Expression of outer surface proteins A and C of Borrelia burgdorferi in Ixodes ricinus ticks removed from humans

A total of 131 Ixodes ricinus (51 females, 1 male and 79 nymphs) removed from persons living in Southern Germany were investigated by immunofluorescence assay for the presence of Borrelia burgdorferi with a polyvalent rabbit immune serum and monoclonal antibodies specific for outer surface proteins (Osp) A or C. Borreliae were detectable in 48 (36.6%) of the ticks. Infection rates of these adults and nymphs were significantly higher than infection rates of unfed ticks from Southern Germany. Borreliae in 31.3% (n = 15) of the infected ticks expressed solely OspA, solely OspC in 12.5% (n = 6), and both OspA and OspC in 39.6% (n = 19) of ticks, while in 16.7% (n = 8) of ticks neither were expressed. Presentation of OspC by B. burgdorferi in I. ricinus was correlated with tick weight: in females, OspC was detectable only in ticks with a minimum weight of about 3.5 mg, and in nymphs weighing at least 1 mg. These results indicate that in I. ricinus removed from humans OspC is up-regulated during the blood meal of the tick, but in most ticks OspA is still detectable and might even be present in the absence of OspC expression in the midgut and salivary glands of nearly fully engorged nymphal ticks. Furthermore, we found strong evidence that borreliae expressing solely OspA while in the salivary glands can cause Lyme borreliosis. Our findings indicate that during tick feeding, humans are exposed to borreliae that may express either OspA or OspC or both, or lack both OspA and C. These findings suggests that, at the minimum, both OspA and C should be considered as vaccine candidates for prophylaxis of Lyme borreliosis in Europe. Received: 28 July 1998     

Influence of saturation deficit and temperature on Ixodes ricinus tick questing activity in a Lyme borreliosis-endemic area (Switzerland)

Questing Ixodes ricinus ticks were sampled monthly in a periurban mixed forest at Neuchatel, Switzerland, in 1996, 1997, and 1998. A total of 5530 I. ricinus nymphs and 1385 adults were collected. The emergence of questing tick populations in spring varied among years in relation to the air temperature. The monthly variation in questing nymph density was negatively correlated with the saturation deficit; there was a strong decrease in tick numbers in late spring and summer as soon as the saturation deficit increased. A variation in tick density in relation to the saturation deficit was also observed between years; the tick density was high when the saturation deficit was low during spring and summer (1997) and was low when the saturation deficit was high (1998). During the 3-year study period, marked climatic differences among years highlighted the influence of temperature and saturation deficit on the phenology of ticks. Received: 17 January 2000 / Accepted: 27 January 2000     

Stage-associated risk of transmission of the lyme disease spirochete by EuropeanIxodes ticks

To more closely define the risk of infection by the agent of Lyme disease in Europe, we determined whether spirochetal prevalence increases throughout the development of theIxodes ricinus vector tick. Of all ticks that could be flagged from vegetation,I. ricinus were by far the most abundant. Spirochetal infection rates in the adult stage of this tick (15%) are no higher than those in nymphs (18%) but greatly exceed those in larvae (0.7%). This tick therefore appears to attain infection mainly from the host of its larval stage, generally feeds on hosts that are noncompetent as reservoirs in its nymphal stage, and rarely inherits infection. Risk of human infection mainly derives from contact with the nymphal stage of the vector tick because the larva is rarely infected and the adult is large enough to be noticed and promptly removed.This work was supported in part by grant AI 19693 from the National Institute of Allergy and Infectious Disease and by grants from the city of Berlin     

Uukuniemi group viruses isolated in Norway

Summary As part of an ecological approach to the question of arboviruses in Norway, ticks(Ixodes ricinus, I. trianguliceps andI. uriae), small mammals and passerine birds were collected from selected localities along the coastal parts of the country in 1973. Some tick collections also were received from ornithological stations. Three Uukuniemi (UUK) virus strains were isolated from the tick material. The ticks yielding viruses originated from three very different biocoenoses. One isolate was made from engorgedI. ricinus nymphs picked up from migrant birds in the spring (SF E1). Another isolate (By E50) was made from unengorgedI. ricinus nymphs collected within an earlier established tick-borne encephalitis (TBE) virus focus. These two strains seemed serologically identical, and also identical to the Finnish prototype strain S23. The third isolate (Ru E82) was made from unengorgedI. uriae collected in a Common Puffin (Fratercula arctica) colony. Although clearly belonging to the UUK group of viruses, this strain demonstrated biological as well as antigenic differences to the other isolates. Some characteristics of the Norwegian UUK viruses are discussed in this paper. An immunoelectroosmophoresis method was used to screen human, bovine, small mammal and passerine bird sera for antibodies against the UUK isolate By E50. The results indicated that UUK virus (es) are wide-spread in the Norwegian coastal areas invaded byI. ricinus. The presence of mixed virus foci of UUK and TBE were indicated.With 7 Figures     

