Cystic echinococcosis in Poland: genetic variability and the first record of Echinococcus granulosus sensu stricto (G1 genotype) in the country.

Cystic echinococcosis is one of the most important zoonotic diseases affecting humans and livestock worldwide, and is endemic in Poland. A set of six isolates on larval stages of Echinococcus granulosus sensu lato tapeworms collected from three humans, two pigs and one sheep from Polish foci of CE was examined by DNA sequencing of two mitochondrial genes (cox1, rrnS). The results demonstrated the presence of E. canadensis and E. granulosus sensu stricto in the investigated hydatid cysts. The former species was found in all five isolates from pigs and humans derived from central Poland. In a sheep hydatid cyst originating from Lesser Poland Voivodeship in southern Poland, E. granulosus s. s. (G1 genotype) was identified. This is the first report of an unambiguously autochthonous infection with E. granulosus s. s. in Poland. The global distribution and host affiliations of the commonly occurring G1 microvariant with nucleotide change 56C/T in cox1, detected here in Polish sheep, are discussed. The finding that sheep harboured E. granulosus s. s. may have important consequences for developing effective hydatid control programmes in Poland due to its longer maturation rate in dogs compared with E. canadensis G7. This may lead to greater expenditures for purchasing anthelmintics to provide an appropriate dosing regime in sheep-raising areas of the country.

Intestinal Helminth Communities of Grey Partridge Perdix perdix and Common Pheasant Phasianus colchicus in Poland.

The aim of this study was to describe the morphology and means of identification of helminths in native partridges (65) and introduced pheasants (32) in Poland and to determine the level of intestinal infection of these birds by helminths using parasitological and ecological indices. The birds were acquired during the hunting season in the years 2015-2017. Nematodes, Capillaria phasianina, cestodes, Railietina friedbergeri, and one trematode, Brachylaima sp. were recorded for the first time in partridges in Poland. Our findings indicate that parasites are more prevalent in pheasants (prevalence 70.4%) than in partridges (prevalence 50.0%). The component community and infracommunity of parasites of partridges are more diverse (Simpson's diversity index: 0.63 and mean Brillouin diversity index: 0.10 ± 0.17) and less dominated by a single parasite species (Capillaria sp., Berger-Parker dominance index: 0.53) than the pheasant parasite community (Simpson's diversity index: 0.07, mean Brillouin diversity index: 0.005 ± 0.02, dominant species Heterakis gallinarum, Berger-Parker dominance index: 0.96). There were statistically significant differences between partridges and pheasants in the Brillouin diversity index and in the prevalence of Heterakis gallinarum (55.6% in pheasants vs. 19.0 in partridges). There were significant differences between wild and farmed partridges in the prevalence of infection by Capillaria sp. (4.3% vs. 37.5%) and H. gallinarum (39.1 vs. 6.2%). In conclusion, the pheasant was shown to be a reservoir, carrier, and shedder of nematodes, which may increase the risk of infection in partridges.

Very low intraspecific sequence variation in selected nuclear and mitochondrial Parascaris univalens genes.

Equines were over decades considered to be infected by two morphologically virtually indistinguishable ascarid species, Parascaris univalens and Parascaris equorum. Reliable species discrimination is only possible using enzyme isoelectric focussing and karyotyping with P. univalens having one and P. equorum two chromosome pairs. However, presumably the complexity of both methods prevented their routine use in nearly all previous studies about prevalence and drug resistance of Parascaris spp. These have barely been performed on the species level although most studies stated presence of one or the other species. Recently, only P. univalens has been identified by karyotyping and the last published study identifying P. equorum dates back to 1989. In order to improve species-specific detection, molecular markers are required. Here, partial 12S rRNA, cytochrome oxidase I (COI) and complete internal transcribed spacer (ITS)-1 and - 2 sequences were obtained from 24 karyotyped Parascaris specimens from Poland and 6 German specimens (not karyotyped) and used in phylogenetic analyses with orthologous sequences from GenBank. All karyotyped specimens were identified as P. univalens. In the phylogenetic analysis, they formed very homogenous clusters for all target genes and in a multi-locus analysis. Within this cluster, almost all sequences from GenBank were also included, no matter if they had been assigned to P. univalens or P. equorum. However, a small number of P. univalens ITS and COI sequences originating from donkeys from a single farm in China formed a highly supported sister cluster suggesting that they might represent another Parascaris genotype or species. Our data also strongly suggest that nearly all ITS and COI sequences previously deposited in GenBank and assigned to P. equorum actually represent P. univalens. The fact that significantly different sequences can be found in Parascaris spp. suggests that PCR-based species diagnosis will be possible once molecular markers have been identified for P. equorum from karyotyped specimens.

