Tropical theileriosis in Tunisia: epidemiology and control

In Tunisia, tropical theileriosis (Theileria annulata infection of cattle) is one of the major diseases affecting cattle in the summer. Each year about 2,500 clinical cases are recorded in the country, mainly in pure-bred animals. An attempt was made in the North of Tunisia to estimate the direct cost related to clinical cases of TT. In the endemic regions, three states of endemicity were described according to cattle age categories at highest disease risk: (i) endemic stability, (ii) low endemic instability, (iii) high endemic instability. The characterisation of these endemic situations, which are conditioned by the levels of the vector tick population and the quantitative aspects of the infection in ticks, are highly relevant for the development of a live attenuated cell line vaccine against TT targeting the cattle population at disease risk in Tunisia. A research programme was set up to develop the attenuation of four local parasite stocks. Two cell lines infected with two distinct T. annulata stocks were tested in the field on pure-bred animals of different age groups from regions with high disease incidence. The vaccination with each of the two cell lines showed a high efficacy. However significant differences in vaccine reactions rates were observed between the two attenuated cell lines emphasising the importance of achieving an optimal balance between protection and vaccine tolerance particularly in pure-bred lactating cows.     

Detection of Theileria annulata by the PCR in ticks (Acari: Ixodidae) collected from cattle in Mauritania

We report on the detection of Theileria annulata in infected Hyalomma ticks by the PCR using primers derived from the gene encoding the 30 kDa major merozoite surface antigen (Tams1–1). No inhibition of the PCR was observed and as little as 0.1 pg of parasite DNA, corresponding to 12 sporozoites, could be detected in non-infected tick DNA samples, spiked with T. annulata genomic DNA. Hyalomma dromedarii ticks, fed on a calf experimentally infected with T. annulata, were used to validate the PCR further. The infection rate in the adult ticks, fed as nymphs during the febrile reaction, was high (62%), dropped to zero for 1 day in tick batches that engorged after treatment with ButalexTM and increased to 30% 2 days later and 38% of the ticks acquired the infection after feeding as nymphs during a carrier state piroplasm parasitaemia of less than 0.1%. As an internal control, 16S tick rDNA sequences could be amplified from T. annulata-negative tick samples. Finally, 202 adult ticks from Mauritania, collected from zebu cattle carrying low levels of Theileria piroplasms, were tested by the PCR. Thirty-eight out of 52 (73%) and 17 out of 30 (57%) H. dromedarii from the Gorgol and Trarza regions, respectively and two out of 30 (7%) Hyalomma marginatum rufipes from the Gorgol region were positive. Hyalomma marginatum rufipes, Rhipicephalus evertsi evertsi and Rhipicephalus guilhoni from the Trarza region were negative. These findings confirm that H. dromedarii is the main vector of T. annulata in Mauritania and that the PCR is a useful method of determining the infection rates in ticks collected from cattle carrying low levels of T. annulata piroplasms.     

Treatment of natural tropical theileriosis with the extract of the plant Peganum harmala

Theileria annulata, a protozoan parasite of cattle and domestic buffaloes, is transmitted by ticks of the genus Hyalomma, and causes a disease named Mediterranean or tropical theileriosis. In this research 50 cattle naturally infected with Theileria annulata were treated with the extract of the plant Peganum harmala. The treatment was continued for 5 days, the dose of the extract being 5 mg/kg per day. After the treatment, 39 cattle responded to the treatment and recovered, but 11 did not respond to the treatment and died. The recovery rate of animals treated with the extract of the plant Peganum harmala was 78%.     

Modelling the transmission dynamics of Theileria annulata: model structure and validation for the Turkish context

A mathematical model that describes the transmission dynamics of Theileria annulata is proposed that consists of 2 host components: the Hyalomma tick population and a compartmental model of T. annulata infection in the cattle population. The model was parameterized using data describing tick infestation and the infection status of cattle in Turkey from 2006 to 2008. The tick attachment rates are highly seasonal and because of the temporal separation of infectious and susceptible ticks virtually all ticks are infected by carrier cattle, so that annual peaks of disease in cattle do not impact on infection in the Hyalomma tick population. The impact of intervention measures that target the tick population both on the host and in the environment and their impact on the transmission of T. annulata were investigated. Interventions that have a limited 'one-off' impact and interventions that have a more permanent impact were both considered. The results from the model show the importance of targeting ticks during the period when they have left their first host as nymphs but have yet to feed on their second host.     

