Search

During feeding, a tick's mouthpart penetrates the host's skin and damages tissues and small blood vessels, triggering the extrinsic coagulation and lectin complement pathways. To elude these defense mechanisms, ticks secrete multiple anticoagulant proteins and complement system inhibitors in their saliva. Here, we characterized the inhibitory activities of the homologous tick salivary proteins tick salivary lectin pathway inhibitor, Salp14, and Salp9Pac from Ixodesscapularis in the coagulation cascade and the lectin complement pathway. All three proteins inhibited binding of mannan-binding lectin to the polysaccharide mannan, preventing the activation of the lectin complement pathway. In contrast, only Salp14 showed an appreciable effect on coagulation by prolonging the lag time of thrombin generation. We found that the anticoagulant properties of Salp14 are governed by its basic tail region, which resembles the C terminus of tissue factor pathway inhibitor alpha and blocks the assembly and/or activity of the prothrombinase complex in the same way. Moreover, the Salp14 protein tail contributes to the inhibition of the lectin complement pathway via interaction with mannan binding lectin-associated serine proteases. Furthermore, we identified BaSO 4 -adsorbing protein 1 isolated from the tick Ornithodoros savignyi as a distant homolog of tick salivary lectin pathway inhibitor/Salp14 proteins and showed that it inhibits the lectin complement pathway but not coagulation. The structure of BaSO 4 -adsorbing protein 1, solved here using NMR spectroscopy, indicated that this protein adopts a noncanonical epidermal growth factor domain-like structural fold, the first such report for tick salivary proteins. These data support a mechanism by which tick saliva proteins simultaneously inhibit both the host coagulation cascade and the lectin complement pathway., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Females, nymphs, and larvae of Ixodes silvanus n. sp. collected from birds and from the vegetation in northwestern Argentina (Yungas Phytogeographic Province) are described herein. The new species belongs to the subgenus Trichotoixodes (Acari: Ixodidae). The female is diagnosed by a combination of the following characters: scutum with setae moderately long and more numerous in central field, fewer and moderately long setae on lateral fields, and inconspicuous setae in anterior field; basis capituli subtriangular dorsally; porose areas large and irregular in shape, lacking distinct margins; auriculae with straight edges diverging posterolaterally and ending with small blunt processes; hypostome narrow and pointed with dental formula 4/4 in the anterior third, then 3/3 and 2/2 near the base; coxae I with two spurs, sub-equal in size, internal slightly slimmer than external. The nymph is diagnosed by notum with numerous and long setae, ventral surface covered by numerous whitish setae, scutum with short scapulae and few and shallow punctations, setae on scutum few, short and irregularly distributed, basis capituli sub-triangular dorsally with posterior margin straight, cornua large and directed postero-laterally, auriculae large and projected laterally, lateral margin of basis capituli above auriculae with a lateral and triangular projection, hypostome pointed with dental formula 3/3 in the anterior third and then 2/2, and coxa I with two short, sub-equal, triangular spurs. The diagnostic characters of the larva are: basis capituli dorsally sub-triangular with lateral angles acute and posterior margin straight, auriculae as large triangular lateral projections, hypostome with apex bluntly pointed and dental formula 3/3 in the anterior third and then 2/2, coxa I with two short, sub-equal, triangular spurs, and pattern of dorsal and ventral body setae. This new species is phylogenetically related to Ixodes brunneus, Ixodes turdus and Ixodes frontalis, and the principal hosts for all its parasitic stages are birds., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

Cases of morphological anomalies in the blacklegged tick, Ixodes scapularis (Acari: Ixodidae), have recently been reported from the Northeastern and upper Midwestern United States, potentially complicating identification of this important vector of human disease-causing pathogens. We hereby report a case of a morphological anomaly in I. scapularis, biting a human host residing in Norwich, Connecticut. Using a dichotomous morphological key, high-resolution and scanning electron microscopy images, as well as DNA sequencing, the tick was identified as an adult female I. scapularis with three legs on the left side of the abdomen versus four on the right side, which we believe is the first case of ectromely in an adult I. scapularis. Using diagnostic genes in polymerase chain reaction, the specimen tested positive for Borrelia burgdorferi sensu lato and Anaplasma phagocytophilum, the causative agents for Lyme disease and anaplasmosis, respectively, and also showed evidence of a rickettsial endosymbiont. Here we discuss recent reports of morphological anomalies in I. scapularis, and emphasize the significance of additional studies of teratology in this important tick species and its potential implications., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

The salivary gland of hard ticks is a highly innervated tissue where multiple intertwined axonal projections enter each individual acini. In the present study, we investigated the ultrastructural architecture of axonal projections within granular salivary gland type II and III acini of Ixodes ricinus female. Using immunogold labeling, we specifically examined the associations of SIFamide neuropeptide, SIFamide receptor (SIFa_R), neuropeptide pigment dispersing factor (PDF), and the invertebrate-specific D1-like dopamine receptor (InvD1L), with acinar cells. In both acini types, SIFamide-positive axons were found to be in direct contact with either basal epithelial cells or a single adlumenal myoepithelial cell in close proximity to the either the acinar duct or its valve, respectively. Accordingly, SIFa_R staining correlated with SIFamide-positive axons in both basal epithelial and myoepithelial cells. Immunoreactivity for both InvD1L and PDF (type II acini exclusively) revealed positive axons radiating along the acinar lumen. These axons were primarily enclosed by the adlumenal myoepithelial cell plasma membrane and interstitial projections of ablumenal epithelial cells. Our study has revealed the detailed ultrastructure of I. ricinus salivary glands, and provides a solid baseline for a comprehensive understanding of the cell-axon interactions and their functions in this essential tick organ.

