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The Veterinary record 03/2008; 162(7):216-7. · 1.25 Impact Factor
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ABSTRACT: Imidocarb treatment of horses infected with Babesia caballi is supposed to eliminate the infection, but data on the efficacy of this treatment is scarce. The study presented here concerns four Paso Fino horses, which were imported into the island of Curacao on the basis of a piroplasmosis negative complement fixation test (CFT). Upon re-testing with an indirect fluorescent antibody test immediately after arrival in Curacao, two horses appeared to have antibodies to B. caballi and all horses had antibodies to Theileria equi. Subsequent testing with polymerase chain reaction combined with a reverse line blot yielded positive results for both agents in all four horses. Treatment with five consecutive doses of imidocarb dipropionate (4.7 mg/kg BW im q 72 h), temporarily resulted in negative results, but B. caballi and T. equi were detected again in the samples taken at 6 and 18 weeks after completion of the treatment. These results confirm that the CFT is not a suitable test for pre-import testing and that even high dose treatment with imidocarb may not be capable of eliminating B. caballi and T. equi infections from healthy carriers.
Veterinary Parasitology 03/2008; 151(2-4):320-2. · 2.58 Impact Factor
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ABSTRACT: Pathogen DNA was isolated from roan antelope (Hippotragus equinus), sable antelope (Hippotragus niger), greater kudu (Tragelaphus strepsiceros), and common gray duiker (Sylvicapra grimmia) in South Africa whose deaths were attributed to either theileriosis or cytauxzoonosis. We developed Theileria species-specific probes used in combination with reverse line blot hybridization assays and identified three different species of Theileria in four African antelope species. The close phylogenetic relationship between members of the genera Theileria and Cytauxzoon, similarities in the morphologies of developmental stages, and confusion in the literature regarding theileriosis or cytauxzoonosis are discussed.
Journal of Clinical Microbiology 01/2006; 43(12):5907-11. · 4.15 Impact Factor
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ABSTRACT: The prevalence of Babesia infections in domestic dogs in South Africa was studied using reverse line blot hybridization and 18S sequence analysis. Babesia canis vogeli was confirmed for the first time in domestic dogs in South Africa. Out of a total of 297 blood samples collected from domestic dogs in Bloemfontein, East London, Johannesburg, Durban and from the Onderstepoort Veterinary Academic Hospital, 31 were positive for Babesia canis rossi, whereas B. c. vogeli was detected in 13 dogs. None of the dogs carried both parasites. The detection of B. c. vogeli has implications with regard to prevalence and varied clinical manifestation of canine babesiosis in South Africa.
Veterinary Parasitology 07/2004; 122(2):119-25. · 2.58 Impact Factor
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ABSTRACT: We report the first case of human babesiosis in Portugal. A 66-year-old splenectomized man was admitted to a Lisbon hospital after 1 week of fever, abdominal pain, anorexia and nausea. A high parasitaemia (30%) of Babesia parasites was found in Giemsa-stained blood smears and, despite treatment, the patient died several weeks later of renal failure. Ethylenediaminetetraacetic acid blood samples were processed for polymerase chain reaction (PCR) and reverse line blot hybridization to confirm and characterize the Babesia infection. The amplified PCR product was cloned and subsequently sequenced. Molecular analysis showed that the infection was caused by Babesia divergens and that other blood parasites were not involved. Phylogenetic analysis showed that the 18 S ribosomal RNA gene sequence was similar to three other European isolates of B. divergens. In view of the high risk for splenectomized individuals, strict measures should be taken to avoid tick bites.
Tropical Medicine & International Health 09/2003; 8(8):760-4. · 2.80 Impact Factor
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ABSTRACT: Background: Rhipicephalus (Boophilus) spp. ticks economically impact on cattle production in Africa and other tropical and subtropical regions of the world. Tick vaccines constitute a costeffective and environmentally friendly alternative to tick control. The R. microplus Bm86 protective antigen has been produced by recombinant DNA technology and shown to protect cattle against tick infestations. Results: In this study, the genes for Bm86 (R. microplus), Ba86 (R. annulatus) and Bd86 (R. decoloratus) were cloned and characterized from African or Asian tick strains and the recombinant proteins were secreted and purified from P. pastoris. The secretion of recombinant Bm86 ortholog proteins in P. pastoris allowed for a simple purification process rendering a final product with high recovery (35–42%) and purity (80–85%) and likely to result in a more reproducible conformation closely resembling the native protein. Rabbit immunization experiments with recombinant proteins showed immune cross-reactivity between Bm86 ortholog proteins. Conclusion: These experiments support the development and testing of vaccines containing recombinant Bm86, Ba86 and Bd86 secreted in P. pastoris for the control of tick infestations in Africa.
