Occurrence of different genospecies ofBorrelia burgdorferi sensu lato in ixodid ticks of Valais,Switzerland

A total of 825 adult ticks (727Ixodes ricinus, 72Dermacentor marginatus and 26Haemaphysalis punctata) was collected from vegetation in Valais (Switzerland) in 1987 to 1992. They were examined for the presence ofBorrelia burgdorferi sensu lato, the etiologic agent of Lyme borreliosis.B. burgdorferi sensu lato was detected by indirect immunofluorescence assay, dark field microscopy and/or culture in 221 out of 727I. ricinus (30.4%) and none in the other two species. From these 221 infected ticks we obtained 50 isolates. Indirect immunofluorescence assay and culture were used for all ticks but dark field examination has also been performed and compared to the two above mentioned methods for 231I. ricinus. Indirect immunofluorescence was found the most efficient method for the detection of Borrelia in ticks with 54 positive out of 231, followed by dark field examination with 35 positive and culture with 12 isolates. We found no site free of Borrelia whereI. ricinus is present. The rate of infection varied from 9.7 to 47.5%, as detected by the addition of the three methods. Typing of the 50 isolates revealed also a nonhomogeneous distribution of the Borrelia species. Based on the electrophoretic mobility of the OspA and B and immunostaining with species specific monoclonal antibodies (H3TS forB. burgdorferi sensu stricto, D6 forB. garinii and J8.3 forB. afzelii) 4 groups could be observed. Half of the isolates (n=26) were typed asB. burgdorferi sensu stricto, 19 asB. garinii, 3 asB. afzelii and 2 as group VS116. This forth group formed of two isolates from one location is genetically distinct from the 3 former species described in Europe so far. The Borreliae of this group are unreactive with any of the three monoclonal antibodies used.     

Extensive examination of different tick species for infestation with Coxiella burnetii in Slovakia

Between 1987–1989 almost 7000 adult Ixodes ricinus, Dermacentor reticulatus, Dermacentor marginatus, Haemaphysalis concinna, Haemaphysalis punctata and Haemaphysalis inermis ticks collected in all 38 districts of Slovakia were screened for the presence of Coxiella burnetii. The proportion of ticks containing C. burnetii as indicated by the haemocyte test was less than 3%. Attempts to recover C. burnetii by inoculation of yolk sacs of embryonated hen eggs from pools of 1–6 specimens of haemocyte test positive ticks resulted in the isolation of 10 rickettsial strains. Six strains were recovered from I. ricinus, the remaining ones from single pools of D. reticulatus, D. marginatus. H. concinna and H. inermis ticks.In addition to the previous recovery of C. burnetii from H. punctata ticks, the agent was thus isolated from all important ticks living in Slovakia. The agent was found in tick habitats regardless of the latitude and altitude in the entire country. These results are not consistent with the negligible number of Q fever cases occurring in past years in Slovakia.Presented at the 4th International Symposium on Rickettsiae and Rickettsial Diseases, Pietany, C.S.F.R., 1–6 October, 1990.     

Detection of Coxiella burnetii in ticks collected in Slovakia and Hungary

A total of 235 adult ticks collected from vegetation in Slovakia and Hungary in 1998–2000 were tested for Coxiella burnetii by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). C. burnetii was identified in six ticks of Ixodes ricinus, Dermacentor marginatus, and Haemaphysalis concinna species.     

A simple method of transmission risk assessment in enzootic foci of Lyme borreliosis

The proposed method consists in examining individual host-seeking vector ticks of the Ixodes ricinus complex by microscopy for borreliae and evaluating the proportion of ticks with more than 100 borreliae. The investigations were carried out in a deciduous oak forest habitat in South Moravia, Czech Republic, over four years (1991–1994). In May (i.e., at the peak of seasonal activity of I. ricinus), about 150 nymphal and 200 adult ticks were examined each year. It was found that annual incidence of human Lyme borreliosis in the region correlated better with the proportion of heavily infected ticks than with the overall infection rate of the ticks.     

