Ultrastructural study of the life cycle of Rickettsia slovaca, wild and standard type, cultivated in L929 and Vero cell lines.

Ultrastructural changes induced by Rickettsia slovaca standard type (ST) and wild type (WT) were examined during their life cycle in L929 and Vero cells. R. slovaca invaded the cytoplasm of the host cell by phagocytosis on the 1st d p.i. Rickettsiae adhering to the cytoplasmic membrane were engulfed by cellular extensions and occurred in phagocytic vacuoles. Binary fission of rickettsia was observed. The nuclear chromatin of eukaryotic cells was rearranged and condensed during 3rd and 6th d p.i. Finally, loss of the plasma membrane integrity, destruction of cytoplasm and nucleus resulted in cell lysis. Degeneration of the host cell caused by WT and ST was observed after 4 and 5 d p.i. in L929 cells and after 3 and 6 d p.i. in Vero cells, respectively. WT type was able to penetrate into the nucleus of the host cell and was responsible for dilatation of the perinuclear space and endoplasmic reticulum, causing more pronounced and different cytopathological changes than the ST.

Incidence of various tick-borne microorganisms in rodents and ticks of central Slovakia.

In this study, we detected Rickettsia helvetica, Candidatus Midichloria mitochondrii, Anaplasma phagocytophilum, Ehrlichia muris, Candidatus Neoehrlichia mikurensis, and Bartonella sp. infections in wild rodents and ticks collected from the vegetation of central Slovakia. The microorganisms were identified by PCR and sequencing. Yellow-necked mice (Apodemus flavicollis) were infected with E. muris and Bartonella sp., while ticks Ixodes ricinus collected from the vegetation were infected with R. helvetica, Candidatus M. mitochondrii, Candidatus N. mikurensis, A. phagocytophilum, and E. muris.

Rickettsial agents in Slovakian ticks (Acarina, Ixodidae) and their ability to grow in Vero and L929 cell lines.

A total of 80 adult ticks (55 Haemaphysalis inermis, 12 Dermacentor reticulatus, 11 D. marginatus, 2 Ixodes ricinus) were collected from vegetation in three areas of Slovakia (forest and pasture habitat) in central Europe. Forty-six (46 ticks) (57.5%) of all species tested were positive by the hemocyte test, PCR assays based on the gltA and ompA genes showed a Rickettsiaceae infection in 77.5% of the ticks, whereas only one H. inermis tick was positive for Anaplasmataceae on a 16S rRNA-based PCR. Isolation of rickettsiae was attempted on all collected ticks by means of the shell vial technique, 52 isolates of which were inoculated into Vero cells and 28 into L929 cells. Rickettsiae were detected in 50% (40/80) of the cell lines using the Gimenez staining method, whereas 33.8% (27/80) of the cell lines were PCR-positive for Rickettsia species. The presence of rickettsiae was shown by PCR to be around 30.8% (16/52) in Vero and 39.3% (11/28) in L929 cell lines. Sequencing results showed that detected infections were Rickettsia sp., R. raoultii, and Anaplasma phagocytophilum in ticks, and R. slovaca in cell lines. This is the first report of R. raoultii in Slovakia. Observations by electron microscopy of the R. slovaca isolate from Vero cell lines showed a microcapsular layer, typical Gram-negative cell wall, and a cytoplasmic membrane.