A new species of co-endangered tick, Ixodes heathi n. sp., is described from specimens of the nymph collected on the critically endangered mountain pygmy possum (Burramys parvus Broom) from the alpine region of Victoria, Australia. Its biology is discussed along with strategies for its conservation.

The Haller's organ plays a crucial role in a tick's ability to detect hosts. Even though this sensory organ is vital to tick survival, the morphology of this organ is not well understood. The objective of this study was to characterize variation in the morphological components of the Haller's organ of three medically important tick species using quantitative methods. The Haller's organs of Ixodes scapularis Say (Ixodida: Ixodidae) (black-legged tick), Amblyomma americanum (L.) (Ixodida: Ixodidae) (lone star tick), and Dermacentor variabilis (Say) (Ixodida: Ixodidae) (American dog tick) were morphologically analyzed using environmental scanning electron microscopy and geometric morphometrics, and the results were statistically interpreted using canonical variate analysis. Our data reveal significant, quantitative differences in the morphology of the Haller's organ among all three tick species and that in D. variabilis the sensory structure is sexually dimorphic. Studies like this can serve as a quantitative basis for further studies on sensor physiology, behavior, and tick species life history, potentially leading to novel methods for the prevention of tick-borne disease.

Morphological anomalies have been reported in the blacklegged tick, Ixodes scapularis (Acari: Ixodidae), from the northeastern and upper Midwestern United States, complicating identification of this important vector of human pathogens. Here, we report a case of a local morphological anomaly in I. scapularis, parasitizing a human living in Connecticut. We used a dichotomous morphological key, high-resolution and scanning electron microscopy images, as well as DNA sequencing for identification. The specimen was identified as an adult female I. scapularis with the left leg II split at the trochanter, resulting in two complete legs from the femur to pretarsus giving the appearance of five legs on the left side versus four on the right side. Here we discuss recent reports of morphological anomalies in I. scapularis, and highlight the need for further studies of teratology in this important tick species and its potential implications., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

Molecular determinants and mechanisms of arthropod-borne flavivirus transmission to the vertebrate host are poorly understood. In this study, we show for the first time that a cell line from medically important arthropods, such as ticks, secretes extracellular vesicles (EVs) including exosomes that mediate transmission of flavivirus RNA and proteins to the human cells. Our study shows that tick-borne Langat virus (LGTV), a model pathogen closely related to tick-borne encephalitis virus (TBEV), profusely uses arthropod exosomes for transmission of viral RNA and proteins to the human- skin keratinocytes and blood endothelial cells. Cryo-electron microscopy showed the presence of purified arthropod/neuronal exosomes with the size range of 30 to 200 nm in diameter. Both positive and negative strands of LGTV RNA and viral envelope-protein were detected inside exosomes derived from arthropod, murine and human cells. Detection of Nonstructural 1 (NS1) protein in arthropod and neuronal exosomes further suggested that exosomes contain viral proteins. Viral RNA and proteins in exosomes derived from tick and mammalian cells were secured, highly infectious and replicative in all tested evaluations. Treatment with GW4869, a selective inhibitor that blocks exosome release affected LGTV loads in both arthropod and mammalian cell-derived exosomes. Transwell-migration assays showed that exosomes derived from infected-brain-microvascular endothelial cells (that constitute the blood-brain barrier) facilitated LGTV RNA and protein transmission, crossing of the barriers and infection of neuronal cells. Neuronal infection showed abundant loads of both tick-borne LGTV and mosquito-borne West Nile virus RNA in exosomes. Our data also suggest that exosome-mediated LGTV viral transmission is clathrin-dependent. Collectively, our results suggest that flaviviruses uses arthropod-derived exosomes as a novel means for viral RNA and protein transmission from the vector, and the vertebrate exosomes for dissemination within the host that may subsequently allow neuroinvasion and neuropathogenesis.

