Sparganosis (Spirometra) in Europe in the Molecular Era.

Sparganosis is a relatively neglected foodborne and waterborne disease caused by species of the tapeworm genus Spirometra, the global distribution of which has not been sufficiently recognized. Known mainly as a zoonosis of East Asia, its species are native to all inhabited continents including Europe. Spirometra has been reported from numerous wildlife species from 17 European countries, and a critical review confirmed 17 autochthonous and 8 imported human clinical cases. We present the first molecular evidence of the coincident presence of 2 species in Europe and review the current distribution to raise awareness of the parasite in this region. Spirometra erinaceieuropaei is restricted to Europe and Spirometra mansoni represents a lineage distributed mainly across Asia and Oceania that reaches Europe. The parasite is common in Eastern Europe and its distribution has potential to expand along with its invasive or migrating mammal hosts, spreading the risks of human infection.

The first case of autochthonous subcutaneous dirofilariasis (Dirofilaria repens) in a dog from Bialowieza (NE Poland) and possible threat posed to inhabitants of Bialowieza Primeval Forest area.

We present the first case of dirofilariasis in a dog from Bialowieza village located in the primeval European forest-Bialowieza Primeval Forest (NE Poland). Molecular analysis of adult nematode specimens isolated from subcutaneous tissue confirmed the infection with Dirofilaria repens. An adult male dog has not travelled out of the Bialowieza village for at least five years; therefore, we assume this is the autochthonous case of the disease. We discuss possible inter- and intra-species transmission routes of dirofilariasis on this territory, which is inhabited by diverse community of wild carnivores, domestic animals, and humans. We also discuss the likely sources of the disease in this, highly biodiverse unique European forest complex. We underline the lack of attention to this problem and its importance for veterinary, wildlife, and human health safety.

The first records of Spirometra erinaceieuropaei (Cestoda: Diphyllobothriidae), a causative agent of human sparganosis, in Latvian wildlife.

Diphyllobothriid tapeworms of the genus Spirometra are causative agents of sparganosis, food-borne zoonotic parasitic disease. They have been recorded in broad spectrum of hosts, including humans, in all continents except Antarctica. Spirometra tapeworms have been intensively studied in several Asian countries; however, they have been rather neglected in Europe. The aim of this study was to provide a pilot screening of Spirometra spp. in Latvia, where data on sparganosis are not available. Tapeworms morphologically identified as diphyllobothriid species were isolated from grey wolves Canis lupus and Eurasian lynxes Lynx lynx from Latvia during the hunting periods 2013-2019. The parasites were subjected to molecular genotyping using sequences of the partial large (LSU rDNA; 615 bp) and small (SSU rDNA; 720 bp) subunits of the nuclear ribosomal RNA gene and complete (1566 bp) cytochrome c oxidase subunit I gene of the mitochondrial DNA (cox1 mtDNA). Analyses of both ribosomal subunits of 13 tapeworms revealed no intraspecific variation within the respective rDNA subunits. On the other hand, sequence analysis of mitochondrial cox1 revealed intraspecific polymorphism displayed by 12 cox1 haplotypes. Comparison of the current data with sequences of the corresponding DNA regions deposited in the GenBank revealed 99.3-99.5% (LSU rDNA), 99.2% (SSU rDNA) and 99.6-100% (cox1 mtDNA) identity of studied tapeworms with Spirometra erinaceieuropaei, which provided the first confirmation of this diphyllobothriid tapeworm in Latvia. Since S. erinaceieuropaei is probably prevalent in Latvian wildlife and may also occur in other potential host species, further studies are needed in order to acquire complex data on its geographic distribution and transmission in the natural environment of Latvia, as well as on the spectrum of its intermediate, paratenic, and definitive hosts.

Alaria spp. mesocercariae in Eurasian badger (Meles meles) and wild boar (Sus scrofa) from the Bialowieza Forest, north-eastern Poland.

