Search

Vaccines based on recombinant Bm86 gut antigen from Boophilus microplus are a useful component of integrated control strategies against B. microplus infestations of cattle. The capacity of such vaccines to control heterologous infestations by two African tick species was investigated. The mean weight of engorged female ticks and mean egg mass per tick were significantly reduced in B. decoloratus infestations, but there was no effect of the vaccine against adult Rhipicephalus appendiculatus. We cloned, sequenced and expressed two Bm86 homologues (Bd86) from B. decoloratus. Amino acid sequence identity between Bd86 homologues (Bd86-1 and Bd86-2) and Bm86 was 86% and 85%, respectively, compared to 93% identity between the variants. Native Bd86 protein in B. decoloratus tick mid-gut sections and recombinant Bd86-1 reacted strongly with sera from TickGARD vaccinated cattle. TickGARD can therefore protect against a heterologous tick species with multiple antigen sequences. Epitope mapping using sera from TickGARD-vaccinated cattle identified two linear peptides conserved between the Bd86 homologues and Bm86. These epitopes represent candidate synthetic peptide vaccines for control of Boophilus spp. and the pathogens transmitted by these tick vectors.

Two novel baculovirus-derived recombinant Theileria parva p67 constructs were tested for their vaccine potential against East Coast fever. Boran calves were immunized with a his-GFP-p67 fusion protein (GFP:p67deltaSS) or with GP64:p67C, a protein fusion between a C-terminal domain of p67 and the baculovirus envelope protein GP64. Both GFP:p67deltaSS and GP64:p67C induced antibodies with high ELISA titers that neutralized T. parva sporozoites with high efficiency. Upon challenge, a correlation was observed between the in vitro neutralizing capacity and the reduction in severe ECF for individual animals. A protection level upto 85% was obtained. This level of protection was achieved with only two inoculations of 100 microg per dose, which is a major improvement over previous recombinant p67 products.

Initial laboratory trials of an experimental subunit vaccine againstTheileria parvabased on the 67-kDa major sporozoite surface antigen revealed a range of responses to challenge. We have analyzed convergence in seven sets of monozygotic twins which suggests that genetic factors may have an influence in determining the degree of protection provided by p67 immunization. In addition, we have examined whether allelic diversity at major histocompatibility complex class II loci influences protection. Analysis of bovine leukocyte antigenDRB3diversity in 201 animals identified significant associations with vaccine success (DRB3*2703;P= 0.027) and vaccine failure (DRB3*1501;P= 0.013). Furthermore,DRB3*2703was associated with the likelihood of immunized animals showing little to no clinical signs of disease following challenge. We discuss the acquired and innate immune mechanisms that may be behind the associations described here.

We developed an F(11) AIL population from an F(1) cross of A/J (susceptible) and C57BL/6J (resistant) mouse strains. One thousand F(11) mice were challenged with P.c. chabaudi 54X, and 340 mice selected from the phenotypic extremes for susceptibility and resistance were genotyped for microsatellite markers on Chromosomes (Chrs) 5, 8, and 17. QTL originally detected in backcross and F(2) populations were confirmed on the three chromosomes within narrower genomic regions, by maximum likelihood and regression analyses. Each of the previously mapped QTL on Chrs 5 and 17 resolved into two linked QTLs. The distal and proximal QTLs on Chrs 5 and 17, respectively, map to the previously reported QTL.

Monoclonal antibodies (MAb) binding to Cowdria ruminantium elementary bodies (EB) were identified by enzyme-linked immunosorbent assay, and surface binding of one MAb (446.15) to intact EB was determined by immunofluorescence, immunogold labeling, and transmission electron microscopy. MAb 446.15 bound an antigen of approximately 43 kDa in immunoblots of eight geographically distinct strains. The MAb did not react with Ehrlichia canis antigens or uninfected bovine endothelial cell lysate and may be useful in diagnostic assays and vaccine development.

