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The most significant processes of arbovirus evolution can be expected to occur in the territories where ticks of different species cohabitate and at the boundaries of virus occurrence, where the probability of the appearance of new virus variants is high due to the possible shift in the main vectors and/or vertebrate hosts. One of the most interesting regions in this regard is the Republic of Tuva. Since most of its territory is covered by mountain ranges and intermountain basins, we were able to study the distribution of vectors and viruses in geographically isolated areas at different altitudes and in various landscapes. From 2008 to 2017, we conducted six expeditions to Tuva and collected 3,077 adult ticks and 24 nymphs. The distribution of tick species was confined to specific landscapes, as follows: Dermacentor nuttalli occurred in steppes, D. silvarum inhabited forest-steppe areas, and Ixodes persulcatus inhabited mixed forests. All three species of ticks were collected on plains and mountain slopes. The range of D. silvarum was shown to be lower than 1300 m above sea level (a.s.l.). Only D. nuttalli and I. persulcatus were collected at higher altitudes. According to our observations, single nymphs of D. nuttalli appear on animals one month before larvae appear. This finding confirms the hypothesis that the immature forms of D. nuttalli are able to overwinter under favourable conditions. We isolated 9 strains and 3 isolates of tick-borne encephalitis virus (TBEV) from I. persulcatus , one strain from D. nuttalli and one strain from D. silvarum. The TBEV strain from D. nuttalli was isolated from the territory inhabited only by Dermacentor ticks. All isolated strains belong to the Siberian subtype of TBEV. TBEV was detected in ticks from all the investigated altitudes. There were no statistically significant differences in the virus prevalence between the Dermacentor and Ixodes ticks. The results of our work provide additional support for the hypothesis of the existence of TBEV foci in areas with an absolute dominance of D. nuttalli. [ABSTRACT FROM AUTHOR]

Abstract Recent discoveries of tick-borne pathogens have raised public health concerns on tick-borne infectious diseases and emphasize the need to assess potential risks of unrecognized tick-borne pathogens. First, to determine the existence of tick-borne phleboviruses (TBPVs), genetic surveillance of phleboviruses in ticks was conducted mainly in Hokkaido, the northernmost island in Japan from 2013 to 2015. Genes of two TBPVs, previously reported as Mukawa virus (MKWV) and a newly identified relative of MKWV, Kuriyama virus (KURV), were detected and the viruses were isolated from Ixodes persulcatus collected in Hokkaido, but not in I. persulcatus collected from other areas of Japan. These viruses were phylogenetically and antigenically similar to each other. Next, to investigate the infection of MKWV in mammals, serum samples from wildlife captured in Hokkaido from 2007 to 2011 were used for serological screening. Neutralizing antibodies against MKWV were detected in both Yezo-deer (Cervus nippon yesoensis) (2/50) and raccoons (Procyon lotor) (16/64). However, no infectious MKWV was recovered from laboratory mice in experimental infections, though viral RNAs were detected in their tissues. Thus, MKWV and KURV may maintain tick-mammalian life cycles in Hokkaido, suggesting their potential as causative agents of tick-borne diseases in mammals. [ABSTRACT FROM AUTHOR]

• Data confirm Ixodes ricinus to be the most common tick in northern Sweden. • Rise in occurrence of Ixodes persulcatus compared with previous reports. • Ticks collected from hosts by the public over a large geographical area. • Updated knowledge on tick distribution is vital for the creation of risk maps. Environmental and climatic changes in northern Europe have shaped a geographical area in which new tick species may become established and introduce new tick-borne pathogens. In recent decades, ticks have expanded their latitudinal and altitudinal range limits in northern Sweden. In this study, ticks were collected in 2018 and 2019 in northern Sweden from different hosts, mainly from dogs, cats and humans. The ticks in 2018 (n = 2141, collected from 65 municipalities in 11 provinces) were identified as Ixodes ricinus (n = 2108, 98.5%), Ixodes persulcatus (n = 18, 0.8%), Ixodes trianguliceps (n = 14, 0.7%) and Hyalomma marginatum (n = 1, 0.05%). The ticks collected in 2019 (n = 519, across a smaller area than in 2018, i.e. Sweden's four northernmost provinces) were identified as I. ricinus (n = 242, 46.6%) and I. persulcatus (n = 277, 53.4%). Among those collected in 2019, the majority of I. ricinus (n = 111, 45.9%) were submitted from the province of Västerbotten, while most I. persulcatus (n = 259, 93.5%) were collected in the province of Norrbotten. This study provides updated figures on the geographical distribution of two Ixodes species in northern Sweden. The results confirmed I. ricinus to be the dominant species and that I. persulcatus has enlarged its distributional area compared with previous reports. Updated knowledge of tick distribution is fundamental for the creation of risk maps and will allow relevant advice to be provided to the general public, suggesting measures to prevent tick bites and consequently tick-borne diseases. [ABSTRACT FROM AUTHOR]