Ixodes pacificus is a host of many bacteria including Rickettsia species phylotypes G021 and G022. As part of the overall goal of understanding interactions of phylotypes with their tick host, this study focused on molecular detection of rickettsiae in ovarian and midgut tissue of I. pacificus by fluorescent in situ hybridization (FISH), PCR, and ultrastructural analysis. Of three embedding media (Technovit 8100, Unicryl, and paraffin) tested for generating thin sections, tissues embedded in paraffin resulted in the visualization of bacteria with low autofluorescence in FISH. Digoxigenin-labeled probes were used in FISH to intensify bacterial hybridization signals using Tyramide Signal Amplification reaction. Using this technique, rickettsiae were detected in the cytoplasm of oocytes of I. pacificus. The presence of rickettsiae in the ovary and midgut was further confirmed by PCR and transmission electron microscopic analysis. Overall, the methods in this study can be used to identify locations of tick-borne bacteria in tick tissues and understand transmission routes of bacterial species in ticks., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

The presence of well-developed, elastic claws on ticks and widely pilose hosts led us to hypothesise that ticks are mostly adapted to attachment and locomotion on rough, strongly corrugated and hairy, felt-like substrates. However, by using a combination of morphological and experimental approaches, we visualised the ultrastructure of attachment devices of Ixodes ricinus and showed that this species adheres more strongly to smooth surfaces than to rough ones. Between paired, elongated, curved, elastic claws, I. ricinus bears a large, flexible, foldable adhesive pad, which represents an adaptation to adhesion on smooth surfaces. Accordingly, ticks attached strongest to glass and to surface profiles similar to those of the human skin, generating safety factors (attachment force relative to body weight) up to 534 (females). Considerably lower attachment force was found on silicone substrates and as a result of thanatosis after jolting., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2017. Published by The Company of Biologists Ltd.)

Background: Taxonomic identification of ticks obtained during a longitudinal survey of the critically endangered marsupial, Bettongia penicillata Gray, 1837 (woylie, brush-tailed bettong) revealed a new species of Ixodes Latrielle, 1795. Here we provide morphological data for the female and nymphal life stages of this novel species (Ixodes woyliei n. sp.), in combination with molecular characterisation using the mitochondrial cytochrome c oxidase subunit 1 gene (cox1). In addition, molecular characterisation was conducted on several described Ixodes species and used to provide phylogenetic context., Results: Ixodes spp. ticks were collected from the two remaining indigenous B. penicillata populations in south-western Australia. Of 624 individual B. penicillata sampled, 290 (47%) were host to ticks of the genus Ixodes; specifically I. woyliei n. sp., I. australiensis Neumann, 1904, I. myrmecobii Roberts, 1962, I. tasmani Neumann, 1899 and I. fecialis Warburton & Nuttall, 1909. Of these, 123 (42%) were host to the newly described I. woyliei n. sp. In addition, 268 individuals from sympatric marsupial species (166 Trichosurus vulpecula hypoleucus Wagner, 1855 (brushtail possum), 89 Dasyurus geoffroii Gould, 1841 (Western quoll) and 13 Isoodon obesulus fusciventer Gray, 1841 (southern brown bandicoot)) were sampled for ectoparasites and of these, I. woyliei n. sp. was only found on two I. o. fusciventer., Conclusions: Morphological and molecular data have confirmed the first new Australian Ixodes tick species described in over 50 years, Ixodes woyliei n. sp. Based on the long-term data collected, it appears this tick has a strong predilection for B. penicillata, with 42% of Ixodes infections on this host identified as I. woyliei n. sp. The implications for this host-parasite relationship are unclear but there may be potential for a future co-extinction event. In addition, new molecular data have been generated for collected specimens of I. australiensis, I. tasmani and museum specimens of I. victoriensis Nuttall, 1916, which for the first time provides molecular support for the subgenus Endopalpiger Schulze, 1935 as initially defined. These genetic data provide essential information for future studies relying on genotyping for species identification or for those tackling the phylogenetic relationships of Australian Ixodes species.

Studies of the phenotypic structure of Ixodes persulcatus (Schulze, 1930) populations in relation to their exoskeleton abnormalities are important in both theoretical and practical respects. The data on the species' population structure in Asiatic part of Russia are fragmentary. The goal of the study was to describe taiga tick population structure based on the pattern of females' exoskeleton abnormalities revealed in Asiatic part of Russia. A total of 3872 I. persulcatus females from 16 geographically remote sites of Far Eastern, Siberian, and Ural Federal Districts (FEFD, SFD, and UFD accordingly) were studied. It was demonstrated that all the populations possessed specimens with exoskeleton abnormalities. The «shagreen skin» abnormality was dominant in all these areas. At the same time, the percentage of abnormalities among the specimens collected to the north of 55°N is considerably higher (63.4 ± 3.39 %) than that of samples from the SFD southward territories (33.1 ± 3.43 %). The frequency of abnormalities in its turn is lower (24.4 ± 1.93 %) in the females from the territories with moderate monsoon and moderate continental climate (FEFD) than that in specimens from SFD and UFD areas with sharp continental climate. Thus, such polymorphism of the females' exoskeleton structure may reflect the natural phenogeographical variability of the character rather than the result of the anthropogenic impact. 403

The aim of the study is co-localization of N-glycans with fucose attached to N-acetylglucosamine in α1,3 linkage, that belong to immunogenic carbohydrate epitopes in humans, and N-glycans with α1,6-core fucose typical for mammalian type of N-linked glycosylation. Both glycan epitopes were labelled in cryosections of salivary glands isolated from the tick Ixodes ricinus. Salivary glands secrete during feeding many bioactive molecules and influence both successful feeding and transmission of tick-borne pathogens. For accurate and reliable localization of labelled glycans in both fluorescence and scanning electron microscopes, we used carbon imprints of finder or indexed EM grids on glass slides. We discuss if the topographical images can provide information about labelled structures, the working setting of the field-emission scanning electron microscope and the influence of the detector selection (a below-the-lens Autrata improved YAG detector of back-scattered electrons; in-lens and conventional Everhart-Thornley detectors of secondary electrons) on the imaging of gold nanoparticles, quantum dots and osmium-stained membranes.

