Identification and characterization of 31 isolates of <Emphasis Type="Italic">Borrelia burgdorferi</Emphasis> (Spirochaetales,Spirochaetaceae) obtained from various hosts and vectors using PCR-RFLP and SDS-PAGE analysis

Borrelia burgdorferi sensu lato, the etiologic agent of Lyme borreliosis, circulates between ticks and vertebrate hosts. Two main genospecies typically occur in the Czech Republic Borrelia garinii and Borrelia afzelii, transmitted generally by Ixodes ricinus (L., 1758) ticks. The aim of our study was to identify spirochaete isolates focusing on Borrelia burgdorferi acquired from different sources: vectors (ticks), potential vectors (mosquitoes, small mites) and hosts (wild rodents). In the years 1996–2001 a total of 2398 ticks, 72 mites (from wild rodents), 2700 mosquito adults, 1798 mosquito larvae and organ parts (kidney and spleen) of 216 wild rodents were collected from seven localities in the Czech Republic. A total of 31 spirochaete strains were isolated: 13 strains from ticks, 1 strain from mite (Haemogamasus sp.), 15 strains from rodents, 1 strain from mosquito adults and 1 strain from mosquito larva. For the genospecies identification of these isolates PCR, PCR-RFLP was used and their characterization was also performed by SDS-PAGE. By nested PCR method all except one isolated strains were detected as Borrelia burgdorferi s.l. Following PCR-RFLP molecular analysis results, tick isolates were identified as B. garinii and B. afzelii, the strain isolated from the mite was identified as B. afzelii. This is the first isolated strain of B.b.s.l. from a different mite of infraorder Parasitiformes than tick. All of rodent isolates were identified as B. afzelii; mosquito adult isolate was identified as B. afzelii. Larval isolate from mosquito is spirochaete, but does not belong to Borrelia burgdorferi sensu lato group.     

Experimental infection of dogs with<Emphasis Type="Italic"> Borrelia burgdorferi</Emphasis> sensu stricto using<Emphasis Type="Italic"> Ixodes scapularis</Emphasis> ticks artificially infected by capillary feeding

Specific pathogen-free dogs were experimentally infected with Borrelia burgdorferi sensu stricto using nymphal or adult female Ixodes scapularis ticks artificially infected with spirochetes by capillary feeding. The ticks were capillary fed B. burgdorferi isolate 610, previously isolated from a dog with Lyme disease and grown in BSK medium. This isolate induced clinical signs in the dogs similar to those for dogs infested with ticks naturally infected with B. burgdorferi. Adult ticks were more efficient than nymphs in transmitting spirochetes to the dogs. One of five dogs infested with nymphal ticks capillary fed B. burgdorferi was skin biopsy culture and serologically positive, and demonstrated lameness. In contrast, all five dogs infested with adult female ticks that had been capillary fed with B. burgdorferi were culture and serologically positive, with one dog developing lameness. The immunoblot profiles of dogs challenged with female ticks infected by capillary feeding (8 weeks post challenge) were similar to immunoblots (4 weeks post challenge) from dogs challenged with naturally infected females collected in the field. These studies demonstrated that B. burgdorferi cultured in BSK medium can be capillary fed to either nymphal or adult female ticks under laboratory controlled conditions for the purpose of transmitting the spirochete to dogs during the tick's blood meal. This tick infection system would be useful for a controlled and defined challenge of vaccinated and non-vaccinated dogs for proper evaluation of vaccine efficacy, which is difficult to achieve using field-collected ticks. Furthermore, this system may also be useful for investigation of the pathogenesis of Lyme disease, evaluation of the pathogenicity of new isolates of B. burgdorferi, or evaluation of antibiotic therapy.     

Wild ruminants in the area of the North-Western Poland as potential reservoir hosts of <Emphasis Type="Italic">Bartonella schoenbuchensis</Emphasis> and <Emphasis Type="Italic">B. bovis</Emphasis>

