Genotyping of Ascaris spp. infecting humans and pigs in Italy, Slovakia and Colombia.

BACKGROUND: The systematics and taxonomy of Ascaris lumbricoides and Ascaris suum, two of the world's most widespread nematodes, still represent a highly debated scientific issue. Two different transmission scenarios have been described according to endemicity: separated host-specific transmission cycles in endemic regions, and a single pool of infection shared by humans and pigs in non-endemic regions. The swine roundworm A. suum is now recognized as an important cause of human ascariasis also in endemic areas such as China, where cross-infections and hybridization have also been reported, as well as in non-endemic regions like Italy. This study aimed to investigate the molecular epidemiology of human and pig ascariasis in three countries representing different epidemiological scenarios: Italy as a non-endemic country, Colombia as an endemic country, and Slovakia as a non-endemic country, but with a poor socio-economic context linked to some focal populations of Roma settlements. MATERIALS AND METHODS: A total of 237 nematodes were analysed: 46 from Colombia (13 from humans, 33 from pigs), 114 from Slovakia (20 from humans, 94 from pigs) and 77 from Italy (17 from humans and 60 from pigs). Genotyping by PCR-RFLP of nuclear (ITS) and sequencing of mitochondrial (cox1) target regions were performed. ITS genotypes were used to estimate the Hardy-Weinberg (HW) equilibrium according to hosts and country of origin. The partial cox1 sequences were used to analyse genetic polymorphisms according to hosts and country of origin, as well as to infer the network of haplotypes, their evolutionary relationships and geographical distribution. RESULTS: 110 quality cox1 sequences were obtained. Haplotype network revealed three main groups corresponding to clade A, B and C. Clade C included most of the human cases from Italy, while those from Slovakia and Colombia were grouped in clade B. Ascaris from Italian and Colombian pigs showed HW equilibrium at the ITS marker, while disequilibrium was found in A. lumbricoides from Slovak pigs, which suggest a high unexpected amount of roundworms of human origin circulating also in pigs. CONCLUSIONS: This study updates and extends the current understanding of Ascaris species and genotypes circulating in different epidemiological scenarios, with particular attention to the inclusion of human-derived Ascaris in the phylogenetic cluster C. Despite the evidence of HW equilibrium in the ITS in pig-derived Italian samples, the amount of genetic variation seems to support the existence of two closely related species.

Cryptosporidium parvum - zoonotic subtype IIdA15G1 in a Slovakian patient.

INTRODUCTION AND OBJECTIVES: The parasite Cryptosporidium spp. is an intracellular protozoa which has a broad range of hosts and zoonotic potential. It presents a serious health risk for agricultural workers and veterinarians. The aim of the study was to identify the species and subtypes of Cryptosporidium occurring in a veterinary student who came into contact with calves on a farm. MATERIAL AND METHODS: The Ziehl-Neelsen staining technique was employed to confirm the presence of Cryptosporidium oocysts. ELISA test was applied to detect coproantigen in faecal specimens. Nested PCR was used to amplify a small ribosomal subunit (SSU rRNA) and sequencing of the GP60 gene served to identify the zoonotic subtypes. RESULTS: The nested PCR allowed to confirm the C. parvum species; subsequently, the IIdA15G1 zoonotic subtype was identified. CONCLUSIONS: This is the first confirmed case in Slovakia of human cryptosporidiosis caused by the unique subtype IIdA15G1.

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.

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.

A frequent roundworm Baylisascaris transfuga in overpopulated brown bears (Ursus arctos) in Slovakia: a problem worthy of attention.