[The case of the nematode Setaria equina found in the vaginal sac of the stallion's scrotum]. / Przypadek obecnosci nicienia Setaria equina w jamie pochwowej worka mosznowego ogiera.

The nematode Setaria equina usually reside in body cavities and do not cause clinical symptoms. From time to time, however, these parasites can be located in the scrotum and spermatic cord inflicting pain and edema in these body parts. The aim of the study was to describe the case of the nematode Setaria equina found in the vaginal sac of the stallion's scrotum. During the study, thorough examination of 50 isolated testicles of 25 stallions was conducted. The horses were obtained post-slaughter from the local slaughterhouse near Krakow. In one of examined stallions, two females of Setaria equina of 80 and 110 mm in length were found attached to the surface of the testicle. A part of one of the nematode was found in the space between the surface of the testicle and the epididymis. During the examination of the testicle sections, chronic inflammation (orchitis chronica) and focal necrosis perivascularis were found.

[The influence of Taenia taeniaeformis larval infection on morphometrical parameters of muskrat (Ondatra zibethicus)]. / Wplyw zarazenia larwami tasiemca Taenia taeniaeformis na parametry morfometryczne pizmaków (Ondatra zibethicus).

An investigation aimed to check the influence of Taenia taeniaeformis larvae on morphometrical parameters of muskrat (Ondatra zibethicus) was carried. A total of 30 animals were hunted down in upper Vistula river basin in south Poland, then measured, weighed and dissected. Statistical comparison were done using U Mann-Whitney test. T. taeniaeformis larvae--cysticercus fasciolaris was found in the liver of 24 muskrats (80%). Significant differences between infected and non infected animals are reported, as regards their body mass, total length, abdomen circumference (p < 0.01) and also in body length (total minus tail length), head length, or chest and neck circumference (p < 0.05). The effect of infection on both muskrat condition and the presence of adult cestodes in definitive hosts are discussed.

The Prevalence of Endoparasites of Free Ranging Cats (Felis catus) from Urban Habitats in Southern Poland.

Growing urbanization leads to an increased risk of parasite spread in densely inhabited areas. Free-ranging cats can be locally numerous and come into frequent contact with both wildlife and humans. Cats are thus expected to contribute to parasitic disease transmission. In our study, we investigated the prevalence of endoparasites in free ranging cats in urban areas of Kraków city, based on necropsy of road-killed cats in relation to sex and diet of cat, season and habitat type. We found that 62% of 81 cats were infected with endoparasites with Toxocara cati being the most prevalent. In total, we identified seven parasite species. The number of parasite species was higher in suburban habitats and aside from Eucoleus aerophilus the prevalence of all parasites was higher in cats from suburban areas than in the individuals living in the city urban core. The prey of examined cats included mostly rodents, followed by soricomorphs and birds, which can all serve as paratenic hosts. Based on our results, we suggest that cats in urban areas should be considered as a serious potential zoonotic threat. Implementation of proper veterinary control and wider education on the topic is recommended.

The P-glycoprotein repertoire of the equine parasitic nematode Parascaris univalens.

