Natural infection of the tick Dermacentor reticulatus (Fabr.) with the microsporidian Nosema slovaca Weiser et Rehácek in Slovakia

A total of 6,199 adult ticks (Haemaphysalis intermis, Dermacentor reticulatus, D. marginatus and Ixodes ricinus) collected in three localities in Slovakia were examined for microsporidian infection. The spores of Nosema slovaca were found in six D. reticulatus (0.097%). The microsporidian experimentally inoculated into the hemocele of half-engorged females of D. reticulatus caused acute infection and death of the host. The infection can be transmitted also to other tick species from the same region. The yield of spores from one tick ranges from 3 X 10(5) to 52 X 10(7) depending on the infection dose and tick species. Peroral infection of some butterfly larvae with the microsporidian pores was unsuccessful. The importance of Nosema infection in ticks in nature is discussed.     

Prevalence of borreliae in Ixodes ricinus ticks from southern Moravia, Czechoslovakia

A total of 378 adult Ixodes ricinus ticks were collected by flagging vegetation in four localities of two districts (Breclav, Znojmo) in south Moravia and examined microscopically. Borreliae were identified in Giemsa-stained midgut smears from 32 (i.e. 8.5%) ticks (9.4% females, 7.2% males); the infection rate varied between 0.0 and 11.4% in the four localities examined. Among female ticks, significantly more were found to be infected in autumn (19.7%) than in spring (5.8%). Dark-field (DF) and Giemsa-stained smears (GS) examinations were compared for their sensitivity in detecting borreliae in 128 field-collected ticks; GS method showed a little higher sensitivity (11.7% ticks were positive) than DF procedure (9.4% ticks positive). Two strains of Borrelia burgdorferi were isolated from a total of 150 adult I. ricinus ticks cultured in BSK medium.     

Bartonella infections in fleas (Siphonaptera: Pulicidae) and lack of bartonellae in ticks (Acari: Ixodidae) from Hungary

Fleas (95 Pulex irritans, 50 Ctenocephalides felis, 45 Ctenocephalides canis) and ixodid ticks (223 ixodes ricinus, 231 Dermacentor reticulatus, 204 Haemaphysalis concinna) were collected in Hungary and tested, in assays based on PCR, for Bartonella infection. Low percentages of P. irritans (4.2%) and C. felis (4.0%) were found to be infected. The groEL sequences of the four isolates from P. irritans were different from all the homologous sequences for bartonellae previously stored in GenBank but closest to those of Bartonella sp. SE-Bart-B (sharing 96% identities). The groEL sequences of the two isolates from C. felis were identical with those of the causative agents of cat scratch disease, Bartonella henselae and Bartonella clarridgeiae, respectively. The pap31 sequences of B. henselae amplified from Hungarian fleas were identical with that of Marseille strain. No Bartonella-specific amplification products were detected in C. canis, I. ricinus, D. reticulatus and H. concinna pools.     

Rickettsiae of the spotted fever group in Hungary

Rickettsiae were detected by haemocyte test in 7.2% of Dermacentor marginatus and 4.7% of D. reticulatus ticks collected in Hungary. Six strains of rickettsiae were isolated from positive specimens. These rickettsiae according to CFR are closely related, probably identical with R. slovaca and they represent the first rickettsiae of the Spotted Fever group found in that country.     

Comparison of virulence of Coxiella burnetii isolates from bovine milk and from ticks.

Laboratory animals (mice and guinea pigs) were infected with the isolates of Coxiella burnetii (Derrick, 1939) obtained from bovine milk (M18 and M35) and the ticks Ixodes ricinus (Linnaeus, 1758) and Dermacentor marginatus (Sulzer, 1776) (Kl3 and Kl6, respectively), and with the reference strain Nine Mile. Neither mortality nor lethality occurred with the mice. Antibody response in mice infected with isolates from milk was lower (1:16-512) than that from ticks (1:32-4096). Onset of seropositivity also occurred later - on the 10th day post-infection (p.i.) for M18 and M35 in comparison with the 7th day for Kl3 and Kl6. In guinea pigs, infection manifested by fever. The fever was less evident in guinea pigs infected with isolates from milk (39.5-40.1 degrees C) than in guinea pigs infected with isolates from ticks (39.5-40.6 degrees C). Partially engorged females of Dermacentor reticulatus (Fabricius, 1794) were inoculated with isolates M18 and Kl3. No differences in the multiplication of C. burnetii in haemocytes between these two isolates were ascertained.     

