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Sample records for cryptosporidium parvum isolates

  1. Molecular characterization of Danish Cryptosporidium parvum isolates

    DEFF Research Database (Denmark)

    Enemark, Heidi L.; Ahrens, Peter; Juel, Cynthia Dawn

    2002-01-01

    was characterized as C. meleagridis. The porcine Cryptosporidium isolates (N = 4) revealed a pattern which was genetically distinct from human and bovine isolates. Cryptosporidium in a hedgehog (Erinaceus europaeus L.) was identified for the first time. By microsatellite sequencing the hedgehog isolate showed...

  2. Isolation of Small Number of Cryptosporidium parvum Oocyst Using Immunochromatography.

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    Elahe Ebrahimzade

    2014-12-01

    Full Text Available Cryptosporidium parvum causes severe gastroenteritis in immunocompromised human and new borne animals. The organism can be transmitted through water. Since small number of C. parvum is infectious, the aim of the present study was to develop a chromatography method for the isolation of C. parvum oocyst in samples with limited number of oocysts.Antibody was prepared against whole antigen from C. parvum oocysts, the achieved Ab bound to the sepharose 4B and used for the isolation of oocysts. Antibody against P23 bound to the sepharose 4B, used also for the isolation of C. parvum oocyst. In comparison to these both methods, 2 traditional methods (Salt floatation and 55% sucrose floatation were also performed.Both chromatography methods could bind oocysts with capacity depends on the column size. The isolated oocysts were free of bacteria. Our results showed that the traditional methods are useful for the isolation of oocysts from feces, in its smear stained with ziehl-nelsen, at least 3 oocyts are detectable in each microscopic field under 1000 X magnification. In contrast to the chromatography methods, the bacterial contamination was always observed in oocysts isolated with traditional methods.Immunochromatography could be used for the successful isolation of C. parvum oocysts from the samples containing limited number of oocysts.

  3. Preliminary molecular characterization of Cryptosporidium parvum isolates of wildlife rodents from Poland.

    Science.gov (United States)

    Bajer, A; Cacciò, S; Bednarska, M; Behnke, J M; Pieniazek, N J; Sinski, E

    2003-10-01

    Isolates of Cryptosporidium were collected from 3 species of woodland and field rodents (Clethrionomys glareolus, Microtus arvalis, and Apodemus flavicollis) and were characterized by polymerase chain reaction amplification and sequencing of fragments of the oocyst wall protein (COWP) gene and of the 18S ribosomal RNA gene. Sequence analysis of these markers revealed that the animals were infected with C. parvum, and that the genotype involved was almost identical to the mouse genotype previously described from Mus musculus. Thus, small rodents should be considered as an important reservoir of C. parvum genotypes closely related to the zoonotic genotype 2 and potentially hazardous to humans.

  4. Biological characterisation of Cryptosporidium parvum isolates of wildlife rodents in Poland.

    Science.gov (United States)

    Bednarska, Malgorzata; Bajer, Anna; Kulis, Karolina; Sinski, Edward

    2003-01-01

    The study was undertaken to characterise the C. parvum isolates originating from naturally infected woodland and field rodents: Clethrionomys glareolus (CG), Apodemus flavicollis (AF) and Microtus arvalis (MA). We found that the measurements of oocyst dimensions and oocyst morphology did not allow distinction between the parasite isolates from the 3 rodent species. The mean dimensions were: for CG 4.67 x 4.21, for AF 4.65 x 4.14 and for MA 4.66 x 4.16. These 3 groups of isolates have produced significantly different pictures of infection in C57BL/6 mice. The overall mean oocysts output was: in CG-mice 41,739, in AF-mice 18,000, in MA-mice 10,384 oocysts/1 g of faeces. From these data we suggest that rodent isolates of C. parvum could represent new subgroups in so-called "mouse" C. parvum strain. The successful cross-transmission from wild hosts to laboratory rodents and the close similarity of COWP sequence between our isolates and "mouse" genotype and between "mouse" and zoonotic genotype of C. parvum (genotype C) inform us that all these isolates should be treated as potentially hazardous for human health.

  5. Quantitative-PCR Assessment of Cryptosporidium parvum Cell Culture Infection

    OpenAIRE

    Di Giovanni, George D.; LeChevallier, Mark W.

    2005-01-01

    A quantitative TaqMan PCR method was developed for assessing the Cryptosporidium parvum infection of in vitro cultivated human ileocecal adenocarcinoma (HCT-8) cell cultures. This method, termed cell culture quantitative sequence detection (CC-QSD), has numerous applications, several of which are presented. CC-QSD was used to investigate parasite infection in cell culture over time, the effects of oocyst treatment on infectivity and infectivity assessment of different C. parvum isolates. CC-Q...

  6. Pathogenicity of Cryptosporidium parvum - evaluation of an animal infection model

    DEFF Research Database (Denmark)

    Enemark, Heidi L.; Bille-Hansen, Vivi; Lind, Peter

    2003-01-01

    With the intention of developing a standardised method for assessment of pathogenicity of Cryptosporidium parvum, the CPB-0 isolate was studied by propagation in 1-day-old calves followed by inoculation into specific pathogen free (SPF) piglets. The experiment was repeated. Diarrhoea and shedding...

  7. Crystal structure of Cryptosporidium parvum pyruvate kinase.

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    William J Cook

    Full Text Available Pyruvate kinase plays a critical role in cellular metabolism of glucose by serving as a major regulator of glycolysis. This tetrameric enzyme is allosterically regulated by different effector molecules, mainly phosphosugars. In response to binding of effector molecules and substrates, significant structural changes have been identified in various pyruvate kinase structures. Pyruvate kinase of Cryptosporidium parvum is exceptional among known enzymes of protozoan origin in that it exhibits no allosteric property in the presence of commonly known effector molecules. The crystal structure of pyruvate kinase from C. parvum has been solved by molecular replacement techniques and refined to 2.5 Å resolution. In the active site a glycerol molecule is located near the γ-phosphate site of ATP, and the protein structure displays a partially closed active site. However, unlike other structures where the active site is closed, the α6' helix in C. parvum pyruvate kinase unwinds and assumes an extended conformation. In the crystal structure a sulfate ion is found at a site that is occupied by a phosphate of the effector molecule in many pyruvate kinase structures. A new feature of the C. parvum pyruvate kinase structure is the presence of a disulfide bond cross-linking the two monomers in the asymmetric unit. The disulfide bond is formed between cysteine residue 26 in the short N-helix of one monomer with cysteine residue 312 in a long helix (residues 303-320 of the second monomer at the interface of these monomers. Both cysteine residues are unique to C. parvum, and the disulfide bond remained intact in a reduced environment. However, the significance of this bond, if any, remains unknown at this time.

  8. Detection of Cryptosporidium parvum in environmental soil and vegetables.

    Science.gov (United States)

    Hong, Semie; Kim, Kyungjin; Yoon, Sejoung; Park, Woo-Yoon; Sim, Seobo; Yu, Jae-Ran

    2014-10-01

    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.

  9. Hydrophobic and electrostatic cell surface properties of Cryptosporidium parvum.

    OpenAIRE

    Drozd, C; Schwartzbrod, J

    1996-01-01

    Microbial adhesion to hydrocarbons and microelectrophoresis were investigated in order to characterize the surface properties of Cryptosporidium parvum. Oocysts exhibited low removal rates by octane (only 20% on average), suggesting that the Cryptosporidium sp. does not demonstrate marked hydrophobic properties. A zeta potential close to -25 mV at pH 6 to 6.5 in deionized water was observed for the parasite. Measurements of hydrophobicity and zeta potential were performed as a function of pH ...

  10. Natural infection with zoonotic subtype of Cryptosporidium parvum in Capybara (Hydrochoerus hydrochaeris) from Brazil.

    Science.gov (United States)

    Meireles, Marcelo Vasconcelos; Soares, Rodrigo Martins; Bonello, Fábio; Gennari, Solange Maria

    2007-06-20

    A total of 145 capybara (Hydrochoerus hydrochaeris) fecal samples from the state of São Paulo, Brazil, were screened for Cryptosporidium spp. oocysts using the malachite green method. Eight samples (5.52%) showed positive results and were further submitted to nested PCR reaction for amplification of fragments of 18S rRNA gene and 60-kDa glycoprotein gene for determination of species, alleles and subtypes of Cryptosporidium. Sequencing of the PCR products of the 18S rRNA gene fragments and 60-kDa glycoprotein gene fragments showed that for both genes all Cryptosporidium isolates from capybara were respectively 100% genetically similar to a bovine isolate of C. parvum and to C. parvum subtype IIaA15G2R1. To the best of our knowledge this is the first report of Cryptosporidium infection in this rodent. The finding of zoonotic C. parvum infection in a semi-aquatic mammal that inhabits anthroponotic habitats raises the concern that human water supplies may be contaminated with zoonotic Cryptosporidium oocysts from wildlife.

  11. Cloning and expression of gene encoding P23 protein from Cryptosporidium parvum

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    Dinh Thi Bich Lan

    2014-12-01

    Full Text Available We cloned the cp23 gene coding P23 (glycoprotein from Cryptosporidium parvum isolated from Thua Thien Hue province, Vietnam. The coding region of cp23 gene from C. parvum is 99% similar with cp23 gene deposited in NCBI (accession number: U34390. SDS-PAGE and Western blot analysis showed that the cp23 gene in E. coli BL21 StarTM (DE3 produced polypeptides with molecular weights of approximately 37, 40 and 49 kDa. These molecules may be non-glycosylated or glycosylated P23 fusion polypeptides. Recombinant P23 protein purified by GST (glutathione S-transferase affinity chromatography can be used as an antigen for C. parvum antibody production as well as to develop diagnostic kit for C. parvum.

  12. Effects of Surfactants on Cryptosporidium parvum Mobility in Agricultural Soils from Illinois and Utah

    Science.gov (United States)

    Darnault, C. J.; Koken, E.; Jacobson, A. R.; Powelson, D.

    2011-12-01

    The occurence of the parasitic protozoan Cryptosporidium parvum in rural and agricultural watersheds due to agricultural activities and wildlife is inevitable. Understanding the behavior of C. parvum oocysts in the environment is critical for the protection of public health and the environment. To better understand the mechanisms by which the pathogen moves through soils and contaminates water resources, we study their mobility under conditions representative of real-world scenarios, where both C. parvum and chemicals that affect their fate are present in soils. Surfactants occur widely in soils due to agricultural practices such as wastewater irrigation and the application of pesticides or soil wetting agents. They affect water tension and, consequently, soil infiltration processes and the air-water interfaces in soil pores where C. parvum may be retained. We investigate the effects of surfactants on the mobility of C. parvum oocysts in agricultural soils from Illinois and Utah under unsaturated flow conditions. As it is critical to examine C. parvum in natural settings, we also developed a quantification method using RT-PCR for monitoring C. parvum oocysts in environmental soil and water samples. We optimized physico-chemical parameters to disrupt C. parvum oocysts and extract their DNA, and developed isolation methods to separate C. parvum oocysts from colloids in natural soil samples. The results of this research will lead to the development of an accurate and sensitive molecular method for the monitoring of C. parvum oocysts in environmental soil and water samples, and will further our understanding of the mechanisms controlling the behavior of C. parvum oocysts in soils, in particular the role of vadose zone processes, sorption to soil and surfactants.

  13. Cryptosporidium parvum, a potential cause of colic adenocarcinoma

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    Pinon Anthony

    2007-11-01

    Full Text Available Abstract Background Cryptosporidiosis represents a major public health problem. This infection has been reported worldwide as a frequent cause of diarrhoea. Particularly, it remains a clinically significant opportunistic infection among immunocompromised patients, causing potentially life-threatening diarrhoea in HIV-infected persons. However, the understanding about different aspects of this infection such as invasion, transmission and pathogenesis is problematic. Additionally, it has been difficult to find suitable animal models for propagation of this parasite. Efforts are needed to develop reproducible animal models allowing both the routine passage of different species and approaching unclear aspects of Cryptosporidium infection, especially in the pathophysiology field. Results We developed a model using adult severe combined immunodeficiency (SCID mice inoculated with Cryptosporidium parvum or Cryptosporidium muris while treated or not with Dexamethasone (Dex in order to investigate divergences in prepatent period, oocyst shedding or clinical and histopathological manifestations. C. muris-infected mice showed high levels of oocysts excretion, whatever the chemical immunosuppression status. Pre-patent periods were 11 days and 9.7 days in average in Dex treated and untreated mice, respectively. Parasite infection was restricted to the stomach, and had a clear preferential colonization for fundic area in both groups. Among C. parvum-infected mice, Dex-treated SCID mice became chronic shedders with a prepatent period of 6.2 days in average. C. parvum-inoculated mice treated with Dex developed glandular cystic polyps with areas of intraepithelial neoplasia, and also with the presence of intramucosal adenocarcinoma. Conclusion For the first time C. parvum is associated with the formation of polyps and adenocarcinoma lesions in the gut of Dex-treated SCID mice. Additionally, we have developed a model to compare chronic muris and parvum

  14. Interactions between Cryptosporidium parvum and the Intestinal Ecosystem

    KAUST Repository

    Douvropoulou, Olga

    2017-04-01

    Cryptosporidium parvum is an apicomplexan protozoan parasite commonly causing diarrhea, particularly in infants in developing countries. The research challenges faced in the development of therapies against Cryptosporidium slow down the process of drug discovery. However, advancement of knowledge towards the interactions of the intestinal ecosystem and the parasite could provide alternative approaches to tackle the disease. Under this perspective, the primary focus of this work was to study interactions between Cryptosporidium parvum and the intestinal ecosystem in a mouse model. Mice were treated with antibiotics with different activity spectra and the resulted perturbation of the native gut microbiota was identified by microbiome studies. In particular, 16S amplicon sequencing and Whole Genome Sequencing (WGS) were used to determine the bacterial composition and the genetic repertoire of the fecal microbial communities in the mouse gut. Following alteration of the microbial communities of mice by application of antibiotic treatment, Cryptosporidium parasites were propagated in mice with perturbed microbiota and the severity of the infection was quantified. This approach enabled the prediction of the functional capacity of the microbial communities in the mouse gut and led to the identification of bacterial taxa that positively or negatively correlate in abundance with Cryptosporidium proliferation.

  15. CP2 gene as a useful viability marker for Cryptosporidium parvum.

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    Lee, Soo-Ung; Joung, Migyo; Ahn, Myoung-Hee; Huh, Sun; Song, Hyunje; Park, Woo-Yoon; Yu, Jae-Ran

    2008-02-01

    The validity of the CP2 gene of Cryptosporidium parvum as a viability marker was evaluated using absolute quantitative real-time polymerase chain reaction (qPCR) assays. Total ribonucleic acid (RNA) was isolated from live and heat-killed C. parvum oocysts, and complementary deoxyribonucleic acid was synthesized and used as a template. The most accurate number of viable C. parvum oocysts was predicted when the CP2 gene was used as a target gene. The lower detection limit of the CP2 gene was ten oocysts, which was the most sensitive among examined target genes. With heat shock induction, only hsp70 messenger RNA (mRNA) was induced, and the predicted viable oocyst number was increased by heat shock for this marker. The CP2, hsp70, Cryptosporidium oocyst wall protein, and beta-tubulin mRNAs were not detected in heat-killed oocysts, but the 18S ribosomal ribonucleic acid (rRNA) showed heat stability until 48 h after heat killing. Although the 18S rRNA demonstrated the fastest response in crossing point (CP) value among the examined primer sets in qPCR, overestimation of viable oocysts was noted in the analysis with this gene. In conclusion, the CP2 gene was identified as the most sensitive, reliable, and accurate candidate of a viability marker of C. parvum by qPCR evaluation.

  16. Cryptosporidium parvum has an active hypusine biosynthesis pathway.

    Science.gov (United States)

    Mittal, Nimisha; Morada, Marie; Tripathi, Pankaj; Gowri, V S; Mandal, Swati; Quirch, Alison; Park, Myung Hee; Yarlett, Nigel; Madhubala, Rentala

    2014-06-01

    The protozoan parasite Cryptosporidium parvum causes severe enteric infection and diarrheal disease with substantial morbidity and mortality in untreated AIDS patients and children in developing or resource-limited countries. No fully effective treatment is available. Hypusination of eIF5A is an important post-translational modification essential for cell proliferation. This modification occurs in a two step process catalyzed by deoxyhypusine synthase (DHS) followed by deoxyhypusine hydroxylase. An ORF of 1086bp was identified in the C. parvum (Cp) genome which encodes for a putative polypeptide of 362 amino acids. The recombinant CpDHS protein was purified to homogeneity and used to probe the enzyme's mechanism, structure, and inhibition profile in a series of kinetic experiments. Sequence analysis and structural modeling of CpDHS were performed to probe differences with respect to the DHS of other species. Unlike Leishmania, Trypanosomes and Entamoeba, Cryptosporidium contains only a single gene for DHS. Phylogenetic analysis shows that CpDHS is more closely related to apicomplexan DHS than kinetoplastid DHS. Important residues that are essential for the functioning of the enzyme including NAD(+) binding residues, spermidine binding residues and the active site lysine are conserved between CpDHS and human DHS. N(1)-guanyl-1,7-diaminoheptane (GC7), a potent inhibitor of DHS caused an effective inhibition of infection and growth of C. parvum in HCT-8 cells. Copyright © 2014 Elsevier B.V. All rights reserved.

  17. Comparative efficacy of conventional primer sets in detection of Cryptosporidium parvum for diagnostic use.

    Science.gov (United States)

    Kar, Sirri; Daugschies, Arwid; Bangoura, Berit

    2010-02-01

    In this study, the sensitivity and specificity of different previously described primer sets for Cryptosporidium parvum detection by polymerase chain reaction (PCR) was evaluated. For this purpose, the primer sets defined by Cacciò et al. (FEMS Microbiol Lett 170(1):173-179, 1999) (tub), Widmer et al. (Appl Environ Microbiol 64(11):4477-4481, 1998) (btub) and Rochelle et al. (Appl Environ Microbiol 63:2029-2037, 1997) (cphsp), respectively, were used. Deoxyribonucleic acid (DNA) was isolated from three different sample materials: (1) from the faeces of an experimentally C. parvum-infected calf, (2) from purified C. parvum oocysts, and (3) from C. parvum-infected HCT-8 cell cultures. The DNA samples were subjected to PCR reactions with each of the three given primer sets to investigate sensitivity and suitability for routine use. The primers described by Cacciò et al. (FEMS Microbiol Lett 170(1):173-179, 1999) (TUB) were superior regarding sensitivity and specificity in terms of detection of C. parvum in faeces, in purified oocysts and also in cell culture, and may thus be applied for routine diagnostic use in common sample materials.

  18. Excystation of Cryptosporidium parvum at temperatures that are reached during solar water disinfection.

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    Gómez-Couso, H; Fontán-Sainz, M; Fernández-Alonso, J; Ares-Mazás, E

    2009-04-01

    Species belonging to the genera Cryptosporidium are recognized as waterborne pathogens. Solar water disinfection (SODIS) is a simple method that involves the use of solar radiation to destroy pathogenic microorganisms that cause waterborne diseases. A notable increase in water temperature and the existence of a large number of empty or partially excysted (i.e. unviable) oocysts have been observed in previous SODIS studies with water experimentally contaminated with Cryptosporidium parvum oocysts under field conditions. The aim of the present study was to evaluate the effect of the temperatures that can be reached during exposure of water samples to natural sunlight (37-50 degrees C), on the excystation of C. parvum in the absence of other stimuli. In samples exposed to 40-48 degrees C, a gradual increase in the percentage of excystation was observed as the time of exposure increased and a maximum of 53.81% of excystation was obtained on exposure of the water to a temperature of 46 degrees C for 12 h (versus 8.80% initial isolate). Under such conditions, the oocyst infectivity evaluated in a neonatal murine model decreased statistically with respect to the initial isolate (19.38% versus 100%). The results demonstrate the important effect of the temperature on the excystation of C. parvum and therefore on its viability and infectivity.

  19. [The ecology of Cryptosporidium parvum infection in small rodent populations].

    Science.gov (United States)

    Bajer, A; Bednarska, M; Siński, E

    2001-01-01

    The prevalence and abundance of Cryptosporidium parvum were studied over a three year period (1997-1999) in three species of rodents sampled from forest and abandoned fields in the Mazury Lake District, Poland. The overall prevalence was consistently higher in voles compared with Apodemus flavicollis (70.6% in Clethrionomys glareolus, 73.0% in Microtus arvalis and 27.8% in A. flavicollis). The prevalence and abundance of infection also varied across the 3 years of the study with 1998 being the year of higher prevalence and abundance of the parasite. Fewer older animals carried the infection, and their infections were relatively mild. We found no consistent pattern of seasonal changes despite the significance of seasonal differences. Host sex did not influence either the prevalence or abundance of infection with C. parvum. A great proportion of recaptured voles developed chronic infections between consecutive trapping sessions and only a small number of animals recovered. However, yellow-necked mice seem to be much more resistant to infection that became self-limiting. Our results firmly establish that the common woodland and grassland wild rodents in the Mazury Lake District constitute a significant and hazardous reservoir of C. parvum for animals and humans.

  20. Occurrence of Cryptosporidium and Giardia in recycled waters used for irrigation and first description of Cryptosporidium parvum and C. muris in Greece.

    Science.gov (United States)

    Spanakos, Gregory; Biba, Anastasia; Mavridou, Athena; Karanis, Panagiotis

    2015-05-01

    Here, we present the first time findings regarding the occurrence of Cryptosporidium and Giardia in sewage waters and the first molecular characterization of Cryptosporidium species in Greece. Biological treatment plants from three regions in Greece have been investigated. The detection of Cryptosporidium oocysts was by modified Ziehl-Neelsen acid fast (MZN-AF) and by immunofluorescence microscopy (IFT) for Cryptosporidium and Giardia (oo)cysts, whereas nested PCR based on the SSU rDNA assay was used for molecular detection of Cryptosporidium followed by sequencing for the genetic characterization of the species. In total, 73 samples (37 raw sewage samples and 38 of treated water samples) were collected and analyzed. Of the 73 water samples, 4 samples were Cryptosporidium-positive by IFT and staining, 12 samples were Cryptosporidium-positive by nested PCR; 9 samples were Giardia-positive by IFT. We showed that Cryptosporidium cysts are found both in the input and the discharge of the biological treatment plants. Molecular characterization of Cryptosporidium based on the small subunit ribosomal DNA gene resulted in the determination of Cryptosporidium parvum and Cryptosporidium muris Greek isolates. This is the first report of Cryptosporidium and Giardia occurrence in wastewaters and the first molecular identification of Cryptosporidium species in Greek environments. As the treated water is used for irrigation, or it is discharged into the sea, our findings indicate that biological treatment facilities constitute a possible risk for public health because the related species are prevalent in humans; the results invite for further epidemiological investigations to evaluate the real public health risk in Greece.

  1. Viability of Cryptosporidium parvum during ensilage of perennial ryegrass.

    Science.gov (United States)

    Merry, R J; Mawdsley, J L; Brooks, A E; Davies, D R

    1997-01-01

    The survival of Cryptosporidium parvum during ensilage of perennial ryegrass was examined in laboratory silos with herbage prepared in one of three different ways; either untreated, inoculated with a strain of Lactobacillus plantarum or by direct acidification with formic acid. The pH values of all silages initially fell below 4.5, but only formic acid-treated silage remained stable at less than pH 4 after 106 d, with the pH of the untreated and inoculant-treated silages rising to above 6. The formic acid-treated silage had a high lactic acid concentration (109 g kg-1 dry matter (DM)) and low concentrations of propionic and butyric acids after 106 d. However, the untreated and inoculant-treated silages showed an inverse relationship, with low lactic acid concentrations and high concentrations of acetic, propionic and butyric acids. These silages also contained ammonia-N concentrations in excess of 9 g kg-1 DM. In terms of the viability of Cryptosporidium parvum oocysts very few differences were seen after 14 d of ensilage with ca 50% remaining viable, irrespective of treatment and total numbers had declined from the initial level of 5.9 x 10(4) to 1 x 10(4) g(-1) fresh matter. Total oocyst numbers remained approximately the same until the end of the ensiling period, with the percentage of viable oocysts declining to 46, 41 and 32% respectively for formic acid, inoculant and untreated silages. The results are discussed in terms of changes occurring during the silage fermentation, in particular the products which may influence the survival of Cryptosporidium and implications for agricultural practice and the health of silage fed livestock.

  2. Hydrophobic and electrostatic cell surface properties of Cryptosporidium parvum.

    Science.gov (United States)

    Drozd, C; Schwartzbrod, J

    1996-04-01

    Microbial adhesion to hydrocarbons and microelectrophoresis were investigated in order to characterize the surface properties of Cryptosporidium parvum. Oocysts exhibited low removal rates by octane (only 20% on average), suggesting that the Cryptosporidium sp. does not demonstrate marked hydrophobic properties. A zeta potential close to -25 mV at pH 6 to 6.5 in deionized water was observed for the parasite. Measurements of hydrophobicity and zeta potential were performed as a function of pH and ionic strength or conductivity. Hydrophobicity maxima were observed at extreme pH values, with 40% of adhesion of oocysts to octane. It also appeared that ionic strength (estimated by conductivity) could influence the hydrophobic properties of oocysts. Cryptosporidium oocysts showed a pH-dependent surface charge, with zeta potentials becoming less negative as pH was reduced, starting at -35 mV for alkaline pH and reaching 0 at isoelectric points for pH 2.5. On the other hand, variation of surface charge with respect to conductivity of the suspension tested in this work was quite small. The knowledge of hydrophobic properties and surface charge of the parasite provides information useful in, for example, the choice of various flocculation treatments, membrane filters, and cleaning agents in connection with oocyst recovery.

  3. AGING OF CRYPTOSPORIDIUM PARVUM OOCYSTS STUDIED BY MALDI-TF MS

    Science.gov (United States)

    Cryptosporidium parvum is an obligate protozoan parasite found in surface waters. It is the etiological agent for cryptosporidiosis, a parasitic infection that causes severe gastrointestinal illness which is potentially fatal among immuno-compromised individuals. This water borne...

  4. MOLECULAR CLONING AND ANALYSIS OF THE CRYPTOSPORIDIUM PARVUM AMINOPEPTIDASE N GENE. (R829180)

    Science.gov (United States)

    Cryptosporidium parvum proteases have been associated with release of infective sporozoites from oocysts, and their specific inhibition blocks parasite excystation in vitro. Additionally, proteases have been implicated in the processing of parasite adhesion molecules fo...

  5. MOLECULAR CLONING AND ANALYSIS OF THE CRYPTOSPORIDIUM PARVUM AMINOPEPTIDASE N GENE. (R828035)

    Science.gov (United States)

    Cryptosporidium parvum proteases have been associated with release of infective sporozoites from oocysts, and their specific inhibition blocks parasite excystation in vitro. Additionally, proteases have been implicated in the processing of parasite adhesion molecules fo...

  6. Structure-activity relationship study of selective benzimidazole-based inhibitors of Cryptosporidium parvum IMPDH

    OpenAIRE

    Kirubakaran, Sivapriya; Gorla, Suresh Kumar; Sharling, Lisa; Zhang, Minjia; Liu, Xiaoping; Ray, Soumya S.; Iain S. MacPherson; Striepen, Boris; Hedstrom, Lizbeth; Gregory D Cuny

    2012-01-01

    Cryptosporidium parasites are important waterborne pathogens of both humans and animals. The C. parvum and C. hominis genomes indicate that the only route to guanine nucleotides is via inosine 5'-monophosphate dehydrogenase (IMPDH). Thus the inhibition of the parasite IMPDH presents a potential strategy for treating Cryptosporidium infections. A selective benzimidazole-based inhibitor of C. parvum IMPDH (CpIMPDH) was previously identified in a high throughput screen. Here we report a structur...

  7. AGING OF CRYPTOSPORIDIUM PARVUM OOCYSTS STUDIED BY MALDI-TOF MS

    Science.gov (United States)

    Cryptosporidium parvum is a protozoan parasite, and it causes a potentially fatal gastrointestinal illness. This water borne pathogen has been the subject of several high profile disease outbreaks in the US and abroad. C. parvum presents challenges for both compliance monitoring ...

  8. Photocatalytic inactivation of Cryptosporidium parvum on nanostructured titanium dioxide films.

    Science.gov (United States)

    Sunnotel, O; Verdoold, R; Dunlop, P S M; Snelling, W J; Lowery, C J; Dooley, J S G; Moore, J E; Byrne, J A

    2010-03-01

    Control of waterborne gastrointestinal parasites represents a major concern to water industries worldwide. In developed countries, pathogens in drinking water supplies are normally removed by sand filtration followed by chemical disinfection. Cryptosporidium spp. are generally resistant to common disinfection techniques and alternative control strategies are being sought. In the current study, the photocatalytic inactivation of C. parvum oocysts was shown to occur in buffer solution (78.4% after 180 min) and surface water (73.7% after 180 min). Viability was assessed by dye exclusion, excystation, direct examination of oocysts and a novel gene expression assay based on lactate dehydrogenase 1 (LDH1) expression levels. Collectively, this confirmed the inactivation of oocysts and scanning electron microscopy (SEM) confirmed cleavage at the suture line of oocyst cell walls, revealing large numbers of empty (ghost) cells after exposure to photocatalytic treatment.

  9. More productive in vitro culture of Cryptosporidium parvum for better study of the intra- and extracellular phases

    Directory of Open Access Journals (Sweden)

    G Perez Cordón

    2007-08-01

    Full Text Available The great difficulties in treating people and animals suffering from cryptosporidiosis have prompted the development of in vitro experimental models. Due to the models of in vitro culture, new extracellular stages of Cryptosporidium have been demonstrated. The development of these extracellular phases depends on the technique of in vitro culture and on the species and genotype of Cryptosporidium used. Here, we undertake the molecular characterization by polymerase chain reaction-restriction fragment lenght polymorphism of different Cryptosporidium isolates from calves, concluding that all are C. parvum of cattle genotype, although differing in the nucleotide at positions 472 and 498. Using these parasites, modified the in vitro culture technique for HCT-8 cells achieving greater multiplication of parasites. The HCT-8 cell cultures, for which the culture had not been renewed in seven days, were infected with C. parvum sporozoites in RPMI-1640 medium with 10% IFBS, CaCl2 and MgCl2 1 mM at pH 7.2. Percentages of cell parasitism were increased with respect to control cultures (71% at 48 h vs 14.5%, even after two weeks (47% vs 1.9%. Also, the percentage of extracellular stages augmented (25.3% vs 1.1% at 96 h. This new model of in vitro culture of C. parvum will enable easier study of the developmental phases of C. parvum in performing new chemotherapeutic assays.

  10. Two geographically separated food-borne outbreaks in Sweden linked by an unusual Cryptosporidium parvum subtype, October 2010.

    Science.gov (United States)

    Gherasim, A; Lebbad, M; Insulander, M; Decraene, V; Kling, A; Hjertqvist, M; Wallensten, A

    2012-11-15

    The number of sporadic cases of Cryptosporidium identified in the Stockholm county area increased above the expected limit during October 2010. Additionally, two food-borne outbreaks of cryptosporidiosis occurred in two other Swedish cities: Umeå (4 October) and Örebro (9 October). The outbreak investigations did not reveal any responsible food item, however fresh herbs were suspected. Thirty stool samples, originating from all three events, tested positive for Cryptosporidium oocysts. Polymerase chain reaction (PCR) and subsequent restriction fragment length polymorphism (RFLP) revealed that 27 individuals were infected with C. parvum, two with C. hominis, and one with C. felis. Using sequence analysis of the GP60 glycoprotein gene, a polymorphic marker with high intra-species diversity, we identified the same C. parvum subtype IIdA24G1 in samples from both the Umeå outbreak and the Stockholm area cases, thus indicating a possible outbreak in the Stockholm area and establishing a link between these two events. C. parvum IIdA24G1 has not previously been described in connection with a food-borne outbreak. For the outbreak in Örebro, another subtype was identified: C. parvum IIdA20G1e. These findings demonstrate that subtyping C. parvum isolates using GP60 gene amplification can be used to link cases in an outbreak investigation and we recommend its use in future similar events.

  11. Triazole inhibitors of Cryptosporidium parvum inosine 5′-monophosphate dehydrogenase

    OpenAIRE

    Maurya, Sushil K.; Gollapalli, Deviprasad R.; Kirubakaran, Sivapriya; Zhang, Minjia; Johnson, Corey R.; Benjamin, Nicole N.; Hedstrom, Lizbeth; Gregory D Cuny

    2009-01-01

    Cryptosporidium parvum is an important human pathogen and potential bioterrorism agent. This protozoan parasite cannot salvage guanine or guanosine and therefore relies on inosine 5′-monophosphate dehydrogenase (IMPDH) for biosynthesis of guanine nucleotides and hence for survival. Since C. parvum IMPDH is highly divergent from the host counterpart, selective inhibitors could potentially be used to treat cryptosporidiosis with minimal effects on its mammalian host. A series of 1,2,3-triazole ...

  12. Structure-activity relationship study of selective benzimidazole-based inhibitors of Cryptosporidium parvum IMPDH

    Science.gov (United States)

    Kirubakaran, Sivapriya; Gorla, Suresh Kumar; Sharling, Lisa; Zhang, Minjia; Liu, Xiaoping; Ray, Soumya S.; MacPherson, Iain S.; Striepen, Boris; Hedstrom, Lizbeth; Cuny, Gregory D.

    2012-01-01

    Cryptosporidium parasites are important waterborne pathogens of both humans and animals. The C. parvum and C. hominis genomes indicate that the only route to guanine nucleotides is via inosine 5'-monophosphate dehydrogenase (IMPDH). Thus the inhibition of the parasite IMPDH presents a potential strategy for treating Cryptosporidium infections. A selective benzimidazole-based inhibitor of C. parvum IMPDH (CpIMPDH) was previously identified in a high throughput screen. Here we report a structure-activity relationship study of benzimidazole-based compounds that resulted in potent and selective inhibitors of CpIMPDH. Several compounds display potent antiparasitic activity in vitro. PMID:22310229

  13. A new in vitro model using small intestinal epithelial cells to enhance infection of Cryptosporidium parvum

    Science.gov (United States)

    To better understand and study the infection of the protozoan parasite Cryptosporidium parvum, a more sensitive in vitro assay is required. In vivo, this parasite infects the epithelial cells of the microvilli layer in the small intestine. While cell infection models using colon,...

  14. INTESTINAL AND PULMONARY INFECTION BY Cryptosporidium parvum IN TWO PATIENTS WITH HIV/AIDS

    Directory of Open Access Journals (Sweden)

    Fábio Tadeu Rodrigues REINA

    2016-01-01

    Full Text Available We describe two patients with HIV/AIDS who presented pulmonary and intestinal infection caused by Cryptosporidium parvum, with a fatal outcome. The lack of available description of changes in clinical signs and radiographic characteristics of this disease when it is located in the extra-intestinal region causes low prevalence of early diagnosis and a subsequent lack of treatment.

  15. INTESTINAL AND PULMONARY INFECTION BY Cryptosporidium parvum IN TWO PATIENTS WITH HIV/AIDS

    Science.gov (United States)

    REINA, Fábio Tadeu Rodrigues; RIBEIRO, Camila Aparecida; de ARAÚJO, Ronalda Silva; MATTÉ, Maria Helena; CASTANHO, Roberto Esteves Pires; TANAKA, Ioshie Ibara; VIGGIANI, Ana Maria Ferreira Sornas; MARTINS, Luciamáre Perinetti Alves

    2016-01-01

    We describe two patients with HIV/AIDS who presented pulmonary and intestinal infection caused by Cryptosporidium parvum, with a fatal outcome. The lack of available description of changes in clinical signs and radiographic characteristics of this disease when it is located in the extra-intestinal region causes low prevalence of early diagnosis and a subsequent lack of treatment. PMID:27007564

  16. Cryptosporidium parvum GP60 subtypes in dairy cattle from Buenos Aires, Argentina

    Science.gov (United States)

    Cryptosporidium parvum from 73 dairy calves less than two months old from Buenos Aires province (Argentina) were molecularly characterized using sequence analysis of the GP60 gene. Seventy five sequences were obtained, and seven different subtypes were identified, all belonging to the IIa subtype f...

  17. Wastewater treatment with Moringa oleifera seed extract: Impact on turbidity and sedimentation of Cryptosporidium parvum oocysts

    DEFF Research Database (Denmark)

    Petersen, Heidi H.; Woolsey, Ian; Dalsgaard, Anders

    produced from seeds of the Moringa oleifera tree (MO) in reducing Cryptosporidium parvum oocysts and turbidity in wastewater. To a total of 5 x 12 glass jars containing 500 ml wastewater samples from a Danish treatment plant, 1.2 x 106 ± 1.2 x 105 oocysts L-1 were added. To half of the wastewater samples 8...

  18. (31)P NMR of apicomplexans and the effects of risedronate on Cryptosporidium parvum growth.

    Science.gov (United States)

    Moreno, B; Bailey, B N; Luo, S; Martin, M B; Kuhlenschmidt, M; Moreno, S N; Docampo, R; Oldfield, E

    2001-06-15

    High-resolution 303.6 MHz (31)P NMR spectra have been obtained of perchloric acid extracts of Plasmodium berghei trophozoites, Toxoplasma gondii tachyzoites, and Cryptosporidium parvum oocysts. Essentially complete resonance assignments have been made based on chemical shifts and by coaddition of authentic reference compounds. Signals corresponding to inorganic pyrophosphate were detected in all three species. In T. gondii and C. parvum, additional resonances were observed corresponding to linear triphosphate as well as longer chain polyphosphates. Spectra of P. berghei and T. gondii also indicated the presence of phosphomonoesters and nucleotide phosphates. We also report that the pyrophosphate analog drug, risedronate (used in bone resorption therapy), inhibits the growth of C. parvum in a mouse xenograft model. When taken together, our results indicate that all the major disease-causing apicomplexan parasites contain extensive stores of condensed phosphates and that as with Plasmodium falciparum and T. gondii, the pyrophosphate analog drug risedronate is an inhibitor of C. parvum cell growth.

  19. Revisiting the reference genomes of human pathogenic Cryptosporidium species: reannotation of C. parvum Iowa and a new C. hominis reference

    Science.gov (United States)

    Isaza, Juan P.; Galván, Ana Luz; Polanco, Victor; Huang, Bernice; Matveyev, Andrey V.; Serrano, Myrna G.; Manque, Patricio; Buck, Gregory A.; Alzate, Juan F.

    2015-01-01

    Cryptosporidium parvum and C. hominis are the most relevant species of this genus for human health. Both cause a self-limiting diarrhea in immunocompetent individuals, but cause potentially life-threatening disease in the immunocompromised. Despite the importance of these pathogens, only one reference genome of each has been analyzed and published. These two reference genomes were sequenced using automated capillary sequencing; as of yet, no next generation sequencing technology has been applied to improve their assemblies and annotations. For C. hominis, the main challenge that prevents a larger number of genomes to be sequenced is its resistance to axenic culture. In the present study, we employed next generation technology to analyse the genomic DNA and RNA to generate a new reference genome sequence of a C. hominis strain isolated directly from human stool and a new genome annotation of the C. parvum Iowa reference genome. PMID:26549794

  20. MODELING CRYPTOSPORIDIUM PARVUM OOCYST INACTIVATION AND BROMATE IN A FLOW-THROUGH OZONE CONTACTOR TREATING NATURAL WATER

    Science.gov (United States)

    A reactive transport model was developed to simultaneously predict Cryptosporidium parvum oocyst inactivation and bromate formation during ozonation of natural water. A mechanistic model previously established to predict bromate formation in organic-free synthetic waters w...

  1. Epidemiology and public health significance of Cryptosporidium isolated from cattle, buffaloes, and humans in Egypt.

    Science.gov (United States)

    Ibrahim, M A; Abdel-Ghany, A E; Abdel-Latef, G K; Abdel-Aziz, S A; Aboelhadid, S M

    2016-06-01

    The epidemiology and public health significance of Cryptosporidium species and genotypes were investigated in Beni-Suef Governorate, Egypt. A total of 610 animal fecal samples (480 from cattle and 130 from buffaloes) beside 290 stool samples from humans were collected in the period between January and December 2014. Based on the microscopic examination, the overall estimated prevalence of Cryptosporidium spp. in cattle, buffaloes, and humans was 10.2, 12.3, and 19 %, respectively. The highest detection rates were in calves less than 2 months of age (17.1 %) and diarrheic animals (13.0 %). Likewise in humans, the highest prevalence of Cryptosporidium was in infants (31.3 %) and diarrheic individuals (21.1 %). The gender distribution in humans denoted that Cryptosporidium was reported more frequently in males (21.7 %) than females (14.5 %). Based on the molecular characterization of Cryptosporidium, Cryptosporidium oocyst wall protein (COWP) and gp60 genes were successfully amplified in 36 out of 50 samples subjected to genotyping. Restriction fragment length polymorphism (RFLP) analysis of the COWP fragments revealed that Cryptosporidium parvum was the only species detected in cattle (12 isolates) and buffaloes (4 isolates), while in humans, the detected species were Cryptosporidium hominis (15 isolates) and C. parvum (5 isolates). Sequence analysis of the gp60 gene identified the subtype IIdA20G1 within C. parvum isolated from both animals and humans. The common occurrence of zoonotic subtypes of C. parvum in cattle and buffaloes highlights the potential role of these animals as significant reservoirs of infection to humans. Also, the presence of C. hominis and C. parvum in humans indicates that both anthroponotic and zoonotic pathways are expected.

  2. Intracellular levels of the viral symbiont CPV in Cryptosporidium parvum correlate with fecundity of the parasite in dairy calves

    Science.gov (United States)

    Previous reports have cited differences in clinical signs and oocyst output among strains of Cryptosporidium parvum. The purpose of this study was to determine if levels of the C. parvum intracellular viral symbiont CPV correlated with observed clinical and parasitological differences. Calves infe...

  3. Removal and fate of Cryptosporidium parvum, Clostridium perfringens and small-sized centric diatoms (Stephanodiscus hantzschii) in slow sand filters

    NARCIS (Netherlands)

    Hijnen, W.A.M.; Dullemont, Y.J.; Schijven, J.F.; Brouwer-Hanzens, Anke J.; Rosielle, M.; Medema, Gerriet Jan

    2007-01-01

    The decimal elimination capacity (DEC) of slow sand filtration (SSF) for Cryptosporidium parvum was assessed to enable quantitative microbial risk analysis of a drinking water production plant. A mature pilot plant filter of 2.56 m2 was loaded with C. parvum oocysts and two other persistent organism

  4. [Cryptosporidium parvum in wild gastropods as bioindicators of fecal contamination in terrestrial ecosystems].

    Science.gov (United States)

    Neira O, Patricia; Muñoz S, Nelson; Stanley V, Bárbara; Gosh C, Marianne; Rosales L, M José

    2010-06-01

    Cryptosporidium sp oocysts were detected in snails (Helix aspersa Miller) and slug (Deroceras reticulatum Miller) from the Valparaiso Region, Chile. Snails and slug were collected from public squares and private domestic gardens. Cryptosporidium sp oocysts were recovered from faeces of both species. Ziehl Neelsen stain, nested PCR, and sequencing analysis demonstrated a profile similar to that described for genotype C or 2 of the parasite. These results demonstrate that snails and slug could act as a reservoir and mechanic vector of C. parvum infection for humans and animals. Moreover, gastropods could serve as bioindicators of fecal soil contamination.

  5. Seven years' experience with Cryptosporidium parvum in Guinea-Bissau, West Africa

    DEFF Research Database (Denmark)

    Perch, M; Sodemann, Morten; Jakobsen, M S;

    2001-01-01

    , exceeded only by Giardia lamblia which was found in 14.8% of the samples. The highest prevalence of cryptosporidium was found in children aged 6-11 months, whereas the prevalence of other enteric parasites increased with age. Cryptosporidiosis showed a marked seasonal variation, with peak prevalences found...... consistently at the beginning of or just before the rainy seasons, May through July. By contrast, no seasonality was found for the enteric parasites Giardia lamblia or Entamoeba histolytica. We conclude that Cryptosporidium parvum is an important pathogen in children with diarrhoea....

  6. Seven years' experience with Cryptosporidium parvum in Guinea-Bissau, West Africa

    DEFF Research Database (Denmark)

    Perch, M; Sodemann, Morten; Jakobsen, M S

    2001-01-01

    , exceeded only by Giardia lamblia which was found in 14.8% of the samples. The highest prevalence of cryptosporidium was found in children aged 6-11 months, whereas the prevalence of other enteric parasites increased with age. Cryptosporidiosis showed a marked seasonal variation, with peak prevalences found...... consistently at the beginning of or just before the rainy seasons, May through July. By contrast, no seasonality was found for the enteric parasites Giardia lamblia or Entamoeba histolytica. We conclude that Cryptosporidium parvum is an important pathogen in children with diarrhoea....

  7. Effect of chlorine, blanching, freezing, and microwave heating on Cryptosporidium parvum viability inoculated on green peppers.

    Science.gov (United States)

    Duhain, G L M C; Minnaar, A; Buys, E M

    2012-05-01

    Cryptosporidium parvum oocysts have been found on the surface of vegetables in both developed and developing countries. C. parvum can contaminate vegetables via various routes, including irrigation water. This study investigated the effect of individual treatments of chlorine, blanching, blast freezing, and microwave heating, as well as combined treatments of chlorine and freezing, and chlorine and microwave heating on the viability of C. parvum oocysts inoculated on green peppers. The viability of the oocysts after the treatments was assessed using propidium iodide and a flow cytometer. Based on the propidium iodide staining, the chlorine treatments did not affect the viability of the oocysts. Blast freezing significantly inactivated 20% of the oocysts. Microwave heating and blanching significantly inactivated 93% of oocysts. Treatment with chlorine followed by blast freezing did not affect the viability of the oocysts significantly. Treatment with chlorine and microwave heating was significantly more effective than microwave heating alone and inactivated 98% of the oocysts. The study indicates that C. parvum oocysts are sensitive to heat and, to some extent, to blast freezing, but are resistant to chlorine. Therefore, the use of chlorine during vegetable processing is not a critical control point for C. parvum oocysts, and the consumption of raw or minimally processed vegetables may constitute a health risk as C. parvum oocysts can still be found viable on ready-to-eat, minimally processed vegetables.

  8. Detection of Cryptosporidium parvum Oocysts on Fresh Produce Using DNA Aptamers.

    Directory of Open Access Journals (Sweden)

    Asma Iqbal

    Full Text Available There are currently no standard methods for the detection of Cryptosporidium spp., or other protozoan parasites, in foods, and existing methods are often inadequate, with low and variable recovery efficiencies. Food testing is difficult due to the low concentrations of parasites, the difficulty in eluting parasites from some foods, the lack of enrichment methods, and the presence of PCR inhibitors. The main objectives of the present study were to obtain DNA aptamers binding to the oocyst wall of C. parvum, and to use the aptamers to detect the presence of this parasite in foods. DNA aptamers were selected against C. parvum oocysts using SELEX (Systematic Evolution of Ligands by EXponential enrichment. Ten rounds of selection led to the discovery of 14 aptamer clones with high affinities for C. parvum oocysts. For detecting parasite-bound aptamers, a simple electrochemical sensor was employed, which used a gold nanoparticle-modified screen-printed carbon electrode. This aptasensor was fabricated by self-assembling a hybrid of a thiolated ssDNA primer and the anti- C. parvum aptamer. Square wave voltammetry was employed to quantitate C. parvum in the range of 150 to 800 oocysts, with a detection limit of approximately 100 oocysts. The high sensitivity and specificity of the developed aptasensor suggests that this novel method is very promising for the detection and identification of C. parvum oocysts on spiked fresh fruits, as compared to conventional methods such as microscopy and PCR.

  9. Effects of ozone, chlorine dioxide, chlorine, and monochloramine on Cryptosporidium parvum oocyst viability.

    OpenAIRE

    Korich, D. G.; Mead, J R; Madore, M S; Sinclair, N A; Sterling, C R

    1990-01-01

    Purified Cryptosporidium parvum oocysts were exposed to ozone, chlorine dioxide, chlorine, and monochloramine. Excystation and mouse infectivity were comparatively evaluated to assess oocyst viability. Ozone and chlorine dioxide more effectively inactivated oocysts than chlorine and monochloramine did. Greater than 90% inactivation as measured by infectivity was achieved by treating oocysts with 1 ppm of ozone (1 mg/liter) for 5 min. Exposure to 1.3 ppm of chlorine dioxide yielded 90% inactiv...

  10. Cryptosporidium parvum (Apicomplexa: Cryptosporidiidae) oocyst and sporozoite antigens recognized by bovine colostral antibodies.

    OpenAIRE

    Tilley, M; Fayer, R; Guidry, A; Upton, S J; Blagburn, B. L.

    1990-01-01

    Colostral whey from seven hyperimmunized and two control cows (hyperimmune bovine colostrum) was examined by Western immunoblotting for the presence of antibody against oocysts and sporozoites of Cryptosporidium parvum, using rabbit anti-bovine immunoglobulin A (IgA), IgG1, IgG2, and IgM antibodies, followed by a horseradish peroxidase goat anti-rabbit polyvalent antibody. Although considerable variation was found in binding activity between cows on different immunization protocols, IgA and I...

  11. Effect of High-Rate Algal Ponds on Viability of Cryptosporidium parvum Oocysts

    Science.gov (United States)

    Araki, S.; Martín-Gomez, S.; Bécares, E.; De Luis-Calabuig, E.; Rojo-Vazquez, F.

    2001-01-01

    The physicochemical conditions of high-rate algal ponds were responsible for a more than 97% reduction in the infectivity of Cryptosporidium parvum oocysts in neonatal mice. The use of semipermeable bags of cellulose showed that pH, ammonia, and/or light seems to be a major factor for the inactivation of oocysts in wastewater, supporting the importance of alga-based systems for safer reuse of treated wastewater. PMID:11425762

  12. Effects of ozone, chlorine dioxide, chlorine, and monochloramine on Cryptosporidium parvum oocyst viability

    Energy Technology Data Exchange (ETDEWEB)

    Korich, D.G.; Mead, J.R.; Madore, M.S.; Sinclair, N.A.; Sterling, C.R. (Univ. of Arizona, Tucson (USA))

    1990-05-01

    Purified Cryptosporidium parvum oocysts were exposed to ozone, chlorine dioxide, chlorine, and monochloramine. Excystation and mouse infectivity were comparatively evaluated to assess oocyst viability. Ozone and chlorine dioxide more effectively inactivated oocysts than chlorine and monochloramine did. Greater than 90% inactivation as measured by infectivity was achieved by treating oocysts with 1 ppm of ozone (1 mg/liter) for 5 min. Exposure to 1.3 ppm of chlorine dioxide yielded 90% inactivation after 1 h, while 80 ppm of chlorine and 80 ppm of monochloramine required approximately 90 min for 90% inactivation. The data indicate that C. parvum oocysts are 30 times more resistant to ozone and 14 times more resistant to chlorine dioxide than Giardia cysts exposed to these disinfectants under the same conditions. With the possible exception of ozone, the use of disinfectants alone should not be expected to inactivate C. parvum oocysts in drinking water.

  13. Effects of ozone, chlorine dioxide, chlorine, and monochloramine on Cryptosporidium parvum oocyst viability.

    Science.gov (United States)

    Korich, D G; Mead, J R; Madore, M S; Sinclair, N A; Sterling, C R

    1990-01-01

    Purified Cryptosporidium parvum oocysts were exposed to ozone, chlorine dioxide, chlorine, and monochloramine. Excystation and mouse infectivity were comparatively evaluated to assess oocyst viability. Ozone and chlorine dioxide more effectively inactivated oocysts than chlorine and monochloramine did. Greater than 90% inactivation as measured by infectivity was achieved by treating oocysts with 1 ppm of ozone (1 mg/liter) for 5 min. Exposure to 1.3 ppm of chlorine dioxide yielded 90% inactivation after 1 h, while 80 ppm of chlorine and 80 ppm of monochloramine required approximately 90 min for 90% inactivation. The data indicate that C. parvum oocysts are 30 times more resistant to ozone and 14 times more resistant to chlorine dioxide than Giardia cysts exposed to these disinfectants under the same conditions. With the possible exception of ozone, the use of disinfectants alone should not be expected to inactivate C. parvum oocysts in drinking water. PMID:2339894

  14. Inactivation of Cryptosporidium parvum Oocysts in Fresh Apple Cider by UV Irradiation

    Science.gov (United States)

    Hanes, D. E.; Worobo, R. W.; Orlandi, P. A.; Burr, D. H.; Miliotis, M. D.; Robl, M. G.; Bier, J. W.; Arrowood, M. J.; Churey, J. J.; Jackson, G. J.

    2002-01-01

    This study evaluated the efficacy of UV irradiation on the inactivation of Cryptosporidium parvum oocysts in fresh apple cider. Cider was inoculated with oocysts and exposed to 14.32 mJ of UV irradiation/cm2. Oocyst viability was assessed with the gamma interferon gene knockout (GKO) mouse and infant BALB/cByJ mouse models. All GKO mice challenged with UV-treated cider demonstrated no morbidity or mortality, and infant BALB/c mice challenged with treated cider were negative for the presence of C. parvum. In contrast, the GKO mice challenged with non-UV-treated inoculated cider died and the parasite was detected in the ileums of all challenged infant mice. This study shows that UV irradiation can be used to inactivate C. parvum in fresh apple cider. PMID:12147528

  15. Chlorine Dioxide Inactivation of Cryptosporidium parvum Oocysts and Bacterial Spore Indicators

    Science.gov (United States)

    Chauret, Christian P.; Radziminski, Chris Z.; Lepuil, Michael; Creason, Robin; Andrews, Robert C.

    2001-01-01

    Cryptosporidium parvum, which is resistant to chlorine concentrations typically used in water treatment, is recognized as a significant waterborne pathogen. Recent studies have demonstrated that chlorine dioxide is a more efficient disinfectant than free chlorine against Cryptosporidium oocysts. It is not known, however, if oocysts from different suppliers are equally sensitive to chlorine dioxide. This study used both a most-probable-number–cell culture infectivity assay and in vitro excystation to evaluate chlorine dioxide inactivation kinetics in laboratory water at pH 8 and 21°C. The two viability methods produced significantly different results (P < 0.05). Products of disinfectant concentration and contact time (Ct values) of 1,000 mg · min/liter were needed to inactivate approximately 0.5 log10 and 2.0 log10 units (99% inactivation) of C. parvum as measured by in vitro excystation and cell infectivity, respectively, suggesting that excystation is not an adequate viability assay. Purified oocysts originating from three different suppliers were evaluated and showed marked differences with respect to their resistance to inactivation when using chlorine dioxide. Ct values of 75, 550, and 1,000 mg · min/liter were required to achieve approximately 2.0 log10 units of inactivation with oocysts from different sources. Finally, the study compared the relationship between easily measured indicators, including Bacillus subtilis (aerobic) spores and Clostridium sporogenes (anaerobic) spores, and C. parvum oocysts. The bacterial spores were found to be more sensitive to chlorine dioxide than C. parvum oocysts and therefore could not be used as direct indicators of C. parvum inactivation for this disinfectant. In conclusion, it is suggested that future studies address issues such as oocyst purification protocols and the genetic diversity of C. parvum, since these factors might affect oocyst disinfection sensitivity. PMID:11425712

  16. Cryptosporidium parvum-induced ileo-caecal adenocarcinoma and Wnt signaling in a mouse model

    Directory of Open Access Journals (Sweden)

    Sadia Benamrouz

    2014-06-01

    Full Text Available Cryptosporidium species are apicomplexan protozoans that are found worldwide. These parasites constitute a large risk to human and animal health. They cause self-limited diarrhea in immunocompetent hosts and a life-threatening disease in immunocompromised hosts. Interestingly, Cryptosporidium parvum has been related to digestive carcinogenesis in humans. Consistent with a potential tumorigenic role of this parasite, in an original reproducible animal model of chronic cryptosporidiosis based on dexamethasone-treated or untreated adult SCID mice, we formerly reported that C. parvum (strains of animal and human origin is able to induce digestive adenocarcinoma even in infections induced with very low inoculum. The aim of this study was to further characterize this animal model and to explore metabolic pathways potentially involved in the development of C. parvum-induced ileo-caecal oncogenesis. We searched for alterations in genes or proteins commonly involved in cell cycle, differentiation or cell migration, such as β-catenin, Apc, E-cadherin, Kras and p53. After infection of animals with C. parvum we demonstrated immunohistochemical abnormal localization of Wnt signaling pathway components and p53. Mutations in the selected loci of studied genes were not found after high-throughput sequencing. Furthermore, alterations in the ultrastructure of adherens junctions of the ileo-caecal neoplastic epithelia of C. parvum-infected mice were recorded using transmission electron microscopy. In conclusion, we found for the first time that the Wnt signaling pathway, and particularly the cytoskeleton network, seems to be pivotal for the development of the C. parvum-induced neoplastic process and cell migration of transformed cells. Furthermore, this model is a valuable tool in understanding the host-pathogen interactions associated with the intricate infection process of this parasite, which is able to modulate host cytoskeleton activities and several host

  17. Chlorine dioxide inactivation of Cryptosporidium parvum oocysts and bacterial spore indicators.

    Science.gov (United States)

    Chauret, C P; Radziminski, C Z; Lepuil, M; Creason, R; Andrews, R C

    2001-07-01

    Cryptosporidium parvum, which is resistant to chlorine concentrations typically used in water treatment, is recognized as a significant waterborne pathogen. Recent studies have demonstrated that chlorine dioxide is a more efficient disinfectant than free chlorine against Cryptosporidium oocysts. It is not known, however, if oocysts from different suppliers are equally sensitive to chlorine dioxide. This study used both a most-probable-number-cell culture infectivity assay and in vitro excystation to evaluate chlorine dioxide inactivation kinetics in laboratory water at pH 8 and 21 degrees C. The two viability methods produced significantly different results (P < 0.05). Products of disinfectant concentration and contact time (Ct values) of 1,000 mg. min/liter were needed to inactivate approximately 0.5 log(10) and 2.0 log(10) units (99% inactivation) of C. parvum as measured by in vitro excystation and cell infectivity, respectively, suggesting that excystation is not an adequate viability assay. Purified oocysts originating from three different suppliers were evaluated and showed marked differences with respect to their resistance to inactivation when using chlorine dioxide. Ct values of 75, 550, and 1,000 mg. min/liter were required to achieve approximately 2.0 log(10) units of inactivation with oocysts from different sources. Finally, the study compared the relationship between easily measured indicators, including Bacillus subtilis (aerobic) spores and Clostridium sporogenes (anaerobic) spores, and C. parvum oocysts. The bacterial spores were found to be more sensitive to chlorine dioxide than C. parvum oocysts and therefore could not be used as direct indicators of C. parvum inactivation for this disinfectant. In conclusion, it is suggested that future studies address issues such as oocyst purification protocols and the genetic diversity of C. parvum, since these factors might affect oocyst disinfection sensitivity.

  18. Putative cis-regulatory elements associated with heat shock genes activated during excystation of Cryptosporidium parvum.

    Directory of Open Access Journals (Sweden)

    Benjamin Cohn

    Full Text Available BACKGROUND: Cryptosporidiosis is a ubiquitous infectious disease, caused by the protozoan parasites Cryptosporidium hominis and C. parvum, leading to acute, persistent and chronic diarrhea worldwide. Although the complications of this disease can be serious, even fatal, in immunocompromised patients of any age, they have also been found to lead to long term effects, including growth inhibition and impaired cognitive development, in infected immunocompetent children. The Cryptosporidium life cycle alternates between a dormant stage, the oocyst, and a highly replicative phase that includes both asexual vegetative stages as well as sexual stages, implying fine genetic regulatory mechanisms. The parasite is extremely difficult to study because it cannot be cultured in vitro and animal models are equally challenging. The recent publication of the genome sequence of C. hominis and C. parvum has, however, significantly advanced our understanding of the biology and pathogenesis of this parasite. METHODOLOGY/PRINCIPAL FINDINGS: Herein, our goal was to identify cis-regulatory elements associated with heat shock response in Cryptosporidium using a combination of in silico and real time RT-PCR strategies. Analysis with Gibbs-Sampling algorithms of upstream non-translated regions of twelve genes annotated as heat shock proteins in the Cryptosporidium genome identified a highly conserved over-represented sequence motif in eleven of them. RT-PCR analyses, described herein and also by others, show that these eleven genes bearing the putative element are induced concurrent with excystation of parasite oocysts via heat shock. CONCLUSIONS/SIGNIFICANCE: Our analyses suggest that occurrences of a motif identified in the upstream regions of the Cryptosporidium heat shock genes represent parts of the transcriptional apparatus and function as stress response elements that activate expression of these genes during excystation, and possibly at other stages in the life

  19. Comparison of in vitro cell culture and a mouse assay for measuring infectivity of Cryptosporidium parvum.

    Science.gov (United States)

    Rochelle, Paul A; Marshall, Marilyn M; Mead, Jan R; Johnson, Anne M; Korich, Dick G; Rosen, Jeffrey S; De Leon, Ricardo

    2002-08-01

    In vitro cell cultures were compared to neonatal mice for measuring the infectivity of five genotype 2 isolates of Cryptosporidium parvum. Oocyst doses were enumerated by flow cytometry and delivered to animals and cell monolayers by using standardized procedures. Each dose of oocysts was inoculated into up to nine replicates of 9 to 12 mice or 6 to 10 cell culture wells. Infections were detected by hematoxylin and eosin staining in CD-1 mice, by reverse transcriptase PCR in HCT-8 and Caco-2 cells, and by immunofluorescence microscopy in Madin-Darby canine kidney (MDCK) cells. Infectivity was expressed as a logistic transformation of the proportion of animals or cell culture wells that developed infection at each dose. In most instances, the slopes of the dose-response curves were not significantly different when we compared the infectivity models for each isolate. The 50% infective doses for the different isolates varied depending on the method of calculation but were in the range from 16 to 347 oocysts for CD-1 mice and in the ranges from 27 to 106, 31 to 629, and 13 to 18 oocysts for HCT-8, Caco-2, and MDCK cells, respectively. The average standard deviations for the percentages of infectivity for all replicates of all isolates were 13.9, 11.5, 13.2, and 10.7% for CD-1 mice, HCT-8 cells, Caco-2 cells, and MDCK cells, respectively, demonstrating that the levels of variability were similar in all assays. There was a good correlation between the average infectivity for HCT-8 cells and the results for CD-1 mice across all isolates for untreated oocysts (r = 0.85, n = 25) and for oocysts exposed to ozone and UV light (r = 0.89, n = 29). This study demonstrated that in vitro cell culture was equivalent to the "gold standard," mouse infectivity, for measuring the infectivity of C. parvum and should therefore be considered a practical and accurate alternative for assessing oocyst infectivity and inactivation. However, the high levels of variability displayed by all

  20. Serological detection and epidemiology of Neospora caninum and Cryptosporidium parvum antibodies in cattle in southern Egypt.

    Science.gov (United States)

    Fereig, Ragab M; AbouLaila, Mahmoud Rezk; Mohamed, Samy G A; Mahmoud, Hassan Y A H; Ali, Alsagher O; Ali, Asmaa F; Hilali, Mosaad; Zaid, Anis; Mohamed, Adel Elsayed Ahmed; Nishikawa, Yoshifumi

    2016-10-01

    Neospora caninum and Cryptosporidium parvum are intracellular protozoan parasites that are distributed worldwide and of major economical concern in cattle industry. N. caninum can cause abortion storms and high culling rates, whereas C. parvum has zoonotic implications and can cause diarrhea in calves. There are currently no data on the prevalence of neosporosis and cryptosporidiosis in humans or animals in southern Egypt. Prevalence of these two infections was determined in a sample of cattle from two different areas in southern Egypt, Sohag and Qena, using enzyme-linked immunosorbent assay. A total 301 cattle were sampled, of which 18.9% were positive for N. caninum, 35.9% were positive for C. parvum and 10.0% were positive for both. Geographical location and breeding system were considered as potential risk factors for C. parvum infection. A higher prevalence of infection was identified on small scale farms, compared with larger, intensive systems, with a prevalence of 50.2% compared with 37.8%, respectively. Animals in Sohag had a significantly higher prevalence compared with Qena, with a seroprevalence of 46.1% compared with 31.6%, respectively. In brief, marked seroprevalence recorded in this study indicates a high incidence of N. caninum and C. parvum infections in cattle, and this necessitates the application of more effective strategies for combating these types of infections on farms in Egypt.

  1. Comparison of Assays for Sensitive and Reproducible Detection of Cell Culture-Infectious Cryptosporidium parvum and Cryptosporidium hominis in Drinking Water

    Science.gov (United States)

    Di Giovanni, George D.; Rochelle, Paul A.