Alaria spp. mesocercariae are commonly found in wild boar and other omnivorous mammals. In Europe, the number of cases presenting Alaria mesocercariae infections has been on the rise in the last years. From October to December 2016, samples of muscle from tongue, neck, and mandibular regions were collected from 1 Eurasian badger (Meles meles) and 14 wild boars (Sus scrofa) hunted in the Bialowieza Forest, north-eastern Poland. Using the Alaria migration technique (AMT), Alaria mesocercariae were isolated and morphologically identified in one badger and one wild boar. To the authors' knowledge, this is the first report of Alaria mesocercariae in paratenic hosts from the Bialowieza Forest.

The first case of genetically confirmed sparganosis (Spirometra erinaceieuropaei) in European reptiles.

Sparganosis is a zoonosis caused by the spargana (larvae) of Spirometra sp. (Diphyllobothriidae). Reptiles are particularly important vectors for the transmission of this parasite in Asia; however, their role in sparganosis spread in European wildlife is unrecognized. We investigated the infection of reptiles with Spirometra sp. in NE Poland, where several mammalian hosts have been identified recently and in the past. Of the 59 dead reptiles, plerocercoids were found in two grass snakes (Natrix natrix) from the Bialowieza Primeval Forest (BPF). The Spirometra erinaceieuropaei species was genetically confirmed using the evolutionary conserved nuclear 18S rRNA gene, and then compared to GenBank deposits. The sequences were identical to previously investigated Spirometra sp. found in Eurasian badger and wild boar from BPF. Our finding is the first genetically confirmed record of Spirometra sp. in reptiles in Europe. Since reptiles are often a component of mammalian diet, they can be a source of Spirometra tapeworm infection in European wildlife; however, further studies are needed to investigate the prevalence of infection in reptiles and other non-mammalian hosts.

Update of the helminth fauna in Eurasian lynx (Lynx lynx) in Poland.

The Eurasian lynx (Lynx lynx) is a strictly protected species of large carnivore in Poland. It inhabits forest complexes in north-eastern Poland and the Carpathian region in southern Poland. The status of the lynx within Poland requires special attention because its range decreased between 1980 and 2001 and has not yet recovered. One of the factors negatively affecting lynx populations is diseases, particularly parasites. The helminth fauna of the Eurasian lynx is not fully known in Poland. Previous coprological studies revealed that Polish lynx have been infected with seven species of nematodes, three species of cestodes, and one species of trematode. In this study, we present new data based on examination of opportunistically collected lynx carcasses. The aim of the study was to complement data on the helminth fauna of Eurasian lynx inhabiting Poland based on morphological and molecular analysis of parasites. Four species of cestodes-Taenia lynciscapreoli, Mesocestoides lineatus, Spirometra sp., and Taenia krabbei-were found for the first time in Eurasian lynx from Poland and three previously reported species of nematodes-Ancylostoma tubaeforme, Toxascaris leonina, and Toxocara cati-were confirmed. Larvae of Trichinella britovi were also detected in Eurasian lynx in Poland for the first time.

Asymptomatic viral infection is associated with lower host reproductive output in wild mink populations.

Many endemic viruses circulate in populations without hosts showing visible signs of disease, while still having the potential to alter host survival or reproduction. Aleutian Mink Disease Virus (AMDV) circulates in many American mink (Neogale vison) populations in its native and introduced ranges. In this study, we analysed how AMDV infection in female American mink affects the reproduction of a feral population. Females infected with AMDV delivered significantly smaller litters (5.8 pups) than uninfected females (6.3 pups), meaning their litter size was reduced by 8%. Larger females and yearling females had larger litters than smaller and older females. There were no significant differences in whole litter survival between infected and uninfected females; however, offspring survival until September or October within litters of infected females was 14% lower than that within those of uninfected females. This negative link between infection and reproductive output means that Aleutian disease could seriously affect the wild mink population. This study increases our understanding of the threats posed by the spread of viruses to wildlife from farm animals or humans, highlighting that viruses circulating in wildlife, even in the absence of clinical manifestation, can be important drivers of population dynamics in wildlife.

The genetic structure of populations of Isthmiophora melis (Schrank, 1788) (Digenea: Echinostomatidae). Does the host's diet matter?

