Prevalence of tick-borne pathogens in questing Ixodes ricinus ticks in urban and suburban areas of Switzerland.

BACKGROUND: Throughout Europe, Ixodes ricinus transmits numerous pathogens. Its widespread distribution is not limited to rural but also includes urbanized areas. To date, comprehensive data on pathogen carrier rates of I. ricinus ticks in urban areas of Switzerland is lacking. RESULTS: Ixodes ricinus ticks sampled at 18 (sub-) urban collection sites throughout Switzerland showed carrier rates of 0% for tick-borne encephalitis virus, 18.0% for Borrelia burgdorferi (sensu lato), 2.5% for Borrelia miyamotoi, 13.5% for Rickettsia spp., 1.4% for Anaplasma phagocytophilum, 6.2% for "Candidatus Neoehrlichia mikurensis", and 0.8% for Babesia venatorum (Babesia sp., EU1). Site-specific prevalence at collection sites with n > 45 ticks (n = 9) significantly differed for B. burgdorferi (s.l.), Rickettsia spp., and "Ca. N. mikurensis", but were not related to the habitat type. Three hundred fifty eight out of 1078 I. ricinus ticks (33.2%) tested positive for at least one pathogen. Thereof, about 20% (71/358) were carrying two or three different potentially disease-causing agents. Using next generation sequencing, we could detect true pathogens, tick symbionts and organisms of environmental or human origin in ten selected samples. CONCLUSIONS: Our data document the presence of pathogens in the (sub-) urban I. ricinus tick population in Switzerland, with carrier rates as high as those in rural regions. Carriage of multiple pathogens was repeatedly observed, demonstrating the risk of acquiring multiple infections as a consequence of a tick bite.

Goats as sentinel hosts for the detection of tick-borne encephalitis risk areas in the Canton of Valais, Switzerland.

BACKGROUND: Tick-borne encephalitis (TBE) is an important tick-borne disease in Europe. Detection of the TBE virus (TBEV) in local populations of Ixodes ricinus ticks is the most reliable proof that a given area is at risk for TBE, but this approach is time-consuming and expensive. A cheaper and simpler approach is to use immunology-based methods to screen vertebrate hosts for TBEV-specific antibodies and subsequently test the tick populations at locations with seropositive animals. RESULTS: The purpose of the present study was to use goats as sentinel animals to identify new risk areas for TBE in the canton of Valais in Switzerland. A total of 4114 individual goat sera were screened for TBEV-specific antibodies using immunological methods. According to our ELISA assay, 175 goat sera reacted strongly with TBEV antigen, resulting in a seroprevalence rate of 4.3%. The serum neutralization test confirmed that 70 of the 173 ELISA-positive sera had neutralizing antibodies against TBEV. Most of the 26 seropositive goat flocks were detected in the known risk areas in the canton of Valais, with some spread into the connecting valley of Saas and to the east of the town of Brig. One seropositive site was 60 km to the west of the known TBEV-endemic area. At two of the three locations where goats were seropositive, the local tick populations also tested positive for TBEV. CONCLUSION: The combined approach of screening vertebrate hosts for TBEV-specific antibodies followed by testing the local tick population for TBEV allowed us to detect two new TBEV foci in the canton of Valais. The present study showed that goats are useful sentinel animals for the detection of new TBEV risk areas.

Evaluation of a new serological test for the detection of anti-Coxiella and anti-Rickettsia antibodies.

