Cryptosporidium, a protozoan parasite that causes watery diarrhea, is found worldwide and is common in areas with low water hygiene. In February 2014, 866 stool samples were collected from the inhabitants of 2 rural areas in White Nile State, Sudan. These stool samples were assessed by performing modified acid-fast staining, followed by examination under a light microscope. The overall positive rate of Cryptosporidium oocysts was 13.3%. Cryptosporidium oocysts were detected in 8.6% stool samples obtained from inhabitants living in the area having water purification systems and in 14.6% stool samples obtained from inhabitants living in the area not having water purification systems. No significant difference was observed in the prevalence of Cryptosporidium infection between men and women (14.7% and 14.1%, respectively). The positive rate of oocysts by age was the highest among inhabitants in their 60s (40.0%). These findings suggest that the use of water purification systems is important for preventing Cryptosporidium infection among inhabitants of these rural areas in Sudan.
Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; Cryptosporidiosis/epidemiology/*parasitology ; Cryptosporidium/genetics/*isolation & purification ; Feces/parasitology ; Female ; Humans ; Infant ; Male ; Middle Aged ; Prevalence ; Rural Population ; Sudan/epidemiology ; Young Adult

In order to examine the prevalence of Cryptosporidium infection in wild rodents and insectivores of South Korea and to assess their potential role as a source of human cryptosporidiosis, a total of 199 wild rodents and insectivore specimens were collected from 10 regions of South Korea and screened for Cryptosporidium infection over a period of 2 years (2012-2013). A nested-PCR amplification of Cryptosporidium oocyst wall protein (COWP) gene fragment revealed an overall prevalence of 34.2% (68/199). The sequence analysis of 18S rRNA gene locus of Cryptosporidium was performed from the fecal and cecum samples that tested positive by COWP amplification PCR. As a result, we identified 4 species/genotypes; chipmunk genotype I, cervine genotype I, C. muris, and a new genotype which is closely related to the bear genotype. The new genotype isolated from 12 Apodemus agrarius and 2 Apodemus chejuensis was not previously identified as known species or genotype, and therefore, it is supposed to be a novel genotype. In addition, the host spectrum of Cryptosporidium was extended to A. agrarius and Crosidura lasiura, which had not been reported before. In this study, we found that the Korean wild rodents and insectivores were infected with various Cryptosporidium spp. with large intra-genotypic variationa, indicating that they may function as potential reservoirs transmitting zoonotic Cryptosporidium to livestock and humans.
Animals ; Animals, Wild/*parasitology ; Cryptosporidiosis/*parasitology ; Cryptosporidium/classification/*genetics/*isolation & purification ; Feces/parasitology ; Genotype ; Insectivora/*parasitology ; Molecular Sequence Data ; Murinae ; Phylogeny ; Republic of Korea ; Rodent Diseases/*parasitology

Various constituents in clinical specimens, particularly feces, can inhibit the PCR assay and lead to false-negative results. To ensure that negative results of a diagnostic PCR assay are true, it should be properly monitored by an inhibition control. In this study, a cloning vector harboring a modified target DNA sequence (approximately375 bp) was constructed to be used as a competitive internal amplification control (IAC) for a conventional PCR assay that detects approximately550 bp of the Cryptosporidium oocyst wall protein (COWP) gene sequence in human feces. Modification of the native PCR target was carried out using a new approach comprising inverse PCR and restriction digestion techniques. IAC was included in the assay, with the estimated optimum concentration of 1 fg per reaction, as duplex PCR. When applied on fecal samples spiked with variable oocysts counts, approximately2 oocysts were theoretically enough for detection. When applied on 25 Cryptosporidium-positive fecal samples of various infection intensities, both targets were clearly detected with minimal competition noticed in 2-3 samples. Importantly, both the analytical and the diagnostic sensitivities of the PCR assay were not altered with integration of IAC into the reactions. When tried on 180 randomly collected fecal samples, 159 were Cryptosporidium-negatives. Although the native target DNA was absent, the IAC amplicon was obviously detected on gel of all the Cryptosporidium-negative samples. These results imply that running of the diagnostic PCR, inspired with the previously developed DNA extraction protocol and the constructed IAC, represents a useful tool for Cryptosporidium detection in human feces.
Cryptosporidiosis/*diagnosis/*parasitology ; Cryptosporidium/genetics/*isolation & purification ; DNA Primers/genetics ; DNA, Protozoan/genetics ; Feces/parasitology ; Humans ; Polymerase Chain Reaction/methods/*standards ; Reference Standards