BACKGROUND: Here we provide a comparative analysis of the genetic structure of populations (based on nad1 mtDNA) of Isthmiophora melis isolated from the American mink (Neogale vison), an introduced invasive species, commonly occurring in the territory of Poland, and from the striped field mouse (Apodemus agrarius). METHODS: A total of 133 specimens of I. melis were obtained from naturally infected N. vison collected from six localities in Poland (108 samples) and 25 individuals of I. melis from A. agrarius. All sequences of the nad1 gene obtained during the present study were assembled and aligned. The standard statistics for haplotype composition, i.e., the number of haplotypes, haplotype diversity, nucleotide diversity, and average number of nucleotide differences, were calculated. Haplotype analysis and visualization of haplotype frequency among populations were performed using a median-joining network. RESULTS: Based on the samples collected from different localities in Poland, our study revealed that the overall genetic diversity of I. melis isolated from the American mink and of the striped field mouse do not differ significantly. The median-joining network showed that the three main haplotypes are in the centre of a star-like structure, with the remaining haplotypes as the satellites, reflecting the recent expansion of the populations. CONCLUSIONS: The overall genetic diversity of I. melis isolated from the American mink and striped field mouse reveals a high level of homogeneity. Moreover, regional differences in the food composition of the definitive hosts play an important role in shaping the genetic structure of the trematode populations.

Effect of Translocation on Host Diet and Parasite Egg Burden: A Study of the European Bison (Bison bonasus).

For the purpose of nature management and species conservation, European bison (Bison bonasus) are being increasingly reintroduced into nature reserves across Europe. The aim of this study was to investigate European bison's adaptability to new areas through the study of their parasite-EPG (eggs per gram feces) and dietary diversity during twelve months after translocation. We compared the parasite-EPG from introduced European bison in Lille Vildmose, Denmark, with the parasite-EPG from populations from Bornholm, Denmark, and Bialowieza Forest, Poland. From March 2021 to February 2022, fecal samples were collected from three populations. Samples from Lille Vildmose were examined through flotation, sedimentation, the Baermann technique, and nanopore sequencing. Fecal samples from Bornholm and Bialowieza were examined through flotation and sedimentation. Nanopore sequencing of DNA from 63 European bison's fecal samples collected during March-September in Lille Vildmose identified 8 species of nematodes within the digestive tract of the European bison, with Haemonchus contortus being the most frequently observed. In Lille Vildmose, a significantly higher excretion of nematode-EPG was observed during the summer period than in the spring, autumn, and winter. In addition, monthly differences in the excretion of nematode eggs were found, with this being significantly higher in June than in the months during autumn and winter (October-February). Significant differences in the nematode-EPG were only found between the excretion of nematode eggs in Bialowieza Forest when compared to that of Lille Vildmose, with significantly higher excretion in Lille Vildmose (October-November). The results indicate that the development rates for nematodes may be affected by changes in temperature, with increasing temperatures speeding up their development time. Independent of this study design, wildlife vets together with the gamekeepers managing the herd found it necessary to treat the herd with antiparasitics for practical and animal welfare reasons in relation to translocation. Furthermore, 79 plant taxa were identified in the diet of the European bison. The broadest diet was observed in March suggesting that the European bison quickly adapted to their new habitat. The results suggest a seasonal shift in their diet, with this being most apparent from March to April.

A tale of two nematodes: Climate mediates mustelid infection by nematodes across the geographical range.

Parasites have the potential to negatively affect host populations, if infection intensity is high. For parasites in which part of life cycle takes place outside the host, host infection intensity is likely affected by climate condition. Therefore, the parasite's impact on the host populations could be related to climatic conditions and may be altered with climate change. The aim of our study was to analyse the prevalence and infection intensity of two nematodes (Aonchotheca putorii and Molineus patens) from the Northern Hemisphere in relation to variations in climatic conditions. We reviewed 54 published studies on the occurrence of these two nematode species in 7 mustelid hosts. For A. putorii, infection parameters were higher when the stomach was included in the analyses compared to M. patens. The seasonality of precipitation influenced the prevalence the most, and the mean temperature of the warmest quarter had the strongest influence on infection intensity. The predicted prevalence of M. patens increased with increasing seasonal variation in precipitation, while the prevalence of A. putorii decreased. The predicted infection intensity of M. patens decreased with increasing precipitation seasonality, whereas the intensity of A. putorii infection did not change much. A. putorii infection intensity significantly decreased with increasing mean temperature of the warmest quarter, while the infection intensity of M. patens was not significantly related to this variable. Prevalence and infection intensity varied over the geographic range for both parasites, broadly with higher levels in northern latitudes for A. putorii and in southern latitudes for M. patens. Our study highlights the differences between these two nematode species and shows that the severity of host infection by these parasites is complex and mediated by climatic conditions. The results suggest that current climate change may potentially modify susceptibility and exposure to parasitic infections in mustelids.

