High Prevalence and Low Diversity of Rickettsia in Dermacentor reticulatus Ticks, Central Europe.

We collected 1,671 Dermacentor reticulatus ticks from 17 locations in the Czech Republic, Slovakia, and Hungary. We found 47.9% overall prevalence of Rickettsia species in ticks over all locations. Sequence analysis confirmed that all tested samples belonged to R. raoultii, the causative agent of tick-borne lymphadenopathy.

Landscape epidemiology of neglected tick-borne pathogens in central Europe.

Studies of tick-borne diseases (TBDs) in Europe focus on pathogens with principal medical importance (e.g. Lyme disease and tick-borne encephalitis), but we have limited epidemiological information on the neglected pathogens, such as the members of the genera Anaplasma, Rickettsia, Babesia and Candidatus Neoehrlichia mikurensis. Here, we integrated an extensive field sampling, laboratory analysis and GIS models to provide first publicly available information on pathogen diversity, prevalence and infection risk for four overlooked zoonotic TBDs in the Czech Republic. In addition, we assessed the effect of landscape variables on the abundance of questing ticks at different spatial scales and examined whether pathogen prevalence increased with tick density. Our data from 13,340 ticks collected in 142 municipalities showed that A. phagocytophilum (MIR = 3.5%) and Ca. Neoehrlichia mikurensis (MIR = 4.0%) pose geographically uneven risks with localized hotspots, while Rickettsia (MIR = 4.9%) and Babesia (MIR = 1.1%) had relatively homogeneous spatial distribution. Landscape variables had significant effect on tick abundance up to the scale of 1 km around the sampling sites. Questing ticks responded positively to landscape diversity and configuration, especially to forest patch density that strongly correlates with the amount of woodland-grassland ecotones. For all four pathogens, we found higher prevalence in places with higher densities of ticks, confirming the hypothesis that tick abundance amplifies the risk of TB infection. Our findings highlight the importance of landscape parameters for tick vectors, likely due to their effect on small vertebrates as reservoir hosts. Future studies should explicitly investigate the combined effect of landscape parameters and the composition and population dynamics of hosts on the host-vector-pathogen system.

Duplex qPCR assay for detection and quantification of Anaplasma phagocytophilum and Rickettsia spp.

Anaplasma phagocytophilum and Rickettsia spp. are vector-borne zoonotic bacteria, which are clinically important especially in immunocompromised patients. There are large gaps in the current knowledge of their geographic distribution and prevalence in both their vectors and hosts. Our aim was to develop reliable and easy detection method for both these pathogens. We made a new hydrolysis probe based duplex Real-Time PCR assay based on previous studies. We optimized the assays and tested them to provide reliable recommended procedures with a sensitivity to a minimum of 10 target DNA copies per sample. The assays were designed to be specific for A. phagocytophilum and in the same reaction detect multiple species of rickettsiae. We designed gBlock quantification standards that provide the option to identify differences in pathogen load among different samples in subsequent studies.

Candidatus Neoehrlichia mikurensis is widespread in questing Ixodes ricinus ticks in the Czech Republic.

Candidatus Neoehrlichia mikurensis, the causative agent of tick-borne "neoehrlichiosis" has recently been reported in humans, mammals and ticks in Europe. The aim of this study was to map the distribution of this bacterium in questing ticks in the Czech Republic. A total of 13,325 Ixodes ricinus including 445 larvae, 5270 nymphs and 7610 adults were collected from vegetation by flagging in 140 Czech towns and villages from every region of the Czech Republic. The ticks were pooled into 2665 groups of 5 individuals respecting life stage or sex and tested for the presence of Ca. Neoehrlichia mikurensis by conventional PCR targeting of the groEL gene. The bacterium was detected in 533/2665 pools and 125/140 areas screened, showing an overall estimated prevalence of 4.4 % in ticks of all life stages. Phylogenetic analysis revealed only small genetic diversity among the strains found. Two pools of questing larvae tested positive, suggesting transovarial transmission. According to this study, Ca. Neoehrlichia mikurensis is another tick-borne pathogen widespread in I. ricinus ticks in the Czech Republic.