Tick-borne diseases are of substantial concern worldwide for animals as well as humans. Dogs have been a human companion for millennia, and their significant impact on human life renders disease in dogs to be of great concern. Tick-borne diseases in dogs represent a substantial diagnostic challenge for veterinarians in that clinical signs are often diffuse and overlapping. In addition, co-infections with two or more pathogens enhance this problem further. Molecular methods are useful to disentangle co-infections and to accurately describe prevalence and geographical distribution of tick-borne diseases. At this point, this information is lacking in many areas worldwide. Romania is one such area, where prevalence and distribution of several important pathogens need to be further investigated. To address this, we screened blood samples from 96 sick dogs with molecular methods for eight different pathogens including Babesia spp., Theileria spp., Hepatozoon spp., Anaplasma spp., Ehrlichia spp., “Candidatus Neoehrlichia mikurensis”, Mycoplasma spp., and Borrelia spp. As many as 45% (43/96) of the dogs in the study were infected with protozoan parasites. Babesia canis was the most frequent of these (28 infected dogs), whereas Hepatozoon canis was detected in 15% (14/96) and Babesia gibsoni was found in a single sample. Bacterial infection with Mycoplasma spp. occurred in 18% (17/96) of the sampled dogs. Obtained bacterial sequences revealed the occurrence of two species: Mycoplasma canis and “Candidatus Mycoplasma haematoparvum”. In several cases co-infection with protozoan parasites and Mycoplasma sp. were detected. All dogs were negative for Anaplasma spp., Ehrlichia spp., “Ca. Neoehrlichia mikurensis”, and for Borrelia spp. The results from the present study reinforce the notion that Babesia canis is an important pathogen in the Romanian dog population. However, more surprisingly, another protozoan species, H. canis, seems to be infecting dogs to a larger extent than previously recognized in Romania. Well-known tick-borne bacterial disease agents such as Anaplasma spp. and Borrelia spp. were not detected. In contrast, less well-studied bacteria such as hemotropic Mycoplasma spp. were detected frequently. Moreover, co-infection might aggravate disease and complicate diagnosis and should be further studied in dogs.

Ticks are transmitting a wide range of bacterial pathogens that cause substantial morbidity and mortality in domestic animals. The full pathogen burden transmitted by tick vectors is incompletely studied in many geographical areas, and extensive studies are required to fully understand the diversity and distribution of pathogens transmitted by ticks. We sampled 824 ticks of 11 species collected in 19 counties in Romania. Ticks were collected mainly from dogs, but also from other domestic and wild animals, and were subjected to molecular screening for pathogens. Rickettsia spp. was the most commonly detected pathogen, occurring in 10.6% (87/824) of ticks. Several species were detected: Rickettsia helvetica, R. raoultii, R. massiliae, R. monacensis, R. slovaca and R. aeschlimannii. A single occurrence of the zoonotic bacterium Bartonella vinsonii berkhoffii was detected in a tick collected from a dog. Anaplasma phagocytophilum occurred in four samples, and sequences similar to Anaplasma marginale/ovis were abundant in ticks from ruminants. In addition, molecular screening showed that ticks from dogs were carrying an Ehrlichia species identical to the HF strain as well as the enigmatic zoonotic pathogen “Candidatus Neoehrlichia mikurensis”. An organism similar to E. chaffeensis or E. muris was detected in an Ixodes ricinus collected from a fox. We describe an abundant diversity of bacterial tick-borne pathogens in ticks collected from animal hosts in Romania, both on the level of species and genotypes/strains within these species. Several findings were novel for Romania, including Bartonella vinsonii subsp. berkhoffii that causes bacteremia and endocarditis in dogs. “Candidatus Neoehrlichia mikurensis” was detected in a tick collected from a dog. Previously, a single case of infection in a dog was diagnosed in Germany. The results warrant further studies on the consequences of tick-borne pathogens in domestic animals in Romania.

