Zuzana Vasilková

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.

The seasonal dynamics of the prevalence, abundance, and mean intensity of egg excretion by the nematode parasite Baylisascaris transfuga in the European brown bear (Ursus arctos) were monitored relative to environmental factors (mean temperature, humidity, and temperature) over three years. The prevalence, abundance, and mean intensity of egg excretion tended to increase from spring to autumn throughout the monitoring period. The seasonal prevalence (84.2–90.6%), abundance (470.2–545.3 eggs per gramme (EPG) of faeces), mean intensity of excretion (558.3–602.1 EPG), and number of eggs (1150 EPG) were highest in autumn. The prevalence of eggs (up to 48.5%), abundance (37.8–60.6 EPG), and mean intensity of excretion (94.4–125.0 EPG) were high in winter, despite the period of hibernation. The seasonal dynamics of B. transfuga abundance in bears, the mean temperature between spring and autumn, and the seasonal trend of increase in intensity of egg excretion with temperature from winter to summer were interrelated. Abundance differed significantly between winter and autumn, spring and autumn, and summer and autumn (p < 0.0001) in all years and between spring and summer in 2016 (p < 0.005), 2017 (p < 0.05). B. transfuga abundance differed significantly between the seasons over the three years only in spring (p ≤ 0.0001).

Summary Dirofilaria immitis, the agent of canine dirofilariosis, is a common parasite of domestic and wild carnivores with zoonotic potential and worldwide distribution, being endemic in many countries. Bulgaria is among European countries recognized as endemic for this heartworm parasite. In the present study, D. immitis adults recovered from pulmonary arteries of domestic dog and golden jackal originating from the Pazardzhik region in southern Bulgaria, and from red fox originating from the Plovdiv region in central-southern Bulgaria, were genetically analyzed in nuclear targets. The first PCR amplification of the internal transcribed region 2 (ITS2) of the ribosomal DNA with previously published D. immitis-specific primers yielded single fragments in size of 302 bp that is characteristic for these heartworms. PCR products of three isolates, resulted from the second amplification of the 5.8S-ITS2 region (235 bp) with pan-filarioid primers, were subjected to direct DNA sequencing. Identical nucleotide composition was detected across the screened target region for these Bulgarian isolates. When the 5.8S-ITS2 sequences were phylogenetically compared to the GenBank-retrieved D. immitis sequences in a worldwide context, the neighbor-joining analysis has shown three discrete clades. The first clade was composed of D. immitis isolates from Europe (including the studied Bulgarian samples), Asia and South America, in the second clade samples from Asia and South America were placed, whereas the third clade was formed by two Brazilian dog isolates originated from the north and southeast part of the country. The purpose of the present study was to verify the taxonomic characterization of D. immitis nematodes from Bulgaria based on morphology and compare their genetic structure with filariae obtained from the different world regions using molecular assays. It also summarizes previous epidemiological and ecological studies on the parasite distribution and prevalences in different hosts and regions undertaken so far in Bulgaria.

Summary: Our study describes changes in haematological parameters in wild ruminants with parasitic infection. Six European mouflons (Ovis musimon), six fallow deer (Dama dama) and six roe deer (Capreolus capreolus) were experimentally infected with the resistant strain of the model parasite 8000 L3 Haemonchus contortus. The blood samples were collected on Day 0, 16, 37, 58, 77, and 99 of the experiment. Mild anaemia was observed in mouflons and roe deer while red blood cells increased in red blood counts (total erythrocytes, haematocrit and haemoglobin). As for the white blood cells count, leucopenia with neutrophilia and lymphopenia was recorded in mouflons, in the fallow deer and roe deer leucocytosis with neutropenia and lymphocytosis were observed. Changes in the dynamics of haematological parameters were statistically insignificant.

Helminth infections caused by Enterobius vermicularis have a cosmopolitan character and most often affect the paediatric pre-school and school age population. The presented study was conducted to determine the prevalence of E. vermicularis in the analyzed population of children in the Eastern Slovakia. The Graham’s scotch tape method was used to investigate the presence of Enterobius vermicularis eggs in 390 specimens. The analyzed set consisted of 218 girls and 172 boys, divided by age into three groups - aged from 5 months to 2 years, aged from 3 to 6 years, and aged from 7 to 15 years. Investigation of perianal scotch tapes of children for the presence of E. vermicularis eggs revealed the prevalence of E. vermicularis was P = 3.59 %. Depending on the incidence of E. vermicularis infection, we detected no statistically signifi cant difference (p> 0.05). The prevalence of E. vermicularis in boys was P = 4.07 %, and in girls P = 3.21 %. The highest prevalence of E. vermicularis was recorded in the group of children aged from 3 to 6 years (P = 5.03 %). Most of the samples were positive at age 4 and 5. The lowest prevalence was in the group of children aged from 5 months to 2 years (P = 0.97 %), and the prevalence of E. vermicularis in the group of children aged from 7 to 15 was P = 3.91 %. The difference in the incidence of E. vermicularis infection among different age groups of children was not statistically significant (p> 0.05). Enterobius vermicularis nematode infection and enterobiasis currently represents a major public health problem in Slovakia. At the present its occurrence is the most frequent in the paediatric population. Therefore it is important to introduce a targeted hygienic-epidemiological measure in children’s collectives, what also should include proper and effective diagnostics and frequent recurrent therapy.

