The zoonotic parasite Cryptosporidium parvum is a global cause of gastrointestinal disease in humans and ruminants. Sequence analysis of the highly polymorphic gp60 gene enabled the classification of C. parvum isolates into multiple groups (e.g., IIa, IIc, Id) and a large number of subtypes. In Europe, subtype IIaA15G2R1 is largely predominant and has been associated with many water- and food-borne outbreaks. In this study, we generated new whole-genome sequence (WGS) data from 123 human- and ruminant-derived isolates collected in 13 European countries and included other available WGS data from Europe, Egypt, China, and the United States (n = 72) in the largest comparative genomics study to date. We applied rigorous filters to exclude mixed infections and analyzed a data set from 141 isolates from the zoonotic groups IIa (n = 119) and IId (n = 22). Based on 28,047 high-quality, biallelic genomic SNPs, we identified three distinct and strongly supported populations: Isolates from China (IId) and Egypt (IIa and IId) formed population 1; a minority of European isolates (IIa and IId) formed population 2; and the majority of European (IIa, including all IIaA15G2R1 isolates) and all isolates from the United States (IIa) clustered in population 3. Based on analyses of the population structure, population genetics, and recombination, we show that population 3 has recently emerged and expanded throughout Europe to then, possibly from the United Kingdom, reach the United States, where it also expanded. The reason(s) for the successful spread of population 3 remain elusive, although genes under selective pressure uniquely in this population were identified.

• MeSH
• Cryptosporidium parvum * genetics   MeSH
• Disease Outbreaks * MeSH
• Phylogeny MeSH
• Genome, Protozoan MeSH
• Genomics methods   MeSH
• Polymorphism, Single Nucleotide MeSH
• Cryptosporidiosis * parasitology   epidemiology   MeSH
• Humans MeSH
• Whole Genome Sequencing methods   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• China epidemiology   MeSH
• Egypt epidemiology   MeSH
• Europe epidemiology   MeSH
• United States epidemiology   MeSH

BACKGROUND: Avian cryptosporidiosis is a common parasitic disease that is caused by five species, which are well characterised at the molecular and biological level, and more than 18 genotypes for which we have limited information. In this study, we determined the occurrence and molecular characteristics of Cryptosporidium spp. in farmed ostriches in the Czech Republic. METHODS: The occurrence and genetic identity of Cryptosporidium spp. were analysed by microscopy and PCR/sequencing of the small subunit rRNA, actin, HSP70 and gp60 genes. Cryptosporidium avian genotype II was examined from naturally and experimentally infected hosts and measured using differential interference contrast. The localisation of the life-cycle stages was studied by electron microscopy and histologically. Infectivity of Cryptosporidium avian genotype II for cockatiels (Nymphicus hollandicus (Kerr)), chickens (Gallus gallus f. domestica (L.)), geese (Anser anser f. domestica (L.)), SCID and BALB/c mice (Mus musculus L.) was verified. RESULTS: A total of 204 individual faecal samples were examined for Cryptosporidium spp. using differential staining and PCR/sequencing. Phylogenetic analysis of small subunit rRNA, actin, HSP70 and gp60 gene sequences showed the presence of Cryptosporidium avian genotype II (n = 7) and C. ubiquitum Fayer, Santín & Macarisin, 2010 IXa (n = 5). Only ostriches infected with Cryptosporidium avian genotype II shed oocysts that were detectable by microscopy. Oocysts were purified from a pooled sample of four birds, characterised morphometrically and used in experimental infections to determine biological characteristics. Oocysts of Cryptosporidium avian genotype II measure on average 6.13 × 5.15 μm, and are indistinguishable by size from C. baileyi Current, Upton & Haynes, 1986 and C. avium Holubová, Sak, Horčičková, Hlásková, Květoňová, Menchaca, McEvoy & Kváč, 2016. Cryptosporidium avian genotype II was experimentally infectious for geese, chickens and cockatiels, with a prepatent period of four, seven and eight days post-infection, respectively. The infection intensity ranged from 1000 to 16,000 oocysts per gram. None of the naturally or experimentally infected birds developed clinical signs in the present study. CONCLUSIONS: The molecular and biological characteristics of Cryptosporidium avian genotype II, described here, support the establishment of a new species, Cryptosporidium ornithophilus n. sp.

