The tick Ixodes ricinus: distribution and climate preferences in the western Palaearctic

In this study, multivariate spatial clustering on monthly normalized difference vegetation index (NDVI) maps is used to classify ecological regions over the western Palaearctic. This classification is then used to delineate the distribution and climate preferences of populations (clades) of the tick Ixodes ricinus L. (Acari: Ixodidae) from a geographically extensive dataset of tick records and a gridded 2.5-km resolution climate dataset. Using monthly layers of the NDVI, regions of similar ecological attributes were defined and nine populations with significant differences in critical climate parameters (P< 0.005) were detected. Grouping of tick records according to other categories, such as political divisions, a 4 degrees x 4 degrees grid overlying the study area, or the CORINE) and USGS) vegetation classification schemes did not provided significantly separated populations (P = 0.094-0.304). Factor analysis and hierarchical tree clustering provided an ecological overview of these tick clades: two Mediterranean and one Scandinavian (western) clades are clearly separated from a node that includes clades of different parts of central Europe and the British Isles, with contrasting affinities between the different clades. The capture records of these ecologically separated clades produce a clear bias when bioclimate envelope modelling is applied to the mapping of habitat suitability for the tick in the western Palaearctic. The best-performing methods (Cohen's kappa = 0.834-0.912) use partial models developed with data from each ecoregion, which are then overlapped over the region of study. It is concluded that the use of ecologically derived ecoregions is an objective step in assessing the presence of ecologically different clades, and provides a guide in the development of data partitioning for habitat suitability modelling.     

Longitudinal surveillance of the tick Ixodes ricinus for borreliae

Host-seeking Ixodes ricinus (L.) (Acari: Ixodidae) were monitored for borreliae (Borrelia burgdorferi s.l.) using dark-field microscopy in South Moravia (Czech Republic) each May from 1991 to 2001 (150 nymphs, 100 females and 100 males each year). This survey revealed a mean annual percentage of infected ticks of 16.8% (range, 11.7-24.2) in nymphs, 24.9% (range, 16.5-33.6) in females and 26.1% (range, 17.1-37.3) in males. Annual incidence of Lyme borreliosis in humans of the area in the same period (range, 8.7-41.7 per 100,000) correlated significantly with the frequency (number of ticks per flag per hour) of nymphs infected with >50 borreliae or all nymphal ticks, but not with the frequency of females, infected females or the infection rate (% of ticks infected) of either nymphal or female ticks. A prediction of the annual incidence of Lyme borreliosis, based on the frequency of heavily infected or all nymphal I. ricinus ticks, is feasible. The infection rate in I. ricinus correlated significantly with the North Atlantic Oscillation winter index of the last year (in nymphs) or of the year before last (in adults).     

Chrological and physiological age of the tick Ixodes ricinus females

Changes of the physiological age in 3 groups of the laboratory reared Ixodes ricinus females were investigated over the period of 6-16 months. The longevity was studied in 27 groups of the tick females from a wild population during 2 seasons of the tick activity. Physiological age of the tick females was determined at the beginning of the study and then monthly till the end of the study. It was established that physiological ageing takes place during the life of the tick, and the ageing of different females is carried out non-uniformly. Physiologically young females (of the second physiological age) in the groups of laboratory reared ticks were occurred right up to 11th month after molting (including 4 months of hibernation in a refrigerator). Ticks of the second physiological age were occurred in the groups from wild populations after one or two periods of hibernation. Hence, in natural conditions some females may be physiologically young both in the first and second seasons of the ticks' activity. On the other hand, physiologically old tick females (of the 4th physiological age) appeared in the groups in a month after molting. The supposition is proposed that the tick population is heterogeneous by the rate of the individual ageing. It means that different specimens differ by the rate of consumption of their storage compounds i.e. by the intensity of metabolism. Such difference may be a cause of the variability in chemical conditions in the gut of the tick being the habitat of Borrelia.     