Borrelia miyamotoi , recently recognized as a human pathogenic spirochete, was isolated from Ixodes persulcatus and I. ovatus in northern Mongolia and Honshu Island, a major island in Japan. Although no human B. miyamotoi infections have been reported in Mongolia, the prevalence of B. miyamotoi in ticks from Mongolia is higher than that in ticks from Hokkaido, Japan, where human cases have been reported. Moreover, the multi-locus sequence analysis of cultured isolates revealed that B. miyamotoi isolates in Mongolia belong to the Siberian type, a sequence type that was originally reported from isolates from I. persulcatus in Hokkaido. Thus, there is a possibility of unrecognized human B. miyamotoi infections in Mongolia. Moreover our data support the hypothesis of clonal expansion of the Siberian type B. miyamotoi . In contrast, although the isolates were found to belong to the Siberian type B. miyamotoi , two isolates from I. persulcatus in Honshu Island were identified to be of a different sequence type. Furthermore, B. miyamotoi isolates from I. ovatus were distinguishable from those from I. ricinus complex ticks, according to genetic analysis. In this study, we show that there may be some genetic diversity among B. miyamotoi in ticks from Honshu Island. [ABSTRACT FROM AUTHOR]

Prostriate ticks (subfamily Ixodinae, genus Ixodes ) can copulate and the females can be inseminated before attachment to the host. In tests with Ixodes persulcatus females collected in the field and fed without males on the host, it was shown that this preprandial insemination is necessary and sufficient for successful engorgement and oviposition if female feeding took place in up to 1 month after collection. A 2-month period between preprandial insemination and female feeding was followed by a significant decrease in the proportion of normally engorged females and significant increase in egg mortality. If a small number of males were added to feeding females in this case, the number of normally engorged females increased but the egg mortality remained as high. Spermatophore destruction during the 2-month period is assumed to have a negative effect on the viability of eggs produced after additional (perprandial) insemination. Prostriate ticks are believed to be an intermediate group between argasid and metastriate ticks. Transition from nidicolous parasitism in argasid ticks to exophily (pasture parasitism) in metastriate ticks determines the change in mating strategy from off-host to on-host copulation. We review the available data concerning mating strategies in representatives of different subgenera of the genus Ixodes in the context of this evolutionary relationship. [ABSTRACT FROM AUTHOR]

Tick-borne encephalitis virus is the etiological agent of a severe human disease transmitted by hard ticks. It occurs in large parts of eastern, central, and western Asia and in Europe with thousands of human cases each year. Here, the discovery of the virus by Soviet scientists in the late 1930s in the Far East is described. The pioneering work involved with this discovery, which resulted in great scientific and epidemiological achievement, was undertaken under the most difficult conditions, and some of the scientists and their technical assistants paid for it with their health and even their lives. This paper briefly outlines the steps on the way that elucidated the basic etiology and eco-epidemiology of the disease, and does not omit that, as one result of the expeditions and the political situation in the former Soviet Union at that time, some scientists were sent to prison. [ABSTRACT FROM AUTHOR]

The number of tick-borne infections in the northern European regions of Russia has increased considerably in the last years. In the present study, 676 unfed adult Ixodes persulcatus ticks were collected in the Komi Republic from 2011 to 2013 to study tick-borne rickettsioses. Rickettsia spp. DNA was detected by PCR in 51 (7.6%) ticks. The nucleotide sequence analysis of gltA fragments (765 bp) from 51 ticks indicated that 60.8% and 39.2% of the ticks were infected with Rickettsia helvetica and Candidatus R. tarasevichiae, respectively. The gltA fragments showed 100% identity with those of Candidatus R. tarasevichiae previously discovered in Siberia and China, whereas R. helvetica showed 99.9% sequence identity with European isolates. The ompB had 8 nucleotide substitutions, 6 of which resulted in amino acid substitutions. In the sca9 gene, 3 nucleotide substitutions were detected, and only one resulted in amino acid substitution. The smpA, ompW , and β-lactamase genes of R. helvetica also showed a high level of sequence identity. [ABSTRACT FROM AUTHOR]

Cystatins are cysteine peptidase inhibitors that in ticks mediate processes such as blood feeding and digestion. The ixodid tick Ixodes persulcatus is endemic to the Eurasia, where it is the principal vector of Lyme borreliosis. To date, no I. persulcatus cystatin has been characterized. In the present work, we describe three novel cystatins from I. persulcatus , named JpIpcys2a, JpIpcys2b and JpIpcys2c. In addition, the potential of tick cystatins as cross-protective antigens was evaluated by vaccination of hamsters using BrBmcys2c, a cystatin from Rhipicephalus microplus , against I. persulcatus infestation. Sequence analysis showed that motifs that are characteristic of cystatins type 2 are fully conserved in JpIpcys2b, while mutations are present in both JpIpcys2a and JpIpcys2c. Protein-protein docking simulations further revealed that JpIpcys2a, JpIpcys2b and JpIpcys2c showed conserved binding sites to human cathepsins L, all of them covering the active site cleft. Cystatin transcripts were detected in different I. persulcatus tissues and instars, showing their ubiquitous expression during I. persulcatus development. Serological analysis showed that although hamsters immunized with BrBmcys2c developed a humoral immune response, this response was not adequate to protect against a heterologous challenge with I. persulcatus adult ticks. The lack of cross-protection provided by BrBmcys2c immunization is perhaps linked to the fact that cystatins cluster into multigene protein families that are expressed differentially and exhibit functional redundancy. How to target such small proteins that are secreted in low quantities remains a challenge in the development of suitable anti-tick vaccine antigens. [ABSTRACT FROM AUTHOR]

