Bartonella and Rickettsia Infections in Haematophagous Spinturnix myoti Mites (Acari: Mesostigmata) and their Bat Host, Myotis myotis (Yangochiroptera: Vespertilionidae), from Poland.

Hematophagous Spinturnix myoti mites and their host, the greater mouse-eared bat (Myotis myotis), were tested for the presence of Bartonella spp., Rickettsia spp., and Anaplasma phagocytophilum. In total, Bartonella spp. DNA was amplified in 28% of 134 mite pools and in 25% of 59 bats tested by PCR targeting a fragment of citrate synthase gltA gen. Adult mites were at least threefold more frequently infected compared to immature stages. The overall infection prevalence among mite pools from cave-dwelling bats was higher than for those collected from attic shelters. Three distinct genotypes were detected. The most prevalent genotype in mites and bats matched closely with Candidatus Bartonella hemsundetiensis identified in bats from Finland and was relatively distant from bat-borne Bartonella strains described in the UK and France. Importantly, most sequences were close to those reported in forest workers from Poland. The presence of identical genotype among S. myoti samples and M. myotis bats suggests that bartonellae can be shared between mites and their bat hosts. In this case, wing mites could serve as vectors, whereas their hosts as reservoirs. One blood sample was positive by PCR for the msp2 gene of A. phagocytophilum. Two mite pools yielded Rickettsia spp. DNA. Both sequences were distinct from any known species but can be classified as spotted fever group Rickettsia spp. Our findings expanded our knowledge on the role of spinturnicid mites in the ecology and epidemiology of bacterial infections associated with vespertilionid bats, especially regarding the genus Bartonella.

Correction to: Bartonella and Rickettsia Infections in Haematophagous Spinturnix myoti Mites (Acari: Mesostigmata) and their Bat Host, Myotis myotis (Yangochiroptera: Vespertilionidae), from Poland.

The original version of this article published online (27 August 2018) unfortunately contained a mistake regarding an affiliation of Dr. Edyta Podsiadly, one of the authors.

Evaluation of knowledge and use of the malaria prevention measures among the patients of the Department of Tropical and Parasitic Diseases University Center of Maritime and Tropical Medicine, Gdynia, based on a questionnaire performed in the years 2012-20

INTRODUCTION: Every year, approximately 125 million travelers visit areas where malaria prevails, located in over 100 countries. Over 10,000 of them suffer from malaria annually. Visitors to these areas may protect themselves against infection by using chemoprophylaxis, insect repellents, appropriate clothing, sleeping in airconditioned and well-screened quarters or using mosquito nets impregnated with insecticides. AIM OF THE STUDY: The aim of this study was to gather and analyze the data about knowledge and usage of pharmacological and non-pharmacological malaria prevention methods among the patients of the University Centre for Maritime and Tropical Medicine (UCMMiT), Gdynia, Poland, in 2012-2013. MATERIAL AND METHODS: A survey was conducted among 245 patients hospitalized in the Department of Tropical and Parasitic Diseases, UCMMiT in Gdynia, Poland in 2012 - 2013. The only criterion for inclusion was a sojourn and consent for participation in the study. The questionnaire included questions concerning mainly the use of chemoprophylaxis, opinion on the medication used for prophylaxis, side effects during its usage, the non-pharmacological prevention methods used against insect bites. RESULTS: Due to travel destination, malaria chemoprophylaxis should have been recommended for 73 (30%) individuals prior to the travel. It should not have been recommended for the group of 129 patients reporting long-term sojourns (over a year) and for 43 persons (17%) due to their travel to non-endemic countries. In fact, chemoprophylaxis in the "recommended" group was used by 32 persons which constituted 44%, while in the "long sojourn" group prophylaxis was used by 7 persons and in the "not recommended group" by 1 person. The number of people who reported proper use of chemoprophylaxis (an appropriate drug and mode of usage) amounted to 26 (36%) in the "recommended" group. Among bite prevention methods, usage of window mosquito nets was reported by 154 people (63%), bed mosquitonets by 39 (16%), insect repellents by 52 (21%) and insecticides by 52 (21%) patients. Adverse effects associated with chemoprophylaxis were reported by 10 persons (26%) but none of them stopped taking the medication. CONCLUSIONS: In spite of the availability of information on the Internet and the access to pre-travel medical counseling, the knowledge of correct prophylaxis against malaria among the people traveling to tropical countries is still unsatisfactory, leading to increased risk of Plasmodium infections. Better understanding of prophylaxis rules among travelers may increase usage of non-pharmacological methods and highlight the factors affecting usage of prevention methods.