    2012-01-01

    This study compared the three most commonly used assays for detecting Cryptosporidium sp. infections in cell culture: immunofluorescent antibody and microscopy assay (IFA), PCR targeting Cryptosporidium sp.-specific DNA, and reverse transcriptase PCR (RT-PCR) targeting Cryptosporidium sp.-specific mRNA. Monolayers of HCT-8 cells, grown in 8-well chamber slides or 96-well plates, were inoculated with a variety of viable and inactivated oocysts to assess assay performance. All assays detected infection with low doses of flow cytometry-enumerated Cryptosporidium parvum oocysts, including infection with one oocyst and three oocysts. All methods also detected infection with Cryptosporidium hominis. The RT-PCR assay, IFA, and PCR assay detected infection in 23%, 25%, and 51% of monolayers inoculated with three C. parvum oocysts and 10%, 9%, and 16% of monolayers inoculated with one oocyst, respectively. The PCR assay was the most sensitive, but it had the highest frequency of false positives with mock-infected cells and inactivated oocysts. IFA was the only infection detection assay that did not produce false positives with mock-infected monolayers. IFA was also the only assay that detected infections in all experiments with spiked oocysts recovered from Envirochek capsules following filtration of 1,000 liters of treated water. Consequently, cell culture with IFA detection is the most appropriate method for routine and sensitive detection of infectious Cryptosporidium parvum and Cryptosporidium hominis in drinking water. PMID:22038611

  2. Effects of hydrogen peroxide on infections of cryptosporidium parvum in vitro

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Effects of hydrogen peroxide (H2O2)on Cryptosporidium parvum infection in vitro were studied in this paper, using an oocyst excystation assay, a cell culture model, and a free radical inhibition technique. H2O2 treatment at 500 and 1 000 μmol/L significantly inhibited excystation of bleach-treated oocysts (P<0.01). Concentrations of H2O2 at 500 and 750 μmol/L resulted in a significant decrease in C. Parvum infection at 35.77% and 58.16% respectively, when compared with the untreated control at 48 hours postinoculation. Surprisingly, C. parvum infection were significantly increased by 22.21% to 39.33% following treatment with 50 (P<0.01), 100 (P<0.01) or 200 (P<0.05) μmol/L H2O2, respectively. Stimulatory effect with treatment of 100 μmol/L H2O2 was most obvious, compared with the untreated control at 48 hours postinoculation. Effects of H2O2 on C. parvum at 24, 48 and 96 hours postinoculation were similar, the highest infection being infection at 48 hours postinoculation, with the maximum inhibitory effect being seen at 96 hours postinoculation. The stimulatory and inhibitory effects of H2O2 treatment on C. parvum infection were, to a certain extent, abolished in the presence of free radical scavengers, reduced glutathione or mannitol. These observations indicate that reactive oxygen species (ROS), such as H2O2, may play a role in the course of C. parvum infection. This is the first time to demonstrate a significant action of ROS in C. parvum infection in vitro.

  3. Prevalence and Multilocus Genotyping Analysis of Cryptosporidium and Giardia Isolates from Dogs in Chiang Mai, Thailand

    Directory of Open Access Journals (Sweden)

    Sahatchai Tangtrongsup

    2017-05-01

    Full Text Available The occurrence and zoonotic potential of Cryptosporidium spp. and Giardia duodenalis isolated from dogs in Chiang Mai, Thailand were determined. Fecal samples were collected from 109 dogs between July and August 2008. Cryptosporidium spp. infection was determined by immunofluorescent assay (IFA, PCR assays that amplify Cryptosporidium heat-shock protein 70 kDa (hsp70, and two PCR assays that amplify a small subunit-ribosomal RNA (SSU-rRNA. Giardia duodenalis infection was identified using zinc sulfate centrifugal flotation, IFA, and four PCR assays that amplify the Giardia glutamate dehydrogenase (gdh, beta-giardin (bg, and generic and dog-specific assays of triosephosphate isomerase (tpi genes. Overall prevalence of Cryptosporidium spp. and G. duodenalis was 31.2% and 45.9%, respectively. Sequence analysis of 22 Cryptosporidium-positive samples and 21 Giardia-positive samples revealed the presence of C. canis in 15, and C. parvum in 7, G. duodenalis Assemblage C in 8, D in 11, and mixed of C and D in 2 dogs. Dogs in Chiang Mai were commonly exposed to Cryptosporidium spp. and G. duodenalis. Cryptosporidium parvum can be isolated from the feces of dogs, and all G. duodenalis assemblages were dog-specific. Dogs could be a reservoir for a zoonotic Cryptosporidium infection in humans, but further studies will be required to determine the clinical and zoonotic importance.

  4. Identification of putative cis-regulatory elements in Cryptosporidium parvum by de novo pattern finding

    Directory of Open Access Journals (Sweden)

    Kissinger Jessica C

    2007-01-01

    Full Text Available Abstract Background Cryptosporidium parvum is a unicellular eukaryote in the phylum Apicomplexa. It is an obligate intracellular parasite that causes diarrhea and is a significant AIDS-related pathogen. Cryptosporidium parvum is not amenable to long-term laboratory cultivation or classical molecular genetic analysis. The parasite exhibits a complex life cycle, a broad host range, and fundamental mechanisms of gene regulation remain unknown. We have used data from the recently sequenced genome of this organism to uncover clues about gene regulation in C. parvum. We have applied two pattern finding algorithms MEME and AlignACE to identify conserved, over-represented motifs in the 5' upstream regions of genes in C. parvum. To support our findings, we have established comparative real-time -PCR expression profiles for the groups of genes examined computationally. Results We find that groups of genes that share a function or belong to a common pathway share upstream motifs. Different motifs are conserved upstream of different groups of genes. Comparative real-time PCR studies show co-expression of genes within each group (in sub-sets during the life cycle of the parasite, suggesting co-regulation of these genes may be driven by the use of conserved upstream motifs. Conclusion This is one of the first attempts to characterize cis-regulatory elements in the absence of any previously characterized elements and with very limited expression data (seven genes only. Using de novo pattern finding algorithms, we have identified specific DNA motifs that are conserved upstream of genes belonging to the same metabolic pathway or gene family. We have demonstrated the co-expression of these genes (often in subsets using comparative real-time-PCR experiments thus establishing evidence for these conserved motifs as putative cis-regulatory elements. Given the lack of prior information concerning expression patterns and organization of promoters in C. parvum we

  5. Cryptosporidium parvum vaccine candidates are incompletely modified with O-linked-N-acetylgalactosamine or contain N-terminal N-myristate and S-palmitate.

    Science.gov (United States)

    Haserick, John R; Klein, Joshua A; Costello, Catherine E; Samuelson, John

    2017-01-01

    Cryptosporidium parvum (studied here) and Cryptosporidium hominis are important causes of diarrhea in infants and immunosuppressed persons. C. parvum vaccine candidates, which are on the surface of sporozoites, include glycoproteins with Ser- and Thr-rich domains (Gp15, Gp40, and Gp900) and a low complexity, acidic protein (Cp23). Here we used mass spectrometry to determine that O-linked GalNAc is present in dense arrays on a glycopeptide with consecutive Ser derived from Gp40 and on glycopeptides with consecutive Thr derived from Gp20, a novel C. parvum glycoprotein with a formula weight of ~20 kDa. In contrast, the occupied Ser or Thr residues in glycopeptides from Gp15 and Gp900 are isolated from one another. Gly at the N-terminus of Cp23 is N-myristoylated, while Cys, the second amino acid, is S-palmitoylated. In summary, C. parvum O-GalNAc transferases, which are homologs of host enzymes, densely modify arrays of Ser or Thr, as well as isolated Ser and Thr residues on C. parvum vaccine candidates. The N-terminus of an immunodominant antigen has lipid modifications similar to those of host cells and other apicomplexan parasites. Mass spectrometric demonstration here of glycopeptides with O-glycans complements previous identification C. parvum O-GalNAc transferases, lectin binding to vaccine candidates, and human and mouse antibodies binding to glycopeptides. The significance of these post-translational modifications is discussed with regards to the function of these proteins and the design of serological tests and vaccines.

  6. Detection of Giardia lamblia and Cryptosporidium parvum by direct immunofluorescence assay in stool specimen.

    Science.gov (United States)

    Rahman, M M; Hossain, M A; Paul, S K; Ahmed, S; Islam, A; Ehsan, M A; Alam, M M; Kabir, M R; Sarkar, S R

    2014-07-01

    Giardia and Cryptosporidium are the pathogens which transmitted through contaminated soil and contaminated water are significant causes of diarrhea and nutritional disorders in institutional and community peoples. Children and immune compromise persons are more vulnerable for these infections. Both Giardiasis and Cryptosporidiosis were included in 2004 as WHO Neglected Disease. So this is a major public health problem in developing countries. The present study was carried out to detect the Giardia and Cryptosporidium from diarrheic or patient having loose stool by Direct Immunofluorescence assay. The study was conducted during July 20012 to February 2013 and the work was done in Mymensingh Medical College in the department of Microbiology and in Bangladesh Agricultural University in the department of Veterinary Medicine. A total of 100 loose stools were collected from school children of different area and hospital under sadar upazilla, Mymensingh. The detection of Giardia lamblia and Cryptosporidium parvum showed the individual prevalence 8% and 4% respectively. The highest cyst/oocyst count was 85,000 and 1,000/gm of stool and the lowest being 100 and 50/gm of stool for Giardiasis and Cryptosporidiosis respectively. The detection rate of Giardia and Cryptosporidium by Immunofluorescence assay was relatively higher than the previous study done in Bangladesh and this was the first report from Bangladesh over human stool specimen using Immunofluorescence assay. So, Immunofluorescence assay could be adapted for rapid and accurate detection of Giardia and Cryptosporidium.

  7. Multiplex-Touchdown PCR to Simultaneously Detect Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the Major Causes of Traveler's Diarrhea.

    Science.gov (United States)

    Shin, Ji-Hun; Lee, Sang-Eun; Kim, Tong Soo; Ma, Da-Won; Chai, Jong-Yil; Shin, Eun-Hee

    2016-10-01

    This study aimed to develop a multiplex-touchdown PCR method to simultaneously detect 3 species of protozoan parasites, i.e., Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the major causes of traveler's diarrhea and are resistant to standard antimicrobial treatments. The target genes included the Cryptosporidium oocyst wall protein for C. parvum, Glutamate dehydrogenase for G. lamblia, and 18S ribosomal RNA (18S rRNA) for C. cayetanensis. The sizes of the amplified fragments were 555, 188, and 400 bps, respectively. The multiplex-touchdown PCR protocol using a primer mixture simultaneously detected protozoa in human stools, and the amplified gene was detected in >1×10(3) oocysts for C. parvum, >1×10(4) cysts for G. lamblia, and >1 copy of the 18S rRNA gene for C. cayetanensis. Taken together, our protocol convincingly demonstrated the ability to simultaneously detect C. parvum, G. lamblia, and C. cayetanenesis in stool samples.

  8. Prevalence of Cryptosporidium parvum in private drinking water cisterns in Bani-Kenanah district, northern Jordan.

    Science.gov (United States)

    Abo-Shehada, Mahmoud; Hindyia, Mona; Saiah, Abbass

    2004-10-01

    Due to water scarcity in Jordan, the water authority only pump the water once or twice a week to the population. Thus people in rural areas, including the Bani-Kenanah district, make the most of their water resources by storing rainwater in private reservoirs for use during periods of water shortage. These reservoirs include; underground cisterns and concrete or metal tanks. The water collected in these reservoirs is at risk of contamination. During the period March-July 2002, the three types of reservoirs from 368 households were surveyed for presence of Escherichia coli and Cryptosporidium parvum, indicators of contamination. The cistern was the most contaminated reservoir with 17% (95% CI: 13,22) for E. coli (significant, P<0.05), and 2% (95% CI: 1,4) for C. parvum. Only 1% (95% CI: 1,6) of the metal reservoirs had E. coli, while concrete reservoirs were free. No C. parvum oocysts were detected in either the concrete or metal reservoirs. Reservoirs opening at floor level and the bucket kept outside the reservoir were significant (P<0.05) enhancing risk factors for contamination with C. parvum.

  9. Cell culture-Taqman PCR assay for evaluation of Cryptosporidium parvum disinfection.

    Science.gov (United States)

    Keegan, Alexandra R; Fanok, Stella; Monis, Paul T; Saint, Christopher P

    2003-05-01

    Cryptosporidium parvum represents a challenge to the water industry and a threat to public health. In this study, we developed a cell culture-quantitative PCR assay to evaluate the inactivation of C. parvum with disinfectants. The assay was validated by using a range of disinfectants in common use in the water industry, including low-pressure UV light (LP-UV), ozone, mixed oxidants (MIOX), and chlorine. The assay was demonstrated to be reliable and sensitive, with a lower detection limit of a single infectious oocyst. Effective oocyst inactivation was achieved (>2 log(10) units) with LP-UV (20 mJ/cm(2)) or 2 mg of ozone/liter (for 10 min). MIOX and chlorine treatments of oocysts resulted in minimal effective disinfection, with disinfection systems for drinking water and recycled water.

  10. Cryptosporidium parvum (Apicomplexa: Cryptosporidiidae) oocyst and sporozoite antigens recognized by bovine colostral antibodies.

    Science.gov (United States)

    Tilley, M; Fayer, R; Guidry, A; Upton, S J; Blagburn, B L

    1990-09-01

    Colostral whey from seven hyperimmunized and two control cows (hyperimmune bovine colostrum) was examined by Western immunoblotting for the presence of antibody against oocysts and sporozoites of Cryptosporidium parvum, using rabbit anti-bovine immunoglobulin A (IgA), IgG1, IgG2, and IgM antibodies, followed by a horseradish peroxidase goat anti-rabbit polyvalent antibody. Although considerable variation was found in binding activity between cows on different immunization protocols, IgA and IgG1 in whey recognized a greater variety of C. parvum antigens than did IgG2 and IgM. A band at 9 to 10 kilodaltons appeared unique in that it was recognized only by IgA.

  11. Immunogold labeling of stages of Cryptosporidium parvum recognized by immunoglobulins in hyperimmune bovine colostrum.

    Science.gov (United States)

    Fayer, R; Barta, J R; Guidry, A J; Blagburn, B L

    1991-06-01

    Ultrathin sections of mouse ileum infected with Cryptosporidium parvum were stained by immunogold techniques. Sections first were stained with polyvalent antibodies in whey from hyperimmune bovine colostrum (HBC), then stained by secondary antibodies in rabbit antibovine IgA, IgM, IgG1, and IgG2, and lastly labeled by goat anti-rabbit gold conjugate. Examination of the immunostained specimens by electron microscopy revealed that each bovine immunoglobulin isotype in the whey recognized antigens in meronts, merozoites, microgametocytes, microgametes, and macrogamonts. Based on these findings it is hypothesized that antigens in all stages of C. parvum provide targets of opportunity for the antiparasitic activity of HBC whey antibodies thereby accounting for its efficacy as an immunotherapeutic agent.

  12. [The co-occurence of Cryptosporidium parvum, Giardia spp. and helminth infections in small rodent populations].

    Science.gov (United States)

    Bajer, Anna; Behnke, Jerzy M; Bednarska, Małgorzata; Kuliś, Karolina; Siński, Edward

    2004-01-01

    During long-term (1998-2000) studies on rodent parasite populations in Mazury lake district there were collected and analyzed data on co-occurrence of intestinal protozoa (Cryptosporidium parvum, Giardia spp.) and helminths. There were performed 178 autopsies of common vole Microtus arvalis, 85 autopsies of yellow-necked mouse Apodemus flavicollis and 386 autopsies of bank vole Clethrionomys glareolus. Positive effect of helminth infections was found in C. glareolus. Voles infected with nematode Heligmosomum mixtum showed higher prevalence of C. parvum and Giardia spp. than voles infected with Heligmosomoides glareoli. The host age took part in these interactions and positive effect of co-occurrence was mainly observed in voles older than 3 months. The other intrinsic (host sex) or extrinsic (season and year of study) factors influenced interactions between parasites. Presented results revealed that helminth infections may facilitate chronic infections of intestinal protozoa in rodent populations.

  13. Viability and fate of Cryptosporidium parvum and Giardia lamblia in tubular anaerobic digesters.

    Science.gov (United States)

    Kinyua, Maureen N; Trimmer, John; Izurieta, Ricardo; Cunningham, Jeffrey; Ergas, Sarina J

    2016-06-01

    In many developing countries where pathogenic diseases of animal waste origin, such as giardiasis and cryptosporidiosis, are often prevalent, facilities are limited to treat livestock waste. However, household-scale anaerobic digesters are currently being promoted for bioenergy production from livestock manure. Since the effluent is often used as a fertilizer for food crops, it is critical to understand the effect of environmental conditions within household-scale digesters on the viability of Cryptosporidium parvum oocysts and Giardia lamblia cysts. In this study, key environmental parameters affecting (oo)cyst inactivation were measured in four tubular anaerobic digesters, which are a type of household-scale digester promoted for treatment of swine waste in rural Costa Rica. Interviews and participant observations were used to understand digester operation and maintenance procedures. Ambient temperatures (21-24°C), near-neutral pH, total ammonia nitrogen (TAN) concentrationsGiardia lamblia and Cryptosporidium parvum were 0.155 ± 0.041 and 0.054 ± 0.006 day(-1), respectively. Temperature and volatile fatty acids were the main factors contributing to Cryptosporidium parvum and Giardia lamblia inactivation. A mathematical model was developed that predicts the concentration of (oo)cysts in the liquid effluent of tubular digesters like those observed in Costa Rica. A mathematical model was developed that predicts the concentration of (oo)cysts in the liquid effluent of tubular digesters like those observed in Costa Rica. Two dimensionless groups can be used to predict the performance of the digesters for inactivating pathogens; both dimensionless groups depend upon the average HRT in the digester. This is the first study to combine mathematical modeling with qualitative analysis, field and laboratory studies to predict the concentrations of (oo)cysts in tubular digester effluents.

  14. Application of an ELISA-type screen printed electrode-based potentiometric assay to the detection of Cryptosporidium parvum oocysts.

    Science.gov (United States)

    Laczka, Olivier; Skillman, Lucy; Ditcham, William G; Hamdorf, Brenton; Wong, Danny K Y; Bergquist, Peter; Sunna, Anwar

    2013-11-01

    We report a novel electrochemical method for the rapid detection of the parasitic protozoan, Cryptosporidium parvum. An antibody-based capture format was transferred onto screen-printed electrodes and the presence of horseradish peroxidase-labelled antibodies binding to the oocysts was potentiometrically detected. This method allowed the detection of 5 × 10(2)Cryptosporidium oocysts per mL in 60 min.

  15. Low-level detection of Cryptosporidium parvum in field water using optical microfluidic biosensors

    Science.gov (United States)

    Angus, Scott V.; Kwon, Hyuck-Jin; Yoon, Jeong-Yeol

    2012-03-01

    Cryptosporidium parvum is a difficult-to-detect protozoan that causes diarrhea in the healthy adults and death in immunocompromised individuals. While it is easy to understand the transmission routes of Cryptosporidium, it is currently difficult to identify low concentrations of Cryptosporidium, especially when following EPA method 1623, which can easily require tens of liters of water to get a positive signal. The current detection method is unacceptable and severely inefficient when taking into account the time that goes into concentrating a sample, actual assays, and training associated with the assays. Using our method, it is possible to use only 15 μL of sample, which is an immunoagglutination assay that uses Mie scatter intensity changes to detect different Cryptosporidium concentrations. In addition to creating a standard curve using a clean sample matrix (i.e., phosphate buffered saline), field samples were collected from a chlorine treated swimming pool, a sump located on a farm, and a turtle pond. Each sample had different intensity changes but the trend represented within the data was the same. This assay has a detection limit of 100-101 oocysts/mL and can be done in as little as 10 minutes.

  16. Detecting Cryptosporidium parvum and Giardia lamblia by coagulation concentration and real-time PCR quantification

    Institute of Scientific and Technical Information of China (English)

    Huining ZHANG; Xiaohu Zhang; Shuting ZHANG; Bo WEI; Qipei JIANG; Xin YU

    2013-01-01

    Rapid and sensitive pathogen detection methods are essential for the effective functioning of the water treatment industry, yet for many pathogens, effective detection and removal methods are still lacking. Cryptosporidium parvum oocysts and Giardia lamblia cysts are two of the most common waterborne pathogens currently infecting the water supply. In this study, a new method was developed for the detection of Cryptosporidium parvum oocysts and Giardia lamblia cysts. The method includes multi-steps as coagulation concentration of (oo)cysts in water, the dissolution of the resulting flocs into a small volume using acid, filtration of the (oo)cysts solution, and DNA extraction, purification, and examination using real-time PCR. The method was tested using spiked tap water and reservoir water as references, and the mean recovery ranged from 19.6% to 97.6% for oocysts, and from 51.4% to 98.7% for cysts. The method is economical and convenient, and is especially suitable for relatively high turbidity surface water.

  17. A unique hexokinase in Cryptosporidium parvum, an apicomplexan pathogen lacking the Krebs cycle and oxidative phosphorylation.

    Science.gov (United States)

    Yu, Yonglan; Zhang, Haili; Guo, Fengguang; Sun, Mingfei; Zhu, Guan

    2014-09-01

    Cryptosporidium parvum may cause virtually untreatable infections in AIDS patients, and is recently identified as one of the top four diarrheal pathogens in children in developing countries. Cryptosporidium differs from other apicomplexans (e.g., Plasmodium and Toxoplasma) by lacking many metabolic pathways including the Krebs cycle and cytochrome-based respiratory chain, thus relying mainly on glycolysis for ATP production. Here we report the molecular and biochemical characterizations of a hexokinase in C. parvum (CpHK). Our phylogenetic reconstructions indicated that apicomplexan hexokinases including CpHK were highly divergent from those of humans and animals (i.e., at the base of the eukaryotic clade). CpHK displays unique kinetic features that differ from those in mammals and Toxoplasma gondii (TgHK) in the preference towards various hexoses and its capacity to use ATP and other NTPs. CpHK also displays substrate inhibition by ATP. Moreover, 2-deoxy-D-glucose (2DG) could not only inhibit the CpHK activity, but also the parasite growth in vitro at concentrations nontoxic to host cells (IC(50) = 0.54 mM). While the exact action of 2-deoxy-D-glucose on the parasite is subject to further verification, our data suggest that CpHK and the glycolytic pathway may be explored for developing anti-cryptosporidial therapeutics.

  18. Adhesion of Cryptosporidium parvum and Giardia lamblia to solid surfaces: the role of surface charge and hydrophobicity

    NARCIS (Netherlands)

    Dai, X.; Boll, J.; Hayes, M.E.; Aston, D.E.

    2004-01-01

    Adhesion of Cryptosporidium parvum and Giardia lamblia to four materials of different surface charge and hydrophobicity was investigated. Glass beads were used with and without three polymer coatings: aminosilines (A0750), fluorosilines (T2494), an amino cationic polymer. Surface charge density and

  19. Adhesion of Cryptosporidium parvum and Giardia lamblia to solid surfaces: the role of surface charge and hydrophobicity

    NARCIS (Netherlands)

    Dai, X.; Boll, J.; Hayes, M.E.; Aston, D.E.

    2004-01-01

    Adhesion of Cryptosporidium parvum and Giardia lamblia to four materials of different surface charge and hydrophobicity was investigated. Glass beads were used with and without three polymer coatings: aminosilines (A0750), fluorosilines (T2494), an amino cationic polymer. Surface charge density and

  20. Wastewater treatment with Moringa oleifera seed extract and impact on turbidity and sedimentation of Cryptosporidium parvum oocysts

    DEFF Research Database (Denmark)

    Petersen, Heidi Huus; Woolsey, Ian David; Dalsgaard, Anders

    produced from seeds of the Moringa oleifera tree (MO) in reducing Cryptosporidium parvum oocysts and turbidity in wastewater. To a total of 5 x 12 glass jars containing 500 ml wastewater samples from a Danish treatment plant, 1.2 x 106 ± 1.2 x 105 oocysts L-1 were added. To half of the wastewater samples 8...

  1. Biofilm roughness determines Cryptosporidium parvum retention in environmental biofilms.

    Science.gov (United States)

    DiCesare, E A Wolyniak; Hargreaves, B R; Jellison, K L

    2012-06-01

    The genus Cryptosporidium is a group of waterborne protozoan parasites that have been implicated in significant outbreaks of gastrointestinal infections throughout the world. Biofilms trap these pathogens and can contaminate water supplies through subsequent release. Biofilm microbial assemblages were collected seasonally from three streams in eastern Pennsylvania and used to grow biofilms in laboratory microcosms. Daily oocyst counts in the influx and efflux flow allowed the calculation of daily oocyst retention in the biofilm. Following the removal of oocysts from the influx water, oocyst attachment to the biofilm declined to an equilibrium state within 5 days that was sustained for at least 25 days. Varying the oocyst loading rate for the system showed that biofilm retention could be saturated, suggesting that discrete binding sites determined the maximum number of oocysts retained. Oocyst retention varied seasonally but was consistent across all three sites; however, seasonal oocyst retention was not consistent across years at the same site. No correlation between oocyst attachment and any measured water quality parameter was found. However, oocyst retention was strongly correlated with biofilm surface roughness and roughness varied among seasons and across years. We hypothesize that biofilm roughness and oocyst retention are dependent on environmentally driven changes in the biofilm community rather than directly on water quality conditions. It is important to understand oocyst transport dynamics to reduce risks of human infection. Better understanding of factors controlling biofilm retention of oocysts should improve our understanding of oocyst transport at different scales.

  2. Triazole inhibitors of Cryptosporidium parvum inosine 5′-monophosphate dehydrogenase

    Science.gov (United States)

    Maurya, Sushil K.; Gollapalli, Deviprasad R.; Kirubakaran, Sivapriya; Zhang, Minjia; Johnson, Corey R.; Benjamin, Nicole N.; Hedstrom, Lizbeth; Cuny, Gregory D.

    2010-01-01

    Cryptosporidium parvum is an important human pathogen and potential bioterrorism agent. This protozoan parasite cannot salvage guanine or guanosine and therefore relies on inosine 5′-monophosphate dehydrogenase (IMPDH) for biosynthesis of guanine nucleotides and hence for survival. Since C. parvum IMPDH is highly divergent from the host counterpart, selective inhibitors could potentially be used to treat cryptosporidiosis with minimal effects on its mammalian host. A series of 1,2,3-triazole containing ether CpIMPDH inhibitors are described. A structure-activity relationship study revealed that a small alkyl group on the alpha-position of the ether was required with the (R)-enantiomer significantly more active than the (S)-enantiomer. Electron-withdrawing groups in the 3- and/or 4-positions of the pendent phenyl ring were best and conversion of the quinoline containing inhibitors to quinoline-N-oxides retained inhibitory activity both in the presence and absence of bovine serum albumin. The 1,2,3-triazole CpIMPDH inhibitors provide new tools for elucidating the role of IMPDH in C. parvum and may serve as potential therapeutics for treating cryptosporidiosis. PMID:19624136

  3. In vitro inhibitory effects of plant-derived by-products against Cryptosporidium parvum

    Directory of Open Access Journals (Sweden)

    Teichmann Klaus

    2016-01-01

    Full Text Available Disposal of organic plant wastes and by-products from the food or pharmaceutical industries usually involves high costs. In the present study, 42 samples derived from such by-products were screened in vitro against Cryptosporidium parvum, a protozoan parasite that may contaminate drinking water and cause diarrhoea. The novel bioassay was previously established in the microtitre plate format. Human ileocaecal adenocarcinoma (HCT-8 cell cultures were seeded with C. parvum oocysts and parasite development was monitored by an indirect fluorescent antibody technique (IFAT and microscopic assessment for clusters of secondary infection (CSI. Minimum inhibitory concentrations (MICs and potential detrimental effects on the host cells were determined. An ethanolic extract from olive (Olea europaea pomace, after oil pressing and phenol recovery, reproducibly inhibited C. parvum development (MIC = 250–500 μg mL−1, IC50 = 361 (279–438 μg mL−1, IC90 = 467 (398–615 μg mL−1. Accordingly, tyrosol, hydroxytyrosol, trans-coniferyl alcohol and oleuropein were selected as reference test compounds, but their contributions to the observed activity of the olive pomace extract were insignificant. The established test system proved to be a fast and efficient assay for identifying anti-cryptosporidial activities in biological waste material and comparison with selected reference compounds.

  4. In vitro inhibitory effects of plant-derived by-products against Cryptosporidium parvum

    Science.gov (United States)

    Teichmann, Klaus; Kuliberda, Maxime; Schatzmayr, Gerd; Pacher, Thomas; Zitterl-Eglseer, Karin; Joachim, Anja; Hadacek, Franz

    2016-01-01

    Disposal of organic plant wastes and by-products from the food or pharmaceutical industries usually involves high costs. In the present study, 42 samples derived from such by-products were screened in vitro against Cryptosporidium parvum, a protozoan parasite that may contaminate drinking water and cause diarrhoea. The novel bioassay was previously established in the microtitre plate format. Human ileocaecal adenocarcinoma (HCT-8) cell cultures were seeded with C. parvum oocysts and parasite development was monitored by an indirect fluorescent antibody technique (IFAT) and microscopic assessment for clusters of secondary infection (CSI). Minimum inhibitory concentrations (MICs) and potential detrimental effects on the host cells were determined. An ethanolic extract from olive (Olea europaea) pomace, after oil pressing and phenol recovery, reproducibly inhibited C. parvum development (MIC = 250–500 μg mL−1, IC50 = 361 (279–438) μg mL−1, IC90 = 467 (398–615) μg mL−1). Accordingly, tyrosol, hydroxytyrosol, trans-coniferyl alcohol and oleuropein were selected as reference test compounds, but their contributions to the observed activity of the olive pomace extract were insignificant. The established test system proved to be a fast and efficient assay for identifying anti-cryptosporidial activities in biological waste material and comparison with selected reference compounds. PMID:27627637

  5. Pilot-Scale Pulsed UV Light Irradiation of Experimentally Infected Raspberries Suppresses Cryptosporidium parvum Infectivity in Immunocompetent Suckling Mice.

    Science.gov (United States)

    Le Goff, L; Hubert, B; Favennec, L; Villena, I; Ballet, J J; Agoulon, A; Orange, N; Gargala, G

    2015-12-01

    Cryptosporidium spp., a significant cause of foodborne infection, have been shown to be resistant to most chemical food disinfectant agents and infective for weeks in irrigation waters and stored fresh vegetal produce. Pulsed UV light (PL) has the potential to inactivate Cryptosporidium spp. on surfaces of raw or minimally processed foods or both. The present study aimed to evaluate the efficacy of PL on viability and in vivo infectivity of Cryptosporidium parvum oocysts present on raspberries, a known source of transmission to humans of oocyst-forming apicomplexan pathogens. The skin of each of 20 raspberries was experimentally inoculated with five 10-μl spots of an oocyst suspension containing 6 × 10(7) oocysts per ml (Nouzilly isolate). Raspberries were irradiated by PL flashes (4 J/cm(2) of total fluence). This dose did not affect colorimetric or organoleptic characteristics of fruits. After immunomagnetic separation from raspberries, oocysts were bleached and administered orally to neonatal suckling mice. Seven days after infection, mice were euthanized, and the number of oocysts in the entire small intestine was individually assessed by immunofluorescence flow cytometry. Three of 12 and 12 of 12 inoculated mice that received 10 and 100 oocysts isolated from nonirradiated raspberries, respectively, were found infected. Four of 12 and 2 of 12 inoculated mice that received 10(3) and 10(4) oocysts from irradiated raspberries, respectively, were found infected. Oocyst counts were lower in animals inoculated with 10(3) and 10(4) oocysts from irradiated raspberries (92 ± 144 and 38 ± 82, respectively) than in animals infected with 100 oocysts from nonirradiated raspberries (35,785 ± 66,221, P = 0.008). PL irradiation achieved oocyst reductions of 2 and 3 log for an inoculum of 10(3) and 10(4) oocysts, respectively. The present pilot-scale evaluation suggests that PL is an effective mode of decontamination for raspberries and prompts further applicability

  6. Genotyping and subtyping of Giardia and Cryptosporidium isolates from commensal rodents in China.

    Science.gov (United States)

    Zhao, Z; Wang, R; Zhao, W; Qi, M; Zhao, J; Zhang, L; Li, J; Liu, A

    2015-05-01

    Cryptosporidium and Giardia are two important zoonotic intestinal parasites responsible for diarrhoea in humans and other animals worldwide. Rodents, as reservoirs or carriers of Cryptosporidium and Giardia, are abundant and globally widespread. In the present study, we collected 232 fecal specimens from commensal rodents captured in animal farms and farm neighbourhoods in China. We collected 33 Asian house rats, 168 brown rats and 31 house mice. 6.0% (14/232) and 8.2% (19/232) of these rodents were microscopy-positive for Giardia cysts and Cryptosporidium oocysts, respectively. All 14 Giardia isolates were identified as Giardia duodenalis assemblage G at a minimum of one or maximum of three gene loci (tpi, gdh and bg). By small subunit rRNA (SSU rRNA) gene sequencing, Cryptosporidium parvum (n = 12) and Cryptosporidium muris (n = 7) were identified. The gp60 gene encoding the 60-kDa glycoprotein was successfully amplified and sequenced in nine C. parvum isolates, all of which belonged to the IIdA15G1 subtype. Observation of the same IIdA15G1 subtype in humans (previously) and in rodents (here) suggests that rodents infected with Cryptosporidium have the potential to transmit cryptosporidiosis to humans.

  7. Pomegranate (Punica granatum) peel is effective in a murine model of experimental Cryptosporidium parvum.

    Science.gov (United States)

    Al-Mathal, Ebtisam M; Alsalem, Afaf M

    2012-07-01

    Cryptosporidiosis, a major health issue for neonatal calves, is caused by the parasite Cryptosporidium parvum, which is highly resistant to drug treatments. To date, many anti-parasitic drugs have been tested, but only a few have been shown to be partially effective in treating cryptosporidiosis. Previous studies have indicated that pomegranate (Punica granatum) possesses anti-plasmodium, anti-cestode, and anti-nematode activities. Therefore, the aim of this study was to evaluate the effect of P. granatum peel on suckling mice infected with experimental C. parvum. At 4days of age, 72 neonatal albino mice were randomly divided into five groups: G1: healthy controls, G2: infected/untreated controls, G3: uninfected/distilled water-treated, G4: uninfected/P. granatum peel-treated, and G5: infected/P. granatum peel-treated. Mice were experimentally-infected by oral administration of 1×10(3)C. parvum oocysts per animal. On day 7 post-inoculation (pi), treated mice received an aqueous suspension of P. granatum peel orally (3g/kg body weight). The presence of diarrhea, oocyst shedding, and weight gain/loss, and the histopathology of ileal sections were examined. Infected mice treated with the P. granatum peel suspension showed improvement in all parameters examined. Additionally, these mice did not exhibit any clinical symptoms and no deaths occurred. Oocyst shedding was very significantly reduced in the P. granatum-treated mice by day 14 pi (Parchitecture of villi from the P. granatum-treated mice on day 14 pi showed visible improvement in comparison with the infected/untreated controls, including renewed brush borders, reduced numbers of C. parvum trophozoites, and reduced lymphatic infiltration. On day 28 pi, tissues of the P. granatum-treated mice were very similar to those of healthy control mice. These results suggest that P. granatum peel is a promising anti-coccidial therapeutic treatment that lacks negative side effects.

  8. Genome-wide upstream motif analysis of Cryptosporidium parvum genes clustered by expression profile.

    Science.gov (United States)

    Oberstaller, Jenna; Joseph, Sandeep J; Kissinger, Jessica C

    2013-07-29

    There are very few molecular genetic tools available to study the apicomplexan parasite Cryptosporidium parvum. The organism is not amenable to continuous in vitro cultivation or transfection, and purification of intracellular developmental stages in sufficient numbers for most downstream molecular applications is difficult and expensive since animal hosts are required. As such, very little is known about gene regulation in C. parvum. We have clustered whole-genome gene expression profiles generated from a previous study of seven post-infection time points of 3,281 genes to identify genes that show similar expression patterns throughout the first 72 hours of in vitro epithelial cell culture. We used the algorithms MEME, AlignACE and FIRE to identify conserved, overrepresented DNA motifs in the upstream promoter region of genes with similar expression profiles. The most overrepresented motifs were E2F (5'-TGGCGCCA-3'); G-box (5'-G.GGGG-3'); a well-documented ApiAP2 binding motif (5'-TGCAT-3'), and an unknown motif (5'-[A/C] AACTA-3'). We generated a recombinant C. parvum DNA-binding protein domain from a putative ApiAP2 transcription factor [CryptoDB: cgd8_810] and determined its binding specificity using protein-binding microarrays. We demonstrate that cgd8_810 can putatively bind the overrepresented G-box motif, implicating this ApiAP2 in the regulation of many gene clusters. Several DNA motifs were identified in the upstream sequences of gene clusters that might serve as potential cis-regulatory elements. These motifs, in concert with protein DNA binding site data, establish for the first time the beginnings of a global C. parvum gene regulatory map that will contribute to our understanding of the development of this zoonotic parasite.

  9. Vaccination with pcDNA3-15/60 Naked DNA Encoding the Surface Proteinof Sporozoites in Cryptosporidium parvum

    Institute of Scientific and Technical Information of China (English)

    HEHong-xuan; ZHANGXi-chen; YINJi-gang; LIJian-hua; YANGJu

    2004-01-01

    The CP15/60 gene encoding the CP15/60 surface protein of sporozoites in Cryptosporidium parvum was obtained by PCR so as to research the nucleic vaccine against C.parvum. The eukaryotic expressing vector pcDNA3-15/60 was constructed by inserting CP15/60 gene into pcDNA3 (+) in Xho Ⅰ and EcoR Ⅰ. A vaccination protocol was the adult pregnant goats inoculated intranasally with the pcDNA3-15/60 plasmid and their offspring were infected with C.parvum oocysts. The results showed that the pcDNA3-15/60 plasmid can induce the immune response of goats and the vaccinated goats can transfer the immunity to offspring conferring protection against C.parvum infection. These suggested that the recombinant plasmid could be a DNA vaccine candidate.

  10. Vaccination with pcDNA3-15/60 Naked DNA Encoding the Surface Protein of Sporozoites in Cryptosporidium parvum

    Institute of Scientific and Technical Information of China (English)

    HE Hong-xuan; ZHANG Xi-chen; YIN Ji-gang; LI Jian-hua; YANG Ju

    2004-01-01

    The CP15/60 gene encoding the CP15/60 surface protein of sporozoites in Cryptosporidium parvum was obtained by PCR so as to research the nucleic vaccine against C.parvum. The eukaryotic expressing vector pcDNA3-15/60 was constructed by inserting CP15/60 gene into pcDNA3 (+) in Xho Ⅰ and EcoR Ⅰ. A vaccination protocol was the adult pregnant goats inoculated intranasally with the pcDNA3-15/60 plasmid and their offspring were infected with C.parvum oocysts. The results showed that the pcDNA3-15/60 plasmid can induce the immune response of goats and the vaccinated goats can transfer the immunity to offspring conferring protection against C.parvum infection. These suggested that the recombinant plasmid could be a DNA vaccine candidate.

  11. Movement of Cryptosporidium parvum Oocysts through Soils without Preferential Pathways: Exploratory Test

    Directory of Open Access Journals (Sweden)

    Christophe J. G. Darnault

    2017-06-01

    Full Text Available Groundwater contamination by oocysts of the waterborne pathogen Cryptosporidium parvum is a significant cause of animal and human disease worldwide. Although research has been undertaken in the past to determine how specific physical and chemical properties of soils affect the risk of groundwater contamination by C. parvum, there is as yet no clear conclusion concerning the range of mobility of C. parvum that one should expect in field soils. In this context, the key objective of this research was to determine the magnitude of C. parvum transport in a number of soils, under conditions in which fast and preferential transport has been successfully prevented. C. parvum oocysts were applied at the surface of different soils and subjected to artificial rainfall. Apparently for the first time, quantitative PCR was used to detect and enumerate oocysts in the soil columns and in the leachates. The transport of oocysts by infiltrating water, and the considerable retention of oocysts in soil was demonstrated for all soils, although differences in the degree of transport were observed with soils of different types. More oocysts were found in leachates from sandy loam soils than in leachates from loamy sand soils and the retention of oocysts in different soils did not significantly differ. The interaction of various processes of the hydrologic system and biogeochemical mechanisms contributed to the transport of oocysts through the soil matrix. Results suggest that the interplay of clay, organic matter, and Ca2+ facilitates and mediates the transfer of organic matter from mineral surfaces to oocysts surface, resulting in the enhanced breakthrough of oocysts through matrices of sandy loam soils compared to those of loamy sand soils. Although the number of occysts that penetrate the soil matrix account for only a small percentage of initial inputs, they still pose a significant threat to human health, especially in groundwater systems with a water table not

  12. Leaching of Salmonella Senftenberg and Cryptosporidium Parvum in Intact Clay Columns

    Science.gov (United States)

    Bech, T. B.; Forslund, A.; Dalsgaard, A.; Jacobsen, O.; Jacobsen, C. S.

    2008-12-01

    Manure application on land has been associated with both environmental and public health problems, even when management is within the current guidelines. Outbreaks of infection have been associated with water or food, including processed fruits and vegetables, contaminated with animal manure. A wide range of pathogenic microorganisms can be found in animal waste, including bacteria, protozoan, and viruses. When animal waste is disposed on agricultural land different factors will influence the risk for contaminating the groundwater. 1) Animal waste application method, rate, volume and frequency will have an effect on contamination. 2) Survival of the pathogens in the soil will e.g. depend on soil water content, temperature and pH. Salmonella species can survive up to 332 days and Cryptosporidium species can remain viable for several years in the soil environment. In the present study we compared the transport between the pathogenic bacteria S. senftenberg and the pathogenic protozoan C. parvum in intact clay columns. Furthermore, we compared the effect from surface and sub-surface manure application on the transport potential. 15 intact clay columns were placed in an outdoor multi-column lysimeter for 36 days. Manure inoculated with S. senftenberg, C. parvum and chloride was added to the soil surface or injected 8 cm into the columns. Drainage water was collected from the soil columns and DNA was extracted to quantify S. senftenberg and C. parvum by quantitative PCR. In addition S. senftenberg was enumerated by plate counting. Acid yellow was applied to selected columns to visualize the pathway down through the soil column. The highest concentration of S. senftenberg was in the first drainage sample ranging from 100-10000 CFU/ml. Breakthrough curves for chloride and S. senftenberg indicates the importance of preferential flow as well as a faster transport for the bacteria compared to chloride. C. parvum is retained to a higher degree in the soil but is still found

  13. Molecular characterization of Cryptosporidium isolates from pre-weaned calves in western France in relation to age.

    Science.gov (United States)

    Rieux, Anaïs; Paraud, Carine; Pors, Isabelle; Chartier, Christophe

    2013-10-18

    Eighteen pre-weaned female calves from a single beef cattle herd in western France were sampled weekly from birth to 21/2 months of age in order to characterize Cryptosporidium oocyst output. 182 fecal samples were screened for the presence of oocysts after concentration using immunofluorescence analysis. DNA was extracted from positive samples and a PCR-RFLP protocol, with the restriction enzyme SspI and MboII, to amplify the partial SSU rRNA gene was performed. For the subtyping of Cryptosporidium parvum, a gp60 PCR was carried out. All animals excreted oocysts at at least one sampling date and 80% of the calves presented a mild diarrhea at at least one occasion, with no mortality. The prevalence of excretion reached 94% when calves were 17-23 days of age. The mean number of oocysts at the peak of excretion (10-16 days) was 5 × 10(5) oocysts per gram of feces. PCR-RFLP analysis was successful for 61 of 84 positive samples: 14 were identified as C. parvum, 15 as Cryptosporidium bovis, and 22 as Cryptosporidium ryanae. Ten mixed infections with all combinations of these species were also identified. Calves excreted the following Cryptosporidium species: C. parvum between 7 and 27 days of age, C. bovis between 11 and 38 days and C. ryanae from 19 to 72 days. The IIaA15G2R1 zoonotic subtype of C. parvum subtype was the only subtype identified. We observed the presence of different Cryptosporidium species depending on the age of the animals. This study showed that C. parvum and C. bovis can infect beef calf neonates at similar levels of oocyst excretion with or without clinical signs and that C. parvum isolates had zoonotic potential. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Simultaneous detection of Giardia lamblia and Cryptosporidium parvum (oo)cysts in soil using immunomagnetic separation and direct fluorescent antibody staining.

    Science.gov (United States)

    Orlofsky, Ezra; Gillor, Osnat; Melli, Ann; Miller, Woutrina; Wuertz, Stefan; Bernstein, Nirit; Shapiro, Karen

    2013-09-01

    An improved approach for simultaneous detection of Cryptosporidium parvum and Giardia lamblia (oo)cysts in soil is described. Recoveries>70% were obtained for concentrations>55 and 21 (oo)cysts g(-1) for C. parvum and G. lamblia, respectively. The limits of detection were determined to be<5 (oo)cysts g(-1) soil.

  15. Risk of Handling as a Route of Exposure to Infectious Waterborne Cryptosporidium parvum Oocysts via Atlantic Blue Crabs (Callinectes sapidus)▿

    Science.gov (United States)

    Graczyk, Thaddeus K.; McOliver, Cynthia; Silbergeld, Ellen K.; Tamang, Leena; Roberts, Jennifer D.

    2007-01-01

    Commercial Atlantic blue crabs (Callinectes sapidus) were exposed to 2.0 × 104 infectious waterborne oocysts of Cryptosporidium parvum. The study demonstrated that blue crabs can transfer C. parvum oocysts to persons involved in handling or preparing crabs and that they may contaminate other surfaces or products during storage. PMID:17449680

  16. Parasites and malignancies, a review, with emphasis on digestive cancer induced by Cryptosporidium parvum (Alveolata: Apicomplexa).

    Science.gov (United States)

    Benamrouz, S; Conseil, V; Creusy, C; Calderon, E; Dei-Cas, E; Certad, G

    2012-05-01

    The International Agency for Research on Cancer (IARC) identifies ten infectious agents (viruses, bacteria, parasites) able to induce cancer disease in humans. Among parasites, a carcinogenic role is currently recognized to the digenetic trematodes Schistosoma haematobium, leading to bladder cancer, and to Clonorchis sinensis or Opisthorchis viverrini, which cause cholangiocarcinoma. Furthermore, several reports suspected the potential association of other parasitic infections (due to Protozoan or Metazoan parasites) with the development of neoplastic changes in the host tissues. The present work shortly reviewed available data on the involvement of parasites in neoplastic processes in humans or animals, and especially focused on the carcinogenic power of Cryptosporidium parvum infection. On the whole, infection seems to play a crucial role in the etiology of cancer.

  17. Parasites and malignancies, a review, with emphasis on digestive cancer induced by Cryptosporidium parvum (Alveolata: Apicomplexa

    Directory of Open Access Journals (Sweden)

    Benamrouz S.

    2012-05-01

    Full Text Available The International Agency for Research on Cancer (IARC identifies ten infectious agents (viruses, bacteria, parasites able to induce cancer disease in humans. Among parasites, a carcinogenic role is currently recognized to the digenetic trematodes Schistosoma haematobium, leading to bladder cancer, and to Clonorchis sinensis or Opisthorchis viverrini, which cause cholangiocarcinoma. Furthermore, several reports suspected the potential association of other parasitic infections (due to Protozoan or Metazoan parasites with the development of neoplastic changes in the host tissues. The present work shortly reviewed available data on the involvement of parasites in neoplastic processes in humans or animals, and especially focused on the carcinogenic power of Cryptosporidium parvum infection. On the whole, infection seems to play a crucial role in the etiology of cancer.

  18. Presence of Cryptosporidium scrofarum, C. suis and C. parvum subtypes IIaA16G2R1 and IIaA13G1R1 in Eurasian wild boars (Sus scrofa).

    Science.gov (United States)

    García-Presedo, Ignacio; Pedraza-Díaz, Susana; González-Warleta, Marta; Mezo, Mercedes; Gómez-Bautista, Mercedes; Ortega-Mora, Luis Miguel; Castro-Hermida, José Antonio

    2013-09-23

    The aim of the present study was to identify the species of Cryptosporidium infecting Eurasian wild boars (Sus scrofa) in Galicia (NW, Spain). A sampling of 209 wild boars shot in different game preserves was carried out during the hunting season in 2009-2010. All samples were examined for Cryptosporidium infection, using both immunological and molecular tools. Cryptosporidium oocysts in faecal samples were identified using a direct immunofluorescence technique with monoclonal antibodies (DFA). The presence of Cryptosporidium DNA was determined using nested PCR involving amplification of a fragment of the small-subunit (SSU) ribosomal RNA gene (SSU rRNA). A total of 35 (16.7%) samples tested positive with both techniques. However, sequencing was only possible in 27 samples. Cryptosporidium scrofarum, Cryptosporidium suis and Cryptosporidium parvum oocysts were identified in 19, 5 and 3 of the samples, respectively. Moreover, C. scrofarum was detected as a dominant species infecting all age groups (juveniles, sub adults and adults). Sequence analyses of the glycoprotein (GP60) gene revealed the presence of C. parvum subtypes IIaA16G2R1 in 2 juveniles and IIaA13G1R1 in 1 sub adult wild boar. These species and subtypes have previously been described in human patients, indicating that isolates from asymptomatic wild boars might have zoonotic potential. This is the first report of the presence of C. scrofarum, C. suis and C. parvum subtypes IIaA16G2R1 and IIaA13G1R1 in wild boars (S. scrofa) in Spain.

  19. Prevalencia del protozoario Cryptosporidium parvum en caninos en tres consultorios veterinarios en Tunja

    Directory of Open Access Journals (Sweden)

    Elkin Yadir Rodríguez-Becerra

    2011-02-01

    Full Text Available Se determinó por primera vez la prevalencia de Cryptosporidium parvum en caninos, en tres consultorios veterinarios de la ciudad de Tunja/Colombia, durante el periodo de tiempo comprendido entre agosto y diciembre de 2006. En el estudio se procesaron 132 muestras de materia fecal de perros, en el laboratorio clínico de la Universidad Pedagógica y Tecnológica de Colombia, UPTC, para lo cual se tuvo en cuenta su edad, raza y sexo, si presentaba diarrea, si se encontraba vacunado y/o desparasitado y, por último, a cada una de las muestras se le realizó la coloración de Ziehl – Nielsen modificada. El análisis estadístico se realizó mediante el programa Epiinfo. Del total de muestras examinadas, el 16,38% de los caninos fueron positivos a C. parvum. Del total de las muestras remitidas, un 47% correspondió a hembras y un 52,3% machos, con edades entre un mes y 13 años, con un promedio de tres años, de los cuales 34,8% presentaron diarrea. Las muestras positivas a C. parvum  se encontraron en perros entre 1 mes y 13 años de edad, con un promedio de cuatro años.

  20. Optimization of benzoxazole-based inhibitors of Cryptosporidium parvum inosine 5'-monophosphate dehydrogenase.

    Science.gov (United States)

    Gorla, Suresh Kumar; Kavitha, Mandapati; Zhang, Minjia; Chin, James En Wai; Liu, Xiaoping; Striepen, Boris; Makowska-Grzyska, Magdalena; Kim, Youngchang; Joachimiak, Andrzej; Hedstrom, Lizbeth; Cuny, Gregory D

    2013-05-23

    Cryptosporidium parvum is an enteric protozoan parasite that has emerged as a major cause of diarrhea, malnutrition, and gastroenteritis and poses a potential bioterrorism threat. C. parvum synthesizes guanine nucleotides from host adenosine in a streamlined pathway that relies on inosine 5'-monophosphate dehydrogenase (IMPDH). We have previously identified several parasite-selective C. parvum IMPDH (CpIMPDH) inhibitors by high-throughput screening. In this paper, we report the structure-activity relationship (SAR) for a series of benzoxazole derivatives with many compounds demonstrating CpIMPDH IC50 values in the nanomolar range and >500-fold selectivity over human IMPDH (hIMPDH). Unlike previously reported CpIMPDH inhibitors, these compounds are competitive inhibitors versus NAD(+). The SAR study reveals that pyridine and other small heteroaromatic substituents are required at the 2-position of the benzoxazole for potent inhibitory activity. In addition, several other SAR conclusions are highlighted with regard to the benzoxazole and the amide portion of the inhibitor, including preferred stereochemistry. An X-ray crystal structure of a representative E·IMP·inhibitor complex is also presented. Overall, the secondary amine derivative 15a demonstrated excellent CpIMPDH inhibitory activity (IC50 = 0.5 ± 0.1 nM) and moderate stability (t1/2 = 44 min) in mouse liver microsomes. Compound 73, the racemic version of 15a, also displayed superb antiparasitic activity in a Toxoplasma gondii strain that relies on CpIMPDH (EC50 = 20 ± 20 nM), and selectivity versus a wild-type T. gondii strain (200-fold). No toxicity was observed (LD50 > 50 μM) against a panel of four mammalian cells lines.

  1. Inactivation of Cryptosporidium parvum oocysts using medium- and low-pressure ultraviolet radiation.

    Science.gov (United States)

    Craik, S A; Weldon, D; Finch, G R; Bolton, J R; Belosevic, M

    2001-04-01

    The effect of ultraviolet radiation from low- and medium-pressure mercury arc lamps on Cryptosporidium parvum oocysts was studied using a collimated beam apparatus. Experiments were conducted using parasites suspended in both filtered surface water and phosphate buffered laboratory water. Inactivation of oocysts was measured as reduction in infectivity using a CD-1 neonatal mouse model and was found to be a non-linear function of UV dose over the range of germicidal doses tested (0.8-119 mJ/cm2). Oocyst inactivation increased rapidly with UV dose at doses less than 25 mJ/cm2 with two and three log-units inactivation at approximately 10 and 25 mJ/cm2, respectively. The cause of significant leveling-off and tailing in the UV inactivation curve at higher doses was not determined. Maximum measured oocyst inactivation ranged from 3.4 to greater than 4.9 log-units and was dependent on different batches of parasites. Water type and temperature, the concentration of oocysts in the suspension, and the UV irradiance did not have significant impacts on oocyst inactivation. When compared on the basis of germicidal UV dose, the oocysts were equally sensitive to low- and medium-pressure UV radiation. With respect to Cryptosporidium, both low- and medium-pressure ultraviolet radiation are attractive alternatives to conventional chemical disinfection methods in drinking water treatment.

  2. Use of a Sentinel System for Field Measurements of Cryptosporidium parvum Oocyst Inactivation in Soil and Animal Waste

    OpenAIRE

    Jenkins, M. B.; Walker, M. J.; Bowman, D. D.; Anthony, L. C.; Ghiorse, W C

    1999-01-01

    A small-volume sentinel chamber was developed to assess the effects of environmental stresses on survival of sucrose-Percoll-purified Cryptosporidium parvum oocysts in soil and animal wastes. Chambers were tested for their ability to equilibrate with external chemical and moisture conditions. Sentinel oocysts were then exposed to stresses of the external environment that affected their viability (potential infectivity), as indicated by results of a dye permeability assay. Preliminary laborato...

  3. Transport and survival of Cryptosporidium Parvum Oocysts in Soil Columns Following Applications of Raw and Separated Liquid Slurry

    DEFF Research Database (Denmark)

    Petersen, H.H.; Enemark, Heidi L.; Olsen, A.;

    The widespread waterborne pathogen Cryptosporidium parvum is primarily transmitted to humans via contaminated drinking and recreational water. Nearly all drinking water in Denmark is groundwater, but this can be contaminated with oocysts from application of contaminated manure to the field. Oocysts...... transport to groundwater requires that the oocysts are transported through soil and bedrock to the water table. The purpose of this study was to determine the potential transport of the protozoan pathogen C. parvum through soil to land drains and, subsequently water courses in a laboratory setup using...... simulated rainfall and six 20 cm long replicate intact soil columns. Two types of contaminated slurry, namely raw slurry and the separated liquid fraction of the slurry were applied ten cm into the soil, following irrigation once a week over a four week period. C. parvum oocysts were detected...

  4. Cryptosporidium parvum genotype IIa and Giardia duodenalis assemblage A in Mytilus galloprovincialis on sale at local food markets.

    Science.gov (United States)

    Giangaspero, Annunziata; Papini, Roberto; Marangi, Marianna; Koehler, Anson V; Gasser, Robin B

    2014-02-03

    To date, there has been no study to establish the genotypic or subgenotypic identities of Cryptosporidium and Giardia in edible shellfish. Here, we explored the genetic composition of these protists in Mytilus galloprovincialis (Mediterranean mussel) purchased from three markets in the city of Foggia, Italy, from May to December 2012. Samples from the digestive glands, gills and haemolymph were tested by nested PCR, targeting DNA regions within the 60 kDa glycoprotein (gp60) gene of Cryptosporidium, and the triose-phosphate isomerase (tpi) and β-giardin genes of Giardia. In total, Cryptosporidium and Giardia were detected in 66.7% of mussels (M. galloprovincialis) tested. Cryptosporidium was detected mostly between May and September 2012. Sequencing of amplicons showed that 60% of mussels contained Cryptosporidium parvum genotype IIa (including subgenotypes A15G2R1, IIaA15G2 and IIaA14G3R1), 23.3% Giardia duodenalis assemblage A, and 6.6% had both genetic types. This is the first report of these types in fresh, edible shellfish, particularly the very commonly consumed M. galloprovincialis from highly frequented fish markets. These genetic types of Cryptosporidium and Giardia are known to infect humans and thus likely to represent a significant public health risk. The poor observance of hygiene rules by vendors, coupled to the large numbers of M. galloprovincialis sold and the eating habits of consumers in Italy, call for more effective sanitary measures pertaining to the selling of fresh shellfish in street markets.

  5. Cloning and Characterization of the Acidic Ribosomal Protein P2 of Cryptosporidium parvum, a New 17-Kilodalton Antigen▿

    Science.gov (United States)

    Priest, Jeffrey W.; Kwon, James P.; Montgomery, Joel M.; Bern, Caryn; Moss, Delynn M.; Freeman, Amanda R.; Jones, Cara C.; Arrowood, Michael J.; Won, Kimberly Y.; Lammie, Patrick J.; Gilman, Robert H.; Mead, Jan R.

    2010-01-01

    Cryptosporidium infection is commonly observed among children and immunocompromised individuals in developing countries, but large-scale outbreaks of disease among adults have not been reported. In contrast, outbreaks of cryptosporidiosis in the United States and Canada are increasingly common among patients of all ages. Thus, it seems likely that residents of regions where Cryptosporidium is highly endemic acquire some level of immunity, while residents of the developed world do not. A new immunodominant Cryptosporidium parvum antigen in the 15- to 17-kDa size range was identified as the Cryptosporidium parvum 60S acidic ribosomal protein P2 (CpP2). We developed a recombinant protein-based enzyme-linked immunosorbent assay for serologic population surveillance for antibodies that was 89% sensitive and 92% specific relative to the results of the large-format Western blot assay. The human IgG response is directed almost exclusively toward the highly conserved, carboxy-terminal 15 amino acids of the protein. Although IgG antibody cross-reactivity was documented with sera from patients with acute babesiosis, the development of an anti-CpP2 antibody response in our Peru study population correlated better with Cryptosporidium infection than with infection by any other parasitic protozoan. In Haiti, the prevalence of antibodies to CpP2 plateaus at 11 to 20 years of age. Because anti-CpP2 IgG antibodies were found only among residents of countries in the developing world where Cryptosporidium infection occurs early and often, we propose that this response may be a proxy for the intensity of infection and for acquired immunity. PMID:20410328

  6. Obtaining hyperimmune anti-Cryptosporidium parvum ovine colostrum. A study of the humoral immune response in immunized sheep.

    Science.gov (United States)

    Martín-Gómez, S; Alvarez-Sánchez, M A; Rojo-Vázquez, F A

    2006-01-01

    Three ewes were immunized five times over a 2-month period prior to giving birth by intramuscular injection, oral administration and intramammary infusion of antigen and viable or freeze-dried Cryptosporidium parvum oocyst solution emulsified with Freund's complete and incomplete adjuvant. Two animals served as controls and another two as adjuvant controls. Serum was collected at first immunization and thereafter every 2 to 4 weeks. Colostrum and milk were collected as well. All samples were assayed for C. parvum-specific antibodies using an enzyme-linked immunosorbent assay methodology, and Western blotting was used to recognize the C. parvum antigens. Hyperimmunization resulted in a progressive and significant increase in specific anti-C. parvum serum IgG, IgA and IgM titres, with the highest values noted at the point of lambing. Titres decreased slightly in milk, although they were in all cases higher than those in the control animals. Moreover, some 30 bands of C. parvum were recognized.

  7. Molecular and parasitological study of cryptosporidium isolates from cattle in ilam, west of iran.

    Directory of Open Access Journals (Sweden)

    Mahmoud Mahami Oskouei

    2014-09-01

    Full Text Available Cryptosporidiosis is one of the most important parasitic infections in human and animals. This study was designed for survey on the prevalence of Cryptosporidium infection in farms of Ilam, west of Iran, using parasitology method and genotyping by Nested PCR-RFLP.Fecal samples of 217 cattle were collected fresh and directly from the rectum of cattle. All of the samples were examined by microscopic observation after staining with modified Ziehl-Neelsen (MZN. Genomic DNA extracted by using EURx DNA kit. A Nested PCR-RFLP protocol amplifying 825 bp fragment of 18s rRNA gene conducted to differentiate species and genotyping of the isolates using SspI and VspI as restriction enzymes.The prevalence of Cryptosporidium infection in cattle using both methods is 3.68%. Most of the positive cattle were calves under six months. Species diagnosis carried out by digesting the secondary PCR product with SspI that C. parvum generated 3 visible bands of 448, 247 and 106 bp and digested by VspI restriction enzyme generated 2 visible bands of 628 and 104bp. In this investigation all of the positive samples were Cryptosporidium parvum.C. parvum (bovine genotype detected in all positive cattle samples in Ilam, west of Iran. The results of the present study can help for public health care systems to prevention and management of cryptosporidiosis in cattle and the assessment of cattle cryptosporidiosis as a reservoir for the human infection.

  8. Expression of Cryptosporidium parvum thioredoxin peroxidase in COS-7 cells confers radioprotection.

    Science.gov (United States)

    Hong, Semie; Kim, Jae-Hwan; Yoon, Sejoung; Kim, Kyoungjin; Sim, Seobo; Park, Woo-Yoon; Yu, Jae-Ran

    2016-04-01

    Cryptosporidium parvum is one of the most radioresistant organisms identified to date. In a previous study, we found that thioredoxin peroxidase (CpTPx) was significantly upregulated in this species following exposure to high dose (10 kGy) of γ-irradiation. To assess the potential of CpTPx to confer radioprotection in mammalian cells, it was expressed in COS-7 African green monkey kidney cells (CpTPx-COS7). For comparison, the thioredoxin peroxidase of Cryptosporidium muris (CmTPx) was also expressed in these cells (CmTPx-COS7 cells), which has been confirmed to have lesser antioxidant activity than CpTPx in the previous study. Notably, the survival rates of CpTPx-COS7 cells were significantly higher (12-22%) at 72 h after 8 Gy irradiation than CmTPx-COS7 or non-transfected COS-7 (ntCOS-7) counterparts. In addition, CpTPx revealed a 50% of ROS reduction in irradiated CpTPx-COS7 cells, while γ-H2AX DNA damage marker expression was not significantly changed. Furthermore, the amount of apoptosis only increased to about 120% after 2-8 Gy irradiation compared to 200-300% increase observed in ntCOS-7 cells. CmTPx was shown to have antioxidant and DNA damage protection activities; however, these activities were always lower than those of CpTPx. These results suggest that the potent antioxidant and protective activities of CpTPx are well conserved in this cell-based system and that CpTPx contributed to the radioprotection of mammalian cells through its exceptional antioxidant activity.

  9. Genotyping of Cryptosporidium isolates from human clinical cases in Poland.

    Science.gov (United States)

    Bajer, Anna; Bednarska, Małgorzata; Cacciò, Simone M; Wolska-Kuśnierz, Beata; Heropolitanska-Pliszka, Edyta; Bernatowska, Ewa; Wielopolska, Małgorzata; Paziewska, Anna; Welc-faleciak, Renata; Siński, Edward

    2008-06-01

    Cryptosporidium spp. infection is usually self-limited in immunocompetent hosts but can be severe and life threatening in children and in immunocompromised individuals including those with primary or acquired immunodeficiencies. One hundred and three faecal samples were collected from 35 hospitalised patients with different symptoms and tested for the presence of the parasite. Cryptosporidium oocysts were found in four of 35 patients (11.4%) using Ziehl-Neelsen staining of faecal smears and immunofluorescence assay, whereas 12 (34.3%) samples tested positive by nested polymerase chain reaction assay. Cryptosporidium DNA was detected in one bile sample but not in a liver tissue biopsy sample collected from a patient who suffered from sclerosing cholangitis. Sequence analysis of oocyst wall protein and beta-tubulin gene fragments revealed three different parasite species (Cryptosporidium hominis, Cryptosporidium meleagridis and Cryptosporidium parvum) in children with primary immunodeficiencies, whereas only C. parvum was found in immunocompetent individuals and in those with secondary immunodeficiencies. This study has revealed a high prevalence of Cryptosporidium infection in hospitalised patients in Poland and confirmed that molecular techniques enable a more sensitive detection of the parasite.

  10. A novel Cryptosporidium parvum antigen, CP2, preferentially associates with membranous structures.

    Science.gov (United States)

    O'Hara, Steven P; Yu, Jae-Ran; Lin, Jim Jung-Ching

    2004-03-01

    The present study addresses the cloning and characterization of a Cryptosporidium parvum antigen, CP2. Sequencing of cDNA and genomic clones revealed a novel gene capable of coding a message of 2,136 nucleotides flanked by 28 and 140 nucleotides of the 5'- and 3'-noncoding regions, respectively. The deduced amino acid sequence suggests that CP2 is a secreted and/or membrane protein. Immunofluorescence microscopy detected CP2 enrichment in sporozoites that subsequently appeared to encase type I meronts in infected HCT-8 cells. Immunogold electron microscopy revealed that CP2 consistently localized to membranous structures throughout development. In addition, progression from macrogametocyte to sporulated oocyst revealed CP2 initially at the periphery of amylopectin-like granules, in the cytoplasm and discrete vesicles, the parasitophorous vacuole, on the surface of sporozoites, and finally on the parasitophorous vacuole membrane (PVM). The observed expression pattern suggests that CP2 may be involved in the invasion process and/or PVM integrity.