Winter activity of Ixodes ricinus in a Berlin forest

Nymphs and adults of the hard tick Ixodes ricinus typically quest in Central Europe from March/April to October/early November. The recent trend towards milder winters raises the question whether I. ricinus will gradually extend its seasonal activity and use mild winter periods for questing. The present study followed the host-seeking activity of I. ricinus from early September 2006 to early March 2007 in a Berlin forest. Field-collected larval and nymphal ticks were allowed to take a full blood meal in the laboratory and to moult. The resultant unfed nymphs (n=125) and adults (n=149) were released onto three field plots (1×1 m²) in early September 2006. A second group of field-collected unfed I. ricinus nymphs (n=79) and adults (n=47) were released onto a field plot in early September 2006. Each plot was covered by an approximately 10 cm thick layer of leaf litter (mostly of oak and beech) to provide a humid microclimate refuge and shelter for non-active ticks. Wooden rods (60 cm high) arranged in a 6×8 grid on these field plots were climbed by questing nymphs and adults. Active ticks were observed on each of 33 observation dates. The winter activity revealed by this experiment was confirmed by flagging for ticks on two occasions in January and February 2007 with remarkably high numbers of collected ticks. This appears to be the first time that extended or even continuous winter activity of I. ricinus nymphs and adults has been demonstrated in Central Europe, most probably caused by the extremely mild winter of 2006/2007. The fact that I. ricinus now can be active during the whole winter, a time of the year when these ticks historically have been dormant when the weather is normal, is relevant to the public because people who enter forest areas should be well aware of the risk of getting tick bites and tick-borne infections in mild winter periods.     

Prevalence of tick‐borne encephalitis virus in Ixodes ricinus ticks from three islands in north‐western Norway

Tick‐borne encephalitis (TBE) is the most important viral tick‐borne disease in Europe and can cause severe disease in humans. In Norway, human cases have been reported only from the southern coast. The aim of this study was to investigate the prevalence of tick‐borne encephalitis virus (TBEV) in questing Ixodes ricinus ticks from the north‐western part of Norway. A total of 4509 ticks were collected by flagging in May and June 2014. A subpopulation of 2220 nymphs and 162 adult ticks were analysed by real‐time PCR and positive samples were confirmed by pyrosequencing. The estimated prevalence of TBEV was 3.08% among adult ticks from Sekken in Møre og Romsdal County and 0.41% among nymphs from both Hitra and Frøya in Sør‐Trøndelag County. This study indicates that TBEV might be more widespread than the distribution of reported human cases suggests.     

Ticks biting humans in the urban area of Istanbul

A passive surveillance for tick bites in humans was undertaken in the city of Istanbul (Turkey) in the summer and autumn of 2006. From 1,054 reported tick bites, most were females of Ixodes ricinus (27%) and nymphs of Hyalomma aegyptium (50%). A few adults of Hyalomma m. marginatum, Rhipicephalus sanguineus and Dermacentor marginatus were also recorded. We investigated potential risk factors for I. ricinus and H. aegyptium with spatial statistics. Climate features at 1-km resolution (monthly minimum temperatures in late summer and autumn and rainfall) and vegetation features at high resolution (density and heterogeneity of forest-type vegetation as well as distance of reporting site to these vegetation features) are useful variables explaining high reporting clusters for both Ixodes and Hyalomma. While Ixodes is highly reported in dense highly heterogeneous vegetation patches, Hyalomma is commonly found in areas far from forest-type features and in the small, relatively dry vegetation patches within the urban fabric.     

Distribution of Rickettsia helvetica in Ixodes ricinus tick populations in Poland

Questing Ixodes ricinus ticks from different regions of Poland were investigated for the presence of spotted fever group (SFG) rickettsiae. A total of 1214 DNA samples of 2813 ticks, including 820 individual adult ticks and 394 pools containing 1993 nymphs, were tested by PCR for a fragment of the rickettsial gltA gene using the primers RpCs.877 and RpCs.1258. Overall, at least 5.5% ticks were found to be positive with the highest prevalence observed in females (10.6%). A sample of 14 positive PCR products was sequenced. Analyzed fragments of 270–370 bp showed 100% similarities to corresponding sequences of Rickettsia helvetica deposited in the GenBank. Results of our investigation confirm the occurrence and wide distribution of R. helvetica in I. ricinus tick populations in Poland. This rickettsia should be added to the list of potentially dangerous pathogens transmitted by ticks in our country.     

