Anaplasma phagocytophilum in European bison (Bison bonasus) and their ticks from Lithuania and Poland.

The increasing population of European bison (Bison bonasus) can contribute to the prevalence of zoonotic pathogens. The aim of the present study was to assess the presence of A. phagocytophilum infection in European bison tissues as well as ticks removed from European bison in Lithuania and Poland. A further objective of this work was to compare the detected A. phagocytophilum strains. A total of 85 tissue samples (spleen) of European bison and 560 ticks belonging to two species, Ixodes ricinus (n = 408) and Dermacentor reticulatus (n = 152) were tested. DNA of A. phagocytophilum was detected based on RT-PCR in 40% of the European bison samples, 8.8% of the I. ricinus and 5.9% of the D. reticulatus ticks. Analysis of the obtained partial 16S rRNA gene sequences of A. phagocytophilum revealed the presence of three variants with two polymorphic sites. Furthermore, phylogenetic analysis with partial msp4 gene sequences grouped A. phagocytophilum variants into three clusters. This study revealed that the groEL gene sequences of A. phagocytophilum from European bison and their ticks grouped into ecotype I and only one sequence from Lithuanian European bison belonged to ecotype II. The results of the present study indicated that European bison may play a role as a natural reservoir of A. phagocytophilum.

Molecular Identification and Phylogenetic Analysis of Laelapidae Mites (Acari: Mesostigmata).

The family Laelapidae (Dermanyssoidea) is morphologically and ecologically the most diverse group of Mesostigmata mites. Although molecular genetic data are widely used in taxonomic identification and phylogenetic analysis, most classifications in Mesostigmata mites are based solely on morphological characteristics. In the present study, eight species of mites from the Laelapidae (Dermanyssoidea) family collected from different species of small rodents in Lithuania, Norway, Slovakia, and the Czech Republic were molecularly characterized using the nuclear (28S ribosomal RNA) and mitochondrial (cytochrome oxidase subunit I gene) markers. Obtained molecular data from 113 specimens of mites were used to discriminate between species and investigate the phylogenetic relationships and genetic diversity among Laelapidae mites from six genera. This study provides new molecular data on Laelaps agilis, Laelaps hilaris, Laelaps jettmari, Haemogamasus nidi, Eulaelaps stabularis, Hyperlaelaps microti, Myonyssus gigas, and Hirstionyssus sp. mites collected from different rodent hosts and geographical regions in Europe.

Changes in the Genetic Structure of Lithuania's Wild Boar (Sus scrofa) Population Following the Outbreak of African Swine Fever.

The emergence of African swine fever (ASF) in Lithuania and its subsequent persistence has led to a decline in the population of wild boar (Sus scrofa). ASF has been spreading in Lithuania since its introduction, therefore it is important to understand any genetic impact of ASF outbreaks on wild boar populations. The aim of this study was to assess how the propensity for an outbreak has shaped genetic variation in the wild boar population. A total of 491 wild boar samples were collected and genotyped using 16 STR markers. Allele richness varied between 15 and 51, and all SSR loci revealed a significant deviation from the Hardy-Weinberg equilibrium. Fixation indices indicated a significant reduction in heterozygosity within and between subpopulations. PCoA and STRUCTURE analysis demonstrated genetic differences between the western region which had had no outbreaks (restricted zone I) and the region with ASF infection (restricted zones II and III). It is concluded that environmental factors may play a particular role in shaping the regional gene flow and influence the genetic structure of the wild boar population in the region with ASF outbreaks.

First Molecular Detection of Bartonella bovis and Bartonella schoenbuchensis in European Bison (Bison bonasus).

Bartonella bacteria infect the erythrocytes and endothelial cells of mammalians. The spread of the Bartonella infection occurs mainly via bloodsucking arthropod vectors. Studies on Bartonella infection in European bison, the largest wild ruminant in Europe, are lacking. They are needed to clarify their role in the maintenance and transmission of Bartonella spp. The aim of this study was to investigate the presence of the Bartonella pathogen in European bison and their ticks in Lithuania. A total of 38 spleen samples from bison and 258 ticks belonging to the Ixodes ricinus and Dermacentor reticulatus species were examined. The bison and tick samples were subjected to ssrA, 16S-23S rRNA ITS, gltA, and rpoB partial gene fragment amplification using various variants of PCR. Bartonella DNA was detected in 7.9% of the tissue samples of European bison. All tick samples were negative for Bartonella spp. The phylogenetic analysis of 16S-23S rRNA ITS, gltA, and rpoB partial gene fragment revealed that European bison were infected by B. bovis (2.6%) and B. schoenbuchensis (5.3%). This is the first report addressing the occurrence of B. bovis and B. schoenbuchensis in European bison in Europe.

The effects of habitat fragmentation on the genetic structure of wild boar (Sus scrofa) population in Lithuania.

