Evaluation of tick protein extracts as anti-tick vaccines targeting Ixodes ricinus in cattle

Abstract

Ticks are obligate haematophagous ectoparasites, which can transmit different pathogens such as bacteria, protozoa or viruses. Tick-borne diseases occur all over the world and threaten the health of humans and animals. A novel and promising tick control strategy is the development of anti-tick vaccines. Therefore, seven European institutes joined forces to develop an anti-tick vaccine, which inhibit transmission of common pathogens in Europe, such as Borrelia, Babesia or the tick-borne encephalitis virus. Within the framework of the ANTIDotE (Anti-tick vaccine to prevent tick-borne diseases in Europe) project, this PhD thesis is focused on immunization experiments with calves and subsequent identification of immunodominant proteins. Chapter 1 gives a general introduction to Ixodes ricinus ticks, highlights the importance of preventing tick-borne diseases and highlights different strategies of tick control. In Chapter 2, the occurrence, abundance as well as clinical symptoms and treatment schemes of common TBDs in Europe transmitted by Ixodes ricinus are summarized and discussed. Two small-scale immunization trials are described in Chapter 3. In these studies, crude protein extracts and recombinant Ferritin 2 were tested as anti-tick vaccine candidates. Ixodes ricinus ticks fed on tick protein extract-vaccinated calves were reduced in number and obtained lower repletion weights. Moreover, pronounced inflammatory signs were observed on the tick bite sites. Tick feeding was not impaired in the Ferritin 2 vaccinated calf. In vitro tick feeding assays using plasma or whole blood from vaccinated animals could not reproduce the in vivo feeding results, indicating that antibodies alone were not responsible for the observed vaccine immunity. In Chapter 4, immune sera from salivary glands or midgut extract vaccinated calves were used for co-immunoprecipitation of tick tissue extracts followed by LC/MS-MS analyses. This resulted in the identification of 46 immunodominant proteins. The relative gene expression in tick tissues of unfed and partially fed females were measured for 15 proteins. Ten out of the 15 candidates were selected for further RNAi studies. The strongest RNAi phenotypes were observed for MG6 (A0A147BXB7), a protein containing eight Fibronectin type III domains predominantly expressed in tick midgut and ovaries of feeding females, and SG2 (A0A0K8RKT7), a glutathione-S-transferase that was found to be upregulated in all investigated tissues upon feeding. The results showed that co-immunoprecipitation of tick proteins with host immune sera, followed by protein identification using LC-MS/MS is a valid approach to identify antigen-antibody interactions and could be integrated in anti-tick vaccine discovery pipelines. The studies of this thesis present possibilities for the evaluation and identification of potential anti-tick vaccine candidates. Furthermore, immunodominant proteins of the species I. ricinus were identified and characterized, which could contribute to the development of an anti-tick vaccine against the vector I. ricinus.