Control of Theileria annulata in Iran

Tropical theileriosis or Mediterranean Coast Fever - caused by Theileria annulata - is a disease of cattle widely distributed across southern Europe, north Africa and central Asia. Its distribution broadly corresponds with that of its main ixodid tick vectors Hyalomma excavatum and H. detritum (Fig. 1). 'Exotic' cattle (Bos taurus) are particularly susceptible with mortalities up to 40-80% in some areas, whereas indigenous cattle (B. indicus) generally suffer much lower mortalities (about 10%) confined mainly to calves. But because imported non-immune cattle are so susceptible, T. annulata represents a major constraint to livestock improvement programmes in many parts of the middle east and Asia. Cattle that recover from T. annulata infection generally show a solid, long-lasting immunity. For many years there have been programmes to protect cattle by inoculation with blood from sick animals, and more recently using live attenuated T. annulata vaccines prepared from cultured schizont-infected lymphoid cells. This article reviews a 14 year immunization programme against T. annulata in Iran.     

Antigens and immunity in Theileria annulata

About 200 million cattle are believed to be at risk from the debilitating and often fatal effects of tropical theileriosis, caused by Theileria annulata. Currently, there is no very cheap effective drug for treatment of T. annulata infections, although the hydroxynophthoquinones parvaquone and buparvaquone are reported to give good results(1-4). Control of the parasite principally involves vector control against the ixodid tick vectors - mainly by cattle dipping and spraying with acaricides - and vaccination using attenuated macroschizont-infected leucocytes (1-17) (see Box I). In this article, Roger Hall discusses the nature of immunity that can be achieved against T. annulata, and progress in identifying the main antigens involved in this immunity.     

Two initial vaccinations with the Bm86-based Gavacplus vaccine against Rhipicephalus (Boophilus) microplus induce similar reproductive suppression to three initial vaccinations under production conditions

Background

The cattle tick, Rhipicephalus (Boophilus) microplus, affects livestock production in many regions of the world. Up to now, the widespread use of chemical acaricides has led to the selection of acaricide-resistant ticks and to environmental contamination. Gavac^plus is a subunit vaccine based on the recombinant Bm86 tick antigen expressed in yeast, capable to control infestations of R. microplus under controlled and production conditions. The vaccine constitutes the core element of broad control programs against this ectoparasite, in which acquired immunity in cattle to Bm86 is combined with a rational use of acaricides. At present, the conventional vaccine scheme consists of three doses that should be administered at weeks 0, 4 and 7, followed by a booster every six months.

Results

In this study we assayed a reduction in the number of the initial doses of Gavac^plus, evaluated the time course and the level of bovine anti-Bm86 antibodies elicited, and analyzed the vaccine effect on ticks engorging on immunized cattle under production conditions. Following three different immunization schemes, the bovines developed a strong and specific immune response characterized by elevated anti-Bm86 IgG titers. A reduction in the weight of engorging female ticks, in the weight of the eggs laid and also in R. microplus viable eggs percentage was obtained by using only two doses of Gavac^plus administered at weeks 0 and 4, followed by a booster six months later. This reduction did not differ from the results obtained on ticks engorging on cattle immunized at weeks 0, 4 and 7. It was also demonstrated that anti-Bm86 antibody titers over 1:640, measured in bovines immunized at weeks 0 and 4, were sufficient to affect weight and reproductive potential of female ticks as compared with ticks engorging on unvaccinated animals. In addition, no statistically significant differences were detected in the average weight of eggs laid by ticks engorged on immunized cattle that showed anti-Bm86 specific titers in the range of 1:640 to 1:81920.

Conclusion

The administration of two initial doses of Gavac^plus containing 100 μg of Bm86 antigen to non-immunized cattle under production conditions is sufficient to affect the weight and the reproductive capacity of R. microplus engorging females. According to these results, cattle herds' manipulation and vaccine costs could be potentially reduced with a positive impact on the implementation of integrated control programs against R. microplus.     