Background: Recently a new hard tick species, Ixodes ariadnae has been discovered, adding to the two known ixodid tick species (I. vespertilionis and I. simplex) of bats in Europe., Findings: Scanning electron microscopic comparison of adult females of these species shows morphological differences concerning the palps, the scutum, the Haller's organ, the coxae, as well as the arrangement and fine structure of setae. Molecular analysis of 10 geographically different isolates revealed 90-95% sequence homology in the 12S and 16S rDNA genes of bat tick species. Based on 12S rDNA sequences, genotypes of I. ariadnae clustered closest to I. simplex, whereas according to their 16S rDNA gene they were closest to I. vespertilionis. The subolesin gene of I. ariadnae had only 91% sequence homology with that of I. ricinus, and is the longest known among hard tick species., Conclusions: The present study illustrates the morphology and clarifies the phylogenetic relationships of the three known bat tick species that occur in Europe. According to its subolesin gene I. ariadnae may have a long evolutionary history.

The three most common Ixodes spp. ticks found on songbirds in Western Europe are Ixodes frontalis, I. arboricola and I. ricinus. As the latter species is a generalist, it shares several avian hosts with the two strictly ornithophilic species. Infestations of the three species can overlap in time and space, implying that tick-borne pathogens maintained by the ornithophilic ticks and their hosts could be bridged by I. ricinus to non-avian hosts. Whereas the endophilic Ixodes arboricola only occurs in cavities, I. frontalis has been collected frequently by flagging methods from understory vegetation, which is also the habitat of the field-dwelling I. ricinus. As the latter two species have rather similar morphological characteristics, they can easily be confused with each other. In this study, we present scanning electron photomicrographs of all developmental stages of I. arboricola and I. frontalis, and provide a differential diagnosis key to distinguish the ornithophilic ticks from I. ricinus. In addition, we interpreted their phylogenetic associations based on mitochondrial 16S rDNA with other Ixodes spp. ticks (I. lividus, I. turdus, I. brunneus, I. vespertilionis, I. trianguliceps, I. hexagonus, I. scapularis)., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

Lyme disease, due to infection with the Ixodes-tick transmitted spirochete Borrelia burgdorferi, is the most common tick-transmitted disease in the northern hemisphere. Our understanding of the tick-pathogen-vertebrate host interactions that sustain an enzootic cycle for B. burgdorferi is incomplete. In this article, we describe a method for imaging the feeding of Ixodes scapularis nymphs in real-time using two-photon intravital microscopy and show how this technology can be applied to view the response of Lyme borrelia in the skin of an infected host to tick feeding.

The tick Ixodes ricinus uses its mouthparts to penetrate the skin of its host and to remain attached for about a week, during which time Lyme disease spirochaetes may pass from the tick to the host. To understand how the tick achieves both tasks, penetration and attachment, with the same set of implements, we recorded the insertion events by cinematography, interpreted the mouthparts' function by scanning electron microscopy and identified their points of articulation by confocal microscopy. Our structural dynamic observations suggest that the process of insertion and attachment occurs via a ratchet-like mechanism with two distinct stages. Initially, the two telescoping chelicerae pierce the skin and, by moving alternately, generate a toehold. Subsequently, a breaststroke-like motion, effected by simultaneous flexure and retraction of both chelicerae, pulls in the barbed hypostome. This combination of a flexible, dynamic mechanical ratchet and a static holdfast thus allows the tick to solve the problem of how to penetrate skin and also remain stuck for long periods of time.

Here, we present an investigation on the spatial distribution of the bacterial symbiont Candidatus Midichloria mitochondrii within Ixodes ricinus, by whole mount fluorescence in situ hybridization (FISH). M. mitochondrii is a peculiar, recently discovered bacterium that resides in the mitochondria of female ticks. We applied a rapid and specific FISH protocol with oligonucleotide probes targeted on the 16S rRNA of M. mitochondrii, 12S rRNA of tick mitochondria, and a probe revealing active mitochondria. In this report that represents the first application of whole mount FISH on ticks, we observed strong, specific fluorescence signals in all the examined life stages, as the optimized protocol allowed us to overcome the autofluorescence interference of the cuticle. Cellular localization and quantification of the symbionts were also assessed with electron microscopy and specific real-time PCR, respectively., (Copyright © 2012 Elsevier GmbH. All rights reserved.)

Ixodes festai Rondelli, 1926 is a poorly known bird parasite tick. Its immature forms have not been described yet, while the adult forms only insufficiently, especially the male. In this note the presence of the male of Ixodes festai for the first time in Sardinia (Italy) is reported and a detailed redescription is provided. Morphometric data as well as photographs performed both with optical and electron microscope (ESEM FEI Quanta 200) are also shown.

