A novel SINE family occurs frequently in both genomic DNA and transcribed sequences in ixodid ticks of the arthropod sub-phylum Chelicerata

Reassociation kinetics and flow cytometry data indicate that ixodid tick genomes are large, relative to most arthropods, containing>or=10(9) base pairs. The molecular basis for this is unknown. We have identified a novel small interspersed element with features of a tRNA-derived SINE, designated Ruka, in genomic sequences of Rhipicephalus appendiculatus and Boophilus (Rhipicephalus) microplus ticks. The SINE was also identified in expressed sequence tag (EST) databases derived from several tissues in four species of ixodid ticks, namely R. appendiculatus, B. (R.) microplus, Amblyomma variegatum and also the more distantly related Ixodes scapularis. Secondary structure predictions indicated that Ruka could adopt a tRNA structure that was, atypically, most similar to a serine tRNA. By extrapolation the frequency of occurrence in the randomly selected BAC clone sequences is consistent with approximately 65,000 copies of Ruka in the R. appendiculatus genome. Real time PCR analyses on genomic DNA indicate copy numbers for specific Ruka subsets between 5800 and 38,000. Several putative conserved Ruka insertion sites were identified in EST sequences of three ixodid tick species based on the flanking sequences associated with the SINEs, indicating that some Ruka transpositions probably occurred prior to speciation within the metastriate division of the Ixodidae. The data strongly suggest that Class I transposable elements form a significant component of tick genomes and may partially account for the large genome sizes observed.     

Ixodid ticks (Acari: Ixodidae) infesting humans in Tokat Province of Turkey: species diversity and seasonal activity

Ixodid ticks (Acari: Ixodidae) are the major vectors of pathogens threatening animal and human health. Tokat Province, Turkey, is a suitable habitat for extended tick activity with its moderate climate and vegetation. In the present study, we surveyed humans visiting health centers to determine the species diversity, geographical distribution, and seasonal activity of ixodid ticks infesting them. Out of 5,999 adult ticks collected from humans from April to September, 2008, 800 ticks were identified to species, while the remaining were identified to genus according to their distinct morphological characteristics. Hyalomma marginatum, Hyalomma detritum, Rhipicephalus bursa, Rhipicephalus (Boophilus) annulatus, Haemaphysalis punctata, Dermacentor marginatus, and Ixodes laguri were the most prevalent tick species among 24 ixodid tick species infesting humans in the region. One of these tick species, Hyalomma isacii was identified as a new record for the ixodid tick fauna of Turkey. Hyalomma species were the most abundant in summer, while Dermacentor and Ixodes species displayed the lowest frequency. Hyalomma aegyptium infestation was very common on humans in the province. Results indicated that a variety of ixodid tick species infest humans depending on the season in the target area. It is possible that a variety of ixodid tick species may contribute to the spread of tick‐borne diseases such as Crimean‐Congo haemorrhagic fever virus (CCHFV), which is endemic in the region.     

Midichloria mitochondrii is widespread in hard ticks (Ixodidae) and resides in the mitochondria of phylogenetically diverse species

The hard tick Ixodes ricinus (Ixodidae) is the sole animal thus far shown to harbour an intra-mitochondrial bacterium, which has recently been named Midichloria mitochondrii. The objectives of this work were (i) to screen ixodid ticks for Midichloria-related bacteria and (ii) to determine whether these bacteria exploit the intra-mitochondrial niche in other tick species. Our main goal was to discover further models of this peculiar form of symbiosis. We have thus performed a PCR screening for Midichloria-related bacteria in samples of ixodid ticks collected in Italy, North America and Iceland. A total of 7 newly examined species from 5 genera were found positive for bacteria closely related to M. mitochondrii. Samples of the tick species Rhipicephalus bursa, found positive in the PCR screening, were analysed with transmission electron microscopy, which revealed the presence of bacteria both in the cytoplasm and in the mitochondria of the oocytes. There is thus evidence that bacteria invade mitochondria in at least 2 tick species. Phylogenetic analysis on the bacterial 16S rRNA gene sequences generated from positive specimens revealed that the bacteria form a monophyletic group within the order Rickettsiales. The phylogeny of Midichloria symbionts and related bacteria does not appear completely congruent with the phylogeny of the hosts.     