Coxiella burnetii and members of the genus Rickettsia are obligate intracellular bacteria. Since cultivation of these organisms requires dedicated techniques, their diagnosis usually relies on serological or molecular biology methods. Immunofluorescence is considered the gold standard to detect antibody-reactivity towards these organisms. Here, we assessed the performance of a new automated epifluorescence immunoassay (InoDiag) to detect IgM and IgG against C. burnetii, Rickettsia typhi and Rickettsia conorii. Samples were tested with the InoDiag assay. A total of 213 sera were tested, of which 63 samples from Q fever, 20 from spotted fever rickettsiosis, 6 from murine typhus and 124 controls. InoDiag results were compared to micro-immunofluorescence. For acute Q fever, the sensitivity of phase 2 IgG was only of 30% with a cutoff of 1 arbitrary unit (AU). In patients with acute Q fever with positive IF IgM, sensitivity reached 83% with the same cutoff. Sensitivity for chronic Q fever was 100% whereas sensitivity for past Q fever was 65%. Sensitivity for spotted Mediterranean fever and murine typhus were 91% and 100%, respectively. Both assays exhibited a good specificity in control groups, ranging from 79% in sera from patients with unrelated diseases or EBV positivity to 100% in sera from healthy patients. In conclusion, the InoDiag assay exhibits an excellent performance for the diagnosis of chronic Q fever but a very low IgG sensitivity for acute Q fever likely due to low reactivity of phase 2 antigens present on the glass slide. This defect is partially compensated by the detection of IgM. Because it exhibits a good negative predictive value, the InoDiag assay is valuable to rule out a chronic Q fever. For the diagnosis of rickettsial diseases, the sensitivity of the InoDiag method is similar to conventional immunofluorescence.

Exposure to low doses of Coxiella burnetii caused high illness attack rates: Insights from combining human challenge and outbreak data.

BACKGROUND: As a major zoonotic pathogen, characterization of the infectivity and pathogenicity of Coxiella burnetii is essential to understand Q-fever epidemiology. OBJECTIVES: We want to extend a recently published human dose response model based on experimental challenge of young adult males to include other age groups and both genders. Additionally, we can estimate the spatial distribution of exposure based on observed outbreak data. METHODS: Dose response assessment based on human challenge, is extended by including outbreak data, using location of cases as a proxy for exposure. This allows estimation of the influence of age and gender on the probability of developing symptoms of acute respiratory illness. RESULTS: In an outbreak in Switzerland, in 1983, exposure to C. burnetii was shown to depend strongly on distance from the source. The susceptibility of males to develop Q-fever decreases with age, while in females, middle-aged women appear to have the lowest risk. CONCLUSIONS: The published dose response model for Q-fever, based on experimental challenge of a small group of human volunteers, has been updated with data from a well studied outbreak. Infectivity estimates remain high, and even low doses (of 10 or fewer organisms) cause a high risk of illness.

Prevalence and phylogenetic analysis of tick-borne encephalitis virus (TBEV) in field-collected ticks (Ixodes ricinus) in southern Switzerland.

BACKGROUND: Tick-borne encephalitis is the most common tick-borne viral infection in Europe with 3,000 human cases reported each year. In Western Europe, the castor bean tick, Ixodes ricinus, is the principal vector of the tick-borne encephalitis virus (TBEV). TBEV appears to be spreading geographically and was recently detected for the first time in Canton Valais in the southern part of Switzerland. The purpose of the present study was to survey the I. ricinus tick populations of Canton Valais for TBEV. METHODS: We collected a total of 19,331 I. ricinus ticks at 45 different sites in Canton Valais between 2010 and 2013. Ticks were processed in pools and tested for TBEV using reverse transcription quantitative PCR. The NS5 gene and the envelope gene of the TBEV isolates were partially sequenced for phylogenetic analysis. RESULTS: TBEV was detected in tick populations at six of the 45 sites. These six sites were all located in a 33 km transect along the Rhône River. TBEV was detected in two sites for three of the four years of the study showing the temporal persistence of the pathogen. Prevalence of TBEV in the six positive sites ranged from 0.16% to 11.11%. Phylogenetic analysis found that all TBEV isolates from Canton Valais belonged to the European subtype. Genetic analysis found two distinct lineages of TBEV suggesting that Canton Valais experienced two independent colonization events. CONCLUSIONS: TBEV appears to be well established at certain locations in Canton Valais.

Presence of Chlamydiales DNA in ticks and fleas suggests that ticks are carriers of Chlamydiae.