As a first step in developing compartment-specific markers for protein trafficking within Theileria parva, we have isolated cDNAs encoding homologues of the small GTP binding proteins Rab1 and Rab4. The T. parva homologue of Rab1 (TpRab1), a protein which regulates vesicular transport between the endoplasmic reticulum and cis golgi in other organisms, was unusual in that it contained a unique 17 amino acid C-terminal extension. The C-terminal motif sequence KCT (XCX) contrasted with the CXC or XCC motifs which act as as signals for isoprenylation by geranylgeranyl in most Rab proteins, including all known Rab1 homologues, in containing only a single cysteine. [C14]mevalonic acid lactone and [H3]geranylgeranyl pyrophosphate were specifically incorporated into recombinant TpRab1 in vitro, demonstrating that the novel motif was functional for isoprenylation. Recombinant TpRab1 bound radiolabeled GTP, and this binding was inhibited by excess unlabeled GTP and GDP and also partially by ATP. The TpRab1 gene contained four short (34–67 bp) introns with a distinct pattern of occurrence within the protein sequence as compared to the introns of other lower eukaryote Rab1 genes. Immunofluorescence microscopy using antiserum specific for the novel C-terminal peptide in combination with labelling of cells using the nucleic acid-staining dye DAPI, indicated that TpRab1 was located in the vicinity of the schizont nucleus within the infected lymphocyte.

The use of tick vaccines in mammalian hosts has been shown to be the most promising alternative tick control method to current use of acaricides, which suffers from a number of limitations. However, the success of this method is dependent on the identification, cloning, and in vitro expression of tick molecules involved in the mediation of key physiological roles with respect to the biological success of a tick as a vector and pest. We have sequenced and characterized a Haemaphysalis longicornis tick salivary gland-associated cDNA coding for a 29-kDa extracellular matrix-like protein. This protein is expressed in both unfed and fed immature and mature H. longicornis ticks. The predicted amino acid sequence of p29 shows high homology to sequences of some known extracellular matrix like-proteins with the structural conservation similar to all known collagen proteins. Immunization with the recombinant p29 conferred a significant protective immunity in rabbits, resulting in reduced engorgement weight for adult ticks and up to 40 and 56% mortality in larvae and nymphs that fed on the immunized rabbits. We speculate that this protein is associated with formation of tick cement, a chemical compound that enables the tick to remain attached to the host, and suggest a role for p29 as a candidate tick vaccine molecule for the control of ticks. We have discussed our findings with respect to the search of tick molecules for vaccine candidates.

The presence of previously uncharacterized antigens (new antigens) on the surface of intact erythrocytes infected with three strains of Babesia bigemina from Kenya and one each from Puerto Rico, Mexico, St. Croix, and Texcoco-Mexico was demonstrated by indirect immunofluorescent antibody (IFA) reactions. These antigens were not strain specific because antibodies in bovine immune serum to either the Mexico or Kenya isolates reacted with all seven strains tested. Homologous and heterologous immune serum antibodies bound a maximum of 83% and 55%, respectively, of intact erythrocytes infected with the Kenya-Ngong strain but not uninfected erythrocytes. Both sera caused agglutination of only infected erythrocytes. Antibodies eluted from the surface of glutaraldehyde (0.25%) fixed infected erythrocytes had IFA reaction patterns among strains similar to those of immune sera before elution. Eluted antibodies were used to determine if these antigens were protein and encoded by B. bigemina. Eluted antibodies bound seven parasite-encoded proteins of 240, 220, 66, 62, 58, 52 and 38 kDa in an erythrocyte surface-specific immunoprecipitation reaction of 35S-methionine labelled proteins. It was concluded that the surface of B. bigemina infected erythrocytes had parasite-encoded proteins and that these proteins had surface exposed epitopes that were conserved among the seven strains examined which were from two continents.