Rickettsia helvetica and R. monacensis infections in immature Ixodes ricinus ticks derived from sylvatic passerine birds in west-central Poland.

The aim of this study was to assess the importance of forest passerine birds in spreading ixodid ticks infected with rickettsiae of spotted fever group (SFG) in sylvatic habitats in western Poland. In total, 834 immature Ixodes ricinus ticks were found on 64 birds of 11 species which were captured during the tick-questing season between May and September of 2006. Ground-foraging passerines hosted most of the ticks compared with arboreal species, and therefore, only the former group was included into a detailed analysis. Significant predominance of larvae over nymphs was observed (581 vs. 253, respectively). Blackbirds and song thrushes hosted 82 % (n = 681) of the ticks collected from all infested passerines. The overall prevalence range of SF rickettsiae (including Rickettsia helvetica and Rickettsia monacensis) in bird-derived ticks was 10.5-26.9 %, exceeding that in questing ticks, and in ticks feeding on rodents and deer reported earlier from the same study area. This high prevalence of infection in immature I. ricinus ticks feeding on passerine birds strongly implies that they are involved in the enzootic maintenance of spotted fever group rickettsiae in the tick vector populations occurring in sylvatic habitats.

Detection of Borrelia burgdorferi sensu lato and spotted fever group rickettsiae in hard ticks (Acari, Ixodidae) parasitizing bats in Poland.

A total of 491 Ixodes vespertilionis and 8 Ixodes ricinus collected from bats and cave walls in southern Poland between 2010 and 2012 were examined by the polymerase chain reaction for tick-transmitted pathogens. PCR analysis for Borrelia burgdorferi s.l., Rickettsia spp., and Anaplasma phagocytophilum yielded negative results for all I. vespertilionis. DNA of Rickettsia helvetica was detected in three specimens of I. ricinus attached to Rhinolophus hipposideros or Myotis myotis, while Borrelia garinii was found in one tick parasitizing Myotis daubentonii. These pathogens were recorded for the first time in hard ticks that parasitized bats.

Kampinos National Park: a risk area for spotted fever group rickettsioses, central Poland?

Ixodid ticks are important vectors of a variety of bacterial and protozoan pathogens which cause infections in humans. In this study, altogether 1041 questing Ixodes ricinus (n = 305) and Dermacentor reticulatus ticks (n = 736), sympatrically occurring in Kampinos National Park (KPN), central-east Poland, were analyzed by PCR for Rickettsia species. Overall, the pathogen prevalence in ticks was 27.5 % for I. ricinus and 42.8 % for D. reticulatus. Sequencing analysis showed that the first tick species was exclusively infected with R. helvetica, whereas the latter was infected with R. raoultii. These organism may pose a threat for populations exposed to ticks. Preliminary results of a serosurvey of 74 KPN employees, inhabitants and visitors from the same area showed a 31.1 % total seroprevalence against SFG rickettsiae compared to 13.3 % seropositive blood donors of the control group. Risk factors significantly associated with IgG seropositivity were: occupational exposure to ticks (p = 0.002), frequency of tick bites (p = 0.02) and male gender (p = 0.005). Seropositive and seronegative individuals occupationally exposed to ticks did not differ significantly with respect to age and years of employment.

Detection and quantification of Anaplasma phagocytophilum and Babesia spp. in Ixodes ricinus ticks from urban and rural environment, northern Poland, by real-time polymerase chain reaction.

