Time-resolved proteomic profile of Amblyomma americanum tick saliva during feeding

Abstract

Amblyomma americanum ticks transmit more than a third of human tick-borne disease (TBD) agents in the United States. Tick saliva proteins are critical to success of ticks as vectors of TBD agents, and thus might serve as targets in tick antigen-based vaccines to prevent TBD infections. We describe a systems biology approach to identify, by LC-MS/MS, saliva proteins (tick = 1182, rabbit = 335) that A. americanum ticks likely inject into the host every 24 h during the first 8 days of feeding, and towards the end of feeding. Searching against entries in GenBank grouped tick and rabbit proteins into 27 and 25 functional categories. Aside from housekeeping-like proteins, majority of tick saliva proteins belong to the tick-specific (no homology to non-tick organisms: 32%), protease inhibitors (13%), proteases (8%), glycine-rich proteins (6%) and lipocalins (4%) categories. Global secretion dynamics analysis suggests that majority (74%) of proteins in this study are associated with regulating initial tick feeding functions and transmission of pathogens as they are secreted within 24-48 h of tick attachment. Comparative analysis of the A. americanum tick saliva proteome to five other tick saliva proteomes identified 284 conserved tick saliva proteins: we speculate that these regulate critical tick feeding functions and might serve as tick vaccine antigens. We discuss our findings in the context of understanding A. americanum tick feeding physiology as a means through which we can find effective targets for a vaccine against tick feeding.

Fig 1. Tick saliva proteins composition in Amblyomma americanum tick saliva every 24 h during feeding.

Pilocarpine induced A. americanum tick saliva was subjected to LC-MS/MS sequencing using the “In-Solution” digestion approach as described in materials and methods. Cumulative normalized spectral abundance factors (NSAF) values as index for relative abundance of detected tick saliva proteins are presented for each functional class. Please note the labels are oriented horizontally and read left to right starting from cytoskeletal proteins (red), antioxidant/detoxification (green), extracellular matrix (orange), then back to glycine rich proteins (light-tan), immune related (pink) and continues this pattern.

Fig 2. Rabbit host proteins composition in Amblyomma americanum tick saliva every 24 h during feeding.

Pilocarpine induced A. americanum tick saliva was subjected to LC-MS/MS sequencing using the “In-Solution” digestion approach as described in materials and methods. Cumulative normalized spectral abundance factors (NSAF) values as index for relative abundance of detected rabbit host proteins are presented for each functional class. Please note the labels are oriented horizontally and read left to right starting from antimicrobial proteins (red), antioxidant/detoxification (green), cytoskeletal (yellow), then back to globin/RBC (light-green), fibrinogen (pink) and continues this pattern.

Fig 3. Overall secretion dynamics of tick saliva protein classes in Amblyomma americanum tick saliva.

Normalized spectral abundance factors (NSAF) values of tick was normalized using the z-score statistics and then used to generate heat maps using heatmap2 function in gplots library using R as described in materials and methods.

Fig 4. Overall secretion dynamics of rabbit host protein classes in Amblyomma americanum tick saliva.

Normalized spectral abundance factors (NSAF) values of rabbit host was normalized using the z-score statistics and then used to generate heat maps using heatmap2 function in gplots library using R as described in materials and methods.