Evaluating Transmission Paths for Three Different Bartonella spp. in Ixodes ricinus Ticks Using Artificial Feeding

Bartonellae are facultative intracellular alpha-proteobacteria often transmitted by arthropods. Ixodes ricinus is the most important vector for arthropod-borne pathogens in Europe. However, its vector competence for Bartonella spp. is still unclear. This study aimed to experimentally compare its vector competence for three Bartonella species: B. henselae, B. grahamii, and B. schoenbuchensis. A total of 1333 ticks (1021 nymphs and 312 adults) were separated into four groups, one for each pathogen and a negative control group. Ticks were fed artificially with bovine blood spiked with the respective Bartonella species. DNA was extracted from selected ticks to verify Bartonella-infection by PCR. DNA of Bartonella spp. was detected in 34% of nymphs and females after feeding. The best engorgement results were obtained by ticks fed with B. henselae-spiked blood (65.3%) and B. schoenbuchensis (61.6%). Significantly more nymphs fed on infected blood (37.3%) molted into adults compared to the control group (11.4%). Bartonella DNA was found in 22% of eggs laid by previously infected females and in 8.6% of adults molted from infected nymphs. The transovarial and transstadial transmission of bartonellae suggest that I. ricinus could be a potential vector for three bacteria

Diverse tick-borne microorganisms identified in free-living ungulates in Slovakia

Background: Free-living ungulates are hosts of ixodid ticks and reservoirs of tick-borne microorganisms in central Europe and many regions around the world. Tissue samples and engorged ticks were obtained from roe deer, red deer, fallow deer, mouflon, and wild boar hunted in deciduous forests of south-western Slovakia. DNA isolated from these samples was screened for the presence of tick-borne microorganisms by PCR-based methods. Results: Ticks were found to infest all examined ungulate species. The principal infesting tick was Ixodes ricinus, identified on 90.4% of wildlife, and included all developmental stages. Larvae and nymphs of Haemaphysalis concinna were feeding on 9.6% of wildlife. Two specimens of Dermacentor reticulatus were also identified. Ungulates were positive for A. phagocytophilum and Theileria spp. Anaplasma phagocytophilum was found to infect 96.1% of cervids, 88.9% of mouflon, and 28.2% of wild boar, whereas Theileria spp. was detected only in cervids (94.6%). Importantly, a high rate of cervids (89%) showed mixed infections with both these microorganisms. In addition to A. phagocytophilum and Theileria spp., Rickettsia helvetica, R. monacensis, unidentified Rickettsia sp., Coxiella burnetii, "Candidatus Neoehrlichia mikurensis", Borrelia burgdorferi (s.l.) and Babesia venatorum were identified in engorged I. ricinus. Furthermore, A. phagocytophilum, Babesia spp. and Theileria spp. were detected in engorged H. concinna. Analysis of 16S rRNA and groEL gene sequences revealed the presence of five and two A. phagocytophilum variants, respectively, among which sequences identified in wild boar showed identity to the sequence of the causative agent of human granulocytic anaplasmosis (HGA). Phylogenetic analysis of Theileria 18S rRNA gene sequences amplified from cervids and engorged I. ricinus ticks segregated jointly with sequences of T. capreoli isolates into a moderately supported monophyletic clade. Conclusions: The findings indicate that free-living ungulates are reservoirs for A. phagocytophilum and Theileria spp. and engorged ixodid ticks attached to ungulates are good sentinels for the presence of agents of public and veterinary concern. Further analyses of the A. phagocytophilum genetic variants and Theileria species and their associations with vector ticks and free-living ungulates are required.Fil: Kazimírová, Mária. Slovak Academy of Sciences. Institute of Zoology; EslovaquiaFil: Hamšíková, Zuzana. Slovak Academy of Sciences. Institute of Zoology; EslovaquiaFil: Spitalská, Eva. Slovak Academy of Sciences. Institute of Virology. Biomedical Research Center,; EslovaquiaFil: Minichová, Lenka. Slovak Academy of Sciences. Institute of Virology. Biomedical Research Center,; EslovaquiaFil: Mahríková, Lenka. Slovak Academy of Sciences. Institute of Zoology; EslovaquiaFil: Caban, Radoslav. Široká ; EslovaquiaFil: Sprong, Hein. National Institute for Public Health and Environment.Laboratory for Zoonoses and Environmental Microbiology; Países BajosFil: Fonville, Manoj. National Institute for Public Health and Environment.Laboratory for Zoonoses and Environmental Microbiology; Países BajosFil: Schnittger, Leonhard. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Patobiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Kocianová, Elena. Slovak Academy of Sciences. Institute of Virology. Biomedical Research Center,; Eslovaqui