Mammals as hosts of Ixodid ticks (Acarina,Ixodidae)

Trophic association between adult ticks of the world fauna with their mammal hosts (511 tick species out of 650 species of Ixodidae of the world fauna) are reviewed. Specific tick species parasitize on representatives of 22 (out of 27) mammal orders, but species of Paucituberculata, Notoryctemorphia, Peramelemorphia, Scandentia, and Dermoptera have no specific parasites. The highest number of tick species was recorded from Artiodactyla and rodents. Mammals, on which representatives of all the 14 genera of the family parasitize, provide the diversity of the ixodid tick fauna. The conclusion on the priority of ecological factors in comparison with phylogenetic status of hosts in the formation of specific characteristics of ticks confirms Balashov’s (1982) concept on the character of host-parasite association between ticks and mammals.     

Variation in genome size of argasid and ixodid ticks

The suborder Ixodida includes many tick species of medical and veterinary importance, but little is known about the genomic characteristics of these ticks. We report the first study to determine genome size in two species of Argasidae (soft ticks) and seven species of Ixodidae (hard ticks) using flow cytometry analysis of fluorescent stained nuclei. Our results indicate a large haploid genome size (1C>1000 Mbp) for all Ixodida with a mean of 1281 Mbp (1.31+/-0.07 pg) for the Argasidae and 2671 Mbp (2.73+/-0.04 pg) for the Ixodidae. The haploid genome size of Ixodes scapularis was determined to be 2262 Mbp. We observed inter- and intra-familial variation in genome size as well as variation between strains of the same species. We explore the implications of these results for tick genome evolution and tick genomics research.     

Megaselia scalaris reared on Rhipicephalus (Boophilus) microplus laboratory cultures

Different laboratory cultures of the acarine tick Rhipicephalus (Boophilus) microplus (Canestrini, 1888) (Ixodida: Ixodidae) were infested by small Megaselia scalaris (Loew, 1866) (Diptera: Phoridae) flies. Larvae of this species exhibited opportunistic parasitism predominantly on engorged female ticks, causing severe damage to their cuticle through which the flies were able to reach R. microplus internal organs, on which they fed until developing into pupae in the tick's remains. The flies were kept by continuous propagation on fresh ticks over six generations during which the same parasitoid behaviour was observed. Here we report on an ixodid tick laboratory culture used for rearing M. scalaris.     

The ixodid tick species attaching to domestic dogs and cats in Great Britain and Ireland

The species of ixodid ticks, attached to dogs and cats presented to veterinary practices in Great Britain and Ireland were identified. Most host animals carried only one tick species with Ixodes ricinus Linné (Acari: Ixodidae) being the most common, identified on 52% of animals, Ixodes hexagonus Leach (Acari: Ixodidae) the second most common (on 39%) and Ixodes canisuga Johnston (Acari: Ixodidae) the third most common (on 11%). A significantly higher proportion of dogs than cats carried I. ricinus, while I. hexagonus was more frequently carried by cats. One animal carried a single specimen of Haemaphysalis punctata Canestrini & Fanzago (Acari: Ixodidae), one carried a Dermacentor reticulatus Fabricius (Acari: Ixodidae) but none carried Rhipicephalus sanguineus Latreille (Acari: Ixodidae). This indicates that the latter two species, vectors of 'exotic' tick-borne diseases, remain at low densities in Great Britain and Ireland. Retrospective information on exposure of the animals to different habitats and geographic regions was collected by questionnaire and subject to contingency table and logistic regression analysis. Woodlands and moorlands were habitats significantly associated with I. ricinus attachment. Exposure to urban parks was significantly associated with I. hexagonus attachment and exposure to boarding kennels and catteries was significantly associated with I. canisuga attachment. Ixodes hexagonus, rather than I. ricinus, was the ixodid tick species most likely to be encountered by urban populations of dogs and cats and, by inference, possibly also humans. The implications of these findings, for the transmission of tick-borne pathogens to dogs, cats and humans are discussed.     