  11. Multiplex-Touchdown PCR to Simultaneously Detect Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the Major Causes of Traveler’s Diarrhea

    Science.gov (United States)

    Shin, Ji-Hun; Lee, Sang-Eun; Kim, Tong Soo; Ma, Da-Won; Chai, Jong-Yil; Shin, Eun-Hee

    2016-01-01

    This study aimed to develop a multiplex-touchdown PCR method to simultaneously detect 3 species of protozoan parasites, i.e., Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the major causes of traveler’s diarrhea and are resistant to standard antimicrobial treatments. The target genes included the Cryptosporidium oocyst wall protein for C. parvum, Glutamate dehydrogenase for G. lamblia, and 18S ribosomal RNA (18S rRNA) for C. cayetanensis. The sizes of the amplified fragments were 555, 188, and 400 bps, respectively. The multiplex-touchdown PCR protocol using a primer mixture simultaneously detected protozoa in human stools, and the amplified gene was detected in >1×103 oocysts for C. parvum, >1×104 cysts for G. lamblia, and >1 copy of the 18S rRNA gene for C. cayetanensis. Taken together, our protocol convincingly demonstrated the ability to simultaneously detect C. parvum, G. lamblia, and C. cayetanenesis in stool samples. PMID:27853120

  12. The Cryptosporidium parvum ApiAP2 gene family: insights into the evolution of apicomplexan AP2 regulatory systems.

    Science.gov (United States)

    Oberstaller, Jenna; Pumpalova, Yoanna; Schieler, Ariel; Llinás, Manuel; Kissinger, Jessica C

    2014-07-01

    We provide the first comprehensive analysis of any transcription factor family in Cryptosporidium, a basal-branching apicomplexan that is the second leading cause of infant diarrhea globally. AP2 domain-containing proteins have evolved to be the major regulatory family in the phylum to the exclusion of canonical regulators. We show that apicomplexan and perkinsid AP2 domains cluster distinctly from other chromalveolate AP2s. Protein-binding specificity assays of C. parvum AP2 domains combined with motif conservation upstream of co-regulated gene clusters allowed the construction of putative AP2 regulons across the in vitro life cycle. Orthologous Apicomplexan AP2 (ApiAP2) expression has been rearranged relative to the malaria parasite P. falciparum, suggesting ApiAP2 network rewiring during evolution. C. hominis orthologs of putative C. parvum ApiAP2 proteins and target genes show greater than average variation. C. parvum AP2 domains display reduced binding diversity relative to P. falciparum, with multiple domains binding the 5'-TGCAT-3', 5'-CACACA-3' and G-box motifs (5'-G[T/C]GGGG-3'). Many overrepresented motifs in C. parvum upstream regions are not AP2 binding motifs. We propose that C. parvum is less reliant on ApiAP2 regulators in part because it utilizes E2F/DP1 transcription factors. C. parvum may provide clues to the ancestral state of apicomplexan transcriptional regulation, pre-AP2 domination. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  13. Direct ELISA aided coprological diagnosis of Cryptosporidium parvum infection in diarrheic neonatal calves in Mosul city, Iraq

    Directory of Open Access Journals (Sweden)

    Israa Al-Robaiee

    2014-03-01

    Full Text Available This study was conducted for the detection of Cryptosporidium (C. parvum infection in neonatal calves. A total of 220 fecal samples (diarrheic 110 and nondiarrheic 110 of neonatal calves were collected from Mosul city, Iraq over a period of 16 months from November 2010 to March 2012. The age of the calves ranged from 1 to 30 days. All the fecal samples were analyzed by capture direct ELISA. The infection was found in 29.0% (n=32/110 of the diarrheic calves. The infection was mostly prevalent (p<0.001 in the calves of three weeks of age. No C. parvum infection could be detected in the nondiarrheic animals.

  14. A multi-locus study of cryptosporidium parasites isolated from patients living in iran, Malawi, Nigeria, the United kingdom, and Vietnam.

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    Salman Ghaffari

    2014-03-01

    Full Text Available Cryptosporidium species are important cause of diarrheal diseases in both developing and developed countries. This study aimed to compare the performance of several molecular methods for identification of Cryptosporidium species, and to detect genetic variation among each of these species isolated from Iran, Malawi, Nigeria, Vietnam and the United Kingdom.The oocysts DNA samples were derived from 106 Cryptosporidium positive feces. Polymerase chain reaction, PCR- restriction fragment length polymorphism and DNA sequence analysis of the 18S rRNA and the Cryptosporidium oocysts wall protein genes; PCR and DNA sequence analysis of a fragment of 70 kDa heat shock protein and 60 kDa glycoprotein genes were carried out.Based on these analysis, three species of Cryptosporidium including C. hominis, C. parvum and C. meleagridis, and both C. hominis and C. parvum were found in Iranian and the UK samples, respectively. Also, three C. hominis (Ib, Ib3& Id and three C. parvum (IIa, IIc & IId subtypes were identified by sequence analysis of the GP60 gene. Of these, C. hominis Ib was predominant and interestingly, one subgenotype (C. hominis Ib A10G2 accounted for the majority of the samples.The current study demonstrates the complex subtypes of Cryptosporidium isolates in both developing and developed countries. This is the first report of C. parvum IId subgenotype and three new subtypes of C. parvum IIa in the UK, a new subtype of C. hominis Id from Malawi; and the first multi-locus study of three species of Cryptosporidium in human from Iran.

  15. Temporal and spatial dynamics of Cryptosporidium parvum infection on dairy farms in the New York City Watershed: a cluster analysis based on crude and Bayesian risk estimates

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    Mohammed Hussni O

    2010-06-01

    Full Text Available Abstract Background Cryptosporidium parvum is one of the most important biological contaminants in drinking water that produces life threatening infection in people with compromised immune systems. Dairy calves are thought to be the primary source of C. parvum contamination in watersheds. Understanding the spatial and temporal variation in the risk of C. parvum infection in dairy cattle is essential for designing cost-effective watershed management strategies to protect drinking water sources. Crude and Bayesian seasonal risk estimates for Cryptosporidium in dairy calves were used to investigate the spatio-temporal dynamics of C. parvum infection on dairy farms in the New York City watershed. Results Both global (Global Moran's I and specific (SaTScan cluster analysis methods revealed a significant (p C. parvum infection in all herds in the summer (p = 0.002, compared to the rest of the year. Bayesian estimates did not show significant spatial autocorrelation in any season. Conclusions Although we were not able to identify seasonal clusters using Bayesian approach, crude estimates highlighted both temporal and spatial clusters of C. parvum infection in dairy herds in a major watershed. We recommend that further studies focus on the factors that may lead to the presence of C. parvum clusters within the watershed, so that monitoring and prevention practices such as stream monitoring, riparian buffers, fencing and manure management can be prioritized and improved, to protect drinking water supplies and public health.

  16. Multicenter Evaluation of BD Max Enteric Parasite Real-Time PCR Assay for Detection of Giardia duodenalis, Cryptosporidium hominis, Cryptosporidium parvum, and Entamoeba histolytica

    Science.gov (United States)

    Relich, R. F.; Doyle, L.; Espina, N.; Fuller, D.; Karchmer, T.; Lainesse, A.; Mortensen, J. E.; Pancholi, P.; Veros, W.; Harrington, S. M.

    2016-01-01

    Common causes of chronic diarrhea among travelers worldwide include protozoan parasites. The majority of parasitic infections are caused by Giardia duodenalis, Entamoeba histolytica, Cryptosporidium parvum, and Cryptosporidium hominis. Similarly, these species cause the majority of parasitic diarrhea acquired in the United States. Detection of parasites by gold standard microscopic methods is time-consuming and requires considerable expertise; enzyme immunoassays and direct fluorescent-antibody (DFA) stains have lowered hands-on time for testing, but improvements in sensitivity and technical time may be possible with a PCR assay. We performed a clinical evaluation of a multiplex PCR panel, the enteric parasite panel (EPP), for the detection of these common parasites using the BD Max instrument, which performs automated extraction and amplification. A total of 2,495 compliant specimens were enrolled, including 2,104 (84%) specimens collected prospectively and 391 (16%) specimens collected retrospectively. Approximately equal numbers were received in 10% formalin (1,273 specimens) and unpreserved (1,222 specimens). The results from the EPP were compared to those from alternate PCR and bidirectional sequencing (APCR), as well as DFA (G. duodenalis and C. parvum or C. hominis) or trichrome stain (E. histolytica). The sensitivity and specificity for prospective and retrospective specimens combined were 98.2% and 99.5% for G. duodenalis, 95.5% and 99.6 for C. parvum or C. hominis, and 100% and 100% for E. histolytica, respectively. The performance of the FDA-approved BD Max EPP compared well to the reference methods and may be an appropriate substitute for microscopic examination or immunoassays. PMID:27535690

  17. Biotin- and Glycoprotein-Coated Microspheres as Surrogates for Studying Filtration Removal of Cryptosporidium parvum in a Granular Limestone Aquifer Medium.

    Science.gov (United States)

    Stevenson, M E; Blaschke, A P; Toze, S; Sidhu, J P S; Ahmed, W; van Driezum, I H; Sommer, R; Kirschner, A K T; Cervero-Aragó, S; Farnleitner, A H; Pang, L

    2015-07-01

    Members of the genus Cryptosporidium are waterborne protozoa of great health concern. Many studies have attempted to find appropriate surrogates for assessing Cryptosporidium filtration removal in porous media. In this study, we evaluated the filtration of Cryptosporidium parvum in granular limestone medium by the use of biotin- and glycoprotein-coated carboxylated polystyrene microspheres (CPMs) as surrogates. Column experiments were carried out with core material taken from a managed aquifer recharge site in Adelaide, Australia. For the experiments with injection of a single type of particle, we observed the total removal of the oocysts and glycoprotein-coated CPMs, a 4.6- to 6.3-log10 reduction of biotin-coated CPMs, and a 2.6-log10 reduction of unmodified CPMs. When two different types of particles were simultaneously injected, glycoprotein-coated CPMs showed a 5.3-log10 reduction, while the uncoated CPMs displayed a 3.7-log10 reduction, probably due to particle-particle interactions. Our results confirm that glycoprotein-coated CPMs are the most accurate surrogates for C. parvum; biotin-coated CPMs are slightly more conservative, while unmodified CPMs are markedly overly conservative for predicting C. parvum removal in granular limestone medium. The total removal of C. parvum observed in our study suggests that granular limestone medium is very effective for the filtration removal of C. parvum and could potentially be used for the pretreatment of drinking water and aquifer storage recovery of recycled water.

  18. DEVELOPMENT OF A CT EQUATION TAKING INTO CONSIDERATION THE EFFECT OF LOT VARIABILITY ON THE INACTIVATION OF CRYPTOSPORIDIUM PARVUM OOCYSTS WITH OZONE

    Science.gov (United States)

    Cryptosporidium parvum oocysts are prevalent in surface water and ground water under the influence of surface water, and are difficult to inactivate using free chlorine, the most common disinfectant currently used for treating drinking water. In contrast, it has been shown...

  19. Biotin- and glycoprotein-coated microspheres: potential surrogates for studying filtration of cryptosporidium parvum in porous media.

    Science.gov (United States)

    Pang, Liping; Nowostawska, Urszula; Weaver, Louise; Hoffman, Gabrielle; Karmacharya, Anjuman; Skinner, Alexandra; Karki, Naveena

    2012-11-06

    Cryptosporidium parvum is a waterborne pathogen, yet no suitable surrogate has been established for quantifying its filtration removal in porous media. Carboxyl polystyrene microspheres with size, density, and shape similar to C. parvum were coated with biotin (free and containing amine, NH(2)) and glycoprotein. These biomolecules have isoelectric points similar to C. parvum (pH ≈ 2), and glycoprotein is a major type of surface protein that oocysts possess. Zeta potential (ζ) and filtration removal of particles in sand of two different grain sizes were examined. Compared to unmodified microspheres, modified microspheres achieved a superior match to the oocysts in ζ, concentration, mass recovery, and collision coefficient. They showed the same log reduction in concentration as oocysts, whereas results from unmodified microspheres deviated by 1 order of magnitude. Of the three types of modified microspheres, glycoprotein-coated microspheres best resembled oocyst concentration, despite having ζ similar to NH(2)-biotin-coated microspheres, suggesting that surface protein also played an important role in particle attachment on solid surfaces. With further validation in environmental conditions, the surrogates developed here could be a cost-effective new tool for assessing oocyst filtration in porous media, for example, to evaluate the performance of sand filters in water and wastewater treatment, water recycling through riverbank filtration, and aquifer recharge.

  20. Capture of water-borne colloids in granular beds using external electric fields: improving removal of Cryptosporidium parvum.

    Science.gov (United States)

    Kulkarni, Pramod; Dutari, Gabriel; Weingeist, David; Adin, Avner; Haught, Roy; Biswas, Pratim

    2005-03-01

    Suboptimal coagulation in water treatment plants often results in reduced removal efficiency of Cryptosporidium parvum oocysts by several orders of magnitude (J. AWWA 94(6) (2002) 97, J. AWWA 93(12) (2001) 64). The effect of external electric field on removal of C. parvum oocysts in packed granular beds was studied experimentally. A cylindrical configuration of electrodes, with granular media in the annular space was used. A negative DC potential was applied to the central electrode. No coagulants or flocculants were used and filtration was performed with and without application of an electric field to obtain improvement in removal efficiency. Results indicate that removal of C. parvum increased from 10% to 70% due to application of field in fine sand media and from 30% to 96% in MAGCHEM media. All other test particles (Kaolin and polystyrene latex microspheres) used in the study also exhibited increased removal in the presence of an electric field. Single collector efficiencies were also computed using approximate trajectory analysis, modified to account for the applied external electric field. The results of these calculations were used to qualitatively explain the trends in the experimental observations.

  1. Calf-level risk factors for neonatal diarrhea and shedding of Cryptosporidium parvum in Ontario dairy calves.

    Science.gov (United States)

    Trotz-Williams, Lise A; Wayne Martin, S; Leslie, Kenneth E; Duffield, Todd; Nydam, Daryl V; Peregrine, Andrew S

    2007-11-15

    This work was conducted to investigate calf-level factors that influence the risk of neonatal diarrhea and shedding of Cryptosporidium parvum oocysts in calves, on dairy farms in Ontario with histories of calf diarrhea or cryptosporidiosis. Fecal samples were collected weekly for 4 weeks from each of 1045 calves under 30 days of age on 11 dairy farms in south-western Ontario during the summer of 2003 and the winter of 2004. A questionnaire designed to gather information on calf-level management factors was administered on farm for each calf in the study. Samples were examined for C. parvum oocysts by microscopy, and a subset of specimens was also tested for enterotoxigenic Escherichia coli, Salmonella, bovine rotavirus and bovine coronavirus. The consistency of each sample was scored and recorded at the time of collection in order to assess the presence or absence of diarrhea. In addition, a blood sample was taken from each calf upon enrollment in the study, for assessment of maternal antibody transfer and for polymerase chain reaction testing for persistent bovine viral diarrhea virus infection. Using the GLLAMM function in Stata 9.0, multilevel regression techniques were employed to investigate associations between management practices and the risk of C. parvum shedding or diarrhea. C. parvum oocysts were detected in the feces of 78% of the 919 calves from which all four fecal samples had been collected. Furthermore, 73% of the 846 calves for which all four fecal consistency scores had been recorded were diarrheic at the time of collection of at least one sample. Significant predictors of the calf-level risk of C. parvum shedding included the use of calf diarrhea prophylaxis in pregnant cows, and the type of maternity facilities in which the calves were born. Factors associated with an increased risk of diarrhea were leaving the calf with the dam for more than an hour after birth, and the birth of a calf in the summer as opposed to winter. Calves shedding C

  2. Evaluation of solar photocatalysis using TiO2 slurry in the inactivation of Cryptosporidium parvum oocysts in water.

    Science.gov (United States)

    Abeledo-Lameiro, María Jesús; Ares-Mazás, Elvira; Gómez-Couso, Hipólito

    2016-10-01

    Cryptosporidium is a genus of enteric protozoan parasites of medical and veterinary importance, whose oocysts have been reported to occur in different types of water worldwide, offering a great resistant to the water treatment processes. Heterogeneous solar photocatalysis using titanium dioxide (TiO2) slurry was evaluated on inactivation of Cryptosporidium parvum oocysts in water. Suspensions of TiO2 (0, 63, 100 and 200mg/L) in distilled water (DW) or simulated municipal wastewater treatment plant (MWTP) effluent spiked with C. parvum oocysts were exposed to simulated solar radiation. The use of TiO2 slurry at concentrations of 100 and 200mg/L in DW yielded a high level of oocyst inactivation after 5h of exposure (4.16±2.35% and 15.03±4.54%, respectively, vs 99.33±0.58%, initial value), representing a good improvement relative to the results obtained in the samples exposed without TiO2 (51.06±9.35%). However, in the assays carried out using simulated MWTP effluent, addition of the photocatalyst did not offer better results. Examination of the samples under bright field and epifluorescence microscopy revealed the existence of aggregates comprising TiO2 particles and parasitic forms, which size increased as the concentration of catalyst and the exposure time increased, while the intensity of fluorescence of the oocyst walls decreased. After photocatalytic disinfection process, the recovery of TiO2 slurry by sedimentation provided a substantial reduction in the parasitic load in treated water samples (57.81±1.10% and 82.10±2.64% for 200mg/L of TiO2 in DW and in simulated MWTP effluent, respectively). Although further studies are need to optimize TiO2 photocatalytic disinfection against Cryptosporidium, the results obtained in the present study show the effectiveness of solar photocatalysis using TiO2 slurry in the inactivation of C. parvum oocysts in distilled water.

  3. Presence of Cryptosporidium parvum and Giardia lamblia in water samples from Southeast Asia: towards an integrated water detection system.

    Science.gov (United States)

    Kumar, Thulasi; Abd Majid, Mohamad Azlan; Onichandran, Subashini; Jaturas, Narong; Andiappan, Hemah; Salibay, Cristina C; Tabo, Hazel A L; Tabo, Norbel; Dungca, Julieta Z; Tangpong, Jitbanjong; Phiriyasamith, Sucheep; Yuttayong, Boonyaorn; Polseela, Raxsina; Do, Binh Nhu; Sawangjaroen, Nongyao; Tan, Tian-Chye; Lim, Yvonne A L; Nissapatorn, Veeranoot

    2016-01-13

    Access to clean and safe drinking water that is free from pathogenic protozoan parasites, especially Cryptosporidium parvum and Giardia lamblia that cause gastrointestinal illness in humans, is still an issue in Southeast Asia (SEA). This study is the first attempt to detect the aforementioned protozoan parasites in water samples from countries in SEA, using real-time polymerase chain reaction (qPCR) assays. A total of 221 water samples of 10 l each were collected between April and October 2013 from Malaysia (53), Thailand (120), the Philippines (33), and Vietnam (15). A physicochemical analysis was conducted. The water samples were processed in accordance with the US Environmental Protection Agency's methods 1622/1623.1, microscopically observed and subsequently screened using qPCR assays. Cryptosporidium oocysts were detected in treated water samples from the Philippines (1/10), with a concentration of 0.06 ± 0.19 oocyst/L, and untreated water samples from Thailand (25/93), Malaysia (17/44), and the Philippines (11/23), with concentrations ranging from 0.13 ± 0.18 to 0.57 ± 1.41 oocyst/L. Giardia cysts were found in treated water samples from the Philippines (1/10), with a concentration of 0.02 ± 0.06 cyst/L, and in untreated water samples from Thailand (20/93), Vietnam (5/10), Malaysia (22/44), and the Philippines (16/23), with concentrations ranging from 0.12 ± 0.3 to 8.90 ± 19.65 cyst/L. The pathogens C. parvum and G. lamblia were detected using using qPCR assays by targeting the 138-bp fragment and the small subunit gene, respectively. C. parvum was detected in untreated water samples from the Philippines (1/23) and Malaysia (2/44), whilst, G. lamblia detected was detected in treated water samples from the Philippines (1/10) and in untreated water samples from Thailand (21/93), Malaysia (12/44), and the Philippines (17/23). Nitrate concentration was found to have a high positive correlation with (oo)cyst (0.993). The presence of

  4. Prevalência do Cryptosporidium parvum em crianças abaixo de 5 anos, residentes na zona urbana de Campo Grande, MS, Brasil, 1996

    OpenAIRE

    Oshiro Elisa Teruya; Dorval Maria Elizabeth Cavalheiros; Nunes Vânia Lúcia Brandão; Silva Marcos Aurélio Almeida; Said Luis Augusto Morelli

    2000-01-01

    O presente estudo visou estabelecer a prevalência de Cryptosporidium parvum em crianças abaixo de 5 anos, residentes na zona urbana de Campo Grande, MS, 1996/97, através de exames coprológicos e avaliar epidemiologicamente os casos diagnosticados. Tratou-se de um estudo transversal com inquérito domiciliar, onde foram examinadas 1051 amostras fecais, processadas segundo a técnica de Blagg, utilizando-se a coloração de Ziehl-Neelsen modificada para a pesquisa de oocistos de C. parvum. Concluiu...

  5. Removal and fate of Cryptosporidium parvum, Clostridium perfringens and small-sized centric diatoms (Stephanodiscus hantzschii) in slow sand filters.

    Science.gov (United States)

    Hijnen, Wim A M; Dullemont, Yolanda J; Schijven, Jack F; Hanzens-Brouwer, Anke J; Rosielle, Martine; Medema, Gertjan

    2007-05-01

    The decimal elimination capacity (DEC) of slow sand filtration (SSF) for Cryptosporidium parvum was assessed to enable quantitative microbial risk analysis of a drinking water production plant. A mature pilot plant filter of 2.56m(2) was loaded with C. parvum oocysts and two other persistent organisms as potential surrogates; spores of Clostridium perfringens (SCP) and the small-sized (4-7microm) centric diatom (SSCD) Stephanodiscus hantzschii. Highly persistent micro-organisms that are retained in slow sand filters are expected to accumulate and eventually break through the filter bed. To investigate this phenomenon, a dosing period of 100 days was applied with an extended filtrate monitoring period of 150 days using large-volume sampling. Based on the breakthrough curves the DEC of the filter bed for oocysts was high and calculated to be 4.7log. During the extended filtrate monitoring period the spatial distribution of the retained organisms in the filter bed was determined. These data showed little risk of accumulation of oocysts in mature filters most likely due to predation by zooplankton. The DEC for the two surrogates, SCP and SSCD, was 3.6 and 1.8log, respectively. On basis of differences in transport behaviour, but mainly because of the high persistence compared to the persistence of oocysts, it was concluded that both spores of sulphite-reducing clostridia (incl. SCP) and SSCD are unsuited for use as surrogates for oocyst removal by slow sand filters. Further research is necessary to elucidate the role of predation in Cryptosporidium removal and the fate of consumed oocysts.

  6. NF-kappaB p65-dependent transactivation of miRNA genes following Cryptosporidium parvum infection stimulates epithelial cell immune responses.

    Science.gov (United States)

    Zhou, Rui; Hu, Guoku; Liu, Jun; Gong, Ai-Yu; Drescher, Kristen M; Chen, Xian-Ming

    2009-12-01

    Cryptosporidium parvum is a protozoan parasite that infects the gastrointestinal epithelium and causes diarrheal disease worldwide. Innate epithelial immune responses are key mediators of the host's defense to C. parvum. MicroRNAs (miRNAs) regulate gene expression at the posttranscriptional level and are involved in regulation of both innate and adaptive immune responses. Using an in vitro model of human cryptosporidiosis, we analyzed C. parvum-induced miRNA expression in biliary epithelial cells (i.e., cholangiocytes). Our results demonstrated differential alterations in the mature miRNA expression profile in cholangiocytes following C. parvum infection or lipopolysaccharide stimulation. Database analysis of C. parvum-upregulated miRNAs revealed potential NF-kappaB binding sites in the promoter elements of a subset of miRNA genes. We demonstrated that mir-125b-1, mir-21, mir-30b, and mir-23b-27b-24-1 cluster genes were transactivated through promoter binding of the NF-kappaB p65 subunit following C. parvum infection. In contrast, C. parvum transactivated mir-30c and mir-16 genes in cholangiocytes in a p65-independent manner. Importantly, functional inhibition of selected p65-dependent miRNAs in cholangiocytes increased C. parvum burden. Thus, we have identified a panel of miRNAs regulated through promoter binding of the NF-kappaB p65 subunit in human cholangiocytes in response to C. parvum infection, a process that may be relevant to the regulation of epithelial anti-microbial defense in general.

  7. NF-kappaB p65-dependent transactivation of miRNA genes following Cryptosporidium parvum infection stimulates epithelial cell immune responses.

    Directory of Open Access Journals (Sweden)

    Rui Zhou

    2009-12-01

    Full Text Available Cryptosporidium parvum is a protozoan parasite that infects the gastrointestinal epithelium and causes diarrheal disease worldwide. Innate epithelial immune responses are key mediators of the host's defense to C. parvum. MicroRNAs (miRNAs regulate gene expression at the posttranscriptional level and are involved in regulation of both innate and adaptive immune responses. Using an in vitro model of human cryptosporidiosis, we analyzed C. parvum-induced miRNA expression in biliary epithelial cells (i.e., cholangiocytes. Our results demonstrated differential alterations in the mature miRNA expression profile in cholangiocytes following C. parvum infection or lipopolysaccharide stimulation. Database analysis of C. parvum-upregulated miRNAs revealed potential NF-kappaB binding sites in the promoter elements of a subset of miRNA genes. We demonstrated that mir-125b-1, mir-21, mir-30b, and mir-23b-27b-24-1 cluster genes were transactivated through promoter binding of the NF-kappaB p65 subunit following C. parvum infection. In contrast, C. parvum transactivated mir-30c and mir-16 genes in cholangiocytes in a p65-independent manner. Importantly, functional inhibition of selected p65-dependent miRNAs in cholangiocytes increased C. parvum burden. Thus, we have identified a panel of miRNAs regulated through promoter binding of the NF-kappaB p65 subunit in human cholangiocytes in response to C. parvum infection, a process that may be relevant to the regulation of epithelial anti-microbial defense in general.

  8. Stress-Induced Hsp70 Gene Expression and Inactivation of Cryptosporidium parvum Oocysts by Chlorine-Based Oxidants▿

    Science.gov (United States)

    Bajszár, George; Dekonenko, Alexander

    2010-01-01

    Our research on the mechanisms of action of chlorine-based oxidants on Cryptosporidium parvum oocysts in water revealed a dual-phase effect: (i) response to oxidative stress, which was demonstrated by induced expression of the Hsp70 heat shock gene, and (ii) oocyst inactivation as a result of long-term exposure to oxidants. The relative biocidal effects of sodium hypochlorite (bleach) and electrolytically generated mixed oxidant solution (MOS) on C. parvum oocysts were compared at identical free chlorine concentrations. Oocyst inactivation was determined by quantitative reverse transcription-PCR (qRT-PCR) amplification of the heat-induced Hsp70 mRNA and compared with tissue culture infectivity. According to both assays, within the range between 25 and 250 mg/liter free chlorine and with 4 h contact time, MOS exhibits a higher efficacy in oocyst inactivation than hypochlorite. Other RNA-based viability assays, aimed at monitoring the levels of β-tubulin mRNA and 18S rRNA, showed relatively slow decay rates of these molecules following disinfection by chlorine-based oxidants, rendering these molecular diagnostic viability markers inappropriate for disinfection efficacy assessment. PMID:20118357

  9. Hydrologic and vegetative removal of Cryptosporidium parvum, Giardia lamblia, and Toxoplasma gondii Surrogate microspheres in coastal wetlands.

    Science.gov (United States)

    Hogan, Jennifer N; Daniels, Miles E; Watson, Fred G; Oates, Stori C; Miller, Melissa A; Conrad, Patricia A; Shapiro, Karen; Hardin, Dane; Dominik, Clare; Melli, Ann; Jessup, David A; Miller, Woutrina A

    2013-03-01

    Constructed wetland systems are used to reduce pollutants and pathogens in wastewater effluent, but comparatively little is known about pathogen transport through natural wetland habitats. Fecal protozoans, including Cryptosporidium parvum, Giardia lamblia, and Toxoplasma gondii, are waterborne pathogens of humans and animals, which are carried by surface waters from land-based sources into coastal waters. This study evaluated key factors of coastal wetlands for the reduction of protozoal parasites in surface waters using settling column and recirculating mesocosm tank experiments. Settling column experiments evaluated the effects of salinity, temperature, and water type ("pure" versus "environmental") on the vertical settling velocities of C. parvum, G. lamblia, and T. gondii surrogates, with salinity and water type found to significantly affect settling of the parasites. The mesocosm tank experiments evaluated the effects of salinity, flow rate, and vegetation parameters on parasite and surrogate counts, with increased salinity and the presence of vegetation found to be significant factors for removal of parasites in a unidirectional transport wetland system. Overall, this study highlights the importance of water type, salinity, and vegetation parameters for pathogen transport within wetland systems, with implications for wetland management, restoration efforts, and coastal water quality.

  10. Serum and colostrum antibody responses induced by jet-injection of sheep with DNA encoding a Cryptosporidium parvum antigen.

    Science.gov (United States)

    Jenkins, M; Kerr, D; Fayer, R; Wall, R

    1995-12-01

    In an effort to generate high titer colostrum for immunotherapy of cryptosporidiosis, a study was conducted to test the efficacy of immunizing sheep with recombinant plasmid DNA (pCMV-CP15/60) encoding epitopes of 15 and 60 kDa surface antigens of Cryptosporidium parvum sporozoites. The plasmid DNA was used to immunize preparturient ewes at three dose levels by jet-injection into either hind limb muscle (IM) or mammary tissue (IMAM). Regardless of route of injection, a dose-dependent anti-CP15/60 immunoglobulin response was observed in sera and colostrum from sheep immunized with pCMV-CP15/60 plasmid DNA. High titer antibody responses were observed in one of three animals per group receiving an IM injection of 100 or 1000 micrograms pCMV-CP15/60. IMAM immunization with 100 or 1000 micrograms pCMV-CP15/60 plasmid DNA elicited higher titer colostrum responses and more consistent serum responses compared to IM injections. A negligible serum and colostrum anti-CP15/60 response was observed in ewes injected IM with 10 micrograms pCMV-CP15/60 or 1000 micrograms control plasmid DNA. Immunoblotting of native C. parvum sporozoite/oocyst protein with hyperimmune serum and colostrum corroborated the increased titers against CP15/60 antigen. Serum and colostrum antibodies from pCMV-CP15/60-immunized sheep were eluted from native CP15 protein and bound a surface antigen of C. parvum sporozoites as indicated by indirect immunofluorescence staining.

  11. Transcriptome analysis reveals unique metabolic features in the Cryptosporidium parvum Oocysts associated with environmental survival and stresses

    Directory of Open Access Journals (Sweden)

    Zhang Haili

    2012-11-01

    Full Text Available Abstract Background Cryptosporidium parvum is a globally distributed zoonotic parasite and an important opportunistic pathogen in immunocompromised patients. Little is known on the metabolic dynamics of the parasite, and study is hampered by the lack of molecular and genetic tools. Here we report the development of the first Agilent microarray for C. parvum (CpArray15K that covers all predicted ORFs in the parasite genome. Global transcriptome analysis using CpArray15K coupled with real-time qRT-PCR uncovered a number of unique metabolic features in oocysts, the infectious and environmental stage of the parasite. Results Oocyst stage parasites were found to be highly active in protein synthesis, based on the high transcript levels of genes associated with ribosome biogenesis, transcription and translation. The proteasome and ubiquitin associated components were also highly active, implying that oocysts might employ protein degradation pathways to recycle amino acids in order to overcome the inability to synthesize amino acids de novo. Energy metabolism in oocysts was featured by the highest level of expression of lactate dehydrogenase (LDH gene. We also studied parasite responses to UV-irradiation, and observed complex and dynamic regulations of gene expression. Notable changes included increased transcript levels of genes involved in DNA repair and intracellular trafficking. Among the stress-related genes, TCP-1 family members and some thioredoxin-associated genes appear to play more important roles in the recovery of UV-induced damages in the oocysts. Our observations also suggest that UV irradiation of oocysts results in increased activities in cytoskeletal rearrangement and intracellular membrane trafficking. Conclusions CpArray15K is the first microarray chip developed for C. parvum, which provides the Cryptosporidium research community a needed tool to study the parasite transcriptome and functional genomics. CpArray15K has been

  12. Molecular characterization of isolates of waterborne Cryptosporidium spp. collected during an outbreak of gastroenteritis in South Burgundy, France.

    Science.gov (United States)

    Dalle, Frédéric; Roz, Pascale; Dautin, Guillaume; Di-Palma, Marc; Kohli, Evelyne; Sire-Bidault, C; Fleischmann, Marie George; Gallay, Anne; Carbonel, Sylvia; Bon, Fabienne; Tillier, Claude; Beaudeau, Pascal; Bonnin, Alain

    2003-06-01

    In September 2001, a waterborne outbreak of gastroenteritis occurred in eastern France. Of 31 fecal samples from symptomatic individuals, 19 tested positive for Cryptosporidium with two PCRs targeting the Hsp70 and the 18S rRNA genes of CRYPTOSPORIDIUM: Sequencing of the PCR fragments produced sequences identical to that of Cryptosporidium parvum genotype 1.

  13. Speeding up the solar water disinfection process (SODIS) against Cryptosporidium parvum by using 2.5l static solar reactors fitted with compound parabolic concentrators (CPCs).

    Science.gov (United States)

    Gómez-Couso, H; Fontán-Sainz, M; Fernández-Ibáñez, P; Ares-Mazás, E

    2012-12-01

    Water samples of 0, 5, and 100 nephelometric turbidity units (NTU) spiked with Cryptosporidium parvum oocysts were exposed to natural sunlight in 2.5l static borosilicate solar reactors fitted with two different compound parabolic concentrators (CPCs), CPC1 and CPC1.89, with concentration factors of the solar radiation of 1 and 1.89, respectively. The global oocyst viability was calculated by the evaluation of the inclusion/exclusion of the fluorogenic vital dye propidium iodide and the spontaneous excystation. Thus, the initial global oocyst viability of the C. parvum isolate used was 95.3 ± 1.6%. Using the solar reactors fitted with CPC1, the global viability of oocysts after 12h of exposure was zero in the most turbid water samples (100 NTU) and almost zero in the other water samples (0.3 ± 0.0% for 0 NTU and 0.5 ± 0.2% for 5 NTU). Employing the solar reactors fitted with CPC1.89, after 10h exposure, the global oocyst viability was zero in the non-turbid water samples (0 NTU), and it was almost zero in the 5 NTU water samples after 8h of exposure (0.5 ± 0.5%). In the most turbid water samples (100 NTU), the global viability was 1.9 ± 0.6% after 10 and 12h of exposure. In conclusion, the use of these 2.5l static solar reactors fitted with CPCs significantly improved the efficacy of the SODIS technique as these systems shorten the exposure times to solar radiation, and also minimize the negative effects of turbidity. This technology therefore represents a good alternative method for improving the microbiological quality of household drinking water in developing countries. Copyright © 2012 Elsevier B.V. All rights reserved.

  14. Wastewater treatment with Moringa oleifera seed extract: Impact on turbidity and sedimentation of Cryptosporidium parvum oocysts

    OpenAIRE

    Petersen, Heidi H.; Woolsey, Ian; Dalsgaard, Anders; Enemark, Heidi; Olsen, Annette

    2014-01-01

    The use of low quality water for irrigation in agriculture is common practise in many countries due to limited freshwater resources. Pathogens may contaminate vegetables when faeces polluted water is used for irrigation. A laboratory study was carried out to investigate the effect of a coagulant produced from seeds of the Moringa oleifera tree (MO) in reducing Cryptosporidium parvum oocysts and turbidity in wastewater. To a total of 5 x 12 glass jars containing 500 ml wastewater samples from ...

  15. Transport and survival of Cryptosporidium parvum oocysts in soil columns following applications of raw and separated liquid slurry

    DEFF Research Database (Denmark)

    Petersen, Heidi H.; Enemark, Heidi; Olsen, Annette;

    The widespread waterborne pathogen Cryptosporidium parvum is frequently transmitted to humans via contaminated drinking and recreational water. Nearly all drinking water in Denmark is groundwater, which can be contaminated with oocysts e.g. from application of contaminated manure to the field....... Oocysts transport to groundwater requires that the oocysts are transported through soil and bedrock to the water table. The purpose of this study was to determine the potential transport of the protozoan pathogen C. parvum through soil to land drains and, subsequently water sources in a laboratory setup...... using simulated rainfall and six 20 cm long replicate intact soil columns. Two types of contaminated slurry: raw slurry and the separated liquid fraction of the slurry were applied ten cm into the soil, which was subsequently irrigated once a week over a four-week period. C. parvum oocysts were detected...

  16. Does the use of tubular digesters to treat livestock waste lower the risk of infection from Cryptosporidium parvum and Giardia lamblia?

    Science.gov (United States)

    Kinyua, Maureen N; Wald, Ileana; Camacho-Céspedes, Fabricio; Izurieta, Ricardo; Haas, Charles N; Ergas, Sarina J

    2016-10-01

    Worldwide, high incidences of cryptosporidiosis and giardiasis are attributed to livestock waste. Quantitative microbial risk assessment can be used to estimate the risk of livestock related infections from Cryptosporidium parvum and Giardia lamblia. The objective of this paper was to assess the occupational and public health risks associated with management of raw and anaerobically digested livestock waste in two rural communities in Costa Rica based on fomite, soil and crop contamination and livestock waste management exposure pathways. Risks related to cattle waste were greater than swine waste due to cattle shedding more (oo)cysts. Cryptosporidium parvum also posed a greater risk than Giardia lamblia in all exposure pathways due to livestock shedding high loads of Cryptosporidium parvum oocysts and oocysts' lower inactivation rates during anaerobic digestion compared with Giardia lamblia cysts. The risk of infection from exposure to contaminated soil and crops was significantly lower for a community using tubular anaerobic digesters to treat livestock waste compared to a community where the untreated waste was applied to soil. The results indicate that treatment of livestock waste in small-scale tubular anaerobic digesters has the potential to significantly decrease the risk of infection below the World Health Organization's acceptable individual annual risk of infection (10(-4)).

  17. Evaluation of the BD MAX Enteric Parasite Panel for the detection of Cryptosporidium parvum/hominis, Giardia duodenalis and Entamoeba histolytica.

    Science.gov (United States)

    Perry, Michael D; Corden, Sally A; Lewis White, P

    2017-08-01

    Conventional laboratory detection methods for gastrointestinal parasites are time consuming, require considerable technical expertise and may suffer from poor analytical sensitivity. This study sought to evaluate the automated BD MAX Enteric Parasite Panel (EPP) for the detection of Cryptosporidium parvum/hominis, Entamoeba histolytica and Giardia duodenalis.Methodolgy. A total of 104 known positive samples (43 Cryptosporidium parvum/hominis and 61 G. duodenalis), 15 simulated samples (E. histolytica and other Entamoeba species) and 745 patient stool samples, submitted for enteric pathogen culture and microscopy, were inoculated into BD MAX EPP sample buffer tubes (SBTs). All specimens were blinded and tested within 7 days of SBT inoculation using the BD MAX EPP assay with results compared to those generated by microscopy.Results/Key findings. Combining the results from the known positive samples and anonymously tested patient samples, the sensitivity of the BD MAX EPP assay was 100 % for both Cryptosporidium spp. and G. duodenalis. Specificities of 99.7 and 98.9 % were calculated for the detection of Cryptosporidium spp. and G. duodenalis respectively. Insufficient clinical specimen data was available to determine the performance of the assay for E. histolytica detection. The findings of this study indicate that the BD MAX EPP is suitable for the detection of Cryptosporidium parvum/hominis and G. duodenalis from clinical specimens with reduced hands-on time and complexity compared to microscopy. Results for the detection of E. histolytica were promising although further work is required to evaluate the assay for the detection of this pathogen.

  18. Removals of cryptosporidium parvum oocysts and cryptosporidium-sized polystyrene microspheres from swimming pool water by diatomaceous earth filtration and perlite-sand filtration.

    Science.gov (United States)

    Lu, Ping; Amburgey, James E; Hill, Vincent R; Murphy, Jennifer L; Schneeberger, Chandra L; Arrowood, Michael J; Yuan, Tao

    2017-06-01

    Removal of Cryptosporidium-sized microspheres and Cryptosporidium parvum oocysts from swimming pools was investigated using diatomaceous earth (DE) precoat filtration and perlite-sand filtration. In pilot-scale experiments, microsphere removals of up to 2 log were obtained with 0.7 kg·DE/m(2) at a filtration rate of 5 m/h. A slightly higher microsphere removal (2.3 log) was obtained for these DE-precoated filters when the filtration rate was 3.6 m/h. Additionally, pilot-scale perlite-sand filters achieved greater than 2 log removal when at least 0.37 kg/m(2) of perlite was used compared to 0.1-0.4 log removal without perlite both at a surface loading rate of 37 m/h. Full-scale testing achieved 2.7 log of microspheres and oocysts removal when 0.7 kg·DE/m(2) was used at 3.6 m/h. Removals were significantly decreased by a 15-minute interruption of the flow (without any mechanical agitation) to the DE filter in pilot-scale studies, which was not observed in full-scale filters. Microsphere removals were 2.7 log by perlite-sand filtration in a full-scale swimming pool filter operated at 34 m/h with 0.5 kg/m(2) of perlite. The results demonstrate that either a DE precoat filter or a perlite-sand filter can improve the efficiency of removal of microspheres and oocysts from swimming pools over a standard sand filter under the conditions studied.

  19. Prevalência do Cryptosporidium parvum em crianças abaixo de 5 anos, residentes na zona urbana de Campo Grande, MS, Brasil, 1996

    Directory of Open Access Journals (Sweden)

    Oshiro Elisa Teruya

    2000-01-01

    Full Text Available O presente estudo visou estabelecer a prevalência de Cryptosporidium parvum em crianças abaixo de 5 anos, residentes na zona urbana de Campo Grande, MS, 1996/97, através de exames coprológicos e avaliar epidemiologicamente os casos diagnosticados. Tratou-se de um estudo transversal com inquérito domiciliar, onde foram examinadas 1051 amostras fecais, processadas segundo a técnica de Blagg, utilizando-se a coloração de Ziehl-Neelsen modificada para a pesquisa de oocistos de C. parvum. Concluiu-se que: a prevalência de C. parvum (1,1% observada não foi estatisticamente significativa; foi relatado diarréia em 58,3% das crianças com diagnóstico positivo, supondo-se associação entre diarréia e a presença do parasita; o C. parvum foi mais freqüente em crianças com idade de 25 a 36 meses (50%, porém sem diferença estatisticamente significativa; o sexo não teve papel diferencial em relação ao parasitismo por C. parvum; entre as 12 crianças com criptosporidiose, 83,3% tiveram contato com animais domésticos (cão e ou gato.

  20. Fluorescent in situ hybridization as a tool to retrospectively identify Cryptosporidium parvum and Giardia lamblia in samples from terrestrial mammalian wildlife.

    Science.gov (United States)

    Bednarska, Malgorzata; Bajer, Anna; Sinski, Edward; Girouard, Autumn S; Tamang, Leena; Graczyk, Thaddeus K

    2007-02-01

    Fecal samples of five terrestrial mammalian wildlife species stored at 4 degrees C or at -20 degrees C for up to 36 months have been tested for human zoonotic enteric parasites (i.e., Cryptosporidium parvum and Giardia lamblia) using combined fluorescent in situ hybridization (FISH) and direct fluorescent antibody techniques. The prevalence of C. parvum and G. lamblia varied from 20 to 63% (mean, 45.8%) and from 13 to 100% (mean, 53.2%), respectively. The prevalence of C. parvum and G. lamblia infections was higher in small rodents (mean, 68.5%) than in other wildlife (mean, 21%). Overall, 31.1% of animals were coinfected, and coinfections were more prevalent in small rodents (mean, 52%) than in other wildlife species (mean, 13.2%). The present study has shown that the FISH assay can be retrospectively applied to fecal samples for the identification of C. parvum oocysts, but is less suitable for the identification of G. lamblia cysts in such samples. Terrestrial mammalian wildlife, particularly small rodents, can contribute to watershed contamination with C. parvum oocysts and G. lamblia cysts. To control contamination, the management of pristine watersheds used for drinking water purposes should incorporate control measures for terrestrial wildlife, especially field rodents residing within such watersheds.

  1. Pomegranate (Punica granatum) peel is effective in a murine model of experimental Cryptosporidium parvum ultrastructural studies of the ileum.

    Science.gov (United States)

    Al-Mathal, Ebtisam M; Alsalem, Afaf A

    2013-08-01

    The current treatments for cryptosporidiosis are ineffective, and there is an urgent need to search for more effective and safer alternatives. One such alternative may be treatments derived from natural resources. The pomegranate peel has been used effectively in traditional medicine to cure diarrhea and dysentery. The purpose of this study was to examine the effectiveness of a Punica granatum (pomegranate) peel suspension as a treatment for Cryptosporidium parvum infection. In this study, the effects of this treatment on the ultrastructure of both the intestinal epithelial layer of infected nursling mice and the parasite were observed with a transmission electron microscope. The histological study focused on the examination of the microvilli, columnar epithelium, goblet cells, lamina propria, and crypts of Lieberkuhn. Examination of the ileums of infected mice that received the pomegranate peel suspension demonstrated that the general structure of the ileal tissue of these mice was similar to that of the control group. In the infected mice treated with the suspension, but not the infected/untreated mice, there was an improvement in all ultrastructure aspects at 28days post-inoculation. The study of the ultrastructure of the parasite (C. parvum) in mice treated with the suspension showed that there was decomposition in the parasite to the extent that in some cases we were unable to identify the stage of the parasite due to the severe degeneration. Significant decomposition of the nutrition organ was also observed. Additionally, microgamonte and macrogamonte were not observed in the suspension-treated group, explaining the disappearance of the sexual phases of the parasite in the lumens of this group. In all, this examination demonstrated the restoration of the normal structures of villi and the disappearance of acute symptoms in the suspension-treated mice and showed that the suspension directly affected the parasite at various stages of its development and led to

  2. Construction of genomic libraries of Cryptosporidium parvum and identification of antigen-encoding genes.

    Science.gov (United States)

    Dykstra, C C; Blagburn, B L; Tidwell, R R

    1991-01-01

    Genomic libraries have been constructed from bovine C. parvum DNA in the lambda ZAP and lambda DASH vectors. Based on an estimated genome size of 2 x 10(4) kilobases (kb), each recombinant library contains greater than 10 genomic equivalents. The average recombinant size for the lambda ZAP library is 2.1 kb and for the lambda DASH library is 14 kb. We have identified genes to major antigens recognized by hyperimmune bovine antiserum. These recombinants are currently being purified and characterized. Limited DNA sequence analysis of random C. parvum clones confirms suggestions that the genome is quite AT-rich. The DNA sequence of random lambda ZAP fusion proteins has identified a potential ATPase, a structural protein and a DNA-binding protein.

  3. Evaluating the transport of bacillus subtilis spores as a potential surrogate for Cryptosporidium parvum Oocysts

    Science.gov (United States)

    The USEPA has recommended the use of aerobic spores as an indicator for Cryptosporidium oocysts when determining groundwater under the direct influence of surface water. Surface properties, interaction energies, transport, retention, and release behavior of B. subtilis spores were measured over a r...

  4. Effects of Enterococcus faecalis CECT 7121 on Cryptosporidium parvum infection in mice

    Science.gov (United States)

    Cryptosporidium is an opportunistic protozoan parasite of humans and animals worldwide, causes diarrheal disease that is typically self-limiting in immunocompetent hosts but often life-threatening to immunocompromised individuals. However, there is a lack of completely efficient therapy available. P...

  5. Modellering van de verwijdering van Cryptosporidium parvum,Giardia lamblia en enterovirussen in spaarbekkens

    NARCIS (Netherlands)

    Ratsak CH; MGB

    1996-01-01

    De protozoa Cryptosporidium en Giardia hebben de laatste decennia in de Verenigde Staten en in Groot-Brittannie een aanzienlijk aantal grote en kleine epidemieen van maag-darminfecties via drinkwater veroorzaakt. Wiskundige modellen kunnen worden gebruikt om de verwijdering van pathogene micro-o

  6. Modellering van de verwijdering van Cryptosporidium parvum,Giardia lamblia en enterovirussen in spaarbekkens

    NARCIS (Netherlands)

    Ratsak CH; MGB

    1996-01-01

    De protozoa Cryptosporidium en Giardia hebben de laatste decennia in de Verenigde Staten en in Groot-Brittannie een aanzienlijk aantal grote en kleine epidemieen van maag-darminfecties via drinkwater veroorzaakt. Wiskundige modellen kunnen worden gebruikt om de verwijdering van pathogene

  7. Molecular and phylogenetic characterization of Cryptosporidium and Giardia from pigs and cattle in Denmark

    DEFF Research Database (Denmark)

    Langkjær, Rikke Breinhold; Vigre, Håkan; Enemark, Heidi L.

    2007-01-01

    . suis, pig genotype II, C. parvum (cattle genotype), C. bovis, Cryptosporidium deer-like genotype and a novel C. suis-like genotype. For both cattle and pigs, a host age-related change in distribution of species/genotypes was observed. The zoonotic C. parvum (cattle genotype) was most prevalent in young...... purely by isolates of the livestock group, Assemblage E....

  8. Influence of organic carbon loading, sediment associated metal oxide content and sediment grain size distributions upon Cryptosporidium parvum removal during riverbank filtration operations, Sonoma County, CA

    Science.gov (United States)

    Metge, D.W.; Harvey, R.W.; Aiken, G.R.; Anders, R.; Lincoln, G.; Jasperse, J.

    2010-01-01

    This study assessed the efficacy for removing Cryptosporidium parvum oocysts of poorly sorted, Fe- and Al-rich, subsurface sediments collected from 0.9 to 4.9 and 1.7-13.9??m below land surface at an operating riverbank filtration (RBF) site (Russian River, Sonoma County, CA). Both formaldehyde-killed oocysts and oocyst-sized (3????m) microspheres were employed in sediment-packed flow-through and static columns. The degree of surface coverage of metal oxides on sediment grain surfaces correlated strongly with the degrees of oocyst and microsphere removals. In contrast, average grain size (D50) was not a good indicator of either microsphere or oocyst removal, suggesting that the primary mechanism of immobilization within these sediments is sorptive filtration rather than physical straining. A low specific UV absorbance (SUVA) for organic matter isolated from the Russian River, suggested that the modest concentration of the SUVA component (0.8??mg??L-1) of the 2.2??mg??L-1 dissolved organic carbon (DOC) is relatively unreactive. Nevertheless, an amendment of 2.2??mg??L-1 of isolated river DOC to column sediments resulted in up to a 35.7% decrease in sorption of oocysts and (or) oocyst-sized microspheres. Amendments (3.2????M) of the anionic surfactant, sodium dodecyl benzene sulfonate (SDBS) also caused substantive decreases (up to 31.9 times) in colloid filtration. Although the grain-surface metal oxides were found to have a high colloid-removal capacity, our study suggested that any major changes within the watershed that would result in long-term alterations in either the quantity and (or) the character of the river's DOC could alter the effectiveness of pathogen removal during RBF operations.

  9. Cryptosporidium parvum infection in SCID mice infected with only one oocyst: qPCR assessment of parasite replication in tissues and development of digestive cancer.

    Directory of Open Access Journals (Sweden)

    Sadia Benamrouz

    Full Text Available Dexamethasone (Dex treated Severe Combined Immunodeficiency (SCID mice were previously described as developing digestive adenocarcinoma after massive infection with Cryptosporidium parvum as soon as 45 days post-infection (P.I.. We aimed to determine the minimum number of oocysts capable of inducing infection and thereby gastrointestinal tumors in this model. Mice were challenged with calibrated oocyst suspensions containing intended doses of: 1, 10, 100 or 10(5 oocysts of C. parvum Iowa strain. All administered doses were infective for animals but increasing the oocyst challenge lead to an increase in mice infectivity (P = 0.01. Oocyst shedding was detected at 7 days P.I. after inoculation with more than 10 oocysts, and after 15 days in mice challenged with one oocyst. In groups challenged with lower inocula, parasite growth phase was significantly higher (P = 0.005 compared to mice inoculated with higher doses. After 45 days P.I. all groups of mice had a mean of oocyst shedding superior to 10,000 oocyst/g of feces. The most impressive observation of this study was the demonstration that C. parvum-induced digestive adenocarcinoma could be caused by infection with low doses of Cryptosporidium, even with only one oocyst: in mice inoculated with low doses, neoplastic lesions were detected as early as 45 days P.I. both in the stomach and ileo-caecal region, and these lesions could evolve in an invasive adenocarcinoma. These findings show a great amplification effect of parasites in mouse tissues after challenge with low doses as confirmed by quantitative PCR. The ability of C. parvum to infect mice with one oocyst and to develop digestive adenocarcinoma suggests that other mammalian species including humans could be also susceptible to this process, especially when they are severely immunocompromised.

  10. Removal of Cryptosporidium parvum oocysts in low quality water using Moringa oleifera seed extract as coagulant

    DEFF Research Database (Denmark)

    Petersen, Heidi Huus; Petersen, T. B.; Enemark, Heidi

    2016-01-01

    The use of different types of low quality water for irrigation in agriculture is common practice in many countries due to limited freshwater resources. Pathogens may contaminate fruit and vegetables when feces contaminated water is used for irrigation or postharvest processing. A laboratory study...... demonstrate that MO seed extract may be used by farmers for treatment of different types of surface water prior to irrigation use. Yet, adding MO seed extract to the low quality water did not successfully remove all oocyst. However, treatment of wastewater with MO seed extract significantly improved the water...... quality with regard to number of oocysts present and turbidity of the water. Further experiments with addition of higher concentrations of MO are needed to establish whether MO seed extract can be used to obtain safe irrigation water free of C. parvum oocysts and other protozoan parasites....

  11. [A new trichromic safranin stain for the detection of Cryptosporidium parvum, Cyclospora cayetanensis, species of Microsporidia and Isospora belli in fecal material].

    Science.gov (United States)

    Ponce de León, P; Flaherty, P; Zdero, M

    1999-01-01

    Cryptosporidium parvum, Isospora belli, Cyclospora cayetanensis and Microsporidia are frequent pathogens in the immunodeficient host, which may cause multiple infections. The above mentioned parasites are found in feces by the application of different specific tintorial techniques. The objective of this work was the development of a stain for the simultaneous detection of these parasites, reducing costs as well as the time taken to make the diagnosis. The safranin-trichrome stain is simple, chip and its results are similar to those of specific tints. All microorganisms are easy to detect and besides being perfectly distinguishable from fungi and faecal elements.

  12. Role of collector alternating charged patches on transport of Cryptosporidium parvum oocyst in a patchwise charged heterogeneous micromodel

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yuanyuan; Zhang, Changyong; Hu, Dehong; Kuhlenschmidt, Mark S.; Kuhlenschmidt, Theresa B.; Mylon, Steven E.; Kong, Rong; Bhargava, Rohit; Nguyen, Thanh H.

    2013-02-04

    The role of collector surface charge heterogeneity on transport of Cryptosporidium parvum oocyst and carboxylate microsphere in 2-dimensional micromodels was studied. The cylindrical silica collectors within the micromodels were coated with 0, 10, 20, 50 and 100% Fe2O3 patches. The experimental values of average single collector removal efficiencies (η) of the Fe2O3 patches and on the entire collectors were determined. In the presence of significant (>3500 kT) Derjaguin–Landau–Verwey–Overbeek (DLVO) energy barrier between the microspheres and the silica collectors at pH 5.8 and 8.1, the values of η determined for Fe2O3 patches were significantly less (p < 0.05, t-test) than that obtained for collectors coated entirely with Fe2O3. However, η on Fe2O3 patches for microspheres at pH 4.4 and for oocysts at pH 5.8 and 8.1, where the DLVO energy barrier was relatively small (ca. 200-360 kT), were significantly greater (p < 0.05, t-test) than that on the collectors coated entirely with Fe2O3. The dependence of η determined for Fe2O3 patches on the DLVO energy barrier indicated the importance of periodic favorable and unfavorable electrostatic interactions between colloids and collectors with alternating Fe2O3 and silica patches. Differences between experimentally determined η and that predicted by a patchwise geochemical heterogeneous model was observed, but can be explained by the model’s lack of consideration for the spatial distribution of charge heterogeneity on the collector surface and colloid migration on patchwise heterogeneous collectors.

  13. Development of Cryptosporidium parvum-Induced Gastrointestinal Neoplasia in Severe Combined Immunodeficiency (SCID) Mice: Severity of Lesions Is Correlated with Infection Intensity

    Science.gov (United States)

    Certad, Gabriela; Creusy, Colette; Ngouanesavanh, Tramy; Guyot, Karine; Gantois, Nausicaa; Mouray, Anthony; Chassat, Thierry; Flament, Nicolas; Fleurisse, Laurence; Pinon, Anthony; Delhaes, Laurence; Dei-Cas, Eduardo

    2010-01-01

    We reported previously that Cryptosporidium parvum was able to induce intestinal tumors in severe combined immunodeficiency (SCID) mice treated with corticoids. To further characterize this Cryptosporidium-induced cell transformation, SCID mice treated with dexamethasone were challenged with C. parvum oocysts, and euthanatized sequentially after infection for histologic examination. Ki-67 was used as a marker of cellular proliferation. Our previous results were confirmed, and it was also found that mice receiving higher inocula (106–107) experienced more severe neoplastic development. Additionally, neoplastic changes were observed not only in the caecum but also in the stomach and duodenum of some animals. Interestingly, SCID mice (6/6) inoculated with 105–107 oocysts showed high grade intraepithelial neoplasia or adenomas with high grade dysplasia in the caecum after Day 46 post-infection (PI). Immunohistochemistry for Ki-67 staining indicated the neoplastic process associated to cryptosporidiosis, and evidenced the first immunohistochemical alterations at early stages of the process, even at 3 weeks PI. PMID:20134002

  14. Comparison of different solar reactors for household disinfection of drinking water in developing countries: evaluation of their efficacy in relation to the waterborne enteropathogen Cryptosporidium parvum.

    Science.gov (United States)

    Gómez-Couso, H; Fontán-Sainz, M; Navntoft, C; Fernández-Ibáñez, P; Ares-Mazás, E

    2012-11-01

    Solar water disinfection (SODIS) is a type of treatment that can significantly improve the microbiological quality of drinking water at household level and therefore prevent waterborne diseases in developing countries. Cryptosporidium parvum is an obligate protozoan parasite responsible for the diarrhoeal disease cryptosporidiosis in humans and animals. Recently, this parasite has been selected by the WHO as a reference pathogen for protozoan parasites in the evaluation of household water treatment options. In this study, the field efficacy of different static solar reactors [1.5 l transparent plastic polyethylene terephthalate (PET) bottles as well as 2.5 l borosilicate glass and 25 l methacrylate reactors fitted with compound parabolic concentrators (CPC)] for solar disinfection of turbid waters experimentally contaminated with C. parvum oocysts was compared. Potential oocyst viability was determined by inclusion/exclusion of the fluorogenic vital dye propidium iodide. The results demonstrate that static solar reactors fitted with CPCs are an excellent alternative to the conventional SODIS method with PET bottles. These reactors improved the efficacy of the SODIS method by enabling larger volumes of water to be treated and, in some cases, the C. parvum oocysts were rendered totally unviable, minimising the negative effects of turbidity.

  15. GIS-based analysis of the fate of waste-related pathogens Cryptosporidium parvum, Giardia lamblia and Escherichia coli in a tropical canal network.

    Science.gov (United States)

    Diallo, Mamadou B C; Anceno, Alfredo J; Tawatsupa, Benjawan; Tripathi, Nitin K; Wangsuphachart, Voranuch; Shipin, Oleg V

    2009-03-01

    Urban canals play a major socio-economic role in many tropical countries and, particularly, Thailand. One of the overlooked functions that they perform is a significant attenuation of waste-related pathogens posing considerable health risk, as well as pollution attenuation in general. The study dealt with a comparison of three canals receiving: (i) municipal, (ii) mainly industrial and (iii) mainly agricultural wastewater, listed in order of progressively decreasing organic loading. The occurrence and fate of waterborne Cryptosporidium parvum, Giardia lamblia and Escherichia coli were monitored in the canals by both real-time PCR and conventionally for 12 months. The pathogens are etiological agents of an estimated 38% and 47% of diarrhea cases worldwide and in Thailand, respectively. The geographic information system (GIS) was used to evaluate and map point and, particularly, non-point pollution sources which allowed differentiating the canal sections in terms of predominant pathogen sources. The flowthrough canals, which can be viewed as waste stabilization ponds, were found to be efficiently removing the pathogens at the following generalized specific rates: 0.3 (C. parvum), 1.2 (G. lamblia), 1.8 (E. coli) log10/km.d in the dry season. The rates decreased in the rainy season for E. coli and G. lamblia, but increased for C. parvum which indicated different removal mechanisms. Data suggest that E. coli and G. lamblia were mainly removed through sedimentation and sunlight (UV) irradiation, while the likely mechanism for C. parvum was predation. Overall, the specific pathogen removal rates positively correlated with the canal organic loading rates in the rainy season. As an important result, an estimate of the municipal pollution mitigation by over 2280 km canals in the Greater Bangkok suggests that concomitant to the pathogens at least 36-95 tons of BOD5 is being removed daily, thereby saving the receiving Chao Phraya River and Bight of Bangkok, by far exceeding

  16. Molecular identification of Giardia and Cryptosporidium from dogs and cats

    Directory of Open Access Journals (Sweden)

    Sotiriadou Isaia

    2013-01-01

    Full Text Available The aim of the present study was to diagnose the presence of Giardia cysts and Cryptosporidium oocysts in household animals using nested polymerase chain reaction (PCR and sequence analysis. One hundred faecal samples obtained from 81 dogs and 19 cats were investigated. The Cryptosporidium genotypes were determined by sequencing a fragment of the small subunit (SSU rRNA gene, while the Giardia Assemblages were determined through analysis of the glutamate dehydrogenase (GDH locus. Isolates from five dogs and two cats were positive by PCR for the presence of Giardia, and their sequences matched the zoonotic Assemblage A of Giardia. Cryptosporidium spp. isolated from one dog and one cat were both found to be C. parvum. One dog isolate harboured a mixed infection of C. parvum and Giardia Assemblage A. These findings support the growing evidence that household animals are potential reservoirs of the zoonotic pathogens Giardia spp. and Cryptosporidium spp. for infections in humans.

  17. Molecular identification of Giardia and Cryptosporidium from dogs and cats.

    Science.gov (United States)

    Sotiriadou, Isaia; Pantchev, Nikola; Gassmann, Doreen; Karanis, Panagiotis

    2013-01-01

    The aim of the present study was to diagnose the presence of Giardia cysts and Cryptosporidium oocysts in household animals using nested polymerase chain reaction (PCR) and sequence analysis. One hundred faecal samples obtained from 81 dogs and 19 cats were investigated. The Cryptosporidium genotypes were determined by sequencing a fragment of the small subunit (SSU) rRNA gene, while the Giardia Assemblages were determined through analysis of the glutamate dehydrogenase (GDH) locus. Isolates from five dogs and two cats were positive by PCR for the presence of Giardia, and their sequences matched the zoonotic Assemblage A of Giardia. Cryptosporidium spp. isolated from one dog and one cat were both found to be C. parvum. One dog isolate harboured a mixed infection of C. parvum and Giardia Assemblage A. These findings support the growing evidence that household animals are potential reservoirs of the zoonotic pathogens Giardia spp. and Cryptosporidium spp. for infections in humans.

  18. Use of a sentinel system for field measurements of Cryptosporidium parvum oocyst inactivation in soil and animal waste.

    Science.gov (United States)

    Jenkins, M B; Walker, M J; Bowman, D D; Anthony, L C; Ghiorse, W C

    1999-05-01

    A small-volume sentinel chamber was developed to assess the effects of environmental stresses on survival of sucrose-Percoll-purified Cryptosporidium parvum oocysts in soil and animal wastes. Chambers were tested for their ability to equilibrate with external chemical and moisture conditions. Sentinel oocysts were then exposed to stresses of the external environment that affected their viability (potential infectivity), as indicated by results of a dye permeability assay. Preliminary laboratory experiments indicated that temperatures between 35 and 50 degrees C and decreases in soil water potential (-0.003 to -3.20 MPa) increased oocyst inactivation rates. The effects of two common animal waste management practices on oocyst survival were investigated on three dairy farms in Delaware County, N.Y., within the New York City watershed: (i) piling wastes from dairy youngstock (including neonatal calves) and (ii) spreading wastes as a soil amendment on an agricultural field. Sentinel containers filled with air-dried and sieved (2-mm mesh) youngstock waste or field soil were wetted and inoculated with 2 million oocysts in an aqueous suspension and then placed in waste piles on two different farms and in soil within a cropped field on one farm. Controls consisted of purified oocysts in either phosphate-buffered saline or distilled water contained in sealed microcentrifuge tubes. Two microdata loggers recorded the ambient temperature at each field site. Sentinel experiments were conducted during the fall and winter (1996 to 1997) and winter (1998). Sentinel containers and controls were removed at 2- to 4-week intervals, and oocysts were extracted and tested by the dye permeability assay. The proportions of potentially infective oocysts exposed to the soil and waste pile material decreased more rapidly than their counterpart controls exposed to buffer or water, indicating that factors other than temperature affected oocyst inactivation in the waste piles and soil. The

  19. Morphological structure and staining characteristics of Cryptosporidium parvum oocysts%微小隐孢子虫卵囊的形态结构与染色观察

    Institute of Scientific and Technical Information of China (English)

    陈甫

    2011-01-01

    Objective To observe the morphological structure and staining characteristics of Cryptosporidium parvum oocysts. Methods The morphological structure of C. Parvum oocysts was observed using a high-resolution imaging a-nalysis system and depicted using Adobe Photoshop CS4. Characteristics of C. Parvum oocysts stained by modified acid-fast staining, Giemsa staining, Alcian blue staining, and HE staining were observed. Results C. Parvum oocysts with thin oocyst walls appeared round and elliptical in shape and contain one to four crescent-shaped sporozoites and a round residual body. Individual oocysts were empty. C. Parvum oocysts appeared rose pink with modified acid-fast staining, blue with Giemsa staining, green with Alcian blue staining, and red with HE staining. Conclusion The morphological structure of C. Parvum oocysts varied, and oocysts were most effectively stained with modified acid-fast staining.%目的 研究隐孢子虫卵囊的形态与染色特点.方法 应用图像分析系统,观察隐孢子虫卵囊的形态与内部结构,并应用Adobe Photoshop CS4软件绘制各种卵囊形态的模拟图.卵囊经改良抗酸染色法、姬姆萨染色法、阿利新蓝染色法和苏木精-伊红染色法染色后观察其染色特点.结果 隐孢子虫卵囊呈圆形或椭圆形,外层有一薄的卵囊壁,内部含有0~4个月牙形子孢子和圆点状残留体,个别卵囊呈空泡状.隐孢子虫卵囊经改良抗酸染色、姬姆萨染色、阿利新蓝染色和HE染色后,分别呈现玫瑰红色、蓝色、绿色和红色.结论 隐孢子虫卵囊形态和内部结构多样,改良抗酸染色法染色效果较好.