BackgroundTheileria parva causes an acute fatal disease in cattle, but infections are asymptomatic in the African buffalo (Syncerus caffer). Cattle can be immunized against the parasite by infection and treatment, but immunity is partially strain specific. Available data indicate that CD8+ T lymphocyte responses mediate protection and, recently, several parasite antigens recognised by CD8+ T cells have been identified. This study set out to determine the nature and extent of polymorphism in two of these antigens, Tp1 and Tp2, which contain defined CD8+ T-cell epitopes, and to analyse the sequences for evidence of selection.Methodology/Principal FindingsPartial sequencing of the Tp1 gene and the full-length Tp2 gene from 82 T. parva isolates revealed extensive polymorphism in both antigens, including the epitope-containing regions. Single nucleotide polymorphisms were detected at 51 positions (~12%) in Tp1 and in 320 positions (~61%) in Tp2. Together with two short indels in Tp1, these resulted in 30 and 42 protein variants of Tp1 and Tp2, respectively. Although evidence of positive selection was found for multiple amino acid residues, there was no preferential involvement of T cell epitope residues. Overall, the extent of diversity was much greater in T. parva isolates originating from buffalo than in isolates known to be transmissible among cattle.Conclusions/SignificanceThe results indicate that T. parva parasites maintained in cattle represent a subset of the overall T. parva population, which has become adapted for tick transmission between cattle. The absence of obvious enrichment for positively selected amino acid residues within defined epitopes indicates either that diversity is not predominantly driven by selection exerted by host T cells, or that such selection is not detectable by the methods employed due to unidentified epitopes elsewhere in the antigens. Further functional studies are required to address this latter point.

DNAs of Theileria parva parva, T. p. lawrencei, T. p. bovis and Theileria mutans stocks, from Kenya, Uganda, Zanzibar and Zimbabwe were digested with either Sfi I or Not I and analysed using contour-clamped homogeneous electric field (CHEF) and field-inversion gel electrophoresis (FIGE). The Sfi I-digested T. parva genomic DNA resolved into approximately 30 fragments while the Not I digestion produced between 4–7 bands. The summation of the sizes of Sfi I fragments gave an estimate of 9–10×10 6 base pairs for the size of the T. parva genome. Heterogeneity within T. p. parva Muguga, Pemba/Mnarani and Mariakani stocks was detected. All the T. parva stocks analysed showed Sfi I and Not I restriction fragment length polymorphisms (RFLP). Hybridisation of 5 Sfi I-digested T. parva DNAs with a Plasmodium berghei telomeric repeat probe suggested that most of the polymorphisms and heterogeneity occurred in the telomeric or sub-telomeric regions of the genome. The recognition by the Plasmodium telomeric probe of 7–8 strongly hybridising Sfi I bands indicates that the T. parva genome may possess at least 4 chromosomes. The T. mutans genome was cut frequently with the above enzymes resulting in large numbers of fragments predominantly below 50 kb, thus suggesting either a much higher G+C content than T. parva or the presence of highly reiterated G+C-rich regions.

Initial laboratory trials of an experimental subunit vaccine against Theileria parva based on the 67-kDa major sporozoite surface antigen revealed a range of responses to challenge. We have analyzed convergence in seven sets of monozygotic twins which suggests that genetic factors may have an influence in determining the degree of protection provided by p67 immunization. In addition, we have examined whether allelic diversity at major histocompatibility complex class II loci influences protection. Analysis of bovine leukocyte antigen DRB3 diversity in 201 animals identified significant associations with vaccine success (DRB3*2703; P = 0.027) and vaccine failure (DRB3*1501; P = 0.013). Furthermore, DRB3*2703 was associated with the likelihood of immunized animals showing little to no clinical signs of disease following challenge. We discuss the acquired and innate immune mechanisms that may be behind the associations described here.