The aim of the research was to determine the prevalence of non-pathogenic protozoa circulating in the human population of Slovakia. We particularly focused on the socially deprived areas with poor sanitation conditions, as they are one of the factors affecting the transmission of these infections. Within this study, 2760 people were coprologically screened for the presence of protozoan cysts. The analyzed group comprised 1173 men and 1587 women from different regions of Slovakia. The total prevalence (2.03%) of non-pathogenic protozoa species was determined. The prevalence of Entamoeba coli was 0.80%, the prevalence of Endolimax nana 0.58%, and the prevalence of Blastocystis hominis was 0.65%. The presence of non-pathogenic protozoa was more frequent in women than that in men, in all age groups. The highest incidence of Entamoeba coli was found in children aged one month – seven years (0.79%), the lowest in the age group of 19–88 years (0.66%). Endolimax nana was most frequent in 8–18 year-olds (0.95%), where the statistical significance was found (p0.05) for Entamoeba coli, and similarly for Blastocystis hominis associated with the sex and age. Although the circulation of non-pathogenic protozoa in the human population is far from being limited to the developing countries, their occurrence is also frequent in the population of developed countries. Despite their controversial pathogenicity, they should not be neglected, particularly in the patients with gastrointestinal symptoms.

Summary: Cystic echinococcosis (CE) and alveolar echinococcosis (AE) are considered as one of the most important zoonotic diseases in Romania, where they are subject to mandatory reporting. To obtain more knowledge about the genetic diversity of Echinococcus causative agents of these diseases, 11 isolates from humans and ungulate intermediate hosts from the two regions of Romania were genotyped using mitochondrial markers. In clinical samples of five patients from north-eastern Romania (Iasi, Botosani, Vaslui counties), Echinococcus multilocularis was identified as causal agent by cox1 sequence analysis. To the best of our knowledge this finding presents the first molecular evidence of E. multilocularis in humans from Romania. Only two cases of AE in patients were previously documented in the country by serological methods. In our four patients the most widespread European variant E5 of E. multilocularis was recorded, whereas in isolate from Vaslui county three nucleotide substitutions were detected as compared to the most related E5 haplotype. One of these mutations (411T/G) matched N1 and N2 haplotypes described previously from North America. In six CE samples retrieved from western Romania (Caras-Severin and Timis counties), two human isolates were diagnosed as Echinococcus canadensis G7, one as E. granulosus s.s. G1 and one as E. granulosus s.s. G3 using atp6 and rrnS sequencing. In ungulates, the cattle isolate was allocated to E. granulosus s.s. G1 and pig isolate to E. canadensis G7. The two G7 findings in humans reinforced the recent view that G7 was underestimated as compared to the E. granulosus s.s. regarding human CE threat that can be further employed for identifying sources of infections and establishing suitable preventive measures.

The present study evaluates the effect of plant extracts of oregano and commercial Xtrakt ^TM administrations on microbial, biochemical, immunological and nutritional indicators and on Eimeria sp. oocyst occurrence in rabbits. Rabbits (5 weeks old, Hy-plus hybrid, n = 66) were divided into experimental group 1 (E1) with oregano extract application, experimental group 2 (E2) with Xtract ^TM application and control group. Natural substances were administered for the first 21 days. The experiment lasted for 42 days. The antibacterial effect was determined by the decrease of coagulase-positive staphylococci in E1 compared to control at day 42. Staphylococcus aureus cells were detected in lower counts in E2 compared to control at day 21. The counts of Clostridium- like bacteria were lower in both experimental groups at day 21 compared to day 7 (difference 1.2 and 1.3 log cycles, respectively) and to control (difference 0.5 and 0.3 log cycles, respectively). At day 7, the counts of coliforms in E1 were significantly lower than in E2 ( P < 0.01). In rabbits fed with oregano, reduction of Eimeria sp. oocysts and higher value of phagocytic activity (21.6 ± 0.51 %) were found compared to Xtract ^TM , and prolonged immuno-stimulatory effect was noted. In the animals of both experimental groups higher final weight, feed conversion ratio and lower mortality were achieved compared to control. The administration of oregano showed antibacterial, anticoccidial, and immunomodulatory effects. The results showed that oregano administration may be used as an alternative prophylactic measure in rabbits.