Non-human primates (NHPs) are confirmed as reservoirs of Cryptosporidium spp., Giardia intestinalis, and Enterocytozoon bieneusi. In this study, 197 fresh fecal samples from 8 NHP species in Qinling Mountains, northwestern China, were collected and examined using multilocus sequence typing (MLST) method. The results showed that 35 (17.8%) samples were positive for tested parasites, including Cryptosporidium spp. (3.0%), G. intestinalis (2.0%), and E. bieneusi (12.7%). Cryptosporidium spp. were detected in 6 fecal samples of Macaca mulatta, and were identified as C. parvum (n=1) and C. andersoni (n=5). Subtyping analysis showed Cryptosporidium spp. belonged to the C. andersoni MLST subtype (A4, A4, A4, and A1) and C. parvum 60 kDa glycoprotein (gp60) subtype IId A15G2R1. G. intestinalis assemblage E was detected in 3 M. mulatta and 1 Saimiri sciureus. Intra-variations were observed at the triose phosphate isomerase (tpi), beta giardin (bg), and glutamate dehydrogenase (gdh) loci, with 3, 1, and 2 new subtypes found in respective locus. E. bieneusi was found in Cercopithecus neglectus (25.0%), Papio hamadrayas (16.7%), M. mulatta (16.3%), S. sciureus (10%), and Rhinopithecus roxellana (9.5%), with 5 ribosomal internal transcribed spacer (ITS) genotypes: 2 known genotypes (D and BEB6) and 3 novel genotypes (MH, XH, and BSH). These findings indicated the presence of zoonotic potential of Cryptosporidium spp. and E. bieneusi in NHPs in Qinling Mountains. This is the first report of C. andersoni in NHPs. The present study provided basic information for control of cryptosporidiosis, giardiasis, and microsporidiosis in human and animals in this area.
Animals ; China ; Cryptosporidiosis/*parasitology ; Cryptosporidium/classification/genetics/*isolation & purification ; Enterocytozoon/classification/genetics/*isolation & purification ; Feces/parasitology ; Female ; Genotype ; Giardia lamblia/classification/genetics/*isolation & purification ; Giardiasis/parasitology/*veterinary ; Male ; Microsporidiosis/parasitology/*veterinary ; Molecular Sequence Data ; Phylogeny ; Primate Diseases/*parasitology ; Primates/classification/parasitology

Cryptosporidium spp., ubiquitous enteric parasitic protozoa of vertebrates, recently emerged as an important cause of economic loss and zoonosis. The present study aimed to determine the distribution and species of Cryptosporidium in post-weaned and adult pigs in Shaanxi province, northwestern China. A total of 1,337 fresh fecal samples of post-weaned and adult pigs were collected by sterile disposable gloves from 8 areas of Shaanxi province. The samples were examined by Sheather's sugar flotation technique and microscopy atx400 magnification for Cryptosporidium infection, and the species in positive samples was further identified by PCR amplification of the small subunit (SSU) rRNA gene. A total of 44 fecal samples were successfully amplified by the nested PCR of the partial SSU rRNA, with overall prevalence of 3.3%. The average prevalence of Cryptosporidium infection in each pig farms ranged from 0 to 14.4%. Species identification by sequencing of SSU rRNA gene revealed that 42 (3.1%) samples were Cryptosporidium suis and 2 (0.15%) were Cryptosporidium scrofarum. C. suis had the highest prevalence (7.5%) in growers and the lowest in breeding pigs (0.97%). C. suis was the predominant species in pre-weaned and adult pigs, while C. scrofarum infected pigs older than 3 months only. A season-related difference of C. suis was observed in this study, with the highest prevalence in autumn (5.5%) and the lowest (1.7%) in winter. The present study provided basic information for control of Cryptosporidium infection in pigs and assessment of zoonotic transmission of pigs in Shaanxi province, China.
Animals ; China/epidemiology ; Cluster Analysis ; Cryptosporidiosis/*epidemiology/*parasitology ; Cryptosporidium/classification/genetics/*isolation & purification ; DNA, Protozoan/chemistry/genetics ; DNA, Ribosomal/chemistry/genetics ; Feces/parasitology ; Molecular Sequence Data ; Phylogeny ; Polymerase Chain Reaction ; Prevalence ; RNA, Ribosomal, 18S/genetics ; Seasons ; Sequence Analysis, DNA ; Swine ; Swine Diseases/*epidemiology/*parasitology