Development of cellular immune response of mice to infection with low doses of Trichinella spiralis, Trichinella britovi and Trichinella pseudospiralis larvae.

The murine cellular immune response to the infection with ten larvae of encapsulating (Trichinella spiralis, Trichinella britovi) and non-encapsulating species (Trichinella pseudospiralis) was studied. Both T. spiralis and T. britovi stimulated the proliferation of splenic T and B lymphocytes during the intestinal phase of infection, but T. spiralis activated the proliferative response also at the muscle phase, particularly in B cells. Non-encapsulating T. pseudospiralis stimulated the proliferation of T and B cells only on day 10 post-infection (p.i.) and later at the muscle phase. The numbers of splenic CD4 and CD8 T cells of T. spiralis infected mice were significantly increased till day 10 p.i., i.e., at the intestinal phase, and then at the late muscle phase, on day 60 p.i. T. britovi infection increased the CD4 and CD8 T cell numbers only on day 30 p.i. Decreased numbers of CD4 and CD8 T cells after T. pseudospiralis infection suggest a suppression of cellular immunity. Both encapsulating Trichinella species induced the Th2 response (cytokines interleukin-5 (IL-5) and interleukin-10) at the intestinal phase and the Th2 dominant response at the advanced muscle phase. Interferon-γ (IFN-γ) production (Th1 type) started to increase with migrating newborn larvae from day 15 p.i. till the end of the experiment. IL-5 production was suppressed during the intestinal phase of T. pseudospiralis infection. The immune response to T. pseudospiralis was directed more to the Th1 response at the muscle phase, the high IFN-γ production was found on day 10 p.i. and it peaked on days 45 and 60 p.i.

Digestive tract nematode infections in non-native invasive American mink with the first molecular identification of Molineus patens.

Parasites may negatively affect hosts condition, especially when infection intensity is high. Species introduced to a new habitat are often less exposed to a parasite pressure but may accumulate parasites in time. American mink (Neovison vison) introduced to Europe, Asia, and South America is an example of such invasive species. We analysed nematode prevalence and digestive tract infection intensity in 796 feral American mink from Poland. The analyses were performed separately for stomach, duodenum, small intestine and large intestine. Parasite species identification was performed using molecular methods based on highly conserved nuclear 18S rRNA gene and supplemented with morphological analysis. In total, we collected 26,852 nematodes and 98.6% of them were isolated from mink stomachs. We found positive association between infection intensity in stomach and other parts of digestive tract. Nematode prevalence was estimated at 63.8% and average infection intensity per one American mink at 52.9 (range from 1 to 1118). If the stomach results were theoretically and intentionally omitted the prevalence was 5 times lower (12.7%) and infection intensity 14 times lower (3.7; range 1-50). We identified two nematode species in digestive tracts of American mink: Aonchotheca putorii and Molineus patens. The 18S rRNA gene sequence of Molineus patens has been reported for the first time. The results showed that Aonchotheca putorii is a dominating nematode in the invasive American mink and that it inhabits stomach intensively and preferably.

Blastocystis occurrence and subtype diversity in wild European terrestrial mammals - The case of Bialowieza Primeval Forest (NE Poland).