East Coast fever (ECF) in cattle is caused by the tick-borne protozoan parasite Theileria parva. The major sporozoite surface antigen of T parva (p67) is an important candidate for inclusion in a subunit vaccine. Recently, we reported the expression and production of different parts of p67 as fusions to either GFP or to the baculovirus GP64 envelope glycoprotein in insect cells, which resulted in stable proteins recognized by a monoclonal specific for native p67. The immunogenicity of these fusion proteins was examined in out-bred mice and cattle. In mice, the full length p67 molecule without its signal peptide and transmembrane region, but fused to GFP (GFP:p67 DeltaSS) was the best immunogen followed by the C-terminus of p67 fused to GP64 (GP64:p67C). These two immunogens also provoked a high level of sero-conversion in cattle when formulated in a water-in-oil or saponin-derived adjuvant with only 100 mug of protein and a single booster. The vaccine-elicited antibodies efficiently inhibited the infectivity of T parva sporozoites in in vitro neutralization assays. This study demonstrated that these new baculovirus-derived p67 vaccines were highly immunogenic, and that in combination with a suitable adjuvant, they have a clear potential to induce protective immunity in cattle. (C) 2004 Elsevier B.V. All rights reserved.

Theileria parva is an intracellular protozoan parasite that causes East Coast fever, a severe lymphoproliferative disease in cattle. Previous attempts to produce recombinant sporozoite surface antigen (p67) in bacterial or insect cells for vaccine purposes have not resulted in a correctly folded protein. Here, we report the expression of N- and C-terminal domains of p67 fused to the baculovirus envelope glycoprotein GP64 by cloning the appropriate p67 cDNA segments between the signal sequence and the major portion of GP64. To further advance the generation of such recombinants, existing surface display techniques were combined with bacmid technology. Chimeric proteins were present on the surface of budded viruses as judged by immunogold labelling and were exposed on the surface of insect cells, as concluded from immunofluorescence studies of infected, non-fixed insect cells. In non-denaturing dot blot experiments, a strong reaction was obtained between monoclonal TpM12 and baculovirus particles displaying the p67N-GP64 chimeric protein. This antibody, raised against native p67, also specifically recognized the surface of recombinant-infected cells. Apparently, a more native conformation was achieved than when p67 was expressed in E.coli or in conventional baculovirus expression systems. The baculovirus surface expression system, therefore, provides an improved way of expressing this T.parva sporozoite surface protein.

Polymorphic, multicopy gene families are frequently located in subtelomeric regions of the genomes of parasitic protozoa. Theileria parva telomere-associated (TA) DNA from two chromosomes contained long open reading frames (ORFs) 54% identical at the N-termini, whose 3′ ends were 2670 and 2680 bp from the telomeric repeats. Probes derived from these ORFs revealed related sequences close to additional telomeres. The 3′ end of an unrelated ORF was approximately 2720 bp from a third telomere. These are among the closest ORFs to telomeres in any organism. Reverse transcription PCR detected transcripts originating within the telomeric multicopy gene family. Additional ORFs, with complex sequence similarities, were located centromeric to the telomere-adjacent ORFs. Transcripts from the schizont stage of T. parva , containing domains with significant amino acid similarity to a 3529 codon ORF located 6900 bp upstream of the telomeric repeats, were mapped to a subtelomeric locus at a fourth telomere. Five telomeric ORFs contained predicted N-terminal signal peptides and one of these signal peptides was functional in a heterologous system. Hybridisation data suggested extensive strain polymorphism between ORFs. Two of the telomere-adjacent ORFs were absent from the genome of a cloned T. parva parasite which can, nonetheless, be passaged through ticks and cattle. T. parva is unusual, among organisms so far studied, in the high density of potential coding sequences located directly adjacent to telomeres and the apparent absence of extensive tracts of repeated sequences within the TA DNA.