Comparison of the protein profiles of salivary gland extracts derived from three species of unfed and partially fed ixodid ticks analysed by SDS-PAGE

Salivary gland extracts (SGE) from unfed and 5 days fed adult female Ixodes ricinus (Linnaeus, 1758); Haemaphysalis inermis (Birula, 1895) and Dermacentor reticulatus (Fabricius, 1794) ticks were prepared. The protein content after feeding increased by 10.6, 8.7 and 6.8 times, respectively. Extracts were equilibrated to the same protein content and submitted to SDS-polyacrylamide gel electrophoresis followed by computer analysis of the scanned gels. Relative differences in protein profiles of extracts obtained from unfed and partially fed ticks were found in all species and some of them were similar in all three species used in the study. Results demonstrate that the increase of the protein content in salivary glands during the feeding does not occur proportionally. Some proteins are synthesised preferentially (67.1 kDa, 13.5 kDa) but other bands (in range of 15-16 kDa) present in the SGE derived from unfed ticks are less discernible in that of fed ticks.     

The prevalence of Borrelia burgdorferi sensu lato in Ixodes persulcatus and I. ricinus ticks in the zone of their sympatry

A total of 7210 unfed adult Ixodes persulcatus Schulze, 1930 and I. ricinus (L., 1758) ticks were collected from the vegetation by flagging in 35 study sites located in the zone of their sympatry (mainly in Leningrad region, Russia). Borrelia infection in ticks was estimated by the dark-field microscopic analysis of gut contents in standard vital preparations at a magnification of x600. No correlation was revealed between the series of parameters characterising the abundance of each tick species (tau = -0.13) and between the series of these parameters and the prevalence of Borrelia in each vector. It is concluded that in the broad zone of I. persulcatus and I. ricinus sympatry, the presence and proportion of one vector in the ecosystem does not have any significant effect on the extensity of infection and on the epizootic and epidemic significance of the other vector. Each tick species has its independent (of the other species) and relatively original functional role in the focal ecosystem.     

Peculiarities of behaviour of taiga (Ixodes persulcatus) and sheep (Ixodes ricinus) ticks (Acarina: Ixodidae) determined by different methods

A comparison of the behavioural peculiarities of Ixodes persulcatus Schulze, 1930 (north-western population, Russia) and Ixodes ricinus (L., 1758) from western Russia and Denmark was determined by using two methods. Method 1 involved a sojourn of ticks on vertical plastic sticks and showed that the questing behaviour of I. ricinus nymphs was dependent on temperature and relative humidity (RH). A significantly greater number of nymphs quested at 22 degrees C and 100% RH than at 18 degrees C. When the humidity was reduced to 30% all of the nymphs departed. In the second method, the activity of ticks on an inclined "ticksdrome" was estimated. The activity of I. ricinus adults from the Danish population was 1.2 times greater than that of ticks from Russia. Females of the species studied and specimens from all study areas were more active than all other stages of development. The locomotor activity of both adult and immature I. ricinus that were infected with Borrelia burgdorferi sensu lato was suppressed when compared with uninfected specimens. The locomotor activity of I. persulcatus females infected by borreliae with exoskeleton anomalies was 1.3 times greater (P<0.05) than that of infected ticks without anomalies. Our data showed that infected females with exoskeleton anomalies could crawl faster on a human and reach uncovered parts of the body that are vulnerable for attachment and feeding. A study of locomotor activity and questing behaviour may be useful for comparing the risk for different tick species and populations to transmit tick-borne pathogens.     

Modulation of human lymphocyte proliferation by salivary gland extracts of ixodid ticks (Acari: Ixodidae): effect of feeding stage and sex

Ixodid ticks remain attached to their hosts for several days to weeks. During this extended feeding process new proteins involved in the modulation of host immune responses are expressed in tick salivary glands. In our study a stimulatory or inhibitory effect of salivary gland extracts (SGE) of unfed and partially fed female Ixodes ricinus (Linnaeus, 1758), female and male Amblyomma variegatum (Fabricius, 1794) and Rhipicephalus appendiculatus Neumann, 1901 ticks on human lymphocyte proliferation induced by Concanavalin A (ConA) and phytohaemagglutinin (PHA), respectively, was investigated. SGE of all female ticks examined suppressed proliferation of ConA-induced lymphocytes; highly significant suppression was observed in the presence of unfed I. ricinus and 9-day fed A. variegatum SGE. SGE of partially fed A. variegatum and I. ricinus females suppressed PHA responses of lymphocytes. Lymphocytes showed reduced PHA and ConA responses in the presence of SGE of unfed and 2-day fed R. appendiculatus females, while SGE of 6-day fed females enhanced PHA responses, but reduced their ConA responses; generally SGE of 2-day fed females displayed the strongest inhibition. Amblyomma variegatum male SGE slightly enhanced PHA, but significantly reduced ConA responses of lymphocytes and their inhibitory effect increased during feeding. SGE of unfed and 2-day fed R. appendiculatus males enhanced PHA and ConA responses and those of 6-day fed males suppressed lymphocyte proliferation. The results suggest that (i) species- and sex-specific differences exist in the effects of tick salivary gland antigens on human lymphocyte proliferation and (ii) effect of SGE on human lymphocyte responses to mitogens varies depending on the tick feeding status.     