Blastocystis cf. Blastocystis hominis is the most common unicellular parasite found in human and animal intestines. Little is known about the life cycle, transmission and mechanisms of pathogenesis for this parasite. The aim of this study was to obtain new data on the genetic diversity of Blastocystis in selected species of mammals found in the Bialowieza Primeval Forest (BPF), Poland, the best preserved lowland forest in temperate Europe. 113 faecal samples were collected in the period 2018-2020 from seven species of wild mammals occurring within the Polish part of BPF and its surroundings. Blastocystis was detected by molecular amplification and sequencing the small subunit rRNA gene. The overall prevalence of Blastocystis in animals was 8.9%. A larger number of stool samples (90) were collected from European bison and only for this species was it possible to calculate the prevalence of infection (5.6%). The isolates obtained from European bison were classified as ST1, ST3, ST5, ST7. Blastocystis was also detected in the grey wolf (Canis lupus) (ST5), wild boar (Sus scrofa) (ST5) and red deer (Cervus elaphus) (ST1). In conclusion, for the first time we have demonstrated the presence of Blastocystis in wild European bison. Preliminary studies have shown that Blastocystis is present in at least 4 species of wild mammals in the BPF area and that it exhibits great genetic diversity.

Diversity and transmission of Aleutian mink disease virus in feral and farmed American mink and native mustelids.

Aleutian mink disease virus (AMDV), which causes Aleutian disease, is widely spread both in farmed mink and wild mustelids. However, only limited data are available on the role of wild animals in AMDV transmission and spread. Our aim was to shed light on AMDV transmission among wild mustelids and estimate the effect of intense farming practices on the virus circulation by studying AMDV prevalence and genetic diversity among wild mustelids in Poland. We compared AMDV seroprevalence and proportion of PCR-positive individuals in American mink, polecats, otters, stone martens, and pine martens and used the phylogenetic analysis of the NS1 region to study transmission. In addition, we used a metagenomic approach to sequence complete AMDV genomes from tissue samples. In eastern Poland, AMDV seroprevalence in wild mustelids varied from 22 per cent in otters to 62 per cent and 64 per cent in stone martens and feral mink, respectively. All studied antibody-positive mink were also PCR positive, whereas only 10, 15, and 18 per cent of antibody-positive polecats, pine martens, and stone martens, respectively, were PCR positive, suggesting lower virus persistence among these animal species as compared to feral mink. In phylogenetic analysis, most sequences from feral mink formed region-specific clusters that have most likely emerged through multiple introductions of AMDV to feral mink population over decades. However, virus spread between regions was also observed. Virus sequences derived from farmed and wild animals formed separate subclusters in the phylogenetic tree, and no signs of recent virus transmission between farmed and wild animals were observed despite the frequent inflow of farmed mink escapees to wild populations. These results provide new information about the role of different mustelid species in AMDV transmission and about virus circulation among the wild mustelids. In addition, we pinpoint gaps of knowledge, where more studies are needed to achieve a comprehensive picture of AMDV transmission.

Aleutian mink disease: Spatio-temporal variation of prevalence and influence on the feral American mink.

Pathogens are one of the factors driving wildlife population dynamics. The spread of pathogens in wildlife is currently highly related to the transmission of pathogens from farmed animals, which has increased with the constant development of farming. Here, we analysed the spatio-temporal variation in the prevalence of Aleutian mink disease virus (AMDV) antibodies in feral American mink (Neovison vison) populations in Poland (1,153 individuals from nine sites) in relation to mink farming intensity. AMDV was detected in feral mink at all study sites and the prevalence ranged from 0.461 in the northern region to 0.826 in the western region. Mink males and adults were infected more often than females and subadults; the infection was also more frequent during the mink breeding season than during non-breeding. The prevalence of AMDV changed non-linearly in consecutive years and the peak of prevalence was every 3-4 years. The predicted AMDV prevalence was low at sites where the number of farmed mink was also low and increased linearly with the increase in the number of mink kept on farms. The predicted AMDV prevalence at sites with low mink farming intensity strongly varied between years, whereas at sites with high mink farming intensity, the predicted prevalence did not change significantly. AMDV infection affected the mink's body condition and caused an increase in the size of the spleen, liver and kidneys. This study shows that Aleutian mink disease strongly affects feral mink but the spatio-temporal variation of its prevalence is complex and partly related to the transmission of the virus from farmed mink to feral populations. The study highlights the complexity of AMDV circulation in feral mink populations and implicates a potential spillover of the virus to native species.

Multispecies reservoir of Spirometra erinaceieuropaei (Cestoda: Diphyllobothridae) in carnivore communities in north-eastern Poland.

