Theileria parva: In Vitro Studies on the Effects of Holding Temperature, pH and Medium on Sporozoite Infectivity (original) (raw)
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A recombinant sporozoite surface antigen of Theileria parva induces protection in cattle
Proceedings of the …, 1992
At present immunization against Theileria parva is by infection with live sporozoites and simultaneous treatment with a long-acting oxytetracycline. This method has major limitations in that live organisms are used and the immunity engendered is parasite stock specific. In an attempt to develop an alternative immunization procedure, the gene encoding p67, a major surface antigen of sporozoites, has been expressed by using the plasmid expression vector pMG1. The gene, which has been characterized previously, encodes 709 amino acid residues, contains a single intron of 29 base pairs, and is only transcribed during sporogony. The recombinant p67 sequences were fused to the first 85 amino acid residues derived from a nonstructural gene (NS1) of influenza virus A. Immunization with a partially purified recombinant antigen emulsified in 3% saponin induced sporozoite neutralizing antibodies in cattle and provided protection in six of nine animals on homologous challenge with T. parva sporozoites. This recombinant antigen is therefore a candidate for development of a vaccine against T. parva. Over 20 million cattle in eastern, central, and southern Africa are at risk of contracting theileriosis caused by the protozoan parasite Theileria parva (1). The disease is of major economic importance because of the high morbidity and mortality it causes and the expensive measures used to control the tick vector (1, 2). T. parva causes East Coast fever (ECF) and is transmitted by the three-host tick Rhipicephalus appendiculatus. The sporozoite develops in the salivary glands of the vector and is introduced into the mammalian host during tick feeding. Sporozoites rapidly enter lymphocytes by a recep
Tropical animal health and production, 2001
An in vitro infectivity assay was used to examine five cryoprotectants for their suitability for preserving Theileria parva sporozoites. All five were capable of preserving T. parva sporozoites through freezing, the optimal concentrations being 7.5% for glycerol, 5% for dimethyl sulphoxide (DMSO), poly (vinylpyrrolidone) (PVP) and poly(ethylene glycol) (PEG), and 2.5% for hydroxyethyl starch (HES). When the five cryoprotectants were compared at their optimal concentrations, using a modification of the standard method of stabilate preparation, glycerol was significantly better than the others (p < 0.05). Measurement of the effects of each cryoprotectant on the osmolality of the media revealed that glycerol and DMSO elevated the osmolality significantly (p < 0.05). Resuscitation of glycerol-preserved sporozoites required the presence of glycerol in the diluent to maintain infectivity. Studies on the effects of equilibration time in glycerol on the infectivity of sporozoites show...
Iranian Journal of Parasitology
Background: The protozoan parasite Theileria annulata is the causative agent of tropical theileriosis in cattle. Vaccination is recommended by administration of attenuated schizont-infected cell lines. The expected protective immunity post-vaccination can be demonstrated by challenge test through inoculation of highly virulent infective sporozoites. The aim of this study was to produce Hyalomma anatolicum anatolicum tick infected with T. annulata (local strain) for preparation of tick-derived sporozoite stabilates for molecular characterization and infectivity test assay. Methods: A local T. annulata strain was used for experimental infection of calves. A field isolate of H. a. anatolicum was isolated, laboratory-reared and infected by blood-feeding on Theileria infected above-mentioned calves. The infectivity of calf, tick and prepared stabilate were confirmed by clinical signs of theileriosis, microscopic inspection, RT-PCR and in vitro cell culture. Results: The tick stabilate wa...
Vaccines against Theileria parva
Annals of the New York Academy of Sciences, 2006
A BSTRACT : Bovine theileriosis caused by Theileria parva continues to be a major economic problem in many parts of Eastern, Southern, and Central Africa. Due to the unsustainable nature of the present control method-using toxic acaricides to kill ticks-alternative control methods are being sought. Live vaccines are being used in many countries in the region. These vaccines are based on the infective sporozoite stage of the parasite. Sporozoites are inoculated in cattle with simultaneous administration of a long-acting formulation of oxytetracycline. These vaccines are poorly adopted in the region, mainly because of problems associated with the use of live parasites. An experimental recombinant vaccine based on a sporozoite surface antigen (p67) has been developed. Immunization with this antigen induces neutralizing antibodies and, under laboratory conditions, this technique protects approximately 70% of the immunized cattle to a defined needle challenge. The efficacy of the vaccine is currently being evaluated under field challenge in Kenya. Since a vaccine based on a single antigen may not be sustainable under field conditions, a search for schizont antigens that induce protective cell-mediated immune responses continues. It is expected that the ultimate vaccine against theileriosis will incorporate a mixture of several antigens derived from both sporozoite and schizont stages, contributing to robust immunity. a Address for correspondence. Voice: +254-2-630743; fax: +254-2-631499. S.Morzaria@cgiar.org
In vitro titration of Theileria parva tick derived stabilates
Parasitology, 2004
Immunization against the protozoan Theileria parva by infection and treatment has proved to be very efficient for the control of East Coast fever, an acute and often-fatal lymphoproliferative tick-borne disease of cattle in Eastern, Central and Southern Africa. The immunizing dose of live T. parva sporozoites used in this method is usually determined by in vivo titration. An alternative in vitro method of quantification of sporozoites in whole tick-derived stabilates is proposed. The method consists of incubating serially diluted T. parva stabilates with bovine peripheral blood lymphocytes, the host cell that is infected naturally. Allowing the cultures to incubate undisturbed for the full cultivation period (10 days) reduced the variability among replicate titrations. Fungal contaminations were avoided by centrifuging stabilates at 400 g prior to the incubation, which did not precipitate sporozoites significantly. Fungistatics, Nystatin and Flucytosine, did not appear to interfere with the in vitro development of T. parva but their effect on fungal growth was limited. In vitro titration data were compared to in vivo infection data for 2 stabilates. In vitro titration of T. parva sporozoites should allow more ethical and efficient research on the preparation and storage of T. parva tick-derived stabilates.
Annals of the New York Academy of Sciences, 1998
Three vaccine stabilates of Theileria parva, of which sporozoites are being used against East Coast fever, were characterized by immunological and molecular biology techniques before being used for a national vaccination campaign in Kenya. T. parva Marikebuni stabilates 316 and 3014, and T. parva Lanet were used in this study and were discriminated from other Kenyan field Theileria isolates. IFAT results showed that all the animals were producing antibodies regardless of the stock used. Primers designed on the TPR1 gene sequence were used for PCR and Decamers were used for RAPD. Specific DNA band patterns (1,877 bp; 1,059 bp, and 443 bp) for the three vaccine stocks were observed. These molecular markers could be used to trace vaccinated animals in Kenya and to identify which isolates are responsible for reactions in animals.
2006
The sporozoite surface antigen, SPAG-1 and the attenuated schizont infected Tunisian line CL1 of Theileria annulata have been shown, in previous studies, to induce variable levels of protection against homologous and heterologous sporozoite challenge, respectively. We report here the result of a vaccination trial comparing the protection level induced by the SPAG-1 antigen (as a recombinant full length His tagged protein) and the attenuated cell line, used singly or in combination. The results, after challenge of immunised calves with a lethal dose of sporozoites, show that SPAG-1 provides limited protection (one out of seven calves surviving), while the attenuated cell line provides moderate protection (three out of seven calves recovered). The combination of SPAG-1 and the attenuated cell line induced the best protection as indicated by the survival of all the vaccinated calves. These results, together with a range of parasitological and clinical parameters, demonstrate the enhanced protection provided by combining sporozoite and schizont antigens in vaccination against tropical theileriosis.