Mitochondrial genetic diversity and phylogenetic relationships of Echinococcus multilocularis in Europe.

The cestode Echinococcus multilocularis is the causative agent of alveolar echinococcosis, a fatal zoonotic parasitic disease of the northern hemisphere. Red foxes are the main reservoir hosts and, likely, the main drivers of the geographic spread of the disease in Europe. Knowledge of genetic relationships among E. multilocularis isolates at a European scale is key to understanding the dispersal characteristics of E. multilocularis. Hence, the present study aimed to describe the genetic diversity of E. multilocularis isolates obtained from different host species in 19 European countries. Based on the analysis of complete nucleotide sequences of the cob, atp6, nad2, nad1 and cox1 mitochondrial genes (4,968 bp), 43 haplotypes were inferred. Four haplotypes represented 62.56 % of the examined isolates (142/227), and one of these four haplotypes was found in each country investigated, except Svalbard, Norway. While the haplotypes from Svalbard were markedly different from all the others, mainland Europe appeared to be dominated by two main clusters, represented by most western, central and eastern European countries, and the Baltic countries and northeastern Poland, respectively. Moreover, one Asian-like haplotype was identified in Latvia and northeastern Poland. To better elucidate the presence of Asian genetic variants of E. multilocularis in Europe, and to obtain a more comprehensive Europe-wide coverage, further studies, including samples from endemic regions not investigated in the present study, especially some eastern European countries, are needed. Further, the present work proposes historical causes that may have contributed to shaping the current genetic variability of E. multilocularis in Europe.

Human alveolar echinococcosis in Slovakia: Epidemiology and genetic diversity of Echinococcus multilocularis, 2000-2023.

Human alveolar echinococcosis (AE) is a serious parasitic disease caused by larval stages of Echinococcus multilocularis. Between January 2000 and October 2023, 137 AE cases were confirmed in Slovakia. The average annual incidence increased from 0.031 per 100,000 inhabitants between 2000 and 2011, to an average of 0.187 since 2012, i.e. about six times. Among patients, 45.3% were men and 54.7% were women; the mean age at the time of diagnosis was 52.8 years. Most cases were diagnosed in the age groups 51-60 years and 61-70 years (33 cases each), and eight patients fell into the age category ≤ 20 years. To better recognize the gene diversity in clinical samples, metacestodes from 21 patients collected between 2013 and 2021 were subjected to DNA sequencing of four mitochondrial genes. Using concatenated sequences of cob (603 bp), nad2 (882 bp) and cox1 (789 bp) gene fragments, 14 isolates (66.7%) were assigned to the European E5 profile of E. multilocularis, two isolates (9.5%) to the E5a subtype, four isolates (19%) to the E4 profile, and one isolate (4.8%) to haplogroup E1/E2. The E5-type profiles and E4 profiles were distributed throughout the country, whereas the E1/E2 profile was found in the patient from western Slovakia. According to the data obtained and GenBank sequences, the E5-type dispersal is so far limited to central-eastern Europe and the variant seems to be indigenous to that region. The admixture with the haplotypes E4 and E1/E2 could have taken place from a historical endemic focus during the fox expansion in the last decades. By employing the nad1 fragment, a typical European haplotype was observed in all 21 resolved Slovak samples. The acceleration in the AE incidence in the last decade suggests the emergence of the disease and the need for further research on human and animal isolates.

Zoonotic Marine Nematode Infection of Fish Products in Landlocked Country, Slovakia.

Fish products in Slovakia have been heavily infected with Anisakis spp. larvae, which causes human anisakiasis. We found larvae in all tested samples of frozen Atlantic herring. Anisakid allergen t-Ani s7 testing revealed 2 positive cases in humans, signaling need for health authorities to closely monitor zoonotic marine parasites, even in inland areas.

Rodents as Sentinels for Toxoplasma gondii in Rural Ecosystems in Slovakia-Seroprevalence Study.

