Tissue samples from nine red foxes (four samples of striated muscle tissue and five samples of heart tissue) that originated from the Michalovce district (Slovakia), an area with endemic occurrence of canine babesiosis were examined by PCR method using primers amplifying a fragment of the 18S rRNA spanning the V4 region of Babesia and Theileria. An unexpected determination of 450 bp DNA fragment of Hepatozoon canis was found in four samples. Partial sequences of the 18S rRNA gene from the H. canis showed 100% similarity with the sequence from Brasil isolate of H. canis from a pampas fox (Pseudalopex gymnocercus) (AY471615) as well as from a fox in Spain (AY150067) and from a dog in Brazil (AY864677). In the present study, we report the first PCR detection of Hepatozoon canis in a naturally infected red fox from Slovakia, a Rhipicephalus sanguineus-free region. We assume that the infection was spread by infected R. sanguineus that might have been brought to Slovakia by travelers, by golden jackals, or by foxes migrating because of expansion of golden jackals and environmental and climate changes. Vector-Borne Zoonotic Dis. 7, 199–202.

Blood parasites of the genus Karyolysus Labbé, 1894 (Apicomplexa: Adeleida: Karyolysidae) represent the protozoan haemogregarines found in various genera of lizards, including Lacerta, Podarcis, Darevskia (Lacertidae) and Mabouia (Scincidae). The vectors of parasites are gamasid mites from the genus Ophionyssus. A total of 557 individuals of lacertid lizards were captured in four different localities in Europe (Hungary, Poland, Romania and Slovakia) and blood was collected. Samples were examined using both microscopic and molecular methods, and phylogenetic relationships of all isolates of Karyolysus sp. were assessed for the first time. Karyolysus sp. 18S rRNA isolates were evaluated using Bayesian and Maximum Likelihood analyses. A total of 520 blood smears were examined microscopically and unicellular protozoan parasites were found in 116 samples (22.3% prevalence). The presence of two Karyolysus species, K. latus and K. lacazei was identified. In total, of 210 samples tested by polymerase chain reaction (PCR), the presence of parasites was observed in 64 individuals (prevalence 30.5%). Results of phylogenetic analyses revealed the existence of four haplotypes, all part of the same lineage, with other parasites identified as belonging to the genus Hepatozoon. Classification of these parasites using current taxonomy is complex - they were identified in both mites and ticks that typically are considered to host Karyolysus and Hepatozoon respectively. Furthermore although distortions to the intermediate host erythrocyte nuclei were observed, the defining characteristic of Karyolysus, the haplotypes were nearly identical to those reported from lizards in the Iberian Peninsula, where such distortions were not reported and which were thus identified as Hepatozoon. Based on the phylogenetic analyses, neither vertebrate host, nor geographical patterns of the studied blood parasites could be established.

Background: Anaplasmataceae and Borrelia burgdorferi s.l. are important tick-borne bacteria maintained in nature by transmission between ticks and vertebrate hosts. However, the potential role of lizards as hosts has not been sufficiently studied. Results: The current study showed that 23 of 171 examined sand lizards Lacerta agilis were PCR positive for Anaplasmataceae. The nucleotide sequences of the several selected PCR products showed 100% homology with Anaplasma spp. found in Ixodes ricinus collected in Tunisia and Morocco (AY672415 - AY672420). 1.2% of lizard collar scale samples were PCR positive for B. lusitaniae. In addition, 12 of 290 examined I. ricinus were PCR positive for B. burgdorferi s.l. and 82 were PCR positive for Anaplasmatacea. The number of ticks per lizard and the number of ticks PCR positive for both microorganisms per lizard were strongly correlated. Moreover, we found a significant correlation between numbers of ticks infected with Anaplasmataceae and with B. burgdorferi s.l. living on the same lizard. However, there was no significant correlation between detection of both bacteria in the same tick. Conclusions: To the best of our knowledge, this is the first report of Anaplasmataceae DNA and additionally the second report of B. burgdorferi s.l DNA detection in the sand lizard.

To investigate the involvement of lizard species in the natural cycle of Borrelia burgdorferi sensu lato (s.l.) in Hungary, a total of 186 reptiles belonging to three species—126 green lizards ( Lacerta viridis ), 40 Balkan wall lizards ( Podarcis taurica ), and 20 sand lizards ( Lacerta agilis )—were captured in 2007 and 2008. All ticks removed from the lizards were Ixodes ricinus , either larvae (324/472; 68.6%) or nymphs (148/472; 31.4%). More than half (66/126; 52.4%) of L. viridis individuals were infested, and the prevalence of tick infestation on both the other two species was 35% each. All 472 I. ricinus ticks and tissue samples collected from 134 collar scales and 62 toe clips of lizards were further analyzed for the presence of B. burgdorferi s.l. with polymerase chain reaction. The amplification of B. burgdorferi s.l. DNA was successful in 8% ( n = 92) of L. viridis , 9% ( n = 32) of P. taurica , and 10% ( n = 10) of L. agilis tissue samples. Restriction fragment length polymorphism genotyping identified the species Borrelia lusitaniae in all tested lizard samples. Prevalence of B. burgdorferi s.l. in ticks collected from L. viridis, P. taurica , and L. agilis was 8%, 2%, and 0%, respectively. Most of the infected ticks carried B. lusitaniae (74% of genotyped positives); however, Borrelia afzelii (5%) and B. burgdorferi sensu stricto (21%) were detected in ticks removed from green lizards and Balkan wall lizards, respectively. We conclude that lizards, particularly L. viridis , can be important hosts for I. ricinus larvae and nymphs; thus, they can be regarded as reservoirs of these important pathogen vectors. The role of green lizards has been confirmed, and the implication of Balkan wall lizards is suggested in the natural cycle of B. lusitaniae at our study site.