Summary The major sporozoite surface antigen of Theileria annulata (SPAG-1) is a candidate for inclusion in a subunit vaccine. In this paper we summarize the results of 4 vaccination experiments using recombinant SPAG-1 expressed in different systems and presented in different adjuvants. The antigen has been presented as either a C terminal 108 amino acid peptide (called SR1) expressed as both β-galactosidase and hepatitis B core antigen fusions or as a full-length form expressed as a GST fusion with an N terminal His6 tag. We used different adjuvants, namely Freund's, saponin, ISCOMs and a proprietary adjuvant supplied by SmithKline Beecham, which we call SKBA. The data point to the conclusion that SPAG-1 can elicit partial protection and is therefore suitable for inclusion in an eventual multicomponent subunit vaccine.

SPAG-1, a sporozoite surface antigen of T. annulata, has previously been shown to elicit partial protection when used, as an hepatitis B core antigen fusion, to immunize cattle. The objective of this study was to try and improve the protective capacity of this antigen by enlisting different vaccine strategies. Cattle were immunized with SPAG-1, as a fusion protein with a His6 tag, either incorporated into ISCOMs, with or without the merozoite antigens TAMS 1-1 and 1-2, or with RWL as adjuvant three times at monthly intervals. Another group of cattle were immunized with p67, the T. parva sporozoite antigen, in RWL to assess whether any cross-protection could be induced. The animals were then challenged with an estimated LD50 of T. annulata sporozoites, and their ability to resist the infection was investigated. Serum responses and T-cell proliferative responses were analyzed throughout the trial. Post-challenge analyses included lymph node biopsies and blood smears to check for the presence of parasites, routine hematological parameters, and observation for clinical manifestations of the disease. The results of this trial will be discussed.

A Theileria parva specific full-length cDNA clone, T7, which encodes a protein with more than 60% homology to heat shock protein 90 (hsp90) of other organisms, has been identified. T7 appears to be a single copy gene. The gene is expressed as a protein of 87 kDa in both the sporozoite and schizont stages of T. parva. The protein was not found in the piroplasm stage, although the corresponding transcript was detected, suggesting post-transcriptional regulation of the gene. In the schizont stage the T7 protein is upregulated upon heat shock and localized in the cytoplasm.

Investigations were undertaken to evaluate the immune response of trypanotolerant N'Dama (Bos taurus) and susceptible Boran (Bos indicus) cattle to two Trypanosoma congolense variable antigen types (VATs) expressed in both breeds following tsetse-transmitted challenge. The VAT-specific antibodies of both IgM and IgG1 isotypes produced by both breeds had similar neutralizing titres. The interaction between immune sera, trypanosomes and freshly isolated peripheral blood leucocytes (PBL) from uninfected N'Dama and Boran animals was studied. It was found that both N'Dama and Boran immune sera were able to induce adherence of trypanosomes to the N'Dama PBL, but not to Boran PBL. The adherence-inducing activity was exhibited by both IgM and IgG1 antibodies, but IgG1 antibodies were more efficient in this respect. These results suggest that there are qualitative and/or quantitative differences in the immunoglobulin receptor function of PBL between the two breeds of cattle.

A neutralizing antiserum, C16, raised against sporozoites of Theileria parva parva was used to screen a lambda gt11 expression library of T. parva parva (Muguga) genomic DNA fragments. Proteins encoded by one phage clone, lambda TpS-17, were reactive with the C16 antiserum. Detailed characterisation of the DNA insert showed it to encode determinants found on four theilerial antigens of approximately 104, 90, 85 and 35 kDa. The sequence encoded by the clone is expressed during sporogony as a single RNA transcript of about 3000 nucleotides. On sequencing a portion of the 5000-bp insert, an open reading frame of 2772 bp was revealed that encoded a 104-kDa protein. Immunoscreening a library of subfragments of the DNA insert with the original antiserum localised sequences encoding the dominant antigenic determinants to an 800-bp stretch of DNA at the 3' end of the open reading frame. Sequence data from three subclones spanning this region show portions of the antigenic domains to be unusually rich in proline residues which are repeated every three amino acids. These repeats often take the form X-S(T)-P or X-K(R)-P. Antibodies directed against each of the three subclones recognize the 104- and 35-kDa antigens and different combinations of the 90- and 85-kDa kDa antigens, suggesting that the smaller proteins are derived from the 104-kDa antigen by limited proteolysis occurring at the carboxyl terminus end of the protein. In immunoelectron micrographs the antigen is associated with the microneme/rhoptry complexes of the sporozoite.