The Ixodes ricinus tick as a vector of Borrelia burgdorferi in Slovenia

In many parts of Slovenia, the vegetation, climate and animals form an ideal natural environment for the persistence of different tick species. In this study, Ixodes ricinus ticks, the prevalent tick species in our country, are identified as the vectors of Borrelia burgdorferi in Slovenia. Ticks were collected by flagging between May and October 1990. All the ticks collected from different parts of Slovenia were I. ricinus and each one was examined individually. The ticks were tested to determine the presence of B. burgdorferi by indirect immunofluorescent assay. Among the 496 ticks examined for borreliae there were 85 adults and 411 nymphs. Twenty (23.5%) adults and 18 (4.4%) nymphs were found positive for Borreliae. This study discusses the importance of I. ricinus ticks in the role of Lyme borreliosis in Slovenia.     

High prevalence of Borrelia lusitaniae in Ixodes ricinus ticks in Tunisia

To investigate whether ticks of the genus Ixodes are infected by Borrelia burgdorferi complex, 490 unfed Ixodes ricinus ticks were collected by flagging in three different areas of Tunisia in 1998. DNAs extracted from 81 adults, 60 nymphs and 38 larvae were analysed after genic amplification of the noncoding spacer between the two copies of the rrl–rrf genes of B. burgdorferi sl. The prevalence of B. burgdorferi sl. in adults, nymphs and larvae was found to be 34, 33.3 and 2.6%, respectively. All DNAs (n = 61) but one were identified as belonging to different genotypes of B. lusitaniae by analysis of the restriction fragment length polymorphism of amplification products. In addition, 290 adults, 14 nymphs and 7 larvae were used to inoculate BSK-H medium to isolate spirochetes. Fifteen strains were isolated from adult ticks in the humid areas of Tunisia, whereas only one was obtained from larvae. Isolates were identified as B. lusitaniae (15/16) and B. garinii (1/16). These results provide new evidence for the existence of Lyme borreliosis in North Africa.     

Advances in investigations of lyme borreliosis in the territory of the former USSR

Whereas late manifestations of Lyme borreliosis were described in Russia more than 100 years ago, early manifestations were described as different disease entities such as tick-borne erythema, etc. In 1985 Lyme borreliosis was first verified serologically and the agent was first identified in 1986. By the beginning of 1992 Lyme borreliosis was serologically confirmed in patients from the Baltics to the Far East.Their geographical patterns in Russia are closely related to areas of the hard ticks Ixodes persulcatus and Ixodes ricinus which are also the main vectors of tick-borne encephalitis virus. As in the case of this infection, in the west of Russia there are ricinus natural foci of Lyme borreliosis, and to the east there are similar persulcatus foci; through large territories in the East Europe there are common foci for both species. Many Borrelia burgdorferi strains of tick origin have been isolated from various regions of Russia and neighbouring republics. Adult tick infection rates vary from several per cent to 30% in I. ricinus and up to 50–60% in I. persulcatus. Double infections of Lyme borreliosis spirochetes and tick-borne encephalitis virus have been recorded for ticks as well as for humans. Lyme borreliosis morbidity and its importance for the Russia territory are discussed.     

Distribution and prevalence of Borrelia burgdorferi sensu lato in Ixodes ricinus ticks of canton Ticino (Switzerland)

Free-living Ixodes ricinus ticks were collected from 12 different sites of canton Ticino, south of the Alps (Switzerland). Each tick was examined for the presence of Borrelia burgdorferi sensu lato (sl), the etiologic agent of Lyme borreliosis using direct fluorescent antibody assay, and isolation of the bacteria. Borreliae were characterized by PCR followed by RFLP. The abundance and infection rates of I. ricinus ticks varied greatly between the areas. Two localities were found free of Borrelia. The prevalence of infected ticks ranged from 5 to 19%. Most ticks (96%) were found infected by <50 spirochetes. Three B. burgdorferi sl species were successfully isolated: B. garinii dominated, followed by B. lusitaniae and B. valaisiana. Additionally, a mixed infection with B. garinii and B. valaisiana was observed. The distribution of the various Borrelia species in the different areas was heterogeneous. This is the first report of the presence of B. lusitaniae in I. ricinus in Switzerland.     