Induction of host resistance toRhipicephalus appendiculatus in rabbits: Effects of immunizing with detergent-solubilized tick tissue proteins

Resistance to the hard tick,Rhipicephalus appendiculatus, was induced in rabbits by immunizing them with tick tissue proteins extracted with a detergent, Triton X-100. There was 25% mortality in female ticks fed on immunized rabbits as compared with those fed on controls. Similarly, there was a 40% and 60% reduction in the engorged weight and the weight of egg batches, respectively, of ticks fed on immunized rabbits. Western blot analysis of detergent-solubilized tick tissue Western blot analysis of detergent-solubilized tick tissue proteins, carried out using immune sera, recognized a complex pattern of proteins. A strong reaction was observed with proteins with apparent molecular weights of 94000 and 40000 daltons.This work was carried out at the Institute of Zoology, University of Neuchâtel, Chantemerle 22, CH-2000 Neuchâtel, Switzerland     

Spatial Lyme borreliosis risk assessment in north-eastern Italy

The risk exposure to Lyme borreliosis in the Friuli Venezia Giulia region, in north-eastern Italy, was investigated on the basis of the abundance of infected ticks. A total of 2620 questing Ixodes ricinus ticks were collected at 25 sampling sites during spring 2005 and 2006, and nymphs and adult females were examined for Borrelia burgdorferi. The distribution of infected ticks was analysed in relation to a set of environmental parameters obtained from satellite, climatic, topographic, and biotic data. A multiple linear regression model was developed to predict the abundance of infected ticks and using GIS facilities, it was possible to elaborate an infected tick distribution map.     

The development ofAmblyomma americanum andDermacentor variabilis (Acari: Ixodidae) fed on rabbits immunized with tick hemolymph

The possibility of interfering with the normal function of tick hemolymph using antihemolymph antibodies taken in with the bloodmeal, was investigated. Cell free hemolymph from repleteAmblyomma americanum andDermacentor variabilis ticks was used to immunize rabbits. Immunized rabbits developed high antihemolymph antibody titers (ca. 10⁵) and had no ill side effects. Rabbits were simultaneously infested with larvae, nymphs, and adult ticks. The biological performance of ticks fed on immunized rabbits was virtually identical to that of ticks fed on nonimmunized rabbits. Usually, the mean engorgement weights of nymphs and females and the weights of the egg masses of both species were slightly higher for ticks fed on the nonimmunized rabbits but differences were not significant (P>0.05) due to a large standard deviation. The possibility of deactivating a single hemolymph component with specific antibodies is discussed.Journal article 4979 of the Agricultural Experiment Station, Oklahoma State University, Stillwater, OK, USA     

Susceptibility of BALB/c mice to nymphs and larvae of Ixodes ricinus after modulation of IgE production with anti-interleukin-4 or anti-interferon-γ monoclonal antibodies

BALB/c mice infested three times with nymphs or larvae of Ixdoes ricinus ticks do not acquire resistance as assessed by evaluation of both tick attachment and the weight of engorged nymphs or larvae. Tick challenge causes a gradual increase in total IgE antibody production from the first to the third infestation. Anti-tick IgG antibodies are never detected. When the mice are treated with anti-interleukin-4 (anti-IL-4) or anti-interferon-gamma (anti-IFN-γ) monoclonal antibodies (mAbs) 1 day before each infestation, they produce fewer or more IgE antibodies, respectively. No effect is observed on IgG antibodies. In IL-4-deficient mice, no IgE or IgG antibody is produced. However, these treatments and the use of IL-4-deficient mice have no negative effect on either tick attachment or the weight of engorged nymphs or larvae. Treatment with anti-IL-4 mAb and the use of IL-4-deficient mice inhibits and abolishes the switching of IgE, respectively, but these are apparently not sufficient to shift the response toward Th1 cells. Received: 18 July 1997 / Accepted: 12 November 1997     

Efficiency of experimental infection of Ixodes ricinus ticks with Borrelia burgdorferi spirochetes

Feeding of an ixodid tick on a Borrelia-infected vertebrate is the natural route of tick infection. To obtain a cohort of nymphs with a high prevalence of infection, the immersion of Ixodes ricinus larvae in the suspension of B. burgdorferi sensu stricto spirochetes was used. Immersed larvae fed on C3H mice until full engorgement, molted to nymphs and 9 of 10 nymphs were shown to be infected by PCR with specific primers. Specific antibodies in the sera of the mice, on which either immersion-infected larvae or infected nymphs had engorged, were not detected. However, nymphs molted from immersed larvae were able to infect naïve mice, and all the resulting 10 adults were infected.     