The Chlamydiales order includes the Chlamydiaceae, Parachlamydiaceae, Waddliaceae, Simkaniaceae, Criblamydiaceae, Rhabdochlamydiaceae, Clavichlamydiaceae, and Piscichlamydiaceae families. Members of the Chlamydiales order are obligate intracellular bacteria that replicate within eukaryotic cells of different origins including humans, animals, and amoebae. Many of these bacteria are pathogens or emerging pathogens of both humans and animals, but their true diversity is largely underestimated, and their ecology remains to be investigated. Considering their potential threat on human health, it is important to expand our knowledge on the diversity of Chlamydiae, but also to define the host range colonized by these bacteria. Thus, using a new pan-Chlamydiales PCR, we analyzed the prevalence of Chlamydiales DNA in ticks and fleas, which are important vectors of several viral and bacterial infectious diseases. To conduct this study, 1340 Ixodes ricinus ticks prepared in 192 pools were collected in Switzerland and 55 other ticks belonging to different tick species and 97 fleas belonging to different flea species were harvested in Algeria. In Switzerland, the prevalence of Chlamydiales DNA in the 192 pools was equal to 28.1% (54/192) which represents an estimated prevalence in the 1340 individual ticks of between 4.0% and 28.4%. The pan-Chlamydiales qPCR was positive for 45% (25/55) of tick samples collected in Algeria. The sequencing of the positive qPCR amplicons revealed a high diversity of Chlamydiales species. Most of them belonged to the Rhabdochlamydiaceae and Parachlamydiaceae families. Thus, ticks may carry Chlamydiales and should thus be considered as possible vectors for Chlamydiales propagation to both humans and animals.

Assessment of the frequency of different Borrelia burgdorferi sensu lato species in patients with Lyme borreliosis from north-east Poland by studying preferential serologic response and DNA isolates.

INTRODUCTION AND OBJECTIVE: Several Borrelia burgdorferi sensu lato species cause Lyme borreliosis throughout Europe and their geographic distribution may influence clinical manifestations of the disease. In Poland, Lyme borreliosis presents mainly with neurologic and cutaneous symptoms, while clinically overt arthritis is rare. The presented study investigates the prevalence of B. burgdorferi s.l. genospecies in a group of patients with different clinical forms and stages of Lyme borreliosis in north-east of Poland. This has not previously been studied. MATERIAL AND METHODS: Preferential reactivity towards different B.burgdorferi s.l. species was investigated with a previously validated immunoblot assay in sera of 33 patients with disseminated Lyme borreliosis: 10 with neuroborreliosis, 6 with acrodermatitis chronica atrophicans and 17 with osteoarticular symptoms. Also typed were B.burgdorferi s.l. DNA isolated from the skin and synovial fluid of 7 patients with erythema migrans, acrodermatitis chronic atrophicans and arthritis. RESULTS: Preferential reactivity was detected in 30 out of 33 serum samples. Of these, 25 reacted preferentially with B.afzelii, 3 with B. garinii and 2 with B. burgdorferi ss. B.burgdorferi DNA was isolated from all studied samples and typed as B.afzelii in 5. In a patient with acrodermatitis chronica atrophicans studied with both methods simultaneously, B.afzelii was identified by both genotyping and serotyping. CONCLUSIONS: Both methods gave consistent results, indicating B.afzelii as the main agent of all the clinical forms of the Lyme borreliosis in the study area.

Tick related diseases in Switzerland, 2008 to 2011.

QUESTION UNDER STUDY: To determine the incidence and determinants of tick related diseases in Switzerland, for example tick bites and Lyme borreliosis in primary care and tick borne encephalitis. METHODS: Analysis of the Swiss data collected by mandatory and facultative surveillance systems for the reporting period of 2008 to 2011. RESULTS: Tick related diseases in Switzerland are common. About 17,000 to 23,000 estimated cases of tick bites lead to a consultation (yearly incidence 254 per 100,000 inhabitants); about 7,000 to 12,000 estimated cases of Lyme borreliosis (yearly incidence 131 per 100,000 inhabitants) and 98 to 172 cases of tick borne encephalitis occur each year (yearly incidence 1.6 per 100,000 inhabitants). The most affected area is the north-eastern part of Switzerland. Whereas cases of tick borne encephalitis are restricted to local endemic areas, cases of Lyme borreliosis and tick bites are spread all over Switzerland. CONCLUSIONS: Tick related diseases are frequent and widespread in Switzerland. They are leading to a considerable usage of the health care system. Thus, tick bite prevention and vaccination against tick borne encephalitis are essential. However, long term follow-up cohort studies with reasonably large study populations after tick bite would be required to elucidate the risk of developing a tick borne disease.