• Keywords
• C. ubiquitum, Cryptosporidium avian genotype II, Cryptosporidium ornithophilus n. sp., Experimental infections, Occurrence, Oocyst size, PCR,
• MeSH
• Cryptosporidium classification   genetics   ultrastructure   MeSH
• Phylogeny MeSH
• Animals, Domestic parasitology   MeSH
• Host Specificity MeSH
• Classification MeSH
• Cryptosporidiosis parasitology   MeSH
• Bird Diseases parasitology   MeSH
• Genes, Protozoan genetics   MeSH
• Birds parasitology   MeSH
• Life Cycle Stages MeSH
• Struthioniformes parasitology   MeSH
• DNA Barcoding, Taxonomic veterinary   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH

BACKGROUND: Cryptosporidium spp., Giardia duodenalis and Enterocytozoon bieneusi are common enteric pathogens in humans and animals. Data on the transmission of these pathogens are scarce from Guangdong, China, which has a subtropical monsoon climate and is the epicenter for many emerging infectious diseases. This study was conducted to better understand the prevalence and identity of the three pathogens in pre-weaned dairy calves in Guangdong. METHODS: The occurrence and genetic identity of three pathogens were analyzed by polymerase chain reaction. PCR-positive products were sequenced to determine the species and genotypes. A Chi-square test was used to compare the prevalence of pathogens among sampling dates, age groups, or clinical signs. RESULTS: The detection rates of Cryptosporidium spp., G. duodenalis and E. bieneusi were 24.0% (93/388), 74.2% (288/388) and 15.7% (61/388), respectively. Three Cryptosporidium species were detected, including C. bovis (n = 73), C. parvum (n = 12) and C. ryanae (n = 7); one animal had concurrence of C. bovis and C. parvum. C. parvum was the dominant species during the first two weeks of life, whereas C. bovis and C. ryanae were mostly seen at 3-9 weeks of age. Sequence analysis identified the C. parvum as subtype IIdA19G1. Assemblage E (n = 282), assemblage A (n = 1), and concurrence of A and E (n = 5) were identified among G. duodenalis-positive animals using multilocus genotyping (MLG). Altogether, 15, 10 and 17 subtypes of assemblage E were observed at the bg, gdh and tpi loci, respectively, forming 49 assemblage E MLGs. The highest detection rate of G. duodenalis was found in winter. Sequence analysis identified genotypes J (n = 57), D (n = 3) and one concurrence of J and D among E. bieneusi-positive animals. The detection rate of E. bieneusi was significantly higher in spring (38.0%; 41/108) than in summer (7.2%; 8/111) and winter (7.1%; 12/169). CONCLUSIONS: These results indicate a common occurrence of C. parvum subtype IIdA19G1, G. duodenalis assemblage E, and E. bieneusi genotype J in pre-weaned dairy calves in Guangdong. More studies are needed to understand the unique genetic characteristics and zoonotic potential of the three enteric pathogens in the province.

• Keywords
• Cryptosporidium, Enterocytozoon bieneusi, Giardia duodenalis, Molecular epidemiology,
• MeSH
• Cryptosporidium genetics   isolation & purification   MeSH
• Enterocytozoon genetics   immunology   MeSH
• Genotype MeSH
• Giardia lamblia genetics   isolation & purification   MeSH
• Giardiasis epidemiology   parasitology   veterinary   MeSH
• Cryptosporidiosis epidemiology   parasitology   MeSH
• Microsporidiosis epidemiology   parasitology   veterinary   MeSH
• Molecular Epidemiology MeSH
• Cattle Diseases epidemiology   MeSH
• Animals, Newborn MeSH
• Prevalence MeSH
• Cross-Sectional Studies MeSH
• Cattle MeSH
• Animals MeSH
• Check Tag
• Cattle MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• China epidemiology   MeSH

IntroductionThis paper reviews the current knowledge and understanding of Cryptosporidium spp. and Giardia spp. in humans, animals and the environment in 10 countries in the eastern part of Europe: Bosnia and Herzegovina, Croatia, Czech Republic, Estonia, Hungary, Latvia, Poland, Romania, Serbia and Slovenia. Methods: Published scientific papers and conference proceedings from the international and local literature, official national health service reports, national databases and doctoral theses in local languages were reviewed to provide an extensive overview on the epidemiology, diagnostics and research on these pathogens, as well as analyse knowledge gaps and areas for further research. Results:Cryptosporidium spp. and Giardia spp. were found to be common in eastern Europe, but the results from different countries are difficult to compare because of variations in reporting practices and detection methodologies used. Conclusion: Upgrading and making the diagnosis/detection procedures more uniform is recommended throughout the region. Public health authorities should actively work towards increasing reporting and standardising reporting practices as these prerequisites for the reported data to be valid and therefore necessary for appropriate control plans.