Development of recombinant antigen vaccines for the control of theileriosis

Immunization against Theileria parva involves infection with sporozoites and simultaneous treatment with a long-acting tetracycline. For T. annulata, immunization is achieved by inoculation of attenuated schizont-infected lymphocytes. The two methods are inadequate because of the use of live organisms and the methods are also bedevilled by the multiplicity of strains, particularly of T. parva. For these reasons, alternative methods of control are being sought. In this review an attempt is made to highlight advances towards subunit vaccines against T. parva and T. annulata. Several candidate antigens which are thought to induce protective responses have been identified and recombinant DNA technology is being employed to produce these antigens in bulk. Relevant antigens may be delivered as subunit vaccines by using recombinant vaccinia virus or attenuated Salmonella spp. as carriers of the genes expressing these antigens. It is likely that effective vaccines against T. parva and T. annulata will have to elaborate immune responses against both the sporozoite and schizont stages of the parasite.     

Bovine babesiosis: Cattle protected in the field with a frozen vaccine containing Babesia bovis and Babesia bigemina cultured in vitro with a serum-free medium

An attenuated live vaccine containing Babesia bovis and B. bigemina cultured in vitro with a serum-free medium was assessed for its clinical protection conferred of naïve cattle, under natural tick-challenge in a high endemicity zone to Babesia spp. Three groups of six animals were treated as follows: group I (GI) received a vaccine derived from parasites cultured with a free-serum medium; group II (GII) were immunized with the standard vaccine, with parasites cultured in a medium supplemented with 40% (v/v) bovine serum; and a control group (GIII) inoculated with non-infected bovine erythrocytes. Inocula were administered by IM route. Experimental animals were kept during 23 days after vaccination in a cattle farm free of ticks and Babesia spp. Thereafter, cattle were moved to a high endemicity farm for natural exposure to Babesia spp. transmitted by Rhipicephalus microplus ticks. Protection against clinical babesiosis was observed in bovines belonging to GI (100%) and GII (83.33%), while the control animals (GIII) were not protected, and showed severe clinical signs, closely related to babesiosis, were observed for at least three consecutive days during the challenge. These were fever, anemia, which were measured simultaneously, and circulating parasites were detected by optic light microscopy. All cattle showed B. bovis and B. bigemina in stained blood films during the challenge; B. bovis antibody titers were higher than those to B. bigemina in GI and GII, and lower titers were determined in GIII. The protective capacity of the vaccine derived from B. bovis and B. bigemina cultured in vitro in a serum-free medium was demonstrated.     

Preliminary studies on the effect of Anaplasma marginale antibodies ingested by Dermacentor andersoni ticks (Acari:Ixodidae) with their blood meal on infections in salivary glands

The effect of Anaplasma marginale antibodies ingested with the tick blood meal was tested on infected male ticks that were allowed to feed on cattle immunized with the erythrocytic stage of A. marginale. The experiments were done in two trials. Trial 1 was done using splenectomized calves (two calves per treated and control groups) while ticks in trial 2 were fed on intact yearling cattle (four cattle per treated and control groups). The cattle were immunized with purified outer membrane proteins of erythrocyte-derived A. marginale using saponin (trial 1) or monophosphoryl lipid-A-trehalose dicorynomycolate adjuvant (trial 2). The corresponding control cattle received adjuvant only. All cattle were challenged using Dermacentor andersoni males infected as adults that were allowed to feed for 7 days. In trial 1, the ticks were allowed to feed a second time on susceptible calves to test whether exposure of ticks to immunized cattle affected their ability to transmit anaplasmosis. Infections in fed ticks were monitored by determining the infection rates in salivary glands with an A. marginale-specific RNA probe and light microscopy. Vaccine-derived antibodies ingested with the tick blood meal did not appear to affect the development of A. marginale in previously infected ticks. The infection rates in the salivary glands were not significantly different among ticks fed on immunized versus adjuvant control cattle. When the vaccine-exposed ticks in trial 1 were allowed to feed a second time on susceptible calves, the resulting clinical symptoms of anaplasmosis were similar to those of the controls. There was no statistically significant effect of tick exposure to the anti-erythrocytic stage antibody on the development of salivary gland infection or transmission of A. marginale by ticks.     