Toxoplasma gondii is a ubiquitous intracellular parasite with felids as definitive hosts and a broad range of intermediate hosts. Rodents are considered suitable sentinels for prevalence studies of many infections, including toxoplasmosis. This study aimed to estimate the seroprevalence of T. gondii in rodents from different localities of Slovakia and investigate the correlation between the seropositivity and the species, age, sex, and sexual activity of animals. Altogether, 1009 wild rodents belonging to 9 species were trapped in 2015 and 2019, and antibodies to T. gondii were detected in 6.7% of the animals. Seropositivity was detected in seven species, ranging from 0.0% in Micromys minutus and Apodemus sylvaticus to 7.7% in A. flavicollis. The females reached significantly higher seropositivity (9.7%) than the males (3.8%), and the adults were positive significantly more often (9.2%) than the subadults (4.9%). The seropositivity differed also among localities, with significantly higher positivity detected in suburban and touristic areas (12.2%) than in localities with a lower level of human activities (5.5%). This study showed that the occurrence of T. gondii varies significantly in rodent species and habitats with various environmental conditions and different levels of anthropic use. Several biological and ecological factors, e.g., soil contamination, soil conditions, the susceptibility of rodent species etc., may influence this variability.

A Study of Oribatid Mites as Potential Intermediate Hosts of Anoplocephalid Tapeworms of Tatra chamois and Tatra marmots from the Tatra Mountains, Central Europe, and Report of a New Intermediate Host for Andrya cuniculi, the Parasite of Leporidae.

Tatra chamois (Rupicapra rupicapra tatrica (Blahout 1972)) and Tatra marmot (Marmota marmota latirostris (Kratochvíl 1961)) are significant endemic subspecies of the subalpine and alpine ranges of the Tatra Mountains in Central Europe. In four studied localities in the range of their typical biotopes in Slovakia and Poland, we investigated intestinal parasites of Tatra chamois and Tatra marmots, with an emphasis on anoplocephalid tapeworms. We also studied the occurrence, species diversity, and abundance of oribatid mites as intermediate hosts thereof, and the prevalence of cysticercoid larval stages of anoplocephalid tapeworms in collected oribatids using morphological and molecular methods. Coprological analyses revealed the average positivity of Moniezia spp. in chamois faeces at 23.5% and Ctenotaenia marmotae in marmot samples at 71.1%, with significant differences between the localities under study. Morphological analyses determined the presence of cysticercoids in five oribatid species: Ceratozetes gracilis, Edwardzetes edwardsi, Scheloribates laevigatus, Trichoribates novus, and Tectocepheus velatus sarekensis. This is the first record of T. v. sarekensis as an intermediate host of anoplocephalid tapeworms, as well as the first report of Andrya cuniculi occurrence in the territory of the Tatra Mountains, confirmed also by molecular methods.

Grassland versus forest dwelling rodents as indicators of environmental contamination with the zoonotic nematode Toxocara spp.

Small mammals are suspected of contributing to the dissemination of Toxocara canis and helping with the parasite survival during periods when there is a temporary absence of suitable definitive hosts. While the primary aim of the current study was the assessment of seroprevalence of Toxocara spp. infections in wild rodents in Poland, we also explored the role of intrinsic (sex, age) and extrinsic factors (study site) influencing dynamics of this infection to ascertain whether grassland versus forest rodents play a greater role as indicators of environmental contamination with T. canis. We trapped 577 rodents belonging to four species (Myodes glareolus, Microtus arvalis, Microtus agrestis, Alexandromys oeconomus) in north-eastern Poland. Blood was collected during the parasitological examination, and serum was frozen at - 80 °C until further analyses. A bespoke enzyme-linked immunosorbent assay was used to detect antibodies against Toxocara spp. We found Toxocara spp. antibodies in the sera of all four rodent species with an overall seroprevalence of 2.8% [1.9-4.1%]. There was a significant difference in seroprevalence between vole species, with the grassland species (M. arvalis, M. agrestis and A. oeconomus) showing a 16-fold higher seroprevalence (15.7% [8.7-25.9%]) than the forest-dwelling M. glareolus (0.98% [0.5-1.8%]). We hypothesise that the seroprevalence of Toxocara spp. differs between forest and grassland rodents because of the higher contamination of grasslands by domestic dogs and wild canids. Our results underline the need for wide biomonitoring of both types of ecosystems to assess the role of rodents as indicators of environmental contamination with zoonotic pathogens.

Systematic Review and Modelling of Age-Dependent Prevalence of Toxoplasma gondii in Livestock, Wildlife and Felids in Europe.