  20. Occurrence of Giardia and Cryptosporidium in pigs on Prince Edward Island, Canada.

    Science.gov (United States)

    Budu-Amoako, Ebo; Greenwood, Spencer J; Dixon, Brent R; Barkema, Herman W; Hurnik, Daniel; Estey, Chelsie; McClure, J T

    2012-02-28

    In a cross-sectional study of 633 pigs from 21 herds on Prince Edward Island, Canada (PEI), the prevalence of infection with Cryptosporidium and Giardia, and the genotypes and species of isolates were determined in order to establish the zoonotic potential of pigs in this region. As determined by direct immunofluorescence microscopy (DFA), 18 herds (86%) and 163 animals (26%; 95% CI: 22-29%) tested positive for Cryptosporidium, while just 3 herds (14%) and 6 animals (1%; 95% CI: 0.4-2%) tested positive for Giardia. Cryptosporidium spp. isolates were detected in 39% (95% CI: 34-44%) of weanlings (1-3 months of age) and 9% (95% CI: 6-13) of sows (>8months of age). Molecular characterization using the 18S rDNA and HSP70 gene fragments revealed the presence of Cryptosporidium sp. pig genotype II, C. suis, C. parvum, and Cryptosporidium sp. mouse genotype. Among the 113 isolates of Cryptosporidium spp. successfully genotyped, pig genotype II (61%) predominated, with C. suis (36%) being the next most prominant isolate. C. parvum (2%; two isolates) and Cryptosporidium sp. mouse genotype (0.9%; one isolate) were only occasionally isolated. The only two Cryptosporidium-positive genotyped isolates from sows included one each of C. suis and Cryptosporidium sp. pig genotype II. All but one of the six Giardia positive isolates were detected in weanling pigs. None of the Giardia-positive isolates was amenable to PCR. This study demonstrates that Cryptosporidium spp. are highly prevalent in pigs on PEI, Canada, are found mostly in weanlings (1-3 months of age). Furthermore, the pigs are primarily infected by the host-specific genotypes and species, Cryptosporidium sp. pig genotype II and C. suis, whereas the zoonotic C. parvum is rare. Giardia duodenalis is only occasionally found in pigs. These findings suggest that domestic pigs on PEI, Canada, likely do not pose a significant health risk to humans from these parasites.

  1. Evaluation of the Solar Water Disinfection Process (SODIS) Against Cryptosporidium parvum Using a 25-L Static Solar Reactor Fitted with a Compound Parabolic Collector (CPC)

    Science.gov (United States)

    Fontán-Sainz, María; Gómez-Couso, Hipólito; Fernández-Ibáñez, Pilar; Ares-Mazás, Elvira

    2012-01-01

    Water samples of 0, 5, and 30 nephelometric turbidity units (NTU) spiked with Cryptosporidium parvum oocysts were exposed to natural sunlight using a 25-L static solar reactor fitted with a compound parabolic collector (CPC). The global oocyst viability was calculated by the evaluation of the inclusion/exclusion of the fluorogenic vital dye propidium iodide and the spontaneous excystation. After an exposure time of 8 hours, the global oocyst viabilities were 21.8 ± 3.1%, 31.3 ± 12.9%, and 45.0 ± 10.0% for turbidity levels of 0, 5, and 30 NTU, respectively, and these values were significantly lower (P 10 times). PMID:22302852

  2. Binding and activation of human and mouse complement by Cryptosporidium parvum (Apicomplexa) and susceptibility of C1q- and MBL-deficient mice to infection.

    Science.gov (United States)

    Petry, Franz; Jakobi, Vera; Wagner, Swen; Tessema, Tesfaye Sisay; Thiel, Steffen; Loos, Michael

    2008-07-01

    Cryptosporidium parvum is a protozoan parasite (Apicomplexa) that causes gastrointestinal disease in animals and humans. Whereas immunocompetent hosts can limit the infection within 1 or 2 weeks, immunocompromised individuals develop a chronic, life-threatening disease. The importance of the adaptive cellular immune response, with CD4+ T-lymphocytes being the major players, has been clearly demonstrated. Several non-adaptive immune mechanisms have been suggested to contribute to the host defence, such as interferon-gamma (IFN-gamma) from NK cells, certain chemokines, beta-defensins and pro-inflammatory cytokines, but the influence of the complement systems has been less well studied. We analysed the in vitro binding and activation of the human and mouse complement systems and tested the susceptibility to infection in complement-deficient mouse strains. We found that C. parvum can activate both the classical and lectin pathways, leading to the deposition of C3b on the parasite. Using real-time PCR, parasite development could be demonstrated in adult mice lacking mannan-binding lectin (MBL-A/C-/-) but not in mice lacking complement factor C1q (C1qA-/-) or in wild type C57BL/6 mice. The contribution of the complement system and the lectin pathway in particular to the host defence against cryptosporidiosis may become apparent in situations of immunodeficiency such as HIV infections or in early childhood.

  3. Disinfection and toxicological assessments of pulsed UV and pulsed-plasma gas-discharge treated-water containing the waterborne protozoan enteroparasite Cryptosporidium parvum.

    Science.gov (United States)

    Hayes, Jennifer; Kirf, Dominik; Garvey, Mary; Rowan, Neil

    2013-09-01

    We report for the first time on the comparative use of pulsed-plasma gas-discharge (PPGD) and pulsed UV light (PUV) for the novel destruction of the waterborne enteroparasite Cryptosporidium parvum. It also describes the first cyto-, geno- and ecotoxicological assays undertaken to assess the safety of water decontaminated using PPGD and PUV. During PPGD treatments, the application of high voltage pulses (16 kV, 10 pps) to gas-injected water (N2 or O2, flow rate 2.5L/min) resulted in the formation of a plasma that generated free radicals, ultraviolet light, acoustic shock waves and electric fields that killed ca. 4 log C. parvum oocysts in 32 min exposure. Findings showed that PPGD-treated water produced significant cytotoxic properties (as determined by MTT and neutral red assays), genotoxic properties (as determined by comet and Ames assays), and ecotoxic properties (as determined by Microtox™, Thamnotox™ and Daphnotox™ assays) that are representative of different trophic levels in aquatic environment (pozone (0.8 mg/L) and/or dissociated nitric and nitrous acid that contributed to the observed disinfection and toxicity. Chemical analysis of PPGD-treated water revealed increasing levels of electrode metals that were present at ≤ 30 times the tolerated respective values for EU drinking water. PUV-treated water did not exhibit any toxicity and was shown to be far superior to that of PPGD for killing C. parvum oocysts taking only 90 s of pulsing [UV dose of 6.29 μJ/cm(2)] to produce a 4-log reduction compared to a similar reduction level achieved after 32min PPGD treatment as determined by combined in vitro CaCo-2 cell culture-qPCR.

  4. THE EFFICACY OF THREE MEDICINAL PLANTS; GARLIC, GINGER AND MIRAZID AND A CHEMICAL DRUG METRONIDAZOLE AGAINST CRYPTOSPORIDIUM PARVUM: II-HISTOLOGICAL CHANGES.

    Science.gov (United States)

    Abouel-Nour, Mohamed F; El-Shewehy, Dina Magdy M; Hamada, Shadia F; Morsy, Tosson A

    2016-04-01

    Cryptosporidiosis parvum is a zoonotic protozoan parasite infects intestinal epithelial cells of man and animals causing a major health problem. This study was oriented to evaluate the protective and curative capacity of garlic, ginger and mirazid in comparison with metronidazole drug (commercially known) against Cryptosporidium in experimental mice. Male Swiss Albino mice experimentally infected with C. parvum were treated with medicinal plants extracts (Ginger, Mirazid, and Garlic) as compared to chemical drug Metronidazole. Importantly, C. parvum-infected mice treated with ginger, Mirazid, garlic and metronidazole showed a complete elimination in shedding oocysts by 9th day PI. The reduction and elimination of shedding oocysts in response to the treatments might be attributable to a direct effect on parasite growth in intestines, sexual phases production and/or the formation of oocysts. The results were evaluated histopathological examination of ideum section of control mice (uninfected, untreated) displayed normal architecture of the villi. Examiination of infected mice ileum section (infected, untreated) displayed histopathological alterations from uninfected groups. Examination of ileum section prepared from mice treated with garlic, ginger, mirazid, and metronidazole displayed histopathological alterations from that of the control groups, and showed marked histologic correction in the pattern with the four regimes used in comparison to control mice. Garlic successfully eradicated oocysts of infected mice from stool and intestine. Supplementation of ginger to infected mice markedly corrected elevation in the inflammatory risk factors and implied its potential antioxidant, anti-inflammatory and immunomodulatory capabilities. Infected mice treated with ginger, mirazid, garlic and metronidazole showed significant symptomatic improvements during treatment.

  5. Comparing the efficacy of chlorine, chlorine dioxide, and ozone in the inactivation of Cryptosporidium parvum in water from Parana State, Southern Brazil.

    Science.gov (United States)

    Pereira, Juliana Tracz; Costa, Adriana Oliveira; de Oliveira Silva, Márcia Benedita; Schuchard, Wagner; Osaki, Silvia Cristina; de Castro, Edilene Alcântara; Paulino, Rosangela Clara; Soccol, Vanete Thomaz

    2008-12-01

    In the present work, assays were performed to compare the efficacy of hypochlorous acid, chlorine dioxide, and ozone in the inactivation of Cryptosporidium oocyst in public water supply from Brazilian South conditions. Experiments were carried out in samples containing 2 x 10(4) oocysts/ml of C. parvum purified from feces of experimentally contaminated calves. An in vitro excystation method was used to evaluate oocysts' viability and to determine the inactivation rates of hypochlorous acid at 2 ppm, chlorine dioxide at 1, 2, and 5 ppm, and ozone at the doses of 0.18, 0.24, 0.36, 0.48, and 1.44 mg/l. By using hypochlorous acid, the maximum inactivation rate obtained was 49.04% after 120 min. Chlorine dioxide at 5 ppm inactivated 90.56% of oocysts after 90 min of contact. Ozone was the most effective product, rendering an inactivation of 100% with the concentration of 24 mg/l. Resistance of Cryptosporidium to the usual disinfectants and the need for more effective water treatments to prevent waterborne diseases in Brazil are discussed in this manuscript.

  6. 猪源隐孢子虫卵囊的分离及基因型鉴定%Isolation and genotype identification of Cryptosporidium oocysts in pigs

    Institute of Scientific and Technical Information of China (English)

    陈甫; 黄克和

    2011-01-01

    To investigate the genotypes of Cryptosporidium parasiting in pigs in Nanjing,Shanghai and Hefei, the total RNA of the oocysts from 6 positive feces was extracted using the TRIZOL reagent,and the partial genes of the small subunit ribosomal RNA (SSU rRNA) were amplified by the reverse transcription polymerase chain reaction (RT-PCR). SSU rRNA gene fragments of the isolates were deposited in the GenBank database under accession NO. DQ855266,DQ855267 and compared with sequences of various Cryptosporidium spp. and genotypes on the NCBI server using the BLAST and DNAStar software by the analysis of the homologous rate and phylogenetic tree. The results showed that the SSU rRNA gene fragments,compared with sequences of various Cryptosporidium spp. and genotypes,belonged to the species C. parvum with 94 %-100 % homologous rate, and were most similar to the C. parvum “mouse” genotype with 99. 8%-100 % homologous rate. The phylogenetic tree and sequence distances gen erated showed that those isolates belong to the C. parvum groups,and lie in the same divarication with C. parvum “mouse” genotype. It is concluded that those isolates are clearly distinct from other sequences as to merit species status, which is C. parvum “mouse” genotype from pigs. There might be a cross-transmission between pigs and mice.%应用RT-PCR方法对南京、上海和合肥猪源隐孢子虫卵囊SSU rRNA部分序列进行扩增,产物测序后提交GenBank,收录号为DQ855266、DQ855267;用BLAST和DNAStar软件与GenBank参考序列进行比较,分析其同源性,绘制系统发育进化树,结合卵囊形态学观察和对小鼠、大鼠、兔、山羊和鸡的传染性试验确定隐孢子虫种类或基因型.结果表明,3地区猪源隐孢子虫分离株与微小隐孢子虫(C.parvum)同源性达94%~100%,与C.parvum"mouse"型有99.8%~100%的同源性,并处于进化树的同一分支.因此,3地区猪源隐孢子虫是C.parvum"mouse"型,提示猪和鼠之间存在交叉传播的可能.

  7. Inhibitory Activities of Epidermal Growth Factor Receptor Tyrosine Kinase-Targeted Dihydroxyisoflavone and Trihydroxydeoxybenzoin Derivatives on Sarcocystis neurona, Neospora caninum, and Cryptosporidium parvum Development

    Science.gov (United States)

    Gargala, G.; Baishanbo, A.; Favennec, L.; François, A.; Ballet, J. J.; Rossignol, J.-F.

    2005-01-01

    Several gene sequences of parasitic protozoa belonging to protein kinase gene families and epidermal growth factor (EGF)-like peptides, which act via binding to receptor tyrosine kinases of the EGF receptor (EGFR) family, appear to mediate host-protozoan interactions. As a clue to EGFR protein tyrosine kinase (PTK) mediation and a novel approach for identifying anticoccidial agents, activities against Sarcocystis neurona, Neospora caninum, and Cryptosporidium parvum grown in BM and HCT-8 cell cultures of 52 EGFR PTK inhibitor isoflavone analogs (dihydroxyisoflavone and trihydroxydeoxybenzoine derivatives) were investigated. Their cytotoxicities against host cells were either absent, mild, or moderate by a nitroblue tetrazolium test. At concentrations ranging from 5 to 10 μg/ml, 20 and 5 analogs, including RM-6427 and RM-6428, exhibited an in vitro inhibitory effect of ≥95% against at least one parasite or against all three, respectively. In immunosuppressed Cryptosporidium parvum-infected Mongolian gerbils orally treated with either 200 or 400 mg of agent RM-6427/kg of body weight/day for 8 days, fecal microscopic oocyst shedding was abolished in 6/10 animals (P of 0.05, respectively). After RM-6427 therapy (200 mg/kg/day for 8 days), the reduction in the ratio of animals with intracellular parasites was nearly significant in ileum (P = 0.067) and more marked in the biliary tract (P < 0.0013) than after nitazoxanide or paromomycin treatment (0.05 < P < 0.004). RM-6428 treatment at a regimen of 400 mg/kg/day for 12 days inhibited oocyst shedding, measured using flow cytometry from day 4 (P < 0.05) to day 12 (P < 0.02) of therapy, when 2/15 animals had no shedding (P < 0.0001) and 11/15 were free of gut and/or biliary tract parasites (P < 0.01). No mucosal alteration was microscopically observed for treated or untreated infected gerbils. To our knowledge, this report is the first to suggest that the isoflavone class of agents has the potential for

  8. Comparison of transport and attachment behaviors of Cryptosporidium parvum oocysts and oocyst-sized microspheres being advected through three minerologically different granular porous media.

    Science.gov (United States)

    Mohanram, Arvind; Ray, Chittaranjan; Harvey, Ronald W; Metge, David W; Ryan, Joseph N; Chorover, Jon; Eberl, D D

    2010-10-01

    In order to gain more information about the fate of Cryptosporidium parvum oocysts in tropical volcanic soils, the transport and attachment behaviors of oocysts and oocyst-sized polystyrene microspheres were studied in the presence of two soils. These soils were chosen because of their differing chemical and physical properties, i.e., an organic-rich (43-46% by mass) volcanic ash-derived soil from the island of Hawaii, and a red, iron (22-29% by mass), aluminum (29-45% by mass), and clay-rich (68-76% by mass) volcanic soil from the island of Oahu. A third agricultural soil, an organic- (13% by mass) and quartz-rich (40% by mass) soil from Illinois, was included for reference. In 10-cm long flow-through columns, oocysts and microspheres advecting through the red volcanic soil were almost completely (98% and 99%) immobilized. The modest breakthrough resulted from preferential flow-path structure inadvertently created by soil-particle aggregation during the re-wetting process. Although a high (99%) removal of oocysts and microsphere within the volcanic ash soil occurred initially, further examination revealed that transport was merely retarded because of highly reversible interactions with grain surfaces. Judging from the slope of the substantive and protracted tail of the breakthrough curve for the 1.8-μm microspheres, almost all (>99%) predictably would be recovered within ∼4000 pore volumes. This suggests that once contaminated, the volcanic ash soil could serve as a reservoir for subsequent contamination of groundwater, at least for pathogens of similar size or smaller. Because of the highly reversible nature of organic colloid immobilization in this soil type, C. parvum could contaminate surface water should overland flow during heavy precipitation events pick up near-surface grains to which they are attached. Surprisingly, oocyst and microsphere attachment to the reference soil from Illinois appeared to be at least as sensitive to changes in pH as was

  9. An unexpected phosphate binding site in Glyceraldehyde 3-Phosphate Dehydrogenase: Crystal structures of apo, holo and ternary complex of Cryptosporidium parvum enzyme

    Directory of Open Access Journals (Sweden)

    Chattopadhyay Debasish

    2009-02-01

    Full Text Available Abstract Background The structure, function and reaction mechanism of glyceraldehyde 3-phosphate dehydrogenase (GAPDH have been extensively studied. Based on these studies, three anion binding sites have been identified, one 'Ps' site (for binding the C-3 phosphate of the substrate and two sites, 'Pi' and 'new Pi', for inorganic phosphate. According to the original flip-flop model, the substrate phosphate group switches from the 'Pi' to the 'Ps' site during the multistep reaction. In light of the discovery of the 'new Pi' site, a modified flip-flop mechanism, in which the C-3 phosphate of the substrate binds to the 'new Pi' site and flips to the 'Ps' site before the hydride transfer, was proposed. An alternative model based on a number of structures of B. stearothermophilus GAPDH ternary complexes (non-covalent and thioacyl intermediate proposes that in the ternary Michaelis complex the C-3 phosphate binds to the 'Ps' site and flips from the 'Ps' to the 'new Pi' site during or after the redox step. Results We determined the crystal structure of Cryptosporidium parvum GAPDH in the apo and holo (enzyme + NAD state and the structure of the ternary enzyme-cofactor-substrate complex using an active site mutant enzyme. The C. parvum GAPDH complex was prepared by pre-incubating the enzyme with substrate and cofactor, thereby allowing free movement of the protein structure and substrate molecules during their initial encounter. Sulfate and phosphate ions were excluded from purification and crystallization steps. The quality of the electron density map at 2Å resolution allowed unambiguous positioning of the substrate. In three subunits of the homotetramer the C-3 phosphate group of the non-covalently bound substrate is in the 'new Pi' site. A concomitant movement of the phosphate binding loop is observed in these three subunits. In the fourth subunit the C-3 phosphate occupies an unexpected site not seen before and the phosphate binding loop remains in

  10. Comparison of transport and attachment behaviors of Cryptosporidium parvum oocysts and oocyst-sized microspheres being advected through three minerologically different granular porous media

    Science.gov (United States)

    Mohanram, A.; Ray, C.; Harvey, R.W.; Metge, D.W.; Ryan, J.N.; Chorover, J.; Eberl, D.D.

    2010-01-01

    In order to gain more information about the fate of Cryptosporidium parvum oocysts in tropical volcanic soils, the transport and attachment behaviors of oocysts and oocyst-sized polystyrene microspheres were studied in the presence of two soils. These soils were chosen because of their differing chemical and physical properties, i.e., an organic-rich (43-46% by mass) volcanic ash-derived soil from the island of Hawaii, and a red, iron (22-29% by mass), aluminum (29-45% by mass), and clay-rich (68-76% by mass) volcanic soil from the island of Oahu. A third agricultural soil, an organic- (13% by mass) and quartz-rich (40% by mass) soil from Illinois, was included for reference. In 10-cm long flow-through columns, oocysts and microspheres advecting through the red volcanic soil were almost completely (98% and 99%) immobilized. The modest breakthrough resulted from preferential flow-path structure inadvertently created by soil-particle aggregation during the re-wetting process. Although a high (99%) removal of oocysts and microsphere within the volcanic ash soil occurred initially, further examination revealed that transport was merely retarded because of highly reversible interactions with grain surfaces. Judging from the slope of the substantive and protracted tail of the breakthrough curve for the 1.8-??m microspheres, almost all (>99%) predictably would be recovered within ~4000 pore volumes. This suggests that once contaminated, the volcanic ash soil could serve as a reservoir for subsequent contamination of groundwater, at least for pathogens of similar size or smaller. Because of the highly reversible nature of organic colloid immobilization in this soil type, C. parvum could contaminate surface water should overland flow during heavy precipitation events pick up near-surface grains to which they are attached. Surprisingly, oocyst and microsphere attachment to the reference soil from Illinois appeared to be at least as sensitive to changes in pH as was observed

  11. Cryptosporidium: from laboratory diagnosis to surveillance and outbreaks

    Directory of Open Access Journals (Sweden)

    Chalmers R.M.

    2008-09-01

    Full Text Available The burden of disease caused by the protozoan parasite Cryptosporidium is unknown. However, routine laboratory diagnosis and surveillance enables the basic epidemiology to be described, changes to be monitored and under-ascertainment to be measured. Although the two main species involved in human disease in developed countries, Cryptosporidium parvum and Cryptosporidium hominis, have differing epidemiologies and risk factors, national surveillance is generally from isolates identified to the genus level only. Enhancing the data by typing, at least to identify the isolates to the species level, removes some of the noise generated and better identifies the risks than when reports are not species-specific. This level of identification is also valuable for outbreak investigations, but further investigation of the population genetics of C. parvum and C. hominis is required for the development of more readily applied subtyping tools.

  12. A new genotype of Cryptosporidium from giant panda (Ailuropoda melanoleuca) in China.

    Science.gov (United States)

    Liu, Xuehan; He, Tingmei; Zhong, Zhijun; Zhang, Hemin; Wang, Rongjun; Dong, Haiju; Wang, Chengdong; Li, Desheng; Deng, Jiabo; Peng, Guangneng; Zhang, Longxian

    2013-10-01

    Fifty-seven fecal samples were collected from giant pandas (Ailuropoda melanoleuca) in the China Conservation and Research Centre for the Giant Panda (CCRCGP) in Sichuan and examined for Cryptosporidium oocysts by Sheather's sugar flotation technique. An 18-year-old male giant panda was Cryptosporidium positive, with oocysts of an average size of 4.60×3.99 μm (n=50). The isolate was genetically analyzed using the partial 18S rRNA, 70 kDa heat shock protein (HSP70), Cryptosporidium oocyst wall protein (COWP) and actin genes. Multi-locus genetic characterization indicated that the present isolate was different from known Cryptosporidium species and genotypes. The closest relative was the Cryptosporidium bear genotype, with 11, 10, and 6 nucleotide differences in the 18S rRNA, HSP70, and actin genes, respectively. Significant differences were also observed in the COWP gene compared to Cryptosporidium mongoose genotype. The homology to the bear genotype at the 18S rRNA locus was 98.6%, which is comparable to that between Cryptosporidium parvum and Cryptosporidium hominis (99.2%), or between Cryptosporidium muris and Cryptosporidium andersoni (99.4%). Therefore, the Cryptosporidium in giant pandas in this study is considered as a new genotype: the Cryptosporidium giant panda genotype.

  13. Comparison of the Triage Micro Parasite Panel and Microscopy for the Detection of Entamoeba histolytica/Entamoeba dispar, Giardia lamblia, and Cryptosporidium parvum in Stool Samples Collected in Kenya

    Directory of Open Access Journals (Sweden)

    Brett Swierczewski

    2012-01-01

    Full Text Available Entamoeba histolytica, Giardia lamblia, and Cryptosporidium parvum are three of the most important parasitic causes of acute diarrhea worldwide. Laboratory diagnosis of these parasites is usually done by ova and parasite examination (O&P examination via microscopy. The sensitivity and specificity of O&P examination varies among laboratories and can be labor intensive and time consuming. The Triage Micro Parasite Panel (BioSite, San Diego, California is an enzyme immunoassay kit that can detect E. histolytica/E. dispar, G. lamblia, and C. parvum simultaneously using fresh or frozen stool. The present study evaluated the Triage Micro Parasite Panel in detecting E. histolytica/E. dispar, G. lamblia, and C. parvum compared to O&P examination in 266 stool samples collected at medical facilities in Kenya. The sensitivity and specificity results for the Triage Micro Parasite Panel were: for E. histolytica/E. dispar: 100%, 100%, G. lamblia: 100%, 100% and C. parvum: 73%, 100%. There was no evidence of cross reactivity using the kit with other parasites identified in the stool specimens. These results indicate that the Triage Micro Parasite Panel is a highly sensitive kit that can be used for screening purposes in large scale studies or outbreak investigations or as a possible alternative to O&P examination.

  14. Prevalence and molecular characterization of Cryptosporidium spp. and Giardia duodenalis in dairy cattle in Beijing, China.

    Science.gov (United States)

    Li, Fuhuang; Wang, Haiyan; Zhang, Zhenjie; Li, Junqiang; Wang, Chenrong; Zhao, Jinfeng; Hu, Suhui; Wang, Rongjun; Zhang, Longxian; Wang, Ming

    2016-03-30

    822 fecal samples from cattle in six areas of Beijing were examined with microscopy for Cryptosporidium oocysts and Giardia cysts. The overall infection rates for Cryptosporidium spp. and Giardia duodenalis were 2.55% and 1.09%, respectively. Cryptosporidium was only detected in calves and heifers, whereas G. duodenalis was found in all age groups. Cryptosporidium spp. were characterized with a PCR-restriction fragment length polymorphism analysis and DNA sequence analysis of the small subunit (SSU) rRNA gene. Two Cryptosporidium species were identified: Cryptosporidium parvum (n=12) and Cryptosporidium andersoni (n=9). Six C. parvum isolates were successfully subtyped with the gp60 gene and three subtypes were detected: IIdA19G1 (n=1), IIdA17G1 (n=1), and IIdA15G1 (n=4). Subtype IIdA17G1 is reported for the first time in cattle worldwide. Nine G. duodenalis isolates were analyzed by sequencing the triosephosphate isomerase (tpi) gene, and only G. duodenalis assemblage E was identified. Therefore, the predominance of C. parvum detected in calves was identical to that found in the Xinjiang Uyghur and Ningxia Hui Autonomous Regions, but differed considerably from that in Henan, Heilongjiang, and Shannxi Provinces. In contrast, the predominance of G. duodenalis assemblage E was more or less similar to its predominance in other areas of China or countries. Our findings confirm the unique character of the C. parvum IId subtypes in China. More systematic studies are required to better understand the transmission of Cryptosporidium and G. duodenalis in cattle in China.

  15. Over-expression and localization of a host protein on the membrane of Cryptosporidium parvum infected epithelial cells.

    Science.gov (United States)

    Yang, Yi-Lin; Serrano, Myrna G; Sheoran, Abhineet S; Manque, Patricio A; Buck, Gregory A; Widmer, Giovanni

    2009-11-01

    The genus Cryptosporidium includes several species of intestinal protozoan parasites which multiply in intestinal epithelial cells. The impact of this infection on the transcriptome of cultured host cells was investigated using DNA microarray hybridizations. The expression of 14 genes found to be consistently up- or down-regulated in infected cell monolayers was validated with RT PCR. Using immunofluorescence we examined the expression of Protease Activated Receptor-2, which is encoded by one of the up-regulated genes. In infected cells this receptor localized to the host cell membrane which covers the intracellular trophozoites and meronts. This observation indicates that the composition of the host cell membrane is affected by the developing trophozoite, a phenomenon which has not been described previously.

  16. 微小隐孢子虫的基因检测实验研究%EXPERIMENTAL STUDY ON GENE DETECTION OF CRYPTOSPORIDIUM PARVUM

    Institute of Scientific and Technical Information of China (English)

    丁慧萍; 卢思奇; 李凤舞; 戎煜; 张西臣; 王凤云

    2002-01-01

    目的探讨聚合酶链反应(PCR)检测微小隐孢子虫(Cryptosporidium parvum)感染的特异性和敏感性. 方法用Sheather's蔗糖梯度离心法从微小隐孢子虫感染者粪便标本中分离纯化卵囊.用酚-氯仿法抽提卵囊总DNA.针对本虫18S rRNA基因片段设计1对引物,对模板DNA进行PCR扩增,同时以蓝氏贾第鞭毛虫(Giardia lamblia)、溶组织内阿米巴(Entamoeba histolytica)、阴道毛滴虫(Trichomonas vaginalis)、刚地弓形虫(Toxoplasma gondii)、日本血吸虫(Schistosoma japonicum)和旋毛虫(Trichinella spiralis),以及人体血细胞等DNA样本为对照.用琼脂糖电泳和溴乙腚染色对PCR产物进行检测. 结果仅微小隐孢子虫DNA被扩增出一 500 bp的DNA片段.其它对照DNA样本均未出现扩增反应. 结论 PCR对隐孢子虫检测的特异性可高达100%,敏感性也很强,可检测到20 pg微小隐孢子虫卵囊的DNA.表明本实验建立的检测微小隐孢子虫的PCR方法有效.

  17. Solar Radiation Induces Non-Nuclear Perturbations and a False Start to Regulated Exocytosis in Cryptosporidium parvum

    Science.gov (United States)

    King, Brendon J.; Hoefel, Daniel; Ee Wong, Pao; Monis, Paul T.

    2010-01-01

    Stratospheric ozone depletion, climate warming and acidification of aquatic ecosystems have resulted in elevated levels of solar radiation reaching many aquatic environments with an increased deleterious impact on a wide range of living organisms. While detrimental effects on living organisms are thought to occur primarily through DNA damage, solar UV can also damage cellular proteins, lipids and signalling pathways. Cryptosporidium, a member of the eukaryotic phylum Apicomplexa, contain numerous vesicular secretory organelles and their discharge via regulated exocytosis is essential for the successful establishment of infection. Using flow cytometric techniques we demonstrate that solar UV rapidly induces sporozoite exocytosis resulting in a significant reduction in the ability of sporozoites to attach and invade host cells. We found that solar UV induced sporozoite membrane depolarization, resulting in reduced cellular ATP and increased cytosolic calcium. These changes were accompanied by a reduction in the internal granularity of sporozoites, indicative of apical organelle discharge, which was confirmed by analysis of sporozoites with an exocytosis-sensitive dye. The precise timing of apical organelle discharge in the presence of a compatible host cell is critical for sporozoite attachment and invasion. Our results demonstrate for the first time how solar UV radiation can interfere with exocytosis, a fundamental cellular process in all eukaryotic cells. We contend that not only may the forecast increases in solar radiation in both aquatic and terrestrial environments significantly affect members of the Apicomplexa, solar UV-induced membrane depolarizations resulting in cytosolic calcium perturbation may affect a wider range of eukaryotic organisms through antagonistic effects on a myriad of calcium dependant cellular functions. PMID:20668710

  18. Solar radiation induces non-nuclear perturbations and a false start to regulated exocytosis in Cryptosporidium parvum.

    Science.gov (United States)

    King, Brendon J; Hoefel, Daniel; Wong, Pao Ee; Monis, Paul T

    2010-07-23

    Stratospheric ozone depletion, climate warming and acidification of aquatic ecosystems have resulted in elevated levels of solar radiation reaching many aquatic environments with an increased deleterious impact on a wide range of living organisms. While detrimental effects on living organisms are thought to occur primarily through DNA damage, solar UV can also damage cellular proteins, lipids and signalling pathways. Cryptosporidium, a member of the eukaryotic phylum Apicomplexa, contain numerous vesicular secretory organelles and their discharge via regulated exocytosis is essential for the successful establishment of infection. Using flow cytometric techniques we demonstrate that solar UV rapidly induces sporozoite exocytosis resulting in a significant reduction in the ability of sporozoites to attach and invade host cells. We found that solar UV induced sporozoite membrane depolarization, resulting in reduced cellular ATP and increased cytosolic calcium. These changes were accompanied by a reduction in the internal granularity of sporozoites, indicative of apical organelle discharge, which was confirmed by analysis of sporozoites with an exocytosis-sensitive dye. The precise timing of apical organelle discharge in the presence of a compatible host cell is critical for sporozoite attachment and invasion. Our results demonstrate for the first time how solar UV radiation can interfere with exocytosis, a fundamental cellular process in all eukaryotic cells. We contend that not only may the forecast increases in solar radiation in both aquatic and terrestrial environments significantly affect members of the Apicomplexa, solar UV-induced membrane depolarizations resulting in cytosolic calcium perturbation may affect a wider range of eukaryotic organisms through antagonistic effects on a myriad of calcium dependant cellular functions.

  19. Molecular characterization of Cryptosporidium spp. in dairy calves from the state of São Paulo, Brazil.

    Science.gov (United States)

    Meireles, Marcelo V; de Oliveira, Fernando P; Teixeira, Weslen Fabrício P; Coelho, William M D; Mendes, Luiz Cláudio N

    2011-09-01

    Cryptosporidiosis is a common protozoan disease observed in a wide range of vertebrate hosts, including ruminants. Cattle can be a potential reservoir of Cryptosporidium spp., leading to environmental contamination with oocysts of zoonotic species. The molecular characterization of Cryptosporidium spp. isolated from cattle from the state of São Paulo, Brazil, was accomplished using nested polymerase chain reaction for amplification of fragments of the 18S rRNA gene and the glycoprotein GP60 gene, following sequencing of amplified fragments. Positivity for Cryptosporidium was found in 10.7% (21/196) of the samples. Four species of Cryptosporidium were identified: C. andersoni, C. bovis, C. parvum subtype IIaA15G2R1, and C. ryanae. To the best of our knowledge, this is the first report of infection by C. ryanae and C. parvum IIaA15G2R1 in cattle from Brazil.

  20. Is real-time PCR-based diagnosis similar in performance to routine parasitological examination for the identification of Giardia intestinalis, Cryptosporidium parvum/Cryptosporidium hominis and Entamoeba histolytica from stool samples? Evaluation of a new commercial multiplex PCR assay and literature review.

    Science.gov (United States)

    Laude, A; Valot, S; Desoubeaux, G; Argy, N; Nourrisson, C; Pomares, C; Machouart, M; Le Govic, Y; Dalle, F; Botterel, F; Bourgeois, N; Cateau, E; Leterrier, M; Le Pape, P; Morio, F

    2016-02-01

    Microscopy is the reference standard for routine laboratory diagnosis in faecal parasitology but there is growing interest in alternative methods to overcome the limitations of microscopic examination, which is time-consuming and highly dependent on an operator's skills and expertise. Compared with microscopy, DNA detection by PCR is simple and can offer a better turnaround time. However, PCR performances remain difficult to assess as most studies have been conducted on a limited number of positive clinical samples and used in-house PCR methods. Our aim was to evaluate a new multiplex PCR assay (G-DiaParaTrio; Diagenode Diagnostics), targeting Giardia intestinalis, Cryptosporidium parvum/Cryptosporidium hominis and Entamoeba histolytica. To minimize the turnaround time, PCR was coupled with automated DNA extraction (QiaSymphony; Qiagen). The PCR assay was evaluated using a reference panel of 185 samples established by routine microscopic examination using a standardized protocol including Ziehl-Neelsen staining and adhesin detection by ELISA (E. histolytica II; TechLab). This panel, collected from 12 French parasitology laboratories, included 135 positive samples for G. intestinalis (n = 38), C. parvum/C. hominis (n = 26), E. histolytica (n = 5), 21 other gastrointestinal parasites, together with 50 negative samples. In all, the G-DiaParaTrio multiplex PCR assay identified 38 G. intestinalis, 25 C. parvum/C. hominis and five E. histolytica leading to sensitivity/specificity of 92%/100%, 96%/100% and 100%/100% for G. intestinalis, C. parvum/C. hominis and E. histolytica, respectively. This new multiplex PCR assay offers fast and reliable results, similar to microscopy-driven diagnosis for the detection of these gastrointestinal protozoa, allowing its implementation in routine clinical practice.

  1. Experimental transmission of Cryptosporidium oocyst isolates from mammals, birds and reptiles to captive snakes.

    Science.gov (United States)

    Graczyk, T K; Cranfield, M R

    1998-01-01

    Groups of four to five, 3-month-old rat snakes (Elaphe obsoleta) were separately gastrically inoculated with 2.0 x 10(6) viable oocysts of Cryptosporidium muris (mice and calves), C. muris-like (Bactrian camels), C. wrairi (guinea pigs), C. baileyi (chickens), C. meleagridis (turkeys), Cryptosporidium sp. (turtles, tortoises, chameleons and lizards) and C. serpentis from clinically (fatal case) and subclinically infected snakes. None of the snakes inoculated with oocysts originating from homothermous vertebrates developed infection as determined by histology and serology, whereas all snakes challenged with reptilian oocyst isolates were infected with Cryptosporidium on weeks 6 and 10 post-inoculation (PI). One week 10 PI, the snakes displayed mild to serve, multifocal to widespread, thinning and disorganization of gastric epithelium and nine out of twelve snakes infected by oocysts originating from reptiles other than snakes displayed severe gastric hyperplasia. Three out of ten snakes infected oocysts originating from snakes had ELISA-detectable Cryptosporidium-specific antibody (Ab) titers on week 6 PI; all snakes were Cryptosporidium-seroconverted on week 10 PI and their serum Ab titer significantly increased. The study demonstrated that Cryptosporidium infections in snakes maintained on the diet of rodents or birds cannot be initiated via ingestion of an infected food item; however, snakes can void ingested oocysts. Lack of host specificity among reptiles to this pathogen, demonstrated for the first time in the present study, indicates that snake-attributed C. serpentis is not distinct from Cryptosporidium sp. infecting reptiles other than snakes, and that clinical manifestations and virulence of Cryptosporidium in snakes in modulated by the species of the host. Housing of snakes with other reptiles can enhance transmission of Cryptosporidium to snakes, and therefore should be avoided.

  2. Occurrence of Cryptosporidium and Giardia on beef farms and water sources within the vicinity of the farms on Prince Edward Island, Canada.

    Science.gov (United States)

    Budu-Amoako, Ebo; Greenwood, Spencer J; Dixon, Brent R; Barkema, Herman W; McClure, J T

    2012-02-28

    The objectives of this study were to determine the prevalence and assemblages of Giardia and species of Cryptosporidium on beef farms in Prince Edward Island (PEI), Canada, including the water sources associated with the farms, and to determine risk factors for infection of cattle with these parasites. Twenty beef farms were selected based on the presence of surface waterGiardia was detected in 42% (95% CI: 38-46%) of fecal samples from 100% farms while Cryptosporidium was detected in 17% (95% CI: 14-19%) of fecal samples from 80% of farms. The most predominant Giardia assemblage isolated was the livestock specific assemblage E (89%). The zoonotic assemblages A and B were found in 4 and 7% of the Giardia isolates that were genotyped, respectively. The Giardia assemblages were detected equally between the cows and calves examined. Overall, the most common Cryptosporidium species detected in this study was Cryptosporidium andersoni (49%), predominantly found in cattle > 6 mo of age, while most Cryptosporidium bovis and Cryptosporidium pestis (previously Cryptosporidium parvum 'bovine genotype') isolates were detected in calves ≤ 6 mo of age. All Cryptosporidium ryanae isolates (four) were found in calves. Giardia cysts and Cryptosporidium oocysts were detected in 14 and 93% of surface water samples of 14 farms, respectively. Cryptosporidium oocysts were detected in three (15%) ground water samples of 20 farms. One Cryptosporidium-positive water sample, which was the only surface water sample amenable to genotyping, contained C. parvum. The farm-level risk factors investigated in this study, age of animals and location of the farm, were not associated with the risk of infection in cattle with either Cryptosporidium spp. or Giardia duodenalis. We conclude that beef cattle are a potential reservoir of Cryptosporidium spp. and G. duodenalis that could contaminate source water. There is the possibility of further transmission to humans on PEI if the source water is not

  3. Genotypic characterization and phylogenetic analysis of Cryptosporidium sp. from domestic animals in Brazil.

    Science.gov (United States)

    Huber, F; da Silva, S; Bomfim, T C B; Teixeira, K R S; Bello, A R

    2007-11-30

    The purpose of the present study was the genetic characterization, sequencing and phylogenetic analysis of 18S rDNA sequences of Cryptosporidium isolates obtained from different animal hosts in Brazil. Fecal samples containing Cryptosporidium oocysts were obtained from chickens, ducks, quails, guinea pigs, dairy calves, dogs and cats. For amplification of 18S rDNA sequences the Secondary-PCR product of the extracted DNA from fecal suspension of each studied animal was utilized. The primary genetic characterization of Cryptosporidium sp. was performed using RFLP with the enzymes SspI and VspI. DNA samples were sequenced and subjected to phylogenetic analysis. The results showed C. baileyi infecting two ducks and one quail and C. melagridis infecting one chicken. The sequences obtained from Cryptosporidium sp. infecting guinea pigs were not identified within groups of known Cryptosporidium species. The isolates found parasitizing cats and one dog were diagnosed as C. felis and C. canis, respectively. One isolate of calf origin was identified as C. parvum. The phylogenetic analysis showed clear distribution of isolates between two Cryptosporidium sp. groups according to their gastric or intestinal parasitism. A great genetic distance was observed between C. felis and C. canis from Brazil when compared to the reference sequences obtained from GenBank. The results obtained during this study constitute the first report of rDNA sequences from C. baileyi, C. meleagridis, C. felis, C. canis and C. parvum isolated in Brazil.

  4. Detection and differentiation of Cryptosporidium by real-time polymerase chain reaction in stool samples from patients in Rio de Janeiro, Brazil

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    Roberta Flávia Ribeiro Rolando

    2012-06-01

    Full Text Available This study reports the first genetic characterisation of Cryptosporidium isolates in Brazil using real-time polymerase chain reaction (RT-PCR. A total of 1,197 faecal specimens from children and 10 specimens from human immunodeficiency virus-infected patients were collected between 1999-2010 and screened using microscopy. Forty-eight Cryptosporidium oocyst-positive isolates were identified and analysed using a generic TaqMan assay targeting the 18S rRNA to detect Cryptosporidium species and two other TaqMan assays to identify Cryptosporidium hominis and Cryptosporidium parvum. The 18S rRNA assay detected Cryptosporidium species in all 48 of the stool specimens. The C. parvum TaqMan assay correctly identified five/48 stool samples, while 37/48 stool specimens were correctly amplified in the C. hominis TaqMan assay. The results obtained in this study support previous findings showing that C. hominis infections are more prevalent than C. parvum infections in Brazil and they demonstrate that the TaqMan RT-PCR procedure is a simple, fast and valuable tool for the detection and differentiation of Cryptosporidium species.

  5. Molecular genotyping and sub-genotyping of Cryptosporidium spp. isolates from symptomatic individuals attending two major public hospitals in Madrid, Spain.

    Science.gov (United States)

    de Lucio, Aida; Merino, Francisco J; Martínez-Ruiz, Rocío; Bailo, Begoña; Aguilera, María; Fuentes, Isabel; Carmena, David

    2016-01-01

    Infections by members of the protozoan genus Cryptosporidium are among the most common causes of human gastrointestinal illness worldwide. In Spain cryptosporidiosis is not a compulsory notifiable disease, so the actual burden of the infection in both clinical and general populations remains largely unknown. We present here data on the diversity and frequency of the Cryptosporidium species and sub-genotypes identified in symptomatic individuals seeking medical care in two major hospitals in Madrid, Spain, between December 2013 and January 2015. Initial detection of the parasite was conducted on a total of 122 stool samples collected from 120 patients by microscopy with modified Ziehl-Neelsen and/or immunochromatographic tests. We used immunofluorescence, PCR-based methods and sequence analyses of the 60-kDa (GP60) glycoprotein and the small subunit ribosomal RNA (SSU rRNA) genes for confirmatory purposes and to characterize Cryptosporidium isolates. A total of 110 patients were confirmed with cryptosporidiosis. Overall, 101 isolates were successfully sub-genotyped at the GP60 locus, and an additional seven at the SSU rRNA locus. The analyses of all amplicons defined 10 distinct sequence types representing the GP60 family sub-genotypes IbA10G2 (78.7%), IeA11G3T3 (3.7%) of C. hominis, and the GP60 family sub-types IIaA15G2R1 (5.6%), IIaA18G6R1 (0.9%), IIcA5G3a (0.9%), IIdA18G1 (0.9%), IIdA19G1 (0.9%), IIdA21G1 (0.9%), and IIdA22G1 (0.9%) of C. parvum. A single isolate was assigned to C. felis (0.9%), two C. parvum isolates (1.9%) could not be characterized at the sub-genotype level and an additional four isolates (3.7%) were not typable. These results strongly suggest that transmission of cryptosporidiosis is mostly anthroponotic in origin in the clinical sample under study. We expect that our molecular epidemiological data will make a significant contribution to unravel the actual epidemiological situation of cryptosporidiosis in Spain, providing health care and

  6. Synthesis and modulation properties of imidazo[4,5-b]pyridin-7-one and indazole-4,7-dione derivatives towards the Cryptosporidium parvum CpABC3 transporter.

    Science.gov (United States)

    Zeinyeh, Waël; Xia, Hexue; Lawton, Philippe; Radix, Sylvie; Marminon, Christelle; Nebois, Pascal; Walchshofer, Nadia

    2010-06-01

    The syntheses of new N-polysubstituted imidazo[4,5-b]pyridine-7-one (IP, 5 and 8a-8f) and indazole-4,7-dione (ID, 9 and 10) derivatives are described. The binding affinity of IP and ID towards the recombinant Nucleotide Binding Domain NBD1 of Cryptosporidium parvum CpABC3 was evaluated by intrinsic fluorescence quenching. IP induced a moderate quenching of the intrinsic fluorescence of H6-NBD1 whereas IDs 9 and 10 showed a binding affinity comparable to the ATP analogue TNP-ATP. In addition, 8d, 8e and 10 were shown to be competitive inhibitors of the ATPase activity, but with low affinity. These compounds could thus act like some flavonoid derivatives, which can partly overlap both the nucleotide-binding site and the adjacent hydrophobic steroid-binding region of mammalian P-glycoproteins.

  7. Cryptosporidium and Giardia as foodborne zoonoses.

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    Smith, H V; Cacciò, S M; Cook, N; Nichols, R A B; Tait, A

    2007-10-21

    Cryptosporidium and Giardia are major causes of diarrhoeal disease in humans, worldwide and are major causes of protozoan waterborne diseases. Both Cryptosporidium and Giardia have life cycles which are suited to waterborne and foodborne transmission. There are 16 'valid'Cryptosporidium species and a further 33+ genotypes described. Parasites which infect humans belong to the Giardia duodenalis "type", and at least seven G. duodenalis assemblages are recognised. Cryptosporidium parvum is the major zoonotic Cryptosporidium species, while G. duodenalis assemblages A and B have been found in humans and most mammalian orders. In depth studies to determine the role of non-human hosts in the transmission of Cryptosporidium and Giardia to humans are required. The use of harmonised methodology and standardised and validated molecular markers, together with sampling strategies that provide sufficient information about all contributors to the environmental (oo)cyst pool that cause contamination of food and water, are recommended. Standardised methods for detecting (oo)cysts in water are available, as are optimised, validated methods for detecting Cryptosporidium in soft fruit and salad vegetables. These provide valuable data on (oo)cyst occurrence, and can be used for species and subspecies typing using appropriate molecular tools. Given the zoonotic potential of these organisms, epidemiological, source and disease tracking investigations involve multidisciplinary teams. Here, the role of the veterinarian is paramount, particularly in understanding the requirement for adopting comprehensive sampling strategies for analysing both sporadic and outbreak samples from all potential non-human contributors. Comprehensive sampling strategies increase our understanding of parasite population biology and structure and this knowledge can be used to determine what level of discrimination is required between isolates. Genetic exchange is frequent in C. parvum populations, leading to

  8. Molecular epidemiology of Cryptosporidium in HIV/AIDS patients in Malaysia.

    Science.gov (United States)

    Asma, I; Sim, B L H; Brent, R D; Johari, S; Yvonne Lim, A L

    2015-06-01

    Cryptosporidiosis is a particular concern in immunocompromised individuals where symptoms may be severe. The aim of this study was to examine the epidemiological and molecular characteristics of Cryptosporidium infections in HIV/AIDS patients in Malaysia in order to identify risk factors and facilitate control measures. A modified Ziehl-Neelsen acid fast staining method was used to test for the presence of Cryptosporidium oocysts in the stools of 346 HIV/AIDS patients in Malaysia. Standard coproscopical methods were used to identify infections with other protozoan or helminths parasites. To identify the species of Cryptosporidium, DNA was extracted and nested-PCR was used to amplify a portion of the SSU rRNA gene. A total of 43 (12.4%) HIV-infected patients were found to be infected with Cryptosporidium spp. Of the 43 Cryptosporidium-positive HIV patients, 10 (23.3%) also harboured other protozoa, and 15 (34.9%) had both protozoa and helminths. The highest rates of cryptosporidiosis were found in adult males of Malay background, intravenous drug users, and those with low CD4 T cell counts (i.e., < 200 cells/mm3). Most were asymptomatic and had concurrent opportunistic infections mainly with Mycobacterium tuberculosis. DNA sequence analysis of 32 Cryptosporidium isolates identified C. parvum (84.3%), C. hominis (6.3%), C. meleagridis (6.3%), and C. felis (3.1%). The results of the present study revealed a high prevalence of Cryptosporidium infection in hospitalized HIV/AIDS patients. The results also confirmed the potential significance of zoonotic transmission of C. parvum in HIV infected patients, as it was the predominant species found in this study. However, these patients were found to be susceptible to a wide range of Cryptosporidium species. Epidemiological and molecular characterization of Cryptosporidium isolates provides clinicians and researchers with further information regarding the origin of the infection, and may enhance treatment and control

  9. Prevalence and abundance of Cryptosporidium parvum and giardia spp. in wild rural rodents from the Mazury Lake District region of Poland.

    Science.gov (United States)

    Bajer, A; Bednarska, M; Pawełczyk, A; Behnke, J M; Gilbert, F S; Sinski, E

    2002-07-01

    Prevalence and abundance of Cryptosporodium parvum and Giardia spp. were studied in 3 species of rodents from forests and abandoned agricultural fields in N.E. Poland (Clethrionomys glareolus n = 459; Microtus arvalis n = 274; Apodemus flavicollis n = 209). Overall prevalence was consistently higher in the voles compared with A. flavicollis (70.6, 73.0 and 27.8% respectively for C. parvum and 93.9, 96.3 and 48.3% respectively for Giardia spp.). Prevalence and abundance of infection also varied markedly across 3 years with 1998 being a year of higher prevalence and abundance with both species. Fewer older animals (especially C. glareolus and M. arvalis) carried infection with C. parvum and infections in these animals were relatively milder. Although seasonal differences were significant, no consistent pattern of changes was apparent. Host sex did not influence prevalence or abundance of infection with C. parvum, but made a small contribution to a 4-way interaction (in 5-way ANOVA) with other factors in the case of Giardia spp. The 2 species co-occurred significantly and in animals carrying both parasites there was a highly significant positive correlation between abundance of infection with each, even with between-year, seasonal, host age, sex and species differences taken into account. Quantitative associations were confined to the 2 vole species in the study. These results are discussed in relation to the importance of wild rodents as reservoir hosts and sources of infection for local human communities.

  10. Effects of sediment-associated extractable metals, degree of sediment grain sorting, and dissolved organic carbon upon cryptosporidium parvum removal and transport within riverbank filtration sediments, Sonoma County, California

    Science.gov (United States)

    Metge, D.W.; Harvey, R.W.; Aiken, G.R.; Anders, R.; Lincoln, G.; Jasperse, J.; Hill, M.C.

    2011-01-01

    Oocysts of the protozoan pathogen Cryptosporidium parvum are of particular concern for riverbank filtration (RBF) operations because of their persistence, ubiquity, and resistance to chlorine disinfection. At the Russian River RBF site (Sonoma County, CA), transport of C. parvum oocysts and oocyst-sized (3 ??m) carboxylate-modified microspheres through poorly sorted (sorting indices, ??1, up to 3.0) and geochemically heterogeneous sediments collected between 2 and 25 m below land surface (bls) were assessed. Removal was highly sensitive to variations in both the quantity of extractable metals (mainly Fe and Al) and degree of grain sorting. In flow-through columns, there was a log-linear relationship (r2 = 0.82 at p < 0.002) between collision efficiency (??, the probability that colloidal collisions with grain surfaces would result in attachment) and extractable metals, and a linear relationship (r2 = 0.99 at p < 0.002) between ?? and ??1. Collectively, variability in extractable metals and grain sorting accounted for ???83% of the variability in ?? (at p < 0.0002) along the depth profiles. Amendments of 2.2 mg L-1 of Russian River dissolved organic carbon (DOC) reduced ?? for oocysts by 4-5 fold. The highly reactive hydrophobic organic acid (HPOA) fraction was particularly effective in re-entraining sediment-attached microspheres. However, the transport-enhancing effects of the riverine DOC did not appear to penetrate very deeply into the underlying sediments, judging from high ?? values (???1.0) observed for oocysts being advected through unamended sediments collected at ???2 m bls. This study suggests that in evaluating the efficacy of RBF operations to remove oocysts, it may be necessary to consider not only the geochemical nature and size distribution of the sediment grains, but also the degrees of sediment sorting and the concentration, reactivity, and penetration of the source water DOC. ?? 2011 American Chemical Society.

  11. Small rodents as reservoirs of Cryptosporidium spp. and Giardia spp. in south-western Poland.

    Science.gov (United States)

    Perec-Matysiak, Agnieszka; Buńkowska-Gawlik, Katarzyna; Zaleśny, Grzegorz; Hildebrand, Joanna

    2015-01-01

    Cryptosporidium spp. and Giardia spp. have been detected in a range of host species, including rodents. The aim of this study was to determine the distribution of these pathogens and recognition of the reservoir role of rodents in the maintenance of these pathogens in south-western Poland. Additionally, preliminary molecular studies were conducted to elucidate the species and genotypes of Cryptosporidium and Giardia identified in this study. Stool samples (n=266) from A. agrarius, A. flavicollis and M. glareolus, were subjected for analyses. Values of prevalence were 61.7, 68.3 and 68.1%, respectively, for Cryptosporidium spp. and 41.7, 24.4 and 38.4%, respectively, for Giardia spp. There was a statistically significant correlation between host species and Giardia infection where A. agrarius was the species of the highest prevalence. Statistically significant differences were not found for comparisons made for study sites and occurrence of Giardia spp. and Cryptosporidium spp. Due to preliminary nested PCR results, specific amplifications of Cryptosporidium COWP and SSU rRNA genes were obtained for several isolates taken from rodent host species. One isolate recovered from A. agrarius (from a semi-aquatic, urban area) was identified as C. parvum and revealed 100% similarity with sequences obtained from humans. To the best of the knowledge of the authors, this is the first record of the C. parvum zoonotic species from the striped field mouse. Also recorded were the first findings of C. ubiquitum from three small rodent species.

  12. Molecular characterization of Cryptosporidium isolates from beef calves under one month of age over three successive years in one herd in western France.

    Science.gov (United States)

    Rieux, Anaïs; Paraud, Carine; Pors, Isabelle; Chartier, Christophe

    2014-05-28

    Cohorts of pre-weaned calves were studied for Cryptosporidium infection over three successive years (2010-2012) in one beef cattle herd in western France. Each year 25-34 calves were sampled weekly from 3 days to one month of age in order to characterize oocyst output, Cryptosporidium species and clinical features associated with infection. Faecal samples were screened for the presence of oocysts using immunofluorescence analysis. DNA was extracted from positive samples and a PCR SSU rRNA followed by RFLP or sequencing was performed. For the subtyping of C. parvum, a gp60 PCR was carried out. Regardless of the year, 92-100% of the animals excreted oocysts on at least one sampling date. Depending on the year of observation, the age of highest prevalence varied. In contrast, the peak of excretion was systematically observed almost at the same age (2nd-3rd week of life) with excretion levels ranging from between 100 and 1.7 × 10(7)oocysts/g of faeces. Differences concerning clinical signs depending on the year of sampling were observed. Different species patterns were observed, with a predominance of C. bovis in the 1st year and a predominance of C. parvum in the last year. Moreover, two zoonotic subtypes of C. parvum, IIaA15G2R1 and IIaA18G2R1, were recorded in different years. This study shows that, in a given farm, the Cryptosporidium species and C. parvum subtypes identified as well as the prevalence of infection and level of excretion may vary greatly and show distinct patterns according to the year. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. Molecular characterisation of Cryptosporidium and Giardia in cats (Felis catus) in Western Australia.

    Science.gov (United States)

    Yang, Rongchang; Ying, Joyce Lau Jie; Monis, Paul; Ryan, Una

    2015-08-01

    Little is known of the prevalence of Cryptosporidium and Giardia in domestic cats in Western Australia and their potential role as zoonotic reservoirs for human infection. In the present study, a total of 345 faecal samples from four different sources were screened for the presence of Cryptosporidium and Giardia by PCR and genotyped by sequence analysis. Oocyst numbers and cyst numbers for Cryptosporidium and Giardia respectively were also determined using quantitative PCR assays. Cryptosporidium and Giardia were detected in 9.9% (95% CI 6.7-13.0) and 10.1% (95% CI 7.0-13.3) of cats in Western Australia respectively. Sequence analysis at the 18S rRNA locus identified five Cryptosporidium species/genotypes; C. felis (n = 8), C. muris (n = 1), C. ryanae (n = 1), Cryptosporidium rat genotype III (n = 5) and a novel genotype most closely related to Cryptosporidium rat genotype III in one isolate. This is the first report of C. ryanae and Cryptosporidium rat genotype III in cats. For Giardia, assemblage F the most commonly identified species, while only 1 assemblage sequence was detected. Since most human cases of cryptosporidiosis are caused by C. parvum and C. hominis and human cases of giardiasis are caused by G. duodenalis assemblage A and B, the domestic cats in the present study are likely to be of low zoonotic risk to pet owners in Perth. Risk analyses identified that elderly cats (more than 6 years) were more prone to Cryptosporidium and Giardia infections than kittens (less than 6 months) (P = 0.009). Clinical symptoms were not associated with the prevalence of Cryptosporidium and Giardia infections in cats.

  14. METHODS FOR DETECTION OF CRYPTOSPORIDIUM SP. AND GIARDIA SP.

    Science.gov (United States)

    There have been several waterborne outbreaks of giardiasis caused by infection with Giardia lamblia, and cryptosporidiosis, caused by infection with Cryptosporidium parvum. These outbreaks have created a need to detect these organisms in source and finished drinking water. The pr...

  15. Development of an IMS-qPCR Method for Detection of Cryptosporidium parvum in Water%IMS-qPCR检测水源性微小隐孢子虫方法的建立

    Institute of Scientific and Technical Information of China (English)

    高姗姗; 吴绍强; 罗静; 王承民; 张敏; 赵宝华; 何宏轩

    2013-01-01

    Objective To develop a detection method for Cryptosporidium parvum oocysts from water samples,which combined immunomagnetic separation (IMS) with Taqman real-time PCR (qPCR).Methods Conditions of separation and enrichment of IMS method by using specific streptavidin magnetic beads coated with monoclonal antibodies Cp23 directed against C.parvum oocysts were optimized.Special primers of PCR and Taqman probes were designed referring to the 18S rDNA gene of C.parvum (GenBank Accession No.AB513881.1).The conserved genes were amplified from genomic DNA of C.parvum,and then cloned into Peasy-T1 vector.Tenfold dilutions of positive plasmids(104-108copy/μl) were used to construct a standard curves by Taqman qPCR.The specificity of the assay was determined using genomic DNA of C.baileyi,Toxoplasma gondii,C.canis and Escherichia coli.The sensitivity of this assay was evaluated by analyzing 10-fold serially dilutions of plasmids ranging from 100 to 108 copy/μl.Both IMS-qPCR assay and indirect immunofluorescent-antibody assay (IFA) were applied to detect 50 water samples collected from the dairy cattle farms in Hebei.Results The optimal incubation concentration and time of antibody Cp23 were 20 ng/ml and 30 min,respectively,and the catching time was 30 min,the recovery rate was more than 95%.PCR product was 272 bp,and identified by restriction enzyme digestion and nucleotide sequencing.There was a good linear relationship between the standard plasmids and Ct value (correlation r2=0.996 1) of the Taqman qPCR.No cross-reactivity was observed with C.baileyi,T.gondii,C.canis and E.coli.The sensitivity of C.parvum-specific assay was 10 copy/μl.Compared with IFA as golden standard method,the specificity and sensitivity of IMS-qPCR for 50 water samples was 100%(18/18) and 93.8%(30/32),respectively.Conclution The IMS-qPCR assay can be used to specifically detect C.parvum oocysts in water samples.%目的 将免疫磁珠分离技术(IMS)和荧光探针定量PCR (qPCR)相结

  16. AIDS-associated diarrhea and wasting in northeast Brazil is associated with subtherapeutic plasma levels of antiretroviral medications and with both bovine and human subtypes of Cryptosporidium parvum

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    Richard K. Brantley

    2003-02-01

    Full Text Available Advanced HIV infection is frequently complicated by diarrhea, disruption of bowel structure and function, and malnutrition. Resulting malabsorption of or pharmacokinetic changes in antiretroviral agents might lead to subtherapeutic drug dosing and treatment failure in individual patients, and could require dose adjustment and/or dietary supplements during periods of diarrheal illness. We determined the plasma levels of antiretroviral medications in patients that had already been started on medication by their physicians in an urban infectious diseases hospital in northeast Brazil. We also obtained blood samples from patients hospitalized for diarrhea or AIDS-associated wasting, and we found reduced stavudine and didanosine levels in comparison with outpatients without diarrhea or wasting who had been treated at the same hospital clinic. There was a predominance of the protozoal pathogens Cryptosporidium and Isospora belli, typical opportunistic pathogens of AIDS-infected humans, in the stool samples of inpatients with diarrhea. We conclude that severe diarrhea and wasting in this population is associated with both protozoal pathogens and subtherapeutic levels of antiretroviral medications.

  17. COMPARISON OF TISSUE CULTURE AND ANIMAL MODELS FOR ASSESSMENT OF CRYPTOSPRIDIUM PARVUM INFECTION

    Science.gov (United States)

    Data from three different disinfection studies using both cell culture and mouse infectivity to assess Cryptosporidium parvum inactivation were evaluated in a total of 35 comparison including process controls and treated samples. C. parvum infectivity in the in vitro FDM-MPN assa...

  18. Comparação entre os métodos de Ziehl-Neelsen modificado e Acid-Fast-Trichrome para a pesquisa fecal de Cryptosporidium parvum e Isospora belli

    Directory of Open Access Journals (Sweden)

    Rigo Carla R.

    2002-01-01

    Full Text Available Devido a crescente importância dos coccídios intestinais (Cryptosporidium, Isospora e Cyclospora como parasitos oportunistas, é fundamental para os laboratórios diferenciar morfologicamente estes protozoários; a técnica de Ziehl-Neelsen modificada (ZNm é amplamente utilizada para este fim; recentemente, foi proposto um novo procedimento, a coloração combinada do ácido tricrômico (Acid-Fast-Trichrome - AFT. O objetivo do presente estudo foi comparar os processos AFT e ZNm para a detecção destes coccídios em amostras fecais de pacientes portadores do vírus VIH. Foram selecionados dois grupos de indivíduos, para inclusão no estudo, segundo a presença (n=60 ou ausência de diarréia (n=60. As amostras de fezes foram coletadas em solução de formalina 10% e os esfregaços fecais preparados i diretamente das fezes e ii após concentração prévia a 500xg (10 minutos, foram submetidos aos diferentes processos de coloração. Considerando-se a positividade por técnica (AFT e ZNm, verificou-se a superioridade do procedimento de ZNm (n=19; 100% dos casos positivos sobre o de AFT (n=8; 42,1%. Ambos possibilitaram a identificação dos 101 casos verdadeiramente negativos. Coccidiose intestinal foi mais frequente entre os pacientes que apresentaram diarréia (26,6% em comparação à positividade observada entre os indíviduos assintomáticos (5% sendo que C. cayetanensis não foi detectada em ambos os grupos. Foi de nosso interesse avaliar a aplicabilidade da técnica AFT para a coloração deste protozoário. Devido à sensibilidade e especificidade obtida neste estudo (100%, conclui-se que o método de ZNm continua sendo o mais indicado para o diagnóstico da criptosporidiose e isosporose, principalmente quando associado ao procedimento de centrífugo-concentração (500xg, 10 minutos. Embora a coloração AFT tenha baixo custo, faz-se necessário o seu aperfeiçoamento pois este procedimento permite o diagnóstico simultâneo dos

  19. Multilocus genotyping of Cryptosporidium hominis associated with diarrhea outbreak in a day care unit in São Paulo Genotipagem de multilocus de Cryptosporidium hominis associado a surto diarréico em creche de São Paulo

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    Elenice Messias do Nascimento Gonçalves

    2006-04-01

    Full Text Available A number of species of Cryptosporidium are associated with diarrhea worldwide. Little data exists regarding the genotypes and species of Cryptosporidium associated with cases of infections in Brazil. PURPOSE: In the present study, we ascertained by molecular methods the species and the genotype of Cryptosporidium sp from a diarrhea outbreak diagnosed in a day care at the Hospital Clínicas, São Paulo University Medical School. MATERIALS AND METHODS: Specific identification and typing of the isolates associated with the outbreak was done by DNA sequencing analysis of fragments amplified by polymerase chain reaction (PCR from 3 different Cryptosporidium loci: the SSUrRNA coding region, the Cryptosporidium oocyst wall protein (COWP gene, and the microsatellite locus 1 (ML1, a tandem GAG-trinucleotide repeat containing substitutions that differentiate the genotypes of Cryptosporidium parvum and Cryptosporidium hominis. RESULTS: A total of 29 positive samples from the outbreak were studied by the molecular methods described. Our study revealed the presence of a single genotype of Cryptosporidium hominis in all samples. CONCLUSION: The molecular analysis reinforced the hypothesis that the transmission of Cryptosporidium hominis during the period the samples were collected occurred in an outbreak pattern, possibly by person-to-person contact through the fecal-oral route. As far as we know, this is the first time that molecular tools have been used to identify the species and the genotype of isolates showing the presence of the ML1 genotype in samples from Brazilian patients.Mundialmente, diferentes espécies de Cryptosporidium estão relacionadas com doenças diarréicas. No Brasil há poucos dados sobre os genótipos das espécies de Cryptosporidium associadas a infecções. OBJETIVO: No presente estudo, caracterizamos, por métodos moleculares, a espécie e o genótipo de Cryptosporidium sp diagnosticado em surto diarréico ocorrido na creche do

  20. Molecular characterization of Cryptosporidium and Giardia occurring in natural water bodies in Poland.

    Science.gov (United States)

    Adamska, Małgorzata

    2015-02-01

    Cryptosporidium and Giardia protozoa are zoonotic parasites that cause human gastroenteritis and can be transmitted to human through the fecal-oral route and water or food. Several species belong to these genera and their resistant forms occur in water, but only some of them are infectious to human. Health risk depends on the occurrence of infectious Cryptosporidium and Giardia species and genotypes in water, and only molecular techniques allow detecting them, as well as enable to identify the contamination source. In this work, genotyping and phylogenetic analysis have been performed on the basis of 18S rDNA and ß-giardin genes sequences of Cryptosporidium and Giardia, respectively, in order to provide the molecular characterization of these parasites detected earlier in five natural water bodies in Poland and to track possible sources of their (oo)cysts in water. Genotyping revealed a high similarity (over 99 up to 100 %) of analyzed sequences to cattle genotype of C. parvum isolated from cattle and human and to G. intestinalis assemblage B isolated from human. The sequences obtained by others originated from patients with clinical symptoms of cryptosporidiosis or giardiasis and/or with the infection confirmed by different methods. The contamination of three examined lakes is probably human-originated, while the sources of contamination of two remaining lakes are wild and domestic animals. Obtained phylogenetic trees support suggestions of other authors that the bovine genotype of C. parvum should be a separate species, as well as A and B assemblages of G. intestinalis.