The extensible alloscutal cuticle of the tick, Ixodes ricinus

The proteins in the distensible alloscutal cuticle of the blood-feeding tick, Ixodes ricinus, have been characterized by electrophoresis and chromatography, two of the proteins were purified and their total amino acid sequence determined. They show sequence similarity to cuticular proteins from the spider, Araneus diadematus, and the horseshoe crab, Limulus polyphemus, and to a lesser extent to insect cuticular proteins. They contain a conserved sequence region, which is closely related to the chitin-binding Rebers-Riddiford consensus sequence present in many insect cuticular proteins. Only a fraction of the alloscutal proteins can be readily dissolved, and the dissolved proteins are difficult to separate by electrophoresis and column chromatography. The insoluble fraction can only be dissolved after degradation to smaller peptides. The mixture of extractable proteins as well as hydrolysates of the insoluble fraction are fluorescent when exposed to ultraviolet light, and the fluorescence corresponds in excitation and emission maxima to the fluorescence of the rubber-like arthropodan protein, resilin, and to the amino acid dityrosine. Small amounts of dityrosine were obtained from ticks in the early phase of a blood meal when the cuticle weighs less than 4 mg; increasing amounts were obtained from animals in the initial period of feeding, during which the cuticular weight increases from 4 to 11 mg, whereas little increase in dityrosine content was observed during the final period of engorgement. Cuticle from fully distended ticks contains about 60-80 nmole dityrosine per tick, corresponding to 2-3 microg/mg cuticle. It is suggested that the major part of the cuticular proteins is made inextractable by cross-linking by dityrosine residues, and that dityrosine plays a role in stabilizing the cuticular structure during the extensive distension occurring during a blood meal. Small amounts of 3-monochlorotyrosine and 3,5-dichlorotyrosine were obtained from the distended tick cuticle, corresponding to chlorination of between 0.5% and 1.5% of the tyrosine residues. It is suggested that the chlorotyrosines are a side-product of oxidative processes in the cuticle.     

Uptake and incorporation of sialic acid by the tick Ixodes ricinus

We describe the detection of sialylated N-linked glycans in partially fed Ixodes ricinus tick females using matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry. Sialylated glycans were detected in salivary glands as well as in tick guts and we propose the host origin of these structures. In addition, we mapped the transport of sialylated structures from the blood meal through the gut to the salivary glands using electron microscopy. Specific localization of sialylated glycans to basement membranes of salivary glands was observed. Finally, the influence of the sample preparation methods for electron microscopy on ultrastructure and immunogold labeling was evaluated.     

A symbiont of the tick Ixodes ricinus invades and consumes mitochondria in a mode similar to that of the parasitic bacterium Bdellovibrio bacteriovorus

We have recently performed molecular characterisation of an intracellular alpha-proteobacterium, named IricES1, which resides in the ovarian tissue of female Ixodes ricinus ticks from Italy. A unique characteristic of this bacterium is its ability to invade the mitochondria of the cells in which it resides. Although some ultrastructural studies have been performed on close relatives of this bacterium from I. ricinus in England and Switzerland, a number of questions remain about its movement within ovarian tissues and mitochondria. We have performed the first detailed ultrastructural examination of IricES1 in engorged female adult I. ricinus. Among our findings was that the bacterium enters mitochondria in a similar way to that employed by the 'predatory' bacterium Bdellovibro bacteriovorus, that is, between the inner and outer membranes. It then appears to multiply, with the new 'colony' consuming the mitochondrial matrix. Despite having many of their mitochondria consumed, oocytes appear to develop normally, and the bacteria are likely to be vertically transferred to all eggs.     