Aggregation in the questing tick, Rhipicephalus pulchellus

ABSTRACT. The ixodid tick, Rhipicephalus pulchellus (Ixodidae) can be observed aggregated on the tips of grass stems in the field in Kenya. From laboratory experiments, Browning (1976) concluded that these aggregations were the result of ticks having first aggregated on the ground if provided with sufficient cover. During our extension of Browning's experiments using several types of cover, the ticks did not exhibit aggregating behaviour. The influence of the presence of a human observer on the activity of the ticks, however, became apparent.     

An updated list of the ticks of Ghana and an assessment of the distribution of the ticks of Ghanaian wild mammals in different vegetation zones

Twenty one species of ticks belonging to five genera of the family Ixodidae (Order Acari, sub-order Ixodida) - Amblyomma, Haemaphysalis, Hyalomma, Ixodes and Rhipicephalus (including the sub-genus Rhipicephalus (Boophilus)) - were collected from 1260 mammals, representing 29 species, 14 families and 6 orders, in four vegetation zones in Ghana during the period 1971-1978. Four other species were collected from humans in 1977. In all, eight species appeared to be new records for Ghana: Amblyomma tholloni Neumann; Dermacentor circumguttatus Neumann; Haemaphysalis houyi Nuttall & Warburton; Ixodes loveridgei Arthur; Ixodes oldi Nuttall; Ixodes vanidicus Schultze; Rhipicephalus complanatus Neumann; Rhipicephalus cuspidatus Neumann. The updated list of tick species in Ghana given here includes 41 species of ixodid ticks and four species of argasid ticks. Most species have been found in neighbouring regions of West Africa but 56 of the 121 different combinations of ixodid tick species and host species found in the collection described here have not apparently been reported before. The new combinations recorded here bring the total number of different combinations of ixodid tick species and mammalian host species now reported in Ghana to 151. The tick species found on wild mammals in Ghana mostly differed from those reported from domestic stock by other authors. The data showed that different tick species occurred in different vegetation zones and that most species displayed a pronounced preference for certain groups of related host species. Some tick species were found in the savanna feeding mainly on large bovids and/or suids; others were found in forests feeding mainly on small bovids, large rodents or small carnivores.     

Birds as hosts of ixodid ticks (Acarina,Ixodidae)

rophic relations between ixodid ticks of the world fauna parasitizing as adults on birds are analyzed. These ticks include 60 out of 650 species of the family Ixodidae and belong to the genera Ixodes, Haemaphysalis, and Amblyomma. Relations between birds and ticks of the genus Ixodes are most ancient. Colonial marine birds seem to be the first hosts of ixodid ticks in the class of birds. Trophic specialization of ticks is not associated with bird taxa, because in most cases no tick specificity is caused by either phylogenetic features of hosts, or specific environmental conditions.     

The interrelations of the causative agents of transmissible diseases in ixodid ticks (Ixodidae) with a mixed infection

The data on interactions between pathogens of human diseases with natural focality in ixodid ticks (Ixodidae) with mixed infection are reviewed. Different variants of mixed infection revealed in nature and accompanied by predominantly extracellular, or both extra- and intracellular location of the agents in tick organism are considered. Natural mixed infection of ticks by different pathogens is a normal and commonly occurring phenomenon; moreover, there is no experimental evidence for the existence of any antagonistic relationships between spirochetes, rickettsiae, and piroplasms in such ticks. The absence of antagonism between different agents in vectors largely provides a relative autonomy of corresponding parasitic systems and creates conditions for coexistence of different parasitic systems in mixed natural foci.     