First record of bacillary rickettsia-like organisms in European tick Dermacentor marginatus (Sulzer).

Bacillary rickettsia-like organisms (BRLO) were found in the tick Dermacentor marginatus. They are gram-negative and differ from common bacteria and reckettsiae both in the cultivation conditions and morphology. BRLO are non-pathogenic for ticks and guinea pigs. In our studies they were isolated on half-engorged females of D. marginatus, on which they are still maintained.     

Effect of fast protein liquid chromatography fractionated salivary gland extracts from different ixodid tick species on interleukin-8 binding to its cell receptors

Interleukin-8 plays a critical role in inflammatory processes. Hence generation of molecules with anti-IL-8 activity is likely to be important for successful feeding and for survival of the ticks. Anti-IL-8 activity was studied in saliva of three ixodid tick species--Dermacentor reticulatus (Fabricius, 1794), Rhipicephalus appendiculatus Neumann, 1901, and Amblyomma variegatum (Fabricius, 1794). The greatest activity was shown in saliva prepared from D. reticulatus. The activity was attributed to tick salivary gland molecules that bind to IL-8, preventing binding of the chemokine to its specific receptor, rather than to occupation of the IL-8 cell receptor by the tick molecules. The distribution of anti-IL-8 activity in fast protein liquid chromatography (FPLC) fractions of salivary gland extracts (SGE) derived from adult female D. reticulatus, R. appendiculatus and A. variegatum was compared directly by both ELISA and receptor-binding inhibition assays. The correspondence in results with fractions of SGE from ELISA is consistent with detection of tick molecules that inhibit IL-8 binding to its receptor. As IL-8 is an important chemoattractant and activator of neutrophils, the presence of an anti-IL-8 activity in tick saliva indicates that neutrophils play an important role in the host response to parasitism by ticks.     

Influence of microclimate on the life cycle of the common tick Ixodes ricinus (L.) in thermophilic oak forest.

Under conditions of the South Moravian thermophilic oak forest (Valtice near Breclav), the life cycle of the common tick Ixodes ricinus was studied by continuous recording of main elements of microclimate (temperature and humidity) in three different biotopes: forest, margin of the forest and meadow. Simultaneously conditions and the process of tick hibernation were studied in four soil layers (surface, depths of 10, 20 and 30 cm). Observations made in the winter and vegetation periods were assessed by mathematical-statistical tests. Results obtained in the forest biotope and at its margin are presented in this paper; results from the meadow biotope will be published separately.     

Occurrence of ixodid ticks--the main vectors of tick-borne encephalitis virus in urbanized territory

The influence of urbanization on ticks I. ricinus and I. persulcatus--the main vectors of tick-borne encephalitis virus is discussed on the basis of concrete examples. It has been revealed that under favourable conditions the populations of these tick species can exist for a long time not only in towns and new housing estates, but in the old residential districts as well. The necessity of studying the ecology of urban populations of ixodid ticks is pointed out.     

Tick salivary gland extract-activated transmission of Borrelia afzelii spirochaetes

Saliva-activated transmission of Borrelia afzelii Canica, Nato, du Merle, Mazie, Baranton et Postic, 1993 was demonstrated using salivary gland extract (SGE) from Ixodes ricinus (L., 1758) ticks and C3H mice. Injection of Borrelia spirochaetes together with SGE increased the level of bacteraemia and accelerated the appearance of bacteria in the urinary bladder, compared with the injection of spirochaetes alone. More I. ricinus nymphs became infected when feeding on mice inoculated with B. afzelii plus SGE. Analysis of cytokines produced by cells of draining lymph nodes from SGE-treated mice showed a suppression of proinflammatory cytokines IFN-gamma, IL-6 and GM-CSF following a transient upregulation in comparison with the control mice infected without SGE.     

