Project description:A 16S rRNA gene approach, including 454 pyrosequencing and quantitative PCR (qPCR), was used to describe the bacterial community in Rhipicephalus turanicus and to evaluate the dynamics of key bacterial tenants of adult ticks during the active questing season. The bacterial community structure of Rh. turanicus was characterized by high dominance of Coxiella and Rickettsia and extremely low taxonomic diversity. Parallel diagnostic PCR further revealed a novel Coxiella species which was present and numerically dominant in all individual ticks tested (n = 187). Coxiella sp. densities were significantly higher in female versus male ticks and were overall stable throughout the questing season. In addition, we revealed the presence of the novel Coxiella sp. in Rh. sanguineus adult ticks, eggs, and hatched larvae, indicating its vertical transmission. The presence of both spotted fever group Rickettsia spp. (SFGR) and non-SFGR was verified in the various individual ticks. The prevalence and density of Rickettsia spp. were very low compared to those of Coxiella sp. Furthermore, Rickettsia sp. densities were similar in males and females and significantly declined toward the end of the questing season. No correlation was found between Coxiella sp. and Rickettsia sp. densities. These results suggest different control mechanisms in the tick over its different bacterial populations and point to an obligatory and facultative association between the two tick species and Coxiella sp. and Rickettsia spp., respectively.
Project description:Rhipicephalus sanguineus belongs to a complex of hard tick species with high veterinary-medical significance. Recently, new phylogenetic units have been discovered within R. sanguineus, which therefore needs taxonomic revision. The present study was initiated to provide new information on the phylogeography of relevant haplotypes from less studied regions of Europe and Africa. With this aim, molecular-phylogenetic analyses of two mitochondrial markers were performed on 50 ticks collected in Hungary, the Balkans, countries along the Mediterranean Sea, Kenya and Ivory Coast.In the "temperate lineage" of R. sanguineus, based on cytochrome c oxidase subunit 1 (cox1) and 16S rRNA genes, Rhipicephalus sp. I was only found in the eastern part of the Mediterranean Basin (with relatively homogenous haplotypes), whereas Rhipicephalus sp. II occurred in the middle-to-western part of this region (with phylogenetically dichotomous haplotypes). Ticks identified as R. leporis (based on morphology and cox1 gene) were found in Kenya and Ivory Coast. These clustered phylogenetically within R. sanguineus (s.l.) ("tropical lineage").In the Mediterranean Basin two mitochondrial lineages of R. sanguineus, i.e. Rhipicephalus sp. I and Rhipicephalus sp. II exist, which show different geographical distribution. Therefore, data from this study confirm limited gene flow between Rhipicephalus sp. I and Rhipicephalus sp. II, but more evidence (analyses of nuclear markers, extensive morphological and biological comparison etc.) are necessary to infer if they belong to different species or not. The phylogenetic relationships of eastern and western African ticks, which align with R. leporis, need to be studied further within R. sanguineus (s.l.) ("tropical lineage").
Project description:The taxonomic status of the brown dog tick Rhipicephalus sanguineus (sensu stricto) is a subject of on-going debate; there is a consensus that populations of this tick species should be referred to as R. sanguineus (sensu lato) until its taxonomic status is resolved. Recent genetic studies revealed the existence of more than one lineage of R. sanguineus (s.l.) in temperate countries. In this study, we assessed the genetic identity of ticks collected from rural dogs living in several areas located in all major geographical regions of Portugal.A total of 347 ticks were collected from rural dogs living in different regions of Portugal. These ticks were morphologically identified and partial mitochondrial 16S rRNA gene sequences (~300 bp) were obtained from representative specimens.The ticks were morphologically identified as Ixodes ricinus (seven males and 27 females), Rhipicephalus bursa (one male), Rhipicephalus pusillus (one female) and R. sanguineus (s.l.) (two larvae, 101 nymphs, 108 males and 100 females). Partial mitochondrial 16S rRNA gene sequences were obtained from 58 R. sanguineus (s.l.) specimens, and all of them were genetically identified as belonging to the so-called temperate lineage of R. sanguineus (s.l.) CONCLUSIONS: These results strongly suggest that the temperate species of R. sanguineus (s.l.) is the only representative of this tick group found on dogs in Portugal. It also adds weight to the hypothesis that Rhipicephalus turanicus is not present in this country, although further investigations are necessary to confirm this.