Spotted fever group rickettsiae identified in Dermacentor marginatus and Ixodes ricinus ticks in Algeria

Our study was carried out using Ixodes ricinus ticks collected from cattle from Tizi-Ouzou and Dermacentor marginatus ticks collected from the vegetation of the Blida region, a tourist site, both regions situated in northern Algeria. The results of real-time quantitative PCR (qPCR) specific for a partial sequence of the citrate synthase gene (gltA) indicate that Rickettsia spp. were present in 11/23 (48%) and 4/9 (44%) of the examined ticks from Tizi-Ouzou and Blida, respectively. The sequences of Rickettsia helvetica and Ri. monacensis were found in I. ricinus ticks using gltA primers. In addition, Ri. slovaca was detected based on the sequences of the gltA and the outer membrane protein (OmpA) genes in D. marginatus ticks. DNA sequencing to identify the species revealed for the first time the presence of Ri. helvetica in I. ricinus ticks and Ri. slovaca in D. marginatus ticks from Algeria and confirmed the presence of Ri. monacensis.     

Tick bite and Lyme borreliosis risk at a recreational site in England

The risk of tick bite and Lyme borreliosis in a forested area in England with public access was studied over a two-year period. Tick infestation levels were high with more than 1000 members of the public reporting for tick removal at a local clinic. Most of the attached ticks were nymphs (82%) and distinct differences in anatomical sites of attachment were observed in children and adults. Children sustained nymphal bites to the head, neck and axilla region much more frequently than adults (48 vs. 10%), whereas adults were bitten on the lower legs more frequently than children (46 vs. 9%). The vegetation was heavily infested with ticks and high numbers were particularly associated with areas used by deer. The average density of nymphs collected from the vegetation was 14.1 per 10 m²(range 5.1–43.6). Infection rates of these nymphs determined by PCR and indirect IFA ranged from 5.2–17.0%, and the genospeciesBorrelia valaisianaand B. gariniiwere detected, suggesting that birds may be important reservoir hosts in this area. It is estimated that, at the level of tick challenge observed here, at least 50 persons per year may be bitten by infected ticks at this site. However, no cases of Lyme borreliosis have been reported through the clinic follow-up procedure, and sera from 19 forest workers were negative for antibody to B. burgdorferisensu lato. Despite the high challenge from tick bites, this particular recreational forest site poses a low risk of infection to the general public, and prophylactic antibiotic treatment or serological testing following a bite is not justified.     

Epidemiological study of a cohort of adult patients with Erythema migrans registered in Slovenia in 1993

Epidemiological characteristics of 892 adult patients with typical Erythema migrans (Em), registered at Lyme borreliosis Outpatients' Clinic, Department of Infectious Diseases, University Medical Centre Ljubljana, Slovenia, in 1993, are presented. Data were gathered prospectively by means of a questionnaire. All the patients were seen by at least one of the four authors of this report. There were 57% females and 43% males aged from 15 to 91 years (median value 44 years, mean 44.4 ± 14.3 years). The highest number of patients was found in age groups from 30 to 59 years. 73% of the patients quoted a tick bite at the site of later Em and 6% an insect bite; 21% didn't remember any bite. 212 out of 638 patients with a tick bite were able to assess the maximal possible duration of tick attachment: two-thirds quoted 24 hours or less. Interval from a tick bite to the appearance of skin changes was one day to six months (median value 15 days, mean 20.3 ± 20.6 days) and from the beginning of Em to registration one day to 10 months (median value eight days, mean 17.1 ± 24.2 days). More than half of the patients were registered in June, July and August. The majority of patients with Em resided in the region of Ljubljana. They were bitten by ticks predominantly near their homes but some of them also quite far away. Slovenia is an endemic region for Lyme borreliosis. The number of cases presented in our report is most likely only a fraction of the real number of patients with Em in the region of Ljubljana and is even further underestimated for other parts of Slovenia.     