Formalin fixation is proved to be the useful tool in the method of age estimation for the taiga ticks collected in field. Formalin does not deform fixed material and does not dissolve fat; hence, it is the best fixing fluid when tick age is estimated by the state of fat reserves in its organism. The ticks preserved in 9% formalin were shown to be suitable for age estimation 1.5-2 months after fixation.

A complicated multicellular gland is situated in tarsi of all the legs, occupying one third to half of the segment. Glandular cells form a single-layer sack; inner surface of the sack (gland cavity) is enveloped by a multilayer membrane. Cuticular rods connecting muscles with the claw pass inside this cavity. Glandular cells are characterized by the presence of numerous apical microvilli associated with accumulations of mitochondria, and secretory vacuoles enclosed in cellular membranes. The basal part of each cell contains lipid vacuoles and glycogen granules. The possibility of the track contact pheromone's production by the tarsal gland is discussed.

The genetic identity of Ixodes granulatus ticks was determined for the first time in Taiwan. The phylogenetic relationships were analyzed by comparing the sequences of mitochondrial 16S ribosomal DNA gene obtained from 19 strains of ticks representing seven species of Ixodes and two outgroup species (Rhipicephalus sanguineus and Haemaphysalis inermis). Four major clades could be easily distinguished by neighbour-joining analysis and were congruent by maximum-parsimony method. All these I. granulatus ticks of Taiwan were genetically affiliated to a monophyletic group with highly homogeneous sequences (92.2-99.3% similarity), and can be discriminated from other Ixodes species and other genera of ticks with a sequence divergence ranging from 11.7 to 30.8%. Moreover, intraspecific analysis revealed that two distinct lineages are evident between the same species of I. granulatus ticks collected from Taiwan and Malaysia. Our results demonstrate that all these I. granulatus ticks of Taiwan represent a unique lineage distinct from the common vector ticks (I. ricinus complex) for Borrelia burgdorferi spirochetes.

By methods of light and electron microscopy, dynamics of functioning of salivary glands has been studied in female ticks of the genus Ixodes. Based on literature data and on our own studies, a comparative analysis of morphofunctional changes and of role of salivary gland secretion has been performed in female ticks of subfamilies Ixodinae and Amblyomminae in the process of nutrition.

Ehrlichiae are small gram-negative obligately intracellular bacteria that multiply within vacuoles of their host cells and are associated for a part of their life cycle with ticks, which serve as vectors for vertebrate hosts. Two morphologically and physiologically different ehrlichial cell types, reticulate cells (RC) and dense-cored cells (DC), are observed during experimental infection of cell cultures, mice, and ticks. Dense-cored cells and reticulate cells in vertebrate cell lines alternate in a developmental cycle. We observed ultrastructure of RC and DC of Ehrlichia muris in morulae in salivary gland cells and coinfection with Borrelia burgdorferi sensu lato (sl), "Candidatus Rickettsia tarasevichiae," and a flavivirus (presumably, tick-borne encephalitis virus [TBEV]) of Ixodes persulcatusticks collected in the Cis-Ural region of Russia. Polymerase chain reaction revealed 326 (81.5%) of 400 ticks carrying at least one infectious agent, and 41.5% (166 ticks) were coinfected with two to four agents. Ehrlichiae and rickettsiae were identified by sequencing of 359 bp of the 16S rRNA gene of E. muris and of 440 bp of the 16S rRNA gene and 385 bp of the gltA gene of "R. tarasevichiae." Different organs of the same tick harbored different microorganisms: TBEV in salivary gland and borreliae in midgut; E. muris in salivary gland; and "R. tarasevichiae" in midgut epithelium. Salivary gland cells contained both RC and DC, a finding that confirmed the developmental cycle in naturally infected ticks. Dense-cored cells in tick salivary glands were denser and of more irregular shape than DC in cell cultures. Ehrlichia-infected salivary gland cells had lysed cytoplasm, suggesting pathogenicity of E. muris for the tick host at the cellular level, as well as potential transmission during feeding. Rickettsiae in the midgut epithelial cells multiplied to significant numbers without altering the host cell ultrastructure. This is the first demonstration of E. muris, "R. tarasevichiae," and the ehrlichial developmental cycle in naturally infected I. persulcatus sticks.

Ixodes schulzei Aragão & Fonseca, 1951 is a tick endemic to Brazil, where nine species of Ixodes Latreille, 1796 are currently known to occur. Larvae, nymphs and females of I. schulzei were obtained from a laboratory colony originating from an engorged female collected on a free-living water rat Nectomys squamipes from the Santa Branca municipality, São Paulo State. Only female ticks were obtained from engorged nymphs. Unfed immature and female adult specimens were measured and the descriptions were based on optical and scanning electron microscopy, as were drawings of some features of the larva. Both immature stages present the very long palpi and basis capituli, and the female has large, contiguous porose areas. However, the basis capituli is triangular, with a slight central elevation in the larva and nymph, whereas in the female this area is depressed. The I. schulzei types deposited at the FIOCRUZ (Instituto Oswaldo Cruz) were also examined, as was other material from collections, such as the IBSP (Coleção Acarológica do Instituto Butantan), CNC-FMVZ/USP (Coleção Nacional de Carrapatos da Faculdade de Medicina Veterinária e Zootecnia da USP) and USNTC (United States National Tick Collection). In addition, the relationship between I. schulzei and other immature neotropical species of Ixodes is discussed.