Real-time PCR for type-specific identification of herpes simplex in clinical samples: evaluation of type-specific results in the context of CNS diseases.

BACKGROUND: HSV-1 and HSV-2 cause CNS infections of dissimilar clinico-pathological characteristics with prognostic and therapeutic implications. OBJECTIVES: To validate a type-specific real-time PCR that uses MGB/LNA Taqman probes and to review the virologico-clinical data of 25 eligible patients with non-neonatal CNS infections. RESULTS: This real-time PCR was evaluated against conventional PCR (26 CSF and 20 quality controls), and LightCycler assay (51 mucocutaneous, 8 CSF and 32 quality controls) and culture/immunofluorescence (75 mucocutaneous) to assess typing with independent methods. Taqman real-time PCR detected 240 HSV genomes per ml CSF, a level appropriate for the management of patients, and provided unambiguous typing for the 104 positive (62 HSV-1 and 42 HSV-2) out the 160 independent clinical samples tested. HSV type diagnosed by Taqman real-time PCR predicted final diagnosis (meningitis versus encephalitis/meningoencephalitis, p<0.001) in 24/25 patients at time of presentation, in contrast to clinical evaluation. CONCLUSIONS: Our real-time PCR, as a sensitive and specific means for type-specific HSV diagnosis, provided rapid prognostic information for patient management.

[Lyme borreliosis: from the erythema migrans to Lyme arthritis]. / La borréliose de Lyme: de l'erythème migrant a l'arthrite de Lyme.

An outbreak of arthritis (Lyme arthritis) initiated the research leading to the description of the Lyme disease in the years 1977-78 as a multisystemic tick-borne disease. Five years later the spirochete responsible for Lyme disease is discovered and will be named Borrelia burgdorferi, in honor to its discoverer, Dr Willy Burgdorfer. Presently in Europe, 6 Borrelia species have been described and current knowledge on epidemiology is reported here. Practical advises for the diagnostic of Lyme arthritis are proposed.

OspA heterogeneity of Borrelia valaisiana confirmed by phenotypic and genotypic analyses.

BACKGROUND: Although European Borrelia burgdorferi sensu lato isolates have been divided into five genospecies, specific tools for the serotype characterization of only three genospecies are available. Monoclonals antibodies (mAbs) H3TS, D6 and I17.3 identify B. burgdorferi sensu stricto (ss.), B. garinii and B. afzelii respectively, but no mAbs are available to identify B. valaisiana. In the same way, specific primers exist to amplify the OspA gene of B. burgdorferi ss., B. garinii and B. afzelii. The aim of the study was to develop species-specific mAb and PCR primers for the phenotypic and genetic identification of B. valaisiana. RESULTS: This study describes a mAb that targets OspA of B. valaisiana and primers targeting the OspA gene of this species. As the monoclonal antibody A116k did not react with strains NE231, M7, M53 and Frank and no amplification was observed with strains NE231, M7 and M53, the existence of two subgroups among European B. valaisiana species was confirmed. CONCLUSIONS: The association of both monoclonal antibody A116k and primers Bval 1F and Bval 1R allows to specific identification of the B. valaisiana isolates belonging to subgroup 1.

African tick bite fever: not a spotless rickettsiosis!

African tick bite fever is caused by Rickettsia africae, a newly recognized species from South Africa. We report the case of a patient with an unusual site of a tick bite and discuss cutaneous differences from other spotted fevers that may help dermatologists with clinical diagnosis.

Rhipicephalus ticks infected with Rickettsia and Coxiella in Southern Switzerland (Canton Ticino).