P-glycoproteins (Pgp) have been proposed as contributors to the widespread macrocyclic lactone (ML) resistance in several nematode species including a major pathogen of foals, Parascaris univalens. Using new and available RNA-seq data, ten different genomic loci encoding Pgps were identified and characterized by transcriptome-guided RT-PCRs and Sanger sequencing. Phylogenetic analysis revealed an ascarid-specific Pgp lineage, Pgp-18, as well as two paralogues of Pgp-11 and Pgp-16. Comparative gene expression analyses in P. univalens and Caenorhabditis elegans show that the intestine is the major site of expression but individual gene expression patterns were not conserved between the two nematodes. In P. univalens, PunPgp-9, PunPgp-11.1 and PunPgp-16.2 consistently exhibited the highest expression level in two independent transcriptome data sets. Using RNA-Seq, no significant upregulation of any Pgp was detected following in vitro incubation of adult P. univalens with ivermectin suggesting that drug-induced upregulation is not the mechanism of Pgp-mediated ML resistance. Expression and functional analyses of PunPgp-2 and PunPgp-9 in Saccharomyces cerevisiae provide evidence for an interaction with ketoconazole and ivermectin, but not thiabendazole. Overall, this study established reliable reference gene models with significantly improved annotation for the P. univalens Pgp repertoire and provides a foundation for a better understanding of Pgp-mediated anthelmintic resistance.

Morphometric identification of equid cyathostome (Nematoda: Cyathostominae) infective larvae.

The prevalence of infection with cyathosthomes of horses is high worldwide. Identification of species on infective larvae based on available morphological keys is not fully accurate and the aim of the present study was to provide a reliable identification key of cyathostomes infective larvae. At the abattoir, horse large intestines were examined and the cyathostomes females extracted and identified. The eggs from uteruses of identified cyathostomes females were used for breeding the infective larvae. Morphological parameters of Cyathostominae infective larvae, i.e. the length of the larvae with sheath, length of oesophagus, intestine, end of the intestine to the end of the larvae body, and finally the body breadth measured and the cells arrangement of the intestine cells (A-D) noted. These morphological parameters of the third infective larvae were determined of the following cyathostomes species: Cylicocyclus nassatus, Cylicostephanus goldi, Cyathostomum catinatum, Cylicostephanus longibursatus, Cyathostomum pateratum, Cylicocyclus ashworti, Cylicocyclus insigne, Cylicostephanus calicatus and Coronocyclus coronatus. The qualitative assignment into A, C and D intestinal type and then the combination of morphological measures allows specific identification for most species.

Tarantobelus arachnicida (Nematoda: Rhabditida) invasion of exotic spiders in Poland

Nematodes of the Brevibuccidae family were stated among spiders of the Theraphosidae family (the South American species), which were bred in Poland. The first sign of infection was anorexia which led to gradually increasing lethargy progressed to a huddled posture. Additionally, a white discharge with nematodes between mouth and chelicerae was noted. All of the derived nematodes were morphologically identified and determined to the species Tarantobelus arachnicida. A molecular analysis covered amplification and sequencing of small subunit ribosomal RNA (18S rRNA). A post mortem examination demonstrated the presence of nematodes not only near the chelicerae, but also inside the intestine, hence the source of infection might be insects used as a food. The research showed that such kind of infection is an important disease, which poses a serious risk to the breeding spiders. To date there is no effective treatment, however, we demonstrated that usage of the Lugol's solution seems to be promising.