Toxoplasma gondii is a zoonotic parasite of importance to both human and animal health. The parasite has various transmission routes, and the meat of infected animals appears to be a major source of human infections in Europe. We aimed to estimate T. gondii prevalence in a selection of animal host species. A systematic literature review resulting in 226 eligible publications was carried out, and serological data were analyzed using an age-dependent Bayesian hierarchical model to obtain estimates for the regional T. gondii seroprevalence in livestock, wildlife, and felids. Prevalence estimates varied between species, regions, indoor/outdoor rearing, and types of detection methods applied. The lowest estimated seroprevalence was observed for indoor-kept lagomorphs at 4.8% (95% CI: 1.8-7.5%) and the highest for outdoor-kept sheep at 63.3% (95% CI: 53.0-79.3%). Overall, T. gondii seroprevalence estimates were highest within Eastern Europe, whilst being lowest in Northern Europe. Prevalence data based on direct detection methods were scarce and were not modelled but rather directly summarized by species. The outcomes of the meta-analysis can be used to extrapolate data to areas with a lack of data and provide valuable inputs for future source attribution approaches aiming to estimate the relative contribution of different sources of T. gondii human infection.

Unveiling the incidences and trends of the neglected zoonosis cystic echinococcosis in Europe: a systematic review from the MEmE project.

The neglected zoonosis cystic echinococcosis affects mainly pastoral and rural communities in both low-income and upper-middle-income countries. In Europe, it should be regarded as an orphan and rare disease. Although human cystic echinococcosis is a notifiable parasitic infectious disease in most European countries, in practice it is largely under-reported by national health systems. To fill this gap, we extracted data on the number, incidence, and trend of human cases in Europe through a systematic review approach, using both the scientific and grey literature and accounting for the period of publication from 1997 to 2021. The highest number of possible human cases at the national level was calculated from various data sources to generate a descriptive model of human cystic echinococcosis in Europe. We identified 64 745 human cystic echinococcosis cases from 40 European countries. The mean annual incidence from 1997 to 2020 throughout Europe was 0·64 cases per 100 000 people and in EU member states was 0·50 cases per 100 000 people. Based on incidence rates and trends detected in this study, the current epicentre of cystic echinococcosis in Europe is in the southeastern European countries, whereas historical endemic European Mediterranean countries have recorded a decrease in the number of cases over the time.

Molecular identification of Taenia hydatigena from domestic and free-living animals in Slovakia, Central Europe.

Taenia hydatigena is a cosmopolitan tapeworm that uses canids or felines as definitive hosts, while the larval stage (metacestode), formerly referred to as cysticercus tenuicollis, infects a wide variety of intermediate hosts, in particular ruminants. In the present study, we used partial nucleotide sequences of the cox1 and nad1 genes of T. hydatigena from different animal species to analyse the intraspecies genetic diversity of this economically important parasite. Twenty-four samples of metacestodes or adults of T. hydatigena from infected sheep, chamois, roe deer, fallow deer, wild boar, and dogs from Slovakia were collected and further analysed. Several haplotypes of T. hydatigena were identified with unique mutations that have not been previously recorded in Slovakia. Analysis of nucleotide polymorphism revealed the existence of 9 and 13 haplotypes, with relatively low nucleotide pairwise divergence ranging between 0.3-1.3 and 0.2-1.8% for the Hcox and Hnad haplotypes, respectively. In general, low nucleotide and high haplotype diversities in the overall population of T. hydatigena from the study indicate a high number of closely related haplotypes within the explored population; nucleotide diversity per site was low for cox1 (Pi = 0.00540) and slightly higher for nad1 (Pi = 0.00898). A molecular study confirmed the existence of genetic variation within T. hydatigena isolates from Slovakia. However, further investigations with more samples collected from different intermediate and definitive hosts are required in order to investigate the epidemiological significance of the apparent genetic differences observed in this study.

Dirofilaria immitis Pulmonary Dirofilariasis, Slovakia.

Dirofilaria immitis is a parasite related to pulmonary dirofilariasis in humans, its accidental hosts. We detected an autochthonous case of D. immitis infection in a woman from Slovakia. The emergence and spread of this parasite in Europe indicates a critical need for proper diagnosis of infection.