Studies on theileriosis in Southern Spain

Mediterranean theileriosis is widely distributed throughout Southern Spain, where it causes economic losses of considerable importance. In recent studies on 221 fighting bulls from 29 locations of different provinces, mainly in Southern Spain, the seroprevalence was 75.1% by IFAT. On the other hand, we are carrying out studies on beef and dairy breeds from eight provinces in the southern half of Spain where the positivity by IFAT ranges from 50 to more than 90% of the sampled cattle. Studies on the isolation of T. annulata strains, the demonstration of the vectorial role of some tick species, and cattle immunisation, have been carried out as well. In order to isolate the causative agent of the disease, infected blood or biopsies of lymph nodes from infected animals were used to establish in vitro cultures of T. annulata-infected lymphoblastoid cells, in six different cell lines. Attenuation of the parasites was achieved by growth in successive passages of a cell line by in vitro culture. Nowadays, a tissue culture vaccine has been developed from a local strain which is tried in experimental calves and under field condition with successful results. Besides, some studies on the transmission of T. annulata to cattle by the tick Hyalomma lusitanicum have been done.     

Could insecticide‐treated cattle reduce Afrotropical malaria transmission? Effects of deltamethrin‐treated Zebu on Anopheles arabiensis behaviour and survival in Ethiopia

Anopheles arabiensis Patton (Diptera: Culicidae) is the most widespread vector of malaria in the Afrotropical Region. Because An. arabiensis feeds readily on cattle as well as humans, the insecticide-treatment of cattle--as employed to control tsetse (Diptera: Glossinidae) and ticks (Acari: Ixodidae)--might simultaneously affect the malaria vectorial capacity of this mosquito. Therefore, we conducted field experiments in southern Ethiopia to establish whether Zebu cattle (Bos indicus L.) treated with a pour-on pyrethroid formulation of 1% deltamethrin, widely used to control ticks and tsetse, would be effective against An. arabiensis or cause the female mosquitoes to feed more frequently on humans, due to behavioural avoidance of insecticide-treated cattle. Contact bioassays (3 min exposure) showed that the insecticide remained effective for about 1 month (kill rate > 50%) against mosquitoes feeding on the flanks of treated cattle. A novel behavioural assay demonstrated that An. arabiensis readily fed on insecticide-treated cattle and were not deflected to human hosts in the presence of treated cattle. DNA-fingerprinting of bloodmeals revealed that An. arabiensis naturally feeds most frequently on older animals, consistent with the established practice of applying insecticide only to older cattle, while allowing younger untreated animals to gain immunity against infections transmitted by ticks. These encouraging results were tempered by finding that > 90% of An. arabiensis, An. pharoensis and An. tenebrosus females feed on the legs of cattle, farthest from the site of pour-on application along the animal's back and where the treatment may be least residual due to weathering. Observations of mosquitoes feeding naturally on insecticide-treated cattle showed that the majority of wild female anophelines alighted on the host animal for less than 1 min to feed, with significantly shorter mean duration of feeding bouts on insecticide-treated animals, and the effective life of the insecticide was only 1 week. Thus the monthly application of deltamethrin to cattle, typically used to control tsetse and ticks, is unlikely to be effective against An. arabiensis populations or their vectorial capacity. Even so, it seems likely that far greater impact on anopheline mosquitoes could be achieved by applying insecticide selectively to the legs of cattle.     