• Keywords
• One Health, cryptosporidiosis, giardiasis, zoonosis,
• MeSH
• Cryptosporidium genetics   isolation & purification   MeSH
• Feces parasitology   MeSH
• Giardia genetics   isolation & purification   MeSH
• Giardiasis epidemiology   parasitology   MeSH
• Cryptosporidiosis epidemiology   parasitology   MeSH
• Real-Time Polymerase Chain Reaction MeSH
• Humans MeSH
• Foodborne Diseases epidemiology   parasitology   MeSH
• Prevalence MeSH
• Public Health * MeSH
• Environment MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Review MeSH
• Geographicals
• Europe, Eastern epidemiology   MeSH

The morphological, biological, and molecular characteristics of Cryptosporidium avian genotype V are described, and the species name Cryptosporidium avium is proposed to reflect its specificity for birds under natural and experimental conditions. Oocysts of C. avium measured 5.30-6.90 μm (mean = 6.26 μm) × 4.30-5.50 μm (mean = 4.86 μm) with a length to width ratio of 1.29 (1.14-1.47). Oocysts of C. avium obtained from four naturally infected red-crowned parakeets (Cyanoramphus novaezealandiae) were infectious for 6-month-old budgerigars (Melopsittacus undulatus) and hens (Gallus gallus f. domestica). The prepatent periods in both susceptible bird species was 11 days postinfection (DPI). The infection intensity of C. avium in budgerigars and hens was low, with a maximum intensity of 5000 oocysts per gram of feces. Oocysts of C. avium were microscopically detected at only 12-16 DPI in hens and 12 DPI in budgerigars, while PCR analyses revealed the presence of specific DNA in fecal samples from 11 to 30 DPI (the conclusion of the experiment). Cryptosporidium avium was not infectious for 8-week-old SCID and BALB/c mice (Mus musculus). Naturally or experimentally infected birds showed no clinical signs of cryptosporidiosis, and no pathology was detected. Developmental stages of C. avium were detected in the ileum and cecum using scanning electron microscopy. Phylogenetic analyses based on small subunit rRNA, actin, and heat shock protein 70 gene sequences revealed that C. avium is genetically distinct from previously described Cryptosporidium species.

• Keywords
• Cryptosporidium avian genotype V, Cryptosporidium avium, Molecular analyses, Morphology, Transmission studies,
• MeSH
• Cecum parasitology   MeSH
• Cryptosporidium classification   genetics   isolation & purification   MeSH
• Feces parasitology   MeSH
• Phylogeny MeSH
• Ileum parasitology   MeSH
• Cryptosporidiosis parasitology   MeSH
• Chickens parasitology   MeSH
• Melopsittacus parasitology   MeSH
• Mice, Inbred BALB C MeSH
• Mice, SCID MeSH
• Mice MeSH
• Poultry Diseases parasitology   MeSH
• Oocysts MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

Proventriculus and intestinal samples from 70 North American red-winged blackbirds (Agelaius phoeniceus; order Passeriformes) were examined for the presence of Cryptosporidium by PCR amplification and sequence analysis of the 18S ribosomal RNA (18S rRNA), actin, and 70-kDa heat shock protein (HSP70) genes. Twelve birds (17.1 %) were positive for the Cryptosporidium 18S rRNA gene: six birds were positive at the proventriculus site only and six birds were positive at the proventriculus and intestinal sites. Sequence analysis of the 18S rRNA, actin and HSP70 genes showed the presence of the gastric species Cryptosporidium galli in a single proventriculus sample and a closely related genotype, which we have named Cryptosporidium avian genotype VI, in all other positive samples. These findings contribute to our understanding of Cryptosporidium diversification in passerines, the largest avian order.

• Keywords
• Avian genotype VI, Cryptosporidium, Cryptosporidium galli, Intestine, Passerines, Proventriculus, Red-winged blackbird,
• MeSH
• Cryptosporidium classification   MeSH
• Genotype MeSH
• Cryptosporidiosis epidemiology   parasitology   MeSH
• Bird Diseases epidemiology   parasitology   MeSH
• Passeriformes * MeSH
• Polymerase Chain Reaction MeSH
• HSP70 Heat-Shock Proteins genetics   metabolism   MeSH
• RNA, Ribosomal, 18S genetics   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• United States epidemiology   MeSH
• Names of Substances
• HSP70 Heat-Shock Proteins MeSH
• RNA, Ribosomal, 18S MeSH