Anaplasma phagocytophilum and Babesia spp. are emerging tick-borne pathogens which can threaten human health. A duplex real-time PCR and qPCRs with primers and probes targeting 97 and 116 bp fragments of 16S rRNA and 18S rRNA genes, respectively, were used for qualitative and quantitative detection of both pathogens in Ixodes ricinus ticks. Altogether 1875 ticks (1084 adults and 791 nymphs) were collected from rural and urban habitats in northern Poland. Of them, at least 0.9% were found to be infected with A. phagocytophilum while 2.5% with Babesia spp. A comparison of the infection rates by the tick stage, the type of area, the collection site, habitats of different tick density and by the month of collection was done. The prevalence of pathogens was significantly lower in nymphs than in adult ticks (p = 0.02) and in rural areas than in urban areas (p = 0.007). Four different 16S rRNA gene variants of A. phagocytophilum were determine, however none of them showed 100% identity with compared sequences isolated from human patients. The dominant Babesia species was B. venatorum. Results of qPCRs with circular and linearized forms of plasmids used as the standards showed significant difference in the pathogen loads (p = 0.001). The copy numbers of A. phagocytophilum and Babesia spp. estimated from the linear plasmids were 28.7 and 5.1 times lower, respectively, when compared with their circular forms, and were accepted as more reliable. The average number of copies of 16S rRNA gene of A. phagocytophilum in the positive I. ricinus samples were 3.39 × 10(5) ± 6.09 × 10(5). The mean copy number of 18S rRNA gene of Babesia spp. was ~2.55 × 10(5) ± 1.04 × 10(6). We confirmed the presence of A. phagocytophilum and Babesia spp. in I. ricinus in both rural and urban environments. The determined low infection rates suggests, however, that the risk for local population and tourists to acquire infection is also low. Moreover, we confirmed recent findings that serious overestimation by circular plasmid DNA makes it less suitable as a standard and that the linear standards should be recommended for qPCR.

Prevalence of tick-borne encephalitis virus (TBEV) RNA in Dermacentor reticulatus ticks from natural and urban environment, Poland.

Tick-borne encephalitis virus (TBEV) (Flaviviridae, Flavivirus) is an arthropod-borne virus, an etiologic agent of tick-borne encephalitis (TBE), a human infection involving the central nervous system. The disease is endemic in a large region in Eurasia, where it is transmitted mainly by Ixodes ricinus and Ixodes persulcatus ticks. It is known that also Dermacentor reticulatus is involved in circulation of TBEV. However, the current knowledge of D. reticulatus importance in TBE epidemiology is still insufficient. A total of 471 adult D. reticulatus ticks were collected by flagging vegetation in the Bialowieza Primeval Forest, Biebrza National Park, Masurian Landscape Park (North-Eastern Poland) and in the city of Warsaw in the years 2007-2010. All collected ticks were examined individually for the presence of RNA of TBEV using nested RT-PCR assay. Positive results were noted in all investigated localities with the infection rate ranging from 0.99 to 12.5 % with a total mean of 2.12 %. The difference in the percentage of infective males and females was not statistically significant.

Wild boars as hosts of human-pathogenic Anaplasma phagocytophilum variants.

To investigate the potential of wild boars to host Anaplasma phagocytophilum, we analyzed bacterial 16S rRNA and ank genes. DNA sequencing identified several A. phagocytophilum variants, including a predominance of strains known to cause human disease. Boars are thus hosts for A. phagocytophilum, notably, strains associated with human granulocytic anaplasmosis.

Fires, vegetation, and human-The history of critical transitions during the last 1000 years in Northeastern Mongolia.