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ABSTRACT: Following an outbreak of autochthonous canine babesiosis in the Netherlands, a request made to veterinarians and the public to collect ticks from companion animals resulted in 4298 ticks submitted between July 2005 and October 2006 to our center. Ticks were identified as Ixodes ricinus adults (2907/4298, 67.6%), Ixodes sp. nymphs (529/4298, 12.3%) and Ixodes sp. larvae (385/4298, 9.0%), I. hexagonus adults (328/4298, 7.6%), Dermacentor reticulatus (72/4298, 1.7%), and several other exotic tick species such as Amblyomma flavomaculatum (formerly Aponomma flavomaculatum), Hyalomma marginatum rufipes, Rhipicephalus sanguineus, and R. turanicus (55/4298, 1.3%). Eight localities were surveyed for the presence of local D. reticulatus, a tick not indigenous to the Netherlands, based on multiple submissions of D. reticulatus ticks from these areas. D. reticulatus was collected from the vegetation in six of these localities, confirming the presence of populations of this tick in the Netherlands. Adult I. ricinus (n=251), I. hexagonus (n=237), and D. reticulatus (n=344) ticks were selected at random and subsequently screened by polymerase chain reaction (PCR) and reverse line blot (RLB) hybridization for the presence of Borrelia, Babesia, Theileria, Anaplasma, Ehrlichia, and Rickettsia species. I. ricinus ticks were infected with Rickettsia helvetica (24.7%), spirochetes belonging to the Borrelia burgdorferi sensu lato group (7.2%), the Ehrlichia-like "Schotii" variant (2.4%), Anaplasma phagocytophilum (1.6%), Babesia sp. (EU1) (1.2%), Babesia divergens (0.4%), and Babesia microti (0.4%). A. phagocytophilum (5.9%) and R. helvetica (0.8%) were also detected in adult I. hexagonus ticks. Spotted fever group Rickettsiae, previously reported as Rickettsia sp. DnS14/RpA4 (14.0%), and Borrelia burgdorferi sensu lato (0.3%) were detected in the D. reticulatus ticks, which appeared to be free from B. canis infection. We concluded that a much broader spectrum of ticks and tick-borne pathogens is present in the Netherlands than previously thought, including several potential zoonotic pathogens.
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A.M. Nijhof
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ABSTRACT: Ticks and tick-borne diseases seriously affect animal and human health worldwide with the highest economic losses occurring in livestock production in the developing world. The control of ticks and the diseases they transmit depends mainly on chemical tick control using acaricides. The development of acaricide resistance, concerns about environmental pollution and pesticide residues in food products result in the need for alternative tick control methods such as anti-tick vaccines. Commercial vaccines based on the recombinant antigen Bm86 have been developed for the control of Rhipicephalus (Boophilus) microplus infestations on cattle. Although they can make an important contribution to an integrated tick control strategy, more efficacious and ideally stand-alone vaccines are required to control multiple tick species in wide geographical areas. There is thus a need for improved vaccine formulations and for the discovery of new tick-protective antigens. This identification of tick-protective antigens is a pivotal step in vaccine development. To study gene function and to identify genes which are important to the tick’s function and survival, RNA interference (RNAi) can be a valuable tool. In this thesis, a strategy for gene silencing by RNAi in the one-host tick R. microplus is outlined and a novel RNAi application for the transovarial silencing of genes in ticks is introduced. Subolesin, a protein which was identified as a tick-protective antigen is further characterized and evidence of the role of subolesin in gene expression in ticks is provided. Cross protection of Bm86 vaccines against tick species other than R. microplus has been reported, but Bm86 vaccines do not protect against Rhipicephalus appendiculatus infestations. One possible explanation is the variation in Bm86 expression levels between R. microplus and R. appendiculatus. To measure the Bm86 gene expression in a reliable and accurate manner, normalization of gene expression data against reference genes is essential and the expression stability of commonly used reference genes was evaluated in both species. The most stable reference genes were subsequently used for normalization of the Bm86 expression profile in all life stages of these ticks to examine the role of antigen abundance in Bm86 vaccine susceptibility. Partial protection of the Bm86 vaccine against other Rhipicephalus (Boophilus) and Hyalomma tick species suggests that the efficacy of a Bm86-based vaccine may be enhanced when based on the orthologous recombinant Bm86 antigen. We therefore identified the Bm86 homologues from species representing the main argasid and ixodid tick genera. A novel protein from metastriate ticks with multiple Epidermal Growth Factor-like domains which is structurally related to Bm86 was discovered and named ATAQ. The vaccine potential of ATAQ proteins against tick infestations is to be evaluated. The expression and purification of Bm86 (R. microplus), Ba86 (R. annulatus) and Bd86 (R. decoloratus) as secreted products from Pichia pastoris is also described. The secretion of recombinant Bm86 ortholog proteins in P. pastoris allowed for a simple purification process rendering a final product with high recovery and purity which is likely to result in a more reproducible conformation closely resembling the native protein and increase the efficacy of these vaccines.