The aim of this work was to examine if the game species from the north-western Poland, roe deer (Capreolus capreolus), red deer (Cervus elaphus) and wild boar (Sus scrofa), may be reservoir hosts of bacteria from the genus Bartonella, and whether the sheep tick (Ixodes ricinus) is their vector. To this end, the prevalence of Bartonella DNA in the tissues of these game species was measured, just as in sheep ticks (I. ricinus) infesting them, and ticks collected from plants in the hunting area. The prevalence of Bartonella DNA was 39% (23/59) in roe deer and 35% (7/20) in red deer. No Bartonella DNA was detected in any of the 21 wild boars. The presence of Bartonella DNAwas detected in 1.9% of ticks infesting roe deer (2/103), while no pathogen DNA was found in the 20 ticks infesting the red deer and the 3 ticks infesting wild boars, or the 200 ticks collected from plants. Amplicons of two different lengths were obtained; 198 bp, characteristic for B. bovis, and 317 bp, characteristic for B. schoenbuchensis, which were confirmed later by sequencing. The examined ruminants are probably the reservoir hosts of B. schoenbuchensis and B. bovis in the biotope of the Puszcza Wkrzańska Forest, and wild boars do not participate in the Bartonella propagation in the environment. I. ricinus is unlikely to be the main vector of Bartonella species detected in the examined roe deer and red deer; probably other bloodsucking arthropods, parasitizing wild ruminants, play this role.     

An<Emphasis Type="Italic"> ospA</Emphasis>-polymerase chain reaction/restriction fragment length polymorphism-based method for sensitive detection and reliable differentiation of all European<Emphasis Type="Italic"> Borrelia burgdorferi</Emphasis> sensu lato species and OspA types

We describe a sensitive and reliable method for detection and differentiation of the five relevant European Borrelia burgdorferi sensu lato species (B. burgdorferi sensu stricto, B. afzelii, B. garinii, B. valaisiana, and B. lusitaniae), based on a heminested ospA-PCR followed by restriction enzyme analysis. Sensitivity was one borrelia per PCR except for B. afzelii, where it was five per PCR. None of seven relapsing fever borreliae, eight Leptospira serovars or two Treponema species were amplified. Except B. garinii, each of the five B. burgdorferi s.l. species is represented by one or two characteristic restriction fragment length polymorphism (RFLP) patterns. Analysis of the heterogeneous group of B. garinii resulted in five different RFLP patterns, corresponding to the OspA types 3–7 associated with this species. In a pilot study on 529 Ixodes ricinus ticks from three different regions in Southern Germany, all species and OspA types were found except B. lusitaniae and B. garinii OspA type 7, arguing for a broad distribution of almost all OspA types. A further notable finding was the focal prevalence of OspA type 4, which has rarely been detected in ticks previously. Thus, the developed method provides a fast and simple tool for epidemiological studies on the heterogeneity of species and OspA types in Europe which has important implications for the development of vaccines and (microbiological) test systems for Europe.     

<Emphasis Type="Italic">Pseudorhabdosynochus inversus</Emphasis> sp. nov. (Monogenea,Diplectanidae) from the halfmoon grouper <Emphasis Type="Italic">Epinephelus rivulatus</Emphasis> (Perciformes,Serranidae) off New Caledonia

Pseudorhabdosynochus inversus sp. nov. is described from three specimens found in a halfmoon grouper, Epinephelus rivulatus, from the external slope of the barrier reef, New Caledonia, South Pacific. The new species is characterised by the structure of its sclerotised vagina, which resembles that of P. epinepheli (Yamaguti, 1938) but has its primary chamber inverted, and by its measurements. The diplectanid fauna of E. rivulatus shows the same pattern as in other groupers, probably belonging to a clade in which fish species harbour both an abundant species of the ‘Pseudorhabdosynochus cupatus group’ (here P. calathus Hinsinger et Justine, 2006) and a rare species (here P. inversus).

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Résumé   Pseudorhabdosynochus inversus sp. nov. est décrit de trois spécimens trouvés chez un mérou demi-lune, Epinephelus rivulatus, de la pente externe du récif barriPre, Nouvelle-Calédonie, Pacifique Sud. La nouvelle espPce est caractérisée par la structure de son vagin sclérifié, qui ressemble B celui de P. epinepheli (Yamaguti, 1938) mais a sa chambre primaire inversée, et par ses mensurations. La faune des Diplectanidae de E. rivulatus montre le mLme patron que chez d.autres mérous, probablement appartenant B un clade, chez lesquels les espPces de poissons hébergent B la fois une espPce abondante du groupe ‘Pseudorhabdosynochus cupatus’ (ici P. calathus Hinsinger et Justine, 2006) et une espPce rare (ici P. inversus).