Phenology of the tick, Ixodes ricinus, in its southern distribution range (central Spain)

The abundance, seasonal activity patterns and development rates of the tick Ixodes ricinus (L.) (Acari: Ixodidae), as well as microclimate features of the site of study, are described for a 9-year-long study (1994-2002) in north-central Spain. According to drag captures, larvae had a unimodal activity pattern, with a maximum observed around July-August, whereas nymphs displayed a bimodal pattern (May-June and August-September) with strong dominance of spring activity. An inversion of this pattern, with larger autumn peak, was observed in years with humid summers. Adults showed a small spring peak and a large autumn one. In the later years of the study, a small increase in the adult spring peak of activity was noticed, correlated with mild winters. Over the entire period of study, a clear increase in the total tick abundance was detected. Statistically significant differences between years were observed for some climate variables (saturation deficit, winter temperatures and number of days with temperatures above 6 degrees C), but a consistent and constant pattern of change was not observed in any climate variable. Temperature requirements for developing stages showed a sharp decrease in weeks 35-51 (eggs) and 38-50 (larvae and nymphs), a feature attributed to the presence of the morphogenetic diapause, beginning around September. Development rates obtained under quasi-natural conditions were almost twice those reported for other sites, suggesting an adaptation of this local, largely isolated I. ricinus population. According to drag captures and field-obtained development rates, interchange of nymphs between the two cohorts is common in this site, and seems to be influenced by the winter temperature and the date of larval engorgement.     

Pasture improvement and the control of sheep tick,Ixodes ricinus L

The sheep tick is confined almost entirely to rough, poor hill and moorland pastures. Agricultural improvement of these pastures involves (among other things) reduction in the depth of the vegetation layer and partial or complete destruction of the moisture-holding underlying 'mat' of decaying vegetable matter. Assessment of the tick populations on improved and on comparable control areas showed a reduction in tick population following upon pasture improvement. But a fairly considerable degree of improvement is required to bring about a satisfactory reduction. Ticks were not completely extirpated even by ploughing. It is probable that pasture improvement affects tick population mainly by reducing cover and thus the humidity.
Unfortunately, effective pasture improvement is not economic on hill and moorland grazings in general. Such improvement as is practicable could only be a supplement to other control measures.
Since the sheep flock supports almost all the female ticks on a pasture, control is most likely to follow from the development of an efficient acaricidal sheep dip.     

Vector competence of the tick Ixodes ricinus for transmission of Bartonella birtlesii

Bartonella spp. are facultative intracellular vector-borne bacteria associated with several emerging diseases in humans and animals all over the world. The potential for involvement of ticks in transmission of Bartonella spp. has been heartily debated for many years. However, most of the data supporting bartonellae transmission by ticks come from molecular and serological epidemiological surveys in humans and animals providing only indirect evidences without a direct proof of tick vector competence for transmission of bartonellae. We used a murine model to assess the vector competence of Ixodes ricinus for Bartonella birtlesii. Larval and nymphal I. ricinus were fed on a B. birtlesii-infected mouse. The nymphs successfully transmitted B. birtlesii to naïve mice as bacteria were recovered from both the mouse blood and liver at seven and 16 days after tick bites. The female adults successfully emitted the bacteria into uninfected blood after three or more days of tick attachment, when fed via membrane feeding system. Histochemical staining showed the presence of bacteria in salivary glands and muscle tissues of partially engorged adult ticks, which had molted from the infected nymphs. These results confirm the vector competence of I. ricinus for B. birtlesii and represent the first in vivo demonstration of a Bartonella sp. transmission by ticks. Consequently, bartonelloses should be now included in the differential diagnosis for patients exposed to tick bites.     

IrAE: an asparaginyl endopeptidase (legumain) in the gut of the hard tick Ixodes ricinus

Ticks are ectoparasitic blood-feeders and important vectors for pathogens including arboviruses, rickettsiae, spirochetes and protozoa. As obligate blood-feeders, one possible strategy to retard disease transmission is disruption of the parasite's ability to digest host proteins. However, the constituent peptidases in the parasite gut and their potential interplay in the digestion of the blood meal are poorly understood. We have characterised a novel asparaginyl endopeptidase (legumain) from the hard tick Ixodes ricinus (termed IrAE), which we believe is the first such characterisation of a clan CD family C13 cysteine peptidase (protease) in arthropods. By RT-PCR of different tissues, IrAE mRNA was only expressed in the tick gut. Indirect immunofluorescence and EM localised IrAE in the digestive vesicles of gut cells and within the peritrophic matrix. IrAE was functionally expressed in Pichia pastoris and reacted with a specific peptidyl fluorogenic substrate, and acyloxymethyl ketone and aza-asparagine Michael acceptor inhibitors. IrAE activity was unstable at pH > or = 6.0 and was shown to have a strict specificity for asparagine at P1 using a positional scanning synthetic combinatorial library. The enzyme hydrolyzed protein substrates with a pH optimum of 4.5, consistent with the pH of gut cell digestive vesicles. Thus, IrAE cleaved the major protein of the blood meal, hemoglobin, to a predominant peptide of 4kDa. Also, IrAE trans-processed and activated the zymogen form of Schistosoma mansoni cathepsin B1 -- an enzyme contributing to hemoglobin digestion in the gut of that bloodfluke. The possible functions of IrAE in the gut digestive processes of I. ricinus are compared with those suggested for other hematophagous parasites.     