  1. Small rodents as reservoirs of Cryptosporidium spp. and Giardia spp. in south-western Poland

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    Agnieszka Perec-Matysiak

    2015-02-01

    Full Text Available [i]Cryptosporidium[/i] spp. and [i]Giardia[/i] spp. have been detected in a range of host species, including rodents. The aim of this study was to determine the distribution of these pathogens and recognition of the reservoir role of rodents in the maintenance of these pathogens in south-western Poland. Additionally, preliminary molecular studies were conducted to elucidate the species and genotypes of [i]Cryptosporidium[/i] and [i]Giardia[/i] identified in this study. Stool samples (n=266 from [i]A. agrarius[/i],[i] A. flavicollis[/i] and [i]M. glareolus[/i], were subjected for analyses. Values of prevalence were 61.7, 68.3 and 68.1%, respectively, for [i]Cryptosporidium[/i] spp. and 41.7, 24.4 and 38.4%, respectively, for [i]Giardia[/i] spp. There was a statistically significant correlation between host species and [i]Giardia[/i] infection where[i] A. agrarius[/i] was the species of the highest prevalence. Statistically significant differences were not found for comparisons made for study sites and occurrence of [i]Giardia[/i] spp. and [i]Cryptosporidium[/i] spp. Due to preliminary nested PCR results, specific amplifications of [i]Cryptosporidium[/i] COWP and SSU rRNA genes were obtained for several isolates taken from rodent host species. One isolate recovered from [i]A. agrarius[/i] (from a semi-aquatic, urban area was identified as [i]C. parvum[/i] and revealed 100% similarity with sequences obtained from humans. To the best of the knowledge of the authors, this is the first record of the [i]C. parvum[/i] zoonotic species from the striped field mouse. Also recorded were the first findings of [i]C. ubiquitum[/i] from three small rodent species.

  2. Ureaplasma parvum undergoes selection in utero resulting in genetically diverse isolates colonizing the chorioamnion of fetal sheep.

    Science.gov (United States)

    Dando, Samantha J; Nitsos, Ilias; Polglase, Graeme R; Newnham, John P; Jobe, Alan H; Knox, Christine L

    2014-02-01

    Ureaplasmas are the microorganisms most frequently isolated from the amniotic fluid of pregnant women and can cause chronic intrauterine infections. These tiny bacteria are thought to undergo rapid evolution and exhibit a hypermutatable phenotype; however, little is known about how ureaplasmas respond to selective pressures in utero. Using an ovine model of chronic intraamniotic infection, we investigated if exposure of ureaplasmas to subinhibitory concentrations of erythromycin could induce phenotypic or genetic indicators of macrolide resistance. At 55 days gestation, 12 pregnant ewes received an intraamniotic injection of a nonclonal, clinical Ureaplasma parvum strain followed by (i) erythromycin treatment (intramuscularly, 30 mg/kg/day, n = 6) or (ii) saline (intramuscularly, n = 6) at 100 days gestation. Fetuses were then delivered surgically at 125 days gestation. Despite injecting the same inoculum into all the ewes, significant differences between amniotic fluid and chorioamnion ureaplasmas were detected following chronic intraamniotic infection. Numerous polymorphisms were observed in domain V of the 23S rRNA gene of ureaplasmas isolated from the chorioamnion (but not the amniotic fluid), resulting in a mosaiclike sequence. Chorioamnion isolates also harbored the macrolide resistance genes erm(B) and msr(D) and were associated with variable roxithromycin minimum inhibitory concentrations. Remarkably, this variability occurred independently of exposure of ureaplasmas to erythromycin, suggesting that low-level erythromycin exposure does not induce ureaplasmal macrolide resistance in utero. Rather, the significant differences observed between amniotic fluid and chorioamnion ureaplasmas suggest that different anatomical sites may select for ureaplasma subtypes within nonclonal, clinical strains. This may have implications for the treatment of intrauterine ureaplasma infections.

  3. Prevalence and genetic characterization of Cryptosporidium spp. and Cystoisospora belli in HIV-infected patients

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    Dnieber Chagas Assis

    2013-06-01

    Full Text Available Cryptosporidium spp. and Cystoisospora belli are monoxenic protozoa that have been recognized as the causative agents of chronic diarrhea in immunocompromised individuals, especially HIV-infected subjects. The objective of this study was to evaluate the frequency of these intestinal protozoa in HIV-positive patients in the Triângulo Mineiro region of Brazil and to correlate the presence of these infections with clinical, epidemiological and laboratory data of the patients. Oocysts were detected in stool samples of 10 (16.9% of the 59 patients studied, while Cryptosporidium spp. were present in 10.1% (6/59 and C. belli in 6.7% (4/59. The frequency of these parasites was higher among patients with diarrheic syndrome and CD4+ T lymphocyte counts < 200 cells/mm 3 , demonstrating the opportunistic characteristic of these infections. A significant association was observed between the lack of adherence to antiretroviral therapy and the presence of Cryptosporidium spp. and/or C. belli. Parasitism with Cryptosporidium spp. was more frequent in February and April, the months following the period of high rainfall. The same was not observed for C. belli. Genetic characterization of two isolates led to the identification of Cryptosporidium parvum, one of the main species associated with the zoonotic transmission of cryptosporidiosis.

  4. Prevalence and Genetic Characterization of Cryptosporidium Spp. In Diarrheic Children from Gonbad Kavoos City, Iran

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    Mitra SHARBATKHORI

    2015-10-01

    Full Text Available Background: Cryptosporidium is an intestinal protozean parasite causing water­borne and foodborne outbreaks of diarrheal diseases. The present study was per­formed in order to find prevalence and subtypes of Cryptosporidium among children with diarrhea in Gonbad Kavoos City, Northern Iran.Methods: Diarrheic samples were collected from 547 children. The initial parasitologi­cal diagnosis was made based on detection of oocysts using the modi­fied Ziehl-Neelsen acid-fast staining method. The positive microscopically samples were selected for sequence analysis of partial 60 kDa glycoprotein (gp60 gene. Results: Out of 547 collected samples, 27 (4.94% were positive for Cryptosporid­ium oocysts. Fifteen from 27 positive samples successfully amplified in PCR. Se­quences analysis of gp60 gene in 15 Cryptosporidium isolates revealed that all of them (100% were C. parvum. The results showed three subtypes of IIa subtype family (7 cases including IIaA16G2R1, IIaA17G1R1, IIaA22G3R1 and one subtype of IId subtype family (8 cases. The most common allele was IId A17G1d (53.3%. Conclusion: The predominance of zoonotic subtype families of C. parvum species (IIa, IId in the present study is in concordance with previous studies in Iran and emphasizes the significance of zoonotic transmission of cryptosporidiosis in the country.

  5. MALDI-MIS INVESTIGATIONS OF DRINKING WATER PATHOGENS--GIARDIA AND CRYPTOSPORIDIUM

    Science.gov (United States)

    The protozoan parasites, Cryptosporidium parvum and Giardia lamblia, have been responsible for numerous waterborne outbreaks of gastrointestinal illness in the United States. The 1993 cryptosporidiosis outbreak in Milwaukee affected approximately 400,000 people and resulted in o...

  6. Literature Reference for Cryptosporidium spp. (Applied and Environmental Microbiology. 1999. 65(9): 3936–3941)

    Science.gov (United States)

    Procedures are described for analysis of animal samples using tissue culture techniques that may be adapted for assessment of solid, particulate, liquid and water samples contaminated with Cryptosporidium parvum.

  7. SPECIES AND GENUS DIFFERENTIATION OF PARASITES (GIARDIA AND CRYPTOSPORIDIUM) BY MALDI - MASS SPECTROMETRY

    Science.gov (United States)

    The protozoan parasites, Cryptosporidium parvum and Giardia lamblia, have been responsible for numerous waterborne outbreaks of gastrointestinal illness in the United States. The 1993 cryptosporidiosis outbreak in Milwaukee affected approximately 400,000 people and resulted in o...

  8. MALDI-MIS INVESTIGATIONS OF DRINKING WATER PATHOGENS--GIARDIA AND CRYPTOSPORIDIUM

    Science.gov (United States)

    The protozoan parasites, Cryptosporidium parvum and Giardia lamblia, have been responsible for numerous waterborne outbreaks of gastrointestinal illness in the United States. The 1993 cryptosporidiosis outbreak in Milwaukee affected approximately 400,000 people and resulted in o...

  9. Prevalence and molecular identification of Cryptosporidium isolates from pet lizards and snakes in Italy

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    Rinaldi L.

    2012-11-01

    Full Text Available In order to acquire prevalence and genetic data on Cryptosporidium infections in captive lizards and snakes kept as pets, a survey was conducted on 150 individual reptiles from southern Italy. Fecal samples were preserved in 5% formalin and analyzed using a commercial immunofluorescence assay (IFA for the detection of Cryptosporidium oocysts and Giardia cysts. IFA revealed the presence of Cryptosporidium oocysts in nine of the 150 samples examined (6.0%, precisely in 6/125 snakes (4.8% and in 3/25 lizards (12.0%; all fecal samples tested negative for the presence of Giardia cysts. Molecular characterization based on nested PCR amplification and sequencing of the SSU-rRNA gene, revealed the presence of Cryptosporidium serpentis in three samples from snakes (Boa constrictor constrictor, Elapheguttata guttata guttata and Python molurus.

  10. Isolation and enumeration of Giardia cysts, cryptosporidium oocysts, and Ascaris eggs from fruits and vegetables.

    Science.gov (United States)

    Robertson, L J; Gjerde, B

    2000-06-01

    Published techniques for recovering parasites from fruit and vegetables are generally inadequate, with low and variable recovery efficiencies. Here we describe an improved methodology for analyzing fruit and vegetables for Giardia cysts, Cryptosporidium oocysts, and Ascaris eggs. The method includes washing procedures, sonication, and, for Giardia and Cryptosporidium, immunomagnetic separation. Identification is by immunofluorescence (Giardia and Cryptosporidium) or brightfield microscopy (Ascaris). Recovery efficiencies from lettuce, Chinese leaves, and strawberries were found to be approximately 67% for Giardia, 42% for Cryptosporidium, and 72% for Ascaris. Recovery efficiencies from bean sprouts tended to be more variable and lower. This could be due to material removed with the parasites during the washing procedures, which, in turn, appeared related to the age of the bean sprouts. It is therefore recommended that fruit and vegetables should be as fresh as possible when analyzed for parasites.

  11. Presence of Cryptosporidium spp. and Giardia duodenalis through drinking water.

    Science.gov (United States)

    Castro-Hermida, José Antonio; García-Presedo, Ignacio; Almeida, André; González-Warleta, Marta; Correia Da Costa, José Manuel; Mezo, Mercedes

    2008-11-01

    To evaluate the presence of Cryptosporidium spp. and Giardia duodenalis in the influent and final effluent of sixteen drinking water treatment plants located in a hydrographic basin in Galicia (NW Spain) - in which the principal river is recognised as a Site of Community Importance (SCI) - estimate the efficiency of treatment plants in removing these protozoans and determine the species and genotypes of the parasites by means of a molecular assay. All plant samples of influent and final effluent (50-100 l) were examined in the spring, summer, autumn and winter of 2007. A total of 128 samples were analysed by method 1623, developed by US Environmental Protection Agency for isolation and detection of both parasites. To identify the genotypes present the following genes were amplified and sequenced: 18S SSU rRNA (Cryptosporidium spp.) and b-giardina (G. duodenalis). The mean concentrations of parasites in the influent were 0.0-10.5 Cryptosporidium spp. oocysts per litre and 1.0-12.8 of G. duodenalis cysts per litre. In the final treated effluent, the mean concentration of parasites ranged from 0.0-3.0 oocysts per litre and 0.5-4.0 cysts per litre. The distribution of results by season revealed that in all plants, the highest numbers of (oo)cysts were recorded in spring and summer. Cryptosporidium parvum, C. andersoni, C. hominis and assemblages A-I, A-II, E of G. duodenalis were detected. Cryptosporidium spp. and G. duodenalis were consistently found at high concentrations in drinking water destined for human and animal consumption in the hydrographic basin under study, in Galicia (NW Spain). It is important that drinking water treatment authorities rethink the relevance of contamination levels of both parasites in drinking water and develop adequate countermeasures.

  12. Elevation and vegetation determine Cryptosporidium oocyst shedding by yellow-bellied marmots (Marmota flaviventris in the Sierra Nevada Mountains

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    Diego Montecino-Latorre

    2015-08-01

    Full Text Available Wildlife are increasingly recognized as important biological reservoirs of zoonotic species of Cryptosporidium that might contaminate water and cause human exposure to this protozoal parasite. The habitat range of the yellow-bellied marmot (Marmota flaviventris overlaps extensively with the watershed boundaries of municipal water supplies for California communities along the foothills of the Sierra Nevada. We conducted a cross-sectional epidemiological study to estimate the fecal shedding of Cryptosporidium oocysts by yellow-bellied marmots and to quantify the environmental loading rate and determine risk factors for Cryptosporidium fecal shedding in this montane wildlife species. The observed proportion of Cryptosporidium positive fecal samples was 14.7% (33/224, positive number relative to total number samples and the environmental loading rate was estimated to be 10,693 oocysts animal-1 day-1. Fecal shedding was associated with the elevation and vegetation status of their habitat. Based on a portion of the 18s rRNA gene sequence of 2 isolates, the Cryptosporidium found in Marmota flaviventris were 99.88%–100% match to multiple isolates of C. parvum in the GenBank.

  13. Flow-through immunomagnetic separation system for waterborne pathogen isolation and detection: Application to Giardia and Cryptosporidium cell isolation

    Energy Technology Data Exchange (ETDEWEB)

    Ramadan, Qasem, E-mail: qasem.alramadan@epfl.ch [Bioelectronics Program, Institute of Microelectronics, 11 Science Park Road, Singapore 117685 (Singapore); Christophe, Lay; Teo, William; ShuJun, Li; Hua, Feng Han [Bioelectronics Program, Institute of Microelectronics, 11 Science Park Road, Singapore 117685 (Singapore)

    2010-07-12

    Simultaneous sample washing and concentration of two waterborne pathogen samples were demonstrated using a rotational magnetic system under continuous flow conditions. The rotation of periodically arranged small permanent magnets close to a fluidic channel carrying magnetic particle suspension allows the trapping and release of particles along the fluidic channel in a periodic manner. Each trapping and release event resembles one washing cycle. The performance of the magnetic separation system (MSS) was evaluated in order to test its functionality to isolate magnetic-labelled protozoan cells from filtered, concentrated tap water, secondary effluent water, and purified water. Experimental protocols described in US Environmental Protection Agency method 1623 which rely on the use of a magnetic particle concentrator, were applied to test and compare our continuous flow cell separation system to the standard magnetic bead-based isolation instruments. The recovery efficiencies for Giardia cysts using the magnetic tube holder and our magnetic separation system were 90.5% and 90.1%, respectively, from a tap water matrix and about 31% and 18.5%, respectively, from a spiked secondary effluent matrix. The recovery efficiencies for Cryptosporidium cells using the magnetic tube holder and our magnetic separation system were 90% and 83.3%, respectively, from a tap water matrix and about 38% and 36%, respectively, from a spiked secondary effluent matrix. Recoveries from all matrices with the continuous flow system were typically higher in glass tubing conduits than in molded plastic conduits.

  14. Cryptosporidium proliferans n. sp. (Apicomplexa: Cryptosporidiidae: Molecular and Biological Evidence of Cryptic Species within Gastric Cryptosporidium of Mammals.

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    Martin Kváč

    Full Text Available 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

  15. Genomic Variation in IbA10G2 and Other Patient-Derived Cryptosporidium hominis Subtypes.

    Science.gov (United States)

    Sikora, Per; Andersson, Sofia; Winiecka-Krusnell, Jadwiga; Hallström, Björn; Alsmark, Cecilia; Troell, Karin; Beser, Jessica; Arrighi, Romanico B G

    2017-03-01

    In order to improve genotyping and epidemiological analysis of Cryptosporidium spp., genomic data need to be generated directly from a broad range of clinical specimens. Utilizing a robust method that we developed for the purification and generation of amplified target DNA, we present its application for the successful isolation and whole-genome sequencing of 14 different Cryptosporidium hominis patient specimens. Six isolates of subtype IbA10G2 were analyzed together with a single representative each of 8 other subtypes: IaA20R3, IaA23R3, IbA9G3, IbA13G3, IdA14, IeA11G3T3, IfA12G1, and IkA18G1. Parasite burden was measured over a range of more than 2 orders of magnitude for all samples, while the genomes were sequenced to mean depths of between 17× and 490× coverage. Sequence homology-based functional annotation identified several genes of interest, including the gene encoding Cryptosporidium oocyst wall protein 9 (COWP9), which presented a predicted loss-of-function mutation in all the sequence subtypes, except for that seen with IbA10G2, which has a sequence identical to the Cryptosporidium parvum reference Iowa II sequence. Furthermore, phylogenetic analysis showed that all the IbA10G2 genomes form a monophyletic clade in the C. hominis tree as expected and yet display some heterogeneity within the IbA10G2 subtype. The current report validates the aforementioned method for isolating and sequencing Cryptosporidium directly from clinical stool samples. In addition, the analysis demonstrates the potential in mining data generated from sequencing multiple whole genomes of Cryptosporidium from human fecal samples, while alluding to the potential for a higher degree of genotyping within Cryptosporidium epidemiology.

  16. AGING OF CRYPTOSPORIDIUM PARVUUM OOCYSTS STUDIED BY MALDI-TOF MS

    Science.gov (United States)

    Cryptosporidium parvum is a protozoan parasite, and it causes a potentially fatal gastrointestinal illness. This water borne pathogen has been the subject of several high profile disease outbreaks in the US and abroad. C. parvum presents challenges for both compliance monitorin...

  17. Multilocus typing and population structure of Cryptosporidium from children in Zaragoza, Spain.

    Science.gov (United States)

    Ramo, Ana; Quílez, Joaquín; Vergara-Castiblanco, Claudia; Monteagudo, Luis; Del Cacho, Emilio; Clavel, Antonio

    2015-04-01

    A multilocus typing approach with eight variable-number tandem-repeat (VNTR) loci and the GP60 gene was used to analyze the inter- and intra-species variation of 44 Cryptosporidium isolates from pediatric patients in Zaragoza city (NE, Spain). Restriction and sequence analyses of the SSU rRNA gene revealed that Cryptosporidium transmission is mostly anthroponotic in this area, with the predominance of Cryptosporidium hominis (n: 41) over Cryptosporidium parvum (n: 3). GP60 subtyping showed limited genetic diversity and four subtypes were identified, including IbA10G2 (n: 35), IaA24R3 (n: 6), IIaA15G1R1 (n: 1) and IIaA15G2R1 (n: 2). Five out of eight VNTR loci showed a discriminatory power higher than the GP60 gene, although each locus had a predominant allele exhibited by more than 50% of isolates. All but four alleles were associated to either C. hominis or C. parvum and linked alleles at different loci were found. Multilocus typing substantially increased the discriminatory power (Hunter-Gaston index: 0.807, 95% CI, 0.683-0.926) and revealed that genetic diversity is much higher than that reported by GP60 sequencing, since 17 multilocus subtypes (MLTs) were identified. Nearly half of the specimens were allocated to a single major MLT. However, no more than three specimens were allocated to each of the remaining MLTs. Both phylogenetic and population analyses revealed a population clustering of C. hominis according to the GP60 subtype, which indicates the robustness of this marker to differentiate genetic subpopulations. Subpopulations had an overall clonal genetic structure, although traces of genetic flow between them were also observed.

  18. Cryptosporidium enteritis

    Science.gov (United States)

    ... page: //medlineplus.gov/ency/article/000617.htm Cryptosporidium enteritis To use the sharing features on this page, please enable JavaScript. Cryptosporidium enteritis is an infection of the small intestine that ...

  19. DETECTION OF PBMC T CELL SUBSETS AND mIL-2R OF THE PERSONS INFECTED BY CRYPTOSPORIDIUM PARVUM BY USING THE SYSTEM OF BSA%应用BSA系统检测隐孢子虫感染者外周血T细胞亚群及mIL-2R

    Institute of Scientific and Technical Information of China (English)

    许礼发; 李朝品; 张荣波; 王晓秋

    2004-01-01

    Objective To explore the changes of cellular immune function in the persons infected by Cryptosporidium parvum .MethodsThe system of biotin streptavidin (BSA) was used to detect T cell subsets and membrane interleukin 2 receptor (mIL 2R) in peripheral blood mononuclear cell (PBMC) of the persons infected byC. parvum . ResultsThe positive rates of CD3+,CD4+,CD8+ and CD4+/CD8+ in the persons in whose stoolsC. parvum oocysts were found were (54.77±7.33)%,(38.17±7.52)%,(30.64±5.87)% and 1.28±0.72 respectively,and the level of mIL 2R in induction period was (32.15±2.42)%. It could be concluded that the positive rates of CD3+,CD4+,CD8+ and CD4+/CD8+ in the persons in whose stoolsC. parvum oocysts were found were significantly different from those with noC. parvum oocysts were found (P <0.05~0.01). The level of mIL 2R in induction period was significantly different from which in resting period (P <0.05~0.01).ConclusionIn the persons infected byC. parvum,cellular immunity participates in resisting the parasite,and T cell subsets and mIL 2R play an important role.%目的探讨隐孢子虫感染者机体细胞免疫功能的变化. 方法采用生物素-链霉亲和素(Biotin-Streptavidin , BSA)法对卵囊阳性患者分别进行外周血T细胞亚群、膜白介素-2受体(mIL-2R)检测. 结果隐孢子虫卵囊阳性者的CD3+、CD4+、CD8+百分率和CD4+/CD8+比值分别为(54.77±7.33)%、(38.17±7.52)%、(30.64±5.87)% 和1.28±0.72,诱导期mIL-2R表达水平为(32.15±2.42)%,两者分别与卵囊阴性者及静息期mIL-2R相比,差异有显著性(P<0.05~0.01). 结论隐孢子虫感染者引起细胞免疫功能的变化,T细胞亚群、mIL-2R在抗隐孢子虫感染中起重要作用.

  20. Molecular characterization of Cryptosporidium spp. from HIV infected patients from an urban area of Brazil Caracterização molecular de Cryptosporidium spp. de pacientes de área urbana do Brasil infectados por HIV

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    Patrícia de Lucca

    2009-12-01

    Full Text Available Cryptosporidium spp. are important cause of enteric disease in humans, but may also infect animals. This study describes the relative frequency of several Cryptosporidium species found in human specimens from HIV infected patients in the São Paulo municipality obtained from January to July 2007. Sequence analysis of the products of nested-PCR based on small subunit rRNA and Cryptosporidium oocyst wall protein coding genes revealed 17 (63.0% isolates of C. hominis, four (14.8% C. parvum, five (18.5% C. felis and one (3.7% C. canis. These findings suggest that, in urban environments of Brazil, the cat adapted C. felis may play a potential role in the zoonotic transmission of cryptosporidiosis whereas the anthroponotic transmission of cryptosporidiosis caused by C. hominis seems to predominate.Cryptosporidium spp. são importantes causas de doenças entéricas em humanos, mas podem também ser encontrados em animais. O presente estudo descreve a frequência relativa de diversas espécies de Cryptosporidium em amostras de humanos da cidade de São Paulo, Brasil, obtidas de janeiro a julho de 2007. Análises de sequências de produtos de nested PCR direcionadas ao genes codificadores da menor unidade ribosomal e da proteina de parede de oocistos revelaram 17 (63,0% isolados de C. hominis, quatro (14,8% C. parvum, cinco (18,5% C. felis, e um (3,7% C. canis. Estes resultados sugerem que, em ambientes urbanos no Brasil, o genótipo adaptado ao gato pode desempenhar potencial papel na transmissão zoonótica de criptosporidiose, enquanto a transmissão antroponótica da criptosporidiose causada pelo C. hominis parece predominar.

  1. Detection and molecular characterization of Giardia and Cryptosporidium in common dolphins (Delphinus delphis) stranded along the Galician coast (Northwest Spain).

    Science.gov (United States)

    Reboredo-Fernández, A; Gómez-Couso, H; Martínez-Cedeira, J A; Cacciò, S M; Ares-Mazás, E

    2014-05-28

    The ubiquitous protozoan parasites Giardia and Cryptosporidium have been detected from many species of captive and free-living wildlife, representing most mammalian orders. Twenty species of marine mammals have been reported to inhabit Galician waters and the region has one of the highest rates of stranding in Europe. Evidence from stranding, reported by-catches and sightings, suggests that the common dolphin (Delphinus delphis) is the most abundant cetacean on the Galician coast (Northwest Spain). The objective of this study was to detect and molecularly characterize isolates of Giardia and Cryptosporidium obtained from common dolphins stranded in this area. Between 2005 and 2012, sections of large intestine from 133 common dolphins stranded along the Galician coast were collected by the personnel of the Galician Stranding Network (Coordinadora para o Estudo dos Mamíferos Mariños, CEMMA). Using direct immunofluorescence antibody test (IFAT) and PCR amplification and sequencing of the SSU-rDNA, β-giardin genes and the ITS1-5.8S-ITS2 region, Giardia and Cryptosporidium were detected in 8 (6.0%) and 12 samples (9.0%), respectively. In two samples, co-infection by both parasites was observed. The molecular characterization revealed the presence of Giardia duodenalis assemblages A (genotypes A1 and A2) and B and Cryptosporidium parvum in these samples. This constitutes the first study in which the presence of Giardia and Cryptosporidium has been investigated in common dolphins on the European Atlantic coast, and it is also the first report of C. parvum in this host. Our findings indicate that these animals could act as reservoir of these waterborne parasites or could be victims of the contamination originated by anthropogenic activities.

  2. Molecular detection and characterization of Cryptosporidium species in household dogs, pet shop puppies, and dogs kept in a school of veterinary nursing in Japan.

    Science.gov (United States)

    Itoh, Naoyuki; Oohashi, Yoshino; Ichikawa-Seki, Madoka; Itagaki, Tadashi; Ito, Yoichi; Saeki, Hideharu; Kanai, Kazutaka; Chikazawa, Seishiro; Hori, Yasutomo; Hoshi, Fumio; Higuchi, Seiichi

    2014-03-01

    Members of Cryptosporidium species, which are protozoan parasites, are prevalent worldwide and can cause diarrhoea in both humans and animals, including dogs. In addition, the Cryptosporidium species harboured in dogs have the potential for zoonotic transmission. The purpose of the present study was to determine the prevalence of Cryptosporidium species infection and perform molecular characterization of isolates in household dogs, pet shop puppies, and dogs kept in a school of veterinary nursing in Japan. Fresh faecal samples were collected once from 529 household dogs (aged from 2 months to 18 years old, from 9 veterinary clinics located in 6 different regions), 471 pet shop puppies (≤ 3 months old, from 4 pet shops located in 2 different regions), and 98 dogs (aged from 2 to 11 years old) kept in a veterinary nursing school. A nested polymerase chain reaction (PCR) assay targeting the 18S rRNA gene was employed for the detection of Cryptosporidium species, and 111 random samples of PCR amplicons (approximately 500-bp) were sequenced for the molecular characterization of the isolates. The prevalences of Cryptosporidium species in household dogs, pet shop puppies, and veterinary nursing school dogs were 7.2%, 31.6%, and 18.4%, respectively. In household dogs, no significant correlation was observed between the prevalence of Cryptosporidium species and the age (≤ 6 months vs. >6 months), living conditions (indoor vs. outdoor), faecal conditions (formed vs. unformed), and location of residence. In pet shop puppies, the prevalence of Cryptosporidium species was not related to faecal condition; however, the prevalence significantly differed among the pet shops. All of the 111 sequence samples (26 from household dogs, 75 from pet shop puppies, and 10 from veterinary nursing school dogs) were identified as Cryptosporidium canis. The present study demonstrates a high prevalence of Cryptosporidium species infections in pet shop puppies and dogs of a veterinary nursing

  3. Molecular characterization of Cryptosporidium spp. in pre-weaned kids in a dairy goat farm in western France.

    Science.gov (United States)

    Rieux, Anais; Paraud, Carine; Pors, Isabelle; Chartier, Christophe

    2013-02-18

    A longitudinal study was undertaken to characterize the course of Cryptosporidium infection in a dairy goat farm located in western France. Two cohorts of twenty-five and fifteen animals, respectively, were sampled once a week from birth to weaning. Each individual fecal sample was screened using direct immunofluorescence (IFT) and if found positive, the Cryptosporidium species was identified using PCR analysis. Seventeen (68% [95% CI: 44-91]) animals were positive at least once during the first study and 14 (93% [95% CI: 80-100]) during the second, after IFT examination. In the first study, the age at first excretion was 17 days and the peak of excretion (mean arithmetic excretion: 22,700 oocysts per gram (opg) of feces) was recorded when kids were between 22 and 28 days old. For the second study, the age at first excretion was 10 days and the peak of excretion (mean arithmetic excretion: 3.4 × 10(6)opg) was recorded in animals aged between 10 and 14 days. Clinical signs were observed only in animals of the second cohort. DNA sequence analysis at the 18S ribosomal RNA locus was successful for 9 of the 27 IFT-positive samples in the first cohort and for 10 of the 34 positive isolates in the second cohort. All isolates were identified as Cryptosporidium xiaoi except one which was identified as Cryptosporidium parvum. Our results confirm that goat kids are hosts for C. parvum and C. xiaoi and that infection by C. xiaoi may be associated with mild clinical signs. Copyright © 2012 Elsevier B.V. All rights reserved.

  4. Identification and genotyping of Giardia spp. and Cryptosporidium spp. isolates in aquatic birds in the Salburua wetlands, Álava, Northern Spain.

    Science.gov (United States)

    Cano, Lourdes; de Lucio, Aida; Bailo, Begoña; Cardona, Guillermo A; Muadica, Aly Salimo Omar; Lobo, Luis; Carmena, David

    2016-05-15

    Aquatic birds are known to be suitable hosts for a number of avian-specific species and genotypes of the enteric protozoan parasites Giardia and Cryptosporidium. Waterbirds have also been reported as sporadic carriers of species of both pathogens from human or domestic animal origin via environmental contamination. Because aquatic birds can shed substantial amounts of infective Giardia and Cryptosporidium (oo)cysts to the environment including surface waters intended for human consumption, this situation may pose a potential risk of waterborne zoonotic disease. A total of 265 waterbird faecal samples were collected from May 2014 to June 2015 at Salburua (Álava), one of the most valued continental wetlands in northern Spain. The detection of Giardia oocyst and Cryptosporidium oocysts was carried out by direct fluorescence microscopy and molecular (PCR and sequence analysis) methods targeting the small subunit ribosomal RNA gene of the parasites. Typing of Giardia duodenalis isolates at the sub-assemblage level was based on the specific amplification and sequencing of a partial fragment of the glutamate dehydrogenase gene. Overall, Giardia cysts and Cryptosporidium oocysts were detected in 22 (8.3%) and 6 (2.3%), respectively, of the 265 faecal samples analysed. The two only Giardia isolates characterized (one novel, one known) were assigned to the sub-assemblage BIV of G. duodenalis, none of them previously reported in Spanish human isolates. This finding raises doubts about the actual origin of the infection and whether waterbirds may serve as potential source of infective Giardia cysts to humans via waterborne transmission or through direct contact. The six Cryptosporidium isolates obtained were characterized as avian genotype III (n=4), duck genotype b (n=1), and goose genotype Id (n=1), all considered avian-specific and therefore of negligible risk of zoonotic infection.

  5. Cryptosporidium and Giardia removal by secondary and tertiary wastewater treatment.

    Science.gov (United States)

    Taran-Benshoshan, Marina; Ofer, Naomi; Dalit, Vaizel-Ohayon; Aharoni, Avi; Revhun, Menahem; Nitzan, Yeshayahu; Nasser, Abidelfatah M

    2015-01-01

    Wastewater disposal may be a source of environmental contamination by Cryptosporidium and Giardia. This study was conducted to evaluate the prevalence of Cryptosporidium oocysts and Giardia cysts in raw and treated wastewater effluents. A prevalence of 100% was demonstrated for Giardia cysts in raw wastewater, at a concentration range of 10 to 12,225 cysts L(-1), whereas the concentration of Cryptosporidium oocysts in raw wastewater was 4 to 125 oocysts L(-1). The removal of Giardia cysts by secondary and tertiary treatment processes was greater than those observed for Cryptosporidium oocysts and turbidity. Cryptosporidium and Giardia were present in 68.5% and 76% of the tertiary effluent samples, respectively, at an average concentration of 0.93 cysts L(-1) and 9.94 oocysts L(-1). A higher detection limit of Cryptosporidium oocysts in wastewater was observed for nested PCR as compared to immune fluorescent assay (IFA). C. hominis was found to be the dominant genotype in wastewater effluents followed by C. parvum and C. andersoni or C. muris. Giardia was more prevalent than Cryptosporidium in the studied community and treatment processes were more efficient for the removal of Giardia than Cryptosporidium. Zoonotic genotypes of Cryptosporidium were also present in the human community. To assess the public health significance of Cryptosporidium oocysts present in tertiary effluent, viability (infectivity) needs to be assessed.

  6. Cryptosporidium infection in infancy as a cause of malnutrition

    DEFF Research Database (Denmark)

    Mølbak, Kare; Andersen, M; Aaby, Peter

    1997-01-01

    Cryptosporidium parvum causes persistent diarrhea in young children in developing countries. To determine the interaction between nutritional status and cryptosporidiosis, an open cohort of 1064 children younger than 3 y of age was followed for 1441 child-years by weekly diarrhea recall visits. A...

  7. Cryptosporidium and Giardia: new challenges to the water industry

    NARCIS (Netherlands)

    Medema, Gerriet Jan

    2001-01-01

    The protozoan parasites Cryptosporidium parvum and Giardia intestinalis have emerged as significant waterborne pathogens over the past decades. Many outbreaks of waterborne cryptosporidiosis and giardiasis have been recorded,primarily in the United States and the United Kingdom.Chapter 1 gives an ov

  8. Cryptosporidium sp. infections in green turtles, Chelonia mydas, as a potential source of marine waterborne oocysts in the Hawaiian Islands

    Science.gov (United States)

    Graczyk, T.K.; Balazs, G.H.; Work, T.M.; Aguirre, A.A.; Ellis, D.M.; Murakawa, Shawn K. K.; Morris, Robert

    1997-01-01

    For the first time, Cryptosporidium sp. oocysts were identified in fecal and intestinal samples from free-ranging marine turtles, Chelonia mydas, from the Hawaiian Islands. The oocysts produced positive reactions with commercial test kits recommended for the detection of human-infectious waterborne oocysts of Cryptosporidium parvum.

  9. Comparação entre os métodos de Ziehl-Neelsen modificado e Acid-Fast-Trichrome para a pesquisa fecal de Cryptosporidium parvum e Isospora belli

    OpenAIRE

    Rigo,Carla R.; Franco,Regina Maura B.

    2002-01-01

    Devido a crescente importância dos coccídios intestinais (Cryptosporidium, Isospora e Cyclospora) como parasitos oportunistas, é fundamental para os laboratórios diferenciar morfologicamente estes protozoários; a técnica de Ziehl-Neelsen modificada (ZNm) é amplamente utilizada para este fim; recentemente, foi proposto um novo procedimento, a coloração combinada do ácido tricrômico (Acid-Fast-Trichrome - AFT). O objetivo do presente estudo foi comparar os processos AFT e ZNm para a detecção de...

  10. Molecular detection of Cryptosporidium spp. infections in water buffaloes from northeast Thailand.

    Science.gov (United States)

    Inpankaew, Tawin; Jiyipong, Tawisa; Wongpanit, Kannika; Pinyopanuwat, Nongnuch; Chimnoi, Wissanuwat; Kengradomkij, Chanya; Xuan, Xuenan; Igarashi, Ikuo; Xiao, Lihua; Jittapalapong, Sathaporn

    2014-02-01

    The objectives of this study were to determine the individual and herd-level prevalence and genotype of Cryptosporidium and to identify putative risk factors associated with Cryptosporidium spp. infections in water buffaloes in northeast Thailand. Fecal samples from 600 water buffaloes of 287 farms in six provinces were collected and tested using DMSO-modified acid-fast staining and polymerase chain reaction. The overall prevalence of Cryptosporidium infections in buffaloes was 5.7 and 8.7% among individual animals and herds, respectively. The provinces with highest infected Cryptosporidium were located in the Sakon Nakhon Basin in the northern part of the region. In addition, higher herd prevalence was observed among farms with more than five buffaloes (30%) than those with five or less animals (16.2%). Thirty (88.2%) of the 34 Cryptosporidium-positive samples were Cryptosporidium parvum and four (11.8%) were Cryptosporidium ryanae.

  11. Cryptosporidium infection in children with cancer undergoing chemotherapy: how important is the prevention of opportunistic parasitic infections in patients with malignancies?

    Science.gov (United States)

    Berahmat, Reza; Mahami-Oskouei, Mahmoud; Rezamand, Azim; Spotin, Adel; Aminisani, Nayyereh; Ghoyounchi, Roghayeh; Madadi, Solmaz

    2017-07-20

    Cryptosporidiosis is a relatively uncommon disease in healthy individuals but could be potentially worrisome in immunocompromised patients. This study aimed to evaluate Cryptosporidium infection in children with cancer undergoing chemotherapy. A case-control study was conducted in 132 children with cancer undergoing chemotherapy and 132 non-cancer controls. The modified Ziehl-Neelsen (MZN) staining and polymerase chain reaction methods were used for the detection of Cryptosporidium parasite. All positive isolates were sequenced for phylogenetic analysis. Statistical analysis was performed using the SPSS version 16 and Fisher exact test. The rate of cryptosporidiosis in children with cancer undergoing chemotherapy was 3.8%, which was higher than that of the control group. Other intestinal parasites detected in patients with cancer included Giardia lamblia (3%), Entamoeba coli (1.5%), and Chilomastix mesnili (0.8%). In the control group, only two (1.5%) cases were positive for G. lamblia. No significant difference was observed between the gender, age, residency, contact with domestic animals, stool appearance, neutropenia, chemotherapy period, and type of malignancy with regard to cryptosporidiosis. Phylogenetic analysis revealed that Cryptosporidium parvum isolates in this study relied on a branch that represents similar sequences from Iran and other countries. Although the rate of Cryptosporidium infection was relatively higher in children with cancer undergoing chemotherapy compared to the control group, any statistically significant difference has not been found between them. These findings should not be contrary to the need for healthcare to prevent opportunistic parasitic infections in malignant and immunocompromised patients.

  12. Identification of Cryptosporidium Species in Fish from Lake Geneva (Lac Leman in France.

    Directory of Open Access Journals (Sweden)

    Gabriela Certad

    Full Text Available Cryptosporidium, a protozoan parasite that can cause severe diarrhea in a wide range of vertebrates including humans, is increasingly recognized as a parasite of a diverse range of wildlife species. However, little data are available regarding the identification of Cryptosporidium species and genotypes in wild aquatic environments, and more particularly in edible freshwater fish. To evaluate the prevalence of Cryptosporidiumspp. in fish from Lake Geneva (Lac Léman in France, 41 entire fish and 100 fillets (cuts of fish flesh were collected from fishery suppliers around the lake. Nested PCR using degenerate primers followed by sequence analysis was used. Five fish species were identified as potential hosts of Cryptosporidium: Salvelinus alpinus, Esox lucius, Coregonus lavaretus, Perca fluviatilis, and Rutilus rutilus. The presence of Cryptosporidium spp. was found in 15 out of 41 fish (37%, distributed as follows: 13 (87% C. parvum, 1 (7% C. molnari, and 1 (7% mixed infection (C. parvum and C. molnari. C. molnari was identified in the stomach, while C. parvum was found in the stomach and intestine. C. molnari was also detected in 1 out of 100 analyzed fillets. In order to identify Cryptosporidium subtypes, sequencing of the highly polymorphic 60-kDa glycoprotein (gp60 was performed. Among the C. parvum positive samples, three gp60 subtypes were identified: IIaA15G2R1, IIaA16G2R1, and IIaA17G2R1. Histological examination confirmed the presence of potential developmental stages of C. parvum within digestive epithelial cells. These observations suggest that C. parvum is infecting fish, rather than being passively carried. Since C. parvum is a zoonotic species, fish potentially contaminated by the same subtypes found in terrestrial mammals would be an additional source of infection for humans and animals, and may also contribute to the contamination of the environment with this parasite. Moreover, the risk of human transmission is strengthened by

  13. Identification of Cryptosporidium Species in Fish from Lake Geneva (Lac Léman) in France

    Science.gov (United States)

    Certad, Gabriela; Dupouy-Camet, Jean; Gantois, Nausicaa; Hammouma-Ghelboun, Ourida; Pottier, Muriel; Guyot, Karine; Benamrouz, Sadia; Osman, Marwan; Delaire, Baptiste; Creusy, Colette; Viscogliosi, Eric; Aliouat-Denis, Cecile Marie; Follet, Jérôme

    2015-01-01

    Cryptosporidium, a protozoan parasite that can cause severe diarrhea in a wide range of vertebrates including humans, is increasingly recognized as a parasite of a diverse range of wildlife species. However, little data are available regarding the identification of Cryptosporidium species and genotypes in wild aquatic environments, and more particularly in edible freshwater fish. To evaluate the prevalence of Cryptosporidiumspp. in fish from Lake Geneva (Lac Léman) in France, 41 entire fish and 100 fillets (cuts of fish flesh) were collected from fishery suppliers around the lake. Nested PCR using degenerate primers followed by sequence analysis was used. Five fish species were identified as potential hosts of Cryptosporidium: Salvelinus alpinus, Esox lucius, Coregonus lavaretus, Perca fluviatilis, and Rutilus rutilus. The presence of Cryptosporidium spp. was found in 15 out of 41 fish (37%), distributed as follows: 13 (87%) C. parvum, 1 (7%) C. molnari, and 1 (7%) mixed infection (C. parvum and C. molnari). C. molnari was identified in the stomach, while C. parvum was found in the stomach and intestine. C. molnari was also detected in 1 out of 100 analyzed fillets. In order to identify Cryptosporidium subtypes, sequencing of the highly polymorphic 60-kDa glycoprotein (gp60) was performed. Among the C. parvum positive samples, three gp60 subtypes were identified: IIaA15G2R1, IIaA16G2R1, and IIaA17G2R1. Histological examination confirmed the presence of potential developmental stages of C. parvum within digestive epithelial cells. These observations suggest that C. parvum is infecting fish, rather than being passively carried. Since C. parvum is a zoonotic species, fish potentially contaminated by the same subtypes found in terrestrial mammals would be an additional source of infection for humans and animals, and may also contribute to the contamination of the environment with this parasite. Moreover, the risk of human transmission is strengthened by the

  14. Identification of Cryptosporidium Species in Fish from Lake Geneva (Lac Léman) in France.

    Science.gov (United States)

    Certad, Gabriela; Dupouy-Camet, Jean; Gantois, Nausicaa; Hammouma-Ghelboun, Ourida; Pottier, Muriel; Guyot, Karine; Benamrouz, Sadia; Osman, Marwan; Delaire, Baptiste; Creusy, Colette; Viscogliosi, Eric; Dei-Cas, Eduardo; Aliouat-Denis, Cecile Marie; Follet, Jérôme

    2015-01-01

    Cryptosporidium, a protozoan parasite that can cause severe diarrhea in a wide range of vertebrates including humans, is increasingly recognized as a parasite of a diverse range of wildlife species. However, little data are available regarding the identification of Cryptosporidium species and genotypes in wild aquatic environments, and more particularly in edible freshwater fish. To evaluate the prevalence of Cryptosporidiumspp. in fish from Lake Geneva (Lac Léman) in France, 41 entire fish and 100 fillets (cuts of fish flesh) were collected from fishery suppliers around the lake. Nested PCR using degenerate primers followed by sequence analysis was used. Five fish species were identified as potential hosts of Cryptosporidium: Salvelinus alpinus, Esox lucius, Coregonus lavaretus, Perca fluviatilis, and Rutilus rutilus. The presence of Cryptosporidium spp. was found in 15 out of 41 fish (37%), distributed as follows: 13 (87%) C. parvum, 1 (7%) C. molnari, and 1 (7%) mixed infection (C. parvum and C. molnari). C. molnari was identified in the stomach, while C. parvum was found in the stomach and intestine. C. molnari was also detected in 1 out of 100 analyzed fillets. In order to identify Cryptosporidium subtypes, sequencing of the highly polymorphic 60-kDa glycoprotein (gp60) was performed. Among the C. parvum positive samples, three gp60 subtypes were identified: IIaA15G2R1, IIaA16G2R1, and IIaA17G2R1. Histological examination confirmed the presence of potential developmental stages of C. parvum within digestive epithelial cells. These observations suggest that C. parvum is infecting fish, rather than being passively carried. Since C. parvum is a zoonotic species, fish potentially contaminated by the same subtypes found in terrestrial mammals would be an additional source of infection for humans and animals, and may also contribute to the contamination of the environment with this parasite. Moreover, the risk of human transmission is strengthened by the

  15. Cryptosporidium infection in a veal calf cohort in France: molecular characterization of species in a longitudinal study

    Directory of Open Access Journals (Sweden)

    Follet Jérôme

    2011-12-01

    Full Text Available Abstract Feces from 142 animals were collected on 15 farms in the region of Brittany, France. Each sample was directly collected from the rectum of the animal and identified with the ear tag number. Animals were sampled three times, at 5, 15 and 22 weeks of age. After DNA extraction from stool samples, nested PCR was performed to amplify partial 18S-rDNA and 60 kDa glycoprotein genes of Cryptosporidium. The parasite was detected on all farms. One hundred out of 142 calves (70.4% were found to be parasitized by Cryptosporidium. Amplified fragments were sequenced for Cryptosporidium species identification and revealed the presence of C. parvum (43.8%, C. ryanae (28.5%, and C. bovis (27%. One animal was infected with Cryptosporidium ubiquitum. The prevalence of these species was related to the age of the animal. C. parvum caused 86.7% of Cryptosporidium infections in 5-week-old calves but only 1.7% in 15-week-old animals. The analysis of the results showed that animals could be infected successively by C. parvum, C. ryanae, and C. bovis for the study period. C. parvum gp60 genotyping identifies 6 IIa subtypes of which 74.5% were represented by IIaA15G2R1. This work confirms previous studies in other countries showing that zoonotic C. parvum is the dominant species seen in young calves.

  16. Distribution of Cryptosporidium and Giardia spp. in selected species of protected and game mammals from North-Eastern Poland.

    Science.gov (United States)

    Paziewska, Anna; Bednarska, Małgorzata; Niewegłowski, Hubert; Karbowiak, Grzegorz; Bajer, Anna

    2007-01-01

    Cryptosporidium spp. and Giardia spp. are wide-spread pathogens of humans and many species of mammals. The ways of transmission are very complex and difficult to define. Both parasites occur in similar environments and share a broad host range. However, in Poland there is still little known about the epidemiology of these parasites due to the paucity of data on human cases and only few studies in wildlife. The aim of our study was to determine the distribution of two intestinal protozoa in a few species of protected and game mammals in North-Eastern Poland. Additionally, we wanted to compare prevalence and abundance of these parasites between wild and farm animals, and to determine the species/genotypes of Cryptosporidium. Fecal samples collected from protected species (European beaver-22, grey wolf-14, European bison-55, Polish Konik (horse)-5) and game mammals (red deer-52, roe deer-22, boar-5) were examined by IFA. We also studied a group of samples collected from farm animals: beaver-30, red deer-66, Polish konik-5. Cryptosporidium oocysts were identified in 5 of 7 studied animal species (prevalence from 9% in roe deer to 36% in wolves), Giardia cysts in 4 of 6 studied species (prevalence from 1.7% in red deer to 7.7% in European beaver). Sequencing analysis of COWP gene fragment revealed that 5 Cryptosporidium isolates from wolves were C. parvum genotype 2 (zoonotic). The results show the important role of examined species in maintaining the natural sources of Cryptosporidium spp. and Giardia spp. infections in the environment.

  17. Classification of Cryptosporidium species from patients with sporadic cryptosporidiosis by use of sequence-based multilocus analysis following mutation scanning.

    Science.gov (United States)

    Jex, Aaron R; Pangasa, Aradhana; Campbell, Bronwyn E; Whipp, Margaret; Hogg, Geoff; Sinclair, Martha I; Stevens, Melita; Gasser, Robin B

    2008-07-01

    In the present study, we analyzed genetic variation in Cryptosporidium species from humans (n = 62) with clinical cryptosporidiosis in South Australia. Sequence variation was assessed in regions within the small subunit of nuclear rRNA (p-SSU), the 70-kDa heat shock protein (p-hsp70), and the 60-kDa glycoprotein (p-gp60) genes by employing single-strand conformation polymorphism analysis and sequencing. Based on the analyses of p-SSU and p-hsp70, Cryptosporidium hominis (n = 38) and Cryptosporidium parvum (n = 24) were identified. The analysis of p-gp60 revealed eight distinct subgenotypes, classified as C. hominis IaA17R1 (n = 3), IbA9G3R2 (n = 14), IbA10G2R2 (n = 20), and IfA12G1R1 (n = 1), as well as C. parvum IIaA18G3R1 (n = 15), IIaA20G3R1 (n = 6), IIaA22G4R1 (n = 2), and IIcA5G3R2 (n = 1). Subgenotypes IaA17R1 and IIaA22G4R1 are new. Of the six other subgenotypes, IbA10G2R2, IIaA18G3R1, IIaA20G3R1, and IIcA5G3R2 were reported previously from the state of Victoria. This is the fourth record in Australia of C. parvum subgenotype IIaA18G3R1 from humans, which, to date, has been isolated only from cattle in other countries. This subgenotype might be a significant contributor to sporadic human cryptosporidiosis and may indicate a greater zoonotic contribution to the infection of humans in the area of study. Comparative analyses revealed, for the first time, the differences in the genetic makeup of Cryptosporidium populations between two relatively close, major metropolitan cities.

  18. Glycoproteins and Gal-GalNAc cause Cryptosporidium to switch from an invasive sporozoite to a replicative trophozoite.

    Science.gov (United States)

    Edwinson, Adam; Widmer, Giovanni; McEvoy, John

    2016-01-01

    The apicomplexan parasite Cryptosporidium causes cryptosporidiosis, a diarrheal disease that can become chronic and life threatening in immunocompromised and malnourished people. There is no effective drug treatment for those most at risk of severe cryptosporidiosis. The disease pathology is due to a repeated cycle of host cell invasion and parasite replication that amplifies parasite numbers and destroys the intestinal epithelium. This study aimed to better understand the Cryptosporidium replication cycle by identifying molecules that trigger the switch from invasive sporozoite to replicative trophozoite. Our approach was to treat sporozoites of Cryptosporidium parvum and Cryptosporidium hominis, the species causing most human cryptosporidiosis, with various media under axenic conditions and examine the parasites for rounding and nuclear division as markers of trophozoite development and replication, respectively. FBS had a concentration-dependent effect on trophozoite development in both species. Trophozoite development in C. parvum, but not C. hominis, was enhanced when RPMI supplemented with 10% FBS (RPMI-FBS) was conditioned by HCT-8 cells for 3h. The effect of non-conditioned and HCT-8 conditioned RPMI-FBS on trophozoite development was abrogated by proteinase K and sodium metaperiodate pretreatment, indicating a glycoprotein trigger. Cryptosporidium parvum and C. hominis trophozoite development also was triggered by Gal-GalNAc in a concentration-dependent manner. Cryptosporidium parvum replication was greatest following treatments with Gal-GalNAc, followed by conditioned RPMI-FBS and non-conditioned RPMI-FBS (PGalNAc (1mM).

  19. An IC-PCR method for detection of Cryptosporidium and Giardia in natural surface waters in Finland.

    Science.gov (United States)

    Rimhanen-Finne, Ruska; Hörman, Ari; Ronkainen, Pilvi; Hänninen, Marja Liisa

    2002-08-01

    We developed an immunocapture-based polymerase chain reaction (PCR) assay for simultaneous detection of Cryptosporidium parvum oocysts and Giardia intestinalis cysts in surface water. Using primer pairs Cry9/Cry15 and LaxA/LaxB for Cryptosporidium and Gdh1/Gdh4 for Giardia, the sensitivity of the entire detection procedure (dealing with concentration, separation, DNA purification and PCR amplification) was at the level of 50-100 oocysts and cysts. Of 54 surface water samples, 4 were positive for Cryptosporidium and 1 for Giardia. Cryptosporidium and Giardia were detected for the first time in surface water in Finland.

  20. Epidemiology of Cryptosporidium infection in cattle in China: a review

    Directory of Open Access Journals (Sweden)

    Gong Chao

    2017-01-01

    Full Text Available The present review discusses the findings of cryptosporidiosis research conducted in cattle in China and highlights the currently available information on Cryptosporidium epidemiology, genetic diversity, and distribution in China, which is critical to understanding the economic and public health importance of cryptosporidiosis transmission in cattle. To date, 10 Cryptosporidium species have been detected in cattle in China, with an overall infection rate of 11.9%. The highest rate of infection (19.5% was observed in preweaned calves, followed by that in juveniles (10.69%, postweaned juveniles (9.0%, and adult cattle (4.94%. The dominant species were C. parvum in preweaned calves and C. andersoni in postweaned, juvenile, and adult cattle. Zoonotic Cryptosporidium species (C. parvum and C. hominis were found in cattle, indicating the possibility of transmission between humans and cattle. Different cattle breeds had significant differences in the prevalence rate and species of Cryptosporidium. This review demonstrates an age-associated, breed-associated, and geographic-related occurrence of Cryptosporidium and provides references for further understanding of the epidemiological characteristics, and for preventing and controlling the disease.

  1. Epidemiology of Cryptosporidium infection in cattle in China: a review

    Science.gov (United States)

    Gong, Chao; Cao, Xue-Feng; Deng, Lei; Li, Wei; Huang, Xiang-Ming; Lan, Jing-Chao; Xiao, Qi-Cheng; Zhong, Zhi-Jun; Feng, Fan; Zhang, Yue; Wang, Wen-Bo; Guo, Ping; Wu, Kong-Ju; Peng, Guang-Neng

    2017-01-01

    The present review discusses the findings of cryptosporidiosis research conducted in cattle in China and highlights the currently available information on Cryptosporidium epidemiology, genetic diversity, and distribution in China, which is critical to understanding the economic and public health importance of cryptosporidiosis transmission in cattle. To date, 10 Cryptosporidium species have been detected in cattle in China, with an overall infection rate of 11.9%. The highest rate of infection (19.5%) was observed in preweaned calves, followed by that in juveniles (10.69%), postweaned juveniles (9.0%), and adult cattle (4.94%). The dominant species were C. parvum in preweaned calves and C. andersoni in postweaned, juvenile, and adult cattle. Zoonotic Cryptosporidium species (C. parvum and C. hominis) were found in cattle, indicating the possibility of transmission between humans and cattle. Different cattle breeds had significant differences in the prevalence rate and species of Cryptosporidium. This review demonstrates an age-associated, breed-associated, and geographic-related occurrence of Cryptosporidium and provides references for further understanding of the epidemiological characteristics, and for preventing and controlling the disease. PMID:28098070

  2. Giardia and Cryptosporidium in cetaceans on the European Atlantic coast.

    Science.gov (United States)

    Reboredo-Fernández, Aurora; Ares-Mazás, Elvira; Martínez-Cedeira, José A; Romero-Suances, Rafael; Cacciò, Simone M; Gómez-Couso, Hipólito

    2015-02-01

    The occurrence of Giardia and Cryptosporidium was investigated in cetacean specimens stranded on the northwestern coast of Spain (European Atlantic coast) by analysis of 65 samples of large intestine from eight species. The parasites were identified by direct immunofluorescence antibody test (IFAT) and by PCR amplification of the β-giardin gene, the ITS1-5.8S-ITS2 region and the SSU-rDNA gene of Giardia and the SSU-rDNA gene of Cryptosporidium. Giardia and Cryptosporidium were detected in 7 (10.8 %) and 9 samples (13.8 %), respectively. In two samples, co-infection with both parasites was observed. Giardia duodenalis assemblages A, C, D and F, and Cryptosporidium parvum were identified. This is the first report of G. duodenalis in Balaenoptera acutorostrata, Kogia breviceps and Stenella coeruleoalba and also the first report of Cryptosporidium sp. in B. acutorostrata and of C. parvum in S. coeruleoalba and Tursiops truncatus. These results extend the known host range of these waterborne enteroparasites.

  3. INFECTIVITY OF A CRYPTOSPORIDIUM PARVUM ISOLATE OF CERVINE ORIGIN FOR HEALTHY ADULTS AND GAMMA INTERFERON-G KNOCKOUT MICE. (R828035)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  4. INFECTIVITY OF A CRYPTOSPORIDIUM PARVUM ISOLATE OF CERVINE ORIGIN FOR HEALTHY ADULTS AND GAMMA INTERFERON-G KNOCKOUT MICE. (R829180)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  5. INFECTIVITY OF A CRYPTOSPORIDIUM PARVUM ISOLATE OF CERVINE ORIGIN FOR HEALTHY ADULTS AND GAMMA INTERFERON-G KNOCKOUT MICE. (R828035)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  6. INFECTIVITY OF A CRYPTOSPORIDIUM PARVUM ISOLATE OF CERVINE ORIGIN FOR HEALTHY ADULTS AND GAMMA INTERFERON-G KNOCKOUT MICE. (R829180)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  7. Molecular Identification of Cryptosporidium Species from Pet Snakes in Thailand

    Science.gov (United States)

    Yimming, Benjarat; Pattanatanang, Khampee; Sanyathitiseree, Pornchai; Inpankaew, Tawin; Kamyingkird, Ketsarin; Pinyopanuwat, Nongnuch; Chimnoi, Wissanuwat; Phasuk, Jumnongjit

    2016-01-01

    Cryptosporidium is an important pathogen causing gastrointestinal disease in snakes and is distributed worldwide. The main objectives of this study were to detect and identify Cryptosporidium species in captive snakes from exotic pet shops and snake farms in Thailand. In total, 165 fecal samples were examined from 8 snake species, boa constrictor (Boa constrictor constrictor), corn snake (Elaphe guttata), ball python (Python regius), milk snake (Lampropeltis triangulum), king snake (Lampropeltis getula), rock python (Python sebae), rainbow boa (Epicrates cenchria), and carpet python (Morelia spilota). Cryptosporidium oocysts were examined using the dimethyl sulfoxide (DMSO)-modified acid-fast staining and a molecular method based on nested-PCR, PCR-RFLP analysis, and sequencing amplification of the SSU rRNA gene. DMSO-modified acid-fast staining revealed the presence of Cryptosporidium oocysts in 12 out of 165 (7.3%) samples, whereas PCR produced positive results in 40 (24.2%) samples. Molecular characterization indicated the presence of Cryptosporidium parvum (mouse genotype) as the most common species in 24 samples (60%) from 5 species of snake followed by Cryptosporidium serpentis in 9 samples (22.5%) from 2 species of snake and Cryptosporidium muris in 3 samples (7.5%) from P. regius. PMID:27658593

  8. Molecular Identification of Cryptosporidium Species from Pet Snakes in Thailand.

    Science.gov (United States)

    Yimming, Benjarat; Pattanatanang, Khampee; Sanyathitiseree, Pornchai; Inpankaew, Tawin; Kamyingkird, Ketsarin; Pinyopanuwat, Nongnuch; Chimnoi, Wissanuwat; Phasuk, Jumnongjit

    2016-08-01

    Cryptosporidium is an important pathogen causing gastrointestinal disease in snakes and is distributed worldwide. The main objectives of this study were to detect and identify Cryptosporidium species in captive snakes from exotic pet shops and snake farms in Thailand. In total, 165 fecal samples were examined from 8 snake species, boa constrictor (Boa constrictor constrictor), corn snake (Elaphe guttata), ball python (Python regius), milk snake (Lampropeltis triangulum), king snake (Lampropeltis getula), rock python (Python sebae), rainbow boa (Epicrates cenchria), and carpet python (Morelia spilota). Cryptosporidium oocysts were examined using the dimethyl sulfoxide (DMSO)-modified acid-fast staining and a molecular method based on nested-PCR, PCR-RFLP analysis, and sequencing amplification of the SSU rRNA gene. DMSO-modified acid-fast staining revealed the presence of Cryptosporidium oocysts in 12 out of 165 (7.3%) samples, whereas PCR produced positive results in 40 (24.2%) samples. Molecular characterization indicated the presence of Cryptosporidium parvum (mouse genotype) as the most common species in 24 samples (60%) from 5 species of snake followed by Cryptosporidium serpentis in 9 samples (22.5%) from 2 species of snake and Cryptosporidium muris in 3 samples (7.5%) from P. regius.

  9. A recombinant DNA vaccine encoding C. andersoni oocyst wall protein induces immunity against experimental C. parvum infection.

    Science.gov (United States)

    Zheng, Jun; Ren, Wenzhi; Pan, Qingshan; Wang, Qiuyue; Elhag, I A Elfaki; Li, Jianhua; Li, Mingying; Gong, Pengtao; Liu, Yingli; Zhang, Xichen

    2011-06-30

    Cryptosporidium andersoni parasited in the abomasum has been demonstrated as a cause of reduction of milk production in dairy cow. In this study, a novel chimeric DNA vaccine pVAX1-AB was constructed and the efficacy against Cryptosporidium parvum was determined. BALB/c mice were divided into 3 groups and immunized with DNA vaccine expressing the oocyst wall protein, AB protein of C. andersoni, the recombinant plasmid containing the AB gene, respectively. After inoculation of 1 × 10(6) oocysts of C. parvum, the humoral and cellular immune responses were detected. Experimental results showed that the recombinant plasmid can induce corresponding specific antibody response, simultaneously influenced cellular immune responses, and provided greater protection rate (48.6%) than the other groups. These results indicated that chimeric DNA vaccine has a potential in Cryptosporidium vaccine development.

  10. Occurrence and potential health risk of Cryptosporidium and Giardia in different water catchments in Belgium.

    Science.gov (United States)

    Ehsan, Amimul; Geurden, Thomas; Casaert, Stijn; Paulussen, Jef; De Coster, Lut; Schoemaker, Toon; Chalmers, Rachel; Grit, Grietje; Vercruysse, Jozef; Claerebout, Edwin

    2015-02-01

    Human wastewater and livestock can contribute to contamination of surface water with Cryptosporidium and Giardia. In countries where a substantial proportion of drinking water is produced from surface water, e.g., Belgium, this poses a constant threat on drinking water safety. Our objective was to monitor the presence of Cryptosporidium and Giardia in different water catchment sites in Belgium and to discriminate between (oo)cysts from human or animal origin using genotyping. Monthly samples were collected from raw water and purified drinking water at four catchment sites. Cryptosporidium and Giardia were detected using USEPA method 1623 and positive samples were genotyped. No contamination was found in purified water at any site. In three catchments, only low numbers of (oo)cysts were recovered from raw water samples (Giardia (92 %) and Cryptosporidium (96 %), especially in winter and spring. Genotyping of Giardia in 38 water samples identified the presence of Giardia duodenalis assemblage AI, AII, BIV, BIV-like, and E. Cryptosporidium andersoni, Cryptosporidium suis, Cryptosporidium horse genotype, Cryptosporidium parvum, and Cryptosporidium hominis were detected. The genotyping results suggest that agriculture may be a more important source of surface water contamination than human waste in this catchment. In catchment sites with contaminated surface water, such as the Blankaart, continuous monitoring of treated water for the presence of Cryptosporidium and Giardia would be justified and (point) sources of surface water contamination should be identified.

  11. Prevalence of the protozoan parasite Cryptosporidium on three organic pig farms in Denmark

    DEFF Research Database (Denmark)

    Petersen, Heidi H.; Jianmin, Wang; Mejer, Helena;

    2013-01-01

    Pigs are a potential source of contamination with Cryptosporidium spp., which can lead to infection in humans. Two species C. parvum and C. hominis can cause an acute diarrheal illness in humans, which can become severe in e.g. patients with HIV. The oocyst can survive for long periods in the env...