Comparison of Preservation Methods of Rhipicephalus appendiculatus (Acari: Ixodidae) for Reliable DNA Amplification by PCR

Five differently preserved groups of adult Rhipicephalus appendiculatus specimens were compared for quality of DNA extracted. Three methods were used to extract DNA from specimens i.e. two simple mosquito validated DNA extraction methods and a tick validated method. Extraction of DNA from tick legs was attempted. The quality of DNA extracted was evaluated by the success of PCR amplification of the ITS2 gene and the mitochondrial COI gene fragment. Fresh specimens (i.e. killed just before extraction) had the highest success of DNA amplification followed by specimens killed in ethanol and subsequently stored in the refrigerator (4 °C). There was no significant difference in amplification success between cryopreserved and 70% ethanol preserved specimens. It was possible to amplify DNA from legs of ticks. Sequenced ITS2 amplicon of template obtained from legs of ticks was as legible as those from whole tick extract. The two mosquito validated DNA extraction methods showed a significantly lower amplification success than the tick validated protocol.     

Species,developmental stage and infection with microbial pathogens of engorged ticks removed from dogs and questing ticks

Research into tick‐borne diseases implies vector sampling and the detection and identification of microbial pathogens. Ticks were collected simultaneously from dogs that had been exposed to tick bites and by flagging the ground in the area in which the dogs had been exposed. In total, 200 ticks were sampled, of which 104 came from dogs and 96 were collected by flagging. These ticks were subsequently examined for DNA of Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum, Rickettsia spp. and Babesia canis. A mixed sample of adult ticks and nymphs of Ixodes ricinus (Ixodida: Ixodidae) and Haemaphysalis concinna (Ixodida: Ixodidae) was obtained by flagging. Female I. ricinus and adult Dermacentor reticulatus (Ixodida: Ixodidae) ticks dominated the engorged ticks removed from dogs. Rickettsia spp. were detected in 17.0% of the examined ticks, A. phagocytophilum in 3.5%, B. canis in 1.5%, and B. burgdorferi s.l. in 16.0%. Ticks with multiple infections were found only among the flagging sample. The ticks removed from the dogs included 22 infected ticks, whereas the flagging sample included 44 infected ticks. The results showed that the method for collecting ticks influences the species composition of the sample and enables the detection of a different pattern of pathogens. Sampling strategies should be taken into consideration when interpreting studies on tick‐borne pathogens.     

Validation and rapid extraction of nucleic acids from alcohol-preserved ticks

A method has been developed to extract DNA from alcohol-preserved ticks (Acari: Ixodidae and Argasidae). The method combines the lysing property of the chaotropic agent guanidinium thiocyanate (GuSCN) and the nucleic acid-binding property of diatomaceous earth (fossilized cell walls of unicellular algae). Debris from the tick is removed in several sequential washing steps. To monitor the efficiency of this method, a polymerase chain reaction (PCR) was designed to amplify the 16S mt rRNA gene of five tick genera (Dermacentor Fabricius, Haemaphysalis Koch, Rhipicephalus Koch, Argas Latreille and Ixodes Latreille). Detection of amplification products from this PCR indicated that DNA had been successfully extracted and that Taq-polymerase inhibitors were absent. The extraction method, therefore, enables purification of DNA such that enzymatic analysis is possible.     

Ticks Feeding on Humans: A Review of Records on Human-Biting Ixodoidea with Special Reference to Pathogen Transmission

In this article, literature records of argasid and ixodid ticks feeding on humans worldwide are provided in view of increased awareness of risks associated with tick bites. Ticks can cause paralyses, toxicoses, allergic reactions and are vectors of a broad range of viral, rickettsial, bacterial and protozoan pathogens. Approximately 12 argasid species (Argas and Ornithodos) are frequently found attached to humans who intrude into tick-infested caves and burrows. Over 20 ixodid tick species are often found on humans exposed to infested vegetation: four of these are Amblyomma species, 7 Dermacentor spp., 3 Haemaphysalis spp., 2 Hyalomma spp. and 6 Ixodes species. Personal protection methods, such as repellents and acaricide-impregnated clothing are advised to minimize contact with infected ticks. Acaricidal control of ixodid ticks is impractical because of their wide distribution in forested areas, but houses infested with soft ticks can be sprayed with acaricidal formulations. Attached ticks should be removed without delay. The best way is to grasp the tick as close to the skin as possible with fine tweezers and pull firmly and steadily without twisting. Finally, despite the fact that most people who are bitten destroy the offending tick in disgust, it is recommended that they preserve specimens in ethanol for taxonomic identification and detection of pathogens by molecular methods.     