Prevalence of tick-borne encephalitis virus in Ixodes ricinus ticks in Finland

Approximately 20 cases of tick‐borne encephalitis (TBE) occur annually in Finland. The known endemic areas are situated mainly in the archipelago and coastal regions of Finland, with highest incidence in Åland islands. Ixodes ricinus panels collected in 1996–1997 from two endemic areas were screened for the presence of RNA. Two distinct RT‐PCR methods were applied, and were shown to have an approximate detection limit of 10 focus forming doses (FFD)/100 μl. One out of 20 pools (a total of 139 ticks) from Helsinki Isosaari Island and one out of 48 pools (a total of 450 ticks) from Åland were positive with both methods, whereas the remaining pools were negative. The observed overall frequency (0.34%) in ticks in endemic areas of Finland, was similar to the low incidence found by virus isolation in mice in the 1960s (0.5%). Viral RNA was detectable in a diluted sample representing 0.005% of a positive pool of ten nymphs suggesting that the viral RNA load within an infected tick pool was approximately equivalent to 20,000–200,000 FFD. Sequence analysis did not show geographical clustering of the Finnish strains, suggesting an independent emergence of different TBE foci from the south. TBE virus RNA positive ticks were not found in I. ricinus panels consisting of 130 pools (726 ticks) from Helsinki city parks or 41 pools (197 ticks) from Võrmsi Island in Estonia. J. Med. Virol. 64:21–28, 2001. © 2001 Wiley‐Liss, Inc.     

Cytokines (IL-4 and IFN-γ) and antibodies (IgE and IgG2a) produced in mice infected with Borrelia burgdorferi sensu stricto via nymphs of Ixodes ricinus ticks or syringe inoculations

Mice were tolerant to tick bites during three infestations with nymphs of Ixodes ricinus infected with Borrelia burgdorferi sensu stricto. To determine whether tick bites influence the immune response against B. burgdorferi, we examined the production of cytokines IL-4 and IFN-γ by lymph node cells of BALB/c mice and IL-4 deficient BALB/c mice after tick inoculation versus syringe inoculation of B. burgdorferi. We also measured IgG2a anti-borrelial antibodies and total IgE in these mice. Results showed that BALB/c mice developed a Th2 immune response against B. burgdorferi after tick inoculation and a mixed Th1/Th2 response after syringe inoculation of B. burgdorferi. IL-4 deficient mice produced a Th1 immune response in both cases. IL-4 produced following tick bites greatly decreased the production of anti-borrelial IgG2a antibodies by comparison with the production of anti-borrelial IgG2a antibodies produced following syringe injection of B. burgdorferi. Received: 23 November 1999 / Accepted: 9 December 1999     

Identification of galactose‐α‐1,3‐galactose in the gastrointestinal tract of the tick Ixodes ricinus; possible relationship with red meat allergy

Patients with IgE antibodies against the carbohydrate epitope galactose‐α‐1,3‐galactose (α‐Gal) have reported severe allergic reactions after consumption of red meat. Investigations have revealed associations between IgE to α‐Gal and tick bites. We provide the first direct evidence that α‐Gal is present within ticks thus potentially explaining the relationship between tick exposure and sensitization to α‐Gal, with development of red meat allergy as a secondary phenomena. Serum from Swedish patients with delayed severe reactions to red meat was included in the study. A dose‐dependent inhibition of IgE responses to α‐Gal by the tick Ixodes ricinus is demonstrated. Furthermore, using cryostat‐cut sections of I. ricinus, we show that both a monoclonal and a polyclonal antibody against α‐Gal stains the gastrointestinal tract of the tick. The same pattern is seen when staining with patient sera IgE positive to α‐Gal. These results confirm that the α‐Gal epitope is present in I. ricinus and imply host exposure to α‐Gal during a tick bite. This provides further evidence that tick bites are associated with IgE responses to α‐Gal and red meat allergy.     

