Project description:The tick midgut is the main tissue involved in blood feeding and the first organ to have contact with pathogens ingested through the blood meal. We characterized the early transcriptional changes in the midgut of O. moubata in the unfed, engorged and Borrelia duttonii-infected state. The aim is to identify transcripts that are differentially expressed in the respective physiological state.
Project description:We developed a new simple method to assess the composition of proteins in the Ornithodoros moubata saliva. To collect naturally expectorated saliva from the ticks, we employed an artificial membrane feeding technique using a simple, chemically defined diet and submitted the collected native saliva samples for liquid chromatography-mass spectrometry (LC-MS) analysis. These experiments were conducted with groups of uninfected ticks as well as with B. duttonii infected ticks. The ticks exhibited a fair feeding response to the tested diet, engorgement rates reaching between 60-100% of ticks per feeding chamber. LC-MS analysis identified a total of 17 and 15 proteins in saliva samples from the uninfected and infected ticks respectively. Importantly, the analysis was sensitive enough to detect up to 9 different proteins in the saliva containing diet upon which as few as 6 nymphal ticks fed. Some of the proteins identified are well known for their immunomodulatory activity in a vertebrate host, whereas others were structural or “housekeeping” proteins and their finding in the naturally expectorated saliva confirms that they can be secreted and having some roles at the tick-host interface. Most notably, some of the proteins that have long been suspected as important for vector-pathogen interactions of Borrelia spirochetes were detected only in the samples from infected ticks, suggesting that their expression was altered by the persistent colonization of the tick’s salivary glands by spirochetes. The simple method described herein is an important addition to the toolbox available to study the vector-host-pathogen interactions in the rapidly feeding soft ticks.
Project description:The specific aim of the present work is to expand the data on the salivary proteome of O. moubata adult ticks, particularly of female ticks, by solving the aforementioned drawbacks, as part of the in depth characterization and analysis of the O. moubata sialome. Knowing this sialome will allow identifying and selecting novel salivary antigens as targets for tick vaccines, and their subsequent testing in animal immunization trials. We pay special attention to female ticks because they are a key developmental stage in the tick life cycle and vaccines may exert on them double deleterious effects, for instance an increase of mortality and a reduction/inhibition of the reproductive performance. With that aim, in the current work we have analysed samples of female and male saliva separately using two different mass spectrometry approaches: data-dependent acquisition (DDA) LC-MS/MS and Sequential Window Acquisition of all Theoretical fragment ion spectra Mass Spectrometry (SWATH MS). This last technique is a specific variant of data-independent acquisition (DIA) methods and is emerging as a technology that combines deep proteome coverage capabilities with quantitative consistency and accuracy.
Project description:The specific aim of the present work is to expand the data on the salivary proteome of O. moubata adult ticks, particularly of female ticks, by solving the aforementioned drawbacks, as part of the in depth characterization and analysis of the O. moubata sialome. Knowing this sialome will allow identifying and selecting novel salivary antigens as targets for tick vaccines, and their subsequent testing in animal immunization trials. We pay special attention to female ticks because they are a key developmental stage in the tick life cycle and vaccines may exert on them double deleterious effects, for instance an increase of mortality and a reduction/inhibition of the reproductive performance. With that aim, in the current work we have analysed samples of female and male saliva separately using two different mass spectrometry approaches: data-dependent acquisition (DDA) LC-MS/MS and Sequential Window Acquisition of all Theoretical fragment ion spectra Mass Spectrometry (SWATH MS). This last technique is a specific variant of data-independent acquisition (DIA) methods and is emerging as a technology that combines deep proteome coverage capabilities with quantitative consistency and accuracy.
Project description:The specific aim of the present work is to expand the data on the salivary proteome of O. moubata adult ticks, particularly of female ticks, by solving the aforementioned drawbacks, as part of the in depth characterization and analysis of the O. moubata sialome. Knowing this sialome will allow identifying and selecting novel salivary antigens as targets for tick vaccines, and their subsequent testing in animal immunization trials. We pay special attention to female ticks because they are a key developmental stage in the tick life cycle and vaccines may exert on them double deleterious effects, for instance an increase of mortality and a reduction/inhibition of the reproductive performance. With that aim, in the current work we have analysed samples of female and male saliva separately using two different mass spectrometry approaches: data-dependent acquisition (DDA) LC-MS/MS and Sequential Window Acquisition of all Theoretical fragment ion spectra Mass Spectrometry (SWATH MS). This last technique is a specific variant of data-independent acquisition (DIA) methods and is emerging as a technology that combines deep proteome coverage capabilities with quantitative consistency and accuracy.
