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.

Tick and Host Derived Compounds Detected in the Cement Complex Substance

“…Some of the tick-derived proteins identified in this study in the R. microplus cementome have been previously reported and characterized in the tick sialome/sialotranscriptome (e.g., Table S5). Similarly, host-derived proteins such as immunoglobulins and apolipoprotein A-I identified here in the cementome have been previously described in the sialome [6,12,54,[79][80][81][82][83][84][85] and in unfed ticks [86,87]. However, the cementome protein content described here supports a highly complex composition of both tick and host derived proteins involved in multiple biological processes such as cement formation, solidification and maintenance, feeding, interference with host immune response, and detachment playing a key role in tick biology.…”

“…Based on a novel methodology for proteomics data analysis using the protein representation profiles after significant differences in pairwise comparisons between time points, proteins were putatively assigned to different categories of developmental processes. This methodology provides a functional relevant alternative to other methodologies such as the analyses using QuiXoT [32] or normalized spectral abundance factors (NSAF) and z-score statistics [54]. Additionally, the first analysis of the chemical elements using SEM combined with EDS allowed the characterization of changes in their composition in tick salivary glands and cement during tick feeding, which correlated with changes in protein profiles.…”

“…Furthermore, the profile of the highly represented Gly (C 2 H 5 NO 2 )-rich superfamily member proteins in the cementome that have been implicated in tick response to oxidative stress [20] showed a decrease in protein representation during tick feeding (e.g., A0A224YEQ4; Data S4), which correlated with changes in the composition of O and N chemical elements ( Figure 8B). Glycine-rich proteins are highly represented in the sialome of different tick species and were also shown to decrease in abundance during feeding of adult female A. americanum [54]. Changes in the composition of cement chemical elements during tick feeding may be also related to physiological mechanisms such as the increase in the intensity of respiratory patterns in R. sanguineus ticks during feeding [55].…”

The Central Role of Salivary Metalloproteases in Host Acquired Resistance to Tick Feeding

“…Insect apolipophorins show a resemblance to tick vitellogenins and carrier proteins (Donohue et al, 2009), all of which are derived from a common ancestor and belong to the large lipid transfer protein superfamily (Smolenaars et al, 2007). This finding is not new, as homologous lipoproteins have previously been identified proteomically in pilocarpine-induced saliva of partially-and fullyfed R. microplus ticks (Tirloni et al, 2014), dopamine-induced saliva of partially-fed D. andersoni (Mudenda et al, 2014), dopamine-and pilocarpine-induced saliva of H. longicornis nymphs and adults (Tirloni et al, 2015), pilocarpine-induced saliva of I. scapularis (Kim et al, 2016), A. americanum (Kim et al, 2020), as well as by antibody-mediated identification in phage display cDNA expression SG libraries from early timepoints of A. americanum feeding (Radulovic et al, 2014). Genetically tick-resistant bovines recognize more tick salivary proteins than sera of susceptible ones.…”

“…Metalloproteases comprise the majority of coding sequences in the tick salivary gland transcriptomes, accounting for more than 5% of the total read of the entire transcriptome (Schwarz et al, 2013), as well as of salivary proteomes of various tick species, e.g. Dermacentor andersoni (Mudenda et al, 2014), Haemaphysalis longicornis (Tirloni et al, 2015), Ixodes scapularis (Kim et al, 2016), A. americanum (Kim et al, 2020). Despite their abundance, the physiological function of most metalloproteases in tick saliva remains unknown, although some were characterized as fibrinogenases (Francischetti et al, 2003).…”