In order to investigate the diversity of spotted fever group (SFG) Rickettsia infection in hard ticks, ticks were harvested from the forest areas in Suifenhe city, along the Chinese–Russian border and conventional PCR was carried out using universal SFG Rickettsia primers targeting glt A and omp A genes to screen for their infection with SFG Rickettsia organisms. Results showed that of the 215 ticks belonging to Ixodes persulcatus , Haemaphysalis concinna and Haemaphysalis japonica Warburton, 1908 species, 138 (64.2%) were positive for SFG Rickettsia . Three species of SFG Rickettsia were detected, Rickettsia raoultii , Rickettsia heilongjiangensis and Candidatus Rickettsia tarasevichiae . No co-infection with different species of SFG Rickettsia was found in any individual tick among the three tick species. We detected more than one SFG Rickettsia species in ticks from each of the three tick species with an overlapping distribution and potentially similar transmission cycles of SFG Rickettsia in the areas surveyed. Consequently, different pathogenic rickettsial species may be involved in human cases of rickettsiosis after a bite of the three above-mentioned tick species in that area Rickettsia [ABSTRACT FROM AUTHOR]

Information about the prevalence and geographical distribution of tick-borne pathogens Anaplasma phagocytophilum , Candidatus Neoehrlichia mikurensis, and Babesia spp. is still rare in Mongolia. We tested 275 Ixodes persulcatus ticks for A. phagocytophilum , Cand . N. mikurensis and Babesia spp. and 125 Dermacentor nuttalli ticks especially for Babesia spp. using different PCR methods. Ticks were collected from three provinces (Selenge, Arkhangai, Khentii) in Mongolia. DNA of A. phagocytophilum , Cand . N. mikurensis and Babesia spp. were found with a prevalence of 6.2%, 1.5% and 3.3% in each case in I. persulcatus ticks. This is the first time Cand . N. mikurensis was found in ticks from Mongolia. Sequence analysis of Babesia spp.-positive amplicons showed exclusively B. venatorum , which had also not been mentioned in Mongolia before. On the contrary, all D. nuttalli ticks tested negatively for Babesia spp. This study demonstrates that all three zoonotic pathogens are present in I. persulcatus ticks in Mongolia, and justify the need for further investigations of a more detailed genetic characterization of these pathogens. [ABSTRACT FROM AUTHOR]

Adult Ixodes persulcatus were collected in highly populated districts in Irkutsk city, Russia, and in popular recreational and professional areas in its neighboring territories. Borrelia miyamotoi infection in I. persulcatus was examined using multiplex Taqman-PCR targeting 16S rDNA, and nested PCR and sequencing analyses targeting flaB and 16S rDNA. B. miyamotoi and Lyme disease Borrelia species were detected in 13 (infection rate, 2.9%) and 77 (17.3%) out of 445 I. persulcatus ticks, respectively, collected from 4 sites around the Baikal Lake. The 16S rDNA and flaB sequences of these amplicons were closely related to those of B. miyamotoi detected and/or isolated from I. persulcatus in Japan and Far Eastern Russia, and clustered separately from those of Europe and North America. These results indicate that additional surveillance for B. miyamotoi infection is needed in order to determine how it affects human health in Irkutsk City and its neighboring territories. [ABSTRACT FROM AUTHOR]

The recently shown phenomenon of natural hybridization between Ixodes persulcatus and Ixodes pavlovskyi ticks ( Kovalev et al., 2015 ) stimulated similar studies in the sympatric zones of other tick species. In the present paper, 265 Ixodes ricinus and I. persulcatus ticks from Estonia were subjected to a search for interspecific hybrids based on nuclear (ITS2) and mitochondrial (cox1) markers as well as morphological features. Surprisingly, only 72.1% of ticks morphologically identified as I. ricinus actually were I. ricinus both at nuclear and mitochondrial markers, while the accuracy of morphological species identification for I. persulcatus was 99.3%. Among ticks morphologically identified as I. ricinus , 24.6% turned out to be interspecific hybrids and 3.3% were I. persulcatus . Generally, about 11% of the individuals studied were shown to be interspecific hybrids with different levels of nuclear DNA introgression. The analysis of hybrid populations proved the mating pair female I. ricinus × male I. persulcatus to form hybrids more efficiently, then female I. persulcatus × male I. ricinus . The same trend can be observed for backcrosses preferentially mating with I. ricinus . Hybridization between I. ricinus and I. persulcatus proved the existing view about their reproductive isolation to be untenable. Interspecific hybridization occurring between both closely ( I. persulcatus and I. pavlovskyi ) and more distantly ( I. ricinus and I. persulcatus ) related Ixodes species could introduce novel alleles that modify vector competence, host use or the ability to exploit diverse microhabitats. [ABSTRACT FROM AUTHOR]

Ixodes ricinus immunosuppressor (Iris) is a tick salivary gland protein derived from I. ricinus . In this study, Iris homolog was identified in the salivary glands of Ixodes persulcatus , which is the specific vector of the Lyme disease agent in Japan. The homolog was named Ipis-1. To investigate the function of Ipis-1, we prepared a recombinant Ipis-1 expressed in COS-7 cells as a rabbit IgG Fc-fused protein (Ipis-1-Ig). Cell proliferation assay and IFN-γ ELISA showed that Ipis-1-Ig inhibits the proliferation and IFN-γ production of bovine peripheral blood mononuclear cells (PBMCs). Notably, Ipis-1-Ig inhibited the cell proliferation and production of IFN-γ in bovine PBMCs even when CD14 + cells were depleted, suggesting that Ipis could directly interact with T cells and inhibit their functions. In conclusion, Ipis could contribute to the establishment of environments suitable for tick blood feeding and pathogen transmission by suppressing the function of immune cells. [ABSTRACT FROM AUTHOR]