Effect of salivary gland extract from Ixodes ricinus ticks on the proliferation of Borrelia burgdorferi sensu stricto in vivo

Saliva-activated transmission (SAT) of Borrelia burgdorferi sensu stricto was demonstrated using real-time PCR and salivary gland extract (SGE) from partially fed Ixodes ricinus ticks. C3H/HeN mice were injected intradermally with 1.5 x 10(3) spirochetes mixed with 40 microg of SGE per mouse. The control group was inoculated with the same dose of spirochetes without SGE. The accelerating effect of SGE on spirochete proliferation was demonstrated on day 1 post infection, when a 4.2-fold increase in spirochetes was found in the skin and a 10-fold increase in the blood, compared with control mice. The data represent the first direct evidence of a SAT effect of I. ricinus SGE on infection with the Lyme disease agent B. burgdorferi.     

Lyme borreliosis: insights into tick-/host-borrelia relations

Lyme borreliosis (LB) is a serious infectious disease of humans and some domestic animals in temperate regions of the Northern Hemisphere. It is caused by certain spirochetes in the Borrelia burgdorferi sensu lato (s.l.) species complex. The complex consists of 11 species (genospecies). Borrelia burgdorferi sensu stricto (s.s.), Borrelia garinii and Borrelia afzelii are the major agents of human disease. Borrelia burgdorferi s.l. species are transmitted mainly by ticks belonging to the Ixodes ricinus species complex plus a few additional species not currently assigned to the complex. B. burgdorferi infections may produce an acute or chronic disease with a wide array of clinical symptoms such as erythema migrans (EM), carditis, arthritis, neuroborreliosis, and acrodermatitis chronica atrophicans (ACA). Differences in LB spirochetes 'genospecies' and strains/isolates determine the occurrence and severity of this multi-system disease. Accurate and reliable identification of the LB spirochetes in ticks as well as knowledge of their prevalence are essential for prevention against the disease and development of an effective vaccine. An overview of the knowledge of molecular factors with emphasis on potential protein-carbohydrate interactions in the tick-borrelia system is the main focus of this review.     

Babesia microti (Piroplasmida: Babesiidae) in nymphal Ixodes ricinus (Acari: Ixodidae) in the Czech Republic

A total of 350 nymphs of the common tick Ixodes ricinus (Linnaeus, 1758) were collected in an endemic focus of Lyme borreliosis (South Moravia, Czech Republic) and examined for the presence of the protozoan Babesia microti (Fran?a, 1909) by polymerase chain reaction (PCR), using primers specific for the B. microti gene encoding small subunit rRNA. The assay revealed five positive pools (out of 70 pools examined); the corresponding prevalence rate was about 1.5%. Sequence analysis of the PCR products confirmed their 100% homology with that of B. microti. The study represents the first evidence of B. microti in ixodid ticks in the Czech Republic.     

Ixodid ticks of Barun Glacier region (the Nepal Himalaya)

During the 3rd Czechoslovak mountaineering expedition to the Himalaya in the spring 1973 ticks of the family Ixodidae were collected from small mammals and additionally from pastured sheep, and humans. The tick material represented the following species: Ixodes (P.) ovatus Neumann, 1899, Ixodes (I.) acutitarsus (Karsch, 1880) and Haemaphysalis (A.) aponommoides Warburton, 1921. Ecological and zoogeographical characteristics of the ticks found are given and their potential medical importance is discussed.     

Detection of Anaplasma dna in Ixodes ricinus ticks: pitfalls

A total of 150 nymphal Ixodes ricinus (L., 1758) (Acari: Ixodidae) from the Czech Republic were examined for Anaplasma phagocytophilum (Foggie, 1949) Dumler et al., 2001 by PCR using EHR521/747 primers: 22 of 50 pools were positive (minimum prevalence, 14.7%). However, sequencing of the PCR products did not show complete homology with A. phagocytophilum (91%) while the closest relationship (95%) was found to "Candidatus Ehrlichia walkefii". The results indicate a need for care in interpretation of Anaplasma PCR results and for PCR optimization for detecting A. phagocytophilum in ticks.     

Isolation of Bhanja virus from ticks of the genus Haemaphysalis in southeast Bulgaria and presence of antibodies in pastured sheep

Bhanja virus was isolated from ticks of the genus Haemaphysalis (H. punctata and H. sulcata) collected from sheep pastured in the surroundings of Akhtopol in southeast Bulgaria, and simultaneously blood sera of 58 sheep were investigated in plaque-reduction neutralization test with B lranja arbovirus. All sera contained antibodies to this virus in titres from 1:32 to 1:4096 (geometrical mean titre 1:494). The district of Akhtopol proved to be a natural focus of Bhanja virus.