Endoparasites of pet reptiles based on coprosopic methods

Due to the growing popularity of reptiles as a household animals and the development of numerous reptile farms, they have become a common sight in veterinary clinics. As parasitic infections represent a serious problem among pet reptiles obtained by captive breeding and from pet shops, the purpose of the present study was to determine the species composition of parasites present in reptiles bred privately or in Cracow Zoological Garden, and those obtained from pet shops. Fecal samples collected from 91 reptiles (30 turtles, 40 lizards, and 21 snakes) were examined using the quantitative McMaster method. Parasite eggs or protozoan oocysts were identified in 59.3% of samples. These included the eggs of the Pharyngodonidae, Ascarididae and Rhabditoidea (Nematoda), and Trematoda, as well as oocysts of Isospora and Eimeria. In addition, pseudoparasites belonging to the Mesostigmata, Demodecidae and Myobiidae were found.

Equine strongyle communities are constrained by horse sex and species dipersal-fecundity trade-off.

BACKGROUND: Equine strongyles are a major health issue. Large strongyles can cause death of horses while cyathostomins (small strongyles) have shown increased resistance to anthelmintics worldwide. Description of strongyle communities have accumulated but little is known about the diversity of these communities and underpinning environmental factors. METHODS: Strongyles were recovered after ivermectin treatment from 48 horses located in six premises in Poland. Correlation between previously published species fecundity and the observed relative abundance and prevalence were estimated. Significance of horse sex was determined at the species level (prevalence, relative abundance) and at the community level (species richness and dissimilarity between communities). RESULTS: Strongyle species fell into two groups, contrasted by their prevalence and relative abundance. Six to nine horses were necessary to sample at least 90% of strongyle community diversity, providing a minimal cut-off to implement sampling trial in the field. Strongyle communities entertained a network of mostly positive interactions and species co-occurrence was found more often than expected by chance. In addition, species fecundity and prevalence were negatively correlated (Pearson's r = -0.71), suggesting functional trade-offs between species dispersal abilities and fecundity. This functional trade-off may underpin species coexistence. Horse sex was also a significant constraint shaping strongyle communities. Indeed, mares generally displayed more similar strongyle communities than stallions (P = 0.003) and Cylicostephanus calicatus was more abundant in stallions suggesting sex-specific interactions (P = 0.006). CONCLUSIONS: While niche partitioning is likely to explain some of the positive interactions between equine strongyle species, coexistence may also result from a functional trade-off between dispersal ability and fecundity. There is significant evidence that horse sex drives strongylid community structure, which may require differential control strategies between mares and stallions.

[The communities of cyathostomes (Cyathostominae) in year-old and two-year-old Pure Blood Arabian mares]. / Zgrupowania slupkowców malych (Cyathostominae) u rocz- nych i dwuletnich klaczek czystej krwi arabskiej.

BACKGROUND: The objective of this study was to evaluate the strongyles species in 34 Pure Blood Arabian Horses: one-year-old mares (n = 12) and two-year old mares (n = 22) from the horse stud in 2005, kept under housed-pastured system and dewormed systematically, twice a year using the ivermectin compounds. MATERIAL AND METHODS: The practical method of in vivo determination of strongyles species in horses is collection of these nematodes from the feaces after treatment. Nematodes were collected 24 hours after deworming from 1000 g sample of feaces, then were classified to appropriate species and relative abundance of strongyles community was determined. RESULTS: In the feaces of horses after treatment 10 species of cyathostomes was determined: Cyathostomum catinatum (CAT), C. pateratum (PAT), Cylicocyclus nassatus (NAS), C. ashworti (ASH), C. leptostomus (LEP), C. insigne (INS), Cylicostephanus longibursatus (LNG), C. goldi (GLD), C. calicatus (CAL) and Coronocyclus coronatus (COR). No large strongyles (Strongylinae) was found. The relative abundance was the highest for following cyathostomes species: NAS (22%), CAT (18.1%), INS (17.7%), PAT (13.8%), LNG (9.1%) and LEP (8.2%). The relative abundance of cyathostomes species was similar in both group of mares. On the other hand CAL (15.8%) was common for the one-year-old mares and INS (25%) for the two-year-old mares. In vivo diagnositic of the infection is important from the perspective of individual small strongyle infections dynamics in horses and because of possible different pathogenicity of individual species of these nematodes.