Ticks of the Ixodes ricinus group are important vectors of human pathogens in both Eurasia and North America; therefore, many studies have focused on their molecular systematics and evolutionary relationships. However, there are species that have not been characterized by molecular genetic methods so far. For the first time, we obtained nucleotide sequences of two nuclear and three mitochondrial genetic markers from four museum specimen of I. kazakstani Olenev et Sorokoumov, 1934, collected in Kyrgyzstan. The phylogenetic analysis showed that I. kazakstani undoubtedly belongs to the I. ricinus group but is not closely related to I. persulcatus, as was expected. Further studies of the genetic features of I. kazakstani, presumably an intermediate between Nearctic and Palearctic species, would elucidate the evolutionary relationships between tick species within the I. ricinus group. [ABSTRACT FROM AUTHOR]

• New Borrelia sp. was found in Ixodes spp. and samples from small mammals. • MLST suggests new Borrelia as a putative species " Candidatus B. sibirica". • Borrelia bavariensis was first found in molted I. trianguliceps and I. apronophorus. • The same B.bavariensis were found in I.persulcatus, I.trianguliceps and I.apronophorus. Four genospecies from the Borrelia burgdorferi sensu lato complex were detected in Ixodes persulcatus and Ixodes pavlovskyi ticks from Siberia and genetically characterized. The presence of Borrelia spp. in Ixodes apronophorus and Ixodes trianguliceps ticks found in Asia has never been studied. In this study, genetic diversity of B. burgdorferi s.l. was investigated in three I. persulcatus / I. trianguliceps / I. apronophorus sympatric habitats in Western Siberia. Three groups of samples were examined: (i) ticks that were taken from rodents and molted in a laboratory; (ii) non-molted ticks collected from rodents; (iii) specimens from small mammals. Expectedly, Borrelia afzelii and Borrelia bavariensis were detected in I. persulcatus and in small mammals from the studied locations. Borrelia bavariensis was first found in molted I. apronophorus and I. trianguliceps. Identical genovariants of B. bavariensis were found in I. apronophorus, I. trianguliceps , and I. persulcatus. In addition, a new Borrelia genovariant was discovered in non-molted and molted I. apronophorus and non-molted I. persulcatus and I. trianguliceps , as well as in small mammals. This new genovariant was genetically characterized using MLST and single locus sequence analysis, which indicated that the new Borrelia genovariant significantly differs from all known Borrelia species. We propose the name " Candidatus Borrelia sibirica" for this putative new species. [ABSTRACT FROM AUTHOR]

Borrelia bavariensis is a recently described agent of Lyme disease within the B. burgdorferi sensu lato species complex and exhibits a strong capacity for human pathogenicity. B. bavariensis strains are widely distributed in Eurasia spanning the distribution range of the tick vectors Ixodes persulcatus and I. ricinus . It has been suggested that B. bavariensis forms two populations, one of which arose through vector adaptation and geographic expansion. We have performed phylogenetic and population genetic analyses with next-generation sequencing data of 26 strains of B. bavariensis targeting the main linear chromosome and two plasmids (lp54, cp26). A very low number of single nucleotide polymorphisms (SNPs) was found in the European population and a deep branching pattern between European and Asian B. bavariensis was observed in all phylogenies. The results confirm the population structure of B. bavariensis and strongly support the hypothesis of clonal expansion of the European population of B. bavariensis . In addition, signals of positive selection identified in the populations further support the hypothesis that the European population of B. bavariensis likely underwent vector adaptation in its recent evolutionary history. Identified genes represent promising candidates for experimental vector adaptation studies. Thus, this species forms a very good model to study vector adaptation, which is known to play an important role in the geographic distribution of B. burgdorferi. Analysis of well known virulence determinants that are attributed to severity of clinical manifestation in B. burgdorferi s.s. revealed no variation within the European population of B. bavariensis , underlining the importance of including various Borrelia species into investigations that aim to understand the pathogenesis of Lyme disease agents. [ABSTRACT FROM AUTHOR]

The objective of the study was detection of hybrid larvae in Ixodes persulcatus and Ixodes ricinus cohabitation sites. To this end, the following three tasks were solved: interspecies crossing of ticks, evaluation of typical morphological signs of the hybrid larvae, and analysis of collected specimens from sites of sympatry. Under experimental conditions, hybrid larvae of I. persulcatus (female) and I. ricinus (male) were obtained that differed from the parental species by the size of setae on the scutum and alloscutum. Discriminant analysis yielded 87.5% classification accuracy for the priory set groups of I. persulcatus , I. ricinus , and hybrids. Of 88 hybrid larvae, 13 (15%) were classified as I. persulcatus and 4 (5%) as I. ricinus . We measured larvae of Ixodes ticks ( n = 141) collected from small mammals in 1950–1970 in Karelia in cohabitation sites of these species that were previously classified as I. persulcatus or I. ricinus . According to the results of discriminant analysis, 31 larvae (22%) were classified as hybrids with probability p ≥ 0.52; for 10 larvae (7%), the probability of placement to the hybrid group was >0.95. [ABSTRACT FROM AUTHOR]