Babesia spp. in ticks and wildlife in different habitat types of Slovakia

Background: Babesiosis is an emerging and potentially zoonotic disease caused by tick-borne piroplasmids of the Babesia genus. New genetic variants of piroplasmids with unknown associations to vectors and hosts are recognized. Data on the occurrence of Babesia spp. in ticks and wildlife widen the knowledge on the geographical distribution and circulation of piroplasmids in natural foci. Questing and rodent-attached ticks, rodents, and birds were screened for the presence of Babesia-specific DNA using molecular methods. Spatial and temporal differences of Babesia spp. prevalence in ticks and rodents from two contrasting habitats of Slovakia with sympatric occurrence of Ixodes ricinus and Haemaphysalis concinna ticks and co-infections of Candidatus N. mikurensis and Anaplasma phagocytophilum were investigated. Results: Babesia spp. were detected in 1.5 % and 6.6 % of questing I. ricinus and H. concinna, respectively. Prevalence of Babesia-infected I. ricinus was higher in a natural than an urban/suburban habitat. Phylogenetic analysis showed that Babesia spp. from I. ricinus clustered with Babesia microti, Babesia venatorum, Babesia canis, Babesia capreoli/Babesia divergens, and Babesia odocoilei. Babesia spp. amplified from H. concinna segregated into two monophyletic clades, designated Babesia sp. 1 (Eurasia) and Babesia sp. 2 (Eurasia), each of which represents a yet undescribed novel species. The prevalence of infection in rodents (with Apodemus flavicollis and Myodes glareolus prevailing) with B. microti was 1.3 % in an urban/suburban and 4.2 % in a natural habitat. The majority of infected rodents (81.3 %) were positive for spleen and blood and the remaining for lungs and/or skin. Rodent-attached I. ricinus (accounting for 96.3 %) and H. concinna were infected with B. microti, B. venatorum, B. capreoli/B. divergens, Babesia sp. 1 (Eurasia), and Babesia sp. 2 (Eurasia). All B. microti and B. venatorum isolates were identical to known zoonotic strains from Europe. Less than 1.0 % of Babesia-positive ticks and rodents carried Candidatus N. mikurensis or A. phagocytophilum.Inst. de PatobiologíaFil: Hamsikova, Zuzana. Slovak Academy of Sciences. Institute of Zoology; EslovaquiaFil: Kazimirová, Mária. Slovak Academy of Sciences. Institute of Zoology; EslovaquiaFil: Harustiakova, Danka. Masaryk University. Faculty of Medicine and Faculty of Science, Institute of Biostatistics and Analyses; República ChecaFil: Mahrikova, Lenka. Slovak Academy of Sciences. Institute of Zoology; EslovaquiaFil: Slovak, Mirko. Slovak Academy of Sciences. Institute of Zoology; EslovaquiaFil: Berthova, Lenka. Slovak Academy of Sciences. Biomedical Research Center. Institute of Virology; EslovaquiaFil: Kocianova, Elena. Slovak Academy of Sciences. Biomedical Research Center. Institute of Virology; EslovaquiaFil: Schnittger, Leonhard. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Patobiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Geographical Distribution and Genetic Diversity of Bank Vole Hepaciviruses in Europe