BACKGROUND: Spirometra erinaceieuropaei is a diphylobothriid tapeworm with a complex life-cycle including definitive, intermediate and paratenic (transport) hosts. Multiple routes of parasite transmission often make it impossible to determine what type of host a specific infected animal is considered to be. Spargana larvae cause sparganosis, a severe food- and water-borne disease mainly found in Asia. In Poland, Spirometra sp. was reported in large carnivores in Bialowieza Primeval Forest for the first time in the 1940s and was recently confirmed as S. erinaceieuropaei in several mammals and snakes using molecular methods. METHODS: In total, 583 carcasses of 9 carnivore species were necropsied between 2013 and 2019 in north-eastern (NE) Poland. The larvae of S. erinaceieuropaei (spargana) were isolated from subcutaneous tissue, counted, and preserved for genetic analyses. We calculated the prevalence and intensity of infection. To assess spatial variation in S. erinaceieuropaei infection probability in NE Poland, we applied a generalized additive model (GAM) with binomial error distribution. To confirm the species affiliation of isolated larvae, we amplified a partial fragment of the 18S rRNA gene (240 bp in length). RESULTS: Spirometra larvae were found in the subcutaneous tissue of 172 animals of 7 species and confirmed genetically as S. erinaceieuropaei. The overall prevalence in all studied hosts was 29.5% with a mean infection intensity of 14.1 ± 33.8 larvae per individual. Native European badgers and invasive raccoon dogs were characterized by the highest prevalence. An analysis of parasite spread showed a spatially diversified probability of infection with the highest values occurring in the biodiversity hot spot, Bialowieza Primeval Forest. CONCLUSIONS: Our study revealed that various mammal species (both native and non-native) can serve as S. erinaceieuropaei reservoirs. The frequency and level of infection may differ between selected hosts and likely depend on host diversity and habitat structure in a given area. Further studies are needed to assess the distribution of the parasite throughout Europe and the environmental and biological factors influencing infection severity in wild mammals.

Seroprevalence of Echinococcus spp. and Toxocara spp. in Invasive Non-native American Mink.

Invasive non-native species can become reservoirs of zoonotic pathogens and cause their spread during colonization, increasing the risk of zoonoses transmission to both wild hosts and humans. American mink (Neovison vison) are considered an important invasive mammal species responsible for carrying endoparasites. The aim of our study was to evaluate the role of feral American mink as a possible transmission vector of Echinococcus spp. and Toxocara spp. in wildlife. We analysed the frequency of American mink exposure to both parasites, the spatial distribution in Poland, and the variability over time on the basis of specific antibody presence using ELISA and Western blot. Alimentary tract analyses revealed that American mink do not serve as definitive hosts for these parasites. Altogether, 1100 American mink were examined. The average seropositivity for American mink was 14.2% for echinococcosis and 21.7% for toxocarosis; dual-seropositivity was detected in only 6.0%. Seroprevalence of both parasites differed between study sites and significantly increased over time in Toxocara spp. Thus, our study revealed that free-living American mink are exposed to parasites and likely to be involved in the maintenance of both Echinococcus spp. and Toxocara spp. in the wild as paratenic hosts.

Large lungworms (Nematoda: Dictyocaulidae) recovered from the European bison may represent a new nematode subspecies.

Although the Dictyocaulus lungworm, the agent of dictyocaulosis, is one of parasitological threats to European bison, its systematic position remains unclear. The aim of the present study was to evaluate the morphological features of the lungworm and the pathological lesions it induces, and to analyse mitochondrial (mt) genetic markers for systematic and molecular epidemiological studies. The morphological findings indicate that Dictyocaulus lungworms of European bison can be distinguished from those of cattle on the basis of differences in buccal capsule wall length, total body length, and spicules length in males, all of which were significantly longer in those of European bison. Nucleotide diversity calculated from pairwise sequence alignments of partial cytochrome c oxidase subunit 1 (cox1), cytochrome B (cytB) and NADH dehydrogenase subunit 5 (nad5) of specimens from cattle and European bison varied from 1.7% for nad5, 2.1% for cytB, to 3.7% for cox1 gene. Thus, among the lungworms of European bison and cattle, nad5 and cytB were the most conserved proteins, whereas cox1 was the most diverse. The mt cytB marker gene may be a suitable candidate for distinguishing between the two genotypes, as nad5 demonstrated the greatest within-genus sequence variation. The lung tissue of infected European bison manifests signs of verminous pneumonia characterized by interstitial pneumonia, bronchitis and bronchiolitis. Therefore, it appears that European bison and cattle are infected with slightly diverged, morphologically-different, genotypes of D. viviparus, indicating they belong to two separate worm populations. We propose, therefore, that the lungworm of European bison should be classified as D. viviparus subsp. bisontis.