Anaplasma phagocytophilum is a causative agent of granulocytic anaplasmosis in different mammals. The presence of A. phagocytophilum was assayed in Ixodes persulcatus, Ixodes trianguliceps ticks and Myodes spp. voles from two I. persulcatus / I. trianguliceps sympatric areas in the Omsk region (Western Siberia, Russia). In total, A. phagocytophilum was found in 42/108 (38.9%) of vole blood samples, 13/34 (38.2%) of I. trianguliceps ticks removed from voles, 1/12 (8.3%) of I. persulcatus removed from voles, and 18/279 (7.2%) of questing I. persulcatus . GroESL operon sequence analysis of positive samples revealed three distinct A. phagocytophilum genetic groups previously identified in ticks and mammals in Russia. Genetic group 1 was found in 6/36 (16.7%) of sequenced positive blood samples; this group was previously revealed in I. persulcatus and Myodes spp. voles in different regions of Russia. Genetic group 2 was found in 30/36 (83.3%) of sequenced positive blood samples and all positive I. trianguliceps ; this group was previously revealed only in Myodes spp. voles and common shrews ( Sorex araneus ) in I. persulcatus / I. trianguliceps sympatric areas in the Northern Ural. Genetic group 3 was found in all positive questing I. persulcatus and one blood sample; this group was previously revealed in I. persulcatus and Siberian chipmunks ( Tamias sibiricus ). We suppose that I. trianguliceps is the most probable vector for A. phagocytophilum of group 2. Analysis of the msp4 gene, intergenic region DOV1, and some other genetic loci has shown that isolates from different genetic groups significantly differ in all studied loci and that A. phagocytophilum of group 2 is closely related to A. phagocytophilum isolates revealed in voles and I. trianguliceps in Europe. A. phagocytophilum of groups 1 and 2 are the most similar to each other, while A. phagocytophilum of group 3 clusters with European A. phagocytophilum isolates from I. ricinus and various mammalian species. [ABSTRACT FROM AUTHOR]

The tick Ixodes persulcatus is the predominant tick species in Northeastern China, and it is a major vector in transmission of tick-borne diseases. By 16S rRNA Illumina sequencing, we investigated the microbiome of I. persulcatus and assessed the variation of the microbiome before and after blood feeding. The prolonged blood meal dramatically altered the composition of the microbiome but did not influence the bacterial diversity. Overall, 373 and 289 bacterial genera were assigned to unfed and fed ticks, respectively. To investigate microbes that were potentially transmitted to vertebrate hosts during a blood meal, we examined the microbiome in rat blood after tick bites. Our data showed that 237 bacterial genera were suspected to be pathogens of vertebrates because they were commonly detected in unfed ticks, fed ticks, and rat blood samples after tick bites. Additionally, the prevalence survey on Borrelia burgdorferi s.l. , Ehrlichia chaffeensis , Anaplasma phagocytophilum and Yersinia pestis was performed. We found that B. garinii and B. afzelii are the predominant genospecies of the Lyme disease spirochete in I. persulcatus ticks. This is the first time that the microbial composition in this tick species and in rat blood transmitted via tick bites has been reported. These data may ultimately assist in identification of novel pathogens transmitted by I. persulcatus ticks. [ABSTRACT FROM AUTHOR]

Abstract: To study the role of wild birds in the transmission of tick borne encephalitis virus (TBEV), we investigated randomly captured wild birds bearing ixodid ticks in a very highly endemic TBE region located in Tomsk city and its suburbs in the south of Western Siberia, Russia. The 779 wild birds representing 60 species were captured carrying a total of 841 ticks, Ixodes pavlovskyi Pom., 1946 (n =531), Ixodes persulcatus P. Sch., 1930 (n =244), and Ixodes plumbeus Leach. 1815 (n =66). The highest average number of ticks per bird in a particular species was found for the fieldfare (Turdus pilaris Linnaeus, 1758) (5.60 ticks/bird) and the tree pipit (Anthus trivialis Linnaeus, 1758) (13.25 ticks/bird). Samples from wild birds and ticks collected in highly endemic periods from 2006 to 2011 were tested for the TBEV markers using monoclonal modified enzyme immunoassay (EIA) and RT-PCR. TBEV RNA and antigen were found in 9.7% and 22.8% samples collected from wild birds, respectively. TBEV markers were also detected in 14.1% I. persulcatus ticks, 5.2% I. pavlovskyi, and 4.2% I. plumbeus ticks collected from wild birds. Two TBEV strains were also isolated on PKE (pig kidney embryo) cells from fieldfare and Blyth's reed warbler (Acrocephalus dumetorum Blyth, 1849). Sequencing of 5′-NCR of TBEV revealed that all TBEV isolates belong to Far Eastern (dominate) and Siberian genotypes. Several phylogenetic subgroups included TBEV sequences novel for the Tomsk region. Our data suggest that wild birds are potential disseminators of TBEV, TBEV-infected ixodid ticks, and possibly other tick-borne infections. [Copyright &y& Elsevier]