     

<Emphasis Type="Italic">Calydiscoides limae</Emphasis> sp. nov. (Monogenea,Diplectanidae) from <Emphasis Type="Italic">Pentapodus aureofasciatus</Emphasis> (Perciformes,Nemipteridae) off New Caledonia

Calydiscoides limae sp. nov. is described from the nemipterid Pentapodus aureofasciatus Russell, 2001 caught along the barrier reef off New Caledonia, South Pacific. The new species is characterised by its male copulatory organ, with a distal blade and a lateral spur, and its female sclerotised organ, with a sphere and a thin tube. Its lamellodiscs always have 7 concentric lamellae, with the 3 internal lamellae complete and the 4 peripheral lamellae progressively less and less complete; measurements of the angles occupied by the lamellae in numerous specimens showed that the lamellodisc structure shows little variation among individuals.

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Résumé   Calydiscoides limae sp. nov. est décrit du Nemipteridae Pentapodus aureofasciatus Russell, 2001 pêché le long du récifbarrière de Nouvelle-Calédonie, Pacifique Sud. La nouvelle espèce est caractérisée par son organe copulateur male, avec une lame distale et un éperon latéral, et par son organe sclérifié femelle, avec une sphère et un tube fin. Ses lamellodisques ont toujours 7 lamelles concentriques, avec les 3 lamelles internes complètes et les 4 lamelles périphériques progressivement de moins en moins complètes. La mesure des angles occupés par les lamelles chez de nombreux spécimens montre que la structure des lamellodisques est peu variable entre les individus.

     

<Emphasis Type="Italic">Pseudopycnadena tendu</Emphasis> sp. nov. (Digenea,Opecoelidae) in the yellow-spotted triggerfish <Emphasis Type="Italic">Pseudobalistes fuscus</Emphasis> (Perciformes,Balistidae) and additional opecoelids parasitizing fishes from the waters off New Caledonia

Pseudopycnadena tendu sp. nov. is described from the balistid Pseudobalistes fuscus from the waters off New Caledonia. It differs from the only other member of the genus P. fischthali Saad-Fares et Maillard, 1986, in its broad cirrus-sac, with the wide field of large gland-cells, its less nearly circular body shape, its dorsal excretory pore, its shorter post-testicular region, its relatively larger ventral sucker and its smaller eggs. The genus is re-defined to take these distinctions into account. Other opecoelid species reported from New Caledonia are Allopodocotyle epinepheli (Yamaguti, 1942) from Epinephelus cyanopodus, E. fasciatus and E. merra, Cainocreadium epinepheli (Yamaguti, 1934) from E. coeruleopunctatus, E. fasciatus and Variola louti, Hamacreadium mutabile (Linton, 1910) from Lutjanus fulviflamma and L. kasmira, Helicometra epinepheli Yamaguti, 1934 from E. fasciatus and E. merra, Orthodena tropica Durio et Manter, 1968 from Lethrinus lentjan, Pacificreadium serrani (Nagaty et Abdel-Aal, 1962) from Plectropomus leopardus and Pseudoplagioporus interruptus Durio et Manter, 1968 from Lethrinus rubrioperculatus.

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Résumé Pseudopycnadena tendu sp. nov. est décrit du baliste Pseudobalistes fuscus pêché en Nouvelle-Calédonie. L’espèce diffère du seul autre membre du genre, P. fischthali Saad-Fares et Maillard, 1986, par son sac du cirre plus épais avec un champ large de cellules glandulaires, sa forme du corps presque circulaire, son pore excréteur dorsal, sa partie post-testiculaire plus courte, sa ventouse ventrale relativement plus grande et ses œufs plus petits. Le genre est redéfini pour prendre en compte ces distinctions. D’autres Opecoelidae sont mentionnés de Nouvelle-Calédonie: Allopodocotyle epinepheli (Yamaguti, 1942) de Epinephelus cyanopodus, E. fasciatus et E. merra, Cainocreadium epinepheli (Yamaguti, 1934) de E. coeruleopunctatus, E. fasciatus et Variola louti, Hamacreadium mutabile (Linton, 1910) de Lutjanus fulviflamma et L. kasmira, Helicometra epinepheli Yamaguti, 1934 de E. fasciatus et E. merra, Orthodena tropica Durio et Manter, 1968 de Lethrinus lentjan, Pacificreadium serrani (Nagaty et Abdel-Aal, 1962) de Plectropomus leopardus et Pseudoplagioporus interruptus Durio et Manter, 1968 de Lethrinus rubrioperculatus.