An intracellular bacterium with the unique ability to enter mitochondria exists in the European vector of Lyme disease, the hard tick Ixodes ricinus. Previous phylogenetic analyses based on 16S rRNA gene sequences suggested that the bacterium formed a divergent lineage within the Rickettsiales (Alphaproteobacteria). Here, we present additional phylogenetic evidence, based on the gyrB gene sequence, that confirms the phylogenetic position of the bacterium. Based on these data, as well as electron microscopy (EM), in situ hybridization and other observations, we propose the name 'Candidatus Midichloria mitochondrii' for this bacterium. The symbiont appears to be ubiquitous in females of I. ricinus across the tick's distribution, while lower prevalence is observed in males (44%). Based on EM and in situ hybridization studies, the presence of 'Candidatus M. mitochondrii' in females appears to be restricted to ovarian cells. The bacterium was found to be localized both in the cytoplasm and in the intermembrane space of the mitochondria of ovarian cells. 'Candidatus M. mitochondrii' is the first bacterium to be identified that resides within animal mitochondria.

The female of Ixodes stilesi Neumann, 1911 (Acari Ixodidae) is redescribed and the male and nymph are described from specimens collected from Pudu puda (Molina) (Artiodactyla: Cervidae) in Chile. Both sexes of I. stilesi have characteristics of the subgenera Ixodes Latreille, 1795 and Ixodiopsis Filippova, 1957. The females of I. stilesi are peculiar in having the combination of the sinuous scutum outline, rounded porose areas with distinct borders separated by the width of one area, slender and long palpi, and two subequal spurs on coxa I. The male is unique in having a combination of a posteriorly wrinkled marginal folder, a basis capituli longer than wide, a non-crenulate hypostome toothed portion, two spurs on coxa II to IV and the presence of a pseudoscutum. The nymph of I. stilesi has blunt anterior and posterior processes on palpal article I (characteristics of the subgenus Ixodiopsis and some Pholeoixodes Schulze, 1942) and a wing-shaped basis capituli with a prominent triangular cornua. Phylogenetic analyses based on 16S mitochondrial rDNA sequences of 12 Neotropical and two Australian Ixodes species, plus three argasids, were carried out to clarify the position of I. stilesi. The results of phylogenetic analyses and morphological characters indicate a close relationships between I. stilesi and two other Neotropical species of uncertain subgeneric status, I. neuquenensis Ringuelet, 1947 and I. sigelos Keirans, Clifford & Corwin, 1976.

Fine structure of salivary glands was investigated in the tick Ixodes persulcatus females. Ultrastructure of II and III acini in unfed and feeding females is described. Changes in size and structure of the secretory products during feeding are demonstrated.

A site-specific glycosylation of salivary glands (SGs) isolated from unfed and partially fed Ixodes ricinus females was identified with the use of lectin affinity labeling on sections and western blots of SDS-PAGE gels. The results revealed that secretory granules of a, b, and c cells of the type II acinus and e and f cells of the type III acinus are glycosylated. In partially engorged tick SGs, 2 subtypes of c cells were distinguished. The granules of c1 cells contained mannose, N-acetyl-D-glucosamine, and sialic acid residues. The granules of b, c2, and e cells exhibited complex glycoconjugates rich in mannose, N-acetyl-D-glucosamine, galactose, N-acetyl-D-galactosamine, and a moderate amount of sialic acid. The granules of f cells contained N-acetyl-D-glucosamine and mannose moieties. Type III acini surfaces were covered with mannose-specific ConA binding sites. Except the granules of salivary cells, sialic acid-specific lectins MAA II and SNA strongly bound cuticular structures of alveolar ducts, and weakly with the cuticular spiral thread of excretory salivary ducts. The total sialic acid level in SG homogenates isolated from partially fed females was determined by the thiobarbituric acid method. Sialic acid, which has been found during the development of a few insect species, has not been reported in ticks as yet.

Ticks of the genus Ixodes are poorly known in the Neotropical zoogeographical region, from where only 45 species have been recorded. In Brazil, the genus is currently represented by eight species, four of which are known only from this country. This paper presents a redescription of all active stages in the life-cycle of I. loricatus , based on optical and scanning electron microscopy. In addition, the relationship of I. loricatus to other Neotropical Ixodes is presented and discussed.

The first tarsal segment of the taiga tick bears 4 general types of sensilla (except for sensilla forming the Haller's sensory organ on the dorsal surface of the tarsus): tactile mechanoreceptor sensilla of 3 types, contact chemo-mechanoreceptor sensilla of 2 types, and double-walled pore sensilla. One of these types, the chemo-mechanoreceptor upper-pore sensilla, was found only in the Ixodinae, and not found in the examined representatives of the Amblyomminae. This type of sensilla was also found in the palpal receptor organ of the ixodid and argasid ticks.