The Nematodes Thelazia gulosa Railiet and Henry, 1910 and Thelazia skrjabini Erschov, 1928 as a Cause of Blindness in European Bison (Bison bonasus) in Poland.

PURPOSE: The nematodes of the genus Thelazia are the cause of eye diseases of wild and domestic ruminants throughout the world. The aim of the study was to describe clinical cases of thelasiosis in European bison (Bison bonasus) in Poland, and provide morphometrical features of Thelazia gulosa Railiet and Henry, 1910 and Thelazia skrjabini Erschov, 1928 regarded as potentially useful for species differentiation METHODS: The conjunctival sacs, tear ducts, the surface of the cornea and nicitating membrane collected from bison were rinsed with saline solution. Any nematodes isolated from the sediment were subjected to morphometric analysis. RESULTS: Thirteen of the 16 examined European bison were infected with Thelazia nematodes, belonging to the species T. gulosa and T. skrjabini. The intensity of infection ranged from one to six (mean intensity 5), and four to 29 (mean intensity 14) nematodes T. skrjabini and T. gulosa respectively. Congestion of conjunctival sac, keratitis and corneal opacity, corneal ulceration and perforation as well as purulent eyeball inflammation were observed in infected animals. CONCLUSIONS: Thelazia gulosa and T. skrjabini can be identified by morphometrical features. As thelasiosis might be a serious threat for protected population of European bison, further studies are needed of the epidemiology and pathology of this emerging parasitosis in Poland.

Rodents as intermediate hosts of cestode parasites of mammalian carnivores and birds of prey in Poland, with the first data on the life-cycle of Mesocestoides melesi.

BACKGROUND: Rodents constitute an important part of the diet of many carnivore species. This predator-prey food chain is exploited by helminth parasites, such as cestodes, whose larval stages develop in rodents and then mature to the adult stage in predators. The main aim of our study was to use molecular techniques for identification of cestode species recovered from both intermediate and definitive hosts, with a particular focus on the genus Mesocestoides. METHODS: Larval cestodes were obtained during our long-term studies on rodent helminth communities in the Mazury Lake District in the north-east Poland in 2000-2018. Cestode larvae/cysts were collected from body cavities or internal organs (e.g. liver) during autopsies. Adult tapeworms were derived from nine red foxes, three Eurasian badgers and one Eurasian lynx. PCR amplification, sequencing and phylogenetic analyses were conducted employing three genetic markers: 18S rDNA, mitochondrial (mt) 12S rDNA and the mt cytochrome c oxydase subunit 1 (cox1) gene fragment. RESULTS: Altogether 19 Mesocestoides samples were analyzed, including 13 adult tapeworms from definitive hosts and six larval samples from 4 bank voles and 2 yellow-necked mice. Phylogenetic analyses revealed three well-supported trees of similar topology. In each case the Mesocestoides samples formed two separate clades. All isolates from foxes, the lynx isolate and two isolates from rodents grouped with Mesocestoides litteratus. Four isolates from rodents and all three isolates from Eurasian badgers were resolved in a separate clade, most similar to North American M. vogae (syn. M. corti). Examination of fixed, stained adult specimens from Eurasian badgers revealed consistency with the morphology of Mesocestoides melesi. Therefore, this clade is likely to represent M. melesi, a species first described in 1985 from the Eurasian badger Meles meles. Molecular analysis allowed also the identification of Taenia crassiceps, Hydatigera kamiyai and Cladotaenia globifera among larvae derived from rodents. CONCLUSIONS: Molecular and phylogenetic analyses support the recognition of M. melesi as a valid species. Our data represent the first record of the larvae of this species in rodents. This is the first report on the occurrence of H. kamiyai in rodents from Poland.