Analysis of the Salivary Gland Transcriptome of Unfed and Partially Fed Amblyomma sculptum Ticks and Descriptive Proteome of the Saliva

Abstract

Ticks are obligate blood feeding ectoparasites that transmit a wide variety of pathogenic microorganisms to their vertebrate hosts. Amblyomma sculptum is vector of Rickettsia rickettsii, the causative agent of Rocky Mountain spotted fever (RMSF), the most lethal rickettsiosis that affects humans. It is known that the transmission of pathogens by ticks is mainly associated with the physiology of the feeding process. Pathogens that are acquired with the blood meal must first colonize the tick gut and later the salivary glands (SG) in order to be transmitted during a subsequent blood feeding via saliva. Tick saliva contains a complex mixture of bioactive molecules with anticlotting, antiplatelet aggregation, vasodilatory, anti-inflammatory, and immunomodulatory properties to counteract both the hemostasis and defense mechanisms of the host. Besides facilitating tick feeding, the properties of saliva may also benefits survival and establishment of pathogens in the host. In the current study, we compared the sialotranscriptome of unfed A. sculptum ticks and those fed for 72 h on rabbits using next generation RNA sequencing (RNA-seq). The total of reads obtained were assembled in 9,560 coding sequences (CDSs) distributed in different functional classes. CDSs encoding secreted proteins, including lipocalins, mucins, protease inhibitors, glycine-rich proteins, metalloproteases, 8.9 kDa superfamily members, and immunity-related proteins were mostly upregulated by blood feeding. Selected CDSs were analyzed by real-time quantitative polymerase chain reaction preceded by reverse transcription (RT-qPCR), corroborating the transcriptional profile obtained by RNA-seq. Finally, high-performance liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analysis revealed 124 proteins in saliva of ticks fed for 96–120 h. The corresponding CDSs of 59 of these proteins were upregulated in SG of fed ticks. To the best of our knowledge, this is the first report on the proteome of A. sculptum saliva. The functional characterization of the identified proteins might reveal potential targets to develop vaccines for tick control and/or blocking of R. rickettsii transmission as well as pharmacological bioproducts with antihemostatic, anti-inflammatory and antibacterial activities.

Wonders of tick saliva

“…Currently, ~70,000 protein sequences deposited on GenBank are assigned to tick salivary glands . Transcriptomic studies indicate ticks of a given species may secrete >500 different proteins and peptides in their saliva over the course of feeding (Giovanni et al, 2014;Esteves et al, 2017;Perner et al, 2018). These are grouped into several multigene families, including lipocalins, Kunitz-domain containing proteins/peptides, metalloproteases, and basic tail secreted proteins, and several families apparently unique to ticks.…”

“…Current sequence-based approaches and bioinformatics reinforce evidence ticks have opted for a 'redundant' system (Ribeiro et al, 2012;Díaz-Martín et al, 2013;Esteves et al, 2017;Hackenberg et al, 2017). Thus, the same host homeostatic mechanism is targeted by more than one tick saliva molecule.…”

Gene Expression in the Salivary Gland of Rhipicephalus (Boophilus) microplus Fed on Tick-Susceptible and Tick-Resistant Hosts

“…microplus belonging to the trypsin inhibitor-like cysteine-rich domain family showed strong inhibitory activity toward elastase, which participates in the inflammatory response and is involved in injury caused by tick fixation on bovines (Sasaki et al, 2008). A secreted cysteine-rich protein containing the TIL domain was upregulated in the proteome of fed Amblyomma sculptum, induced by blood feeding, compared with non-fed ticks (Esteves et al, 2017). In R.…”

An economic dilemma between weapon systems may explain an arachno-atypical venom in wasp spiders (Argiope bruennichi)

“…The A. bruennichi IGFBP is closely related to a protein found in the venom of the tiger wandering spider Cupiennius salei (Kuhn-Nentwig et al, 2011). Such proteins are commonly found in arachnid venoms but also in scorpions (Superstitia donensis, Hadrurus spadix and Centruroides hentzi) and ticks of the genus Amblyomma (Santibanez-Lopez et al, 2016;Esteves et al, 2017;Rokyta and Ward, 2017;Ward et al, 2017). A related protein is present in the venom of A.…”