     

Identification of<Emphasis Type="Italic"> Borrelia burgdorferi</Emphasis> sensu lato,<Emphasis Type="Italic"> Anaplasma</Emphasis> and<Emphasis Type="Italic"> Ehrlichia</Emphasis> Species,and Spotted Fever Group Rickettsiae in Ticks from Southeastern Europe

Prevalence data for tick-borne pathogens are used to assess the risk for human health. In this study the presence and identity of Borrelia burgdorferi sensu lato, Ehrlichia, Anaplasma, and Rickettsia species in Bulgarian Ixodes ricinus ticks and in non-Ixodes ticks from Turkey and Albania was determined by polymerase chain reaction (PCR) and reverse line blot hybridization. In the adult Bulgarian ticks, the prevalence of Borrelia burgdorferi sensu lato infection was approximately 40%, while Borrelia afzelii was the predominant species, representing more than half of all Borrelia-positive ticks. Ehrlichia and Anaplasma species were detected in 35% of the adult Ixodes ricinus ticks and in 10% of the nymphs. Sequence analysis of PCR products reacting with the Anaplasma phagocytophila probe revealed a 16S rRNA gene identical to that of the Anaplasma phagocytophila prototype strain. Ehrlichia and Anaplasma species were found in approximately 7% of the non-Ixodes ticks. Sequence analysis of some of these samples revealed the presence of Anaplasma ovis, Ehrlichia canis, and a species closely resembling Ehrlichia chaffeensis. About half of all adult ticks examined and approximately 20% of all nymphs were infected with Rickettsia species. In Ixodes ricinus ticks, Rickettsia helvetica and a Rickettsia species designated as IRS3 were found in high prevalence. Rickettsia conorii was found in virtually all non-Ixodes tick species from Albania and Turkey. The results of this study show that many tick-borne diseases are most probably endemic in the Balkan area. Furthermore, the results suggest that there is a considerable chance for simultaneous transmission of tick-borne pathogens to human beings.     

<Emphasis Type="Italic">Demodex folliculorum</Emphasis> in patients with rheumatoid arthritis

The objective of this study was to investigate the incidence and density of Demodex folliculorum in the patients with rheumatoid arthritis (RA). Forty-one patients with RA and twenty-seven age and sex matched healthy controls were enrolled in this study. Disease Activity Score (DAS 28) was used for the assessment of disease activity. Out of 41 patients, 33 were females and 8 males. The mean disease duration was 10.9 ± 8.2 years. The mean DAS 28 was 3.8 ± 1.2. No statistically significant differences in the incidence and density of Demodex mites were found between patients with RA and controls. Although immunosuppression is thought to be a risk factor for the D. folliculorum infestation no such correlations could be found in the 41 immunosuppressed patients with RA, therefore, further studies with larger groups are needed.     

Rate of infection ofIxodes ricinus ticks withBorrelia burgdorferi sensu stricto,Borrelia garinii,Borrelia afzelii and group VS116 in an endemic focus of Lyme disease in Italy

A study to evaluate the natural rate of infection ofIxodes ricinus withBorrelia burgdorferi sensu lato was carried out in an endemic focus of Lyme disease in the Trieste area in northern Italy. Two-hundred and twenty-seven ticks collected in ten different stations were tested individually for the presence of the spirochetes using polymerase chain reaction techniques able to identify bothBorrelia burgdorferi sensu lato and the four genospecies (Borrelia burgdorferi sensu stricto,Borrelia garinii, Borrelia afzelii and group VS116). Multiple infection of individual ticks was found. The infection rate ranged from 0–70%. Infection ofIxodes ricinus withBorrelia burgdorferi group VS116 was found for the first time in Italy in both a high and a low endemic focus of Lyme disease.     

Genotyping <Emphasis Type="Italic">Borrelia burgdorferi sensu lato</Emphasis> isolates from <Emphasis Type="Italic">Ixodes ricinus</Emphasis> ticks in Russia and Ukraine

The four Borrelia burgdorferi sensu lato isolates obtained from I. ricinus ticks collected from the natural loci in Russia and Ukraine that had an unusual RFLP MseI pattern were studied using the sequencing of the rrfA-rrlB spacer and the rrs gene. Isolate Ir-5215 from a tick collected in southern Ukraine represented the recently described genospecies B. spielmanii, which is pathogenic for humans. The three atypical isolates Ir-3519, Ir-4721, and Ir-4812 had 100% identity with the sequence of atypical European B. burgdorferi sensu stricto strains; they constituted a subgroup of B. burgdorferi sensu stricto based on multilocus sequence analysis (MLSA). These data are indicative of the genetic heterogeneity of the current group B. burgdorferi sensu stricto.     

