonosis in cats with parvaquone and buparvaquone

The efficacy of parvaquone (Clexon) and buparvaquone (Butalex) in treating experimentally induced feline cytauxzoonosis was explored. Domestic cats were inoculated subcutaneously with blood from a cat infected with Cytauxzoon felis and treated daily with either 20 or 30 mg kg-1 parvaquone, or 5 or 10 mg kg-1 buparvaquone, beginning on either the first day parasites were detected in peripheral blood, or 2 days after the onset of parasitemia. Fifteen cats were treated and all but one died due to the infection. Unexpectedly, one of two non-treated, infected control cats survived. Although parvaquone and buparvaquone are the treatments of choice for a related hemoprotozoan parasite causing theileriosis in African cattle, wer concluded that at the dosages and regimes tested, these drugs are not effective treatments for feline cytauxzoonosis. Blood from the two surviving cats was inoculated into naive cats and in these animals clinical disease or death were not observed. The latter two naive recipient cats were then inoculated with a lethal dose of viable, frozen C. felis and both died, thereby indicating that blood from surviving cats did not induce an infectious state that resulted in immunity. The two cats that survived the acute infection were subsequently challenged with a lethal inoculum of C. felis; they showed no clinical signs of cytauxzoonosis and were obviously immune to reinfection.     

Clinical, haematological and therapeutic studies on tropical theileriosis in water buffaloes (Bubalus bubalis) in Egypt

Thirty buffaloes naturally infected with Theileria annulata and 10 parasitologically free controls were used to determine the potential clinical, haematological and therapeutic impact of tropical theileriosis in Egypt. The clinical signs in the infected buffaloes were pyrexia (40.5-41.5 degrees C), enlargement of superficial lymph nodes, slight nasal and ocular discharges, salivation, anaemia and respiratory distress. Eye lesions also were recorded. There was a significant decrease in erythrocyte counts and haemoglobin content and a significant decrease in total leucocyte counts in infected buffaloes compared to controls. Early treatment with buparvaquone was 100% effective in eliminating the protozoan parasites from the blood and lymph nodes and led to an improvement in the clinical state whereas treatment in the later stages of the disease whilst eliminating the parasites failed to improve the clinical condition of the animal.     

Molecular approaches to elucidating innate and acquired immune responses to Babesia bovis, a protozoan parasite that causes persistent infection.

For many vector-transmitted protozoal parasites, immunological control of acute infection leads to a state of persistent infection during which parasitemias may cycle unnoticed in infected but otherwise clinically healthy animals. Achieving persistent infection is a strategy that favors parasitism, since both host and, therefore, parasite survive, and endemically infected animal populations provide a reservoir of parasites continually available for subsequent transmission. Examples of the major economically important protozoan pathogens that cause persistent infection in mammals include the related Theileria and Babesia parasites as well as Trypanosoma species. Control of acute infection and maintenance of clinical immunity against subsequent infection are determined by the interplay of innate and acquired immune responses. This review will focus on approaches taken to gain an understanding of the molecular basis for innate and acquired immunity against the hemoprotozoan parasite of cattle, Babesia bovis. Knowledge of mechanisms used by the parasite to survive within infected cattle from acute to persistent infection combined with definition of the correlates of protective immunity in cattle should be applicable to designing effective vaccines.     

Incidence and prevalence of tick-borne haemoparasites in domestic ruminants in Ghana

Giemsa-stained thin blood smears prepared monthly from cattle, sheep and goats in the Greater Accra region of Ghana between May 1994 and December 1996 were examined for presence of tick-borne haemoparasites. The majority of animals were less than 2 months old at the start of the survey. Monthly and cumulative incidences are presented of Anaplasma sp., Babesia bigemina, Borrelia sp., Eperythrozoon sp., Theileria mutans and Theileria velifera in cattle, Anaplasma sp., Borrelia sp., and Theileria sp. in sheep, and Anaplasma sp. in goats. T. mutans was the commonest parasite in cattle, with 100% incidence in calves by 10 months of age, and Anaplasma was commonest in small ruminants. The relative prevalence of these haemoparasites in blood smears from cattle, sheep and goats sampled on a single occasion at sites in all 10 regions of Ghana was found to be similar, though actual infection rates were lower. Packed cell volume (PCV) measurements from the sampled animals are also presented; no seasonal trends were evident in the PCV of the cattle, sheep and goats sampled monthly. In animals sampled on a single occasion, mean PCV was significantly higher in cattle and sheep without detectable haemoparasite infection, and in cattle was lowest in animals positive for both Babesia and Anaplasma, while there was no difference in mean PCV levels between parasitised and non-parasitised goats.     