Objective: To observe the ultrastructural changes in the midgut epithelium of Ixodes sinensis after infesting rabbits immunized with Mr 105000 purified tick antigen., Methods: New Zealand rabbits were inoculated with Mr 105000 purified antigen by means of mutiple intradermal injection in foot pad, groin and back. Each immunized rabbit was infested by 30 female Ixodes sinensis. At 24 hours, 48 hours, 72 hours, 5 days and 8 days after infestation, three Ixodes sinensis in each group were observed for ultrastructural changes in the epithelium of their midgut., Results: Histological examinations showed that with the time going, digestive cells of the ticks after infesting hosts became more and larger with dense and regularly arranged microvilli, enriched organella, distinct unit-membrane structure, and the appearance of tubli, small vacuole, numerous lipid droplets and hematin granules. These cells also developed a highly infolded basal lamina, forming a labyrinth system. The digestive cells of immunized group were however greatly damaged, whose number and volume were significantly different from control groups. From 24 to 48 hours after infestation, the midgut epithelium of Ixodes snenss showed pathological changes with the basal lamina becoming thinner, looser and broken; digestive cells damaged and vacuolated; microvilli decreased, shortened and irregularly arranged; the mitochondria swollen and its crests reduced, shortened and even with myeloid changes; the rough endoplasmic reticulum dilated; lipid droplets and hematin granules decreased; phagocytic and pinocytic activity weakened; and basal labyrinth system vacuolated. From 72 hours to 8 days after infestation, cells were severely damaged, organella were denatured and necrotic, nuclei showed pyknosis and cells lysed., Conclusion: The rabbits immunized with Mr 105000 purified ixodic protein have acquired the adoptive immunity against Ixodes sinensis; in the anti-tick immunity described above, the midgut of Ixodes sinensis is the major affected site.

Extract of steer wool odor was found to excite olfactory receptor(s) in a wall-pore olfactory sensillum on the distal knoll of the Haller's organ. Three active volatile compounds were revealed in this odor by gas chromatography. Electrophysiological experiments revealed two types of receptors (sensory neurons) within the sensilla examined. One type of receptor responded only to phenolic derivatives, such as o-chlorophenol, o-bromophenol, o-methylphenol, 2,6-dichlorophenol, 2,6-dibromophenol, 2,4,6-trichlorophenol, but not to o-nithrophenol, p-methylphenol, 2,5-dichlorophenol, 3,5-dichlorophenol, 2,6-dinithrophenol, 2,6-dimethylphenol, and pentachlorophenol. The other type of receptor responded only to gamma-valerolactone. It is assumed that these cells play an important role in perception of a host from long distances (10-15 m), which is typical of Ixodes ricinus ticks.

The formation of the peritrophic matrix in the midgut of females of 5 ixodid tick species (Ixodes pacificus, I. pavlovskyi, I. persulcatus, I. ricinus and I. scapularis) was studied by means of light and electron microscopy in different periods of the feeding and after detachment. The formation of the peritrophic matrix started when the first food portions came into the gut lumen, 9-12 hours after the attachment. Renovation of the peritrophic matrix took place during the whole feeding period; every new generation of midgut cells synthesized their own matrix. It was deposited on the apical surface of every midgut cell in the beginning of differentiation, and was functioning during the life of the cell. The peritrophic matrix separates spaces of the cavitary and cytozoic digestions.

The behaviour of Ixodes ricinus nymphs was recorded in 10-day experiments using computer-assisted video-tracking, in the absence of any host stimuli. These ticks switch spontaneously from questing in a desiccating atmosphere to quiescence in a water-saturated atmosphere after dark. Quantification of both questing and quiescence duration demonstrates that questing duration is inversely related to saturation deficit whereas quiescence duration is not. Distance walked after quiescence increased with desiccating conditions, while the distance walked after questing remained unchanged. Almost all locomotor activities of I. ricinus occurred during darkness under either a 14 h:10 h L:D or a 8 h:4 h L:D cycle. We established that all life stages of I. ricinus are equipped to sense shifts in light intensity with bilaterally placed strings of photoreceptors. This permits I. ricinus to use onset of darkness to trigger mobility when desiccation risk is reduced in nature.

We examined a parthenogenetic strain of the hard tick Ixodes woodi Bishopp for the presence of endosymbiotic bacteria. Electron microscopic examination revealed the ovarian tissues and Malpighian tubules were infected with pleomorphic bacteria. Two basic types were observed: a larger granular cell and a smaller condensed cell. Cloning and sequence analysis of polymerase chain reaction (PCR) amplified 16S rRNA gene yielded a single sequence from bacteria present in I. woodi tissues. Phylogenetic analysis of the nearly complete 16S rDNA indicated that the ticks were infected with an endosymbiont belonging to the gamma subdivision of the Proteobacteria. It clustered with the insect pathogenic species Rickettsiellagrylli (Vago and Martoja 1963) and the animal pathogen Coxiella burnetii (Derrick 1939) Philip 1948. Our results suggest that the I. woodi females harbored a single endosymbiotic bacterium related to selected Rickettsiella species and to C burnetii.