Characterisation of <Emphasis Type="Italic">Zygosaccharomyces rouxii</Emphasis> centromeres and construction of first <Emphasis Type="Italic">Z. rouxii</Emphasis> centromeric vectors

Zygosaccharomyces rouxii is a hemiascomycetous yeast known for its high osmotolerance, the basis of which still remains unknown. By exploring the Génolevures I database, four Z. rouxii fragments homologous to Saccharomyces cerevisiae centromeres were identified. Two of them were subjected to further analysis. Their function as centromeres in Z. rouxii was proved, and they were localized to Z. rouxii chromosomes II and VII, respectively. The species-specificity of centromeres was observed; plasmids with a Z. rouxii centromere were not recognized as centromeric in S. cerevisiae, and a S. cerevisiae centromere did not function as a centromere in Z. rouxii. Constructed plasmids bearing Z. rouxii centromeres serve as the first specific centromeric plasmids, and thus contribute to the so-far limited set of genetic tools needed to study the Z. rouxii specific features.     

Stimulation of biofilm formation by insertion of <Emphasis Type="Italic">Tetrahymena pyriformis</Emphasis> wells within <Emphasis Type="Italic">Burkholderia cenocepacia</Emphasis> biofilms

Biofilm formation is an important part of the bacterial life cycle. Biofilms provide bacterial resistance to external stresses and protozoan grazing. Biofilm formation by the wild type of B. cenocepacia strain 370 in the presence of the free-living ciliate Tetrahymena pyriformis was studied. T. pyriformis grazed on planktonic bacteria and reduced the planktonic bacterial subpopulation while it noticeably stimulated biofilm formation. When cultivated alone, T. pyriformis did not form visible biofilms. Confocal laser scanning microscopy was used to demonstrate the inclusion and further destruction of protozoan cells within the biofilms formed by the bacteria. The destruction of protozoan cells was accompanied by the exit of bacteria from vacuoles and intracytoplasmic multiplication; changes in the form of protozoan cells; the demolition of internal structures; and the visual exit of the cytoplasmic content from destructing cells. Microcolonies of a characteristic round shape were revealed in the biofilms formed by B. cenocepacia in the presence of T. pyriformis. These structures were absent in the biofilms formed by B. cenocepacia alone. Insertion of protozoan cells within biofilms seems to be a driving force that promotes biofilm proliferation and influences their structure. The mortality of protozoan cells in the biofilms caused a decrease in the T. pyriformis population under conditions advantageous to B. cenocepacia biofilm formation. The mutant B. cenocepacia strain Bcb-1, which is unable to form biofilms, was isolated by plasposon mutagenesis. In contrast to the parental strain, the cocultivation with Bcb-1 bacteria improved the growth of T. pyriformis. A mutation was mapped in the ompR gene. The text was submitted by the authors in English.     

Heterogeneity of the <Emphasis Type="Italic">ospA</Emphasis> gene structure from isolates of <Emphasis Type="Italic">Borrelia garinii</Emphasis> and <Emphasis Type="Italic">Borrelia afzelii</Emphasis> from Western Siberia and Mongolia

In this work, identification and analyses of 48 full-length sequences of the ospA gene isolates of Borrelia garinii and Borrelia afzelii from Western Siberia and Mongolia has been made. It was shown that B. garinii isolates was of its high genetic heterogeneity of the ospA gene. Four genetic groups of the ospA gene from the Ixodes persulcatus tick collected in of Western Siberia and Mongolia were defined. The basic differences in the genetic variants of the ospA gene considered are seen in regions which code for antibody determinants of thhe OspA protein.     