Unusual features in the epidemiology of Lyme borreliosis

In this study two cases of Lyme borreliosis are presented. First, the author describes how he contracted Lyme borreliosis 24 hours after he visited an endemic area. The second case described is that of a woman who developed Lyme borreliosis symptoms, when intestinal content of an infected tick came into contact with her conjunctiva. In both cases the diagnosis is based on clinical picture and positive serological tests. The first case shows the probability of contracting Lyme borreliosis when the duration of the tick's attachment to the skin is less than 24 hours. The second case, described demonstrates transmission of B. burgdorferi by contact.     

Detection of Crimean-Congo hemorrhagic fever virus in ticks collected from South Russia

Crimean-Congo hemorrhagic fever (CCHF) has a central role among tick-borne infections in southern Russia. Multiple cases of disease are recorded annually in most regions with the СCHF foci; moreover, an expansion of the geographic range of the disease has been noted. Since 1999, more than 2300 people have fallen ill in Russia. Ticks are the main vectors and reservoirs of the Crimean-Congo hemorrhagic fever virus (CCHFV). As it is currently known, CCHFV has been detected in ticks of 33 species. In the period 2012–2019 in the south of the European part of Russia, more than 38,000 ticks of 14 species of the genera Hyalomma, Rhipicephalus, Dermacentor, Haemaphysalis and Ixodes were tested for the presence of CCHFV. Among 4,188 tick pools studied, the virus was detected in 252 (6%). The main vector, as expected, is H. marginatum (81% of positive pools). The same species is the most numerous among the studied ticks (66%). Apart from H. marginatum, the virus was detected in the following species: H. scupense, R. rossicus, R. turanicus, R. bursa, R. annulatus, D. marginatus, Haem. punctata and Ixodes ricinus. Most of the positive results were obtained from ticks collected from vertebrate hosts. As for H. marginatum, R. rossicus, D. marginatus, and Haem. punctata, the virus was detected in questing unfed specimens collected from vegetation and soil surface, which indicates the participation of these ticks in the circulation of the virus.     

Prevalence of Borrelia burgdorferi (sensu lato) in Ixodes ricinus (L.) populations in France, according to a phytoecological zoning of the territory

Ixodes ricinus is considered as the main vector of Lyme Borreliosis in France. The aim of our investigations was to obtain a comprehensive view of the spatial risk linked to the distribution of the species in our country. Previous studies [1] have provided strong evidence that the species populations are widely distributed, so the objective of the present work was to ascertain the bacteriological infection of the tick by the agent of the zoonosis (Borrelia burgdorferi, sensu lato), over the French territory, whatever the ecological conditions may be. For this purpose, we kept the same framework as that used in our acarological investigation, a phytoecological zoning of the territory into 54 geographically separate and ecologically distinct units distributed into three climatic zones. Batches of ticks, picked up in these different phytoecological units (only two thirds of which were sampled), were submitted for bacteriological investigation. A total of 4,673 ticks were examined, individually, for the presence of Borellia burgdorferi, by immunofluorescence. Percentages of infection according to the various stages of Ixodes ricinus free stages, collected by flagging, were as follows: 4.95% in 3,247 nymphs, 11.2% in 699 males, 12.5% in 727 females. Larvae were ignored. Practically all the tested units harboured the bacterium. The percentage of tick samples (25 ticks or more) absolutely free of Borrelia, wherever they came from, is very low (not exceeding 10 percent of the sampled forests). Our study confirms the assessment of a widespread distribution of the zoonosis in France which was, until now, exclusively based on an approximate distribution of limited human cases observed in the country.     