Palmer G. H., Barbet A. F., Musoke A. J., Katende J. M., Rurangirwa F., Shkap V., Pipano E., Davis W. C. and Mcguire T. C. 1988. Recognition of conserved surface protein epitopes on Anaplasma centrale and Anaplasma marginale isolates from Israel, Kenya and the United States. International Journal for Parasitology18: 33–38. Field isolates of A. marginale from Israel, Kenya and the United States and Anaplasma centrale were shown to be antigenically distinct using a panel of 1:1 isolaterestricted monoclonal antibodies. One monoclonal antibody reacted with all A. marginale isolates but not A. centrale and may be species-specific. A second panel of seven antibodies detected epitopes common to all isolates and both species. These common epitopes were present on two major Anaplasma surface proteins of Mr 36,000 and Mr 105,000. Monoclonal antibodies capable of neutralizing heterologous A. marginale isolate infectivity recognized 100% of the organisms in each isolate including A. centrale. In addition, immune sera from cattle recovered from infection with each of the isolates contained high titer antibodies to the isolated surface proteins bearing the common epitopes. These results demonstrate that antigenically distinct Anaplasma isolates and species contain common surface epitopes that may be important in developing effective immunoprophylaxis.

SUMMARY Cattle were infected with two different clones of Trypanosoma hrurei (MITat 1.2 and ILTat 1.3) and antibody response was followed by radioimmunoassay. In four of the seven animals there were at least two peaks of antibody activity to the infecting clones, with the second peak much higher than the first. Specific antibodies (IgG1 and IgM but not IgG2) were eluted from the immunoabsorbent columns on which the variant surface glycoproteins (VSGs) were coupled. By neutralization of infectivity tests, IgM antibodies from the first peak of antibody activity were more efficient in neutralizing trypanosomes than IgGl but the reverse was true of the antibodies isolated from the second peak. By absorption with multiple variable antigen types isolated during the course of infection, all the IgM and IgG1 in the first 3 weeks of infection were shown to be specific. It is suggested that polyclonal B cell stimulation leading to dysfunction in the control of IgM and IgG production may not be responsible for the high levels of these immunoglobulins in bovine trypanosomiasis.

Groups of cattle were inoculated intravenously with cloned populations of bloodstream forms of Trypanosoma brucei or Trypanosoma congolense. All five steers infected with T. brucei ILTat 2.1 and six of the eight steers infected with T. congolense IL 13-E14 became aparasitemic within 16 and 32 weeks postinfection, respectively. Examination of sera from animals infected with T. brucei by indirect immunofluorescence and neutralization assays revealed the presence of antibodies against all the metacyclic variable antigen types (VATs) of the infecting clone. The neutralizing capacity of the sera increased with the course of infection from 1:10 at 2 months to 1:100 at 3 to 4 months postinfection. The recovered animals were completely immune to challenge by Glossina morsitans subsp. centralis infected with clone IL Tat 2.1, which had initiated the infection, as well as with another clone (IL Tat 2.2) belonging to the same serodeme, but they were susceptible to a tsetse-transmitted heterologous challenge with isolate STIB 367-H. Similar results were obtained with sera from T. congolense IL 13-E14-infected steers. The six steers infected with a different T. congolense ILNat 3.1 clone did not recover spontaneously; however, 2 months postinfection, sera from five of them also contained neutralizing antibodies against ILNat 3.1 metacyclic VATs. These results indicate that some of the bloodstream VATs that arise during the course of a chronic infection possess surface epitopes in their variable surface glycoproteins that are identical to those of the metacyclic VATs. It is suggested that in chronic infection, the infecting trypanosomes could exhaust their VAT repertoire, including those that cross-react with metacyclics, thereby leading to both "self-cure" and subsequent immunity to homologous cyclically transmitted challenge.