Molecular Study of Echinococcus granulosus Cestodes in Ukraine and the First Genetic Identification of Echinococcus granulosus Sensu Stricto (G1 Genotype) in the Country.

INTRODUCTION: Cystic echinococcosis is a globally distributed zoonotic disease of great medical and veterinary importance, which is caused by the cestode Echinococcus granulosus sensu lato. In Ukraine, two areas of the prominent circulation of the parasite are established, the southern steppe zone with sheep as the main transmitter, and the northern forest-steppe zone and Polissia, where pigs are mainly responsible for maintaining the E. granulosus transmission. METHODS: Given that only a few studies have so far addressed the genetic diversity of the parasite in Ukraine, we have sequenced partial mitochondrial genes of cytochrome c oxidase 1 (789 bp), NADH dehydrogenase 1 (602 bp) and 12S rRNA (333-334 bp) in pig metacestodes from the Sumy region (farms close to Sumy, northeastern Ukraine) and the Kyiv region (a farm in Bila Tserkva, central Ukraine). RESULTS: Four isolates from four pigs in the Sumy region were identified as E. canadensis (G7 genotype), the major E. granulosus s.l. species circulating in Eastern Europe, including the three microvariants (G7A, G7B, G7C). Three isolates from the two pigs in the Kyiv region were classified as E. granulosus s.s. (G1 genotype), including one microvariant (G1A). CONCLUSION: To our knowledge, this is the first genetic record of E. granulosus s.s. with the presumed highest infectivity and virulence among the E. granulosus s.l. species in Ukraine. The finding has implications for public health as local control programmes should take into consideration different development rate of this parasite in dogs and the greater risk of the species for human infection.

Parasitic and Vector-Borne Infections in HIV-Positive Patients in Slovakia-Evidence of an Unexpectedly High Occurrence of Anaplasma phagocytophilum.

In HIV (human immunodeficiency virus) infected people, the immunodeficiency caused by a reduced level of CD4 (cluster of differentiation 4) T-lymphocytes increases the risk of infectious diseases. Additionally, in individuals with immunologically compromising conditions, tick-borne or some parasitic pathogens may cause chronic, debilitating opportunistic infections and even death. The study aimed at determining the IgG seropositivity of HIV-infected patients to Toxoplasma gondii, Toxocara spp., Echinococcus multilocularis, and E. granulosus s.l. and performing the molecular identification of T. gondii and some tick-borne pathogens, namely, Borrelia spp., Babesia spp., Anaplasma phagocytophilum, Rickettsia spp., and Bartonella spp. Out of 89 HIV-positive patients, specific IgG antibodies to T. gondii were detected in 17 (19.1%) and to Borrelia spp. in 12 (13.5%) individuals. Seropositivity to Toxocara spp., E. multilocularis, and E. granulosus s.l. was not recorded. Molecular approaches showed positivity to T. gondii in two (2.2%) patients, and 11 (12.4%) individuals had positive PCR signal for the msp2 gene of A. phagocytophilum. Relatively high prevalence of A. phagocytophilum in HIV-positive patients suggests that these people are more susceptible to some vector-borne pathogens. The presence of opportunistic infections may pose a health risk for patients with weakened immune systems, and should not be neglected during the regular monitoring of the patient's health status.

A Survey of Intestinal Helminths of Dogs in Slovakia with an Emphasis on Zoonotic Species.

Dogs are the most popular pets worldwide; however, close contact with people increases the risk of transmission of different zoonotic parasites. This study aims to determine the prevalence of gastrointestinal helminths in dogs in Slovakia. A total of 495 faecal samples collected from pet, shelter, guard, working (police), and hunting dogs, as well as dogs from segregated Roma settlements between 2016 and 2021, were examined using flotation and molecular methods. Eggs of intestinal helminths were detected in 134 (27.1%) samples. Microscopically, six different species/genera/families, namely, Toxocara canis (14.7%), Toxascaris leonina (1.6%), Trichuris vulpis (6.3%), Capillaria spp. (1.4%), Ancylostoma/Uncinaria spp. (8.3%), and taeniid eggs (4.0%), were recorded. Molecular analyses revealed infection with Echinococcus multilocularis in 2.2% of dogs and 0.4% of the animals were infected with Taenia hydatigena. The results showed a correlation between the occurrence of intestinal helminths and the availability of veterinary care, as dogs from Roma settlements and shelter dogs were the most often infected (66.7% and 39.2%, respectively). On the other hand, working animals were in the best health condition, with only 2.5% being positive. The relatively frequent occurrence of zoonotic species points to the constant need for preventive measures and regular deworming of dogs.

