Lyme borreliosis is a tick-transmitted infectious disease. The causative agents are spiral-shaped bacteria and the most common sign of infection is a skin rash at the site of the tick bite. If not treated with antibiotics, the bacteria can disseminate and cause a variety of different manifestations including arthritis, carditis or neurological problems. The disease is a zoonosis and the bacteria are maintained in nature by different vertebrate reservoir host animals. In Europe, three different Borrelia genospecies cause Lyme borreliosis: B. burgdorferi, B. afzelii and B. garinii. The latter depends in part on birds as its reservoir host. B. garinii bacteria have been found in a marine enzootic infection cycle worldwide and also among terrestrial birds. This thesis suggests that passerine birds and seabirds constitute an important reservoir for B. garinii bacteria also with clinical importance. We have found bacteria very similar to Lyme borreliosis causing isolates in ticks infesting migrating passerine birds. The birds not only transport infected ticks, but are competent reservoir hosts, as measured by their ability to infect naïve ticks. Their role as a reservoir host is dependent on their foraging behavior, where ground-dwelling birds are of greater importance than other species. When comparing B. garinii isolates from Europe, the Arctic and North Pacific, and including isolates from seabirds, passerine birds, Ixodes ricinus ticks and Lyme borreliosis patients, we found that phylogenetic grouping was not necessarily dependent on geographical or biological origin. B. garinii from seabirds were very heterogeneous and found in all different groups. Therefore, the marine and the terrestrial infection cycles are likely to overlap. This was supported by the fact that B. garinii isolated from seabirds can establish a long-term infection in mice. Bacteria from the genospecies B. garinii are overrepresented among neuroborreliosis patients. Interestingly, many clinical B. garinii isolates are sensitive to human serum and have shown weak binding to the complement inhibitor protein factor H. By transforming a serum-sensitive B. garinii isolate with a shuttle vector containing the gene for the factor H binding protein OspE from complement-resistant B. burgdorferi, serum resistance could be increased. In addition, neurovirulent B. garinii strains recently isolated from neuroborreliosis patients were shown to express a factor H binding protein, not found in bacteria that had been kept in culture for a long time. This protein may contribute to the virulence of neuroborreliosis-causing B. garinii strains. When testing B. garinii isolates from Lyme borreliosis patients and seabirds for resistance to human serum, all members of the latter group were sensitive to even low levels of serum. This suggests that seabird isolates are not capable of infecting humans. In agreement with this, B. garinii isolated from seabirds do not appear to bind human factor H.

The topic of this thesis is a logical continuation of the bachelor project. In our previous project, using the method of Borrelia cultivation on solid medium, we succeeded obtaining a monoclonal population of B. garinii from the american isolate in which the presence of more borrelia species was confirmed. By this we confirmed the presence so-called "european" Borrelia species in the southeastern United States (specifically in South Carolina). By multilocus analysis of eight housekeeping genes of american isolate it has been shown that the obtained strain of B. garinii is different from the previously described european and asian strains of this species. The main objective of the proposed project, is the analysis of the pathogenic potential of the isolated clone of B. garinii. Validation of the pathogenic potential of American B. garinii isolate in laboratory mice model could be important for the public health of the southeast american population, as this will confirm the possibility of existence of Lyme disease, caused by this spirochete species, with significantly different clinical signs in this region. For this purpose, the method of direct diagnosis (infecting laboratory animals and transferring of infection from infected animals to the tick and vice versa) was used. Laboratory mice were infected subcutaneously, and the spirochete infection was confirmed in ear tissue by PCR. Uninfected Ixodes ricinus larvae were fed on infected mice. After the confirmation of infection in newly hatched nymphs they were fed on naive mice. The ability of transmitted spirochetes to disseminate into different mice tissues was confirmed by PCR. The presence another so-called "European" species, B. afzelii, was confirmed in original rodent associated American isolate. Another aim of the project was to obtain monoclonal populations B. afzelii from co-infected northamerican sample confirming by this way the transoceanic migration of spirochtes.