Fires are natural phenomena that impact human behaviors, vegetation, and landscape functions. However, the long-term history of fire, especially in the permafrost marginal zone of Central Asia (Mongolia), is poorly understood. This paper presents the results of radiocarbon and short-lived radionuclides (210Pb and 137Cs) dating, pollen, geochemical, charcoal, and statistical analyses (Kohonen's artificial neural network) of sediment core obtained from Northern Mongolia (the Khentii Mountains region). Therefore, we present the first high-resolution fire history from Northern Mongolia covering the last 1000 years, based on a multiproxy analysis of peat archive data. The results revealed that most of the fires in the region were likely initiated by natural factors, which were probably related to heatwaves causing prolonged droughts. We have demonstrated the link between enhanced fires and "dzud", a local climatic phenomenon. The number of livestock, which has been increasing for several decades, and the observed climatic changes are superimposed to cause "dzud", a deadly combination of droughts and snowy winter, which affects fire intensity. We observed that the study area has a sensitive ecosystem that reacts quickly to climate change. In terms of changes in the vegetation, the reconstruction reflected climate variations during the last millennium, the degradation of permafrost and occurrence of fires. However, more sites with good chronologies are needed to thoroughly understand the spatial relationships between changing climate, permafrost degradation, and vegetation change, which ultimately affect the nomadic societies in the region of Central and Northern Mongolia.

Detection of Anaplasma phagocytophilum in Amblyomma flavomaculatum ticks (Acari: Ixodidae) collected from lizard Varanus exanthematicus imported to Poland.

Adults and nymphs of Amblyomma exornatum, A. flavomaculatum, A. latum, Amblyomma spp. and Hyalomma aegyptium, were collected from savannah monitors, royal python (Africa, Ghana) and marginated tortoises (Europe, Greece) imported to Poland, in 2004-2007. Altogether 345 ticks were examined by polymerase chain reaction for rickettsial agents. None of the tested ticks was positive for the gltA gene of Rickettsia spp. DNA of 16S rRNA gene from Anaplasma phagocytophilum was amplified and sequenced from two adult A. flavomaculatum ticks attached to two different specimens Varanus exanthematicus from Ghana. Both PCR amplicons obtained (GQ305134) had 100% sequence homology with A. phagocytophilum sequences deposited in GenBank. This results suggests for the first time that A. flavomaculatum may be vector of this pathogen in Africa. It could be expected that ticks distributed on their hosts may introduce pathogens not yet known to Polish or even European fauna. This study contributes to the overall recognition of the scarcely explored fauna of the exotic ticks transferred to Poland on reptiles from remote destinations.

Diversity of Borrelia burgdorferi sensu lato species in Ixodes ticks (Acari: Ixodidae) associated with cave-dwelling bats from Poland and Romania.

Bats comprise one quarter of the world's mammal species. In Europe, three nidicolous Ixodes tick species, I. vespertilionis, I. simplex and I. ariadnae are specifically associated with cave-dwelling bats, but their role as potential vectors of zoonotic agents is unknown. In this study, we used PCR-based methods to provide the first evidence of Borrelia burgdorferi sensu lato (s.l.) infections in the three bat-associated tick species collected from ten bat species sampled in Poland and Romania. B. burgdorferi s.l. was detected in 24% (64/266) of tick samples, and 40.3% (60/149) of the bats carried infected chiropterophilic ticks. In Poland, the B. burgdorferi s.l. infection prevelance of I. ariadnae ticks parasitizing Myotis species was four times higher compared to the I. vespertilionis ticks derived from Rhinolophus hipposideros bats (44.4% vs.10%, respectively). The observed differences in infection prevalence could be explained by differences in reservoir potential between bat species. Bats from the genus Myotis and Miniopterus schreibersii carried more infected ticks than R. hipposideros regardless of the tick species. Analysis of the flaB gene sequences revealed seven species from the B. burgdorferi s.l. complex (B. afzelii, B. carolinensis, B. garinii, B. lanei, B. spielmanii, B. burgdorferi s.s., and B. valaisiana), of which five are considered as human pathogens. This large diversity of Borrelia species may reflect differences in susceptibility of chiropteran hosts and/or the tick vectors. Generally, mammal-associated B. burgdorferi s.l. species were more common than bird-associated species. Our study provides evidence for new enzootic transmission cycles of B. burgdorferi s.l. spirochetes involving nidicolous Ixodes tick species and cave-dwelling bats.