Occurrence of roundworm (Parascaris equorum) in horses from small farms based on necropsy.

BACKGROUND: The objective of this study was to evaluate the level of roundworm infection in working horses slaughtered for meat. In these horses anthelmintics had not been used. MATERIAL AND METHODS: Roundworms were collected post-mortem from small intestines of 83 horses. RESULTS: The mean prevalence of roundworm infection was 12% and intensity--46.7 specimens per horse. Infection was found only in foals; 10 animals were infected (45.4%) among the 22 examined. Due to common occurrence of roundworms (Parascaris equorum) in foals this nematode control should focus particularly on regular anthelmintic treatment of young horses, providing proper hygienic conditions for them and checking the infection level by coproscopical methods.

Cephenemyia stimulator (Diptera) infection in roe deer (Capreolus capreolus) from Kraków area, southern Poland.

UNLABELLED: The aim of the study was to determine the prevalence of botfly (Diptera: Oestridae) larvae infection in roe deer populations (Capreolus capreolus) in the Kraków area on the basis of necropsy and questionnaire surveys. Hunters were surveyed about the age and sex of hunted animals, and the origin of their habitat. All parasite specimens were identified to species in the laboratory. The parasites were found in the nasal cavities, esophagus, and larynx of male roe deer aged 3-8 years, living in forest habitats. The level of infection was relatively low (13%), with the intensity ranging from 1 to 10 larvae per host. Although no fly larvae were found during the dissection of roe deer carcasses, the parasites received from the hunters were identified as Cephenemyia stimulator botflies. These findings are supported by the evidence drawn from the questionnaires completed by the hunters. KEY WORDS: parasites, Cephenemyia stimulator, Capreolus capreolus, necropsy, questionnaire study, Poland.

Endoparasites of exotic ungulates from the Giraffidae and Camelidae families kept ex situ.

Giraffes and camels are popular attractions at zoological gardens. In order to present the diversity of parasites infecting exotic ungulates from zoos, faecal samples from three giraffes and six camels from both the Silesian Zoological Garden in Chorzów, and Kraków Zoological Garden, were examined. The research was carried out over a ten-month period in 2013 and 2014. In total, 100 faecal samples from 18 animals were analysed with the use of the McMaster method. Moreover, coccidian oocysts were incubated to investigate their development and larvoscopic examination was conducted to detect the presence of nematode species. Giraffes were infected with coccidia from the genus Eimeria, and gastrointestinal nematodes from the Strongylida order, and Trichuris and Aonhotheca genera. One male giraffe was uninfected. The level of infection in giraffes was low when compared to camels kept in both of the zoos. Limited contact with other animal species contributed greatly to the lower level of infection in camels from Kraków Zoo than those from Chorzów, which were kept in the same enclosure as alpacas and Shetland ponies.

Lungworm (Nematoda: Protostrongylidae) infection in wild and domestic ruminants from Malopolska region of Poland.

The conducted study has focused on domestic, as well as wild ruminant species. The post mortem examination was carried out on 68 animals, including three wild species: roe deer (Capreolus capreolus) (25 indyviduals), red deer (Cervus elaphus) (6), fallow deer (Dama dama) (5) and two domestic: sheep (Ovis aries) (14) and cattle (Bos taurus) (18). Some of the species have also been investigated in the field by the coproscopical analyses. The faecal samples from roe deer (27), fallow deer (20), red deer (36) and moose (Alces alces) (10) were collected from the environment, while from sheep (10) and goat (Capra hircus) (10)--per rectum. Based on the obtained results the following values were calculated: prevalence, mean intensity and intensity range. The post mortem examination did not reveal pulmonary nematodes neither in domestic nor in wild ungulates, however, the larvae of aforementioned parasites were often stated in the stool samples taken from the environment. All wild species, except fallow deer were infected. Consequently, six species of lungworms have been identified. The first stage larvae of Varestrongylus capreoli occurred in 11 samples of roe deer and Varestrongylus alces in one moose. The larvae of Elaphostrongylus cervi were found in 19 red deer and Varestrongylus sagittatus in 3. Furthermore, Elaphostrongylus alces larvae were noted in 6 moose. Within domestic ruminants only one sheep and two goats were infected by Muellerius capillaris.

