[EN] The development of successful anti-tick vaccines depends largely on the identification of tick-derived antigens capable of eliciting protective immune responses in the host. This requires a thorough understanding of the molecular interactions occurring at the tick-host interface to facilitate the selection of promising antigen candidates. In other tick species, various molecular and cellular components-including proteins, non-coding RNAs, and the microbiome-have been shown to play critical roles at the tick-host-pathogen interface influencing tick development, reproduction and pathogen transmission.
In O. moubata, knowledge of these molecular and microbial components and their involvement in host-parasite interactions remains limited. The application of omics technologies to study these components may provide valuable insights to inform the rational design of efficient anti-O. moubata vaccines.
This Doctoral Thesis investigates the O. moubata-host interface by focusing on three key components-salivary miRNAs, salivary proteins, and the microbiome-using omics-based approaches to enhance our understanding of the molecular mechanisms underlying these interactions.
Given the lack of prior information on the identity of O. moubata miRNAs and their potential roles in tick physiology and tick-host interactions, in the first chapter of this thesis, we have characterised the miRNAs present in the saliva of O. moubata and Ornithodoros erraticus, the Mediterranean vector of ASF and TBRF, and have predicted their putative target genes in the swine host (Sus scrofa), including an in silico functional analysis.
To this end, small RNAs were sequenced using next-generation sequencing (NGS), and a total of 379 miRNAs were identified by mapping to the RNAcentral database. Functional analysis of the predicted host target genes suggested that O. moubata salivary miRNAs may be involved in key processes at the tick-host interface, including signal transduction, transcriptional regulation, synapse modulation, immune response, angiogenesis, and vascular development (Cano-Argüelles et al., 2023. Ticks and Tick-borne Diseases 14, no 6, p. 102249).
In the second chapter, we investigated the regulatory roles of miRNAs in the salivary glands of O. moubata and O. erraticus. For this purpose, previously generated sialotranscriptomes of O. moubata and O. erraticus were used to predict putative miRNA target genes in silico. Subsequently, the in silico predictions for O. moubata were experimentally validated for several well-characterised miRNAs using antagomirs to induce miRNA knockdown in adult females. This approach enabled us to identify differentially expressed salivary genes in response to miRNA knockdown, thereby confirming them as miRNA targets, and to detect significant impairments in tick feeding, reproduction, and survival associated with miRNA silencing (Cano-Argüelles et al., 2025. Pathogens 14, no 6, p. 595).
The results exposed in these two chapters underscore the importance of miRNAs in tick physiology and tick-host interactions, offering new insights into this complex interface. These findings may contribute to the more precise selection of tick-derived molecules for the development of therapeutic or immunological strategies aimed at controlling tick infestations and the transmission of tick-borne pathogens.
In the third chapter, a vaccinomics approach was employed to identify potential vaccine targets among O. moubata salivary proteins recently characterised in the O. moubata sialome. Seven secreted salivary proteins, predicted to be antigenic and involved in blood-feeding and modulation of host immune response, were selected for further evaluation. The proteins were expressed as recombinants, formulated with Montanide adjuvant, and administered to rabbits. Vaccine efficacy ranged from approximately 8% to 32%, with four candidates demonstrating potential for inclusion in multi-antigen vaccine formulations. These results validate the vaccinomics pipeline applied and support the selection of 57% of the tested candidates as promising antigens for the development of anti-O. moubata vaccines (Cano-Argüelles et al., 2025. Ticks and Tick-borne Diseases 16, no 3, p. 102483).
Finally, considering the critical role of the tick microbiome in physiology, reproduction, vector competence, nutrition, immunity, and host-pathogen interactions-as demonstrated in other tick species-the fourth chapter implemented a combined in silico and in vivo approach to investigate whether microbiota-targeted vaccination could alter keystone taxa and affect the microbial community structure and fitness of O. moubata. Vaccination targeting Pseudomonas spp. (identified as a keystone taxon in O. moubata) and Lactobacillus spp. (a non-keystone taxon) revealed that the former significantly reduced female survival, while the latter impaired oviposition and fertility. Co-occurrence network analysis of the O. moubata midgut microbiome following vaccination indicated shifts in microbial diversity and community composition, underscoring the influence of the vaccine target on microbiome structure. Further analysis of community assembly dynamics and network robustness emphasized the pivotal role of keystone taxa in maintaining microbiome structure and stability. These results offer valuable insights into the microbial network dynamics of O. moubata and support the potential of microbiota-targeted vaccines as a novel strategy for tick control (Cano-Argüelles et al., 2024. Molecular Ecology 33, no 18, p. e17506).
The results of this thesis advance our understanding of the complex molecular mechanisms underlying tick-host interactions and identify novel antigen candidates for the development of anti-Ornithodoros vaccines. This work supports the reliability of two complementary omics-based approaches for designing innovative vaccines targeting tick populations and provides a foundation for future strategies aimed at controlling ticks and the diseases they transmit.