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Z. Zivkovic,
A Torina,
R. Mitra,
A Alongi,
S Scimeca,
K M Kocan,
R C Galindo,
C. Almazan,
E F Blouin,
M. Villar, A.M. Nijhof,
R. Mani,
G. La Barbera,
S Caracappa,
F Jongejan,
J de la Fuente
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ABSTRACT: BACKGROUND: Ticks (Acari: Ixodidae) are vectors of pathogens worldwide that cause diseases in humans and animals. Ticks and pathogens have co-evolved molecular mechanisms that contribute to their mutual development and survival. Subolesin was discovered as a tick protective antigen and was subsequently shown to be similar in structure and function to akirins, an evolutionarily conserved group of proteins in insects and vertebrates that controls NF-kB-dependent and independent expression of innate immune response genes. The objective of this study was to investigate subolesin expression in several tick species infected with a variety of pathogens and to determine the effect of subolesin gene knockdown on pathogen infection. In the first experiment, subolesin expression was characterized in ticks experimentally infected with the cattle pathogen, Anaplasma marginale. Subolesin expression was then characterized in questing or feeding adult ticks confirmed to be infected with Anaplasma, Ehrlichia, Rickettsia, Babesia or Theileria spp. Finally, the effect of subolesin knockdown by RNA interference (RNAi) on tick infection was analyzed in Dermacentor variabilis males exposed to various pathogens by capillary feeding (CF). RESULTS: Subolesin expression increased with pathogen infection in the salivary glands but not in the guts of tick vector species infected with A. marginale. When analyzed in whole ticks, subolesin expression varied between tick species and in response to different pathogens. As reported previously, subolesin knockdown in D. variabilis infected with A. marginale and other tick-borne pathogens resulted in lower infection levels, while infection with Francisella tularensis increased in ticks after RNAi. When non-tick-borne pathogens were fed to ticks by CF, subolesin RNAi did not affect or resulted in lower infection levels in ticks. However, subolesin expression was upregulated in D. variabilis exposed to Escherichia coli, suggesting that although this pathogen may induce subolesin expression in ticks, silencing of this molecule reduced bacterial multiplication by a presently unknown mechanism. CONCLUSIONS: Subolesin expression in infected ticks suggested that subolesin may be functionally important for tick innate immunity to pathogens, as has been reported for the akirins. However, subolesin expression and consequently subolesin-mediated innate immunity varied with the pathogen and tick tissue. Subolesin may plays a role in tick innate immunity in the salivary glands by limiting pathogen infection levels, but activates innate immunity only for some pathogen in the guts and other tissues. In addition, these results provided additional support for the role of subolesin in other molecular pathways including those required for tissue development and function and for pathogen infection and multiplication in ticks. Consequently, RNAi experiments demonstrated that subolesin knockdown in ticks may affect pathogen infection directly by reducing tick innate immunity that results in higher infection levels and indirectly by affecting tissue structure and function and the expression of genes that interfere with pathogen infection and multiplication. The impact of the direct or indirect effects of subolesin knockdown on pathogen infection may depend on several factors including specific tick-pathogen molecular interactions, pathogen life cycle in the tick and unknown mechanisms affected by subolesin function in the control of global gene expression in ticks.
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A.M. Nijhof,
M.R. de la Fuente,
P Willadsen,
A. Neitz,
A. Taoufik,
M Canales,
S. Hope,
L. Jiang,
E B M Koney,
A. Agyei,
A. Gaspar,
C. Maritz-Olivier,
C. Stutzer,
M. Darghouth,
Y. Galai,
L.J. Bell-Sakyi,
V Naranjo,
J.M.J. Perez,
F Jongejan