Cryptosporidium spp., ubiquitous enteric parasitic protozoa of vertebrates, recently emerged as an important cause of economic loss and zoonosis. The present study aimed to determine the distribution and species of Cryptosporidium in post-weaned and adult pigs in Shaanxi province, northwestern China. A total of 1,337 fresh fecal samples of post-weaned and adult pigs were collected by sterile disposable gloves from 8 areas of Shaanxi province. The samples were examined by Sheather's sugar flotation technique and microscopy atx400 magnification for Cryptosporidium infection, and the species in positive samples was further identified by PCR amplification of the small subunit (SSU) rRNA gene. A total of 44 fecal samples were successfully amplified by the nested PCR of the partial SSU rRNA, with overall prevalence of 3.3%. The average prevalence of Cryptosporidium infection in each pig farms ranged from 0 to 14.4%. Species identification by sequencing of SSU rRNA gene revealed that 42 (3.1%) samples were Cryptosporidium suis and 2 (0.15%) were Cryptosporidium scrofarum. C. suis had the highest prevalence (7.5%) in growers and the lowest in breeding pigs (0.97%). C. suis was the predominant species in pre-weaned and adult pigs, while C. scrofarum infected pigs older than 3 months only. A season-related difference of C. suis was observed in this study, with the highest prevalence in autumn (5.5%) and the lowest (1.7%) in winter. The present study provided basic information for control of Cryptosporidium infection in pigs and assessment of zoonotic transmission of pigs in Shaanxi province, China.
Animals ; China/epidemiology ; Cluster Analysis ; Cryptosporidiosis/*epidemiology/*parasitology ; Cryptosporidium/classification/genetics/*isolation & purification ; DNA, Protozoan/chemistry/genetics ; DNA, Ribosomal/chemistry/genetics ; Feces/parasitology ; Molecular Sequence Data ; Phylogeny ; Polymerase Chain Reaction ; Prevalence ; RNA, Ribosomal, 18S/genetics ; Seasons ; Sequence Analysis, DNA ; Swine ; Swine Diseases/*epidemiology/*parasitology

Cryptosporidium parvum is a zoonotic protozoan parasite that causes cryptosporidial enteritis. Numerous outbreaks of cryptosporidiosis have been reported worldwide. Cryptosporidium is transmitted to hosts via consumption of contaminated water and food but also by direct contact with contaminated soil or infected hosts. The present study investigated farm soil collected from 34 locations along the western Korean peninsula and 24 vegetables purchased from local grocery markets in Seoul. The soil and vegetable samples were examined by real-time polymerase chain reaction (qPCR) to estimate the risk of infection. Eleven of 34 locations (32.4%) and 3 of 24 vegetable samples (12.5%) were contaminated with Cryptosporidium parvum, as confirmed by TaqI enzyme digestion of qPCR products and DNA sequencing. It is suggested that Cryptosporidium infection can be mediated via farm soil and vegetables. Therefore, it is necessary to reduce contamination of this organism in view of public health.
Base Sequence ; Cryptosporidiosis/parasitology ; Cryptosporidium parvum/*genetics/*isolation & purification ; DNA, Protozoan/analysis/genetics ; Enteritis/parasitology ; Foodborne Diseases/*parasitology ; Humans ; Sequence Alignment ; Sequence Analysis, DNA ; Soil/*parasitology ; Vegetables/*parasitology