Redescription of the tapeworm <Emphasis Type="Italic">Monticellia amazonica</Emphasis> de Chambrier et Vaucher, 1997 (Cestoda,Proteocephalidea), a parasite of <Emphasis Type="Italic">Calophysus macropterus</Emphasis> (Siluriformes,Pimelodidae) from the Amazon River

The proteocephalidean cestode Monticellia amazonica de Chambrier et Vaucher, 1997 (nom. nov. for Nomimoscolex piracatinga Woodland, 1935), a parasite of Calophysus macmpterus (Lichtenstein, 1819) (Siluriformes, Pimelodidae), is redescribed on the basis of type specimens and recently collected material from the Amazon River near the type localities in Brazil, and from Peru. This insufficiently known taxon is most similar to M. ventrei de Chambrier et Vaucher, 1999 from Pinirampus spp. from Paraguay and Brazil in having almost indistinguishable internal longitudinal musculature formed by a very few (5–15) tiny bundles of muscle fibres. Monticellia amazonica differs from M. ventrei in the shape of proglottids (much more elongate in the former taxon), absence of medially situated vitelline follicles at the ovarian level (present in M. ventrei) and the number of testes (175–233 in M amazonica vs. 222–325 in M ventrei).     

Larval <Emphasis Type="Italic">Hysterothylacium</Emphasis> sp. (Nematoda,Anisakidae) and trematode metacercariae from the amphipod <Emphasis Type="Italic">Paracorophium excavatum</Emphasis> (Corphiidae) in New Zealand

Previously undescribed fourth-stage larvae of anisakid nematodes were found in the haemocoel of the amphipod Paracorophium excavatum (Thomson, 1884) (Corophiidae) in New Zealand. Morphological examination by light microscopy showed that the worms belonged to a species of Hysterothylacium Ward et Magath, 1917, based on several characters including the presence of interlabia, the location of the excretory pore posterior to the nerve ring, and the characteristics of the intestinal caecum and ventricular appendix. Interestingly, several male specimens showed precocious sexual development. This is the first record of fourth larval stage and precocious adult male specimens of Hysterothylacium in an invertebrate host, as well as the first record of anisakid larvae in New Zealand crustaceans. In addition, metacercariae of two trematode species, Coitocaecum parvum and Microphallus sp., are recorded for the first time from the amphipod P. excavatum.     

Use of cross-species <Emphasis Type="Italic">in-situ</Emphasis> hybridization (ZOO-FISH) to assess chromosome abnormalities in day-6 <Emphasis Type="Italic">in-vivo</Emphasis>- or <Emphasis Type="Italic">in-vitro</Emphasis>-produced sheep embryos

Causes of chromosomal differences such as mosaicism between embryos developed in vivo and in vitro may be resolved using animal models to compare embryos generated in vivo with those generated by different production systems. The aims of this study were: (1) to test a ZOO-FISH approach (using bovine painting probes) to detect abnormal chromosome make-up in the sheep embryo model, and (2) to examine the extent of chromosome deviation in sheep embryos derived in vivo and in vitro. Cytogenetic analysis was performed on day 6 in-vivo and in-vitro derived sheep embryos using commercially available bovine chromosome painting probes for sex chromosomes X–Y and autosomes 1–29. A total of 8631 interphase and metaphase nuclei were analyzed from 49 in-vitro-derived and 51 in-vivo-derived embryos. The extent of deviation from normal ovine chromosome make-up was higher (p < 0.05) in in-vitro-produced embryos relative to in-vivo-derived embryos (65.3% vs. 19.6% respectively) mainly due to diploid–polyploid mosaicism. Polyploid cells ranged from 3n to 8n with tetraploids most predominant among non-diploid cells. The proportions of polyploid cells per mixoploid embryo in in-vitro-produced embryos ranged from 1.4% to 30.3%, in contrast to less than 10% among the in-vivo-derived embryos. It was concluded that in-vitro-derived embryos are vulnerable to ploidy change compared to their in-vivo counterparts. The application of ZOO-FISH to domestic animal embryos is an effective approach to study the chromosome complement of species for which DNA probes are unavailable.     

Characterization of Borrelia burgdorferi sensu lato strains isolated from Ixodes ricinus in Mecklenburg-Vorpommern,Germany

Epidemiological studies in Mecklenburg-Vorpommern have shown a high prevalence ofBorrelia burgdorferi-infected ticks. A total of 17B. burgdorferi sensu lato strains were isolated from ticks and investigated by Western blots (immunoblot) with eight monoclonal antibodies against different epitopes of the outer surface protein A (OspA). Except for one, all strains could be classified using this system. The majority of strains belonged to theB. garinii-associated OspA serotypes 3, 5 and 6. Three isolates were classified as OspA serotype 2 (B. afzelii).B. burgdorferi sensu stricto strains (Ospa serotype 1) as well asB. garinii-associated OspA serotype 4 were not present.