Seroepidemiological survey forBorrelia burgdorferi (Lyme disease) in dogs from northwestern of Spain

A random epidemiological study was undertaken to estimate the prevalence of canine borreliosis (Lyme disease) in Castilla y León, the largest region in Spain. The presence of antibodies was determined by indirect immunofluorescence assay (IFA), using theBorrelia burgdorferi B31 strain as antigen. Sera from 308 dogs from 7 provinces in the region were tested. Of all the animals sampled, 37 (21%) were seropositive (titres of 1/64 or above). Almost all the provinces had seroprevalence of 20% or below, except for an important focus of borreliosis in the Zamora province (42.8%). Potential risk factors (age, sex, use, habitat, season, and presence of ticks on the animals) relating to the presence of antibodies were evaluated. Those dogs which had at some time had ticks were more often seropositive, at 24.2%, than those which had never had them, at only 6.2% (p<10^–n). No significant differences were discovered for the remaining factors studied. This work indicates that dogs in Castilla y Leon, even if in most cases they do not develop the disease, are exposed to the Lyme disease agent.     

Coinfections of Rickettsia slovaca and Rickettsia helvetica with Borrelia lusitaniae in ticks collected in a Safari Park,Portugal

Borrelia and Rickettsia bacteria are the most important tick-borne agents causing disease in Portugal. Identification and characterization of these circulating agents, mainly in recreational areas, is crucial for the development of preventive measures in response to the gradually increasing exposure of humans to tick vectors. A total of 677 questing ticks including Dermacentor marginatus, Rhipicephalus sanguineus, Ixodes ricinus, Hyalomma lusitanicum, H. marginatum, and Haemaphysalis punctata were collected in a Safari Park in Alentejo, Portugal, to investigate the prevalences of infection and characterize Borrelia and Rickettsia species. From a total of 371 ticks tested by PCR for Borrelia burgdorferi sensu lato (s.l.), of which 247 were tested for Rickettsia, an infection prevalence of 18.3% was found for B. lusitaniae and 55.1% for Rickettsia spp. Sequence analysis of positive amplicons identified the presence of B. lusitaniae (18.3%), R. monacensis strain IRS3 (51.7%), and R. helvetica (48.3%) in I. ricinus. R. slovaca (41.5%), R. raoultii (58.5%), and also B. lusitaniae (21%) were identified in D. marginatus ticks. One (5.9%) H. lusitanicum was infected with B. lusitaniae, and R. massiliae was found in one Rhipicephalus sanguineus. Coinfection was found in 7 (20%) I. ricinus and 34 (23.3%) D. marginatus ticks. We report, for the first time, simultaneous infection with R. helvetica and B. lusitaniae and also R. slovaca, the agent of TIBOLA/DEBONEL, with B. lusitaniae. Additionally, 6 isolates of B. lusitaniae were established, and isolates of Rickettsia were also obtained for the detected species using tick macerates cultured in mammalian and mosquito cell lines. This report describes the detection and isolation of tick-borne agents from a Portuguese Safari Park, highlighting the increased likelihood of infection with multiple agents to potential visitors or staff.     

Prevalence of ticks infected withFrancisella tularensis in natural foci of tularemia in western Slovakia

The prevalence of ticks infected withF. tularensis was followed during a systematic surveillance in endemic area of tularemia in western Slovakia over the years 1984–93. Ticks were collected from vegetation in localities of Podunajské Biskupice, in the vicinity of the capital of Slovakia, Bratislava, near the river Danube. In total 6033 ticks, mostly adults ofDermacentor reticulatus andIxodes ricinus (4994 and 1004, respectively) and 35 nymphs ofHaemaphysalis concinna, were examined for the presence ofF. tularensis. Out of 4542 starving ticks, 34F. tularensis strains were isolated predominantly fromD. reticulatus (30), and to a smaller extent also fromI. ricinus (3) andH. concinna (1). Natural infection withF. tularensis was further proved from 1491 adults ofD. reticulatus fed on laboratory animals, rabbits and white mice, together in 27 cases. From that, 21 times it was by positive isolation either from suspensions of partly or fully engorged ticks and their feaces, or from spleens of animals dead after the feeding of ticks. In addition, solely the development of antibodies against the agent was confirmed in 6 rabbit hosts. The presence ofF. tularensis in all the above mentioned tick species and namely the relatively high and permanent infestation ofD. reticulatus adults, ranging between 0.5–2% during the followed time period, demonstrated the maintenance of active natural focus of tularemia in the area under study. The present paper also emphasizes the epidemiologic consequence of various species of ticks in endemic foci of tularemia and the aspect of possible ways of transmission of the agent to humans.     