The first detection of Babesia EU1 and Babesia canis canis in Ixodes ricinus ticks (Acari, Ixodidae) collected in urban and rural areas in northern Poland.

Ixodes ricinus, the most commonly observed tick species in Poland, is a known vector of such pathogenic microorganisms as TBE viruses, Borrelia burgdoferi sensu lato, Anaplasma phagocytophilum, Rickettsia helvetica, Babesia divergens and B. microti in our country. Our study aimed to find out whether this tick can also transmit other babesiae of medical and veterinary importance. DNA extracts of 1392 ticks (314 nymphs, 552 male and 526 female ticks) collected in urban and rural areas in the Pomerania province (northern Poland), were examined by nested PCR for the detection of Babesia spp., using outer primers: 5-22F and 1661R, and inner primers: 455-479F and 793-772R, targeting specific fragment of 18S rRNA gene. Overall, at least 1.6% ticks were found to be infected with babesial parasites. In the case of nymphs, the minimal prevalence was 0.6%, and it was approx. 3-times lower than in adults (1.9%). Percentages of infected males and females were comparable (2.0% vs. 1.7%). Sequences of 15/22 PCR-derived fragments of 18S rRNA gene demonstrated 100% similarities with the sequence of Babesia EUI (proposed name B. venatorum) (acc. no. AY046575) (n = 13) and with B. canis canis (acc. no. AY321119) (n = 2), deposited in the GenBank database. The partial 18S rDNA sequences of Babesia EUI and B. c. canis obtained by us from I. ricinus have been deposited in GenBank, accession nos. GQ325619 and GQ325620, respectively. The results obtained suggest the possible role of I. ricinus as a source of microorganisms, which have been identified as agents of human and canine babesiosis, respectively, in Europe. To our knowledge this is the first report on the occurrence of Babesia EUI and B. c. canis in I. ricinus in Poland.

[The occurrence of Cryptosporidium spp. in synanthropic flies in urban and rural environments]. / Wystepowanie Cryptosporidium spp. u muchówek synantropijnych na wybranych stanowiskach miejskich i wiejskich.

This study was carried out to determine the role of non-biting synanthropic flies as carriers of Cryptosporidium spp. oocysts in the vicinity of the city of Gdansk (NE Poland). In 2001-2003, flies were collected from three breeding sites: cow sheds and meadows in the Bystra cattle farm and municipal landfill Szadólki using inhaustors (aspirators) and entomologic nets. A total of 2358 specimens of the families: Muscidae (n = 1598), Calliphoridae (n = 739) and Sarcophagidae (n =21) were collected and analysed in 249 pools consisted of 9.5 insects, in average. Microscopic examination was used to detect Cryptosporidium spp. oocysts in the fly faeces deposited on the glass microscope slides and stained by Zhiel-Nielsen method. The mean number of faecal droplets per one glass slide was 11.5. Ooocysts of Cryptosporidium spp., stained from light pink to bright red, were found in fly faeces deposited on 25 (27.5%) of 91 glass slides checked. The highest prevalence of the pathogen was observed in faecal droplets deposited by flies collected in municipal landfill (50% investigated slides). DNA of Cryptosporidium spp. was extracted from the surface eluants of flies and/or their gut homogenates and purified. Then extracts were examined by PCR using CPB-DIAGF and CPB-DIAGR primers amplifying a variable region SSU-rRNA of all Cryptosporidium species. Altogether 387 isolates, 228 from surfaces and 159 from gut homogenates, were obtained from 249 pools of flies and analyzed. A specific 435 bp fragment of DNA was obtained in 49 (12.7%) lysates tested. In 10.4% pools, DNA of the pathogen was detected only in the surface eluants while in 7.6% only in gut extracts. In the case of two pooled samples (0.8%) Cryptosporidium spp. was found in both types of lysates. In total, Cryptosporidium spp. was detected in 47/249 pools of flies (18.9%). Assumed that each positive pool contained just one infected fly, the percentage of specimens able to oocysts transmission were calculated at the minimal level 2.0% (n = 47/2358). The result confirm that synanthropic flies can harbour oocysts of Cryptosporidium spp. both externally and internally, and disseminate them mechanically in the environment. Therefore, under unsanitary conditions they could be involved in the transmission of human and animal cryptosporidiosis.

