Abstract
BackgroundCalreticulin proteins (CRTs) are important components of tick saliva, which is involved in the blood meal success, pathogen transmission and host allergic responses. The characterization of the genes encoding for salivary proteins, such as CRTs, is pivotal to understand the mechanisms of tick-host interaction during blood meal and to develop tick control strategies based on their inhibition. In hard ticks, crt genes were shown to have only one intron with conserved position among species. In this study we investigated the exon-intron structure and variation of the crt gene in Rhipicephalus spp. ticks in order to assess the crt exon-intron structure and the potential utility of crt gene as a molecular marker.MethodsWe sequenced the exon-intron region of crt gene in ticks belonging to so-called tropical and temperate lineages of Rhipicephalus sanguineus (sensu lato), Rhipicephalus sp. I, Rhipicephalus sp. III, Rhipicephalus sp. IV, R. guilhoni, R. muhsamae and R. turanicus. Genetic divergence and phylogenetic relationships between the sequences obtained were estimated.ResultsAll individuals belonging to the tropical lineage of R. sanguineus (s.l.), R. guilhoni, R. muhsamae, R. turanicus, Rhipicephalus sp. III and Rhipicephalus sp. IV analysed showed crt intron-present alleles. However, both crt intron-present and intron-absent alleles were found in Rhipicephalus sp. I and the temperate lineage of R. sanguineus (s.l.), showing the occurrence of an intraspecific intron presence-absence polymorphism. Phylogenetic relationships among the crt intron-present sequences showed distinct lineages for all taxa, with the tropical and temperate lineages of R. sanguineus (s.l.) being more closely related to each other.ConclusionsWe expanded previous studies about the characterization of crt gene in hard ticks. Our results highlighted a previously overlooked variation in the crt structure among Rhipicephalus spp., and among hard ticks in general. Notably, the intron presence/absence polymorphism observed herein can be a candidate study-system to investigate the early stages of intron gain/loss before fixation at species level and some debated questions about intron evolution. Finally, the sequence variation observed supports the suitability of the crt gene for molecular recognition of Rhipicephalus spp. and for phylogenetic studies in association with other markers.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-016-1909-3) contains supplementary material, which is available to authorized users.
Highlights
Calreticulin proteins (CRTs) are important components of tick saliva, which is involved in the blood meal success, pathogen transmission and host allergic responses
Several studies were focused on the genetic characterization of CRTs and to date crt genes have been sequenced in several hard tick species [23,24,25,26]
Comparative analysis of the exon-intron structure of crt genes in 28 hard tick species belonging to different genera, including Amblyomma, Dermacentor, Ixodes and Rhipicephalus showed that (i) two exons and only one intron are present in tick crt genes, contrary to what was observed in other invertebrate and vertebrate species; (ii) the intron position is conserved in hard ticks, the intron size and nucleotide sequences vary among species [24, 25]
Summary
Calreticulin proteins (CRTs) are important components of tick saliva, which is involved in the blood meal success, pathogen transmission and host allergic responses. CRTs are important components of tick saliva, which plays a pivotal role in tick feeding and pathogen transmission to hosts by acting as anti-haemostatic, anti-inflammatory and immunomodulatory molecules [7,8,9,10,11,12,13,14,15] In this context, CRTs have been suggested to play a modulating role in host haemostasis [9, 16, 17] and have been shown to be highly immunogenic to tick mammalian hosts [18,19,20,21]. Crt genes could be good target sites for control strategies based on gene expression silencing approaches such as RNA interference (RNAi), as their silencing could affect the success of tick blood feeding and pathogen transmission and tick physiology and fitness [27, 28]
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