Theileria buffeli infection of a Michigan cow confirmed by small subunit ribosomal RNA gene analysis

Theileria buffeli, generally a benign parasite of cattle, has been reported in animals from Texas, Missouri and North Carolina. To date, there have been no reports of the parasite in cattle residing in northern portions of the US. An 8-year-old cow (Maine Anjou x Angus cross-bred) in Michigan presented with hemoglobinuria and a packed cell volume of 9. Blood films stained with Giemsa showed numerous intraerythrocytic parasites morphologically consistent with T. buffeli. The parasite was confirmed to be T. buffeli by SSU rRNA gene sequence analysis (SSU rRNA sequence, Type A). This represents the first report of this parasite in an animal in Michigan.     

Transformation of Theileria parva derived from African buffalo (Syncerus caffer) by tick passage in cattle and its use in infection and treatment immunization.

A sporozoite stabilate (St. 199) of Theileria parva was obtained by feeding nymphal Rhipicephalus appendiculatus on an African buffalo (Syncerus caffer) and was used to immunize cattle by the infection and treatment method. Nymphal ticks were applied to one of the steers 90 days later and it was shown that the resultant adult tick had become infected. Using tick/cattle passage, two passage lines of T. parva were established. By the fifth tick/cattle passage, the parasite stocks had changed their behaviour to that of T. parva derived from cattle as the parasite produced relatively high schizont parasitosis and piroplasm parasitaemia in cattle, and had become highly infective to ticks. At various passage levels the parasite populations were characterized by behaviour and by monoclonal antibodies against T. parva schizonts using infected cell culture isolates from cattle during acute infections. The monoclonal antibody profile showed little evidence of antigen change of the parasite during passage through cattle, which was confirmed in a two-way cross-immunity experiment using sporozoite stabilate derived from ticks obtained from the buffalo and fourth passage in cattle. The implication of these results, particularly in relationship to immunization of cattle against T. parva derived from buffalo, is discussed.     

Evaluation of buparvaquone (BUTA-Kel KELA, Belgium) as a treatment of East Coast fever in cattle, in the peri-urban of Dar Es Salaam city, Tanzania

Evaluation trials of the efficacy of buparvaquone (BUTA-kel KELA Laboratoria, N.V. Belgium), as a treatment of field cases of Theileria parva infection (East Coast fever - ECF) were carried out on 63 cattle in the peri-urban of Dar Es Salaam city, Tanzania, during the period November 2004 to August 2005. Thirty-two cattle (56%) received single-dose treatment (2.5 mg buparvaquone per kg body weight), while two and three-dose treatment with interval(s) of 48 h was given to 33% and 11% of total treated cattle, respectively; 38 cattle (60.3%) were treated at an early stage of the disease, while 25 cattle (39.7%) were treated at an advanced stage of the disease. The rectal body temperature of 90.5% of buparvaquone-treated cattle dropped to normal values (37.5-39.5 degrees C) by day 7 of treatment, and by day 15 of treatment 96.8% of treated cattle showed normal values. Pulmonary signs were observed in 8/68 (11.8%) of total ECF diagnosed cattle and were successfully treated, albeit with parvaquone plus frusemide (Fruvexon); were not included in final evaluation of the efficacy of BUTA-kel. The present evaluation trials record a recovery rate of 95.2%. Buparvaquone (BUTA-kel KELA Laboratoria, N.V. Belgium), therefore, records another efficacious and valuable alternative treatment against East Coast fever in Tanzania.