[A study on the occurrence of West Nile virus in mosquitoes (Diptera: Culicidae) on the selected areas in Poland]. / Badania nad wystepowaniem wirusa goraczki Zachodniego Nilu u komarów (Diptera: Culicidae) na wybranych terenach Polski.

West Nile virus (WNV), the etiologic agent of West Nile Fever (WNF), an emerging infectious disease that lately has been rapidly extending its range of occurrence from Africa to Middle East, and to Asia and Southern Europe. In Europe, cases of isolating WNV from mosquitoes representing four genera have been reported from Romania, Portugal, France, southern Russia and what is the most important, from Poland's neighbouring countries as the Czech Republic, Slovakia and southern Ukraine. These data, as well as human and equine cases in the Czech Republic and Belarus, support hypothesis that WNV has already been present also in Poland, the more so, specific antibodies were detected in the sera of birds collected at the Kampinos and Bialowieza Primeval Forests and in human from the vicinity of Bialystok. Mosquitoes were collected in 2004-2009 at indoor and outdoor collection sites in districts: Kujawsko-Pomorskie, Mazowieckie, Podlaskie and Warminsko-Mazurskie. In total 15400 female mosquitoes were collected in the cow sheds and overwintering in the cellars, and from human bait and CO2 traps. Mosquitoes were sorted by the collecting site, species and sex, and placed in pools of not in more than 50 specimens in special mixture of phenol and chaotropic salts and frozen in -20 degrees C. Altogether, 15400 females were examined. Total RNA were extracted according to protocol of A&A Biotechnology. RT reaction was performed with random primers and 217-nucleotide fragment from the NS5 coding sequence was amplified by heminested PCR. PCR products were analysed on 1.5% agarose gel. The predominant species was Culex pipiens, accounting for over 42% of total insects collection. All obtained results were negative. Further investigations are needed.

[Occurrence of Toxoplasma gondii specific antibodies in group of forestry workers from pomorskie and warminsko-mazurskie provinces]. / Wystepowanie specyficznych przeciwcial anty-Toxoplasma gondii w grupie pracowników lesnych z województwa pomorskiego i warminsko-mazurskiego.

Toxoplasma gondii infection is one of the most common human zoonosis. Humans acquire toxoplasmosis by consuming raw or undercooked meat, from environments and from via vertical transmission to the foetus through the placenta during pregnancy. In the present study, a total of 784 persons (forestry workers), from north-eastern region of Poland (pomorskie and warminsko-mazurskie provinces), were tested by using commercially available ELISA test (DiaSorin) to determine the frequency of Toxoplasma gondii infection. The study was based on detection of IgG and IgM specific antibodies in collected serum samples. The T. gondii-specific IgG were found in 490 samples (62.5%), while the IgM antibodies were noted only in 18 persons (3.7%). High prevalence of seropositive results in forestry workers may suggest an occupational exposure of this group to T. gondii.

The role of particular ticks developmental stages in the circulation of tick-borne pathogens in Central Europe. 5. Borreliaceae

The developmental cycles of all B. burgdorferi s.l. genospecies present typical, main pattern described in the 90thies. The simple scheme might be modified according to the biology of species and hosts preference. Central European genospecies of B. burgdorferi s.l. can be associated with four groups of hosts playing the role of animal reservoirs. The group 1 contains genospecies associated with rodents as primary animal reservoir ­ B. afzelii, B. garinii, B. burgdorferi sensu stricto, strains B. bavariensis (B. garinii OspA serotype 4). The group 2 involves B. valaisiana and most of B. garinii strains, associated with birds. The group 3 involves B. spielmanii, the reservoir hosts are Gliridae, and hedgehogs. The group 4 includes B. lusitaniae, the hosts are lizards. B. miyamotoi enzootic cycle seems to be similar to B. burgdorferi complex, however, differs by the transovarial transmission possibility. The divisions are not extreme; in the hosts group, infected with appropriate Borrelia genospecies, very often are found the specimens infected with other genospecies.