Seroprevalence of Toxoplasma gondii among Sylvatic Rodents in Poland.

Toxoplasma gondii is an intracellular Apicomplexan parasite with a broad range of intermediate hosts, including humans and rodents. Rodents are considered to be reservoirs of infection for their predators, including cats, felids, pigs, and wild boars. We conducted a multi-site, long-term study on T. gondii in northeastern Poland. The study aimed to monitor the seroprevalence of T. gondii in the four abundant vole species found in the region (Myodes glareolus, Microtus arvalis, Microtus agrestis, and Alexandromys oeconomus) and to assess the influence of both extrinsic (year of study and study site) and intrinsic (host sex and host age) factors on seroprevalence. A bespoke enzyme-linked immunosorbent assay was used to detect antibodies against T. gondii. We examined 577 rodent individuals and detected T. gondii antibodies in the sera of all four rodent species with an overall seroprevalence of 5.5% [4.2-7.3] (3.6% [2.6-4.9] for M. glareolus and 20% [12-30.9] for M. arvalis, M. agrestis, and A. oeconomus). Seroprevalence in bank voles varied significantly between host age and sex. Seroprevalence increased with host age and was higher in females than males. These results contribute to our understanding of the distribution and abundance of T. gondii in voles in Poland and confirm that T. gondii also circulates in M. glareolus and M. arvalis, M. agrestis and A. oeconomus. Therefore, they may potentially play a role as reservoirs of this parasite in the sylvatic environment.

Trichinellosis in Slovakia - epidemiological situation in humans and animals (2009-2018).

INTRODUCTION: Trichinella spp. are zoonotic parasitic nematodes with almost worldwide distribution. The infection can be transmitted through the foodborne route and can cause serious health problems in infected human patients. It is also an economically important issue due to the high financial cost connected with prevention of the disease. OBJECTIVE: The study aimed to discover trends in the epidemiological situation in people and animals in Slovakia in 2009-2018. MATERIAL AND METHODS: Data on human trichinellosis originated from the Public Health Authority of the Slovak Republic, and data on Trichinella infection in animals from the State Veterinary and Food Administration of the Slovak Republic. A seroepidemiological study on 655 voluntary respondents was performed. RESULTS: Altogether, 29 human cases were reported between 2009-2018, with a maximum of 13 cases in 2011. Males were affected more often (19 cases) than females (10 cases); the average age of patients was 45.1 years. Antibodies to Trichinellawere detected in two sera (0.3%): in a serum from one hunter and one veterinarian. In the monitored period, the average prevalence of Trichinella infection was 9.6% in 2,295 red foxes, and 0.04% in 165,643 wild boars. Three (1.7%) of 178 surveyed brown bears were positive. Within the compulsory monitoring of trichinellosis in domestic pigs, none from a total of 1,632,688 pigs were positive. The presence of three species, Trichinellabritovi, T. spiralis and T. pseudospiralis, was documented, with T. britovi representing 93.9% of identified isolates. CONCLUSIONS: The study indicates that the prevalence of Trichinella infection has not changed considerably with time in Slovakia, and the risk of human infection outbreaks is connected mainly with the consumption of wild boar meat.

Ocular Dirofilariasis after Clinically Manifested Subcutaneous Migration of the Parasite: A Case Report.