Three groups of cattle, each group comprising six animals, were inoculated intravenously with populations of bloodstream forms of Trypanosoma vivax. The first group received T. vivax clone ILDat 1.3 derived from an isolate from Nigeria, while the other two received T. vivax stocks IL 1875 or IL 2133 isolated from Coast Province, Kenya. One animal from the group that was infected with IL 1875 died 8 weeks postinfection. The remaining 17 animals became aparasitemic in 8 to 12 weeks without intervention by drug therapy. The recovered animals developed serodeme-specific immunity against Glossina morsitans subsp. centralis-transmitted challenge. There was complete cross-protection between the two East African T. vivax stocks, although they were isolated from areas 80 to 90 km apart, indicating that they belong to the same serodeme. Antibodies to the homologous metacyclic variable antigen types (VATs) were not detected in sera from recovered animals, suggesting that the immunity displayed by the recovered animals was directed at the bloodstream and not the metacyclic VATs. It is thus suggested that recovery in these animals is due to exhaustion of the repertoire of bloodstream VATs expressed in the animals by the infecting T. vivax clone or stocks.

ANTICOMPLEMENT factors have been shown to occur in certain parasitic infections of man and animals. Hammerberg et al.1 demonstrated the presence of a factor from Taenia taeniaeformis capable of activating complement through the alternate pathway. Similar findings have been reported for a substance in the cercarial coat of Schistosoma mansoni2. In trypanosomiasis, no such parasitic activity has been reported although hypocomplementaemia has been recognised as a consistent feature of the disease. Marked reductions in complement C4 and C3 were observed in infections with Trypanosoma lewisi3, Tr. rhodesiense4 and Tr. congolense5. One possible mechanism is that the decrease of these complement components results from activation by antigen–antibody complexes. Deposits of immunoglobulins and complement have been demonstrated in kidneys of mice infected with Trypanosoma brucei6,7 and monkeys infected with Tr. rhodesiense4. We present here evidence for an alternative mechanism operating through direct activation of human complement, via the classical pathway, by an isolated variant-specific surface antigen of Tr. brucei.

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.

The p67 sporozoite antigen of Theileria parva has been fused to the C-terminal secretion signal of Escherichia coli hemolysin and expressed in secreted form by attenuated Salmonella dublin aroA strain SL5631. The recombinant p67 antigen was detected in the supernatant of transformed bacterial cultures. Immunization trials in cattle revealed that SL5631 secreting the antigen provoked a 10-fold-higher antibody response to p67 than recombinant SL5631 expressing but not secreting p67. Immunized calves were challenged with a 80% lethal dose of T. parva sporozoites and monitored for the development of infection. Two of three calves immunized intramuscularly with the p67-secreting SL5631 strain were found to be protected, whereas only one of three animals immunized with the nonsecreting p67-expressing SL5631 strain was protected. This is the first demonstration that complete eukaryotic antigens fused to the C-terminal portion of E. coli hemolysin can be exported from attenuated Salmonella strains and that such exported antigens can protect cattle against subsequent parasite challenge.

Seven out of 11 bovines infected with different clones of Trypanosoma brucei showed 2 peaks of antibody activity against the infecting clone within 7 weeks, as measured by immunofluorescence, radioimmunoassay, and neutralization of infectivity tests. Using other clones from an unrelated Stock, antibodies to these clones were not detectable, indicating that the antibodies produced were specific to the infecting organisms. These results suggest that there was a reappearance or increase in numbers of the infecting organisms or of organisms with variable surface antigens similar to those of the infecting clones. The reappearance of variable antigen types in the presence of specific antibodies would imply that antibody plays a selective rather than an inductive role in the process of antigenic variation in African trypanosomes.