The role of particular ticks developmental stages in the circulation of tick-borne pathogens in Central Europe. 6. Babesia.

In the Central European conditions, three species of Babesia have epidemiological significance as human pathogens ­ Babesia divergens, B. microti and B. venatorum. Tick Ixodes ricinus is considered as their main vector, wild mammals as the animal reservoir. The zoonotic cycles of small and large Babesia differ in details. Due to the lack of transovarial mode transmission in small species B. microti, the circulation goes mainly between immature ticks and vertebrate hosts; pathogen circulates primarily in the cycle: infected rodent → the tick larva → the nymph → the mammal reservoir →the larva of the tick. The tick stages able to effectively infect human are nymphs and adult females, males do not participate in the follow transmission. For large Babesia ­ B. divergens and B. venatorum, the transovarial and transstadial transmission enable the presence of the agent in adult ticks, moreover, that larvae and nymphs feed on not-susceptible hosts. The tick stages able to effectively infect cattle and other ruminants are adult females. Resuming, pathogen circulates primarily in the cycle the ruminant host ­ adult female tick ­ the larva ­ the nymph ­ adult female of the next generation ­ the ruminant. Due to the compound developmental transmission has place after the outflow of a tick began feeding.

Prevalence of different Rickettsia spp. in Ixodes ricinus and Dermacentor reticulatus ticks (Acari: Ixodidae) in north-eastern Poland.

In two surveys conducted in April and May 2013, a total of 1148 unfed ticks belonging to two species, Ixodes ricinus and Dermacentor reticulatus, were collected by flagging lower vegetation in 15 different localities throughout the Bialowieza Primeval Forest (Podlaskie voivodship; north-eastern Poland) and in its buffer zone. In order to establish their infection rate with Rickettsia spp. individual adult ticks and pooled nymphs were tested by real-time PCR targeting the gltA gene. For the further identification of rickettsial species, positive samples were subjected for nested and semi-nested PCR targeting ompA and 16S rRNA genes, respectively, followed by sequencing analysis. Rickettsial DNA was detected in at least 279 ticks (minimum infection rate [MIR], 23.9%), including 52 nymphal and adult I. ricinus (MIR 8.6%) and 222 adult D. reticulatus (41%). Three species of SFG rickettsiae were identified: Rickettsia helvetica and 'Candidatus R. mendelii' in I. ricinus and R. raoultii in D. reticulatus and I. ricinus. Moreover, unidentified Rickettsia spp. which showed 99.4% identity, among others, with the uncultured Rickettsia sp. isolated from Cicadella viridis leafhopper, Rickettsia endosymbiont of Lasioglossum semilucens bee and R. bellii, were detected in I. ricinus, while Rickettsia sp. 98.3-98.4% homologous to Rickettsia secondary endosymbionts of Curculio spp. weevils was found in D. reticulatus. These results confirm the diversity of rickettsiae occurring in Poland. Further studies are needed to expand the knowledge on the species spectrum, prevalence and epidemiology of SFG rickettsiae in the country.

Seroprevalence of TBEV in bank voles from Poland-a long-term approach.

Rodents are known to play a significant role as reservoir hosts for TBEV. During three sequential expeditions at 4-year intervals to three ecologically similar study sites in NE Poland, we trapped bank voles (Myodes glareolus) and then tested their blood for the presence of specific antiviral antibodies to TBEV. The strongest effects on seroprevalence were the extrinsic factors, site of capture of voles and year of sampling. Seroprevalence increased markedly with increasing host age, and our analysis revealed significant interactions among these three factors. Seroprevalence did not differ between the sexes. Therefore, based on the seroprevalence results, the dynamics of TBEV infection differ significantly in time, between local sub-populations of bank voles and with increasing host age. To fully understand the circulation of the virus among these reservoir hosts and in the environment, long-term monitoring is required and should employ a multi-site approach, such as the one adopted in the current study.