Female ixodid ticks grow endocuticle during the rapid phase of engorgement

Lees (Proc Zool Soc Lond 121:759–772, 1952) concluded that the ixodid tick Ixodes ricinus grows endocuticle during the slow but not during the rapid, phase of engorgement, a conclusion supported by Andersen and Roepstorff (Insect Biochem Mol Biol 35:1181–1188, 2005) for the same species. In this study analysis of dimensional data and cuticle weight measurements from female ixodid ticks (Amblyomma hebraeum) were used to test this hypothesis. Both approaches showed that endocuticle growth continues during the rapid phase, tapering to zero at a fed/unfed weight ratio of ~60. Of the total mass of cuticle in the engorged tick 32–43% was formed during the rapid phase. We demonstrate that if cuticle growth stopped at the end of the slow phase, there would not be sufficient cuticle to account for the thickness of cuticle observed at the end of engorgement. This finding is consistent with prior studies of Rhipicephalus (Boophilus) microplus, and with a dimensional analysis of the cuticle thickness data of Lees for I. ricinus, in contradiction to his conclusion from an analysis of tick cuticle weight measurements. An examination of cuticle weight measurements for I. ricinus by Andersen and Roepstorff similarly supports the finding of cuticle growth during the rapid phase. All ixodid ticks undergo major body expansion, typically tenfold or more, during a rapid phase of engorgement and require sufficient cuticle at the end of that process to contain their body. The fact that cuticle grows during the rapid phase of engorgement in three species suggests that this is a general characteristic of the family Ixodidae.     

Evaluating levels of PCR efficiency and genotyping error in DNA extracted from engorged and non‐engorged female Dermacentor variabilis ticks

Polymerase chain reaction (PCR)‐based methods are increasingly used to elucidate tick biology. However, DNA extracted from ticks may provide poor PCR templates as a result of PCR inhibition by mammalian blood or contamination by male DNA (in fertilized females). In this study, the effects of removing the bloodmeal and reproductive organs were evaluated through paired DNA extractions in engorged and non‐engorged Dermacentor variabilis (Say) (Acari: Ixodidae), prior to PCR amplification at 12 microsatellites. The first extraction utilized only mouthparts and legs (‘mouthpart’ samples) and the second utilized tick bodies (‘body’ samples). The results indicated that contamination by male DNA was an unlikely source of genotyping error in mouthpart and body samples. Engorged females showed higher levels of PCR inhibition in body vs. mouthpart samples, with a 29% decrease in amplification success rates per PCR and a 10‐fold increase in levels of missing genotypes in body samples. By contrast, non‐engorged females showed little difference in amplification success rates or numbers of missing genotypes in body vs. mouthpart samples. We discuss analytical concerns related to this systematic bias in PCR problems and recommend the removal of the bloodmeal and reproductive organs prior to DNA extraction, especially in engorged female ticks.     

Phylogeographic relationships of Ixodes uriae (Acari: Ixodidae) and their significance to transequatorial dispersal of Borrelia garinii

The seabird tick Ixodes uriae (Acari: Ixodidae) has a bi- and circumpolar distribution and is commonly infected with Lyme disease Borrelia. Identical Borrelia flagellin gene sequences have been detected in I. uriae from both the Northern and Southern Hemispheres, suggesting a transequatorial transport of Borrelia. Parsimony analysis of the internal transcribed spacer 2 (ITS2) and a part of 16S rDNA of I. uriae from the Northern and Southern Hemispheres indicated that northern and southern I. uriae might be reproductively separated. We hypothesize that Borrelia is probably not dispersed from one hemisphere to the other by ticks attached to seabirds.