Borrelia burgdorferi infection prevalences in questing Ixodes ricinus ticks (Acari: Ixodidae) in urban and suburban Bonn, western Germany

From March to October 2003, a total of 2,518 host-seeking Ixodes ricinus ticks (1,944 nymphs, 264 females, 310 males) were collected by blanket dragging at 45 sites all over the city area of Bonn, western Germany, to be checked for Borrelia burgdorferi infection. The collection sites included 20 private gardens, nine public recreational parks, the boundaries of 14 sylvatic suburban areas and two footpaths between suburban farmed fields. Generally, numbers of specimens collected along sylvatic suburban areas and at urban sites with dense tree populations were significantly higher than at the other collection sites. Out of 1,394 specimens (865 nymphs, 241 females, 288 males) that were randomly chosen for Borrelia analysis by a simple PCR, 250 (17.9 %) were found to be infected with B. burgdorferi sensu lato. While the infection prevalences varied significantly between females (26.6%), males (12.5%) and nymphs (17.3%), there were no striking differences between sylvatic and unwooded sites. A total of 92.8% of the ticks Borrelia-positive by the simple PCR were also positive in a diagnostic nested PCR. Using genospecies-specific oligonucleotide probes, single Borrelia genospecies infections (91.4%) could be assigned to B. afzelii (39.5%), B. garinii (27.9%), B. burgdorferi sensu stricto (15.6%) and B. valaisiana (8.6%) by DNA hybridization. Various combinations of double infections were observed in 4.3% of the infected ticks. Another 4.3% of the Borrelia infections were untypeable. The B. burgdorferi genospecies distribution in the city area was shown to be variable from site to site and, even more, it was distinct from rural collection sites near Bonn. This is ascribed to a different spectrum of reservoir hosts. Taking into account the infection prevalences of host-seeking ticks in the forested surroundings of Bonn, our study demonstrates that the risk of acquiring Lyme disease after a tick bite in urban/suburban areas is comparably as high as in woodlands outside of the city.     

Attempted transmission of Borrelia burgdorferi (Spirochaetales: Spirochaetaceae) (JDI strain) by Ixodes scapularis (Acari: Ixodidae), Dermacentor variabilis, and Amblyomma americanum.

Laboratory-reared Ixodes scapularis Say, Amblyomma americanum (L.), and Dermacentor variabilis (Say) were fed on New Zealand white rabbits experimentally infected with Borrelia burgdorferi (JDI strain). At repletion, spirochetes could be detected by dark-field microscopy only in I. scapularis. Acquisition rates were 18 and 21%. When previously exposed nymphs of each species were fed on susceptible rabbits, I. scapularis was the only tick of the three species that transmitted B. burgdorferi. When a single rabbit was experimentally infected with B. burgdorferi and infested at 7-d intervals with I. scapularis, A. americanum, D. variabilis, and a second time with I. scapularis, B. burgdorferi was detected again only in cultures from the two groups of I. scapularis. When molted nymphs from each tick species were allowed to feed on susceptible rabbits, spirochetes again were isolated only at necropsy from the rabbits on which the two groups of I. scapularis fed.     

Winter infestation of wild birds by ticks and chiggers (Acari: Ixodidae,Trombiculidae) in the Czech Republic

In winter months during 2003–2006, wild birds were captured and examined for ticks and chiggers at two sites near Brno, Czech Republic. In total, 1,362 birds, mostly passerines, were examined. The tick Ixodes arboricola Schulze et Schlottke, 1929 was found on 47 (3%) birds of six species. Ixodes ricinus Linnaeus, 1758 was found on 11 (1%) birds of five species. Larvae of chiggers Ascoschoengastia latyshevi (Schluger 1955) were found on 13 (1%) birds of six species. I. arboricola and A. latyshevi associated with hole-nesting birds can appear on birds rather frequently even during winter months. I. ricinus occurs on birds in winter sporadically.     

Detection ofTheileria parva in the salivary glands ofRhipicephalus appendiculatus: evaluation of staining methods

A comparison of ten methods for staining tick salivary glands for detection ofTheileria parva infection from ticks fed on rabbits for various periods was undertaken. Staining with azure without hydrochloric acid hydrolysis was found to be the most reliable method for detection of the presporozoite stages (sporoblasts) ofT. parva in the salivary gland acini of unfedRhipicephalus appendiculatus and could be applied to field ticks. All the stains proved suitable for the detection and quantitation of sporozoites in ticks fed for 4 days on rabbits. The capacity of the stains to allow detection of early stages ofT. parva differed, but it became more reliable during tick feeding as sporoblasts developed and matured. Giemsa's stain and Feulgen's stain followed by superimposition of Giemsa's stain were superior to other stains for the detection and quantitation of immature salivary gland stages in feeding ticks.