Project Number: 3094-32000-031-00-D
Project Type: In-House Appropriated
Start Date: Jun 1, 2009
End Date: May 31, 2014
Objective 1: Develop and test anti-tick vaccines for immunization of deer. Sub-objective 1A. Refine understanding of white-tailed deer immune system. Sub-objective 1B. Define quantitative and qualitative gene and protein responses in R. microplus during feeding on B. bovis-infected deer. Sub-objective 1C. Evaluation of candidate vaccine antigens. Objective 2: Identify sensory, physiological, and biological targets for development of novel acaricides and drugs for use as chemical control technology. Sub-objective 2A. Identify neuroregulatory processes in tick pharyngeal muscles and salivary glands. Sub-objective 2B. Identify inhibitors of pharyngeal pump function and tick feeding. Sub-objective 2C. Identify GPCRs and agonists/antagonists as candidates for novel acaricide development.
To design effective vaccines and vaccination protocols for this project, we must first better understand the nature of the immune system of the white-tailed deer, specifically the suitability of deer as hosts for R. microplus, the deer immune response upon exposure to tick antigens, and the ability of deer to serve as reservoirs for the transmission of Babesia to cattle. This will help define the role of deer in tick distribution and population maintenance. Infestation of the deer with R. microplus and B. bovis induces responses in the deer but also in the tick at the gene, protein and immunochemical level. We will determine these Babesia-induced responses in R. microplus using functional genomics and proteomics. Differentially expressed genes/proteins will be prioritized as candidates for vaccine development. To help identify these genes and proteins, we will use an established in vitro feeding system adapted for use with R. microplus females. Quantitative gene expression associated with ingestion of Babesia-infected blood will be analyzed using available R. microplus microarrays probed with RNA isolated from dissected tick tissues. Tick proteins will also be purified from the dissected tissues and SDS polyacrylamide gel electrophoresis used for comparisons between infected and uninfected samples. Candidate vaccine antigens will be evaluated for their effectiveness in controlling R. microplus infestations on deer and cattle and their capacity to block transmission of B. bovis between individual animals. Neurotransmitters and neuromodulators, including dopamine, GABA, and acetylcholine, play key roles in many tick physiological processes. We will identify these neuroregulators in tick synganglia and neurons innervating pharyngeal muscles and salivary glands. Literature-derived protocols will also be applied to study the neuromuscular organization of the pharyngeal pump. We will test effects and determine the mechanisms of action of various pharmacological agents, peptides, and vaccine candidates on pharyngeal pump function and tick feeding. Functional studies, including gene silencing studies, will confirm target identity and target validation. This information would facilitate development of novel acaricides that target genes that are critical to feeding success. It is necessary to identify muscular components involved in blood sucking and salivation to understand the physiology of feeding and test the pharmacology of molecules that regulate the tick pharyngeal pump. We will use the electrical pharyngeal graph to record muscle contractions associated with blood ingestion and/or salivation and to test effects of neuroactive compounds. Additionally, we will identify feeding-induced changes in R. microplus gene expression with a functional genomics approach. We will identify candidate tick-specific G-protein coupled receptor genes in our R. microplus expressed gene database and agonist/antagonists that affect the function of these GPCRs. Our database of sequenced expressed genes, BACs, and genomic DNA will serve as the foundation for bioinformatic and analytical approaches aimed at finding genes encoding R. microplus GPCRs.