Borrelia miyamotoi infection is an emerging tick-borne disease that causes hard tick-borne relapsing fever. B. miyamotoi is transmitted through the bite of ticks, including Ixodes persulcatus. Although accumulating evidence suggests that tick salivary proteins enhance the infectivity of other tick-borne pathogens, the association of B. miyamotoi with tick-derived proteins remains unknown. In this study, the effect of I. persulcatus sialostatin L2 (Ip-sL2), a tick-derived cystatin, on specific immunity to B. miyamotoi was preliminarily investigated in vitro. Mice were immunized with heat-killed B. miyamotoi and in vitro analyses of the splenocytes of the immunized mice indicated that the expression levels of the activation markers of CD11c+ and CD3+ cells were significantly upregulated by B. miyamotoi stimulation. Spleen cells from B. miyamotoi -immunized mice were used to determine whether Ip-sL2 regulates murine immune responses against B. miyamotoi. Treatment with Ip-sL2 in vitro inhibited the activation of CD11c+ and CD3+ cells as well as inflammatory cytokine production by cultured splenocytes. These findings show that Ip-sL2 has modulatory effects on murine immune responses to B. miyamotoi. Therefore, it is necessary to clarify in the future whether Ip-sL2 is involved in the enhanced infectivity of B. miyamotoi. [ABSTRACT FROM AUTHOR]

Abstract: In Mongolia, Lyme borreliosis was first reported in 2003. To determine which Borrelia species may contribute to the occurrence of Lyme borreliosis in Mongolia, real-time PCR was conducted on 372 adult Ixodes persulcatus ticks collected in Selenge Aimag, the province with the highest incidence of human Lyme borreliosis. 24.5% of ticks were identified to be positive for Borrelia burgdorferi sensu lato DNA. Species differentiation using an SNP-based real-time PCR and multi-locus sequence analysis revealed that strains phylogenetically closely related to B. bavariensis (previously known as B. garinii OspA serotype 4) is the most prevalent species, showing an unexpectedly high genetic diversity. [Copyright &y& Elsevier]

Abstract: The northern boundary of the sympatric zone of Ixodes persulcatus and I. ricinus (Acari: Ixodidae) passes through Karelia. Studies carried out in the 1950s showed that these Ixodes species were mostly found in southern Karelia. I. ricinus inhabited the west of the region, I. persulcatus the east, with a zone of sympatry in the centre. Here, we describe the present distribution of these species in southern Karelia and provide a retrospective assessment of potential changes in the sympatric zone. Tick distribution and abundance were investigated during transect samplings, conducted in May–June 2006–2010. Additional information was obtained during examination of pet dogs and cats. Overall, 4561 adult ticks were collected. Since the 1950s, there has been a significant increase in the abundance of I. persulcatus, and a decrease in I. ricinus. Currently, southern Karelia can be considered as a zone of sympatry for I. ricinus and I. persulcatus, without a clear geographic boundary between the 2 species. In most areas, except to the west of Lake Ladoga, I. persulcatus is more abundant. Possible reasons for this trend are discussed. [Copyright &y& Elsevier]

Abstract: Since the year 2005, clinical patterns resembling tick-borne rickettsioses have been noticed in Mongolia. Epidemiological data regarding species of the aetiological agent, tick vector, prevalence, and distribution as well as incidence of human cases throughout Mongolia are still sparse to date. In order to identify Rickettsia species occurring in Mongolia, we investigated Dermacentor nuttalli (n =179) and Ixodes persulcatus (n =374) collected in 4 selected provinces. Rickettsia raoultii was the predominant Rickettsia (82% prevalence) found in D. nuttalli and was also detected in I. persulcatus (0.8%). The Rickettsia prevalence in D. nuttalli from different provinces varied between 70% and 97%. In addition, R. sibirica was identified in approximately 4% of D. nuttalli, but solely from Arkhanghai province. The results of this study extend the common knowledge about the geographic distribution of R. raoultii and its high prevalence in D. nuttalli. Although the pathogenicity of this Rickettsia is still unclear, it should be considered in Mongolian patients suspected of having tick-borne rickettsiosis. [Copyright &y& Elsevier]

Abstract: The tick Ixodes persulcatus Schulze, 1930, has a wide distribution from the Baltic to the Far East and is a vector of a number of human pathogens. Thus, the study of the genetic structure and evolution of this species is of great epidemiological importance. rRNA genes were used as genetic markers to identify the phylogeographical structure of the ticks. The sequences of gene fragments of 28S (expansion segment D3) and mitochondrial 12S rRNA for 25 and 76 ticks, respectively, that had been collected in various regions of Russia in 2007–2011, were obtained. The sequences of the 28S rRNA D3 segment were identical for all ticks within the studied area. Analysis of the sequences of the mitochondrial 12S rRNA fragment revealed 4 haplotypes with one occurring at a frequency of 0.96. It is shown that the ‘deep’ population structure of I. persulcatus () was erroneous because of the inclusion of contaminating fungi sequences of 28S rRNA in the phylogenetic analysis. This was, possibly, due to the use of universal PCR primers that amplify the DNA of a wide range of eukaryotes, particularly of fungi which are common in samples of ticks. The influence of PCR conditions on the preferential amplification of the DNA of different organisms is also demonstrated. [Copyright &y& Elsevier]