Ticks of the Rhipicephalus sanguineus species complex may be vector of various pathogens including Rickettsia conorii (the etiological agent of the Mediterranean spotted fever) and Coxiella burnetii (cause of the Query (Q) fever). R. sanguineus ticks have been imported in several parts of central and northern Europe, especially in environments such as kennels and houses providing the appropriate microclimatic conditions and the blood source necessary for their survival. Since 1940 these ticks have occasionally been recorded in Switzerland. In Ticino (the southern part of Switzerland), they have been reported since 1980 and their probable establishment in this area has been suggested in the '90s. By means of PCR and direct sequencing, we tested the identity of these ticks (using 12S rDNA gene) and the occurrence of Rickettsia spp. (using 16S rDNA, gltA and OmpA genes) as well as Coxiella sp. (using 16S rDNA). The results indicated that in Ticino, two different tick species coexist, i.e. R. sanguineus sensu stricto and Rhipicephalus turanicus. A few individuals of R. sanguineus sensu stricto are infected with Rickettsia massiliae/Bar29, which are strains of unknown pathogenicity. Coxiella sp., an endosymbiont of Rhipicephalus ticks, has also been identified in both tick species. Due to climatic changes towards global warming, imported tick species may therefore adapt to new area and might be considered as epidemiological markers for a number of infectious agents transmitted by them.

Interpretation of immunoblots for Lyme borreliosis using a semiquantitative approach.

OBJECTIVE: To test the performances of new Borrelia garinii immunoblots specific for Borrelia burgdorferi sensu lato with a selected panel of sera from patients with various clinical presentations of Lyme borreliosis. METHODS: In order to establish the sensitivity and the specificity of these immunoblots, we tested serum samples obtained from patients with early- and late-stage Lyme disease (erythema migrans n=35, neuroborreliosis n=61, acrodermatitis chronica atrophicans (ACA) n=27 and arthritis n=41), from patients with diagnoses and laboratory findings associated with serologic cross-reactivity to Lyme disease (syphilis n=12, Epstein-Barr infection n=9, autoimmune markers n=29) and from blood donors residing in regions of low and medium endemicity (n=80, n=100). RESULTS: The combined sensitivity (IgG and IgM) of the tests was 90% for patients with erythema migrans, 92% for neuroborreliosis, 96% for ACA and 100% for Lyme arthritis. The specificity of the IgG immunoblot was 94%, and that of the IgM immunoblot was 97%, taking into account the prevalence of borrelia antibodies in the overall population. Interpretation of these immunoblots is based on scores allocated to different specific borrelia antigens. CONCLUSIONS: The Western blot technology is extremely useful in dissecting the immune response to borrelia infections, which develops gradually over a period of weeks to years and which involves the appearance of IgM and IgG antibodies directed against a number of borrelia-associated proteins.

Association of distinct species of Borrelia burgdorferi sensu lato with neuroborreliosis in Switzerland.

OBJECTIVE: To evaluate by species-specific immunoblots the association of Borrelia burgdorferi sensu stricto, B. garinii and B. afzelii with neuroborreliosis in Switzerland. METHODS: Borrelia strains isolated from the cerebrospinal fluid (CSF) of three children with neuroborreliosis were typed by phenotypic and genotypic analysis. The serologic reactions (IgG) of these three patients as well as those of 28 patients, including one of these three children, with confirmed neuroborreliosis were characterized and scored by immunoblots on the three individual Borrelia species antigens. Twenty patients with typical erythema migrans served as a control group. RESULTS: Phenotypic and genotypic analysis confirmed that all three CSF isolates were B. garinii. In the 28 patients with neuroborreliosis, the comparatively strongest reactions were as follows: 18 to B. garinii, three to B. burgdorferi sensu stricto and two to B. afzelii; five were inconclusive. In the control group (erythema migrans), the comparatively strongest reactions were as follows: six B. garinii, one to B. burgdorferi sensu stricto and five to B. afzelii; eight were indeterminate. CONCLUSIONS: Typing of these three CSF isolates and characterization by immunoblots of the antibody reactions of patients with neuroborreliosis give additional evidence of the association of B. garinii and neuroborreliosis. Our serologic results suggest that B. burgdorferi sensu stricto and B. afzelii are also responsible for some neuroborreliosis cases in Switzerland. Our immunoblots and the scoring system proved particularly useful for the serologic typing of patients with late Lyme borreliosis.