Dirofilaria repens is the causative agent of human subcutaneous or, less often, ocular dirofilariasis. The work presents a rare case of ocular dirofilariasis manifested by previous subcutaneous migration accompanied by severe headache symptoms. In February 2017, a 58-yr-old man from Trnava region, western Slovakia, noticed red and itchy stripes on his left leg. Inflamed but painless stripes disappeared and showed up again every 5-7 days, migrating gradually towards the head. Approximately one month after the first skin's alterations, strong pain in the left temple, with the swelling of the left face and the enlargement of mandibular lymph nodes appeared. Several days later, the patient felt excruciating pain of the right eyeball accompanied by strong nausea and subsequent vomiting. Ocular examination revealed the presence of a live worm in the subconjunctival space and morphological and molecular analyses of extracted helminth confirmed D. repens as etiological agent of the infection. According to clinical manifestation of the infection, it could be supposed that ocular form of the disease was the result of the migration of a parasite through the subcutaneous tissues. Moreover, a rare phenomenon of lymphadenitis of underlying lymph nodes and the swelling of left face accompanied the migration.

Human Taeniases in Slovakia (2010-2019): Genetic Analysis of Taenia saginata Isolates.

Taenia saginata and Taenia solium, known as beef and pork tapeworm, are foodborne pathogens of global importance having a substantial impact on human health and economy. The study aimed to summarize the occurrence of human Taenia spp. infection in past 10 years in Slovakia based on reports of clinicians and diagnostic laboratories to Public Health Authority of the Slovak Republic. Altogether, 19 human cases were reported in Slovakia in 2010-2019, with the incidence of infection ranging from 0.00 to 0.12/100,000 inhabitants per year. In two patients T. saginata infection was confirmed molecularly. Nucleotide sequences of the analyzed nad1 gene fragments derived from both patients were identical and in a phylogenetic tree clustered together with T. saginata Tsa isolate (AM503345), as well as with a homologous sequence of the completely sequenced mitochondrial genome of T. saginata. Similarly, cox1 nucleotide sequences derived from one of the patients and isolates from Asia and/or Europe were identical. A cluster for cox1 partial gene sequence was placed separately from closely related Taenia asiatica and/or T. solium isolates in the phylogenetic tree.

The cross-sectional study of Toxoplasma gondii seroprevalence in selected groups of population in Slovakia.

Toxoplasma gondii is an obligate intracellular protozoan parasite with domestic or free-living members of family Felidae known as the only definitive hosts of the parasite. The study aimed to evaluate the seroprevalence of toxoplasmosis in selected population groups in Slovakia and to analyse the infection risk factors. Totally 1536 serum samples of voluntary respondents were examined by EIA Toxoplasma IgG (Test-Line Ltd., Clinical Diagnostics, Brno, Czech Republic). IgG antibodies to T. gondii were detected in 322 (21.0%) sera, with the highest seropositivity recorded among farmers (42.5%) and hunters (28.5%). According to age, the highest seropositivity was recorded in 12-18-year-old Roma children (46.2%), what confirms increased risk of infection in this minority that often lives in inadequate hygiene conditions. When only adult participants were taken into account, the highest seroprevalence was observed in the age group of ≥ 70 years. Higher risk of infection (p < 0.05) was connected with cat ownership, living in rural residency and consumption of raw meat and meat processing. The results underline the importance of preventive measures and ongoing need to improve the knowledge on toxoplasmosis in both professionals and public.

Seroepidemiology of human toxocariasis in selected population groups in Slovakia: A cross-sectional study.

BACKGROUND: Human toxocariasis is a cosmopolitan helminthic zoonosis caused by roundworms of the genus Toxocara. The study aimed at a cross-sectional analysis of seropositivity to Toxocara spp. in selected groups of population in Slovakia and an estimate of the risk factors of infection. METHODS: For the detection of anti-Toxocara antibodies, an enzyme-linked immunosorbent assay (ELISA) based on an excretory-secretory (E/S) antigen of Toxocara canis was used. RESULTS: The overall seropositivity to Toxocara in 1489 evaluated sera was 3.7%. The seropositivity of Roma children/young people (40.3%) was significantly higher when compared with all other analysed groups. Among adult respondents, the positivity of farmers and hunters (5.5% and 5.1%, respectively) was significantly higher (p<0.001) than in the other analysed groups (veterinarians and veterinary assistants, soldiers, students of veterinary medicine, and students of other universities). Analysis of infection risk factors showed that living in a rural locality, everyday contact with soil, and everyday outdoor activities (12.7%, 8.8% and 8.3% positivity, respectively) can increase the risk of infection, but the observed differences were not significant. CONCLUSION: The results underline the importance of preventive measures and the need for improving of knowledge of toxocariasis among professionals, as well as the public.