BACKGROUND: Wild boar (Sus scrofa) is a widely distributed ungulate whose success can be attributed to a variety of ecological features. The genetic variation and population structure of Lithuania's wild boar population have not yet been thoroughly studied. The purposes of this study were to investigate the genetic diversity of S. scrofa and assess the effects of habitat fragmentation on the population structure of wild boar in Lithuania. A total of 96 S. scrofa individuals collected from different regions of Lithuania were genotyped using fifteen microsatellite loci. RESULTS: The microsatellite analysis of the wild boars indicated high levels of genetic diversity within the population. Microsatellite markers showed evidence of a single panmictic wild boar population in Lithuania according to STRUCTURE's highest average likelihood, which was K = 1. This was supported by pairwise Fst values and AMOVA, which indicated no differentiation between the four sampling areas. The results of the Mantel test revealed a weak isolation by distance and geographic diversity gradients that persisted between locations. Motorway fencing and heavy traffic were not an effective barrier to wild boar movement. CONCLUSIONS: There was limited evidence of population genetic structure among the wild boar, supporting the presence of a single population across the study area and indicating that there may be no barriers hindering wild boar dispersal across the landscape. The widespread wild boar population in Lithuania, the high level of genetic variation observed within subpopulations, and the low level of variation identified between subpopulations suggest migration and gene flow between locations. The results of this study should provide valuable information in future for understanding and comparing the detailed structure of wild boar population in Lithuania following the outbreak of African swine fever.

Molecular detection of Babesia spp. in European bison (Bison bonasus) and their ticks.

Babesia spp. are tick-borne haemoparasites that infect a wide range of domestic and wild mammals. Free-ranging ungulates are considered to be important reservoir hosts of Babesia parasites. The European bison (Bison bonasus) is a large and rare ungulate species, reintroduced into the forests of Central Europe after an absence of several decades. Owing to their protected status, studies of tick-borne pathogens in European bison have so far been rare and fragmented. The aim of this study was to investigate the presence of Babesia infection in free-ranging and captive herds of European bison and their ticks. Tissue samples obtained from 37 European bison individuals and 242 ticks belonging to two species, Ixodes ricinus and Dermacentor reticulatus, collected from bison were subjected to PCR analysis of the 18S rRNA gene followed by sequencing. Babesia spp. were detected in 8% of the samples from European bison and in 11% of the ticks. Sequence analysis of partial 18S rRNA gene indicated the presence of B. divergens and B. capreoli in European bison, while B. divergens, B. microti and B. venatorum were detected in ixodid ticks. To the best of authors' knowledge, this is the first molecular detection and characterization of Babesia spp. in European bison and their ticks.

Prevalence and co-infection with tick-borne Anaplasma phagocytophilum and Babesia spp. in red deer (Cervus elaphus) and roe deer (Capreolus capreolus) in Southern Norway.

Anaplasma phagocytophilum and Babesia spp. are causative agents of tick-borne infections that are increasingly considered as a threat to animal and public health. To assess the role of cervids in the maintenance of zoonotic pathogens in Norway, we investigated the prevalence of A. phagocytophilum and Babesia spp. in free-ranging roe deer and red deer. Initial screening of spleen samples of 104 animals by multiplex real-time PCR targeting the major surface protein (msp2) gene and 18S rRNA revealed the presence of A. phagocytophilum infection in 81.1% red deer (Cervus elaphus) and 88.1% roe deer (Capreolus capreolus), and Babesia spp. parasites in 64.9% red deer and 83.6% roe deer, respectively. Co-infections were found in 62.2% red deer and 79.9% roe deer. Nested PCR and sequence analysis of partial msp4 and 18S rRNA genes were performed for molecular characterization of A. phagocytophilum strains and Babesia species. A total of eleven A. phagocytophilum msp4 gene sequence variants were identified: five different variants were 100% identical to corresponding A. phagocytophilum sequences deposited in the GenBank database, while other six sequence variants had unique nucleotide polymorphisms. Sequence analysis of the 18S rRNA gene demonstrated the presence of multiple Babesia species, including Babesia capreoli, Babesia divergens, Babesia venatorum and Babesia odocoilei/Babesia cf. odocoilei. This study is the first report demonstrating the prevalence and molecular characterization of A. phagocytophilum strains and Babesia species in roe deer and red deer in Norway. The high infection and co-infection rates with A. phagocytophilum and Babesia spp. in red deer and roe deer suggest that these cervids may play an important role in the transmission of single and multiple pathogens.

Impact of anthropogenic pressure on the formation of population structure and genetic diversity of raccoon dog Nyctereutes procyonoides.

The raccoon dog Nyctereutes procyonoides experienced an active introduction and acclimatization in the European part of Russia followed by its migration to and colonization in the neighboring countries. Eventually, it has spread rapidly into many European countries. N. procyonoides probably invaded Lithuania from the neighboring countries of Belarus and Latvia where the species was introduced. However, the data on genetic diversity and population structure of the raccoon dogs in the recently invaded territories are still scarce. The objectives of this study were to investigate genetic diversity of N. procyonoides in Lithuania after acclimatization, and to assess the impact of anthropogenic pressure on the formation of population structure. A total of 147 N. procyonoides individuals collected from different regions of Lithuania were genotyped using 17 microsatellite markers. The microsatellite analysis of raccoon dogs indicated high levels of genetic diversity within the population. The Bayesian clustering analysis in STRUCTURE identified 4 genetic clusters among sampled raccoon dogs that could not reveal a clear separation between subpopulations. The widespread distribution of raccoon dogs in Lithuania, high level of genetic variation observed within subpopulations, and low level of variation portioned among subpopulations suggest migration and gene flow among locations. The significant correlation between genetic and geographic distances indicated isolation that reflected the distance between locations. The fencing of highways and very intensive traffic could be barriers to gene flow between the western and eastern sampling areas of raccoon dogs.