Tick-borne encephalitis (TBE) is a severe neurologic disease in Europe and Asia. Disease expression ranges from asymptomatic to severe neurological clinical pictures, involving meningitis, encephalitis, meningoencephalitis and potentially fatal outcome. Humans mostly become infected with TBE virus (TBEV) by the bite of an infected tick. Gastrointestinal (GI) symptoms in humans are mainly attributed to the first viremic phase of TBEV infection with unspecific symptoms and/or resulting from severe neurological impairment of the central nervous system (CNS). We used the subcutaneous TBEV-infection of C57BL/6 mice as a model to analyze GI complications of TBE. We observed the acute distension and segmental dilation of the intestinal tract in 10 of 22 subcutaneously infected mice. Histological analysis revealed an intramural enteric ganglioneuritis in the myenteric and submucosal plexus of the small and large intestine. The numbers of infiltrating macrophages and CD3⁺ T lymphocytes correlated with the severity of ganglioneuritis, indicating an immune-mediated pathogenesis due to TBEV-infection of the enteric plexus. Our study demonstrates that the inflammation of enteric intramural ganglia presents to be a common feature in TBEV-infected mice. Accordingly, the results of this mouse model emphasize that GI disease manifestation and consequences for long-term sequelae should not be neglected for TBEV-infections in humans and require further investigation.

(1) Background: Tick-borne encephalitis (TBE) is the most important tick-borne viral disease in Eurasia, although effective vaccines are available. Caused by the tick-borne encephalitis virus (TBEV, syn. Orthoflavivirus encephalitidis), in Europe, it is transmitted by ticks like Ixodes ricinus and Dermacentor reticulatus. TBEV circulates in natural foci, making it endemic to specific regions, such as southern Germany and northeastern Poland. Our study aimed to identify new TBEV natural foci and genetically characterize strains in ticks in previously nonendemic areas in Eastern Germany and Western Poland. (2) Methods: Ticks were collected from vegetation in areas reported by TBE patients. After identification, ticks were tested for TBEV in pools of a maximum of 10 specimens using real-time RT-PCR. From the positive TBEV samples, E genes were sequenced. (3) Results: Among 8400 ticks from 19 sites, I. ricinus (n = 4784; 56.9%) was predominant, followed by D. reticulatus (n = 3506; 41.7%), Haemaphysalis concinna (n = 108; 1.3%), and I. frontalis (n = 2; <0.1%). TBEV was detected in 19 pools originating in six sites. The phylogenetic analyses revealed that TBEV strains from Germany and Poland clustered with other German strains, as well as those from Finland and Estonia. (4) Conclusions: Although there are still only a few cases are reported from these areas, people spending much time outdoors should consider TBE vaccination.

Tick-borne encephalitis virus (TBEV), like other arthropod-transmitted viruses, depends on specific vectors to complete its enzootic cycle. It has been long known that Ixodes ricinus ticks constitute the main vector for TBEV in Europe. In contrast to the wide distribution of the TBEV vector, the occurrence of TBEV transmission is focal and often restricted to a small parcel of land, whereas surrounding areas with seemingly similar habitat parameters are free of TBEV. Thus, the question arises which factors shape this focal distribution of TBEV in the natural habitat. To shed light on factors driving TBEV-focus formation, we used tick populations from two TBEV-foci in Lower Saxony and two TBEV-foci from Bavaria with their respective virus isolates as a showcase to analyze the impact of specific virus isolate-tick population relationships. Using artificial blood feeding and field-collected nymphal ticks as experimental means, our investigation showed that the probability of getting infected with the synonymous TBEV isolate as compared to the nonsynonymous TBEV isolate was elevated but significantly higher only in one of the four TBEV foci. More obviously, median viral RNA copy numbers were significantly higher in the synonymous virus–tick population pairings. These findings may present a hint for a coevolutionary adaptation of virus and tick populations.

Tick-borne encephalitis (TBE) vaccines are highly effective in preventing TBE and vaccine failures (VF) are rare events. In this study, we compared the age distribution of TBE cases and TBE VF in three endemic countries: Sweden, Southern Germany, and Latvia. While the age distribution of TBE cases was similar for those <50 years versus those ≥50 years in all three countries, in Sweden, a higher proportion of VF cases was ≥50 years, whereas most VF cases in Latvia were <50 years of age and more evenly distributed between those <50 years versus those ≥50 in Southern Germany. Here, theoretical explanations were provided, including differences in diagnostic practices, vaccine uptake between age groups, behavioral patterns and underlying medical conditions, as to why VF were generally older in Sweden than the other countries. There is no scientific rationale to give an extra priming dose of TBE vaccine to subjects ≥50 years of age.