Experience in the study of the peculiarities of the distribution of the tick Ixodes ricinus in a large territory]

Attempts were undertaken to study the character of the distribution of I. ricinus over a large territory. Methodic principles of the land survey of arthropods distribution and laboratory processing of its results, that were used before for I. persulcatus, were found to be quite suitable for I. ricinus. It was established that hungry adults of I. ricinus have at least three types of the distribution throughout the forests of Lithuania. Types of the distribution can be ascertained by the results of the record of I. ricinus nymphs with a flag.     

A comparison of Chryseobacterium indologenes pathogenicity to the soft tick Ornithodoros moubata and hard tick Ixodes ricinus

A yellow-pigmented Gram-negative bacterium, Chryseobacterium indologenes, was found in the gut contents of about 65% of soft ticks Ornithodoros moubata from a perishing laboratory colony. The isolated putative pathogen, C. indologenes, was susceptible to cotrimoxazol and addition of this antibiotic (Biseptol 480) to the blood meal significantly decreased the tick mortality rate. The artificial infection of healthy O. moubata by membrane feeding on blood contaminated with C. indologenes was lethal to all ticks at concentrations 10(6) bacteria/ml. On the contrary, a similar infection dose applied to the hard tick Ixodes ricinus by capillary feeding did not cause significant mortality. Examination of guts dissected from infected O. moubata and I. ricinus revealed that C. indologenes was exponentially multiplied in the soft tick but were completely cleared from the gut of the hard ticks within 1 day. In both tick species, C. indologenes were found to penetrate from the gut into the hemocoel. The phagocytic activity of hemocytes from both tick species was tested by intrahaemocoelic microinjection of C. indologenes and evaluated by indirect fluorescent microscopy using antibodies raised against whole bacteria. Hemocytes from both tick species displayed significant phagocytic activity against C. indologenes. All O. moubata injected with C. indologenes died within 3 days, whereas the increase of the mortality rate of I. ricinus was insignificant. Our results indicate that hard ticks possess much more efficient defense system against infection with C. indologenes than the soft ticks. Thus, C. indologenes infection has the potential to be a relevant comparative model for the study of tick immune reactions to transmitted pathogens.     

A family of putative metalloproteases in the salivary glands of the tick Ixodes ricinus

Ticks are obligate blood-feeding arachnids. During their long-lasting blood meal, they have to counteract the protective barriers and defense mechanisms of their host. These include tissue integrity, pain, hemostasis, and the inflammatory and immune reactions. Here, we describe a multigene family coding for five putative salivary metalloproteases induced during the blood meal of Ixodes ricinus. The evolutionary divergence inside the family was driven by positive Darwinian selection. This came together with individual variation of expression, functional heterogeneity, and antigenic diversification. Inhibition of the expression of some of these genes by RNA interference prevented completion of the tick blood meal and affected the ability of the tick saliva to interfere with host fibrinolysis. This family of proteins could therefore participate in the inhibition of wound healing after the tick bite, thereby facilitating the completion of the blood meal.     