Parasite infections in red deer Cervus elaphus from Krakow area, southern Poland.

The study describes the parasitofauna of red deer Cervus elaphus from Kraków area (southern Poland). The study was done in years 2011-2012 and included altogether 6 animals. Five deer originated from the forest areas (hunted) and one came from the urban site (it died in a road accident). In addition to post-mortem examination, coproscopic analyses with sporulation were performed to define Eimeria species diversity. The deer were infected with four species of nematodes: Ashworthius sidemi, Spiculopteragia spiculoptera, Oesophagostomum venulosum and O. sikae, and two species of coccidia: E. elaphi and E. austriaca. The level of parasitie infections was low, but the results indicate the important role of deer as possible transmitter of A. sidemi to other ungulates in examined area.

Zatorska goose - a subject of parasitological research.

The aim of the study was to determine the level of gastrointestinal parasites in a native breed of geese - Zatorska goose - based on coproscopic testing. Faecal samples were collected from 90 young geese in three age groups (5, 7 and 9 weeks old) in 2014. The geese were kept indoors on deep litter and pastured from spring to autumn. The area of the pastures around the buildings where the geese grazed was about 1 hectare, divided into quarters for different age groups. Before grazing, the birds were dewormed with fenbendazole (Fenbenat powder 4%, Naturan). As additional treatment for coccidiosis, coccidiostats were added to the feed. The study was conducted using the McMaster quantitative method with centrifugation (flotation liquid: NaCl and glucose). The birds were shown to be infected with coccidia and nematodes. The prevalence of Eimeria sp. infection (mean 40%) and the number of oocysts per gram of faeces (reaching 5,300 OPG) were highest in the youngest age group of geese. The level of Amidostomum anseris infection was similar in the three age groups, with prevalence from 40% to 50% (nematode egg output ranged from 50 to 350 eggs per gram of faeces, EPG). Capillaria anatis was observed only in 5- and 7-week-old geese.

The Alice - "Follow the White Rabbit" - parasites of farm rabbits based on coproscopy.

The aim of the study, conducted in the years 2011-2013, was to determine the level of gastrointestinal parasites infection in New Zealand White rabbits, kept at the Experimental Station of the University of Agriculture in Krakow. The study showed rabbits protozoan infection with the genus Eimeria, belonging - based on the sporulation method - to the following species: E. magna, E. media, E. perforans, E. stiedae and E. irresidua. The highest prevalence of infection, as well as the intensity of oocysts output (OPG - oocysts per gram of faeces), was noted for E. magna and E. media - respectively 31.4 % (19477.3 OPG), and 40.0 % (14256.07 OPG). The infection of rabbits with Eimeria spp. differed significantly between years. With regard to oocysts output, the level of infection was strongly connected with the age of rabbits, being higher in young animals. However, the range of infection was highest among adults. Among nematodes, Passalurus ambiguus pinworm was regularly found (prevalence reached 21.9%), other species - Trichuris leporis, and Graphidium strigosum were rarely noted. The overall infection with nematodes did not differ between years. Similarly, as in the case of Eimeria older individuals were more often infected by nematodes. We observed some trends in parasite oocysts/eggs output; the protozoan oocysts were recorded more often in faecal samples collected in the evenings, whereas the nematodes eggs occurred frequently in the mornings. This situation may be related to the phenomenon of coprophagy occurring in the mammals of Lagomorpha order. The results of the study indicate that especially coccidiosis constitute permanently throughout the years an important problem in the rabbitry examined.