Immunoprophylaxis of Dermatophytosis in Animals

Dermatophytosis is a relatively common disease in many countries occurring endemically both in companion and food animals. Fungi belonging to the genera Trichophyton and Microsporum are most often isolated from clinical cases. Measures to control and prevent dermatophytosis include sanitation, hygienic measures and treatment. In some countries, successful control and eradication have been achieved by mass vaccination of cattle and fur-bearing animals. Vaccines containing live attenuated cells of the fungus stimulate a cell-mediated immune response conferring long-lasting protection against subsequent challenge by the homologous fungus. A delayed type hypersensitivity (DTH) skin test using appropriate dermatophyte antigens is suitable to assess the response. Inactivated dermatophyte vaccines are available for use in cattle, horse, dog, and cat in some countries. However, the scientific literature is scarce making it difficult to conclude on efficacy and appropriate use. Current vaccines are all first generation vaccines. Attempts have been made to prepare subunit vaccines based on new knowledge about virulence factors like the keratinases, so far with limited success. Candidate antigens must be able to stimulate a strong T helper 1 cell response and future research should focus on identification of major T-cell epitopes that specifically elicit a DTH reaction. Dermatophytosis is a zoonotic disease. In Norway and a few other countries, systematic vaccination against cattle ringworm has almost eliminated the disease, and ringworm in man caused by T. verrucosum is almost nonexistent. A similar benefit could be expected if a safe and efficacious vaccine was available for Microsporum canis infection in cats and dogs.     

Systemic activity of closantel for control of lone star ticks,Amblyomma americanum (L.), on cattle

Cattle were treated once at 5 mg/kg orally or subcutaneously or daily at 0.1–5 mg/kg orally or 0.1–1 mg/kg subcutaneously with closantel, N-[5-chloro-4-[(4-chlorophenyl) cyanomethyl]-2-methylphenyl]-2-hydroxy-3,5-diiodobenzamide, and numbers and weights of engorged females, weights of egg masses and hatch of eggs of lone star ticks,Amblyomma americanum, were recorded.Effectiveness of treatments on reproduction was determined by comparing total estimated larvae (EL) (EL=wt. egg mass×est. % hatch×20000) or ticks from treated cattle with that of ticks from untreated cattle. With certain treatments, we also determined the effect of manure of treated cattle on survival of larvae of the horn fly,Haematobia irritans, or effect on survival and of fecundity of adult horn flies or stable flies,Stomoxys calcitrans, fed on blood from treated animals.The single oral treatment afforded essentially complete control of total EL only of ticks placed on the animal on the day of treatment, while the single subcutaneous treatment afforded >92% control of total EL of ticks placed on animal on treatment day and for 6 weeks posttreatment. Daily treatments of 0.5 mg/kg or greater orally and 0.1 mg/kg or greater subcutaneously afforded essentially complete control of total EL of ticks throughout the treatment period (3–12 weeks) and for 1–7 weeks after treatment was discontinued. An estimated concentration of >9 g/ml of blood was calculated by probit analysis to be necessary to provide >90% control of total EL of lone star ticks; that same concentration also provided >90% control of hatch of eggs laid by treated females. A higher concentration (40 g/ml) was necessary to prevent engorging of the females. No treatments tested were effective against larvae of the horn fly or adult horn flies or stable flies.     

Control of lymphoproliferation by Theileria annulata

The economic importance of bovine theilerioses has prompted several new approaches to understanding the diseases in the hope of developing more efficient methods of control. Most Theileria species that infect cattle cause a lymphoproli ferative disease. Sporozoites, injected into the host bloodstream by the tick vectors, rapidly invade host lymphocytes and stimulate rapid division of infected cells. As these rupture, merozoites are released which invade red blood cells ready to infect feeding ticks again. The process by which Theileria parasites can control host lymphocytes, and induce them to divide in synchrony with the parasites themselves, is poorly understood but seems to be the key to pathogenesis. In this article, Michael Dyer and Andrew Tait discuss the possible mechanisms of cellular control in the light of recent work revealing sequences homologous to oncogenes in the DNA of T. annulata.     