High-throughput sequencing (HTS) technology has profoundly been involved in sequencing whole genomes of several organisms in a fast and cost-effective manner. Although HTS provides an alternative biomonitoring method to the time-consuming and taxonomy-expertise dependent morphological approach, still we cannot rule out the possibility of the impediment and misidentification biases. In this article we aim to retrieve whole mitochondrial genome (mitogenome) sequences from publicly available raw sequencing data for phylogenetic comparison of Ixodes persulcatus. For this comparison, we sequenced whole mitogenomes of four I. persulcatus ticks from Japan and constructed mitogenomes from raw sequencing data of 74 I. persulcatus ticks from China. Bayesian phylogenetic trees were inferred by the concatenated fifteen mitochondrial genes. We further tested our results by the phylogenetic analysis of cytochrome c oxidase subunit 1 (cox1) gene and internal transcribed spacer 2 (ITS2) sequences. Our findings showed that 70 constructed mitogenomes from China were clustered with the sequenced four mitogenomes of I. persulcatus from Japan. We also revealed that mitogenome sequences retrieved from two data sets CRR142297 and CRR142298 were clustered with Ixodes nipponensis. Moreover, other two mitogenome sequences from CRR142310 and CRR142311 formed a clade with Ixodes pavlovskyi. The phylogenetic analysis of cox1 gene and ITS2 sequences confirmed the identification errors of these four samples. The overall phylogenetics in our study concluded that accurate morphological identification is necessary before implementing HTS to avoid any misidentification biases. [ABSTRACT FROM AUTHOR]

Francisella tularensis subsp. holarctica is the causative agent of tularaemia in Europe. Finland is a high-incidence region for tularaemia, with mosquito bites as the most common sources of infection. However, in Central and Western Europe, ticks (Acari: Ixodidae) have been suggested as the main vectors. Indeed, several studies have reported the pathogen from the locally most common human-biting tick species, Ixodes ricinus. In Finland, the occurrence of the pathogen in ticks has started receiving attention only recently. Here, we collate previous tick screening data from Finland regarding F. tularensis as well as present the results from a novel screening of roughly 15 000 I. ricinus and I. persulcatus collected from across the country. In total, 14 878 ticks collected between 2015 and 2020 were screened for F. tularensis using a TaqMan-based qPCR assay targeting the 23 KDa gene. The combined screening efforts of the current and previous studies, encompassing roughly 20 000 ticks, did not find any positive ticks. Given the negative results despite the considerable sample size, it appears that the pathogen is not circulating in local tick populations in Finland. We discuss some possible reasons for the lack of the bacterium in ticks in this high-incidence region of tularaemia. [ABSTRACT FROM AUTHOR]

Kemerovo group viruses are tick-transmitted members of Orbivirus genus of the Reoviridae family that can cause infections of the central nervous system of humans. In this work, Kemerovo virus (KEMV) RNA was detected for the first time in Novosibirsk region of Western Siberia, Russia, in Ixodes pavlovskyi and Ixodes persulcatus ticks. [ABSTRACT FROM AUTHOR]

Borreliosis is one of the most common vector-borne zoonotic diseases in the world. Limited data are available regarding Borrelia spp. and their genotypes in Kazakhstan. The goal of this study was to investigate the prevalence of Borrelia spp. in ixodid ticks collected in the southeastern region of Kazakhstan. A total of 1907 ixodid ticks were collected by flagging vegetation at three collection areas in the Almaty oblast between 2015 and 2018. They were grouped into 407 pools and examined by qPCR for Borrelia burgdorferi sensu lato (s.l.). A conventional PCR with specific primers targeting 16S rRNA gene was used to differentiate B. burgdorferi s.l. genospecies. Sequence analysis of the PCR products was performed for sixteen samples. Lyme borreliosis agents were only detected in adult questing Ixodes persulcatus. The overall B. burgdorferi s.l. prevalence in I. persulcatus estimated as the minimum infection rate reached 10.7 %. Borrelia burgdorferi sensu stricto was not detected in any of the tick pools. Partial 16S rRNA gene sequencing revealed the presence of B. miyamotoi , B. afzelii, and B. garinii. Borrelia afzelii was the dominant genospecies in Almaty oblast. A significantly lower proportion of B. garinii positive tick pools was detected in the Zailiyskiy Alatau as compared to the Dzungarian Alatau (χ2 = 16.243; p = 0.0001) and Yenbekshikazakh district (χ2 = 7.4156; p = 0.0065). The obtained results indicate the epidemiological significance of B. afzelii and B. garinii in southeastern Kazakhstan. These new data aim to improve the diagnostics of Lyme borreliosis and monitoring of tick-borne infections in Kazakhstan. [ABSTRACT FROM AUTHOR]

Abstract: Ixodes persulcatus Schulze, 1930 (I. persulcatus) is distributed from the western to the eastern borders of Russia where it is found in the taiga and in mixed forests of the European type. This tick is a known vector of viral (tick-borne encephalitis virus), spirochetal (Borrelia spp.), and protozoan (Babesia spp.) agents. In this work, we analyzed the phylogenetic position of I. persulcatus within the Prostriata based on its 18S rRNA gene. Furthermore, we review the relationship of this tick with 9 Alphaproteobacteria from the genera Rickettsia, Ehrlichia, and Anaplasma which are found in I. persulcatus in the territory of Russia. These observations suggest that I. persulcatus may be one of the more important vectors and reservoirs of Alphaproteobacteria and other microorganisms in Russia. [Copyright &y& Elsevier]