  12. Prevalence and molecular characterization of bovine Cryptosporidium in Qazvin province, Iran.

    Science.gov (United States)

    Keshavarz, Akbar; Haghighi, Ali; Athari, Amid; Kazemi, Bahram; Abadi, Alireza; Nazemalhosseini Mojarad, Ehsan

    2009-03-23

    The aim of this study was to determine the prevalence, variability with host age, and the genotypes of species of Cryptosporidium in cattle from 15 dairy farms in Qazvin province, Iran. Fecal samples, collected from 272 cattle during May 2006 to December 2007, were characterized microscopically. Oocysts from 51 positive samples were analyzed using PCR assay of 18S SSU rRNA, restriction fragment length polymorphism (RFLP) and sequencing. We identified 72.6% of the positive samples as Cryptosporidium parvum, 17.7% as Cryptosporidium andersoni, 7.8% as Cryptosporidium bovis and 1.9% as a novel genotype of C. parvum possessing a single mutation on MboII restriction. An infection rate of 19.5% of C. parvum among 174 pre-weaned calves was significantly higher than the 3.1% among 98 post-weaned calves (P<0.0006). This is the first report of C. bovis and the new subgenotype of C. parvum in Iranian cattle.

  13. The effects of Moringa lieifera seed powder on turbidity and sedimentation of Cryptosporidium spp. in wastewater

    DEFF Research Database (Denmark)

    Petersen, H. H.; Wolsey, I.; Dalsgaard, A.

    2013-01-01

    or water used for postharvest washing of the produce is contaminated. A laboratory study was carried out to investigate the effect of a coagulant from the seeds of Moringa oleifera (MO) in reducing Cryptosporidium parvum oocysts and turbidity in Danish wastewater. To each of five time points, 12 replicates...

  14. Multiplex PCR detection of waterborne intestinal protozoa: microsporidia, Cyclospora, and Cryptosporidium.

    Science.gov (United States)

    Lee, Seung-Hyun; Joung, Migyo; Yoon, Sejoung; Choi, Kyoungjin; Park, Woo-Yoon; Yu, Jae-Ran

    2010-12-01

    Recently, emerging waterborne protozoa, such as microsporidia, Cyclospora, and Cryptosporidium, have become a challenge to human health worldwide. Rapid, simple, and economical detection methods for these major waterborne protozoa in environmental and clinical samples are necessary to control infection and improve public health. In the present study, we developed a multiplex PCR test that is able to detect all these 3 major waterborne protozoa at the same time. Detection limits of the multiplex PCR method ranged from 10(1) to 10(2) oocysts or spores. The primers for microsporidia or Cryptosporidium used in this study can detect both Enterocytozoon bieneusi and Encephalitozoon intestinalis, or both Cryptosporidium hominis and Cryptosporidium parvum, respectively. Restriction enzyme digestion of PCR products with BsaBI or BsiEI makes it possible to distinguish the 2 species of microsporidia or Cryptosporidium, respectively. This simple, rapid, and cost-effective multiplex PCR method will be useful for detecting outbreaks or sporadic cases of waterborne protozoa infections.

  15. Molecular Characterization of Cryptosporidium Species and Giardia duodenalis from Symptomatic Cambodian Children.

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    Catrin E Moore

    2016-07-01

    Full Text Available In a prospective study, 498 single faecal samples from children aged under 16 years attending an outpatient clinic in the Angkor Hospital for Children, northwest Cambodia, were examined for Cryptosporidium oocysts and Giardia cysts using microscopy and molecular assays.Cryptosporidium oocysts were detected in 2.2% (11/498 of samples using microscopy and in 7.7% (38/498 with molecular tests. Giardia duodenalis cysts were detected in 18.9% (94/498 by microscopy and 27.7% (138/498 by molecular tests; 82% of the positive samples (by either method were from children aged 1-10 years. Cryptosporidium hominis was the most common species of Cryptosporidium, detected in 13 (34.2% samples, followed by Cryptosporidium meleagridis in 9 (23.7%, Cryptosporidium parvum in 8 (21.1%, Cryptosporidium canis in 5 (13.2%, and Cryptosporidium suis and Cryptosporidium ubiquitum in one sample each. Cryptosporidium hominis and C. parvum positive samples were subtyped by sequencing the GP60 gene: C. hominis IaA16R6 and C. parvum IIeA7G1 were the most abundant subtypes. Giardia duodenalis was typed using a multiplex real-time PCR targeting assemblages A and B. Assemblage B (106; 76.8% of all Giardia positive samples was most common followed by A (12.3% and mixed infections (5.1%. Risk factors associated with Cryptosporidium were malnutrition (AOR 9.63, 95% CI 1.67-55.46, chronic medical diagnoses (AOR 4.51, 95% CI 1.79-11.34 and the presence of birds in the household (AOR 2.99, 95% CI 1.16-7.73; specifically C. hominis (p = 0.03 and C. meleagridis (p<0.001 were associated with the presence of birds. The use of soap was protective against Giardia infection (OR 0.74, 95% CI 0.58-0.95.This is the first report to describe the different Cryptosporidium species and subtypes and Giardia duodenalis assemblages in Cambodian children. The variety of Cryptosporidium species detected indicates both anthroponotic and zoonotic transmission in this population. Interventions to improve

  16. Investigating source water Cryptosporidium concentration, species and infectivity rates during rainfall-runoff in a multi-use catchment.

    Science.gov (United States)

    Swaffer, Brooke A; Vial, Hayley M; King, Brendon J; Daly, Robert; Frizenschaf, Jacqueline; Monis, Paul T

    2014-12-15

    Protozoan pathogens present a significant human health concern, and prevention of contamination into potable networks remains a key focus for drinking water providers. Here, we monitored the change in Cryptosporidium concentration in source water during high flow events in a multi-use catchment. Furthermore, we investigated the diversity of Cryptosporidium species/genotypes present in the source water, and delivered an oocyst infectivity fraction. There was a positive and significant correlation between Cryptosporidium concentration and flow (ρ = 0.756) and turbidity (ρ = 0.631) for all rainfall-runoff events, despite variable source water pathogen concentrations. Cell culture assays measured oocyst infectivity and suggested an overall source water infectious fraction of 3.1%. No infectious Cryptosporidium parvum or Cryptosporidium hominis were detected, although molecular testing detected C. parvum in 7% of the samples analysed using PCR-based molecular techniques. Twelve Cryptosporidium species/genotypes were identified using molecular techniques, and were reflective of the host animals typically found in remnant vegetation and agricultural areas. The inclusion of molecular approaches to identify Cryptosporidium species and genotypes highlighted the diversity of pathogens in water, which originated from various sources across the catchment. We suggest this mixing of runoff water from a range of landuses containing diverse Cryptosporidium hosts is a key explanation for the often-cited difficulty forming strong pathogen-indicator relationships.

  17. Molecular characterization of Cryptosporidium spp. from fecal samples of birds kept in captivity in Brazil.

    Science.gov (United States)

    Nakamura, Alex Akira; Simões, Daniel Castendo; Antunes, Rômulo Godik; da Silva, Deuvânia Carvalho; Meireles, Marcelo Vasconcelos

    2009-12-03

    The aim of this study was to determine the prevalence of Cryptosporidium species and genotypes in birds kept in captivity in Brazil. A total of 966 samples from 18 families of birds was collected and stored in 5% potassium dichromate solution at 4 degrees C until processing. Oocysts were purified in Sheather sugar solution following extraction of genomic DNA. Molecular analyses were performed using nested-PCR for amplification of fragments of the 18S subunit of rRNA gene and of the actin gene. Amplification of Cryptosporidium DNA fragments was obtained in 47 (4.86%) samples. Sequencing of amplified fragments and phylogenetic analyses allowed the identification of Cryptosporidium baileyi in a black vulture (Coragyps atratus), a domestic chicken (Gallus gallus domesticus) and a saffron finch (Sicalis flaveola); Cryptosporidium galli in canaries (Serinus canaria), a cockatiel (Nymphicus hollandicus) and lesser seed-finches (Oryzoborus angolensis); Cryptosporidium meleagridis in a domestic chicken (G. g. domesticus); Cryptosporidium parvum in a cockatiel (N. hollandicus); Cryptosporidium avian genotype I in a canary (S. canaria) and an Indian peafowl (Pavo cristatus); Cryptosporidium avian genotype II in ostriches (Struthio camelus) and Cryptosporidium avian genotype III in a cockatiel (N. hollandicus) and a peach-faced lovebird (Agapornis roseicolis).

  18. Evidence for a new species of Cryptosporidium infecting tortoises: Cryptosporidium ducismarci

    Directory of Open Access Journals (Sweden)

    Traversa Donato

    2010-03-01

    Full Text Available Abstract Cryptosporidiosis affects the gastrointestinal and respiratory tract of humans as well as of a wide range of companion, farm, laboratory and wild animals. In the past few years, three independent studies have provided strong evidence for the existence of a distinct Cryptosporidium species affecting tortoises and likely circulating in other reptile species as well. A new Cryptosporidium genotype was firstly detected and genetically characterized in a marginated tortoise in Italy in 2007 and named Cryptosporidium sp. ex Testudo marginata CrIT-20. The phylogenetic analysis of this isolate indicated that this Cryptosporidium was unique and belonged to the intestinal clade. These findings were later on confirmed by the detection of genetic homologies of isolates from a python and a chameleon from Spain and by recent research in the United States. The latter study presented both the occurrence of intestinal lesions in a pancake tortoise and a Russian tortoise and the genetic characterization of the isolates, together with the first pictures of the endogenous stages of Cryptosporidium CrIT-20. Phylogenetic inference based on the sequences representing small subunit of the nuclear ribosomal RNA gene (SSU of these isolates confirmed the pathological findings because this Cryptosporidium was related to the intestinal group and supported previous results in T. marginata from Italy. The present scientific data on the Cryptosporidium CrIT-20 support its classification as a new species of Cryptosporidium causing intestinal diseases in tortoises. Although further morphological (i.e. exogenous stages and biological aspects (i.e. complete host range are yet to be elucidated, it is proposed that this Cryptosporidium is designated Cryptosporidium ducismarci.

  19. Occurrence and molecular characterization of Cryptosporidium spp. isolated from domestic animals in a rural area surrounding Atlantic dry forest fragments in Teodoro Sampaio municipality, State of São Paulo, Brazil Ocorrência e caracterização molecular de Cryptosporidium spp. isolados de animais domésticos de propriedades rurais circunvizinhas a fragmentos de Floresta Atlântica Seca do Estado de São Paulo, Brasil

    Directory of Open Access Journals (Sweden)

    Anaiá da Paixão Sevá

    2010-12-01

    Full Text Available The aim of this study was to assess the occurrence of Cryptosporidium in domestic animals in rural properties surrounding rain forest fragments within the municipality of Teodoro Sampaio, southeastern Brazil. Conventional sucrose flotation method followed by molecular characterization of the parasites by sequencing PCR products amplified from SSU rRNA gene were used. Stool samples were collected from domestic animals raised as pets and livestock in all rural properties surrounding three forest fragments. Samples from cattle (197, equine (63, pigs (25, sheep (11, and dogs (28 were collected from 98 rural properties. The frequency of occurrence of Cryptosporidium within each animal species was 3.0% (6/197 among cattle and 10.7% (3/28 among dogs. Cryptosporidium was not detected in stool samples from equine, sheep, and pigs. All sequences obtained from the six samples of calves showed molecular identity with Cryptosporidium andersoni while all sequences from dog samples were similar to C. canis. The frequency of occurrence of Cryptosporidium in these domestic animal species was low. The absence of C. parvum in the present study suggests that the zoonotic cycle of cryptosporidiosis may not be relevant in the region studied. The presence of Cryptosporidium species seldom described in humans may be, otherwise, important for the wild fauna as these animals are a source of infection and dissemination of this protozoan to other animal species. The impact and magnitude of infection by C. andersoni in wild ruminants and C. canis in wild canids have to be assessed in future studies to better understand the actual importance of these species in this region.O objetivo deste estudo foi avaliar a ocorrência de Cryptosporidium, em animais domésticos de propriedades rurais ao redor de fragmentos de mata Atlântica de interior no município de Teodoro Sampaio, por exame convencional de flutuação em solução de sacarose, seguido de caracterização molecular

  20. Molecular Insights for Giardia, Cryptosporidium, and Soil-Transmitted Helminths from a Facility-Based Surveillance System in Guatemala

    Science.gov (United States)

    Velasquez, Daniel E.; Arvelo, Wences; Cama, Vitaliano A.; López, Beatriz; Reyes, Lissette; Roellig, Dawn M.; Kahn, Geoffrey D.; Lindblade, Kimberly A.

    2011-01-01

    We molecularly characterized samples with Giardia, Cryptosporidium, and soil-transmitted helminths from a facility-based surveillance system for diarrhea in Santa Rosa, Guatemala. The DNA sequence analysis determined the presence of Giardia assemblages A (N = 7) and B (N = 12) and, Cryptosporidium hominis (N = 2) and Cryptosporidium parvum (N = 2), suggestive of different transmission cycles. All 41 samples with soil-transmitted helminths did not have the β-tubulin mutation described for benzimidazole resistance, suggesting potential usefulness in mass drug administration campaigns. PMID:22144459

  1. Cryptosporidium parvum: infectivity and pathogenicity of the 'porcine' genotype

    DEFF Research Database (Denmark)

    Enemark, Heidi L.; Ahrens, Peter; Bille-Hansen, Vivi

    2003-01-01

    mild clinical signs in piglets despite the excretion of high numbers of oocysts. Concomitant infection with rotavirus, however, caused a dramatic aggravation of the clinical signs, and 5 of 6 experimentally infected piglets died. CPP-13 appeared to be adapted to porcine hosts as illustrated by the lack...

  2. Bioaccumulation of Toxoplasma and Cryptosporidium by the crustacean Gammarus fossarum: involvement in biomonitoring survey and trophic transfer

    Science.gov (United States)

    The protozoans Toxoplasma gondii and Cryptosporidium parvum are public health priorities and their oocysts can persist in environment for long time. They are present in various watercourses as recreational, surface, drinking, river, and seawater and could interact with organisms. To evaluate the cap...

  3. Drug treatment and novel drug target against Cryptosporidium

    Directory of Open Access Journals (Sweden)

    Gargala G.

    2008-09-01

    Full Text Available Cryptosporidiosis emergence triggered the screening of many compounds for potential anti-cryptosporidial activity in which the majority were ineffective. The outbreak of cryptosporidiosis which occurred in Milwaukee in 1993 was not only the first significant emergence of Cryptosporidium spp. as a major human pathogen but also a huge waterborne outbreak thickening thousands of people from a major city in North America. Since then, outbreaks of cryptosporidiosis are regularly occurring throughout the world. New drugs against this parasite became consequently urgently needed. Among the most commonly used treatments against cryptosporidiosis are paromomycin, and azithromycin, which are partially effective. Nitazoxanide (NTZ’s effectiveness was demonstrated in vitro, and in vivo using several animal models and finally in clinical trials. It significantly shortened the duration of diarrhea and decreased mortality in adults and in malnourished children. NTZ is not effective without an appropriate immune response. In AIDS patients, combination therapy restoring immunity along with antimicrobial treatment of Cryptosporidium infection is necessary. Recent investigations focused on the potential of molecular-based immunotherapy against this parasite. Others tested the effects of probiotic bacteria, but were unable to demonstrate eradication of C. parvum. New synthetic isoflavone derivatives demonstrated excellent activity against C. parvum in vitro and in a gerbil model of infection. Newly synthesized nitroor non nitro- thiazolide compounds, derived from NTZ, have been recently shown to be at least as effective as NTZ against C. parvum in vitro development and are promising new therapeutic agents.

  4. Prevalence and genetic characteristics of Cryptosporidium, Enterocytozoon bieneusi and Giardia duodenalis in cats and dogs in Heilongjiang province, China.

    Science.gov (United States)

    Li, Wei; Li, Yijing; Song, Mingxin; Lu, Yixin; Yang, Jinping; Tao, Wei; Jiang, Yanxue; Wan, Qiang; Zhang, Siwen; Xiao, Lihua

    2015-03-15

    This study investigated 319 fecal specimens of cats (n=52) and dogs (n=267) from Heilongjiang province, China for the prevalence and genetic characteristics of Cryptosporidium, Enterocytozoon bieneusi, and Giardia duodenalis. PCR and DNA sequence analysis of the small subunit rRNA gene identified C. felis and C. parvum in one cat each (3.8%) and C. canis and C. ubiquitum in 6 dogs (2.2%). Polymorphisms in the ribosomal internal transcribed spacer and phylogenetic analysis characterized zoonotic E. bieneusi genotypes D, EbpC, NED1, and NED2 and host-adapted ones NED3, NED4, and PtEb IX in 18 dogs (6.7%) and human-pathogenic genotypes D and IV in 3 cats (5.8%). Genotyping based on the hypermutation of G. duodenalis triosephosphate isomerase gene (TPI) facilitated identification of assemblage F in a cat (1.9%) and assemblages C and E in 12 dogs (4.5%). Subtypes of G. duodenalis isolates were determined by measuring the diversity of both TPI nucleotide and amino acid sequences. C. canis, C. felis, C. parvum, E. bieneusi genotypes D, EbpC, and IV, and G. duodenalis assemblage C identified herein have been documented in human infections in China. C. canis, C. parvum, C. ubiquitum, and E. bieneusi genotypes D, EbpC, and IV carried by cats or dogs also existed in wastewater in China. The finding suggested pet animals could be reservoirs for human cryptosporidiosis, microsporidiosis, and giardiasis and potential sources of water contamination in China.

  5. Molecular characterization of Cryptosporidium and Giardia in farmers and their ruminant livestock from the Coastal Savannah zone of Ghana.

    Science.gov (United States)

    Squire, Sylvia Afriyie; Yang, Rongchang; Robertson, Ian; Ayi, Irene; Ryan, Una

    2017-09-21

    Cryptosporidium and Giardia are major causes of diarrhoea in developing countries including Ghana, however, nothing is known about the species and subtypes of Cryptosporidium and Giardia in farmers and their ruminant livestock in this country. A total of 925 faecal samples from humans (n=95), cattle (n=328), sheep (n=217) and goats (n=285), were screened for Cryptosporidium and Giardia by quantitative PCR (qPCR) at the 18S rRNA and glutamate dehydrogenase (gdh) loci respectively. Cryptosporidium positives were typed by sequence analysis of 18S and 60kDa glycoprotein (gp60) loci amplicons. Giardia positives were typed at the triose phosphate isomerase (tpi), beta-giardin (bg) and gdh loci. The prevalence of Cryptosporidium and Giardia by qPCR was 8.4% and 10.5% in humans, 26.5% and 8.5% in cattle, 34.1% and 12.9% in sheep, and 33.3% and 12.3% in goat faecal samples, respectively. G. duodenalis assemblages A and B were detected in humans and assemblage E was detected in livestock. Cryptosporidium parvum was the only species identified in humans; C. andersoni, C. bovis, C. ryanae and C. ubiquitum were identified in cattle; C. xiaoi, C. ubiquitum and C. bovis in sheep; and C. xiaoi, C. baileyi and C. parvum in goats. This is the first molecular study of Cryptosporidium and Giardia in livestock in Ghana. The identification of zoonotic species and the identification of C. parvum subtype IIcA5G3q in livestock, which has previously been identified in children in Ghana, suggests potential zoonotic transmission. Further studies on larger numbers of human and animal samples, and on younger livestock are required to better understand the epidemiology and transmission of Cryptosporidium and Giardia in Ghana. Copyright © 2017. Published by Elsevier B.V.

  6. Molecular characterization of Cryptosporidium spp. in children from Mexico.

    Science.gov (United States)

    Valenzuela, Olivia; González-Díaz, Mariana; Garibay-Escobar, Adriana; Burgara-Estrella, Alexel; Cano, Manuel; Durazo, María; Bernal, Rosa M; Hernandez, Jesús; Xiao, Lihua

    2014-01-01

    Cryptosporidiosis is a parasitic disease caused by Cryptosporidium spp. In immunocompetent individuals, it usually causes an acute and self-limited diarrhea; in infants, infection with Cryptosporidium spp. can cause malnutrition and growth retardation, and declined cognitive ability. In this study, we described for the first time the distribution of C. parvum and C. hominis subtypes in 12 children in Mexico by sequence characterization of the 60-kDa glycoprotein (GP60) gene of Cryptosporidium. Altogether, 7 subtypes belonging to 4 subtype families of C. hominis (Ia, Ib, Id and Ie) and 1 subtype family of C. parvum (IIa) were detected, including IaA14R3, IaA15R3, IbA10G2, IdA17, IeA11G3T3, IIaA15G2R1 and IIaA16G1R1. The frequency of the subtype families and subtypes in the samples analyzed in this study differed from what was observed in other countries.

  7. Molecular characterization of Cryptosporidium spp. in children from Mexico.

    Directory of Open Access Journals (Sweden)

    Olivia Valenzuela

    Full Text Available Cryptosporidiosis is a parasitic disease caused by Cryptosporidium spp. In immunocompetent individuals, it usually causes an acute and self-limited diarrhea; in infants, infection with Cryptosporidium spp. can cause malnutrition and growth retardation, and declined cognitive ability. In this study, we described for the first time the distribution of C. parvum and C. hominis subtypes in 12 children in Mexico by sequence characterization of the 60-kDa glycoprotein (GP60 gene of Cryptosporidium. Altogether, 7 subtypes belonging to 4 subtype families of C. hominis (Ia, Ib, Id and Ie and 1 subtype family of C. parvum (IIa were detected, including IaA14R3, IaA15R3, IbA10G2, IdA17, IeA11G3T3, IIaA15G2R1 and IIaA16G1R1. The frequency of the subtype families and subtypes in the samples analyzed in this study differed from what was observed in other countries.

  8. Efficacy of chlorine dioxide tablets on inactivation of cryptosporidium oocysts.

    Science.gov (United States)

    Murphy, Jennifer L; Haas, Charles N; Arrowood, Michael J; Hlavsa, Michele C; Beach, Michael J; Hill, Vincent R

    2014-05-20

    The ability of chlorine dioxide (ClO2) to achieve 2-log inactivation of Cryptosporidium in drinking water has been documented. No studies have specifically addressed the effects of ClO2 on C. parvum oocyst infectivity in chlorinated recreational water venues (e.g., pools). The aim of this research was to determine the efficacy of ClO2 as an alternative to existing hyperchlorination protocols that are used to achieve a 3-log inactivation of Cryptosporidium in such venues. To obtain a 3-log inactivation of C. parvum Iowa oocysts, contact times of 105 and 128 min for a solution containing 5 mg/L ClO2 with and without the addition of 2.6 mg/L free chlorine, respectively, were required. Contact times of 294 and 857 min for a solution containing 1.4 mg/L ClO2 with and without the addition of 3.6 mg/L free chlorine, respectively, were required. The hyperchlorination control (21 mg/L free chlorine only) required 455 min for a 3-log inactivation. Use of a solution containing 5 mg/L ClO2 and solutions containing 5 or 1.4 mg/L ClO2 with the addition of free chlorine appears to be a promising alternative to hyperchlorination for inactivating Cryptosporidium in chlorinated recreational water venues, but further studies are required to evaluate safety constraints on use.

  9. Molecular Characterization of Cryptosporidium spp. in Children from Mexico

    Science.gov (United States)

    Valenzuela, Olivia; González-Díaz, Mariana; Garibay-Escobar, Adriana; Burgara-Estrella, Alexel; Cano, Manuel; Durazo, María; Bernal, Rosa M.; Hernandez, Jesús; Xiao, Lihua

    2014-01-01

    Cryptosporidiosis is a parasitic disease caused by Cryptosporidium spp. In immunocompetent individuals, it usually causes an acute and self-limited diarrhea; in infants, infection with Cryptosporidium spp. can cause malnutrition and growth retardation, and declined cognitive ability. In this study, we described for the first time the distribution of C. parvum and C. hominis subtypes in 12 children in Mexico by sequence characterization of the 60-kDa glycoprotein (GP60) gene of Cryptosporidium. Altogether, 7 subtypes belonging to 4 subtype families of C. hominis (Ia, Ib, Id and Ie) and 1 subtype family of C. parvum (IIa) were detected, including IaA14R3, IaA15R3, IbA10G2, IdA17, IeA11G3T3, IIaA15G2R1 and IIaA16G1R1. The frequency of the subtype families and subtypes in the samples analyzed in this study differed from what was observed in other countries. PMID:24755606

  10. Presence and molecular characterisation of Giardia and Cryptosporidium in alpacas (Vicugna pacos) from Peru.

    Science.gov (United States)

    Gómez-Couso, Hipólito; Ortega-Mora, Luis M; Aguado-Martínez, Adriana; Rosadio-Alcántara, Raúl; Maturrano-Hernández, Lenin; Luna-Espinoza, Luis; Zanabria-Huisa, Víctor; Pedraza-Díaz, Susana

    2012-07-06

    The presence of Giardia and Cryptosporidium was investigated in 274 faecal samples of alpacas (Vicugna pacos) from 12 herds from Peru by immunofluorescence microscopy and PCR amplification and sequencing of fragments of the ssu-rRNA and β-giardin genes from Giardia spp., as well as the ssu-rRNA gene from Cryptosporidium spp. A total of 137 samples (50.0%) were positive for Giardia spp., and 12 samples (4.4%) for Cryptosporidium spp. In ten samples (3.6%), co-infection by both pathogens was found. Herd prevalence was found to be 91.7% (11/12 herds) for Giardia and 58.3% (7/12 herds) for Cryptosporidium. Regarding the age of the animals, although Giardia was detected in animals as young as 1 week, the prevalence increased with age, reaching 80% by 8 weeks. Similarly, the highest percentage of Cryptosporidium detection (20%) was also found in the 8 week-old group. By PCR, 92 of the 274 analysed samples were positive for Giardia. Sequencing of the amplicons showed the existence of Giardia duodenalis assemblage A in 67 samples; G. duodenalis assemblage E in 24 samples; and inconsistent results between the two molecular markers used in a further sample. Cryptosporidium was only detected by PCR in 3 of the 274 samples; Cryptosporidium parvum was identified in two samples and Cryptosporidium ubiquitum in one sample. This study is the first performing molecular characterisation of both parasites in Peruvian alpacas, and the first report of C. ubiquitum in this host. The identification of G. duodenalis assemblage A, C. parvum and C. ubiquitum, suggests that zoonotic transmission of these enteropathogens between alpacas and humans is possible.

  11. A new set of primers directed to 18S rRNA gene for molecular identification of Cryptosporidium spp. and their performance in the detection and differentiation of oocysts shed by synanthropic rodents.

    Science.gov (United States)

    Silva, Sheila O S; Richtzenhain, Leonardo J; Barros, Iracema N; Gomes, Alessandra M M C; Silva, Aristeu V; Kozerski, Noemila D; de Araújo Ceranto, Jaqueline B; Keid, Lara B; Soares, Rodrigo M

    2013-11-01

    Cryptosporidium spp. are cosmopolitan protozoa that infect fishes, reptiles, amphibians, birds and mammals. More than 20 species are recognized within this genus. Rodents are a group of abundant and ubiquitous organisms that have been considered reservoirs of Cryptosporidium for humans and livestock. The aim of this study was to design specific primers for the gene encoding 18S rRNA, potentially capable of amplifying any species or genotype of Cryptosporidium spp. and evaluate the diagnostic attributes of the nested-PCR based on such probes. The primers were designed to amplify the shortest segment as possible to maximize the sensitivity of the test, but preserving the discriminatory potential of the amplified sequences for phylogenetic inferences. The nested-PCR standardized in this study (nPCR-SH) was compared in terms of sensitivity with another similar assay (nPCR-XIAO) that has been largely used for the detection and identification of Cryptosporidium spp. worldwide. We also aimed to molecularly characterize samples of Cryptosporidum spp. isolated from synanthropic rodents using these probes. Forty-five rodents were captured in urban areas of the municipality of Umuarama, Paraná State, Brazil. Fecal samples were submitted to three molecular tests (nested-PCRs), two of them targeted to the 18S rDNA gene (nPCR-SH and nPCR-XIAO) and the third targeted to the gene encoding actin (nPCR-actin). The nPCR-SH was tested positive on samples of Cryptosporidum parvum, Cryptosporidum andersoni, Cryptosporidum meleagridis, Cryptosporidum hominis, Cryptosporidum canis, and Cryptosporidum serpentis. Sixteen samples of rodents were positive by nPCR-SH, six by nPCR-XIAO and five by nPCR-actin. Sequencing of amplified fragments allowed the identification of Cryptosporidum muris in three samples of Rattus rattus, and two genotypes of Cryptosporidium, the genotypes mouse II and III. Cryptosporidium genotype mouse II was found in one sample of Mus musculus and genotype mouse III

  12. The Structural Basis of Cryptosporidium-Specific IMP Dehydrogenase Inhibitor Selectivity

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    MacPherson, Iain S.; Kirubakaran, Sivapriya; Gorla, Suresh Kumar; Riera, Thomas V.; D’Aquino, J. Alejandro; Zhang, Minjia; Cuny, Gregory D.; Hedstrom, Lizbeth (BWH); (Brandeis)

    2010-03-29

    Cryptosporidium parvum is a potential biowarfare agent, an important AIDS pathogen, and a major cause of diarrhea and malnutrition. No vaccines or effective drug treatment exist to combat Cryptosporidium infection. This parasite relies on inosine 5{prime}-monophosphate dehydrogenase (IMPDH) to obtain guanine nucleotides, and inhibition of this enzyme blocks parasite proliferation. Here, we report the first crystal structures of CpIMPDH. These structures reveal the structural basis of inhibitor selectivity and suggest a strategy for further optimization. Using this information, we have synthesized low-nanomolar inhibitors that display 10{sup 3} selectivity for the parasite enzyme over human IMPDH2.

  13. Identification and differentiation of Cryptosporidium species by capillary electrophoresis single-strand conformation polymorphism.

    Science.gov (United States)

    Power, Michelle L; Holley, Marita; Ryan, Una M; Worden, Paul; Gillings, Michael R

    2011-01-01

    Cryptosporidium species generally lack distinguishing morphological traits, and consequently, molecular methods are commonly used for parasite identification. Various methods for Cryptosporidium identification have been proposed, each with their advantages and disadvantages. In this study, we show that capillary electrophoresis coupled with single-strand conformation polymorphism (CE-SSCP) is a rapid, simple and cost-effective method for the identification of Cryptosporidium species and genotypes. Species could be readily differentiated based on the SSCP mobility of amplified 18S rRNA gene molecules. Clones that differed by single-nucleotide polymorphisms could be distinguished on CE-SSCP mobility. Profiles of species known to have heterogenic copies of 18S rRNA gene contained multiple peaks. Cloning and sequencing of Cryptosporidium parvum, Cryptosporidium hominis, Cryptosporidium fayeri and Cryptosporidium possum genotype 18S rRNA gene amplicons confirmed that these multiple peaks represented type A and type B 18S rRNA gene copies. CE-SSCP provides a reliable and sensitive analysis for epidemiological studies, environmental detection and diversity screening.

  14. Household Socioeconomic and Demographic Correlates of Cryptosporidium Seropositivity in the United States.

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    Daniel J Becker

    Full Text Available Cryptosporidium are parasitic protozoa that infect humans, domestic animals, and wildlife globally. In the United States, cryptosporidiosis occurs in an estimated 750,000 persons annually, and is primarily caused by either of the Cryptosporidium parvum genotypes 1 and 2, exposure to which occurs through ingestion of food or water contaminated with oocytes shed from infected hosts. Although most cryptosporidiosis cases are caused by genotype 1 and are of human origin, the zoonotic sources of genotype 2, such as livestock, are increasingly recognized as important for understanding human disease patterns. Social inequality could mediate patterns of human exposure and infection by placing individuals in environments where food or water contamination and livestock contact is high or through reducing the availability of educational and sanitary resources required to avoid exposure.We here analyzed data from the National Health and Nutritional Examination Survey (NHANES between 1999 and 2000, and related seropositivity to Cryptosporidium parvum to correlates of social inequality at the household and individual scale. After accounting for the complex sampling design of NHANES and confounding by individual demographics and household conditions, we found impaired household food adequacy was associated with greater odds of Cryptosporidium seropositivity. Additionally, we identified individuals of non-white race and ethnicity and those born outside the United States as having significantly greater risk than white, domestic-born counterparts. Furthermore, we provide suggestive evidence for direct effects of family wealth on Cryptosporidium seropositivity, in that persons from low-income households and from families close to the poverty threshold had elevated odds of seropositivity relative to those in high-income families and in households far above the poverty line.These results refute assertions that cryptosporidiosis in the United States is independent of

  15. First molecular characterisation of Cryptosporidium and Giardia from Bubalus bubalis (water buffalo) in Victoria, Australia.

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    Abeywardena, Harshanie; Jex, Aaron R; von Samson-Himmelstjerna, Georg; Haydon, Shane R; Stevens, Melita A; Gasser, Robin B

    2013-12-01

    We conducted a molecular epidemiological survey of Cryptosporidium and Giardia from Bubalus bubalis (water buffalo) on two extensive farms (450 km apart) in Victoria, Australia. Faecal samples (n=476) were collected from different age groups of water buffalo at two time points (six months apart) and tested using a PCR-based mutation scanning-targeted sequencing-phylogenetic approach, employing markers within the small subunit of ribosomal RNA (designated pSSU) and triose phosphate isomerase (ptpi) genes. Based on pSSU data, Cryptosporidium parvum, Cryptosporidium bovis and Cryptosporidium genotypes 1, 2 (each 99% similar genetically to Cryptosporidium ryanae) and 3 (99% similar to Cryptosporidium suis) were detected in two (0.4%), one (0.2%), 38 (8.0%), 16 (3.4%) and one (0.2%) of the 476 samples tested, respectively. Using ptpi, Giardia duodenalis assemblages A and E were detected in totals of 56 (11.8%) and six (1.3%) of these samples, respectively. Cryptosporidium was detected on both farms, whereas Giardia was detected only on farm B, and both genera were detected in 1.5% of all samples tested. The study showed that water buffaloes on these farms excreted C. parvum and/or G. duodenalis assemblage A, which are consistent with those found in humans, inferring that these particular pathogens are of zoonotic significance. Future work should focus on investigating, in a temporal and spatial manner, the prevalence and intensity of such infections in water buffaloes in various geographical regions in Australia and in other countries.

  16. Adhesive-tape recovery combined with molecular and microscopic testing for the detection of Cryptosporidium oocysts on experimentally contaminated fresh produce

    Science.gov (United States)

    Cryptosporidium parvum suspended in water were applied to the surface of apples, cucumbers, peaches, and tomatoes at concentrations of 100, 50 or 10 oocysts within circles drawn with a permanent marker. Approximately 18 hr later, skin containing uncontaminated and contaminated circles from each prod...

  17. Assessment of zoonotic transmission of Giardia and Cryptosporidium between cattle and humans in rural villages in Bangladesh.

    Science.gov (United States)

    Ehsan, Amimul M; Geurden, Thomas; Casaert, Stijn; Parvin, Sonia M; Islam, Taohidul M; Ahmed, Uddin M; Levecke, Bruno; Vercruysse, Jozef; Claerebout, Edwin

    2015-01-01

    Giardia and Cryptosporidium are important causes of diarrhoea in Bangladesh. The high prevalence of both parasites in humans and cattle in rural Bangladesh and the common use of water ponds by village inhabitants and their animals suggest a potential for zoonotic transmission. Direct transmission of Giardia and Cryptosporidium between cattle and their handlers and indirect transmission through water ponds was investigated. Faecal/stool samples were collected from 623 calves and 125 calf handlers in a cross-sectional survey. In two villages, water samples were collected monthly from water ponds and faecal/stool samples were collected monthly from inhabitants and their cattle. Giardia cysts and Cryptosporidium oocysts were detected in water samples and in faecal/stool samples and positive samples were genotyped, to determine their human or animal origin. The prevalence of Giardia and Cryptosporidium in calves was 22% and 5% respectively. In calf handlers, the prevalence of Giardia and Cryptosporidium was 11.2% and 3.2% respectively. Both in the cross-sectional survey and in the longitudinal study in the villages, G. duodenalis assemblage E was most prevalent in calves, while in humans assemblage AII, BIII and BIV were found. In cattle, Cryptosporidium parvum, C. bovis and C. andersoni were identified, but no Cryptosporidium sequences were obtained from humans. Giardia and Cryptosporidium were detected in 14/24 and 12/24 water samples respectively. G. duodenalis assemblage E and BIV (-like), as well as C. andersoni and C. hominis were identified. Although the presence of Giardia and Cryptosporidium in both water ponds suggests that water-borne transmission of Giardia and Cryptosporidium is possible, the genotyping results indicate that there is no significant direct or indirect (water-borne) transmission of Giardia between cattle and people in this area of rural Bangladesh. No conclusions could be drawn for Cryptosporidium, because of the low number of sequences that

  18. Ureaplasma parvum infection alters filamin a dynamics in host cells

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    Brown Mary B

    2011-04-01

    Full Text Available Abstract Background Ureaplasmas are among the most common bacteria isolated from the human urogenital tract. Ureaplasmas can produce asymptomatic infections or disease characterized by an exaggerated inflammatory response. Most investigations have focused on elucidating the pathogenic potential of Ureaplasma species, but little attention has been paid to understanding the mechanisms by which these organisms are capable of establishing asymptomatic infection. Methods We employed differential proteome profiling of bladder tissues from rats experimentally infected with U. parvum in order to identify host cell processes perturbed by colonization with the microbe. Tissues were grouped into four categories: sham inoculated controls, animals that spontaneously cleared infection, asymptomatic urinary tract infection (UTI, and complicated UTI. One protein that was perturbed by infection (filamin A was used to further elucidate the mechanism of U. parvum-induced disruption in human benign prostate cells (BPH-1. BPH-1 cells were evaluated by confocal microscopy, immunoblotting and ELISA. Results Bladder tissue from animals actively colonized with U. parvum displayed significant alterations in actin binding proteins (profilin 1, vinculin, α actinin, and filamin A that regulate both actin polymerization and cell cytoskeletal function pertaining to focal adhesion formation and signal transduction (Fisher's exact test, P U. parvum perturbed the regulation of filamin A. Specifically, infected BPH-1 cells exhibited a significant increase in filamin A phosphorylated at serine2152 (P ≤ 0.01, which correlated with impaired proteolysis of the protein and its normal intracellular distribution. Conclusion Filamin A dynamics were perturbed in both models of infection. Phosphorylation of filamin A occurs in response to various cell signaling cascades that regulate cell motility, differentiation, apoptosis and inflammation. Thus, this phenomenon may be a useful

  19. Cryptosporidium species from human immunodeficiency-infected patients with chronic diarrhea in Jakarta, Indonesia.

    Science.gov (United States)

    Kurniawan, Agnes; Dwintasari, Sri W; Connelly, Lisa; Nichols, Rosely A B; Yunihastuti, Evy; Karyadi, Teguh; Djauzi, Samsuridjal

    2013-11-01

    Cryptosporidium is an opportunistic parasite that manifests as chronic and severe diarrhea in the immune-compromised subject. We investigated the species of Cryptosporidium to understand the epidemiology, mode of transmission, response to treatment, and prevention. Polymerase chain reaction/restriction fragment length polymorphism of the 18 S rRNA gene and sequencing were performed on 41 Cryptosporidium-positive stools from 36 patients with HIV AIDS, which comprised 36 pretreatment stools and 5 stools after treatment with Paromomycin. C. hominis, C. meleagridis, C. felis, and C. parvum were detected; 28 of 36 (77.7%) patients were infected with C. hominis and two (5.5%) patients with multiple species of Cryptosporidium. Treatment with Paromomycin resulted in different outcomes, perhaps because patients harbored other intestinal parasitic infections. Multiple infection with various Cryptosporidium species in the presence of other intestinal parasites occurs in patients with HIV AIDS suffering from chronic diarrhea who are severely immune-compromised. Common transmission of Cryptosporidium is anthroponotic. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. Occurrence of Giardia and Cryptosporidium in wild birds in Galicia (Northwest Spain).

    Science.gov (United States)

    Reboredo-Fernández, Aurora; Ares-Mazás, Elvira; Cacciò, Simone M; Gómez-Couso, Hipólito

    2015-06-01

    Faecal samples were obtained from 433 wild birds being treated in wildlife recovery centres in Galicia (Northwest Spain), between February 2007 and September 2009. The birds belonged to 64 species representing 17 different orders. Giardia cysts and Cryptosporidium oocysts were detected by an immunofluorescence antibody test and identified at the molecular level by established PCR-sequencing methods. The overall prevalence of Giardia was 2·1% and that of Cryptosporidium, 8·3%. To our knowledge, this is the first description of Giardia sp. in Tyto alba and Caprimulgus europaeus; and of Cryptosporidium sp. in Apus apus, Athene noctua, C. europaeus, Falco tinnunculus, Morus bassanus, Parabuteo unicinctus and Strix aluco. Furthermore, the first PCR-sequence confirmed detection of Giardia duodenalis assemblage B in, Buteo buteo, Coturnix coturnix and Pica pica; G. duodenalis assemblage D in Garrulus glandarius; and G. duodenalis assemblage F in Anas platyrhynchos; Cryptosporidium parvum in Accipiter nisus, B. buteo, Milvus migrans, Pernis apivorus and P. pica; and Cryptosporidium meleagridis in Streptopelia turtur. The study findings demonstrate the wide spread of Giardia and Cryptosporidium between wild birds.

  1. Molecular-based investigation of Cryptosporidium and Giardia from animals in water catchments in southeastern Australia.

    Science.gov (United States)

    Nolan, Matthew J; Jex, Aaron R; Koehler, Anson V; Haydon, Shane R; Stevens, Melita A; Gasser, Robin B

    2013-04-01

    There has been no large-scale systematic molecular epidemiological investigation of the waterborne protozoans, Cryptosporidium or Giardia, in southeastern Australia. Here, we explored, for the first time, the genetic composition of these genera in faecal samples from animals in nine Melbourne Water reservoir areas, collected over a period of two-years. We employed PCR-based single-strand conformation polymorphism (SSCP) and phylogenetic analyses of loci (pSSU and pgp60) in the small subunit (SSU) of ribosomal RNA and 60-kDa glycoprotein (gp60) genes to detect and characterise Cryptosporidium, and another locus (ptpi) in the triose-phosphate isomerase (tpi) gene to identify and characterise Giardia. Cryptosporidium was detected in 2.8% of the 2009 samples examined; the analysis of all amplicons defined 14 distinct sequence types for each of pSSU and pgp60, representing Cryptosporidium hominis (genotype Ib - subgenotype IbA10G2R2), Cryptosporidium parvum (genotype IIa - subgenotypes IIaA15G2R1, IIaA19G2R1, IIaA19G3R1, IIaA19G4R1, IIaA20G3R1, IIaA20G4R1, IIaA20G3R2 and IIaA21G3R1), Cryptosporidium cuniculus (genotype Vb - subgenotypes VbA22R4, VbA23R3, VbA24R3, VbA25R4 and VbA26R4), and Cryptosporidium canis, Cryptosporidium fayeri, Cryptosporidium macropodum and Cryptosporidium ubiquitum as well as six new pSSU sequence types. In addition, Giardia was identified in 3.4% of the samples; all 28 distinct ptpi sequence types defined were linked to assemblage A of Giardia duodenalis. Of all 56 sequence types characterised, eight and one have been recorded previously in Cryptosporidium and Giardia, respectively, from humans. In contrast, nothing is known about the zoonotic potential of 35 new genotypes of Cryptosporidium and Giardia recorded here for the first time. Future work aims to focus on estimating the prevalence of Cryptosporidium and Giardia genotypes in humans and a wide range of animals in Victoria and elsewhere in Australia. (Nucleotide sequences reported in

  2. Mussels (Perna perna as bioindicator of environmental contamination by Cryptosporidium species with zoonotic potential

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    Geisi Ferreira Mariné Oliveira

    2016-04-01

    Full Text Available Sources of contamination such as animal feces runoff, organic fertilizer application, and the release of partially treated or untreated sewage can lead to the contamination of aquatic environments by Cryptosporidium spp. The quality of mussels as food is closely related to the sanitary conditions of the marine environment where these bivalves are found. Marine mollusks are filter feeders that are able to retain Cryptosporidium oocysts in their tissue, thus functioning as bioindicators. A total of 72 pooled mussel samples of the species Perna perna were collected at two sites (A and B in the municipality of Mangaratiba, Rio de Janeiro State, Brazil. Sampling involved removal of 30 mussels, from each collection site every month for one year. The 30 mussels from each sampling were then allocated into three groups of 10. Two Cryptosporidium spp. genes (18S and GP60 were targeted for DNA amplification from the samples obtained. After purification, all of the products obtained were sequenced and phylogenetic analyses were performed. Of the 72 samples analyzed using the nested-PCR for the 18S gene target, 29.2% were positive for the presence of Cryptosporidium spp. Of these samples, 52.4% were collected at site A (ie 11/21 and 47.6% at site B (ie 10/21. The 18S genes of all the samples considered positive for Cryptosporidium spp. were sequenced, and the following three species were identified: Cryptosporidium parvum, C. meleagridis, and C. andersoni. Three distinct C. parvum subtypes (IIaA19G2R2; IIaA20G2R2; IIaA20G3R2 were identified using the GP60 gene. More studies to evaluate the zoonotic potential of this species should be performed as both sampling locations contain human and/or animal fecal contaminants.

  3. Mussels (Perna perna) as bioindicator of environmental contamination by Cryptosporidium species with zoonotic potential.

    Science.gov (United States)

    Mariné Oliveira, Geisi Ferreira; do Couto, Melissa Carvalho Machado; de Freitas Lima, Marcelo; do Bomfim, Teresa Cristina Bergamo

    2016-04-01

    Sources of contamination such as animal feces runoff, organic fertilizer application, and the release of partially treated or untreated sewage can lead to the contamination of aquatic environments by Cryptosporidium spp. The quality of mussels as food is closely related to the sanitary conditions of the marine environment where these bivalves are found. Marine mollusks are filter feeders that are able to retain Cryptosporidium oocysts in their tissue, thus functioning as bioindicators. A total of 72 pooled mussel samples of the species Perna perna were collected at two sites (A and B) in the municipality of Mangaratiba, Rio de Janeiro State, Brazil. Sampling involved removal of 30 mussels, from each collection site every month for one year. The 30 mussels from each sampling were then allocated into three groups of 10. Two Cryptosporidium spp. genes (18S and GP60) were targeted for DNA amplification from the samples obtained. After purification, all of the products obtained were sequenced and phylogenetic analyses were performed. Of the 72 samples analyzed using the nested-PCR for the 18S gene target, 29.2% were positive for the presence of Cryptosporidium spp. Of these samples, 52.4% were collected at site A (ie 11/21) and 47.6% at site B (ie 10/21). The 18S genes of all the samples considered positive for Cryptosporidium spp. were sequenced, and the following three species were identified: Cryptosporidium parvum, C. meleagridis, and C. andersoni. Three distinct C. parvum subtypes (IIaA19G2R2; IIaA20G2R2; IIaA20G3R2) were identified using the GP60 gene. More studies to evaluate the zoonotic potential of this species should be performed as both sampling locations contain human and/or animal fecal contaminants.

  4. First genetic analysis of Cryptosporidium from humans from Tasmania, and identification of a new genotype from a traveller to Bali.

    Science.gov (United States)

    Koehler, Anson V; Whipp, Margaret; Hogg, Geoff; Haydon, Shane R; Stevens, Melita A; Jex, Aaron R; Gasser, Robin B

    2014-09-01

    Little is known about the molecular composition of Cryptosporidium species from humans living in the insular state of Tasmania, Australia. In the present study, we genetically characterized 82 samples of Cryptosporidium from humans following conventional coproscopic testing in a routine, diagnostic laboratory. Using a PCR-coupled single-strand conformation polymorphism (SSCP) technique, targeting portions of the small subunit rRNA (SSU), and 60 kDa glycoprotein (gp60) loci, we identified two species of Cryptosporidium, including C. hominis (subgenotypes IbA10G2, IdA16, IeA12G3T3, and IfA19G1) and C. parvum (IIaA16G1R1 and IIaA18G3), and a new operational taxonomic unit (OTU) that genetically closely resembled C. wrairi. This OTU was further characterized using markers in the actin, Cryptosporidium oocyst wall protein (COWP), and 70 kDa heat shock protein (hsp70) genes. This study provides the first characterization of species and genotypes of Cryptosporidium from Tasmania, and presents clear genetic evidence, using five independent genetic loci, for a new genotype or species of Cryptosporidium in a Tasmanian person with a recent history of travelling to Bali, Indonesia. It would be interesting to undertake detailed molecular-based studies of Cryptosporidium in Indonesia and neighbouring countries, in conjunction with morphological and experimental investigations of new genotypes.

  5. Prevalence, environmental loading, and molecular characterization of Cryptosporidium and Giardiaisolates from domestic and wild animals along the Central California Coast

    Science.gov (United States)

    Oates, Stori C; Miller, Melissa A.; Hardin, Dane; Conrad, Patricia A.; Melli, Ann; Jessup, David A.; Dominik, Clare; Roug, Annette; Tinker, M. Tim; Miller, Woutrina A.

    2012-01-01

    The risk of disease transmission from waterborne protozoa is often dependent on the origin (e.g., domestic animals versus wildlife), overall parasite load in contaminated waterways, and parasite genotype, with infections being linked to runoff or direct deposition of domestic animal and wildlife feces. Fecal samples collected from domestic animals and wildlife along the central California coast were screened to (i) compare the prevalence and associated risk factors for fecal shedding of Cryptosporidium and Giardia species parasites, (ii) evaluate the relative importance of animal host groups that contribute to pathogen loading in coastal ecosystems, and (iii) characterize zoonotic and host-specific genotypes. Overall, 6% of fecal samples tested during 2007 to 2010 were positive for Cryptosporidium oocysts and 15% were positive for Giardia cysts. Animal host group and age class were significantly associated with detection of Cryptosporidium and Giardia parasites in animal feces. Fecal loading analysis revealed that infected beef cattle potentially contribute the greatest parasite load relative to other host groups, followed by wild canids. Beef cattle, however, shed host-specific, minimally zoonotic Cryptosporidium and Giardia duodenalis genotypes, whereas wild canids shed potentially zoonotic genotypes, including G. duodenalis assemblages A and B. Given that the parasite genotypes detected in cattle were not zoonotic, the public health risk posed by protozoan parasite shedding in cattle feces may be lower than that posed by other animals, such as wild canids, that routinely shed zoonotic genotypes.

  6. Cryptosporidium genotypes in children and calves living at the wildlife or livestock interface of the Kruger National Park, South Africa.

    Science.gov (United States)

    Abu Samra, Nada; Jori, Ferran; Cacciò, Simone M; Frean, John; Poonsamy, Bhavani; Thompson, Peter N

    2016-05-20

    Cryptosporidium infection is one of the most common causes of parasitic diarrhoea worldwide in cattle and humans. In developing countries, human cryptosporidiosis is most prevalent during early childhood and links between zoonotic infection and animal related activities have been demonstrated. This study investigated the prevalence and species/genotype distribution of Cryptosporidium among children (Kruger National Park in South Africa, where interactions between humans and wild and domestic animals are known to occur. Cryptosporidium oocysts were detected in 8/143 stool samples of children recruited within the hospital system (5.6%; 95% CI 2.4%, 10.7%) and in 2/352 faecal samples of calves (0.6%; 95% CI 0.1%, 2.0%) using the modified Ziehl-Neelsen (MZN) staining technique. Microscopy positive samples from children were further analysed by PCR targeting the 18S rRNA gene and identified as Cryptosporidium hominis (3/4) and Cryptosporidium meleagridis (1/4). Regardless of the microscopy outcome, randomly selected samples (n = 36) from calves 0-4 months of age were amplified and sequenced at the 18S rRNA gene using nested PCR. Two calves tested positive (5.6%; 95% CI 1.7%, 18.7%), and revealed the presence of Cryptosporidium parvum and Cryptosporidium bovis. The detection of only two zoonotic species (C. parvum in one calf and C. meleagridis in one child) suggests that zoonotic cryptosporidiosis is not currently widespread in our study area; however, the potential exists for amplification of transmission in an immunocompromised population.

  7. Identification of Cryptosporidium species and genotypes in Scottish raw and drinking waters during a one-year monitoring period.

    Science.gov (United States)

    Nichols, R A B; Connelly, L; Sullivan, C B; Smith, H V

    2010-09-01

    We analyzed 1,042 Cryptosporidium oocyst-positive slides (456 from raw waters and 586 from drinking waters) of which 55.7% contained 1 or 2 oocysts, to determine species/genotypes present in Scottish waters. Two nested PCR-restriction fragment length polymorphism (RFLP) assays targeting different loci (1 and 2) of the hypervariable region of the 18S rRNA gene were used for species identification, and 62.4% of samples were amplified with at least one of the PCR assays. More samples (577 slides; 48.7% from raw water and 51.3% from drinking water) were amplified at locus 1 than at locus 2 (419 slides; 50.1% from raw water and 49.9% from drinking water). PCR at loci 1 and 2 amplified 45.4% and 31.7% of samples containing 1 or 2 oocysts, respectively. We detected both human-infectious and non-human-infectious species/genotype oocysts in Scottish raw and drinking waters. Cryptosporidium andersoni, Cryptosporidium parvum, and the Cryptosporidium cervine genotype (now Cryptosporidium ubiquitum) were most commonly detected in both raw and drinking waters, with C. ubiquitum being most common in drinking waters (12.5%) followed by C. parvum (4.2%) and C. andersoni (4.0%). Numerous samples (16.6% total; 18.9% from drinking water) contained mixtures of two or more species/genotypes, and we describe strategies for unraveling their identity. Repetitive analysis for discriminating mixtures proved useful, but both template concentration and PCR assay influenced outcomes. Five novel Cryptosporidium spp. (SW1 to SW5) were identified by RFLP/sequencing, and Cryptosporidium sp. SW1 was the fourth most common contaminant of Scottish drinking water (3%).

  8. The Burden of Cryptosporidium Diarrheal Disease among Children < 24 Months of Age in Moderate/High Mortality Regions of Sub-Saharan Africa and South Asia, Utilizing Data from the Global Enteric Multicenter Study (GEMS.

    Directory of Open Access Journals (Sweden)

    Samba O Sow

    2016-05-01

    Full Text Available The importance of Cryptosporidium as a pediatric enteropathogen in developing countries is recognized.Data from the Global Enteric Multicenter Study (GEMS, a 3-year, 7-site, case-control study of moderate-to-severe diarrhea (MSD and GEMS-1A (1-year study of MSD and less-severe diarrhea [LSD] were analyzed. Stools from 12,110 MSD and 3,174 LSD cases among children aged <60 months and from 21,527 randomly-selected controls matched by age, sex and community were immunoassay-tested for Cryptosporidium. Species of a subset of Cryptosporidium-positive specimens were identified by PCR; GP60 sequencing identified anthroponotic C. parvum. Combined annual Cryptosporidium-attributable diarrhea incidences among children aged <24 months for African and Asian GEMS sites were extrapolated to sub-Saharan Africa and South Asian regions to estimate region-wide MSD and LSD burdens. Attributable and excess mortality due to Cryptosporidium diarrhea were estimated.Cryptosporidium was significantly associated with MSD and LSD below age 24 months. Among Cryptosporidium-positive MSD cases, C. hominis was detected in 77.8% (95% CI, 73.0%-81.9% and C. parvum in 9.9% (95% CI, 7.1%-13.6%; 92% of C. parvum tested were anthroponotic genotypes. Annual Cryptosporidium-attributable MSD incidence was 3.48 (95% CI, 2.27-4.67 and 3.18 (95% CI, 1.85-4.52 per 100 child-years in African and Asian infants, respectively, and 1.41 (95% CI, 0.73-2.08 and 1.36 (95% CI, 0.66-2.05 per 100 child-years in toddlers. Corresponding Cryptosporidium-attributable LSD incidences per 100 child-years were 2.52 (95% CI, 0.33-5.01 and 4.88 (95% CI, 0.82-8.92 in infants and 4.04 (95% CI, 0.56-7.51 and 4.71 (95% CI, 0.24-9.18 in toddlers. We estimate 2.9 and 4.7 million Cryptosporidium-attributable cases annually in children aged <24 months in the sub-Saharan Africa and India/Pakistan/Bangladesh/Nepal/Afghanistan regions, respectively, and ~202,000 Cryptosporidium-attributable deaths (regions combined. ~59

  9. Identifying host sources, human health risk and indicators of Cryptosporidium and Giardia in a Canadian watershed influenced by urban and rural activities.

    Science.gov (United States)

    Van Dyke, Michele I; Ong, Corinne S L; Prystajecky, Natalie A; Isaac-Renton, Judith L; Huck, Peter M

    2012-06-01

    Cryptosporidium and Giardia were characterized in a watershed in southern Ontario, Canada, over a 2½ year period. River samples were collected every two weeks, primarily near a municipal drinking water treatment plant intake. Cryptosporidium and Giardia were frequently detected with an overall occurrence rate of 88 and 97%, respectively. Giardia concentrations were higher than Cryptosporidium, with median values of 80 cysts 100 L(-1) and 12 oocysts 100 L(-1), respectively. Although pathogens rarely show a significant relationship with fecal or water quality indicators, this study determined that Cryptosporidium, but not Giardia, was significantly correlated with Escherichia coli, turbidity and river flow. There was no correlation between the two types of protozoa, and only Giardia showed a seasonal trend with higher concentrations at cold water temperatures. Cryptosporidium genotyping of all samples found that farm animals and wildlife were an important contributor of oocysts in the watershed, and that Cryptosporidium strains/genotypes of medium to high risk for human infection (C. hominis, C. parvum and C. ubiquitum) were detected in 16% of samples. This study was able to identify Cryptosporidium host sources and human health risk, and to identify differences between Cryptosporidium and Giardia occurrence in the watershed.

  10. Prevalence and risk factors associated with Cryptosporidium spp. infection in young domestic livestock in India.

    Science.gov (United States)

    Maurya, Prem Sagar; Rakesh, Radhamma Lakshmipathy; Pradeep, Balaraju; Kumar, Saroj; Kundu, Krishnendu; Garg, Rajat; Ram, Hira; Kumar, Ashok; Banerjee, Partha Sarathi

    2013-04-01

    A total of 938 faecal samples (461 cattle calves, 264 buffalo calves, 55 lambs, 116 kids and 42 piglets) from different livestock farms and individual small holdings in six targeted states of India were collected and screened by modified Ziehl-Neelsen staining technique to determine the prevalence of Cryptosporidium spp. and its association with age, sex, season and faecal consistency in domesticated animals. Overall, 16.2 % of the animals were positive for Cryptosporidium infection with prevalence of 16.3, 24.2, 1.8, 3.5 and 19.1 % in cattle calves, buffalo calves, lambs, kids and piglets, respectively. The prevalence of infection was significantly higher (p0.05) was recorded in females than in males. Seasons had a significant effect (p0.05) in post-monsoon than in monsoon season. A high degree of association was noticed between Cryptosporidium infection and diarrhoea in ruminants screened during the present study. But, in case of pigs, the prevalence was higher in non-diarrhoeic than in diarrhoeic animals. Genotyping of Cryptosporidium spp. based on nested PCR amplification of partial 18S rRNA and its subsequent digestion with SspI, VspI and MboII restriction enzymes revealed prevalence of Cryptosporidium parvum in representative number of positive samples of cattle, buffalo and goats.

  11. Analytical sensitivity of staining and molecular techniques for the detection of Cryptosporidium spp. oocysts isolated from bovines in water samples: a preliminary study

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    A Díaz-Lee

    2015-01-01

    Full Text Available Cryptosporidium spp. is a globally distributed protozoan that causes digestive disease in different animals including humans. Excreted oocysts contaminate water and soil, constituting a public health threat. Sensitive and fast methods to detect oocysts in water samples are necessary due to the small number of oocysts present in the environment and their low infectious dose. This study compared the analytical sensitivity of two staining techniques, modified Ziehl-Neelsen and Auramine versus a nested PCR that amplifies a region of ~520bp from 18S rDNA gene, to detect Cryptosporidium spp. in water samples. Water was inoculated with oocysts using serial dilutions, and then a water filtration method was used to recover the parasite oocysts. The staining techniques had similar analytical sensitivity, detecting 8 oocysts/mL, while the nested PCR detected down to 6 oocysts/mL. In conclusion, all of these methods are effective for Cryptosporidium spp. detection in water samples, contributing to the implementation of standardized diagnostic methods for environmental water infectious agents.

  12. Diversity of Cryptosporidium species occurring in sheep and goat breeds reared in Poland.

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    Kaupke, Agnieszka; Michalski, Mirosław M; Rzeżutka, Artur

    2017-03-01

    The aim of this study was molecular identification of Cryptosporidium species and assessment of their prevalence in different breeds of sheep and goat reared in Poland. In addition, the relationship between animal age, breed type, and the frequency of Cryptosporidium infections was determined. Fecal samples from 234 lambs and 105 goat kids aged up to 9 weeks, representing 24 breeds and their cross-breeds were collected from 71 small ruminant farms across Poland. The identification of Cryptosporidium species was performed at the 18 SSU ribosomal RNA (rRNA) and COWP loci followed by subtyping of C. parvum and C. hominis strains at GP60 gene locus. The presence of Cryptosporidium DNA at the 18 SSU rRNA locus was detected in 45/234 (19.2%) lamb feces samples and in 39/105 (37.1%) taken from goats. The following Cryptosporidium species: C. xiaoi, C. bovis, C. ubiquitum, C. parvum, and C. hominis were detected in small ruminants. Infections caused by C. xiaoi were predominant without favoring any tested animal species. Subsequent GP60 subtyping revealed the presence of C. parvum IIaA17G1R1 subtype in sheep and IIdA23G1 subtype in goats. IIdA23G1 subtype was detected in a goat host for the first time. There were no significant differences found in frequency of infections between the age groups ( 0.05) or goat kids (P = 0.06, α > 0.05). In addition, there was no correlation observed between the frequency in occurrence of particular parasite species and breed type in relation to native sheep breeds (F = 0.11; P = 0.990 > 0.05). In the case of goats, more breed-related differences in parasite occurrence were found. The results of this study improve our knowledge on the breed-related occurrence of Cryptosporidium infections in the population of small ruminants reared in Poland.

  13. Cryptosporidium genotypes in children and calves living at the wildlife or livestock interface of the Kruger National Park, South Africa

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    Nada Abu Samra

    2016-03-01

    Full Text Available Cryptosporidium infection is one of the most common causes of parasitic diarrhoea worldwide in cattle and humans. In developing countries, human cryptosporidiosis is most prevalent during early childhood and links between zoonotic infection and animal related activities have been demonstrated. This study investigated the prevalence and species/genotype distribution of Cryptosporidium among children (< 5 years and calves (< 6 months living in a rural farming area adjacent to the Kruger National Park in South Africa, where interactions between humans and wild and domestic animals are known to occur. Cryptosporidium oocysts were detected in 8/143 stool samples of children recruited within the hospital system (5.6%; 95% CI 2.4%, 10.7% and in 2/352 faecal samples of calves (0.6%; 95% CI 0.1%, 2.0% using the modified Ziehl–Neelsen (MZN staining technique. Microscopy positive samples from children were further analysed by PCR targeting the 18S rRNA gene and identified as Cryptosporidium hominis (3/4 and Cryptosporidium meleagridis (1/4. Regardless of the microscopy outcome, randomly selected samples (n = 36 from calves 0–4 months of age were amplified and sequenced at the 18S rRNA gene using nested PCR. Two calves tested positive (5.6%; 95% CI 1.7%, 18.7%, and revealed the presence of Cryptosporidium parvum and Cryptosporidium bovis. The detection of only two zoonotic species (C. parvum in one calf and C. meleagridis in one child suggests that zoonotic cryptosporidiosis is not currently widespread in our study area; however, the potential exists for amplification of transmission in an immunocompromised population.Keywords: Cryptosporidium; children; calves; South Africa; genotyping; GP60 subtyping

  14. Evolution of mitosome metabolism and invasion-related proteins in Cryptosporidium.

    Science.gov (United States)

    Liu, Shiyou; Roellig, Dawn M; Guo, Yaqiong; Li, Na; Frace, Michael A; Tang, Kevin; Zhang, Longxian; Feng, Yaoyu; Xiao, Lihua

    2016-12-08

    The switch from photosynthetic or predatory to parasitic life strategies by apicomplexans is accompanied with a reductive evolution of genomes and losses of metabolic capabilities. Cryptosporidium is an extreme example of reductive evolution among apicomplexans, with losses of both the mitosome genome and many metabolic pathways. Previous observations on reductive evolution were largely based on comparative studies of various groups of apicomplexans. In this study, we sequenced two divergent Cryptosporidium species and conducted a comparative genomic analysis to infer the reductive evolution of metabolic pathways and differential evolution of invasion-related proteins within the Cryptosporidium lineage. In energy metabolism, Cryptosporidium species differ from each other mostly in mitosome metabolic pathways. Compared with C. parvum and C. hominis, C. andersoni possesses more aerobic metabolism and a conventional electron transport chain, whereas C. ubiquitum has further reductions in ubiquinone and polyisprenoid biosynthesis and has lost both the conventional and alternative electron transport systems. For invasion-associated proteins, similar to C. hominis, a reduction in the number of genes encoding secreted MEDLE and insulinase-like proteins in the subtelomeric regions of chromosomes 5 and 6 was also observed in C. ubiquitum and C. andersoni, whereas mucin-type glycoproteins are highly divergent between the gastric C. andersoni and intestinal Cryptosporidium species. Results of the study suggest that rapidly evolving mitosome metabolism and secreted invasion-related proteins could be involved in tissue tropism and host specificity in Cryptosporidium spp. The finding of progressive reduction in mitosome metabolism among Cryptosporidium species improves our knowledge of organelle evolution within apicomplexans.