From July to November 2012, preliminary coprological examinations were carried out on 85 pooled faecal samples of different aged ring-necked pheasants (Phasianus colchicus) (hatches from May until July) from an intensive artificial breeding programme in the Czech Republic. Cryptosporidium oocysts were detected in 12 samples (14.1 %) of ages >12 weeks (August-September). These results were supported by findings of Cryptosporidium baileyi and Cryptosporidium meleagridis oocysts in intestinal, or cloacal contents, and/or the bursa of Fabricius in 9 from 36 examined dead pheasants (prevalence 25 %). We describe in detail the various age groups of pheasants after hatching and present graphically the overall results of coprological examinations, showing pathways of infection of C. baileyi and C. meleagridis during the full rearing seasons of 2013 and 2014. We found very similar mean proportions of Cryptosporidium-positive samples over the entire 2013 period in pheasantry (173 pooled samples tested, 25 positive, 14.5 %) and 2014 (238 samples tested, 43 positive, 18.1 %). All tests were verified as being Cryptosporidium positive in 9 from 219 (prevalence 4.1 %) and 4 from 168 (prevalence 2.4 %) post-mortem examinations. Significantly, C. baileyi was found more frequently in faeces, with positivities ranging from 11.1 to 100 % (4->16-week-old pheasants). Oocysts of C. meleagridis were detected at ages 6->15 weeks ranging from 7.1 to 100 % in faeces during the rearing seasons. The burdens of C. baileyi (7 of 14 and 10 of 16) and C. meleagridis (5 of 14 and 7 of 16) for each year, in monitored brooder houses, flight pens and spread across all open areas were recorded. Oocysts of C. baileyi and C. meleagridis obtained from this study, and Cryptosporidium galli (obtained in another aviary from 36-week-old pheasants), were sequenced, and we characterized the highly variable 60-kDa glycoprotein gene of C. meleagridis. These results highlight the real risk of transmission of Cryptosporidium to susceptible wild birds and other potential hosts after termination of rearing and release.

• Keywords
• C. baileyi, C. galli, Cryptosporidium meleagridis, Dynamics and transmission, GP60, Prevalence,
• MeSH
• Cryptosporidium genetics   MeSH
• Feces parasitology   MeSH
• Galliformes * MeSH
• Cryptosporidiosis epidemiology   parasitology   MeSH
• Poultry Diseases epidemiology   parasitology   MeSH
• Oocysts MeSH
• Aging MeSH
• Intestines parasitology   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Czech Republic epidemiology   MeSH

The morphological, biological, and molecular characteristics of Cryptosporidium muris strain TS03 are described, and the species name Cryptosporidium proliferans n. sp. is proposed. Cryptosporidium proliferans obtained from a naturally infected East African mole rat (Tachyoryctes splendens) in Kenya was propagated under laboratory conditions in rodents (SCID mice and southern multimammate mice, Mastomys coucha) and used in experiments to examine oocyst morphology and transmission. DNA from the propagated C. proliferans isolate, and C. proliferans DNA isolated from the feces of an African buffalo (Syncerus caffer) in Central African Republic, a donkey (Equus africanus) in Algeria, and a domestic horse (Equus caballus) in the Czech Republic were used for phylogenetic analyses. Oocysts of C. proliferans are morphologically distinguishable from C. parvum and C. muris HZ206, measuring 6.8-8.8 (mean = 7.7 μm) × 4.8-6.2 μm (mean = 5.3) with a length to width ratio of 1.48 (n = 100). Experimental studies using an isolate originated from T. splendens have shown that the course of C. proliferans infection in rodent hosts differs from that of C. muris and C. andersoni. The prepatent period of 18-21 days post infection (DPI) for C. proliferans in southern multimammate mice (Mastomys coucha) was similar to that of C. andersoni and longer than the 6-8 DPI prepatent period for C. muris RN66 and HZ206 in the same host. Histopatologicaly, stomach glands of southern multimammate mice infected with C. proliferans were markedly dilated and filled with necrotic material, mucus, and numerous Cryptosporidium developmental stages. Epithelial cells of infected glands were atrophic, exhibited cuboidal or squamous metaplasia, and significantly proliferated into the lumen of the stomach, forming papillary structures. The epithelial height and stomach weight were six-fold greater than in non-infected controls. Phylogenetic analyses based on small subunit rRNA, Cryptosporidium oocyst wall protein, thrombospondin-related adhesive protein of Cryptosporidium-1, heat shock protein 70, actin, heat shock protein 90 (MS2), MS1, MS3, and M16 gene sequences revealed that C. proliferans is genetically distinct from C. muris and other previously described Cryptosporidium species.