The genus Baylisascaris (order Ascaridida) includes numerous relatively host-specific nematodes, which are common in intestines of wild mammals. Some of them may have impact on veterinary and public health, as their larvae have the potential to cause visceral, ocular, and/or neural larva migrans in a wide range of mammals, birds, and humans. Baylisascaris transfuga is a parasite occurring in a range of bear species throughout the world. We present the current data on B. transfuga occurrence in brown bears from a relatively restricted territory of the Polana Protected Landscape Area in Central Slovakia, obtained by traditional methods (faecal examination, morphology). Species affiliation was confirmed by employing molecular markers generating nuclear 28S and mitochondrial cox1 sequences in adult worms. Based on 17 examined samples (15 excrements and two intestines of young bear females), the occurrence of B. transfuga in the surveyed area was assessed as 52.9%. Both bear females were infected with adult and juvenile worms. Due to the high density of bears in the locality, the high infection rate with ascarids, and the huge number of eggs produced by the parasites, it is apparent that the respective environment, including the inhabited areas, might be markedly contaminated by Baylisascaris eggs. The ability of B. transfuga to serve as a zoonotic agent has not been unambiguously proved; however, this attribute should be considered and subjected to further research.

Dog's genotype of Giardia duodenalis in human: first evidence in Europe.

The unicellular parasite Giardia duodenalis has been divided to eight assemblages (A-H) from which A and B have the most important zoonotic potential. All remaining genotypes have a strong commitment to various host animals. We present here the first clinical case of a human infection with the dog-specific genotype C of G. duodenalis in Slovakia. The patient, 44-year-old woman, suffered from long-term diarrhoea, abdominal pain, anorexia, weight loss, severe itching and dermatitis in the perianal area. The initial microscopic diagnosis was completed by a nested polymerase chain reaction (PCR) which revealed the first evidence of human giardiasis caused by the dog-specific genotype of G. duodenalis on a European scale. A possible role of dogs in zoonotic transmission of giardiasis and its epidemiological and public health relevance is accentuated.

Molecular identification and genotyping of Microsporidia in selected hosts.

The work is described by microscopic analysis, the serological analysis (IFAT) and the molecular analysis of isolates from clinical samples (blood, faeces and urine) from ten domestic rabbits (Oryctolagus cuniculus), breed Malický, four New Zealand domestic rabbits, 11 sows of breed Slo0076akian Improved White and 15 clinically healthy laboratory BALB/c mice. The aim of the study was to validate the suitability of species-unspecific primer pairs 530F and 580R for genotype determination of the Microsporidia strain and species-specific primer pairs ECUNF and ECUNR, SINTF and SINTR and EBIER1 and EBIEF1 for the determination of E ncephalitozoon cuniculi, Encephalitozoon intestinalis and Enterocytozoon bieneusi species for diagnostic purposes. Sequences of animals were compared with those from the GenBank database. In rabbits, two murine genotypes II and four canine genotypes III were identified. Genotype II was identified in mice. The Encephalitozoon intestinalis identified in the sample from swine showed no genetic heterogeneity.

Occurrence of Cryptosporidium spp. in red foxes and brown bear in the Slovak Republic.

Wild animals can be involved in epidemiology of many important diseases and often act as reservoirs of pathogens which cause disease in domestic animals and humans. This paper aims the role of red fox (Vulpes vulpes) and brown bear (Ursus arctos) in the circulation of coccidian parasites from the genus Cryptosporidium. Cryptosporidiosis is known as an important enteric pathogen, clinical symptoms in particular in immune-compromised individuals range from mild to severe diarrhoea and dehydration, which could be fatal. Fecal samples from 62 red foxes shot during September 2010 to February 2011 and 63 brown bears collected during June 2010 to March 2011 in central and eastern Slovakia were examined for the qualitative determination of Cryptosporidium spp. antigens in faeces by sandwich ELISA kit. Overall, 38.7% (24/62) of faecal samples of red foxes and 55.6% (35/63) of faecal samples of brown bear were positive. Our preliminary results emphasize prevalence of Cryptosporidium spp. amongst brown bears and red foxes in Slovakia and highlight the potential risk for transmission of cryptosporidiosis to humans using the countryside for professional or recreational purposes.