  15. First molecular characterization of Cryptosporidium from three different points of two main rivers in Kuantan, Malaysia using 18S rRNA gene nested PCR

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    Mohd Aiman Barudin

    2017-09-01

    Full Text Available Objective: To identify 18S ribosomal RNA (18S rRNA partial sequences from three different points, namely, downstream, midstream and upstream of two major rivers in Kuantan, Pahang, Malaysia. Methods: In this study, six river water samples were collected from three different points of both Kuantan River and Balok River. Each water sample was processed and concentrated using continuous flow centrifuge machine and purified using immunomagnetic separation technique. Nested PCR was applied to detect 18S rRNA sequence in those samples after DNA extraction. Genotyping and phylogenetic analysis were carried out based on the gene of 18S rRNA sequence of Cryptosporidium. Results: A total of five samples from six different points of both Kuantan River and Balok River were contaminated with Cryptosporidium. Only river water sample from the upstream point of Kuantan River was negative for Cryptosporidium. In this study, the sequencing results of five samples from both Kuantan River and Balok River belonged to Cryptosporidium parvum. Conclusions: This is the first study of Cryptosporidium parvum genotyping in both Kuantan River and Balok River by using molecular approach nested PCR on 18S rRNA gene.

  16. Pathological features of Cryptosporidium andersoni-induced lesions in SCID mice.

    Science.gov (United States)

    Masuno, Koichi; Yanai, Tokuma; Sakai, Hiroki; Satoh, Masaaki; Kai, Chieko; Nakai, Yutaka

    2013-07-01

    To assess the infectivity and the istopathological features of Cryptosporidium andersoni (C. andersoni) in laboratory animals, SCID mice were orally inoculated with oocysts of C. andersoni. Starting one week after inoculation, the SCID mice began shedding oocysts, and this continued for ten weeks. Histopathologically, myriads of C. andersoni were observed on the apical surface of the epithelium in the gastric pit of the glandular stomach. There were few lesions in the gastric epithelium except C. andersoni adhesion. In the lamina propria of the affected mucosa, minimum infiltration of inflammatory cells was observed. Immunohistochemically, C. andersoni demonstrated a positive reaction to a number of primary antibodies of Cryptosporidium parvum. In the experiment described here, few increases were seen in apoptotic epithelial cells in the affected mucosas of the SCID mice, and the nuclear augmentation was not enhanced. It was hypothesized that the absence of apoptosis and cell division were due to a lack of inflammatory cell reaction in the lamina propria.

  17. The first report of Cryptosporidium andersoni in horses with diarrhea and multilocus subtype analysis.

    Science.gov (United States)

    Liu, Aiqin; Zhang, Jia; Zhao, Jingmin; Zhao, Wei; Wang, Rongjun; Zhang, Longxian

    2015-09-22

    Horses interact with humans in a wide variety of sport competitions and non-competitive recreational pursuits as well as in working activities. Cryptosporidium spp are one of the most important zoonotic pathogens causing diarrhea of humans and animals. The reports of Cryptosporidium in horses and the findings of zoonotic Cryptosporidium species/genotypes show a necessity to carry out molecular identification of Cryptosporidium in horses, especially in diarrheic ones. The aim of the present study was to understand Cryptosporidium infection and species/genotypes in diarrheic horses, and to trace the source of infection of horse-derived Cryptosporidium isolates at a subtype level. Fecal specimens of 29 diarrheic adult horses were collected in Taikang County in northeastern China's Heilongjiang Province. Cryptosporidium oocysts were concentrated by Sheather's sugar flotation technique, and then examined by a bright-field microscope. Meanwhile, all the specimens were subjected to PCR amplification of the small subunit (SSU) rRNA gene of Cryptosporidium. C. andersoni isolates were further subtyped by multilocus sequence typing (MLST) at the four microsatellite/minisatellite loci (MS1, MS2, MS3 and MS16). One and two Cryptosporidium-positive isolates were obtained in horses by microscopy and by PCR, respectively. The two C. andersoni isolates were identified by sequencing of the SSU rRNA gene of Cryptosporidium. Both of them were identical to each other at the MS1, MS2, MS3 and MS16 loci, and MLST subtype A4,A4,A4,A1 was found here. This is the first report of C. andersoni in horses. The fact that the MLST subtype A4,A4,A4,A1 was reported in cattle suggests a large possibility of transmission of C. andersoni between cattle and horses.

  18. Genetic diversity of Cryptosporidium identified in clinical samples from cities in Brazil and Argentina

    Science.gov (United States)

    Peralta, Regina Helena Saramago; Velásquez, Jorge Néstor; Cunha, Flavia de Souza; Pantano, María Laura; Sodré, Fernando Campos; da Silva, Sidnei; Astudillo, Osvaldo Germán; Peralta, José Mauro; Carnevale, Silvana

    2016-01-01

    The identification and characterisation of Cryptosporidiumgenotypes and subtypes are fundamental to the study of cryptosporidiosis epidemiology, aiding in prevention and control strategies. The objective was to determine the genetic diversity ofCryptosporidium in samples obtained from hospitals of Rio de Janeiro, Brazil, and Buenos Aires, Argentina. Samples were analysed by microscopy and TaqMan polymerase chain reaction (PCR) assays forCryptosporidium detection, genotyped by nested-PCR-restriction fragment length polymorphism (RFLP) analysis of the 18S rRNA gene and subtyped by DNA sequencing of the gp60 gene. Among the 89 samples from Rio de Janeiro, Cryptosporidium spp were detected in 26 by microscopy/TaqMan PCR. In samples from Buenos Aires,Cryptosporidium was diagnosed in 15 patients of the 132 studied. The TaqMan PCR and the nested-PCR-RFLP detected Cryptosporidium parvum, Cryptosporidium hominis, and co-infections of both species. In Brazilian samples, the subtypes IbA10G2 and IIcA5G3 were observed. The subtypes found in Argentinean samples were IbA10G2, IaA10G1R4, IaA11G1R4, and IeA11G3T3, and mixed subtypes of Ia and IIa families were detected in the co-infections. C. hominis was the species more frequently detected, and subtype family Ib was reported in both countries. Subtype diversity was higher in Buenos Aires than in Rio de Janeiro and two new subtypes were described for the first time. PMID:26814641

  19. Genetic diversity of Cryptosporidium identified in clinical samples from cities in Brazil and Argentina

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    Regina Helena Saramago Peralta

    2016-01-01

    Full Text Available The identification and characterisation of Cryptosporidiumgenotypes and subtypes are fundamental to the study of cryptosporidiosis epidemiology, aiding in prevention and control strategies. The objective was to determine the genetic diversity ofCryptosporidium in samples obtained from hospitals of Rio de Janeiro, Brazil, and Buenos Aires, Argentina. Samples were analysed by microscopy and TaqMan polymerase chain reaction (PCR assays forCryptosporidium detection, genotyped by nested-PCR-restriction fragment length polymorphism (RFLP analysis of the 18S rRNA gene and subtyped by DNA sequencing of the gp60 gene. Among the 89 samples from Rio de Janeiro, Cryptosporidium spp were detected in 26 by microscopy/TaqMan PCR. In samples from Buenos Aires,Cryptosporidium was diagnosed in 15 patients of the 132 studied. The TaqMan PCR and the nested-PCR-RFLP detected Cryptosporidium parvum, Cryptosporidium hominis, and co-infections of both species. In Brazilian samples, the subtypes IbA10G2 and IIcA5G3 were observed. The subtypes found in Argentinean samples were IbA10G2, IaA10G1R4, IaA11G1R4, and IeA11G3T3, and mixed subtypes of Ia and IIa families were detected in the co-infections. C. hominis was the species more frequently detected, and subtype family Ib was reported in both countries. Subtype diversity was higher in Buenos Aires than in Rio de Janeiro and two new subtypes were described for the first time.

  20. Eimeria and Cryptosporidium in Estonian dairy farms in regard to age, species, and diarrhoea.

    Science.gov (United States)

    Lassen, Brian; Viltrop, Arvo; Raaperi, Kerli; Järvis, Toivo

    2009-12-23

    Eimeria and Cryptosporidium are among the most common bovine parasites in the world, but little is known about them in Estonia. Basic field research is needed to gain insight into pathogen dynamics, providing knowledge for veterinarians and research. A survey of 45 Estonian dairy farms in 15 counties was carried out between 2006 and 2007. Three age groups: 12 months old animals were sampled. Collected faeces were examined by quantitative flotation and Ziehl-Neelsen contrast staining, and species examined morphologically. Selected samples containing Cryptosporidium were additionally examined by polymerase-chain-reaction (PCR) and sequencing to determine genotypes. Twelve species of Eimeria were identified, seven previously unknown in Estonia. Main species in samples were E. bovis (30%), E. zuernii (23%), and E. ellipsoidalis (14%). All herds were infected and animals aged 3-12 months were more commonly infected with Eimeria oocysts (63%) than any other group. Calves Eimeria and the age category. Cryptosporidium were detected in 84% of the farms, and C. andersoni and C. parvum were successfully identified. Though prevalences of Cryptosporidium in the age groups were similar to the sample prevalence (30%) an increase in the infections was found with increasing age (pEimeria spp. infection (slope=-0.08, pEimeria species.

  1. Development of a Cryptosporidium oocyst assay using an automated fiber optic-based biosensor

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    Kramer Marianne F

    2007-10-01

    Full Text Available Abstract An intestinal protozoan parasite, Cryptosporidium parvum, is a major cause of waterborne gastrointestinal disease worldwide. Detection of Cryptosporidium oocysts in potable water is a high priority for the water treatment industry to reduce potential outbreaks among the consumer populace. Anti-Cryptosporidium oocyst polyclonal and monoclonal antibodies were tested as capture and detection reagents for use in a fiber optic biosensor assay for the detection of Cryptosporidium oocysts. Antibodies were validated using enzyme-linked immunosorbent assays, flow cytometry, Western blotting and fluorescent microscopy. Oocysts could be detected at a concentration of 105 oocysts/ml when the polyclonal antibodies were used as the capture and detection reagents. When oocysts were boiled prior to detection, a ten-fold increase in sensitivity was achieved using the polyclonal antibody. Western blotting and immunofluorescence revealed that both the monoclonal and polyclonal antibodies recognize a large (>300 kDa molecular weight mucin-like antigen present on the surface of the oocyst wall. The polyclonal antibody also reacted with a small (105 kDa molecular weight antigen that was present in boiled samples of oocysts. Preliminary steps to design an in-line biosensor assay system have shown that oocysts would have to be concentrated from water samples and heat treated to allow detection by a biosensor assay.

  2. Identification of Cryptosporidium species and genotypes in dairy cattle in Brazil Identificação de espécies e genótipos de Cryptosporidium em bovinos leiteiros no Brasil

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    Flavio Medeiros Paz e Silva

    Full Text Available In this study, we identified Cryptosporidium species and genotypes present in dairy cattle in the central region of São Paulo state, Brazil. Fecal specimens were collected from 200 animals (100 calves and 100 cows in ten dairy farms. Fecal samples were examined using microscopic examination (ME, enzyme immunoassay (EIA and polymerase chain reaction (PCR. Cryptosporidium species and genotypes were determined by restriction fragment length polymorphism (RFLP or DNA sequencing analysis of the SSU-rRNA and GP60 genes. The occurrence of Cryptosporidium spp. infection was 14% (28/200. The occurrence in calves (26% was significantly higher than in cows (2%. Of the 27 Cryptosporidium-positive specimens submitted to genotyping, C. andersoni was identified in 23 (85.1%, C. bovis in three (11.1%, and the zoonotic C. parvum subtype IIaA15G2R1 in one (3.7%. The study demonstrates that Cryptosporidium spp. infection was common and widespread in dairy cattle in this region and that calves have a high prevalence of C. andersoni. Furthermore, the presence of C. parvum subtype IIaA15G2R1 indicates that dairy calves from this region should be considered a potential source of zoonotic Cryptosporidium oocysts.No presente estudo foram identificadas espécies e genótipos de Cryptosporidium originadas de bovinos leiteiros na região central do estado de São Paulo, Brasil. Amostras fecais foram coletadas de 200 animais (100 bezerros e 100 vacas em 10 propriedades leiteiras. As amostras foram examinadas utilizando os métodos de microscopia óptica (MO, ensaio imunoenzimático (EI e reação em cadeia da polimerase (PCR. As espécies e genótipos de Cryptosporidium foram determinados pelo método de polimorfismo no tamanho dos fragmentos de restrição (RFLP ou sequenciamento dos genes SSU-rRNA e GP60. A infecção por Cryptosporidium spp. teve ocorrência de 14% (28/200. A ocorrência em bezerros (26% foi significativamente maior do que em vacas (2%. Do total de 27

  3. Cryptosporidium muris: Infectivity and Illness in Healthy Adult Volunteers

    Science.gov (United States)

    Chappell, Cynthia L.; Okhuysen, Pablo C.; Langer-Curry, Rebecca C.; Lupo, Philip J.; Widmer, Giovanni; Tzipori, Saul

    2015-01-01

    Although Cryptosporidium parvum and C. hominis cause the majority of human cryptosporidiosis cases, other Cryptosporidium species are also capable of infecting humans, particularly when individuals are immunocompromised. Ten C. muris cases have been reported, primarily in human immunodeficiency virus (HIV) -positive patients with diarrhea. However, asymptomatic cases were reported in two HIV-negative children, and in another case, age and immune status were not described. This study examines the infectivity of C. muris in six healthy adults. Volunteers were challenged with 105 C. muris oocysts and monitored for 6 weeks for infection and/or illness. All six patients became infected. Two patients experienced a self-limited diarrheal illness. Total oocysts shed during the study ranged from 6.7 × 106 to 4.1 × 108, and the number was slightly higher in volunteers with diarrhea (2.8 × 108) than asymptomatic shedders (4.4 × 107). C. muris-infected subjects shed oocysts longer than occurred with other species studied in healthy volunteers. Three volunteers shed oocysts for 7 months. Physical examinations were normal, with no reported recurrence of diarrhea or other gastrointestinal complaints. Two persistent shedders were treated with nitazoxanide, and the infection was resolved. Thus, healthy adults are susceptible to C. muris, which can cause mild diarrhea and result in persistent, asymptomatic infection. PMID:25311695

  4. Detection of Giardia lamblia, Cryptosporidium spp. and Entamoeba histolytica in clinical stool samples by using multiplex real-time PCR after automated DNA isolation.

    Science.gov (United States)

    Van Lint, P; Rossen, J W; Vermeiren, S; Ver Elst, K; Weekx, S; Van Schaeren, J; Jeurissen, A

    2013-01-01

    Diagnosis of intestinal parasites in stool samples is generally still carried out by microscopy; however, this technique is known to suffer from a low sensitivity and is unable to discriminate between certain protozoa. In order to overcome these limitations, a real-time multiplex PCR was evaluated as an alternative approach for diagnosing Giardia lamblia, Cryptosporidium spp. and Entamoeba histolytica in stool samples.Therefore, a total of 631 faecal samples were analysed both by microscopy as well as by real-time PCR following automated DNA extraction. Results showed that real-time PCR exhibited sensitivity and specificity of both 100%, whereas traditional microscopy exhibited sensitivity and specificity of 37.5% and 99.8% respectively. As real-time PCR provides simple, sensitive and specific detection of these three important pathogenic protozoan parasites, this technique, rather than microscopy, has become our diagnostic method of choice for the detection of enteric protozoan parasites for the majority of patients.

  5. Chemodiversity of Ladder-Frame Prymnesin Polyethers in Prymnesium parvum.

    Science.gov (United States)

    Rasmussen, Silas Anselm; Meier, Sebastian; Andersen, Nikolaj Gedsted; Blossom, Hannah Eva; Duus, Jens Øllgaard; Nielsen, Kristian Fog; Hansen, Per Juel; Larsen, Thomas Ostenfeld

    2016-09-23

    Blooms of the microalga Prymnesium parvum cause devastating fish kills worldwide, which are suspected to be caused by the supersized ladder-frame polyether toxins prymnesin-1 and -2. These toxins have, however, only been detected from P. parvum in rare cases since they were originally described two decades ago. Here, we report the isolation and characterization of a novel B-type prymnesin, based on extensive analysis of 2D- and 3D-NMR data of natural as well as 90% (13)C enriched material. B-type prymnesins lack a complete 1,6-dioxadecalin core unit, which is replaced by a short acyclic C2 linkage compared to the structure of the original prymnesins. Comparison of the bioactivity of prymnesin-2 with prymnesin-B1 in an RTgill-W1 cell line assay identified both compounds as toxic in the low nanomolar range. Chemical investigations by liquid chromatography high-resolution mass spectrometry (LC-HRMS) of 10 strains of P. parvum collected worldwide showed that only one strain produced the original prymnesin-1 and -2, whereas four strains produced the novel B-type prymnesin. In total 13 further prymnesin analogues differing in their core backbone and chlorination and glycosylation patterns could be tentatively detected by LC-MS/HRMS, including a likely C-type prymnesin in five strains. Altogether, our work indicates that evolution of prymnesins has yielded a diverse family of fish-killing toxins that occurs around the globe and has significant ecological and economic impact.

  6. Molecular investigation of Cryptosporidium in small caged pets in northeast China: host specificity and zoonotic implications.

    Science.gov (United States)

    Li, Qiao; Li, Lu; Tao, Wei; Jiang, Yanxue; Wan, Qiang; Lin, Yongchao; Li, Wei

    2016-07-01

    This study screened 151 pet-derived fecal specimens randomly collected from four commercial markets in northeast China for the presence of Cryptosporidium by genus-specific nested PCRs of the small subunit rRNA gene. Of these, 14 specimens (9.3 %) from nine species of birds, two types of rodents, and a hedgehog were positive for Cryptosporidium. Sequence analysis on the PCR-positive isolates facilitated identification of three Cryptosporidium species (C. baileyi, C. galli, and C. ubiquitum) and two Cryptosporidium genotypes (ferret genotype and avian genotype V). The study birds were affected predominantly with bird-specific C. baileyi (Atlantic canary, budgerigar, crested myna, rock dove, and silky fowl), C. galli (Chinese hwamei), and Cryptosporidium avian genotype V (Fischer's lovebird and rosy-faced lovebird). Cryptosporidium ferret genotype previously considered rodent-adapted was identified in three specimens from budgerigar, chipmunk, and red squirrel. Two specimens collected from common hill myna and hedgehog were positive for C. ubiquitum. The species of birds that can be colonized by Cryptosporidium were extended. Moreover, the data expanded the host range of Cryptosporidium ferret genotype and C. ubiquitum, especially the birds. The carriage of zoonotic C. ubiquitum in small caged pets is of public health importance.

  7. Cryptosporidium oocysts and giardia cysts on salad products irrigated with contaminated water.

    Science.gov (United States)

    Amorós, Inmaculada; Alonso, José L; Cuesta, Gonzalo

    2010-06-01

    A field study in Valencia, Spain, was done to determine the occurrence of Giardia and Cryptosporidium on salad products that are frequently eaten raw, such as lettuces and Chinese cabbage, and in irrigation waters. Four water samples were taken weekly 1 month before harvesting the vegetables. All water samples were analyzed using techniques included in the U.S. Environmental Protection Agency Method 1623. Standard methods for detecting protozoan parasites on salad vegetables are not available. Published techniques for the isolation of parasites from vegetables generally have low and variable recovery efficiencies. In this study, vegetables were analyzed using a recently reported method for detection of Cryptosporidium oocysts and Giardia cysts on salad products. The waters tested were positive for both Cryptosporidium and Giardia. Of 19 salad products studied, we observed Cryptosporidium in 12 samples and Giardia in 10 samples. Recoveries of the Texas Red-stained Cryptosporidium and Giardia, which were used as internal controls, were 24.5% +/- 3.5% for Cryptosporidium and 16.7% +/- 8.1% for Giardia (n = 8). This study provides data on the occurrence of Cryptosporidium and Giardia in salad products in Spain. The method was useful in the detection of Cryptosporidium oocysts and Giardia cysts on the vegetables tested, and it provides a useful analytical tool for occurrence monitoring.

  8. The prevalence of Cryptosporidium spp. oocysts in wild mammals in the Kruger National Park, South Africa.

    Science.gov (United States)

    Abu Samra, Nada; Jori, Ferran; Samie, Amidou; Thompson, Peter

    2011-01-10

    This study determined the prevalence of Cryptosporidium spp. oocysts in faecal samples from elephant (Loxodonta africana), buffalo (Syncerus caffer) and impala (Aepyceros melampus) in the Kruger National Park (KNP) and an adjacent game reserve in South Africa. Two of the study areas were in close proximity to rural communities on the western KNP boundary and the third study area was located in the centre of the KNP. Fresh stool samples (n=445) were collected and tested using an immunofluorescent antibody test (IFA) for Cryptosporidium parvum. A total of 278 of these were randomly selected (approximately 90 samples per wildlife species) and tested with the modified Ziehl Neelsen staining technique (ZN) for Cryptosporidium spp. The prevalence of Cryptosporidium spp. was highest in elephants (25.8% [95% confidence interval: 17.3, 35.9]), compared to buffalo (5.5% [1.8, 12.4]) and impala (4.3% [1.2, 10.5]). C. parvum showed similar patterns, being most prevalent in elephants (4.2% [1.5, 8.8]), compared to buffalo (1.4% [0.2, 5.1]) and impala (1.9% [0.4, 5.3]). 29 samples, including ZN positive and IFA positive samples, were retested using a real time PCR (rtPCR) technique. Of the 28 ZN-positive samples, 14 (50%) were positive with rtPCR and of the 9 IFA-positive samples 6 (67%) were confirmed positive by rtPCR. The prevalence of Cryptosporidium oocysts was significantly higher in both of the two study areas adjacent to the western KNP boundary compared to the area in the centre of the KNP (OR=3.2 [1.2, 9.0]; P=0.024). Our study demonstrates for the first time the presence of Cryptosporidium spp. in wildlife in South Africa. The transmission of this parasite between wildlife, domestic animals and humans is a plausible hypothesis and represents a potential risk for immunodeficient human populations.

  9. Genotyping Cryptosporidium andersoni in cattle in Shaanxi Province, Northwestern China.

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    Guang-Hui Zhao

    Full Text Available The present study examined the prevalence and genotypes of Cryptosporidium andersoni in cattle in Shaanxi province, China. A total of 2071 fecal samples (847 from Qinchuan cattle and 1224 from dairy cattle were examined for the presence of Cryptosporidium oocysts, and 70 samples (3.4% were C. andersoni-positive and those positive samples were identified by PCR amplification of the small subunit ribosomal RNA (SSU rRNA and the Cryptosporidium oocyst wall protein (COWP genes. C. andersoni was the only species found in the examined cattle in this province. Fifty-seven C. andersoni isolates were characterized into 5 MLST subtypes using multilocus sequence typing analysis, including a new subtype in the native beef breed Qinchuan cattle. All of these C. andersoni isolates presented a clonal genetic structure. These findings provide new insights into the genetic structure of C. andersoni isolates in Shaanxi province and basic data of Cryptosporidium prevalence status, which in turn have implications for controlling cryptosporidiosis in this province.

  10. Development of a PCR Diagnostic Kit for Cryptosporidium andersoni in Dairy Cow

    Institute of Scientific and Technical Information of China (English)

    ZHOU Rong-qiong; LI Guo-qing; XIAO Shu-min; LI Wei-hua

    2007-01-01

    Cryptosporidiosis is an important zoonosis caused by the Cryptosporidium species. To develop a PCR diagnostic kit for molecular detection and differential diagnosis of Cryptosporidiurn spp., a portion of ITS-1 sequence of Cryptosporidium. Andersoni was chosen as the target DNA for designing the species-specific primers (ZRQF/ZR). The kit components were determined after the PCR amplification conditions were serially optimized. A series of tests were conducted in the specificity, sensitivity, stability, reproducibility, and stored period of the kit, respectively. The results showed that only C. Andersoni were amplified specific band of about 500 bp, while Cryptosporidium. Parvum, Cryptosporidium. Baileyi, Eimeria sp of dairy cow, Toxoplasma gondii, Eimeria sp of pig, Ascaris suum, Cyclospora sp, and E. Coli could not be amplified. 254 oocysts of C. Andersoni was the lowest number that could be detected using the kit. The kit worked well after being stored at room temperature, 4 and -20℃ for nine months. Fecal specimens, which were collected from a total of 243 calves on four different dairy farms in Guangdong Province, China, and one dairy farm in Henan Province, China, were examined using the kit; the positive rate of the kit was 2-13% higher than that of the routine methods. The results indicated that the kit can detect fecal samples faster, more sensitively, and conveniently, and can provide a useful tool for the identification and differentiation of C. Andersoni from the other Cryptosporidium species; it also has implications for further studies on molecular epidemiology and differential diagnostics of cryptosporidiosis in animals.

  11. PROPHYLACTIC EFFECT OF BOVINE ANTI-CRYPTOSPORIDIUM HYPERIMMUNE COLOSTRUM IN HEALTHY VOLUNTEERS CHALLENGED WITH CRYPTOSPORIDIUM PARVUM. (R824759)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  12. Alternate phase variation in expression of two major surface membrane proteins (MBA and UU376) of Ureaplasma parvum serovar 3.

    Science.gov (United States)

    Zimmerman, Carl-Ulrich R; Stiedl, Thomas; Rosengarten, Renate; Spergser, Joachim

    2009-03-01

    Ureaplasma urealyticum and Ureaplasma parvum are commensals and pathogens of the human urogenital tract and of newborn infants. There are four distinct U. parvum serovars and 10 distinct U. urealyticum serovars. Both species possess a distinct immunodominant variable surface protein, the multiple banded antigen (MBA), which shows size variability among isolates as a result of changes in the number of C-terminal repeating units. Adjacent to the MBA gene (UU375) lies UU376, which was annotated as 'Ureaplasma-specific conserved hypothetical gene'. In four different strains of U. parvum serovar 3, we demonstrated expression of UU376 by Western blot analysis and phase variation between UU376, here designated Upvmp376 (Ureaplasma phase-variable membrane protein 376), and MBA after application of selective pressure with hyperimmune antisera directed against either protein. By Southern blot analysis, we found that the switch between MBA and Upvmp376 expression is associated with a DNA inversion event in which the nonrepetitive region of the MBA gene and its putative promoter region are opposed to either the repetitive region of MBA or UU376. We propose that in U. parvum serovar 3, and presumably in all U. parvum and U. urealyticum, an inversion event at specific sites effects an alternate ON/OFF switching of the genes UU375 and UU376.

  13. COMPARAÇÃO DA EFICIÊNCIA DAS COLORAÇÕES DE ZIEHL- NEELSEN MODIFICADO E SAFRANINA MODIFICADA NA DETECÇÃO DE OOCISTOS DE Cryptosporidium spp. (EUCOCCIDIORIDA, CRYPTOSPORIDIIDAE A PARTIR DE AMOSTRAS FECAIS DE BEZERROS DE 0 A 3 MESES

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    Renata Dias Rodrigues

    2016-01-01

    Full Text Available Bovine cryptosporidiosis is caused by four differents s pecies: C ryptosporidium parvum, Cryptosporidium bovis, Cryptosporidium andersoni and Cryptosporidium ryanae. The species Cryptosporidium parvum (Order: Eucoccidiorida, Family: Cryptosporidiidae is considered of high zoonotic potential and it can infect humans through the elimination of oocysts by both cattle and by humans. The objective of this research was to detect oocysts of the genus Cryptosporidium spp. in fecal contents of calves (75 males and 77 females. We collected 152 stool samples from animals aged between 0 day and 3 months. The material was subjected to modified Ziehl-Neelsen and modified Safranin techniques, the slides were observed in its entire length by optical microscopy to verify the presence of oocysts of this parasitic infections. The results showed 17.1% (26/152 positivity in the samples examined, and the statistical analysis showed no difference between sex and the staining techniques used in this study. We concluded the infection by Cryptosporidium spp. is present in the evaluated properties, but more studies are needed, so that the risk of infection is measured properly and prophylactic measures are implemented.

  14. Cryptosporidium infections in children in Durban

    African Journals Online (AJOL)

    1991-03-16

    Mar 16, 1991 ... diarrhoea were found to be passing Cryptosporidium oocysts compared with only 2,4% ... this increase in morbidity and mortality in Cryptosporidium- associated .... taminated water.34 It is possible that contamination of rivers.

  15. Giardia and Cryptosporidium in Pristine Protected Catchments in Central Eastern Australia%澳大利亚中东部原始汇水保护区里的表吮贾第虫Giardia和隐小孢子虫Crptosporidium

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    @@ 我们首次证实在澳大利亚中东部没有都市和农业污染的原始汇水保护区的偏远小溪里存在着致病的原生动物表吮贾第虫(肠兰伯氏鞭毛虫)Giardia lamblia和隐小孢子虫Cryptosporidium parvum.

  16. [Molecular Detection of Giardia lamblia and Cryptosporidium Species in Pet Dogs].

    Science.gov (United States)

    GU, You-fang; WANG, Kai; LIU, De-yi; MEI, Nan; CHEN, Cheng; CHEN, Tao; HAN, Min-min; ZHOU, Li; CAO, Jia-tong; ZHANG, Heng; ZHANG, Xue-liang; FAN, Zi-lai; LI, Wen-chao

    2015-10-01

    To determine the prevalence of Giardia lamblia and Cryptosporidium species infection in pet dogs, and identify the G. lamblia assemblages and Cryptosporidium species. A total of 315 fresh fecal samples were collected from pet clinics in five counties of Anhui Province and in Hangzhou of Zhejiang Province. Hemi-nested-PCR targeting the GDH gene of G. lamblia and nested-PCR targeting the SSU rRNA gene of Cryptosporidium were performed in all the fecal samples. The PCR products were sequenced and analyzed using bioinformatics methods to identify the G. lamblia assemblages and Cryptosporidium species. The positive rates of G. lamblia and Cryptosporidium spp. infections in the 315 fecal samples were 3.2% (10/315) and 1.6% (5/315), respectively. Specifically, the two indicators were both significantly higher in dogs ≤12 months (17.8% and 11.1%, respectively) than in adult dogs (0.7% and 0.0%)(Plamblia assemblages were identified, assemblages B (n=6) and D (n=4). Sequence analysis of PCR products of the SSU rRNA gene showed that the five Cryptosporidium isolates were C. canis (n =5). The prevalences of G. lamblia and Cryptosporidium infection in pet dogs in Anhui and Zhejiang Provinces were 3.2 % and 1.6 %, respectively. The assemblages of G. lamblia in this study are of types B and D.

  17. Age-related Infection with Cryptosporidium Species and Genotype in Pigs in China

    Institute of Scientific and Technical Information of China (English)

    YIN Jian Hai; YUAN Zhong Ying; CAI Hui Xia; SHEN Yu Juan; JIANG Yan Yan; ZHANG Jing; WANG Yan Juan; CAO Jian Ping

    2013-01-01

    Objective Pigs, as hosts of zoonotic Cryptosporidium species/genotypes, are domestic animals with public health significance. The present study was to characterize the infection rate and species/genotype of Cryptosporidium in pre-weaned and post-weaned pigs from Shanghai and Shaoxing, China. Methods A total of 208 fecal samples (42 from pre-weaned piglets, and 166 from post-weaned pigs) were examined by nested PCR of the 18S rRNA gene and analyzed by phylogenetic DNA fragment sequencing of secondary PCR products. Results Infection was detected in 79 samples (19/42 pre-weaned piglets, and 60/166 post-weaned pigs). C. suis (14/79) and Cryptosporidium pig genotype II (65/79) were identified; piglets were more susceptible to the former (13/14) and post-weaned pigs to the latter (59/65). Conclusion Infection of Cryptosporidium spp. in pigs was age-specific;piglets were more susceptible to C. suis while pigs were more susceptible to Cryptosporidium pig genotype II. These findings combined with the isolation of the two Cryptosporidium from water suggest that pigs may be a source of zoonotic Cryptosporidium water pollution. Improvements in pig feeding practices, sewage discharge, feces disposal and field worker protection are therefore important to prevent potential public health problems.

  18. Developing risk models of Cryptosporidium transport in soils from vegetated, tilted soilbox experiments.

    Science.gov (United States)

    Harter, Thomas; Atwill, Edward R; Hou, Lingling; Karle, Betsy M; Tate, Kenneth W

    2008-01-01

    Transport of Cryptosporidium parvum through macroporous soils is poorly understood yet critical for assessing the risk of groundwater contamination. We developed a conceptual model of the physics of flow and transport in packed, tilted, and vegetated soilboxes during and immediately after a simulated rainfall event and applied it to 54 experiments implemented with different soils, slopes, and rainfall rates. Using a parsimonious inverse modeling procedure, we show that a significant amount of subsurface outflow from the soilboxes is due to macropore flow. The effective hydraulic properties of the macropore space were obtained by calibration of a simple two-domain flow and transport model that accounts for coupled flow in the matrix and in the macropores of the soils. Using linear mixed-effects analysis, macropore hydraulic properties and oocyst attenuation were shown to be associated with soil bulk density and rainfall rate. Macropore flow was shown to be responsible for bromide and C. parvum transport through the soil into the underlying pore space observed during the 4-h experiments. We confirmed this finding by conducting a pair of saturated soil column studies under homogeneously repacked conditions with no macropores in which no C. parvum transport was observed in the effluent. The linear mixed-effects and logistic regression models developed from the soilbox experiments provide a basis for estimating macropore hydraulic properties and the risk of C. parvum transport through shallow soils from bulk density, precipitation, and total shallow subsurface flow rate. The risk assessment is consistent with the reported occurrence of oocysts in springs or groundwater from fractured or karstic rocks protected only by shallow overlying soils.

  19. [Cryptosporidium hominis diarrhea outbreak and transmission linked to diaper infant use].

    Science.gov (United States)

    Teresa Ortega, María; Vergara, Alberto; Guimbao, Joaquín; Clavel, Antonio; Gavín, Patricia; Ruiz, Andrés

    2006-11-04

    On the basis of several cases of cryptosporidiosis detected in a child day-care center, we stablished the extent of the outbreak and investigated causes of parasite transmission. A retrospective cohort study was designed on all children attending day-care center and care givers to determine their infection status and identify risk factors associated to the outbreak. 24 cases of cryptosporidiosis were detected, with an attack rate of 0.46 (24/52); 12 of them were parasitologycally confirmed. All care givers were negative for Cryptosporidium and none of them reported symptoms of acute gastroenteritis. Transmission pattern was compatible with person to person modes. Among the factors investigated, two were associated with the risk of disease: diaper wear (relative risk = 2.06; p = 0.059); and diarrhea in relatives (relative risk = 2.05; p = 0.01). In all confirmed cases, Cryptosporidium hominis (previously known as C. parvum, genotype 1), was identified. Cryptosporidiosis should be considered as a possible cause of outbreaks of gastroenteritis at day-care centers. Increasing care on diaper changing practices, specially over children with diarrhea, may be the key factor to prevent transmission of Cryptosporidium.

  20. Effective Concentration and Detection of Cryptosporidium, Giardia, and the Microsporidia from Environmental Matrices

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    Joseph A. Moss

    2014-01-01

    Full Text Available Cryptosporidium spp., Giardia spp., and members of Microsporidia are enteropathogenic parasites of humans and animals, producing asymptomatic to severe intestinal infections. To circumvent various impediments associated with current detection methods, we tested a method providing multistage purification and separation in a single, confined step. Standard real-time PCR was used as a detection method. Samples spiked with C. parvum and G. intestinalis were split for comparison to standard Method 1623. Results were equivalent to immunomagnetic procedures for Cryptosporidium, and Giardia. Overall percent recovery for Cryptosporidium with Method 1623 averaged 26.89% (std 21.44%; min = 0%; max = 73% and was similar but less variable for qPCR method at an estimated average of 27.67 (std 17.65%; min = 5%; max = 63%. For Giardia, Method 1623 had an overall average recovery of 27.11% (std 17.98%; min = 1%; max = 58%, while multistage purification and qPCR had an estimated lower overall recovery at 18.58% (std 13.95%; min = 0%; max = 35%. Microsporidia were also readily detected with an estimated recovery of 46.81% overall (std 17.66%; min = 18%; max = 70% for E. intestinalis and 38.90% (std 14.36%; min = 13%; max = 62% for E. bieneusi.

  1. In vitro activity of five quinolones and analysis of the quinolone resistance-determining regions of gyrA, gyrB, parC, and parE in Ureaplasma parvum and Ureaplasma urealyticum clinical isolates from perinatal patients in Japan.

    Science.gov (United States)

    Kawai, Yasuhiro; Nakura, Yukiko; Wakimoto, Tetsu; Nomiyama, Makoto; Tokuda, Tsugumichi; Takayanagi, Toshimitsu; Shiraishi, Jun; Wasada, Kenshi; Kitajima, Hiroyuki; Fujita, Tomio; Nakayama, Masahiro; Mitsuda, Nobuaki; Nakanishi, Isao; Takeuchi, Makoto; Yanagihara, Itaru

    2015-04-01

    Ureaplasma spp. cause several disorders, such as nongonococcal urethritis, miscarriage, and preterm delivery with lung infections in neonates, characterized by pathological chorioamnionitis in the placenta. Although reports on antibiotic resistance in Ureaplasma are on the rise, reports on quinolone-resistant Ureaplasma infections in Japan are limited. The purpose of this study was to determine susceptibilities to five quinolones of Ureaplasma urealyticum and Ureaplasma parvum isolated from perinatal samples in Japan and to characterize the quinolone resistance-determining regions in the gyrA, gyrB, parC, and parE genes. Out of 28 clinical Ureaplasma strains, we isolated 9 with high MICs of quinolones and found a single parC gene mutation, resulting in the change S83L. Among 158 samples, the ParC S83L mutation was found in 37 samples (23.4%), including 1 sample harboring a ParC S83L-GyrB P462S double mutant. Novel mutations of ureaplasmal ParC (S83W and S84P) were independently found in one of the samples. Homology modeling of the ParC S83W mutant suggested steric hindrance of the quinolone-binding pocket (QBP), and de novo prediction of peptide structures revealed that the ParC S84P may break/kink the formation of the α4 helix in the QBP. Further investigations are required to unravel the extent and mechanism of antibiotic resistance of Ureaplasma spp. in Japan.

  2. Genetic characterisation of Cryptosporidium and Giardia from dairy calves: discovery of species/genotypes consistent with those found in humans.

    Science.gov (United States)

    Abeywardena, Harshanie; Jex, Aaron R; Nolan, Matthew J; Haydon, Shane R; Stevens, Melita A; McAnulty, Robin W; Gasser, Robin B

    2012-12-01

    Cryptosporidium and Giardia are important genera of parasitic protists that can cause significant diarrhoeal diseases in humans and other animals. Depending on the species/genotype of parasite, human infection may be acquired via anthroponotic or zoonotic transmission routes. Here, we undertook a molecular epidemiological investigation of these two genera of parasites in pre- and post-weaned calves from eight locations in Canterbury, New Zealand, by PCR-coupled sequencing and phylogenetic analysis of sequence data for regions in the 60 kDa glycoprotein (pgp60) gene of Cryptosporidium and/or the triose-phosphate isomerase (ptpi) gene of Giardia. The pgp60 and ptpi regions were specifically amplified from 15 (8.3%) and 11 (6.1%) of the 180 individual faecal samples, respectively. The sequences derived from all of the amplicons were aligned with homologous reference sequences and subjected to phylogenetic analysis by Bayesian inference. For Cryptosporidium, three samples contained Cryptosporidium parvum genotype IIa, subgenotypes IIaA15G3R1, IIaA19G3R1 and IIaA23G4. Twelve samples contained Cryptosporidium hominis genotype Ib, subgenotype IbA10G2R2. While subgenotypes IIaA15G3R1 and IIaA23G4 are new records, IIaA19G3R1 and IbA10G2R2 are commonly found in humans in various countries. For Giardia, two samples contained Giardia duodenalis assemblage A, also common in humans. In contrast, nine samples contained G. duodenalis assemblage E, which is the first report of this assemblage in cattle in New Zealand. Therefore, the present results indicate that dairy calves on the South Island of New Zealand harbour 'zoonotic' genotypes of Cryptosporidium and Giardia, which is likely to have significant public health implications.

  3. Occurrence of Cryptosporidium spp. and Giardia spp. in a public water-treatment system, Paraná, Southern Brazil

    Directory of Open Access Journals (Sweden)

    Jonatas Campos Almeida

    Full Text Available The purpose of this study was to investigate the occurrence of Cryptosporidium spp. and Giardia spp. in a public water-treatment system. Samples of raw and treated water were collected and concentrated using the membrane filtration technique. Direct Immunofluorescence Test was performed on the samples. DNA extraction using a commercial kit was performed and the DNA extracted was submitted to a nested-PCR reaction (n-PCR and sequencing. In the immunofluorescence, 2/24 (8.33% samples of raw water were positive for Giardia spp.. In n-PCR and sequencing, 2/24 (8.33% samples of raw water were positive for Giardia spp., and 2/24 (8.33% samples were positive for Cryptosporidium spp.. The sequencing showed Cryptosporidium parvum and Giardia duodenalis DNA. In raw water, there was moderate correlation among turbidity, color and Cryptosporidium spp. and between turbidity and Giardia spp.. The presence of these protozoans in the water indicates the need for monitoring for water-treatment companies.

  4. Cryptosporidiosis in Haiti: surprisingly low level of species diversity revealed by molecular characterization of Cryptosporidium oocysts from surface water and groundwater

    Science.gov (United States)

    Damiani, Céline; Balthazard-Accou, Ketty; Clervil, Elmyre; Diallo, Aïssata; Da Costa, Cécilia; Emmanuel, Evens; Totet, Anne; Agnamey, Patrice

    2013-01-01

    The protozoan parasite Cryptosporidium sp. has emerged as one of the most important water contaminants, causing waterborne outbreaks of diarrhoeal diseases worldwide. In Haiti, cryptosporidiosis is a frequent cause of diarrhoea in children under the age of five years, HIV-infected individuals, and people living in low socioeconomic conditions, mainly due to the consumption of water or food polluted by Cryptosporidium oocysts. The aim of this study was to detect and identify Cryptosporidium oocysts present in 12 water samples collected in Port-au-Prince and 4 water samples collected in Cap Haïtien. Initial detection consisted of immunomagnetic separation – immunofluorescence assay (IMS-IFA), which was confirmed by nested PCR, targeting the most polymorphic region of the 18S rRNA gene in 15/16 samples. Genotyping was performed by PCR-restriction fragment length polymorphism (RFLP) analysis and DNA sequencing. Under our working conditions, neither nested PCR-RFLP nor direct DNA sequencing revealed the expected species diversity, as only Cryptosporidium parvum was identified in the water samples studied. This study highlights the difficulty of detecting mixed populations of Cryptosporidium species in environmental samples. PMID:24252814

  5. Chemodiversity of Ladder-Frame Prymnesin Polyethers in Prymnesium parvum

    DEFF Research Database (Denmark)

    Rasmussen, Silas Anselm; Meier, Sebastian; Andersen, Nikolaj Gedsted;

    2016-01-01

    Blooms of the microalga Prymnesium parvum cause devastating fish kills worldwide, which are suspected to be caused by the supersized ladder-frame polyether toxins prymnesin-1 and -2. These toxins have, however, only been detected from P. parvum in rare cases since they were originally described two...

  6. Assessing calves as carriers of Cryptosporidium and Giardia with zoonotic potential on dairy and beef farms within a water catchment area by mutation scanning.

    Science.gov (United States)

    Abeywardena, Harshanie; Jex, Aaron R; Firestone, Simon M; McPhee, Sandra; Driessen, Nicole; Koehler, Anson V; Haydon, Shane R; von Samson-Himmelstjerna, Georg; Stevens, Melita A; Gasser, Robin B

    2013-08-01

    In the present study, we undertook a molecular epidemiological survey of Cryptosporidium and Giardia in calves on three dairy and two beef farms within an open drinking water catchment area (Melbourne, Australia). Faecal samples (n = 474) were collected from calves at two time points (5 months apart) and tested using a PCR-based mutation scanning-targeted sequencing phylogenetic approach, employing regions within the genes of small subunit (SSU) of ribosomal RNA (designated partial SSU), 60 kDa glycoprotein (pgp60) and triose phosphate isomerase (ptpi) as genetic markers. Using partial SSU, the C. bovis, C. parvum, C. ryanae and a new genotype of Cryptosporidium were characterised from totals of 74 (15.6%), 35 (7.3%), 37 (7.8%) and 9 (1.9%) samples, respectively. Using pgp60, C. parvum genotype IIa subgenotype A18G3R1 was detected in 29 samples. Using ptpi, G. duodenalis assemblages A and E were detected in totals of 10 (2.1%) and 130 (27.4%) samples, respectively. The present study showed that a considerable proportion of dairy and beef calves in this open water catchment region excreted Cryptosporidium (i.e. subgenotype IIaA18G3R1) and Giardia (e.g. assemblage A) that are consistent with those infecting humans, inferring that they are of zoonotic importance. Future work should focus on exploring, in a temporal and spatial way, whether these parasites occur in the environment and water of the catchment reservoir.

  7. Cryptosporidium cuniculus and Giardia duodenalis in rabbits: genetic diversity and possible zoonotic transmission.

    Directory of Open Access Journals (Sweden)

    Weizhe Zhang

    Full Text Available BACKGROUND: Cryptosporidium and Giardia are the two important zoonotic pathogens causing diarrhea of humans and animals worldwide. Considering the human cryptosporidiosis outbreak and sporadic cases caused by C. cuniculus, the important public health significance of G. duodenalis and little obtained information regarding rabbit infected with Cryptosporidium and Giardia in China, the aim of this study is to determine the prevalence and molecularly characterize Cryptosporidium and Giardia in rabbits in Heilongjiang Province, China. METHODOLOGY/PRINCIPAL FINDINGS: 378 fecal samples were obtained from rabbits in Heilongjiang Province. Cryptosporidium oocysts and Giardia cysts were detected using Sheather's sugar flotation technique and Lugol's iodine stain method, respectively. The infection rates of Cryptosporidium and Giardia were 2.38% (9/378 and 7.41% (28/378, respectively. Genotyping of Cryptosporidium spp. was done by DNA sequencing of the small subunit rRNA (SSU rRNA gene and all the nine isolates were identified as Cryptosporidium cuniculus. The nine isolates were further subtyped using the 60-kDa glycoprotein (gp60 gene and two subtypes were detected, including VbA32 (n = 3 and a new subtype VbA21 (n = 6. G. duodenalis genotypes and subtypes were identified by sequence analysis of the triosephosphate isomerase (TPI gene. The assemblage B (belonging to eight different subtypes B-I to B-VIII was found in 28 G. duodenalis-positive samples. CONCLUSIONS/SIGNIFICANCE: The rabbits have been infected with Cryptosporidium and Giardia in Heilongjiang Province. The results show that the rabbits pose a threat to human health in the studied areas. Genotypes and subgenotypes of C. cuniculus and G. duodenalis in this study might present the endemic genetic characterization of population structure of the two parasites.

  8. Isolation of parasites on fruits and vegetables.

    Science.gov (United States)

    Bier, J W

    1991-12-01

    The current FDA method to recover parasites from fruits and vegetables is derived from procedures used to isolate parasitic protozoa from water. A 1kg portion of fruit or vegetable is divided into 200 g subportions. The subportions are sequentially processed in a sonic cleaning bath with 1.5 liters of detergent solution (1% sodium dodecyl sulfate, 0.1% Tween 80) and sonicated for 10 minutes. As each subsample is removed, it is thoroughly drained. After this sonic treatment, the wash water is collected in a polypropylene beaker, transferred to 50 ml polypropylene centrifuge tubes and centrifuged for 15 min at 1500 x g. The sediment is consolidated into one tube along with two rinsings of each tube. The final sediment is fixed in 4% formaldehyde for 10 minutes before examination for parasites. Indirect fluorescent antibody is applied to stain the parasites (Giardia spp. and/or Cryptosporidium spp.) by using commercial kits when available. If a large quantity of extraneous matter is contained in the sediment it may be reduced by layering on Sheather's fluid and centrifuging at 1500 x g for 15 minutes. The supernatant is collected and washed twice in distilled water. This procedure is adequate for protozoa and nonoperculate helminth eggs; operculate helminth eggs may be cleaned by extraction with ethyl acetate. When cabbage and lettuce were seeded at 1 organism/g, the rate of recovery for Cryptosporidium parvum with the FDA method was 1%. When cabbage was seeded at 1 egg/g and 10 eggs/g, the average rate of recovery of decorticated eggs of Ascaris sp. or untreated Trichuris sp. was 10%.(ABSTRACT TRUNCATED AT 250 WORDS)

  9. Transport of Cryptosporidium parvum oocysts in soil columns following applications of raw and separated liquid slurry

    DEFF Research Database (Denmark)

    Petersen, Heidi Huus; Enemark, Heidi L.; Olsen, Annette;

    2012-01-01

    . Among leachate samples containing oocysts, 44/72 samples yielded viable oocysts as determined by a dye permeability assay (DAPI/PI) with the majority (41%) of viable oocysts found in leachate from soil columns with added liquid slurry. The number of viable oocysts was positively correlated (r=0...

  10. ESTIMATING MAXIMUM POSSIBLE ENVIRONMENTAL LOADING AMOUNTS OF CRYPTOSPORIDIUM PARVUM ATTRIBUTABLE TO BEEF CATTLE. (R828038)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  11. Assessment of drugs against Cryptosporidium parvum using a simple in vitro screening method.

    Science.gov (United States)

    Armson, A; Meloni, B P; Reynoldson, J A; Thompson, R C

    1999-09-15

    A rapid semi-quantitative screening method was devised for assessing the anticryptosporidial and cytotoxic effects of putative chemotherapeutic compounds. The method is suitable as an initial rapid screening procedure from which compounds demonstrating anticryptosporidial activity can be identified for further analysis. It has the advantages of speed, low cost and concurrent assessment of anticryptosporidial and cytotoxic effects and allows accurate determination of minimum lethal concentrations. Of the 71 compounds screened, six completely inhibited cryptosporidial growth at 1 microM (monensin, salinomycin, alborixin, lasalocid, trifluralin and nicarbazin) and a further eight showed significant anticryptosporidial activity at 1 or 20 microM (halquinol, bleomycin, suramin, mitomycin, doxycycline hydrochloride, toltrazuril, chloroquine phosphate and teniposide). Twelve compounds were found to have some degree of cytotoxicity at 1 microM and a further 12 at 20 microM.

  12. PATHOGENESIS OF HUMAN AND BOVINE CRYPTOSPORIDIUM PARVUM IN GNOTOBIOTIC PIGS. (R826138)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  13. PATHOGENESIS OF BOVINE (GENOTYPE 2) CRYPTOSPORIDIUM PARVUM STRAINS IN NEONATAL AND AGED GNOTOBIOTIC PIGS (R826138)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  14. EFFECT OF LACTOBACILLUS AND BIFIDOBACTERIUM ON CRYPTOSPORIDIUM PARVUM OOCYST VIABILITY. (R826138)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  15. The General Biology of Cryptosporidium

    Science.gov (United States)

    The protozoan parasites belonging to the genus Cryptosporidium infect all classes of vertebrate animals. Of the sixteen valid species and nearly forty genotypes, three species are responsible for the majority of economic losses to livestock and morbidity and mortality to humans. This chapter summari...

  16. Isolation and amplification of mRNA within a simple microfluidic lab on a chip.

    Science.gov (United States)

    Reinholt, Sarah J; Behrent, Arne; Greene, Cassandra; Kalfe, Ayten; Baeumner, Antje J

    2014-01-07

    The major modules for realizing molecular biological assays in a micro-total analysis system (μTAS) were developed for the detection of pathogenic organisms. The specific focus was the isolation and amplification of eukaryotic mRNA within a simple, single-channel device for very low RNA concentrations that could then be integrated with detection modules. The hsp70 mRNA from Cryptosporidium parvum was used as a model analyte. Important points of study were surface chemistries within poly(methyl methacrylate) (PMMA) microfluidic channels that enabled specific and sensitive mRNA isolation and amplification reactions for very low mRNA concentrations. Optimal conditions were achieved when the channel surface was carboxylated via UV/ozone treatment followed by the immobilization of polyamidoamine (PAMAM) dendrimers on the surface, thus increasing the immobilization efficiency of the thymidine oligonucleotide, oligo(dT)25, and providing a reliable surface for the amplification reaction, importantly, without the need for blocking agents. Additional chemical modifications of the remaining active surface groups were studied to avoid nonspecific capturing of nucleic acids and hindering of the mRNA amplification at low RNA concentrations. Amplification of the mRNA was accomplished using nucleic acid sequence-based amplification (NASBA), an isothermal, primer-dependent technique. Positive controls consisting of previously generated NASBA amplicons could be diluted 10(15) fold and still result in successful on-chip reamplification. Finally, the successful isolation and amplification of mRNA from as few as 30 C. parvum oocysts was demonstrated directly on-chip and compared to benchtop devices. This is the first proof of successful mRNA isolation and NASBA-based amplification of mRNA within a simple microfluidic device in relevant analytical volumes.

  17. Cryptosporidium huwi n. sp. (Apicomplexa: Eimeriidae) from the guppy (Poecilia reticulata).

    Science.gov (United States)

    Ryan, Una; Paparini, Andrea; Tong, Kaising; Yang, Rongchang; Gibson-Kueh, Susan; O'Hara, Amanda; Lymbery, Alan; Xiao, Lihua

    2015-03-01

    The morphological, biological, and molecular characteristics of Cryptosporidium piscine genotype 1 from the guppy (Poecilia reticulata) are described, and the species name Cryptosporidium huwi n. sp. is proposed to reflect its genetic and biological differences from gastric and intestinal Cryptosporidium species. Oocysts of C.huwi n. sp. over-lap in size with Cryptosporidium molnari, measuring approximately 4.4-4.9 µm (mean 4.6) by 4.0-4.8 µm (mean 4.4 µm) with a length to width ratio of 1.04 (0.92-1.35) (n = 50). Similar to C.molnari, C.huwi n. sp. was identified in the stomach only and clusters of oogonial and sporogonial stages were identified deep within the epithelium. However, phylogenetic analysis of 18S rRNA sequences indicated that C. huwi n. sp. exhibited 8.5-9.2% and 3.5% genetic distance from C.molnari isolates and piscine genotype 7 respectively. At the actin locus, the genetic distance between C.huwi n. sp. and C.molnari was 16.6%. The genetic distance between C.huwi n. sp. and other Cryptosporidium species at the 18S locus was 13.2%-17% and at the actin locus was 18.9%-26.3%. Therefore C. huwi n. sp. is genetically distinct from previously described Cryptosporidium species.

  18. Cryptosporidium andersoni from a Danish cattle herd: identification and preliminary characterisation

    DEFF Research Database (Denmark)

    Enemark, Heidi L.; Ahrens, Peter; Lowery, C.J.

    2002-01-01

    In November 1997, Cryptosporidium andersoni, for the first time, was isolated from a Danish heifer. The isolate was characterised morphologically, molecularly, and furthermore inoculated into mice and one calf. Data on the distribution of cryptosporidia in the herd of origin were obtained at two ...

  19. Genotyping of Cryptosporidium spp. from free-living wild birds from Brazil.

    Science.gov (United States)

    Sevá, Anaiá da Paixão; Funada, Mikaela Renata; Richtzenhain, Leonardo; Guimarães, Marta Brito; Souza, Sheila de Oliveira; Allegretti, Luciana; Sinhorini, Juliana Anaya; Duarte, Vanessa Vertematti; Soares, Rodrigo Martins

    2011-01-10

    In wild and domestic birds, cryptosporidiosis is often associated with infections by Cryptosporidium galli, Cryptosporidium baileyi and Cryptosporidium meleagridis. In addition to these species, a number of avian Cryptosporidium species yet to be fully characterized are commonly found among exotic and wild avian isolates. The present study aimed to detect and identify samples of Cryptosporidium spp. from free-living wild birds, in order to contribute to the knowledge of the variability of this parasite in the free-living population of Brazil. Stool samples were collected from 242 birds, with the following proportions of individuals: 50 Emberizidae (20.7%), 112 Psittacidae (46.3%), 44 Cardinalidae (18.2%), 12 Turdidae (5.0%), eight Ramphastidae (3.3%), seven Icteridae (2.9%), three Estrilididae (1.2%), two Contigidae (0.8%), two Thraupidae (0.8%) and two Fringilidae (0.8%). Among the 242 fecal samples from wild birds, 16 (6.6%) were positive for the presence of oocysts of Cryptosporidium. Molecular characterization of the 16 samples of Cryptosporidium, were performed with phylogenetic reconstructions employing 292 positions of 18S rDNA. None of the samples of birds was characterized as C. meleagridis. C. galli was identified in one rufous-bellied thrush (Turdus rufiventris), five green-winged saltators (Saltator similis), one slate-coloured seedeater (Sporophila schistacea), one goldfinch (Carduelis carduelis) and three saffron finches (Sicalis flaveola). One goldfinch isolate, one buffy-fronted seedeater (Sporophila frontalis), one red-cowled cardinal (Paroaria dominicana) and one other saffron finch (S. flaveola) were identified as C. baileyi. Avian genotype II was found in an isolate from a white-eyed parakeet (Aratinga leucophthalma). Clinical symptoms of cryptosporidiosis in birds have already been described and the number of wild birds which were shedding parasites was high. Therefore, further epidemiological research and disease surveillance of birds in the

  20. Cryptosporidium spp. and Giardia duodenalis as pathogenic contaminants of water in Galicia, Spain: the need for safe drinking water.

    Science.gov (United States)

    Castro-Hermida, José Antonio; González-Warleta, Marta; Mezo, Mercedes

    2015-01-01

    The objectives of this cross-sectional study were to detect the presence of Cryptosporidium spp. and Giardia duodenalis in drinking water treatments plants (DWTPs) in Galicia (NW Spain) and to identify which species and genotype of these pathogenic protozoans are present in the water. Samples of untreated water (surface or ground water sources) and of treated drinking water (in total, 254 samples) were collected from 127 DWTPs and analysed by an immunofluorescence antibody test (IFAT) and by PCR. Considering the untreated water samples, Cryptosporidium spp. were detected in 69 samples (54.3%) by IFAT, and DNA of this parasite was detected in 57 samples (44.8%) by PCR, whereas G. duodenalis was detected in 76 samples (59.8%) by IFAT and in 56 samples (44.0%) by PCR. Considering the treated drinking water samples, Cryptosporidium spp. was detected in 52 samples (40.9%) by IFAT, and the parasite DNA was detected in 51 samples (40.1%) by PCR, whereas G. duodenalis was detected in 58 samples (45.6%) by IFAT and in 43 samples (33.8%) by PCR. The percentage viability of the (oo)cysts ranged between 90.0% and 95.0% in all samples analysed. Cryptosporidium andersoni, C. hominis, C. parvum and assemblages A-I, A-II, E of G. duodenalis were identified. The results indicate that Cryptosporidium spp. and G. duodenalis are widespread in the environment and that DWTPs are largely ineffective in reducing/inactivating these pathogens in drinking water destined for human and animal consumption in Galicia. In conclusion, the findings suggest the need for better monitoring of water quality and identification of sources of contamination.

  1. Chemodiversity of Ladder-Frame Prymnesin Polyethers in Prymnesium parvum

    DEFF Research Database (Denmark)

    Rasmussen, Silas Anselm; Meier, Sebastian; Andersen, Nikolaj Gedsted;

    2016-01-01

    Blooms of the microalga Prymnesium parvum cause devastating fish kills worldwide, which are suspected to be caused by the supersized ladder-frame polyether toxins prymnesin-1 and -2. These toxins have, however, only been detected from P. parvum in rare cases since they were originally described two......-HRMS) of 10 strains of P. parvum collected worldwide showed that only one strain produced the original prymnesin-1 and -2, whereas four strains produced the novel B-type prymnesin. In total 13 further prymnesin analogues differing in their core backbone and chlorination and glycosylation patterns could...

  2. Transport of Cryptosporidium oocysts in porous media: Role of straining and physicochemical filtration

    Science.gov (United States)

    Tufenkji, N.; Miller, G.F.; Ryan, J.N.; Harvey, R.W.; Elimelech, M.

    2004-01-01

    The transport and filtration behavior of Cryptosporidium parvum oocysts in columns packed with quartz sand was systematically examined under repulsive electrostatic conditions. An increase in solution ionic strength resulted in greater oocyst deposition rates despite theoretical predictions of a significant electrostatic energy barrier to deposition. Relatively high deposition rates obtained with both oocysts and polystyrene latex particles of comparable size at low ionic strength (1 mM) suggest that a physical mechanism may play a key role in oocyst removal. Supporting experiments conducted with latex particles of varying sizes, under very low ionic strength conditions where physicochemical filtration is negligible, clearly indicated that physical straining is an important capture mechanism. The results of this study indicate that irregularity of sand grain shape (verified by SEM imaging) contributes considerably to the straining potential of the porous medium. Hence, both straining and physicochemical filtration are expected to control the removal of C. parvum oocysts in settings typical of riverbank filtration, soil infiltration, and slow sand filtration. Because classic colloid filtration theory does not account for removal by straining, these observations have important implications with respect to predictions of oocyst transport.

  3. Chitosan and metal salt coagulant impacts on Cryptosporidium and microsphere removal by filtration.

    Science.gov (United States)

    Brown, Trevor J; Emelko, Monica B

    2009-02-01

    Maintenance of appropriate chemical pretreatment is a critical component of ensuring proper filtration performance. Pilot-scale in-line filtration studies were performed to investigate the relative impacts of chitosan, alum, and FeCl(3) coagulation on the removal of Cryptosporidium parvum oocysts and oocyst-sized polystyrene microspheres by granular media filtration. Similar removals of oocysts and microspheres were achieved when optimal coagulant doses were utilized. Sub-optimal alum and FeCl(3) coagulation resulted in a deterioration filter effluent turbidity (0.2-0.3NTU) and total particle counts (30-100 total particles > or =2microm/mL) that were accompanied by reduced (by approximately 2-3-log) median oocyst and microsphere removals by filtration. At all doses investigated, chitosan coagulation resulted in excellent turbidity and particle reductions by filtration. Nonetheless, chitosan coagulation at doses of 0.1, 0.5, and 1.0mg/L did not result in appreciable improvements in C. parvum oocyst removal relative to complete coagulation failure (median oocyst removals were alum and FeCl(3) coagulation, but not chitosan coagulation.

  4. First description of Cryptosporidium ubiquitum XIIa subtype family in farmed fur animals.

    Science.gov (United States)

    Kellnerová, Klára; Holubová, Nikola; Jandová, Anna; Vejčík, Antonín; McEvoy, John; Sak, Bohumil; Kváč, Martin

    2017-06-01

    This study investigated the prevalence of Cryptosporidium in farmed fur animals in the Czech Republic and Poland. A total of 480 faecal samples were collected from fur animals, including 300 American mink (Mustela vison), 60 silver foxes (Vulpes vulpes), 50 long-tailed chinchillas (Chinchilla lanigera), and 70 nutrias (Myocastor coypus), at 14 farms. Samples were examined for the presence of Cryptosporidium using microscopy (following aniline-carbol-methyl violet staining) and sequence analysis of PCR amplified products. Three mink and two chinchillas from two different farms tested positive for Cryptosporidium ubiquitum DNA. The presence of C. ubiquitum DNA was not associated with diarrhoea. Subtyping of C. ubiquitum isolates by sequence analysis of the 60-kDa glycoprotein gene showed that isolates belonged to the XIIa subtype family, which was previously restricted to humans and ruminants. This suggests that C. ubiquitum subtype XIIa has a broader host range than previously reported. Copyright © 2017 Elsevier GmbH. All rights reserved.

  5. A study of risk factors associated with the prevalence of Cryptosporidium in villages around Lake Atitlan, Guatemala

    Directory of Open Access Journals (Sweden)

    H. E. Laubach

    2004-08-01

    Full Text Available Cryptosporidium parvum is an endemic, zoonotic coccidian parasitosis that is highly prevalent in third-world countries where waterborne fecal contamination of food and drink or person-to-person contact with oocysts are the most common methods of transmission of the enteric protozoan. This type of transmission of the parasite made the villages around Lake Atitlan, Guatemala a unique site to compare environmental risk factors with the level of Cryptosporidium infections in the local residents. The study was carried out in two villages, San Antonio Palopo and Santa Catarina Palopo, located in the highlands above the shores of the lake. Smears from stool specimens of patients with gastroenteritis were processed using Kinyoun's modified acid-fast stain and observed with light microscopy. Of the 100 residents examined from the two villages, 32% had Cryptosporidium infections. Female children had the highest prevalence of infection (44% in San Antonio Palopo and 46% in Santa Catarina Palopo, p<0.05, and they also had significantly higher infection rates than males, 50% vs. 17%, respectively. The prevalence rate was not influenced by the season of the year or by the location of the residents. We found differences in prevalence rates due to age and gender, and we suggest that the high infection rates of specific groups are associated with their exposure to the contaminated water supply from Lake Atitlan.

  6. Seasonal variation and potential sources of Cryptosporidium contamination in surface waters of Chao Phraya River and Bang Pu Nature Reserve pier, Thailand.

    Science.gov (United States)

    Koompapong, Khuanchai; Sukthana, Yaowalark

    2012-07-01

    Using molecular techniques, a longitudinal study was conducted with the aims at identifying the seasonal difference of Cryptosporidium contamination in surface water as well as analyzing the potential sources based on species information. One hundred forty-four water samples were collected, 72 samples from the Chao Phraya River, Thailand, collected in the summer, rainy and cool seasons and 72 samples from sea water at Bang Pu Nature Reserve pier, collected before, during and after the presence of migratory seagulls. Total prevalence of Cryptosporidium contamination in river and sea water locations was 11% and 6%, respectively. The highest prevalence was observed at the end of rainy season continuing into the cool season in river water (29%) and in sea water (12%). During the rainy season, prevalence of Cryptosporidium was 4% in river and sea water samples, but none in summer season. All positive samples from the river was C. parvum, while C. meleagridis (1), and C. serpentis (1) were obtained from sea water. To the best of our knowledge, this is the first genetic study in Thailand of Cryptosporidium spp contamination in river and sea water locations and the first report of C. serpentis, suggesting that humans, household pets, farm animals, wildlife and migratory birds may be the potential sources of the parasites. The findings are of use for implementing preventive measures to reduce the transmission of cryptosporidiosis to both humans and animals.