• MeSH
• Cryptosporidium classification   genetics   MeSH
• Phylogeny MeSH
• Cryptosporidiosis parasitology   MeSH
• Mole Rats MeSH
• Mice, SCID MeSH
• Mice MeSH
• Oocysts metabolism   MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Faecal samples were collected from 352 horses on 23 farms operating under six different management systems in the Czech Republic and Poland during 2011 and 2012. Farms were selected without previous knowledge of parasitological status. All faecal samples were screened for Cryptosporidium spp. presence using microscopy, following aniline-carbol-methyl violet staining and PCR analysis of the small-subunit (SSU) rRNA and the 60-kDa glycoprotein (gp60) genes. Cryptosporidium muris-positive samples were additionally genotyped at four minisatellite markers: MS1 (encoding a hypothetical protein), MS2 (encoding a 90-kDa heat shock protein), MS3 (encoding a hypothetical protein) and MS16 (encoding a leucine-rich repeat family protein). Cryptosporidium spp. was detected by PCR in 12/352 (3.4%) samples from 4 out of 13 farms. None of the samples tested by microscopy was positive. There was no relationship between Cryptosporidium prevalence and age, sex, diarrhoea or management system; however, Cryptosporidium was found only on farms where horses were kept on pasture during the day and in a stable overnight. Sequence analyses of SSU and gp60 genes revealed the presence of C. muris RN66 (n = 9), Cryptosporidium parvum IIaA15G2R1 (n = 1), Cryptosporidium tyzzeri IXbA22R9 (n = 1), and Cryptosporidium horse genotype VIaA15G4 (n = 1). The C. muris subtypes were identified as MS1-M1, MS2-M4, novel MS2-M7 and MS16-M1 by multilocus sequence of three minisatellite loci. The MS3 locus was not amplified from any isolate. This is the first report of C. tyzzeri and C. muris subtypes from horses.

• MeSH
• Cryptosporidium classification   genetics   isolation & purification   MeSH
• Feces parasitology   MeSH
• Genetic Variation * MeSH
• Genotype MeSH
• Horses MeSH
• Cryptosporidiosis epidemiology   MeSH
• Horse Diseases parasitology   MeSH
• Prevalence MeSH
• Diarrhea parasitology   veterinary   MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Czech Republic epidemiology   MeSH
• Poland epidemiology   MeSH

BACKGROUND: Infectious diseases represent the greatest threats to endangered species, and transmission from humans to wildlife under increased anthropogenic pressure has been always stated as a major risk of habituation. AIMS: To evaluate the impact of close contact with humans on the occurrence of potentially zoonotic protists in great apes, one hundred mountain gorillas (Gorilla beringei beringei) from seven groups habituated either for tourism or for research in Volcanoes National Park, Rwanda were screened for the presence of microsporidia, Cryptosporidium spp. and Giardia spp. using molecular diagnostics. RESULTS: The most frequently detected parasites were Enterocytozoon bieneusi found in 18 samples (including genotype EbpA, D, C, gorilla 2 and five novel genotypes gorilla 4-8) and Encephalitozoon cuniculi with genotype II being more prevalent (10 cases) compared to genotype I (1 case). Cryptosporidium muris (2 cases) and C. meleagridis (2 cases) were documented in great apes for the first time. Cryptosporidium sp. infections were identified only in research groups and occurrence of E. cuniculi in research groups was significantly higher in comparison to tourist groups. No difference in prevalence of E. bieneusi was observed between research and tourist groups. CONCLUSION: Although our data showed the presence and diversity of important opportunistic protists in Volcanoes gorillas, the source and the routes of the circulation remain unknown. Repeated individual sampling, broad sampling of other hosts sharing the habitat with gorillas and quantification of studied protists would be necessary to acquire more complex data.

• MeSH
• Cryptosporidium classification   genetics   isolation & purification   MeSH
• Encephalitozoon classification   genetics   isolation & purification   MeSH
• Encephalitozoonosis epidemiology   microbiology   MeSH
• Phylogeny MeSH
• Giardia classification   genetics   isolation & purification   MeSH
• Giardiasis epidemiology   parasitology   MeSH
• Hominidae MeSH
• DNA, Intergenic genetics   MeSH
• Cryptosporidiosis epidemiology   parasitology   MeSH
• Molecular Sequence Data MeSH
• Ape Diseases epidemiology   microbiology   parasitology   MeSH
• Parks, Recreational MeSH
• Zoonoses epidemiology   microbiology   parasitology   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Geographicals
• Rwanda epidemiology   MeSH
• Names of Substances
• DNA, Intergenic MeSH