Stefancíková A

Occurrence of Borrelia burgdorferi sensu stricto, Borrelia garinii and Borrelia afzelii in the Ixodes ricinus ticks from Eastern Slovakia.

A total of 2816 unfed adults nymphs of Ixodes ricinus ticks were collected from vegetation in Košice (Eastern Slovakia) from 1994 to 1997. Prevalence of Borrelia burgdorferi s. 1. in I. ricinus ticks, detected by dark field microscopy, varied and depended upon the year and the habitat of the collected ticks. The lowest prevalence was observed in 1994 (4.8%). During 1995 it increased to 17.2% and during the next two years decreased to 15.5% and 14.2%. The rate of infection varied from 2.1 to 23.3% within 10 examined habitats of the Košice area. A different value of relative density of ticks was observed in various habitats. It ranged from 9–212 ticks per collecting hour within one flagged area (600 m2) which is 1.5–35.5 ticks per 100 m2. Eight isolates were obtained from the infected ticks. Electrophoresis and immunoblotting with 6 monoclonal antibodies were used for the identification of Borrelia strains. Three tick isolates were identified as B. burgdorferi s. s. and the other three isolates were found to be B. garinii. One strain reacted as a mixed culture of B. burgdorferi s. s., and B. garinii. The strain originated from the Vihorlat Mountains habitat and was detected by PCR-SSCP as B. burgdorferi s. s. with a small amount of B. afzelii. The obtained results emphasize the epidemiological importance not only of B. garinii and B. afzelii but also of B. burgdorferi s. s. in Central Europe.

Emergence and genetic variability of Anaplasma species in small ruminants and ticks from Central Europe.

Anaplasmoses are common tick-borne zoonotic bacterial diseases of livestock and free-living ungulates from the genus Anaplasma that are recently emerging in Central Europe. The main aim of this study was to analyze the prevalence and genetic variability of Anaplasma phagocytophilum and Anaplasma ovis in small ruminants and questing ticks from six different sites in Slovakia and the Czech Republic using the PCR of the msp4 gene followed by the sequence analysis. At two farms from southeastern Slovakia, 66.1% small ruminants were infected with A. ovis in contrast to one positive animal from both sites in northern Slovakia. It was represented by two different genotypes. A. phagocytophilum was present in all tested flocks with the infection prevalence ranging from 0.9% to 5.7%. None of the tested questing ticks carried A. ovis. A. phagocytophilum was detected in 1.1% and 7.8% of questing Ixodes ricinus ticks collected around the farms located in southeastern and northern Slovakia, respectively. A. phagocytophilum revealed higher intraspecific diversity than A. ovis.

Serological and molecular detection of Borrelia burgdorferi sensu lato and Anaplasmataceae in rodents

The presence of Anaplasma spp. and Borrelia burgdorferi sensu lato in rodents from Eastern Slovakia were followed by serological and molecular methods. The seroprevalence for Borrelia was detected in 16.6 %, for Anaplasmataceae (APT) in 13.2 % and co-occurrence of Borrelia and APT in 7.5 %. Out of 110 ear biopsies of rodents, 5 were B. afzelii-positive. Five biopsies tested positive with the Ehr521-Ehr747 primers amplifying all the members of the family APT. A. phagocytophilum was detected in 1.8 %, 2.7 % were infected with Anaplasma-like organisms. Co-occurrence of Borrelia and Anaplasma in ear biopsies was found in 1.8 %. The circulation of both Borrelia and Anaplasma in the region of Eastern Slovakia was confirmed.

Evaluation of Recombinant Line Immunoblot for Detection of Lyme Disease in Slovakia: Comparison with Two Other Immunoassays

In the present study the sensitivity and the specificity of three serological tests (enzyme-linked immunosorbent assay [ELISA], indirect fluorescent antibody test [IFA], and recombinant line immunoblot) were compared by examining 74 sera from patients diagnosed with Lyme disease in Eastern Slovakia. In addition, the reactivity to each of the recombinant proteins in the immunoblot was examined in order to evaluate their diagnostic value. Generally, the immunoblot (93.2%) and the ELISA (90.5%) were significantly more sensitive than the IFA (64.9%; df = 1; p < or = 0.001). Correlation between results of the ELISA, IFA, and immunoblot for IgM or IgG, when two tests were always compared, one to the other, ranged from r(s) = 0.673 to r(s) = 0.905. In the immunoblot, the highest sensitivity was observed in DbpA and VlsE proteins (76.9% and 84.6%, respectively) in IgG testing of the sera from the patient group of Lyme arthritis. VlsE proteins, together with OspC proteins, were also shown to be useful for IgM antibody detection in erythema migrans patients (up to 44.4% and 53.7% sensitivity, respectively). Our results indicate that both the ELISA and the recombinant immunoblot test were more satisfactory for seroconfirmation of Lyme disease than IFA. Moreover, the reseach confirmed diagnostic value of the in-vivo expressed proteins (VlsE and DbpA), which might have the potential to play an important role in improving whole-cell antigen-based testing.

Some epidemiological and epizootiological aspects of Lyme borreliosis in Slovakia with the emphasis on the problems of serological diagnostics

The paper analyses the results of serological examinations of domestic, farm and free-living animals from different regions of Slovakia, Southern Moravia, Southern Bohemia and Southern Poland using ELISA, indirect hemagglutination assay (IHA) and Western blot (WB). In Slovakia, significantly higher seroprevalence was recorded in dogs (33.5%) than in horses (26.5%), cattle (22.5%), sheep (16.6%) and rodents (17.8%) by using a mixture of Borrelia garinii, B. afzelii, B. burgdorferi sensu stricto (s.s.) antigens in ELISA. Seroprevalence in horses was significantly higher than in sheep and rodents, and seroprevalence in cattle was significantly higher than in rodents. By using IHA in free-living species, the highest seropositivity rates were detected in fallow deer (40.7%) compared with moufflons (16.6%), pheasants (8.0%) and pigeons (1.2%). When testing sera of horses, dogs and cattle from Slovakia by using different Slovak B. burgdorferi sensu lato (s.l.) isolates as antigens in ELISA, significantly higher seroprevalence of anti-Borrelia IgG antibodies and consistency of positive and negative findings was detected in comparison when American isolates were used. In WB analyses using the Eurocarpathian antigens, dog sera from Eastern Slovakia and Southern Moravia showed statistically insignificant differences in sensitivity and consistency of positive and negative findings. By using different methods and antigens in the same group of dog sera, significant differences in seroprevalence were only found in IHA with a mixture of Euroamerican B.b.s.l and WB CB26 B.b.s.s. In addition to other factors, the complexity of the standardization of the assay system with regard to the genetic and geographical heterogeneity of B. burgdorferi s.l. isolates used as antigens is also discussed.

Immune complexes in early Lyme disease

The study investigated the presence of Borrelia-specific antibodies captured in immune complexes (ICs) in patients with early Lyme disease manifested by erythema migrans. Out of 18 patients, 15 (83.3%) tested positive for polyethylene glycol-precipitated ICs containing IgM antibodies, while only 4 (22.2%) were IgG positive. These results are in accordance with our findings obtained by standard ELISA and recombinant blot, which indicated that ICs might be used for serological diagnosis of the early disease.