Initial attachment of Borrelia burgdorferi spirochetes to Vero cells.

BACKGROUND: Adhesion is the initial process in the establishment of any infection and can contribute to bacterial pathogenesis. Without the ability to adhere to host cell surface, there is no invasion, dissemination, or persistence and host colonization by many bacterial pathogens, including B. burgdorferi. During the infection, B. burgdorferi cells interact with cells of various origins. We are having limited information and knowledge regarding the borrelial invasion, intracellular existence and the host cell damage and the pathological effects to the host. We have investigated by electron microscope the adherence of motile Borrelia burgdorferi s.l. to Vero cells derived from the kidney of an African green monkey by electronmicroscopy. These cells have been shown as an interesting model for study of the toxic potential of many bacterial pathogens. METHODS: Adherence of the long-term in vitro passaged Borrelia burgdorferi sensu lato strains to a 24-hour monolayer of primate kidney epithelial Vero cells was studied using transmission electronmicroscopy. The reaction was read after incubation at 1-hour intervals. RESULTS: A vertical contact between borreliae and Vero cells was confirmed already after one hour of in vitro incubation. A cytotoxic effect of borreliae could be observed when the time of incubation was extended to 4 hour. The extent of attachment varied between the two borrelia strains tested. CONCLUSION: The optimal time for spirochetal adhesion in our model was 1 h postinoculation. Our results suggest that borrelia attaches to the tested cells by length and by the tip. The data showed that the extent of attachment varied between the two borrelia strains tested (Tab. 1, Fig. 4, Ref. 21).

[Evidence of Borrelia burgdorferi spirochetes in patients with early disseminated Lyme borreliosis]. / Dôkaz spirochét Borrelia burgdorferi u pacientov s vcasnou diseminovanou formou lymeskej boreliózy.

Fifteen patients with clinically documented early disseminated lyme borreliosis were screened for the presence of spirochetes. In vitro trials of isolation of the pathogen were positive in 8 blood and 1 cerebrospinal fluid (CSF) samples. The positivity was further confirmed by electron microscopy, Western blot analysis and PCR. However, succesfull DNA extraction after using Borrelia specific primers proved the positivity in 9 blood and 1 CSF samples (67 %). Restriction fragment length polymorphism (RFLP) method using MseI restriction of PCR products of the amplified rrf-rrl region, allowed the identification of three species, namely the Borrelia garinii, B. afzelii and B. burgdorferi s.s.

Detection of Borrelia burgdorferi sensu lato and Chlamydophila psittaci in throat and cloacal swabs from birds migrating through Slovakia.

We have screened 91 migratory birds representing 32 species during the autumn of 2003 for the presence of the zoonotic pathogens Borrelia and Chlamydophila. Using polymerase chain reaction (PCR), B. burgdorferi sensu stricto was detected in cloacal swabs and, in two causes, also in throat swabs in 8 individuals (8.7 %) representing 7 birds species; B. garinii and B. afzelii were not detected. C. psittaci was detected only in cloacal swabs; 6 birds (6.6 %) from four species were found to be positive. The PCR products were sequenced and the sequences were compared phylogenetically with the gene sequences of 14 Chlamydophila strains retrieved from nucleotide databases; although the sequenced DNA was only 110 bp long, all obtained sequences created a new cluster with sublines branching from a position close to the periphery of the genus. All tested samples appear distinct within the known species and were most similar to C. felis or C. abortis.

Combined infection of Ixodes ricinus with three Borrelia burgdorferi sensu lato genotypes.

Ixodes ricinus ticks were collected by random collections from western and central Slovakia during the years 1996-98. The midgut content of 240 ticks was examined by dark-field microscopy and submitted for cultivation for the presence of borrelias. Spirochetes were found in 21 unfed and host-seeking adults and nymphs (8.8%). By the analysis of restriction fragment length polymorphism (RFLP) one sample from unfed I. ricinus male from western Slovakia was identified as a triple infection of Borrelia burgdorferi sensu stricto, B. garinii and B. afzelii. The simultaneous presence of different B. burgdorferi genospecies in one midgut sample (triple infection in the tick) could be observed only after the multipart amplification of denaturated DNA and subsequent pooling of the products for further analysis.

Immunochemical analysis of lipopolysaccharide-like component extracted from Borrelia burgdorferi sensu lato.

Immunoelectrophoresis and its modifications were applied to analysis of a lipopolysaccharide-like component (LPS-LC) extracted from Borrelia garinii strains K24 and K48 isolated from Ixodes ricinus and Borrelia burgdorferi sensu stricto strain B31. A modification of the hot phenol-water method was used for isolation of LPS. Immunoelectrophoresis (IE) and crossed immunoelectrophoresis (CIE) of LPS-LC with polyclonal rabbit antisera revealed a pattern and properties partially similar to LPS from other Gram-negative bacteria. B. garinii LPS-LC formed in CIE a diffuse band extending from the start to the anode. Similarly, the shape and position of the band in IE did not show major differences from LPS of other Gram-negative bacteria. The LPS-LC extracted from the three genomic groups of B. burgdorferi sensu lato were found to have similar immunochemical properties irrespective of their genotype origin.

Spirochetal non-Borrelia-microorganism isolated from Ixodes ricinus.

Spirochetal microorganisms were isolated from female Ixodes ricinus in Slovakia. Morphological, immunochemical and molecular biological analysis showed that the microorganism shared several common antigens with Borrelia species while other genetic traits were distinct and not related to Borrelia burgdorferi sensu lato. Lyme disease patient's serum contained antibodies reacting with antigens of this microorganism. On the one hand the cross-reacting antigens represent a risk of false positive results in laboratory diagnostics, while on the other hand they have a certain potential for vaccine development against Lyme disease.

[Lyme borreliosis: review of present knowledge]. / Lymeská borrelióza: prehl'ad doterajsích poznatkov.

The author reviews hitherto assembled knowledge on a bacterial disease, Lyme borreliosis transmitted by ticks. Initial information on Lyme borreliosis appeared at the beginning of the 20th century. In Czechoslovakia attention to the disease was paid since cca 1960. The infection occurs as a rule in the summer months during the period when ticks are parasitic and at that time the causal agent of the disease is transmitted to hosts. Information on the prevalence and incidence of Lyme borreliosis in Europe is not complete and so far we do not possess a standardized diagnostic method for assessment of circulating antibodies in the patients' serum and cerebrospinal fluid. The infectious disease is caused by the gram-negative spirochete Borrelia burgdorferi. The Borrelia cell has a similar morphological structure as cells of other gram-negative bacteria. Chemical analysis of the external membrane of B. burgdorferi revealed the presence of 46% proteins, 51% lipids and 3% carbohydrates. The typical shape of borrelias indicates marked ondulation of 8-14 periplasmatic flagellae along the cell body. Borrelias can be cultivated in vitro in modified Barbour-Stoenner-Kelly medium at an optimal temperature of 30-37 degrees C. The change of morphology during cultivation is typical for B. burgdorferi. Clinically Lyme borreliosis is manifested in two stages. A typical manifestation of the early stage is a skin lesion--erythema migrans. The later stage is characterized above all by relapsing arthritis, CNS infection and chronic acrodermatitis chronica atrophicans. The disease is treated by administration of a number of antibiotics either by the oral route or by injection.(ABSTRACT TRUNCATED AT 250 WORDS)