• Methionine aminopeptidase of Ixodes persulcatus (Ip MetAP) was cloned and identified. • Ip MetAP1b has high sequence and structure homology with MetAPs from other species. • The enzyme has preference for N-terminal methionine and Mn2+ and Co2+ as cofactors. • Ip MetAP1b has optimal activity at physiological pH and temperature. • The enzyme is sensitive to aminopeptidase inhibitors such as bestatin. Methionine aminopeptidases (MetAPs), which remove the initiator methionine from nascent peptides, are essential in all organisms and considered to be a valuable targets for the treatment of various diseases, including cancer, malaria, and bacterial infections. However, MetAPs have not been reported in hard ticks (family Ixodidae), and their bioinformatics characterisation in tick's genome sequences is limited. In this study, we cloned, identified, and characterised a novel MetAP from Ixodes persulcatus , a vector for pathogens causing Lyme borreliosis and tick-borne encephalitis. The sequence analysis showed that I. persulcatus MetAP was a type 1 enzyme carrying C-terminal motifs conserved in the M24A family of metallopeptidases. Protein–protein docking simulations using human MetAP revealed conservation of substrate and metal-binding residues in the catalytic site cleft of the novel enzyme, which was designated Ip MetAP. Recombinant Ip MetAP expressed in Escherichia coli revealed its significant enzymatic activity with the synthetic substrate H-Met-4-methyl-coumaryl-7-amide at pH 7.5 with K m of 0.014 mM, k cat of 0.25 s−1, and overall catalytic efficiency (k cat /K m) of 18.36 mM−1 s−1. The activity of Ip MetAP was enhanced by the addition of divalent cations Mn2+ and Co2+ and significantly inhibited by EDTA and bestatin. Site-directed mutagenesis of conserved amino acids indicated that the substitution of metal-binding residues D226 and H288 completely abolished the Ip MetAP enzymatic activity, whereas that of the other sites had only moderate effects on substrate hydrolysis. The catalytic properties of Ip MetAP suggest that the enzyme behaves similar to other MetAPs and such characterization expands our knowledge of aminopeptidases and protein metabolism of tick. [ABSTRACT FROM AUTHOR]

Genetic variability of I. apronophorus from Western Siberia, Russia was examined using the nuclear internal transcribed spacer 2 (ITS2) and mitochondrial 16S rRNA and cytochrome c oxidase subunit 1 (cox 1) genes and compared to those of Ixodes persulcatus and Ixodes trianguliceps from the same site. The I. apronophorus sequences demonstrated the highest nucleotide and haplotype diversity for both mitochondrial genes, whereas I. persulcatus was more variable in the nuclear ITS2. Phylogenetic analysis of the molecular sequence data showed that I. apronophorus differed from other Ixodes species, including Romanian I. apronophorus. The level of identity between 16S rRNA gene sequences of Siberian and Romanian I. apronophorus was only 91%; these sequences did not form a monophyletic group, indicating that I. apronophorus from Siberia and Romania could be different tick species. The analysis of morphological features of the Siberian I. apronophorus confirmed their consistency with those for the previously described I. apronophorus species. Based on the 16S rRNA and ITS2 sequences, Siberian I. apronophorus clustered together with Ixodes kazakstani and Ixodes scapularis , which are the recognized members of the Ixodes ricinus-I. persulcatus species complex within the subgenus Ixodes , and can be assigned to this complex. [ABSTRACT FROM AUTHOR]

Ixodes persulcatus and Ixodes pavlovskyi ticks, two closely related species of the I. ricinus - I. persulcatus group, are widely distributed in the southern part of Western Siberia. Recently, the existence of natural hybrids of I. persulcatus and I. pavlovskyi ticks has been demonstrated. The aim of this study was to evaluate the abundance of I. persulcatus / pavlovskyi hybrids in several locations with different ratios of parental tick species and to investigate the prevalence and genetic variability of a wide range of infectious agents in these hybrids compared to the parental tick species. Natural hybrids of I. persulcatus and I. pavlovskyi ticks were identified in all examined locations in Altai and Novosibirsk, Western Siberia, Russia. The abundance of hybrids varied from 7% to 40% in different locations and was maximal in a location with similar proportions of I. persulcatus and I. pavlovskyi ticks. For the first time, it was shown that hybrids can be infected with the same agents as their parental tick species: tick-borne encephalitis and Kemerovo viruses, Borrelia afzelii , Borrelia bavariensis, Borrelia garinii , Borrelia miyamotoi , Rickettsia helvetica , Rickettsia raoultii , Rickettsia sibirica , " Candidatus Rickettsia tarasevichiae", Anaplasma phagocytophilum, Ehrlichia muris , " Candidatus Neoehrlichia mikurensis", and Babesia microti. The prevalence of most bacterial agents in hybrids was intermediate compared to their parental tick species. Most genetic variants of the identified agents have been previously found in the parental tick species. Wide distribution of I. persulcatus / pavlovskyi natural hybrids implies that I. persulcatus , I. pavlovskyi and their hybrids coexist in all I. persulcatus - I. pavlovskyi sympatric areas. [ABSTRACT FROM AUTHOR]