Feeding of Ticks on Animals for Transmission and Xenodiagnosis in Lyme Disease Research

Transmission of the etiologic agent of Lyme disease, Borrelia burgdorferi, occurs by the attachment and blood feeding of Ixodes species ticks on mammalian hosts. In nature, this zoonotic bacterial pathogen may use a variety of reservoir hosts, but the white-footed mouse (Peromyscus leucopus) is the primary reservoir for larval and nymphal ticks in North America. Humans are incidental hosts most frequently infected with B. burgdorferi by the bite of ticks in the nymphal stage. B. burgdorferi adapts to its hosts throughout the enzootic cycle, so the ability to explore the functions of these spirochetes and their effects on mammalian hosts requires the use of tick feeding. In addition, the technique of xenodiagnosis (using the natural vector for detection and recovery of an infectious agent) has been useful in studies of cryptic infection. In order to obtain nymphal ticks that harbor B. burgdorferi, ticks are fed live spirochetes in culture through capillary tubes. Two animal models, mice and nonhuman primates, are most commonly used for Lyme disease studies involving tick feeding. We demonstrate the methods by which these ticks can be fed upon, and recovered from animals for either infection or xenodiagnosis.     

The Ivomec SR Bolus for control of Boophilus annulatus (Acari: Ixodidae) on cattle in South Texas.

When Hereford heifers infested with Boophilus annulatus (Say) were treated with a single Ivomec SR Bolus, the concentration of ivermectin in the serum of the treated cattle reached a maximum of 8.8 +/- 0.9 ppb at 2 wk posttreatment. The single bolus treatment resulted in 84.4% control of standard engorging B. annulatus females on treated cattle over the 20-wk trial. Although fewer engorged ticks were collected from the sentinel heifers exposed in the treated pasture than those in the control pasture at weeks 4, 10, and 16 posttreatment, none of the differences was statistically significant. Each exposure of sentinel cattle found free-living ticks in both the treated and control pastures, indicating the infestation was not eliminated by the treatment. When the trial was repeated using two Ivomec SR Boluses/heifer, the concentration of ivermectin in the serum of the treated cattle reached a maximum level of 31.2 +/- 3.9 ppb at week 13 posttreatment. The use of two boluses/heifer resulted in 99.6% control of standard engorging B. annulatus females over the 20-wk trial. No ticks were found on sentinels placed in the treated pasture after week 9 posttreatment, an indication that the treatment had eliminated the free-living population in the treated pasture. From these studies, we conclude that a single Ivomec SR Bolus is incapable of sufficient control of B. annulatus to meet the rigid requirements of the Cattle Fever Tick Eradication Program in South Texas. Although two boluses per animal did eliminate the ticks from treated heifers and the pasture they were in, the treatment would not be sufficiently efficacious for mature cattle (>400 kg) for it to be useful in the program.     

Immunity against Boophilus annulatus induced by the Bm86 (Tick-GARD) vaccine

Friesian cattle were immunized with two inoculations of anti-tick Bm86 (Tick-GARD) vaccine and were challenged 30 or 90 d later with Boophilus annulatus larvae derived from 1.2 g of eggs. No nymphs or adult ticks were found on the immunized cattle during four weeks after challenge. Repeated infestations (2 to 4) with larvae on three other calves during a period of 160 and 390 d after the immunization did not result in development of nymphal and adult stages. In control, non-immunized cattle infested with corresponding batches of larvae 1380 to 4653 replete adult female ticks were collected. Larvae issued from Babesia bovis-infected female ticks transmitted the infection to Bm86-immunized cattle, but the progeny of B. bigemina-infected females did not. Since B. bigemina is transmitted exclusively by nymphal stages of Bo. annulatus these results support the observation that immunity induced by Bm86 affects the larval stage of this tick.     

Neospora caninum - How close are we to development of an efficacious vaccine that prevents abortion in cattle?

Neospora caninum is a protozoan parasite that causes abortion in cattle around the world. Although the clinical signs of disease in both dogs and cattle have now been recognised for over 20 years, treatment and control options are still limited, despite the availability of a commercial vaccine in some countries of the world. The case for an efficacious vaccine has not been convincingly waged by farmers, veterinarians and other members of the agricultural and rural communities. In recent times, however, economic modelling has been used to estimate the industry losses due to Neospora-associated abortion, providing, in turn, the business case for forms of control for this parasite, including the development of vaccines. In this review, we document progress in all areas of the vaccine development pipeline, including live, killed and recombinant forms and the animal models available for vaccine evaluation. In addition, we summarise the main outcomes on the economics of Neospora control and suggest that the current boom in the global dairy industry increases the specific need for a vaccine against N. caninum-associated abortion.