The aim of the present study was twofold. First, in general, to reveal new aspects of the potential vector role of ixodid ticks and fleas by screening large numbers of specimens with recently developed molecular biological methods. Second, to evaluate the occurrence of vector-borne infectious agents in a geographical context. Altogether 3442 unfed hard ticks (Ixodes ricinus, Dermacentor marginatus, D. reticulatus, Haemaphysalis inermis, H. concinna, H. punctata) and 939 fleas of cats and dogs (Ctenocephalides felis, C. canis, Pulex irritans) were collected in Hungary. DNA was extracted and analyzed in pools for representatives of the orders Rickettsiales and Mycoplasmatales. H. inermis was newly identified as the most important potential vector for Rickettsia helvetica in the study region. A novel Rickettsia genotype (designated 'Candidatus R. hungarica') was also detected in the same tick species, with a maximum of 95.8% gltA gene sequence identity to known rickettsiae. In addition, P. irritans tested positive for Rickettsia sp. RF2125, which has not been previously described in Europe. The human pathogen R. felis and the feline pathogen 'Candidatus Mycoplasma turicensis' were shown for the first time to occur in Central-Eastern Europe. Further novel findings include the presence of Spiroplasma spp. in D. marginatus and fleas. In conclusion, this molecular study extends the geographic range and vector spectrum of several arthropod-borne agents, some of which have zoonotic potential.