From May to June 2012, a waterborne outbreak of 124 cases of cryptosporidiosis occurred in the plumbing systems of an older high-rise apartment complex in Seoul, Republic of Korea. The residents of this apartment complex had symptoms of watery diarrhea and vomiting. Tap water samples in the apartment complex and its adjacent buildings were collected and tested for 57 parameters under the Korean Drinking Water Standards and for additional 11 microbiological parameters. The microbiological parameters included total colony counts, Clostridium perfringens, Enterococcus, fecal streptococcus, Salmonella, Shigella, Pseudomonas aeruginosa, Cryptosporidium oocysts, Giardia cysts, total culturable viruses, and Norovirus. While the tap water samples of the adjacent buildings complied with the Korean Drinking Water Standards for all parameters, fecal bacteria and Cryptosporidium oocysts were detected in the tap water samples of the outbreak apartment complex. It turned out that the agent of the disease was Cryptosporidium parvum. The drinking water was polluted with sewage from a septic tank in the apartment complex. To remove C. parvum oocysts, we conducted physical processes of cleaning the water storage tanks, flushing the indoor pipes, and replacing old pipes with new ones. Finally we restored the clean drinking water to the apartment complex after identification of no oocysts.
Cryptosporidiosis/*epidemiology/*parasitology ; Cryptosporidium parvum/genetics/growth & development/*isolation & purification ; Disease Outbreaks ; Drinking Water/*parasitology ; Housing ; Humans ; Oocysts/growth & development ; Republic of Korea/epidemiology ; Water Supply/analysis

Fecal samples of 2,056 dairy cattle from 14 farms were collected in three geographical regions of China and stained using a modified acid-fast staining technique to identify Cryptosporidium oocysts. A total of 387 (18.82%) positive samples were identified and further analyzed by polymerase chain reaction (PCR) using primers designed to amplify DNA fragments from the small subunit ribosomal RNA. The PCR products were sequenced and the sequences were deposited in the GenBank database under accession numbers EU369377-84 and GU070730-33. Phylogenetic analysis was performed and a distances matrix generated from these sequences confirmed the existence of Cryptosporidium (C.) parvum 'mouse' genotype, C. bovis, C. andersoni, C. hominis, and C. serpentis in cattle. These results represent the first report on the prevalence and genetic identification of Cryptosporidium species, and may contribute to a better understanding of the epidemiology of Cryptosporidium in cattle in China.
Animals ; Base Sequence ; Cattle ; Cattle Diseases/epidemiology/*parasitology ; Chi-Square Distribution ; China/epidemiology ; Cryptosporidiosis/epidemiology/parasitology/*veterinary ; Cryptosporidium/genetics/*isolation & purification ; DNA, Protozoan/chemistry/genetics ; Feces/parasitology ; Female ; Molecular Sequence Data ; Phylogeny ; Polymerase Chain Reaction/veterinary ; Prevalence ; RNA, Ribosomal, 18S/chemistry/genetics ; Sequence Alignment ; Sequence Analysis, DNA

OBJECTIVECryptosporidium spp. are prevalent globally and sheep are an important zoonotic reservoir. Little data regarding the rates of Cryptosporidium infections in ovines in China are available. This study assessed the prevalence of Cryptosporidium spp. in pre-weaned ovines from Aba Tibetan and Qiang Autonomous Prefecture in the Sichuan province of China.

METHODSA total of 213 fecal samples were collected from pre-weaned ovines and were examined microscopically (following modified acid fast staining). In addition, 18S rRNA genetic sequences were amplified from fecal samples by nested PCR and phylogenetically analyzed.

RESULTSThe prevalence of Cryptosporidium in the collected samples was at 14.6% (31/213) and four isolates identified by PCR belonged to the Cryptosporidium cervine genotype (Cryptosporidium ubiquitum) demonstrating that this species was the primary sheep species found in sheep in China.

CONCLUSIONThe present study suggested that the high incidence of Cryptosporidium in sheep poses a significant public health threat and that surveillance practices must be established to prevent zoonotic disease of humans.

Animals ; China ; Cryptosporidium ; genetics ; isolation & purification ; Feces ; parasitology ; Oocysts ; microbiology ; Polymerase Chain Reaction ; Sheep ; Weaning