Structural characterization of tick cement cones collected from in vivo and artificial membrane blood-fed Lone Star ticks (Amblyomma americanum)

Abstract

The Lone Star tick, Amblyomma americanum, is endemic to the southeastern United States and capable of transmitting pathogenic diseases and causing non-pathogenic conditions. To remain firmly attached to the host, the tick secretes a proteinaceous matrix termed the cement cone which hardens around the tick’s mouthparts to assist in the attachment of the tick as well as to protect the mouthparts from the host immune system. Cement cones collected from ticks on a host are commonly contaminated with host skin and hair making analysis of the cone difficult. To reduce the contamination found in the cement cone, we have adapted an artificial membrane feeding system used to feed long mouthpart ticks. Cones collected from in vivo and membrane fed ticks are analyzed to determine changes in the cone morphology. Comparisons of the cement cones using light microscopy shows similar structures and color however using scanning electron microscopy the cones have drastically different structures. The in vivo cones contain fibrils, sheets, and are heavily textured whereas cones from membrane fed ticks are remarkably smooth with no distinct structures. Analysis of the secondary protein structures using FTIR-ATR show both in vivo and membrane fed cement cones contain β sheets but only in vivo cement cones contain helical protein structures. Additionally, proteomic analysis using LC–MS/MS identifies many proteins including glycine rich proteins, metalloproteases, and protease inhibitors. Proteomic analysis of the cones identified both secreted and non-secreted tick proteins. Artificial membrane feeding is a suitable model for increased collection of cement cones for proteomic analysis however, structurally there are significant differences.

Tick attachment cement – reviewing the mysteries of a biological skin plug system

“…Another approach in cement analysis was to obtain material free of host skin tissue (Bullard et al ., 2016) by in vitro feeding of ticks on artificial membranes (Fig. 1).…”

“…1). Compared to in vivo feeding, this method allows monitoring the progress of cement deposition and contamination‐free collection of samples (Bullard et al ., 2016). …”

“…Bullard et al . (2016) reported that microlitre amounts of saliva can dissolve tick cement rapidly. Due to the high protein content of the cement, it might be expected that a protease is involved.…”

“…The introduction of artificial feeding systems on silicone membranes allowed the isolation of cement cones of Amblyomma americanum free from host skin tissue for comparison with cement from in vivo ‐fed ticks of this species (Bullard et al ., 2016). Cement originating from in vitro feeding allowed the identification of more proteins than cones harvested from in vivo systems.…”

“…Bullard et al . (2016) hypothesised that GRPs may stabilize and strengthen the cement cone and contribute to its insolubility.…”