Project description:BACKGROUND:The brown dog tick Rhipicephalus sanguineus (sensu stricto) is reputed to be the most widespread tick of domestic dogs worldwide and has also been implicated in the transmission of many pathogens to dogs and humans. For more than two centuries, Rh. sanguineus (s.s.) was regarded as a single taxon, even considering its poor original description and the inexistence of a type specimen. However, genetic and crossbreeding experiments have indicated the existence of at least two distinct taxa within this name: the so-called "temperate" and "tropical" lineages of Rh. sanguineus (sensu lato). Recent genetic studies have also demonstrated the existence of additional lineages of Rh. sanguineus (s.l.) in Europe and Asia. Herein, we assessed the biological compatibility between two lineages of Rh. sanguineus (s.l.) found in southern Europe, namely Rhipicephalus sp. I (from Italy) and Rhipicephalus sp. II (from Portugal). METHODS:Ticks morphologically identified as Rh. sanguineus (s.l.) were collected in southern Portugal and southern Italy. Tick colonies were established and crossbreeding experiments conducted. Morphological, biological and genetic analyses were conducted. RESULTS:Crossbreeding experiments confirmed that ticks from the two studied lineages were able to mate and generate fertile hybrids. Hybrid adult ticks always presented the same genotype of the mother, confirming maternal inheritance of mtDNA. However, larvae and nymphs originated from Rhipicephalus sp. I females presented mtDNA genotype of either Rhipicephalus sp. I or Rhipicephalus sp. II, suggesting the occurrence of paternal inheritance or mitochondrial heteroplasmy. While biologically compatible, these lineages are distinct genetically and phenotypically. CONCLUSIONS:The temperate lineages of Rh. sanguineus (s.l.) studied herein are biologically compatible and genetic data obtained from both pure and hybrid lines indicate the occurrence of paternal inheritance or mitochondrial heteroplasmy. This study opens new research avenues and raises question regarding the usefulness of genetic data and crossbreeding experiments as criteria for the definition of cryptic species in ticks.
Project description:Brown dog ticks morphologically identifiable as Rhipicephalus sanguineus sensu lato, are distributed world-wide and their systematics is controversial. Results of genetic and reproductive compatibility studies of geographically distinct populations of R. sanguineus s.l. indicate that the R. sanguineus complex is paraphyletic. To further elucidate systematic relationships within R. sanguineus s.l. and geographic boundaries of its lineages, we conducted a phylogeographical study of 136 tick specimens from 23 countries. Voucher specimens were morphologically identified. A phylogenetic tree was constructed using concatenated partial mitochondrial 12S and 16S rDNA gene sequences and analyzed by the Neighbor-Joining method. A set of 19 bioclimatic variables within the WorldClim dataset were extracted and analyzed to assess correlations between distribution of R. sanguineus s.l. lineages and climatic variables. The following four branches are clearly recognized on the phylogenetic tree: R. sanguineus s.l.-tropical and temperate clades, R. leporis, and R. turanicus. DNA sequences of Rhipicephalus ticks from Israel differ from those of other groups. Strong association between geographical locations of major clades of R. sanguineus s.l. and temperature was identified. The tropical clade of R. sanguineus s.l. occupies areas with the annual mean temperature >20 °C, whereas the temperate clade is present in areas with the annual mean temperature <20 °C. Our results indicate that ticks in two closely related phylogenetic clades are adapted to different environmental conditions and support proposals for re-classification of R. sanguineus complex. Differences in R. sanguineus s.l. ecology and human/animal pathogens transmitted by different taxa of brown dog tick need to be studied.
Project description:Calreticulin 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.We 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.All 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.We 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.
Project description:The brown dog tick (Rhipicephalus sanguineus) is the most widespread tick in the world and a well-recognized vector of many pathogens affecting dogs and occasionally humans. This tick can be found on dogs living in both urban and rural areas, being highly adapted to live within human dwellings and being active throughout the year not only in tropical and subtropical regions, but also in some temperate areas. Depending on factors such as climate and host availability, Rh. sanguineus can complete up to four generations per year. Recent studies have demonstrated that ticks exposed to high temperatures attach and feed on humans and rabbits more rapidly. This observation suggests that the risk of human parasitism by Rh. sanguineus could increase in areas experiencing warmer and/or longer summers, consequently increasing the risk of transmission of zoonotic agents (e.g., Rickettsia conorii and Rickettsia rickettsii). In the present article, some aspects of the biology and ecology of Rh. sanguineus ticks are discussed including the possible impact of current climate changes on populations of this tick around the world.
Project description:Obligatory hematophagous arthropods such as lice, bugs, flies, and ticks harbor bacterial endosymbionts that are expected to complement missing essential nutrients in their diet. Genomic and some experimental evidence support this expectation. Hard ticks (Acari: Ixodidae) are associated with several lineages of bacterial symbionts, and very few were experimentally shown to be essential to some aspects of tick's fitness. In order to pinpoint the nature of interactions between hard ticks and their symbionts, we tested the effect of massive elimination of Coxiella-like endosymbionts (CLE) by antibiotics on the development and fitness of the brown dog tick (Rhipicephalus sanguineus). Administration of ofloxacin to engorged (blood fed) nymphs resulted in significant and acute reduction of their CLE loads - an effect that also persisted in subsequent life stages (aposymbiotic ticks). As a result, the post-feeding development of aposymbiotic female (but not male) nymphs was delayed. Additionally, aposymbiotic adult females needed a significantly prolonged feeding period in order to replete (detach from host), and had reduced engorgement weight and a lower capacity to produce eggs. Consequently, their fecundity and fertility were significantly reduced. Eggs produced by aposymbiotic females were free of CLE, and the resulting aposymbiotic larvae were unable to feed successfully. Our findings demonstrate that the observed fitness effects are due to CLE reduction and not due to antibiotic administration. Additionally, we suggest that the contribution of CLE is not mandatory for oocyte development and embryogenesis, but is required during feeding in females, when blood meal processing and tissue buildup are taking place. Presumably, under these extreme physiological demands, CLE contribute to R. sanguineus through supplementing essential micro- and macronutrients. Further nutrient complementary studies are required to support this hypothesis.