The cleaning of ixodid ticks for microscopy can be achieved quickly and efficiently using a combination of a wax solvent and an ultrasonic cleaner. The technique involves minimum handling of specimens, produces no detectable damage and is suitable for cleaning many specimens at the same time.

The chalcid wasp Ixodiphagus hookeri (Howard) is a parasitoid of several ixodid ticks including the blacklegged tick, Ixodes scapularis Say. We evaluated effects of the feeding process of nymphal I. scapularis on the embryonic development of I. hookeri. Potentially wasp-parasitized nymphal I. scapularis were collected on Prudence Island, RI. Subsamples of the questing nymph cohort were allowed to feed on laboratory white mice. Both the body length and the scutal length of ticks were measured individually for questing nymphs and for feeding nymphs that were removed from hosts at time intervals of 12, 24, 36, 48, and 72 h after attachment. The diameters of wasp eggs they contained were also measured for each designated time interval. There was a positive relationship between the mean scutal index (ratio between body length and scutal length) of ticks and the mean diameter of wasp eggs during 72 h of tick feeding (P < 0.05). Moreover, it appeared that within 24 h of tick attachment, the scutal index of ticks remained unchanged. However, after that period, the scutal index increased significantly (P < 0.05). Diameters of wasp eggs increased continuously during tick feeding and at 72 h after attachment, enclosed eggs and completely formed larvae were found in several ticks. We conclude that factors related to the feeding process of nymphal I. scapularis are necessary to initiate the embryonic development of wasps.

The entomopathogenic fungus Metarhizium anisopliae is highly pathogenic to the black-legged tick, Ixodes scapularis. Spore concentrations of 10(8)/ml for engorged larvae and 10(7)/ml for engorged females resulted in 100% tick mortality, 2 wk postinfection. The LC50 value for engorged larvae (concentration to kill 50% of ticks) was 10(7) spores/ml. Metarhizium anisopliae shows considerable potential as a microbial control agent for the management of Ixodes scapularis.

The internal organs of Ixodes ovatus and the ears of wild rodents (Apodemus speciosus, Eothenomys smithii) and an insectivore (Crocidura dsinezumi) were cultured to isolate borreliae; positive samples were examined for the distribution and dissemination of spirochetes in the host tissues using electron microscopy. Seven isolates were derived from the unfed ticks and the three species of mammals. These isolates were identified as Borrelia japonica judging from the outer surface protein profile using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reactivity to a B. japonica-specific monoclonal antibody. Borreliae were found only in the midgut lumen of the tick in close contact with the microvilli on the midgut epithelium; on the other hand, borreliae found in the ears of mammals existed freely in the collagenous intercellular substances of connective tissues or in close contact with fibrocytes. The ultrastructural disparities between the borreliae in ticks and mammals appeared to correspond to differences in motility. Interestingly, the borrelia which invaded through the perineurium appeared to contact the basement membrane of a Schwann cell that enclosed several nonmyelinated nerve fibers. This may offer important information regarding the involvement of the nervous system in Lyme disease.

Nymphal and larval stages of Ixodes (Ixodes) jellisoni Cooley & Kohls and I. (I.) neotomae Cooley are described for the first time. These 2 tick species occur only in the western United States, predominantly in California. The primary host for I. jellisoni is the California kangaroo rat, Dipodomys californicus (Merriam); that for I. neotomae is the dusky-footed woodrat, Neotoma fuscipes Baird. The etiologic agent of Lyme disease Borrelia burgdorferi Johnson, Schmidt, Hyde, Steigerwalt & Brenner has recently been isolated from both tick species, and I. neotomae was proven a competent enzootic vector of the Lyme disease spirochete.

Lectins have been suggested to function as pattern-recognition molecules in invertebrate immune mechanisms. A lectin from the hemolymph of the tick Ixodes ricinus with main specificity for sialic acid was characterized and antibodies directed against this lectin were prepared. In this study, these antibodies were used to localize the lectin in the tissues of I. ricinus. Immunoreactivity with poly- and monoclonal antibodies was detected in the granules of both types of granular hemocytes, at the membrane of hemocytes, and at the basal laminae surrounding the hemocoel. Furthermore, cells attached to the midgut, invaginations of Géné's organ, and granular inclusions of nephrocytes were labeled. The immunoreactivity detected in hemocytes and the hemocoel lining supports the idea that the hemolymph lectin may function as a recognition molecule in the immune system of I. ricinus. Another function could be protection of eggs that are coated with secretions by Géné's organ. The lectin activity could also be involved in transmission of Borrelia burgdorferi, the causative agent of Lyme borreliosis, and the tick-borne encephalitis virus.

A 58-year old housewife consulted us about 1 cm sized, dark-brownish, bean-like mass which was dropped spontaneously from indurated skin lesion on her abdomen. The mass was identified morphologically as an engorged female Ixodes nipponensis. Nine days earlier, she had an excursion collecting edible sprouts of wild grass. Spontaneous retreat has been unusual in clinical tick bites in Korea. Fourteen cases of tick bite described in the Korean literature were reviewed briefly in relation to Lyme borreliosis.