Antigens and glycoproteins of larvae, nymphs and adults of the tick Ixodes ricinus

Abstract Protein components of homogenates of unfed larvae and nymphs of Ixodes ricinus (L.), and of ovary, haemolymph, Malpighian tubules, rectal ampulla, fat body, integument, salivary glands and midgut of partially fed adult females were studied for their antigenicity and carbohydrate moieties using immunoblotting and lectin affinity blotting (LAB) techniques. Comparing the individual anti-larval, anti-nymphal and anti-adult immune sera for their capacity to recognize the specific and trans-stadially cross-reactive antigenic proteins, larval feeding induced the most effective humoral response. The majority of immunogens recognized by rabbit anti-tick immune sera are glycoproteins. Most of the glycosylated antigens were modified with N-type glycans; however, O-type glycans were also demonstrated in some antigens. The correlation of the type of glycosylation with antigenicity, and the sharing of common antigenic epitopes by various tissues, are discussed.     

Risk indicators for the tick Ixodes ricinus and Borrelia burgdorferi sensu lato in Sweden

The distributional area of the tick Ixodes ricinus (L.), the primary European vector to humans of Lyme borreliosis spirochaetes (Borrelia burgdorferi sensu lato) and tick‐borne encephalitis virus, appears to be increasing in Sweden. It is therefore important to determine which environmental factors are most useful to assess risk of human exposure to this tick and its associated pathogens. The geographical distribution of I. ricinus in Sweden was analysed with respect to vegetation zones and climate. The northern limit of I. ricinus and B. burgdorferi s.l. in Sweden corresponds roughly to the northern limit of the southern boreal vegetation zone, and is characterized climatically by snow cover for a mean duration of 150 days and a vegetation period averaging 170 days. The zoogeographical distribution of I. ricinus in Sweden can be classified as southerly–central, with the centre of the distribution south of the Limes Norrlandicus. Ixodes ricinus nymphs from 13 localities in different parts of Sweden were examined for the presence of B. burgdorferi s.l. and found to be infected with Borrelia afzelii and Borrelia garinii. Tick sampling localities were characterized on the basis of the density of Borrelia‐infected I. ricinus nymphs, presence of specific mammals, dominant vegetation and climate. Densities of I. ricinus nymphs and Borrelia‐infected nymphs were significantly correlated, and nymphal density can thus serve as a general indicator of risk for exposure to Lyme borreliosis spirochaetes. Analysis of data from this and other studies suggests that high densities of Borrelia‐infected nymphs typically occur in coastal, broadleaf vegetation and in mixed deciduous/spruce vegetation in southern Sweden. Ixodes ricinus populations consistently infected with B. burgdorferi s.l. can occur in: (a) biotopes with shrews, rodents, hares and birds; (b) biotopes with shrews, rodents, hares, deer and birds, and (c) island locations where the varying hare (Lepus timidus) is the only mammalian tick host.     

Anti-hemostatic effects of a serpin from the saliva of the tick Ixodes ricinus

Serpins (serine protease inhibitors) are a large family of structurally related proteins found in a wide variety of organisms, including hematophagous arthropods. Protein analyses revealed that Iris, previously described as an immunomodulator secreted in the tick saliva, is related to the leukocyte elastase inhibitor and possesses serpin motifs, including the reactive center loop (RCL), which is involved in the interaction between serpins and serine proteases. Only serine proteases were inhibited by purified recombinant Iris (rIris), whereas mutants L339A and A332P were found devoid of any protease inhibitory activity. The highest Ka was observed with human leukocyte-elastase, suggesting that elastase-like proteases are the natural targets of Iris. In addition, mutation M340R completely changed both Iris substrate specificity and affinity. This likely identified Met-340 as amino acid P1 in the RCL. The effects of rIris and its mutants were also tested on primary hemostasis, blood clotting, and fibrinolysis. rIris increased platelet adhesion, the contact phase-activated pathway of coagulation, and fibrinolysis times in a dose-dependent manner, whereas rIris mutant L339A affected only platelet adhesion. Taken together, these results indicate that Iris disrupts coagulation and fibrinolysis via the anti-proteolytic RCL domain. One or more other domains could be responsible for primary hemostasis inhibition. To our knowledge, this is the first ectoparasite serpin that interferes with both hemostasis and the immune response.