The development of new diagnostic methods resulted in the discovery of novel hepaciviruses in wild populations of the bank vole (Myodes glareolus, syn. Clethrionomys glareolus). The naturally infected voles demonstrate signs of hepatitis similar to those induced by hepatitis C virus (HCV) in humans. The aim of the present research was to investigate the geographical distribution of bank vole-associated hepaciviruses (BvHVs) and their genetic diversity in Europe. Real-time reverse transcription polymerase chain reaction (RT-qPCR) screening revealed BvHV RNA in 442 out of 1838 (24.0%) bank voles from nine European countries and in one of seven northern red-backed voles (Myodes rutilus, syn. Clethrionomys rutilus). BvHV RNA was not found in any other small mammal species (n = 23) tested here. Phylogenetic and isolation-by-distance analyses confirmed the occurrence of both BvHV species (Hepacivirus F and Hepacivirus J) and their sympatric occurrence at several trapping sites in two countries. The broad geographical distribution of BvHVs across Europe was associated with their presence in bank voles of different evolutionary lineages. The extensive geographical distribution and high levels of genetic diversity of BvHVs, as well as the high population fluctuations of bank voles and occasional commensalism in some parts of Europe warrant future studies on the zoonotic potential of BvHVs.Peer reviewe

Emerging infectious disease implications of invasive mammalian species : the greater white-toothed shrew (Crocidura russula) is associated with a novel serovar of pathogenic Leptospira in Ireland

The greater white-toothed shrew (Crocidura russula) is an invasive mammalian species that was first recorded in Ireland in 2007. It currently occupies an area of approximately 7,600 km2 on the island. C. russula is normally distributed in Northern Africa and Western Europe, and was previously absent from the British Isles. Whilst invasive species can have dramatic and rapid impacts on faunal and floral communities, they may also be carriers of pathogens facilitating disease transmission in potentially naive populations. Pathogenic leptospires are endemic in Ireland and a significant cause of human and animal disease. From 18 trapped C. russula, 3 isolates of Leptospira were cultured. However, typing of these isolates by standard serological reference methods was negative, and suggested an, as yet, unidentified serovar. Sequence analysis of 16S ribosomal RNA and secY indicated that these novel isolates belong to Leptospira alstonii, a unique pathogenic species of which only 7 isolates have been described to date. Earlier isolations were limited geographically to China, Japan and Malaysia, and this leptospiral species had not previously been cultured from mammals. Restriction enzyme analysis (REA) further confirms the novelty of these strains since no similar patterns were observed with a reference database of leptospires. As with other pathogenic Leptospira species, these isolates contain lipL32 and do not grow in the presence of 8-azagunaine; however no evidence of disease was apparent after experimental infection of hamsters. These isolates are genetically related to L. alstonii but have a novel REA pattern; they represent a new serovar which we designate as serovar Room22. This study demonstrates that invasive mammalian species act as bridge vectors of novel zoonotic pathogens such as Leptospira

Survival time of Leptospira kirschneri serovar Grippotyphosa under different environmental conditions.

Leptospirosis is a re-emerging zoonotic disease of high medical importance that affects humans worldwide. Humans or animals acquire an infection with pathogenic leptospires either by direct contact with infected animals or by indirect contact to contaminated environment. Survival of Leptospira spp. in the environment after having been shed via animal urine is thus a key factor to estimate the risk of infection, but not much is known about the tenacity of pathogenic leptospires. Here, the survival time of both a laboratory strain and a field strain of L. kirschneri serovar Grippotyphosa in animal urine and their tenacity while drying was investigated and compared at different temperatures (15°C-37°C). Leptospira spp. are also often found in rivers and ponds. As the infection risk for humans and animals also depends on the spreading and survival of Leptospira spp. in these environments, the survival of L. kirschneri serovar Grippotyphosa was investigated using a 50-meter-long hose system simulating a water stream. Both strains did not survive in undiluted cattle or dog urine. Comparing different temperatures and dilution media, the laboratory strain survived the longest in diluted cattle urine with a slightly alkaline pH value (3 days), whilst the field strain survived in diluted dog urine with a slightly acid pH value up to a maximum of 24 h. Both strains did not survive drying on a solid surface. In a water stream, leptospires were able to move faster or slower than the average velocity of the water due to their intrinsic mobility but were not able to survive the mechanical damage caused by running water in the hose system. From our results we conclude, that once excreted via animal urine, the leptospires immediately need moisture or a water body to survive and stay infectious