Borrelia burgdorferi in rodents (Apodemus flavicollis andA. sylvaticus): Duration and enhancement of infectivity forIxodes ricinus ticks

Ixodes ricinus is an important vector ofBorrelia burgdorferi in Europe, and small rodents (Apodemus flavicollis, A. sylvaticus andClethrionomys glareolus) are important sources for infecting ticks. In this study, we examined their reservoir role by studying the duration of their infectivity for ticks.A. flavicollis andA. sylvaticus mice captured in nature were exposed to uninfectedI. ricinus larvae at different times after their capture: 10 days, and 2, 7, 11, 14 and 40 months. Ticks were examined for spirochaetes after moulting using direct immunofluorescence. All animals remained infective for ticks their life long but the efficiency of transmission from hosts to ticks varied from one individual to the other, presenting a three-fold variation (26.5% to 81.4%). Rodents continously exposed to successive infestations by larvalI. ricinus ticks over a period of one month showed an enhancement of infectivity for larval ticks during this period.     

Detection and molecular typing of Borrelia burgdorferi sensu lato in Ixodes ricinus ticks and in different patient samples from southwest Germany

The prevalence of different genospecies of Borrelia burgdorferi sensu lato in infected ticks could be a determinant for the risk of acquiring Lyme borreliosis (LB) and its clinical presentation. A total of 7373 ticks and 2761 samples from LB patients from the same area in southwest Germany were analyzed by PCR to assess the frequency of the occurrence of LB-associated genospecies. Fifteen percent of the tick samples and 19% of the human samples were found positive for the presence of B. burgdorferi sensu lato. Further identification of 1106 B. burgdorferi sensu lato positive tick samples by reverse line blotting and 125 positive patient samples by nested PCR using species-specific primers revealed the occurrence of B. afzelii, B. burgdorferi sensu stricto, B. garinii and B. valaisiana. Both single-species and mixed infections were noted and a similar distribution of the different genospecies was found in ticks compared with human samples. It was also the purpose of this study to obtain more information about a possible correlation between the distribution of Borrelia species and clinical syndromes of LB. Skin biopsies of 59 patients with acrodermatitis chronica atrophicans and cerebrospinal fluid samples from 78 patients with possible neuroborreliosis were analyzed. In conclusion, the distribution of the different genospecies in ticks is the decisive factor for the occurrence of the different Borrelia genospecies in samples from LB patients. Borrelia afzelii is the predominant genospecies in all kind of samples from the observed area and there seems to be no association of particular Borrelia genospecies with distinct clinical manifestations of LB.     

Efficacy of Hyalomma scupense (Hd86) antigen against Hyalomma excavatum and H. scupense tick infestations in cattle

The Rhipicephalus microplus recombinant Bm86-based tick vaccines have shown their efficacy for the control of several Hyalomma cattle ticks genera, namely H. dromedarii and H. anatolicum. However, H. scupense species, the most important tick in North Africa has never been studied. Vaccination trials using either a recombinant Bm86-based vaccine or a recombinant Hd86-based vaccine (the Bm86 ortholog in H. scupense) were conducted in cattle against immature and adult H. scupense ticks and adult H. excavatum ticks. The results showed a 59.19% reduction in the number of scupense nymphs engorging on Hd86 vaccinated cattle. However, cattle vaccination with Bm86 or Hd86 did not have an effect on H. scupense or H. excavatum adult ticks infestations. These results showed that Hd86 vaccines are selectively effective against H. scupense immature instars and emphasize on an integrated anti-tick vaccine control in North Africa.