  7. Prevalence and genetic characterizations of Cryptosporidium spp. in pre-weaned and post-weaned piglets in Heilongjiang Province, China.

    Directory of Open Access Journals (Sweden)

    Weizhe Zhang

    Full Text Available BACKGROUND: Cryptosporidium spp. are common intestinal protozoa of humans and animals. There have been few studies conducted on the molecular characterizations of pig-derived Cryptosporidium isolates worldwide, especially in China. Thus, the aim of the present study was to understand the prevalence, distribution and genotypes of Cryptosporidium in pigs in Heilongjiang Province, China. METHODOLOGY/PRINCIPAL FINDINGS: A total of 568 fecal samples from pre-weaned and post-weaned piglets were collected from eight pig farms from four areas of Heilongjiang Province. The average infection rate of Cryptosporidium was 1.6% (9/568 by microscopy. 113 samples were subjected to PCR amplification of the small subunit (SSU rRNA gene of Cryptosporidium, with 55.8% (63/113 being positive for Cryptosporidium. Cryptosporidium suis (n = 31 and C. scrofarumn (n = 32 were identified by DNA sequencing of the SSU rRNA gene. Three types of C. scrofarumn were found at the SSU rRNA locus, with one novel type being detected. Using species/genotype-specific primers for pig-adapted Cryptosporidium spp., 22 and 23 respectively belonged to C. suis and C. scrofarum mono-infections, with 18 co-infections detected. The infection peaks for C. suis (60%, 24/40 and C. scrofarum (51.2%, 21/41 were respectively found in the piglets of 5 to 8 weeks and more than 8 weeks. CONCLUSION/SIGNIFICANCE: The detection of C. suis and C. scrofarum in pre-weaned and post-weaned piglets has public health implications, due to the fact that the two species are both zoonotic Cryptosporidium. The novel C. scrofarum type detected may be endemic to China.

  8. Cryptosporidium en Giardia in Nederlandse zwembaden

    NARCIS (Netherlands)

    Schets FM; Engels GB; Leenen EJTM; MGB

    2003-01-01

    Swimming-pool associated outbreaks of cryptosporidiosis have been frequently reported in the UK and USA. Cryptosporidium oocysts could sometimes be detected in the pool water or the filter backwash water in cases where the source of the outbreak was confirmed. The occurrence of Cryptosporidium and G

  9. Emissie van Cryptosporidium en Giardia door landbouwhuisdieren

    NARCIS (Netherlands)

    Schrijven JF; Bruin HAM de; Engels GB; Leenen EJTM; MGB

    1999-01-01

    In this study, the relative contributions of the pathogenic protozoa Cryptosporidium and Giardia by manure of farm animals in The Netherlands to the total yearly environmental load was studied. Manure of veal calves forms a very large source of Cryptosporidium (1.5 m 10 square 16 oocysts per year) a

  10. A perspective on Cryptosporidium and Giardia, with an emphasis on bovines and recent epidemiological findings.

    Science.gov (United States)

    Abeywardena, Harshanie; Jex, Aaron R; Gasser, Robin B

    2015-04-01

    Cryptosporidium and Giardia are two common aetiological agents of infectious enteritis in humans and animals worldwide. These parasitic protists are usually transmitted by the faecal-oral route, following the ingestion of infective stages (oocysts or cysts). An essential component of the control of these parasitic infections, from a public health perspective, is an understanding of the sources and routes of transmission in different geographical regions. Bovines are considered potential sources of infection for humans, because species and genotypes of Cryptosporidium and Giardia infecting humans have also been isolated from cattle in molecular parasitological studies. However, species and genotypes of Cryptosporidium and Giardia of bovids, and the extent of zoonotic transmission in different geographical regions in the world, are still relatively poorly understood. The purpose of this article is to (1) provide a brief background on Cryptosporidium and Giardia, (2) review some key aspects of the molecular epidemiology of cryptosporidiosis and giardiasis in animals, with an emphasis on bovines, (3) summarize research of Cryptosporidium and Giardia from cattle and water buffaloes in parts of Australasia and Sri Lanka, considering public health aspects and (4) provide a perspective on future avenues of study. Recent studies reinforce that bovines harbour Cryptosporidium and Giardia that likely pose a human health risk and highlight the need for future investigations of the biology, population genetics and transmission dynamics of Cryptosporidium and Giardia in cattle, water buffaloes and other ruminants in different geographical regions, the fate and transport of infective stages following their release into the environment, as well as for improved strategies for the control and prevention of cryptosporidiosis and giardiasis, guided by molecular epidemiological studies.

  11. Detection of Cryptosporidium spp. Oocysts in raw sewage and creek water in the city of São Paulo, Brazil

    Directory of Open Access Journals (Sweden)

    Farias Eveline Wilma Coutinho

    2002-01-01

    Full Text Available The protozoan parasite Cryptosporidium has emerged as one of the most important contaminants of water, causing waterborne outbreaks of gastroenteritis worldwide. To monitor and understand the public health significance of this pathogen in environmental samples, several methods have been developed to isolate and detect Cryptosporidium oocysts. The purpose of this study was to perform the first investigation on the presence of Cryptosporidium spp. oocysts in raw sewage and creek water in the city of São Paulo, Brazil. The oocysts were concentrated by flocculation and membrane filtration. The results showed the occurrence of Cryptosporidium spp. in all wastewater samples analyzed, indicating a possible risk for dissemination of these pathogens in aquatic environment and in the community.

  12. Cryptosporidium in pet snakes from Italy: molecular characterization and zoonotic implications.

    Science.gov (United States)

    Díaz, P; Rota, S; Marchesi, B; López, C; Panadero, R; Fernández, G; Díez-Baños, P; Morrondo, P; Poglayen, G

    2013-10-18

    To provide information on the occurrence of Cryptosporidium species and genotypes in captive snakes from Italy, faecal specimens from 120 snakes belonging to 13 different genera of the families Boidae, Colubridae and Pythonidae were collected. Faecal samples were taken from the ground of the terrarium when available; otherwise cloacal cotton swabs were used. No clinical signs of cryptosporidiosis were observed in any animal at the time of sampling. Samples were examined for the presence of Cryptosporidium by using a direct immunofluorescence antibody test (IFAT) and two-step nested PCR at the small subunit (SSU) rRNA locus. PCR-positive samples were genotyped by restriction fragment length polymorphism (RFLP) analysis with the endonucleases SspI and VspI. By IFAT, 42 out of 120 snakes (35.0%) were found to be shedding Cryptosporidium oocysts. A significant higher percentage of positive ophidians were detected by using faecal specimens obtained from the terrarium (55.5%) than by cloacal cotton swabs (29.0%). SSU rRNA gene products were obtained from 25 isolates. Twenty samples tested positive to both microscopy and molecular techniques. Our data reveal a wide extent of cryptosporidial infections in snake-food animals since most of the identified isolates belonged to Cryptosporidium species, some of them with zoonotic potential, considered specific for rodents and resulting from ingestion of infected preys. The reptilian-specific species Cryptosporidium serpentis was identified in only one isolate. The common presence of reptile non-specific and, in some cases, zoonotic Cryptosporidium oocysts in snake faeces should to be taken into consideration in order to avoid the misidentification of the protozoan as well as the possible public health implications.

  13. Prymnesium parvum exotoxins affect the grazing and viability of the calanoid copepod Eurytemora affinis

    DEFF Research Database (Denmark)

    Sopanen, S.; Koski, Marja; Uronen, P.

    2008-01-01

    The calanoid copepod Eurytemora affinis from the northern Baltic Sea was exposed to cell-free filtrates of the toxic haptophyte Prymnesium parvum as well as to cell mixtures of P. parvum and Rhodomonas salina. To test the effects of P. parvum exudates and allelopathy on selective grazers, copepods...

  14. Identification and molecular characterization of Cryptosporidium and Giardia in children and cattle populations from the province of Álava, North of Spain.

    Science.gov (United States)

    Cardona, Guillermo A; Carabin, Hélène; Goñi, Pilar; Arriola, Larraitz; Robinson, Guy; Fernández-Crespo, Juan C; Clavel, Antonio; Chalmers, Rachel M; Carmena, David

    2011-12-15

    The prevalence of and factors associated with the protozoan enteropathogens Cryptosporidium and Giardia have been investigated in selected children and cattle populations from the province of Álava (Northern Spain). The presence of these organisms was detected in fecal samples using commercially available coproantigen-ELISA (CpAg-ELISA) and immunochromatographic (ICT) assays. A total of 327 caregivers of children participants were asked to answer questions on risk factors potentially associated to the prevalence of Cryptosporidium and Giardia, including water-use practices, water sports and contact with domestic or pet animals. Molecular analyses were conducted using a nested-PCR technique to amplify the small-subunit (SSU) rRNA gene of Cryptosporidium and the triosephosphate isomerase (tpi) gene of Giardia. Cryptosporidium oocysts and Giardia cysts were found in 3 and 16 samples using the CpAg-ELISA, and in 5 and 9 samples using the ICT test, respectively. Cryptosporidium and Giardia were also found in 7 and 17 samples by CpAg-ELISA, and 4 and 14 samples by ICT, respectively, of 227 cattle fecal samples. The overall Cryptosporidium and Giardia infection prevalences, based on a Bayesian approach accounting for the imperfect sensitivities and specificities of both diagnostic tests, were estimated to 1.0% (95% BCI: 0.2%-2.8%) and 3.1% (1.5%-5.3%) in children and 3.0% (0.5%-9.2%) and 1.4% (0.0%-6.4%) in cattle, respectively. In humans, a single Cryptosporidium isolate was characterized as C. hominis. Of seven Giardia isolates, four were identified as assemblage B, two as assemblage A-II and one was a mixed assemblage B+A-II infection. No Cryptosporidium or Giardia isolates could be obtained from cattle samples. Although limited, these results seem to suggest that cattle are unlikely to be an important reservoir of zoonotic Cryptosporidium and/or Giardia in the province of Álava.

  15. Identification and molecular characterization of Cryptosporidium and Giardia in children and cattle populations from the province of Alava, North of Spain

    Energy Technology Data Exchange (ETDEWEB)

    Cardona, Guillermo A. [Livestock Laboratory, Regional Government of Alava, Ctra. de Azua 4, 01520 Vitoria-Gasteiz (Spain); Carabin, Helene [Department of Biostatistics and Epidemiology, College of Public Health, Oklahoma University Health Sciences Center, 801 Northeast 13th Street, Room 309, Oklahoma City, OK 73104 (United States); Goni, Pilar [Department of Microbiology, Preventive Medicine and Public Health, Faculty of Medicine, University of Zaragoza, Domingo Miral s/n, 50009 Zaragoza (Spain); Arriola, Larraitz [Epidemiology Unit, Public Health Division of Guipuzcoa, Basque Government, Av. Navarra 4, 2013 San Sebastian (Spain); Robinson, Guy [UK Cryptosporidium Reference Unit, Public Health Wales, Microbiology ABM, Swansea, Singleton Hospital, Swansea SA2 8QA (United Kingdom); Fernandez-Crespo, Juan C. [Sub-direction of Public Health of Alava, Department of Health, Basque Government, Avda. Santiago 11, 01002 Vitoria-Gasteiz (Spain); Clavel, Antonio [Department of Microbiology, Preventive Medicine and Public Health, Faculty of Medicine, University of Zaragoza, Domingo Miral s/n, 50009 Zaragoza (Spain); Chalmers, Rachel M. [UK Cryptosporidium Reference Unit, Public Health Wales, Microbiology ABM, Swansea, Singleton Hospital, Swansea SA2 8QA (United Kingdom); Carmena, David, E-mail: d.carmena@imperial.ac.uk [MRC Clinical Sciences Centre, Faculty of Medicine, Imperial College, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN (United Kingdom)

    2011-12-15

    The prevalence of and factors associated with the protozoan enteropathogens Cryptosporidium and Giardia have been investigated in selected children and cattle populations from the province of Alava (Northern Spain). The presence of these organisms was detected in fecal samples using commercially available coproantigen-ELISA (CpAg-ELISA) and immunochromatographic (ICT) assays. A total of 327 caregivers of children participants were asked to answer questions on risk factors potentially associated to the prevalence of Cryptosporidium and Giardia, including water-use practices, water sports and contact with domestic or pet animals. Molecular analyses were conducted using a nested-PCR technique to amplify the small-subunit (SSU) rRNA gene of Cryptosporidium and the triosephosphate isomerase (tpi) gene of Giardia. Cryptosporidium oocysts and Giardia cysts were found in 3 and 16 samples using the CpAg-ELISA, and in 5 and 9 samples using the ICT test, respectively. Cryptosporidium and Giardia were also found in 7 and 17 samples by CpAg-ELISA, and 4 and 14 samples by ICT, respectively, of 227 cattle fecal samples. The overall Cryptosporidium and Giardia infection prevalences, based on a Bayesian approach accounting for the imperfect sensitivities and specificities of both diagnostic tests, were estimated to 1.0% (95% BCI: 0.2%-2.8%) and 3.1% (1.5%-5.3%) in children and 3.0% (0.5%-9.2%) and 1.4% (0.0%-6.4%) in cattle, respectively. In humans, a single Cryptosporidium isolate was characterized as C. hominis. Of seven Giardia isolates, four were identified as assemblage B, two as assemblage A-II and one was a mixed assemblage B + A-II infection. No Cryptosporidium or Giardia isolates could be obtained from cattle samples. Although limited, these results seem to suggest that cattle are unlikely to be an important reservoir of zoonotic Cryptosporidium and/or Giardia in the province of Alava.

  16. Cryptosporidium testudinis sp. n., Cryptosporidium ducismarci Traversa, 2010 and Cryptosporidium tortoise genotype III (Apicomplexa: Cryptosporidiidae) in tortoises.

    Science.gov (United States)

    Jezkova, Jana; Horcickova, Michaela; Hlaskova, Lenka; Sak, Bohumil; Kvetonova, Dana; Novak, Jan; Hofmannova, Lada; McEvoy, John; Kvac, Martin

    2016-10-14

    Understanding of the diversity of species of Cryptosporidium Tyzzer, 1910 in tortoises remains incomplete due to the limited number of studies on these hosts. The aim of the present study was to characterise the genetic diversity and biology of cryptosporidia in tortoises of the family Testudinidae Batsch. Faecal samples were individually collected immediately after defecation and were screened for presence of cryptosporidia by microscopy using aniline-carbol-methyl violet staining, and by PCR amplification and sequence analysis targeting the small subunit rRNA (SSU), Cryptosporidium oocyst wall protein (COWP) and actin genes. Out of 387 faecal samples from 16 tortoise species belonging to 11 genera, 10 and 46 were positive for cryptosporidia by microscopy and PCR, respectively. All samples positive by microscopy were also PCR positive. Sequence analysis of amplified genes revealed the presence of the Cryptosporidium tortoise genotype I (n = 22), C. ducismarci Traversa, 2010 (n = 23) and tortoise genotype III (n = 1). Phylogenetic analyses of SSU, COWP and actin gene sequences revealed that Cryptosporidium tortoise genotype I and C. ducismarci are genetically distinct from previously described species of Cryptosporidium. Oocysts of Cryptosporidium tortoise genotype I, measuring 5.8-6.9 µm × 5.3-6.5 µm, are morphologically distinguishable from C. ducismarci, measuring 4.4-5.4 µm × 4.3-5.3 µm. Oocysts of Cryptosporidium tortoise genotype I and C. ducismarci obtained from naturally infected Russian tortoises (Testudo horsfieldii Gray) were infectious for the same tortoise but not for Reeve's turtles (Mauremys reevesii [Gray]), common garter snake (Thamnophis sirtalis [Linnaeus]), zebra finches (Taeniopygia guttata [Vieillot]) and SCID mice (Mus musculus Linnaeus). The prepatent period was 11 and 6 days post infection (DPI) for Cryptosporidium tortoise genotype I and C. ducismarci, respectively; the patent period was longer than 200 days for both cryptosporidia

  17. Review of Cervi Cornu Parvum Pharmacopuncture in Korean Medicine

    Directory of Open Access Journals (Sweden)

    Lee Dong-Jin

    2013-06-01

    Full Text Available Objective: The endpoint of this review is to investigate existing studies of Cervi cornu parvum (CCP pharmacopuncture within Korean medicine journals in order to present a better research method in the future. Methods: We searched all the papers through six Korean electrical databases that included the title of "Cervi cornu parvum pharmacopuncture" or "Cervi cornu parvum aqua-acupuncture". Articles that had been published until December 2012 were largely divided into experimental studies and clinical studies. Results: Fifty-three (53 experimental studies and six clinical studies were found. The number of published articles has been constantly increasing. Many of the experimental studies demonstrated anti-inflammatory effects for arthritis, and most of the clinical studies dealt with musculoskeletal problems. Conclusion: Various therapeutically significant effects of the CCP pharmacopuncture have been found through this review; however, more systematic clinical studies on the CCP pharmacopuncture seem to be necessary to substantially support its clinical effects.

  18. Cryptosporidium Infections Among Children in Peru

    Centers for Disease Control (CDC) Podcasts

    2008-09-25

    Cryptosporidium is a waterborne bacteria that can cause severe diarrhea and vomiting. In this podcast, Dr. Vita Cama, CDC microbiologist, discusses an article in the October 2008 issue of Emerging Infectious Diseases. The paper examines Cryptosporidium infections among children in Peru, including the number of infections, symptoms experienced, and what species of Crypto were responsible.  Created: 9/25/2008 by Emerging Infectious Diseases.   Date Released: 9/25/2008.

  19. 40 CFR 141.712 - Unfiltered system Cryptosporidium treatment requirements.

    Science.gov (United States)

    2010-07-01

    ... Cryptosporidium, Giardia lamblia, or viruses. ... systems must meet the combined Cryptosporidium inactivation requirements of this section and Giardia lamblia and virus inactivation requirements of § 141.72(a) using a minimum of two disinfectants, and...

  20. Bioaccumulation of Toxoplasma and Cryptosporidium by the freshwater crustacean Gammarus fossarum: Involvement in biomonitoring surveys and trophic transfer.

    Science.gov (United States)

    Bigot-Clivot, Aurélie; Palos Ladeiro, Mélissa; Lepoutre, Alexandra; Bastien, Fanny; Bonnard, Isabelle; Dubey, Jitender P; Villena, Isabelle; Aubert, Dominique; Geffard, Olivier; François, Adeline; Geffard, Alain

    2016-11-01

    The protozoa Toxoplasma gondii and Cryptosporidium parvum are public health priorities because their oocysts can persist in recreational, surface, drinking, river, and sea water sources for a long time. To evaluate the capacity of the freshwater crustacean Gammarus fossarum to accumulate T. gondii and C. parvum oocysts, gammarids were exposed to 200, 2000 or 20,000 oocysts per gammarid and per day for 21 days followed by 5 days of depuration. C. parvum DNA was detected by qPCR in G. fossarum in only one out of four pools for the highest concentration and after 14 days of exposure, and T. gondii DNA was detected after 7 days of exposure to the two highest concentrations. Our results document the capacity of G. fossarum to accumulate T. gondii in its tissues proportionally to the ambient concentration; the maximum number of oocysts was detected in gammarid tissues after exposure to 20,000 oocysts per day. Mean values of 3.26 (±3), 21.71 (±15.18), and 17.41 (±10.89) oocysts were detected in gammarids after 7, 14, and 21 days, respectively, and after 5 days of depuration, T. gondii oocysts were still present in gammarid tissues. These results show for the first time that a freshwater crustacean can bioaccumulate T. gondii oocysts, suggesting that G. fossarum is a potential effective bioindicator of protozoan contamination in biomonitoring studies. Moreover, due to its key position in freshwater food webs, G. fossarum could also play a role in the trophic transfer of protozoa. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. Potential role of beavers (Castor fiber in contamination of water in the Masurian Lake District (north-eastern Poland with protozoan parasites Cryptosporidium spp. and Giardia duodenalis

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    Sroka Jacek

    2015-06-01

    Full Text Available The purpose of this study was to assess the possible influence of beavers on the contamination of lake water with zoonotic parasites Giardia duodenalis and Cryptosporidium spp., with respect to the risk to human health. A total of 79 water samples were taken around the habitats of beavers from 14 localities situated in the recreational Masurian Lake District (north-eastern Poland. Water was sampled in the spring and autumn seasons, at different distances from beavers’ lodges (0-2, 10, 30, and 50 m. The samples were examined for the presence of (oocysts of zoonotic protozoa Giardia duodenalis and Cryptosporidium spp. by direct fluorescence assay (DFA and by nested and real time PCR. By DFA, the presence of Giardia cysts was found in 36 samples (45.6% and the presence of Cryptosporidium oocysts in 26 samples (32.9%. Numbers of Giardia cysts, Cryptosporidium oocysts, and summarised (oocysts of both parasites showed a significant variation depending on locality. The numbers of Giardia cysts significantly decreased with the distance from beavers’ lodges while the numbers of Cryptosporidium oocysts did not show such dependence. The amount of Giardia cysts in samples collected in spring was approximately 3 times higher than in autumn. Conversely, a larger number of Cryptosporidium oocysts were detected in samples collected in autumn than in spring. By PCR, Giardia DNA was found in 38 samples (48.1% whereas DNA of Cryptosporidium was found in only 7 samples (8.9%. Eleven Giardia isolates were subjected to phylogenetic analysis by restriction fragment length polymorphism PCR or sequencing which evidenced their belonging to zoonotic assemblages: A (3 isolates and B (8 isolates. In conclusion, water in the vicinity of beavers’ lodges in the tested region was markedly contaminated with (oocysts of Giardia duodenalis and Cryptosporidium spp., which confirms the potential role of beavers as a reservoir of these parasites and indicates a need for

  2. Assessment of molecular methods as a tool for detecting pathogenic protozoa isolated from water bodies.

    Science.gov (United States)

    Adamska, M; Sawczuk, M; Kolodziejczyk, L; Skotarczak, B

    2015-12-01

    Several species belong to the Cryptosporidium and Giardia genus, the main parasitic protozoa occurring in water, but only some of them are infectious to humans. We investigated the occurrence of Cryptosporidium and Giardia and identified their species in the water samples collected from natural water bodies in north-western Poland. A total of 600 samples from water bodies used for bathing, sewage discharge, as drinking water sources and watering places for animals were screened. The samples were collected during a 3-year period in each of the four seasons and filtered using Filta-Max (IDEXX Laboratories, USA). Genomic DNA was extracted from all samples and used as a target sequence for polymerase chain reaction (PCR) and TaqMan real-time PCR, as well as for reverse line blotting (RLB) methods. PCR methods seem to be more sensitive to detect Giardia and Cryptosporidium DNA in water samples than RLB methods. All PCR products were sequenced and three were identified as C. parvum and four as G. intestinalis. The overall prevalence of C. parvum (0.5%) and G. intestinalis (0.6%) in the samples suggests that the risk of Cryptosporidium and Giardia infections in north-western Poland is minimal.

  3. Growth and extracellular laccase production in liquid cultures of Minimidochium parvum LPSC # 548 Strain Crecimiento y producción de lacasa extracelular en cultivos líquidos de Minimidochium parvum cepa LPSC # 548

    OpenAIRE

    Saparrat, Mario C. N.; Arambarri, Angélica M.; Balatti, Pedro A.

    2007-01-01

    Minimidochium parvum LPSC # 548, a fungus isolated from litter floating on waters of Río Santiago (Provincia de Buenos Aires, Argentina) polluted with industrial effluents and crude-oil, was grown as a shaking culture on a C-limited medium to evaluate its ability to produce extracellular laccase. The effect of anthracene, CuSO4 · 5H2O, ethanol, guaiacol, humic acids, Kraft lignin, MnSO4· H2O, Tween 20 and veratryl alcohol on its growth and extracellular laccase activity levels was also analyz...

  4. Global modelling of Cryptosporidium in surface water

    Science.gov (United States)

    Vermeulen, Lucie; Hofstra, Nynke

    2016-04-01

    Introduction Waterborne pathogens that cause diarrhoea, such as Cryptosporidium, pose a health risk all over the world. In many regions quantitative information on pathogens in surface water is unavailable. Our main objective is to model Cryptosporidium concentrations in surface waters worldwide. We present the GloWPa-Crypto model and use the model in a scenario analysis. A first exploration of global Cryptosporidium emissions to surface waters has been published by Hofstra et al. (2013). Further work has focused on modelling emissions of Cryptosporidium and Rotavirus to surface waters from human sources (Vermeulen et al 2015, Kiulia et al 2015). A global waterborne pathogen model can provide valuable insights by (1) providing quantitative information on pathogen levels in data-sparse regions, (2) identifying pathogen hotspots, (3) enabling future projections under global change scenarios and (4) supporting decision making. Material and Methods GloWPa-Crypto runs on a monthly time step and represents conditions for approximately the year 2010. The spatial resolution is a 0.5 x 0.5 degree latitude x longitude grid for the world. We use livestock maps (http://livestock.geo-wiki.org/) combined with literature estimates to calculate spatially explicit livestock Cryptosporidium emissions. For human Cryptosporidium emissions, we use UN population estimates, the WHO/UNICEF JMP sanitation country data and literature estimates of wastewater treatment. We combine our emissions model with a river routing model and data from the VIC hydrological model (http://vic.readthedocs.org/en/master/) to calculate concentrations in surface water. Cryptosporidium survival during transport depends on UV radiation and water temperature. We explore pathogen emissions and concentrations in 2050 with the new Shared Socio-economic Pathways (SSPs) 1 and 3. These scenarios describe plausible future trends in demographics, economic development and the degree of global integration. Results and

  5. Molecular characterization of Cryptosporidium species at the wildlife/livestock interface of the Kruger National Park, South Africa.

    Science.gov (United States)

    Abu Samra, Nada; Jori, Ferran; Xiao, Lihua; Rikhotso, Oupa; Thompson, Peter N

    2013-05-01

    Molecular characterization of Cryptosporidium spp. was done on isolates from African elephant (Loxodonta africana), African buffalo (Syncerus caffer), impala (Aepyceros melampus) and native domestic calves collected during May and June 2008 at the wildlife/livestock interface of the Kruger National Park (KNP), South Africa. A polymerase chain reaction (PCR) restriction fragment length polymorphism (RFLP) analysis of the 18S rRNA gene was used in feces from 51 calves (3-12 months of age), 71 buffalo, 71 impala and 72 elephant, and sequencing of the 18S rRNA gene was done on PCR-RFLP-positive wildlife samples. Cryptosporidium spp. were detected in 8% (4/51) of the calves and identified as C. andersoni (2/4) and C. bovis (2/4). Four of the 214 wildlife samples were positive for Cryptosporidium with a prevalence of 2.8% each in impala and buffalo. Cryptosporidium ubiquitum was detected in two impala and one buffalo, and C. bovis in one buffalo. A concurrent questionnaire conducted among 120 farmers in the study area investigated contacts between wildlife species and livestock. Buffalo and impala had the highest probability of contact with cattle outside the KNP. Despite the fairly low prevalence found in wildlife and cattle, the circulation of zoonotic Cryptosporidium spp., such as C. ubiquitum, should be investigated further, particularly in areas of high HIV infection prevalence. Further studies should target younger animals in which the prevalence is likely to be higher.

  6. Canker and decline caused by Neofusiccocum parvum on Acacia melanoxylon in Italy

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    Sidoti A

    2016-12-01

    Full Text Available In the spring of 2012, in reforested areas of Peloritani Mountains (Sicily, Italy a severe dieback of Acacia melanoxylon R. Brown was observed. The main symptoms on both young and adults plants consisted of elongated cankers on the trunks and epicormic shoots, wilt of the canopy and dieback interested mostly aged trees. The woody tissues showed browning beyond the cankers. Sapwood and heartwood appeared decayed with a brown to gray-greenish discoloration. One fungal species was consistently isolated from infected woody tissues, which was morphologically attributed to Neofusiccocum sp. The sequencing of the ITS regions of a representative isolate allowed to identify (99% similarity the species Neofusiccocum parvum (Pennycook & Samuels Crous, Slippers and Phillips, teleomorph Botryosphaeria parva Pennycook & Samuels. The pathogenicity tests have reproduced symptoms similar to those observed in the field. N. parvum is the aetiologic agent of mortality of australian blackwood observed in Sicily and to our knowledge this is the first report of this fungus on Acacia melanoxylon. It is a generalist pathogen, cosmopolitan, present in many temperate areas, Mediterranean and subtropical. The older Peloritani Mountains populations of australian blackwood seem particularly susceptible to the pathogen, the latter favored by the lack of silvicultural interventions that generate interspecific and intraspecific competition, as well as the increase and spread of the fungus. To minimize the consequential damage is necessary to adopt sanitation measures that would lower the fungal inoculum and program substitutions of this exotic species with others that have multiple functions suited to environments (e.g., Chestnut or encouraging the establishment and development of native species, such as the holm oak and shrub.

  7. Gene expression in the mixotrophic prymnesiophyte, Prymnesium parvum, responds to prey availability

    Directory of Open Access Journals (Sweden)

    Zhenfeng eLiu

    2015-04-01

    Full Text Available The mixotrophic prymnesiophyte, Prymnesium parvum, is a widely distributed alga with significant ecological importance. It produces toxins and can form ecosystem disruptive blooms that result in fish kills and changes in planktonic food web structure. However, the relationship between P. parvum and its prey on the molecular level is poorly understood. In this study, we used RNA-Seq technology to study changes in gene transcription of P. parvum in three treatments with different microbial populations available as potential prey: axenic P. parvum (no prey, bacterized P. paruvm, and axenic P. parvum with ciliates added as prey. Thousands of genes were differentially expressed among the three treatments. Most notably, transcriptome data indicated that P. parvum obtained organic carbon, including fatty acids, from both bacteria and ciliate prey for energy and cellular building blocks. The data also suggested that different prey provided P. parvum with macro- and micronutrients, namely organic nitrogen in the form of amino acids from ciliates, and iron from bacteria. However, both transcriptomic data and growth experiments indicated that P. parvum did not grow faster in the presence of prey despite the gains in nutrients, although algal abundances attained in culture were slightly greater in the presence of prey. The relationship between phototrophy, heterotrophy and growth of P. parvum is discussed.

  8. Cryptosporidium avian genotype III as a possible causative agent of chronic vomiting in peach-faced lovebirds (Agapornis roseicollis).

    Science.gov (United States)

    Makino, Ikuko; Abe, Niichiro; Reavill, Drury R

    2010-09-01

    In the present study, Cryptosporidium oocysts were found, by light microscopy, in 37 fecal samples of peach-faced lovebirds (Agapornis roseicollis). Cryptosporidium avian genotype III was isolated in 13 of the 37 infected birds by sequence analysis of the small subunit ribosomal RNA and the actin genes. All of the birds showed chronic vomiting and weight loss with enlargement of isthmi, narrowed proventricular lumens, and thickened proventricular walls radiographically. Cryptosporidium parasites were found only in the ductal epithelium of the proventricular glands in three of the tissue samples provided for necropsy. To date, there have been no reports concerning the pathogenicity, nor the location, of avian genotype III in avian hosts. Our report confirms, for the first time, the presence of avian genotype III in peach-faced lovebirds in Japan and also reveals the location in the avian host.

  9. Occurrence and molecular characterization of Giardia duodenalis and Cryptosporidium spp. in sheep and goats reared under dairy husbandry systems in Greece☆

    Science.gov (United States)

    Tzanidakis, Nikolaos; Sotiraki, Smaragda; Claerebout, Edwin; Ehsan, Amimul; Voutzourakis, Nikolaos; Kostopoulou, Despoina; Stijn, Casaert; Vercruysse, Jozef; Geurden, Thomas

    2014-01-01

    Giardia duodenalis and Cryptosporidium spp. are gastro-intestinal protozoa known to infect small ruminants. Both protozoa are also considered as a potential public health concern. The objective of this study was to determine their prevalence in lambs and goat kids kept under common Mediterranean dairy husbandry systems and to identify the species and genotypes infecting these small ruminants. In total, 684 faecal samples (429 from lambs and 255 from goat kids) were collected on 21 farms in Greece and examined using a quantitative immunofluorescence assay. G. duodenalis was detected in 37.3% of the lambs and 40.4% of the goat kids. On all but one of the farms G. duodenalis was detected. Most samples were typed as a mono-infection with G. duodenalis assemblage E, both on the β-giardin gene and the triose phosphate isomerase gene. Only 10% of samples were typed as mixed assemblage A and E infections. The prevalence of Cryptosporidium spp. was 5.1% in lambs and 7.1% in goat kids. In total, 8 out of the 14 farms with a sheep flock and 7 out of the 14 farms with a goat flock were positive. Cryptosporidium parvum (subtype IId), C. ubiquitum and C. xiaoi were identified, the latter especially in goat kids. In conclusion, the results of the present study illustrate that G. duodenalis and Cryptosporidium spp. occur frequently on both sheep and goats farms. The prevalence of zoonotic genotypes or species was low, indicating a limited but existing risk for zoonotic infections. PMID:25187088

  10. Cryptosporidium rubeyi n. sp. (Apicomplexa: Cryptosporidiidae in multiple Spermophilus ground squirrel species

    Directory of Open Access Journals (Sweden)

    Xunde Li

    2015-12-01

    Full Text Available Previously we reported the unique Cryptosporidium sp. “c” genotype (e.g., Sbey03c, Sbey05c, Sbld05c, Sltl05c from three species of Spermophilus ground squirrel (Spermophilus beecheyi, Spermophilus beldingi, Spermophilus lateralis located throughout California, USA. This follow-up work characterizes the morphology and animal infectivity of this novel genotype as the final step in proposing it as a new species of Cryptosporidium. Analysis of sequences of 18S rRNA, actin, and HSP70 genes of additional Cryptosporidium isolates from recently sampled California ground squirrels (S. beecheyi confirms the presence of the unique Sbey-c genotype in S. beecheyi. Phylogenetic and BLAST analysis indicates that the c-genotype in Spermophilus ground squirrels is distinct from Cryptosporidium species/genotypes from other host species currently available in GenBank. We propose to name this c-genotype found in Spermophilus ground squirrels as Cryptosporidium rubeyi n. sp. The mean size of C. rubeyi n. sp. oocysts is 4.67 (4.4–5.0 μm × 4.34 (4.0–5.0 μm, with a length/width index of 1.08 (n = 220. Oocysts of C. rubeyi n. sp. are not infectious to neonatal BALB/c mice and Holstein calves. GenBank accession numbers for C. rubeyi n. sp. are DQ295012, AY462233, and KM010224 for the 18S rRNA gene, KM010227 for the actin gene, and KM010229 for the HSP70 gene.

  11. Cryptosporidium suis and Cryptosporidium scrofarum in Eurasian wild boars (Sus scrofa) in Central Europe.

    Science.gov (United States)

    Němejc, Karel; Sak, Bohumil; Květoňová, Dana; Hanzal, Vladimír; Janiszewski, Paweł; Forejtek, Pavel; Rajský, Dušan; Ravaszová, Petra; McEvoy, John; Kváč, Martin

    2013-11-08

    From 2011 to 2012, to identify Cryptosporidium spp. occurrence in Eurasian wild boars (Sus scrofa) 29 randomly selected localities (both forest areas and enclosures) across the Central European countries of Austria, the Czech Republic, Poland, and the Slovak Republic were investigated. Cryptosporidium oocysts were microscopicaly detected in 11 out of 460 faecal samples examined using aniline-carbol-methyl violet staining. Sixty-one Cryptosporidium infections, including the 11 infections that were detected by microscopy, were detected using genus- or species-specific nested PCR amplification of SSU rDNA. This represents a 5.5 fold greater sensitivity for PCR relative to microscopy. Combining genus- and species-specific PCR tools significantly changes the perspective on the occurrence of Cryptosporidium spp. in wild boars. While RFLP and direct sequencing of genus specific PCR-amplified products revealed 56 C. suis (20) and C. scrofarum (36) monoinfections and only 5 mixed infections of these species, species-specific molecular tools showed 44 monoinfections and 17 mixed infections with these species. PCR analysis of the gp60 gene did not reveal any other Cryptosporidium infections. Similar to domestic pigs, C. scrofarum was detected as a dominant species infecting adult Eurasian wild boars (Sus scrofa). Cryptosporidium infected wild boars did not show signs of clinical disease. This report is perhaps the most comprehensive survey of cryptosporidial infection in wild boars.

  12. Molecular Epidemiology of Giardia, Blastocystis and Cryptosporidium among Indigenous Children from the Colombian Amazon Basin

    Science.gov (United States)

    Sánchez, Angie; Munoz, Marina; Gómez, Natalia; Tabares, Juan; Segura, Laura; Salazar, Ángela; Restrepo, Cristian; Ruíz, Miguel; Reyes, Patricia; Qian, Yuchen; Xiao, Lihua; López, Myriam C.; Ramírez, Juan D.

    2017-01-01

    The incidence and prevalence of intestinal parasites in children is most likely due to lack of natural or acquired resistance and differences in behavior and habits closely related to environmental and socioeconomic determinants. The most important protozoa that parasitize humans are Giardia, Entamoeba, Blastocystis, and Cryptosporidium. These parasites present wide intraspecific genetic diversity and subsequently classified into assemblages and subtypes. The Amazon basin is the largest in the world and is the fifth freshwater reserve on the planet. Contradictorily, people living in these areas (Indigenous populations) have poor quality of life, which favors the infection of diseases of fecal-oral transmission. The aim of this work was to unravel the molecular epidemiology of Giardia, Blastocystis and Cryptosporidium across four communities (Puerto Nariño, San Juan del Soco, Villa Andrea and Nuevo Paraíso). We obtained 284 fecal samples from children under 15 years old that were analyzed by direct microscopy (261 samples) and Real Time PCR (qPCR) (284 samples). The positive samples for these protozoa were further characterized by several molecular markers to depict assemblages and subtypes. We observed a frequency of Giardia infection by microscopy of 23.7% (62 samples) and by qPCR of 64.8% (184 samples); for Blastocystis by microscopy of 35.2% (92 samples) and by qPCR of 88.7% (252 samples) and for Cryptosporidium only 1.9% (5 samples) were positive by microscopy and qPCR 1.8% (5 samples). Regarding the Giardia assemblages, using the glutamate dehydrogenase (gdh) marker we observed AI, BIII and BIV assemblages and when using triose phosphate isomerase (tpi) we observed assemblages AI, AII, BIII and BIV. In contrast, Blastocystis STs detected were 1, 2, 3, 4, and 6. Lastly, the species C. viatorum, C. hominis (with the subtypes IdA19 and IaA12R8) and C. parvum (with the subtype IIcA5G3c) were identified. We observed a high profile of zoonotic transmission

  13. Ruling out nosocomial transmission of Cryptosporidium in a renal transplantation unit: case report.

    Science.gov (United States)

    Brunet, J; Lemoine, J P; Pesson, B; Valot, S; Sautour, M; Dalle, F; Muller, C; Borni-Duval, C; Caillard, S; Moulin, B; Pfaff, A W; Razakandrainibe, R; Abou-Bacar, A; Favennec, L; Candolfi, E

    2016-08-02

    Cryptosporidium spp. is a ubiquitous parasite affecting humans as well as domestic and wild vertebrates, causing diarrhea in both immunocompetent and immunocompromised hosts worldwide. Its transmission occurs primarily by the fecal-oral route. In humans, C. parvum and C. hominis are the most prevalent species, whereas immunocompetent and immunocompromised individuals can also be infected by other zoonotic species. Renal transplant patients are prone to develop cryptosporidiosis, which can induce severe and life-threatening diarrhea. We report here a series of nearly concomitant cases of acute symptomatic cryptosporidiosis in three renal transplant patients attending the Strasbourg University Hospital Nephrology Unit. The clinical presentation was persistent diarrhea and acute renal failure. The diagnosis was confirmed by microscopic stool examination using a modified Ziehl-Neelsen staining method and species identification by molecular tools. All patients were treated with nitazoxanide and recovered from diarrhea after 14 days of therapy. Genotypic species identification was not consistent with an epidemic context, thus underlining the need for genotyping to monitor at risk patients.

  14. Emissie van Cryptosporidium en Giardia door landbouwhuisdieren

    NARCIS (Netherlands)

    Schrijven JF; Bruin HAM de; Engels GB; Leenen EJTM; MGB

    1999-01-01

    Het hier gepresenteerde deelonderzoek richt zich op de relatieve bijdrage van verschillende populaties landbouwhuisdieren via mest en afvalwater aan de totale emissie van Cryptosporidium en Giardia in Nederland. Vleeskalveren vormen per jaar in Nederland via hun mest een grote emissiebron van be

  15. Cryptosporidium en Giardia in Nederlandse zwembaden

    NARCIS (Netherlands)

    Schets FM; Engels GB; Leenen EJTM; MGB

    2003-01-01

    Zwembad gerelateerde explosies van cryptosporidiose zijn regelmatig gerapporteerd in Groot-Brittannie en de Verenigde Staten. De bron van de explosie kon soms achterhaald worden doordat Cryptosporidium oocysten in het zwembadwater of in het terugspoelwater van de zwembadfilters konden worden gedete

  16. The first report on Cryptosporidium suis and Cryptosporidium pig genotype II in Eurasian wild boars (Sus scrofa) (Czech Republic).

    Science.gov (United States)

    Němejc, Karel; Sak, Bohumil; Květoňová, Dana; Hanzal, Vladimír; Jeníková, Martina; Kváč, Martin

    2012-03-23

    A total of 193 faecal samples of adult Eurasian wild boars were collected at 12 enclosures across the Czech Republic and examined for Cryptosporidium infection using both microscopic and molecular tools. Cryptosporidium oocysts were not detected in any of the 193 faecal samples examined using the aniline-carbol-methyl violet staining method. Thirty-two positive cases of Cryptosporidium infection were detected using either genus- or species-specific nested PCR. Mono-infection with Cryptosporidium suis and Cryptosporidium pig genotype II were found in 13 and 7 cases, respectively. Five mixed infections of C. suis and Cryptosporidium pig genotype II were detected using PCR/RFLP with genus specific primers. The number of detected mixed infections increased 2.4 fold when a species-specific PCR was employed. No other Cryptosporidium spp. was detected. Unlike cryptosporidiosis of domestic pigs, C. suis was detected as a dominant species infecting adult Eurasian wild boars. There was no association between diarrhoea and the presence of Cryptosporidium infection in the Eurasian wild boars studied. This is the first report on the Cryptosporidium infection caused by C. suis and Cryptosporidium pig genotype II in Eurasian wild boars (Sus scrofa).

  17. Assessment of Giardia and Cryptosporidium spp. as a microbial source tracking tool for surface water: application in a mixed-use watershed.

    Science.gov (United States)

    Prystajecky, Natalie; Huck, Peter M; Schreier, Hans; Isaac-Renton, Judith L

    2014-04-01

    Knowledge of host specificity, combined with genomic sequencing of Giardia and Cryptosporidium spp., has demonstrated a microbial source tracking (MST) utility for these common waterborne microbes. To explore the source attribution potential of these pathogens, water samples were collected in a mixed rural-urban watershed in the Township of Langley, in southwestern British Columbia (BC), Canada, over a 2-year period. Cryptosporidium was detected in 63% of surface water samples at concentrations ranging from no positive detection (NPD) to 20,600 oocysts per 100 liters. Giardia was detected in 86% of surface water samples at concentrations ranging from NPD to 3,800 cysts per 100 liters of water. Sequencing at the 18S rRNA locus revealed that 50% of Cryptosporidium samples and 98% of Giardia samples contained species/genotypes (Cryptosporidium) or assemblages (Giardia) that are capable of infecting humans, based on current knowledge of host specificity and taxonomy. Cryptosporidium genotyping data were more promising for source tracking potential, due to the greater number of host-adapted (i.e., narrow-host-range) species/genotypes compared to Giardia, since 98% of Giardia isolates were zoonotic and the potential host could not be predicted. This report highlights the benefits of parasite genomic sequencing to complement Method 1623 (U.S. Environmental Protection Agency) and shows that Cryptosporidium subtyping for MST purposes is superior to the use of Giardia subtyping, based on better detection limits for Cryptosporidium-positive samples than for Giardia-positive samples and on greater host specificity among Cryptosporidium species. These additional tools could be used for risk assessment in public health and watershed management decisions.

  18. Comparative diagnostic techniques for cryptosporidium infection.

    Science.gov (United States)

    Omoruyi, Beauty E; Nwodo, Uchechukwu U; Udem, Chukwuneke S; Okonkwo, Francis O

    2014-02-24

    Diarrhoea caused by Cryptosporidium is usually mild in immune competent individuals but severe in the young and those with underlying disease leading to compromised immunity. The conventional diagnosis of Cryptosporidium requires observation of the infective oocysts however, their tiny size yields indistinct results, thus limiting the effectiveness of the conventional diagnostic technique, modified Ziehl-Neelsen (ZN) differential staining. Consequent to the abovementioned limitation, ZN staining, sandwich antigen detection enzyme linked immunosorbent assay (sad-ELISA) and a direct polymerase chain reaction (PCR) assay techniques were evaluated for diagnostic efficacy. Stool samples were collected from 180 consenting adult patients attending outpatient and inpatient clinics at Victoria Hospital, Alice, Eastern Cape Province of South Africa. Subjects were stratified as; 35 HIV-positive and diarrhoeagenic, 125 HIV-negative diarrhoeagenic and 20 apparently healthy controls. Cryptosporidium incidence following diagnostic techniques were 13 (37.1%; ZN staining), 26 (74.3%; sad-ELISA) and 23 (65.7%; PCR), respectively, among HIV-positive diarrhoeagenic patients and 34 (27.2%; ZN staining), 96 (76.8%; sad-ELISA) and 89 (71.2%; PCR) among HIV-negative diarrhoeagenic patients. Sensitivity, specificity and predictive values of the diagnostic techniques' efficiency were: sensitivity: 46.2% (HIV-positive) and 32.3% (HIV-negative) against the ZN technique and 96.9% against sad-ELISA and PCR, respectively, for both HIV-positive and -negative patients; specificity was 88.9% (HIV-positive) and 96.6% (HIV-negative) against the ZN technique. Lastly, the predictive values were 92.3% (HIV-positive) and 96.9% (HIV-negative), respectively, following ZN staining. The sad-ELISA technique proved more suitable for the determination of the presence of Cryptosporidium oocysts. The high incidence of Cryptosporidium in HIV-positive subjects as compared to the HIV-negative population accentuates

  19. Comparative Diagnostic Techniques for Cryptosporidium Infection

    Directory of Open Access Journals (Sweden)

    Beauty E. Omoruyi

    2014-02-01

    Full Text Available Diarrhoea caused by Cryptosporidium is usually mild in immune competent individuals but severe in the young and those with underlying disease leading to compromised immunity. The conventional diagnosis of Cryptosporidium requires observation of the infective oocysts however, their tiny size yields indistinct results, thus limiting the effectiveness of the conventional diagnostic technique, modified Ziehl-Neelsen (ZN differential staining. Consequent to the abovementioned limitation, ZN staining, sandwich antigen detection enzyme linked immunosorbent assay (sad-ELISA and a direct polymerase chain reaction (PCR assay techniques were evaluated for diagnostic efficacy. Stool samples were collected from 180 consenting adult patients attending outpatient and inpatient clinics at Victoria Hospital, Alice, Eastern Cape Province of South Africa. Subjects were stratified as; 35 HIV-positive and diarrhoeagenic, 125 HIV-negative diarrhoeagenic and 20 apparently healthy controls. Cryptosporidium incidence following diagnostic techniques were 13 (37.1%; ZN staining, 26 (74.3%; sad-ELISA and 23 (65.7%; PCR, respectively, among HIV-positive diarrhoeagenic patients and 34 (27.2%; ZN staining, 96 (76.8%; sad-ELISA and 89 (71.2%; PCR among HIV-negative diarrhoeagenic patients. Sensitivity, specificity and predictive values of the diagnostic techniques’ efficiency were: sensitivity: 46.2% (HIV-positive and 32.3% (HIV-negative against the ZN technique and 96.9% against sad-ELISA and PCR, respectively, for both HIV-positive and -negative patients; specificity was 88.9% (HIV-positive and 96.6% (HIV-negative against the ZN technique. Lastly, the predictive values were 92.3% (HIV-positive and 96.9% (HIV-negative, respectively, following ZN staining. The sad-ELISA technique proved more suitable for the determination of the presence of Cryptosporidium oocysts. The high incidence of Cryptosporidium in HIV-positive subjects as compared to the HIV-negative population

  20. Equine cryptosporidial infection associated with Cryptosporidium hedgehog genotype in Algeria.

    Science.gov (United States)

    Laatamna, Abd Elkarim; Wagnerová, Pavla; Sak, Bohumil; Květoňová, Dana; Aissi, Miriem; Rost, Michael; Kváč, Martin

    2013-10-18

    Faecal samples from two horse farms in Algeria keeping Arabian, Thoroughbred, and Barb horses were examined for the presence of Cryptosporidium in 2010-2011. A total of 138 faecal samples (16 from a farm keeping 50 animals and 122 from a farm with 267 horses) were screened for Cryptosporidium spp. infection using molecular tools. DNA was extracted from all samples. Nested PCR was performed to amplify fragments of the SSU rDNA and gp60 genes to determine the presence of Cryptosporidium species and genotypes. Sequence analyses of SSU and gp60 genes revealed four animals positive for the presence of subtype XIIIa A22R9 of the Cryptosporidium hedgehog genotype. The infections were not associated with diarrhoea. This study reports, for the first time, the occurrence of Cryptosporidium in Algeria and the first occurrence of the hedgehog genotype in horses. These findings support the potential role of infected horses in sylvatic-domestic transmission of Cryptosporidium.

  1. Stability of the intra- and extracellular toxins of Prymnesium parvum using a microalgal bioassay

    DEFF Research Database (Denmark)

    Blossom, Hannah Eva; Andersen, Nikolaj Gedsted; Rasmussen, Silas Anselm

    2014-01-01

    Prymnesium parvum produces a variety of toxic compounds, which affect other algae, grazers and organisms at higher trophic levels. Here we provide the method for development of a sensitive algal bioassay using a microalgal target, Teleaulax acuta, to measure strain variability in P. parvum toxicity...

  2. New system for higher recovery rate of water borne Cryptosporidium oocysts and Giardia cysts

    DEFF Research Database (Denmark)

    Al-Sabi, Mohammad Nafi Solaiman; Gad, Jens; Klinting, Mette

    2012-01-01

    motivate the development of systems with higher recovery rates. Materials and methods: Five replicates of IMS purified Cryptosporidium oocysts and Giardia cysts (N=2x103) were injected into a specially coated filter unit with a carefully chosen pore size. Following filtration, sonication was performed...... at predetermined time intervals to loosen all parasites and other particles from the filter and break up clusters. The concentrated parasite suspension was backwashed and counted after immuno-fluorescence staining. Results and discussion: Without sonication the recovery rates of both Cryptosporidium and Giardia...... chamber further facilitated the concentration of parasites by ensuring a backwash volume of less than 1ml. The presented design of the filter system can be used as a cheap and time effective method of isolating water borne parasites in the laboratory....

  3. Removal of Cryptosporidium by wastewater treatment processes: a review.

    Science.gov (United States)

    Nasser, Abidelfatah M

    2016-02-01

    Cryptosporidium is a protozoan parasite that infects humans and various animal species. The environmental stability and the low infectious dose of Cryptosporidium facilitate its transmission by water and food. Discharge of untreated wastewater may result in waterborne or foodborne Cryptosporidium outbreaks, therefore a suitable treatment may prevent its dissemination. Most studies on the prevalence of Cryptosporidium oocysts in wastewater have reported a concentration range between 10 and 200 oocysts/L and a prevalence of 6 to 100%. Activated sludge has been found to be ineffective for the removal of Cryptosporidium oocysts. Stabilization ponds and constructed wetlands are efficient for the reduction of Cryptosporidium from wastewater, especially when the retention time is longer than 20 days at suitable sunlight and temperature. High rate filtration and chlorine disinfection are inefficient for the reduction of Cryptosporidium from effluents, whereas ultrafiltration and UV irradiation were found to be very efficient for the reduction of Cryptosporidium oocysts. Adequate tertiary treatment may result in high quality effluent with low risk of Cryptosporidium for unrestricted irrigation and other non-potable applications.

  4. Growth and extracellular laccase production in liquid cultures of Minimidochium parvum LPSC # 548 Strain Crecimiento y producción de lacasa extracelular en cultivos líquidos de Minimidochium parvum cepa LPSC # 548

    Directory of Open Access Journals (Sweden)

    Mario C. N. Saparrat

    2007-07-01

    Full Text Available Minimidochium parvum LPSC # 548, a fungus isolated from litter floating on waters of Río Santiago (Provincia de Buenos Aires, Argentina polluted with industrial effluents and crude-oil, was grown as a shaking culture on a C-limited medium to evaluate its ability to produce extracellular laccase. The effect of anthracene, CuSO4 · 5H2O, ethanol, guaiacol, humic acids, Kraft lignin, MnSO4· H2O, Tween 20 and veratryl alcohol on its growth and extracellular laccase activity levels was also analyzed. The cultures grown on basal medium produced maximum biomass (over 420 mg/100 ml and maximum extracellular laccase activity (351.7 ±53.3 pkat/ml after 5 days of incubation. Among the different factors tested, only the humic acids at 0.1 % (w/v were found to stimulate the growth of M. parvum . However, Tween 20 (0.1 %, v/v was the only one that produced an increase of laccase activity levels up to 2.5-fold compared to the control.Minimidochium parvum LPSC # 548, un hongo aislado de materia orgánica colectada en aguas de Río Santiago (Provincia de Buenos Aires, Argentina contaminadas con efluentes industriales y crudo de petróleo, se cultivó en un medio líquido limitante en carbono bajo agitación para evaluar su habilidad para producir lacasa extracelular. Se analizó también el efecto de ácidos húmicos, alcohol veratrílico, antraceno, CuSO4 · 5H2O, etanol, guaiacol, lignina Kraft, MnSO4· H2O y Tween 20 sobre el crecimiento fúngico y los niveles de actividad lacasa extracelular. Los cultivos sobre medio basal produjeron máximos niveles de biomasa (superior a 420 mg/100 ml y actividad lacasa extracelular (351,7 ±53,3 pkat/ml después de 5 días de incubación. Entre los diferentes agentes químicos testeados, sólo los ácidos húmicos al 0,1 % (p/v estimularon el crecimiento de M. parvum . No obstante, sólo el Tween 20 (0,1 %, v/v produjo un incremento de los niveles de actividad lacasa (2,5 veces comparado a cultivos control.

  5. Subtyping of Cryptosporidium cuniculus and genotyping of Enterocytozoon bieneusi in rabbits in two farms in Heilongjiang Province, China

    Directory of Open Access Journals (Sweden)

    Yang Ziyin

    2016-01-01

    Full Text Available Cryptosporidium spp. and Enterocytozoon bieneusi are two prevalent opportunistic pathogens in humans and animals. Currently, few data are available on genetic characterization of both pathogens in rabbits in China. The aim of the present study was to understand prevalence and genetic characterization of Cryptosporidium spp. and E. bieneusi in rabbits. We collected 215 fecal samples from 150 Rex rabbits and 65 New Zealand White rabbits on two different farms in Heilongjiang Province, China. Cryptosporidium spp. and E. bieneusi were tested by polymerase chain reaction (PCR and sequencing the partial small subunit of ribosomal DNA (SSU rDNA and the internal transcribed spacer (ITS region of rDNA, respectively. Cryptosporidium was detected in 3.3% (5/150 of Rex rabbits and 29.2% (19/65 of New Zealand White rabbits. All the 24 Cryptosporidium isolates were identified as C. cuniculus. Enterocytozoon bieneusi was only found in 14.7% (22/150 of Rex rabbits. Five known genotypes: CHN-RD1 (n = 12, D (n = 3, Type IV (n = 2, Peru6 (n = 1, and I (n = 1, and three novel ones CHN-RR1 to CHN-RR3 (one each were detected. By analyzing the 60-kDa glycoprotein (gp60 gene sequences of C. cuniculus isolates, three subtypes were obtained: VbA28 (n = 2, VbA29 (n = 16, and VbA32 (n = 3. All these three C. cuniculus subtypes were reported previously in humans. Four known E. bieneusi genotypes have been found to be present in humans. The three novel ones fell into zoonotic group 1. The results suggest zoonotic potential of C. cuniculus and E. bieneusi isolates in rabbits.

  6. Subtyping of Cryptosporidium cuniculus and genotyping of Enterocytozoon bieneusi in rabbits in two farms in Heilongjiang Province, China.

    Science.gov (United States)

    Yang, Ziyin; Zhao, Wei; Shen, Yujuan; Zhang, Weizhe; Shi, Ying; Ren, Guangxu; Yang, Di; Ling, Hong; Yang, Fengkun; Liu, Aiqin; Cao, Jianping

    2016-01-01

    Cryptosporidium spp. and Enterocytozoon bieneusi are two prevalent opportunistic pathogens in humans and animals. Currently, few data are available on genetic characterization of both pathogens in rabbits in China. The aim of the present study was to understand prevalence and genetic characterization of Cryptosporidium spp. and E. bieneusi in rabbits. We collected 215 fecal samples from 150 Rex rabbits and 65 New Zealand White rabbits on two different farms in Heilongjiang Province, China. Cryptosporidium spp. and E. bieneusi were tested by polymerase chain reaction (PCR) and sequencing the partial small subunit of ribosomal DNA (SSU rDNA) and the internal transcribed spacer (ITS) region of rDNA, respectively. Cryptosporidium was detected in 3.3% (5/150) of Rex rabbits and 29.2% (19/65) of New Zealand White rabbits. All the 24 Cryptosporidium isolates were identified as C. cuniculus. Enterocytozoon bieneusi was only found in 14.7% (22/150) of Rex rabbits. Five known genotypes: CHN-RD1 (n = 12), D (n = 3), Type IV (n = 2), Peru6 (n = 1), and I (n = 1), and three novel ones CHN-RR1 to CHN-RR3 (one each) were detected. By analyzing the 60-kDa glycoprotein (gp60) gene sequences of C. cuniculus isolates, three subtypes were obtained: VbA28 (n = 2), VbA29 (n = 16), and VbA32 (n = 3). All these three C. cuniculus subtypes were reported previously in humans. Four known E. bieneusi genotypes have been found to be present in humans. The three novel ones fell into zoonotic group 1. The results suggest zoonotic potential of C. cuniculus and E. bieneusi isolates in rabbits. © Z. Yang et al., published by EDP Sciences, 2016.

  7. Potential cryptosporidium surrogates and evaluation of compressible oocysts

    Energy Technology Data Exchange (ETDEWEB)

    Li, S.Y.; Goodrich, J.A.; Owens, J.H. [Environmental Protection Agency, Cincinnati, OH (United States)] [and others

    1995-10-01

    Cryptosporidium has been recognized as an important waterborne agent of gastroenteritis and a biological contaminant in drinking water. The widespread presence of Cryptosporidium in surface source water and either untreated or insufficiently treated drinking water has led to Cryptosporidium outbreaks in the United States and worldwide. Among the conventional control practices, filtration and high temperature distillation appear to be the potentially viable technologies for protection against Cryptosporidium in drinking water. As employed in many water plants, filtration is likely to be the most practical treatment technology utilized for Cryptosporidium removal in the near future. Consequently, accurate and reliable methods for evaluation of Cryptosporidium removal rates for filtration-based systems are necessary to assist States in determining drinking water quality and complying with the up-coming national standard for Cryptosporidium in drinking water. Furthermore, searching for reliable and non-hazardous surrogates for evaluation of treatment plant efficiency has been intensified because of the potential health risk associated with Cryptosporidium. Additionally, during the filtration procedure Cryptosporidium may squeeze and fold through pores size of the filtration systems that are smaller than the diameter of the organism; a fraction of these Cryptosporidium oocysts may still remain a certain degree of viability. These uncertainties are critical for the evaluation and optimization of filtration-based physical treatment systems. The in-house research studies described below consist of two parts. One is a potential surrogate study using bag filtration systems at the US EPA Test & Evaluation Facility in Cincinnati, Ohio. The second is Cryptosporidium compressibility and viability investigation.

  8. Isolation and characterization of species-specific DNA probes from Taenia solium and Taenia saginata and their use in an egg detection assay.

    Science.gov (United States)

    Chapman, A; Vallejo, V; Mossie, K G; Ortiz, D; Agabian, N; Flisser, A

    1995-05-01

    Cysticercosis results from ingestion of the eggs of the tapeworm Taenia solium. Reduction of the incidence of human and swine cysticercosis requires identification and treatment of individuals who carry the adult tapeworm. T. solium and Taenia saginata eggs cannot be differentiated on the basis of morphology; thus, in order to improve existing methods for the diagnosis of taeniasis, we have developed highly sensitive, species-specific DNA probes which differentiate T. solium and T. saginata. Recombinant clones containing repetitive DNA sequences which hybridize specifically with genomic DNAs from either species were isolated and characterized. T. solium-specific DNA sequences contained complete and truncated forms of a tandemly repeated 158-bp DNA sequence. An unrelated T. saginata DNA sequence was also characterized and shown to encode a portion of the mitochondrial cytochrome c oxidase I gene. T. solium- and T. saginata-specific DNA probes did not hybridize in dot blot assays either with genomic DNA from the platyhelminths Taenia hydatigena, Taenia pisiformis, Taenia taeniaeformis, Echinococcus granulosus, and Schistosoma mansoni or with genomic DNA from other eukaryotes, including Saccharomyces cerevisiae, Candida albicans, Cryptosporidium parvum, Entamoeba histolytica, Trypanosoma gambiense, Trypanosoma brucei, and Giardia lamblia, Caenorhabditis elegans, and human DNA. By using these T. solium and T. saginata DNA probes, a rapid, highly sensitive and specific dot blot assay for the detection of T. solium eggs was developed.

  9. Prevalence of Cryptosporidium oocysts and Giardia cysts in raw and treated sewage sludges.

    Science.gov (United States)

    Amorós, Inmaculada; Moreno, Yolanda; Reyes, Mariela; Moreno-Mesonero, Laura; Alonso, Jose L

    2016-11-01

    Treated sludge from wastewater treatment plants (WWTPs) is commonly used in agriculture as fertilizers and to amend soils. The most significant health hazard for sewage sludge relates to the wide range of pathogenic microorganisms such as protozoa parasites.The objective of this study was to collect quantitative data on Cryptosporidium oocysts and Giardia cysts in the treated sludge in wastewater treatment facilities in Spain. Sludge from five WWTPs with different stabilization processes has been analysed for the presence of Cryptosporidium and Giardia in the raw sludge and after the sludge treatment. A composting plant (CP) has also been assessed. After a sedimentation step, sludge samples were processed and (oo)cysts were isolated by immunomagnetic separation (IMS) and detected by immunofluorescence assay (IFA). Results obtained in this study showed that Cryptosporidium oocysts and Giardia cysts were present in 26 of the 30 samples (86.6%) of raw sludge samples. In treated sludge samples, (oo)cysts have been observed in all WWTP's analysed (25 samples) with different stabilization treatment (83.3%). Only in samples from the CP no (oo)cysts were detected. This study provides evidence that (oo)cysts are present in sewage sludge-end products from wastewater treatment processes with the negative consequences for public health.

  10. Effect of imbalanced nutrients and immigration on Prymnesium parvum community dominance and toxicity: Results from in-lake microcosm experiments

    Science.gov (United States)

    Errera, R.M.; Roelke, D.L.; Kiesling, R.L.; Brooks, B.W.; Grover, J.P.; Schwierzke, L.; Urena-Boeck, F.; Baker, J.W.; Pinckney, J.L.

    2008-01-01

    Prymnesium parvum, a haptophyte species, forms harmful blooms, including those that have caused severe fish kills in Texas, USA, over the past 6 yr. We studied P. parvum dynamics using in situ microcosm experiments at Lake Possum Kingdom, Texas, during 3 seasons (fall 2004, winter and spring 2005). Experimental treatments included full and partial nutrient enrichment (encompassing nitrogen [N] and phosphorus [P] deficient treatments), P. parvum immigration and combinations of these factors. In the control and N and P deficient treatments, P. parvum populations dominated the community, but only in the N deficient treatments did P. parvum experience a significant growth in the population. In contrast, when nutrients were not limiting, P. parvum tended to lose its competitive edge to other taxa such as chlorophytes, euglenophytes and diatoms, which then dominated the community. Population growth of P. parvum was also stimulated through immigration, but only during the winter experiment, a period of the year when bloom initiation is common. This finding suggests that movement into the water column may be an important process leading to P. parvum bloom initiation. Toxicity of P. parvum to fish was also affected by the nutrient changes: during conditions of no nutrient addition P. parvum was most toxic; intermediate toxicity was observed under N and P deficient conditions, and full nutrient enrichments resulted in nearly non-toxic conditions. ?? Inter-Research 2008.

  11. Prevalence of Cryptosporidium sp. in patients with colorectal cancer.

    Science.gov (United States)

    Sulżyc-Bielicka, Violetta; Kołodziejczyk, Lidia; Jaczewska, Sylwia; Bielicki, Dariusz; Kładny, Józef; Safranow, Krzysztof

    2012-07-01

    Parasitic protozoans of the Cryptosporidium genus are intracellular intestinal parasites of mammals, causing cryptosporidiosis. Clinically, cryptosporidiosis manifests as chronic diarrhoea. Individuals with immune disorders, including those with neoplasms, are at risk of symptomatic invasion. Was the evaluation of Cryptosporidium sp. prevalence in patients with diagnosed colorectal cancer. The studied group encompassed 87 patients with diagnosed colorectal cancer, undergoing surgery at the Department of General and Oncological Surgery, Pomeranian Medical University, in the years 2009-2010. Immunoenzymatic tests for Cryptosporidium sp. on faeces samples were performed with the use of commercial test kit, ProSpecT(®)Cryptosporidium Microplate Assay (Remel Inc). The presence of Cryptosporidium sp. was found in 12.6% of studied patients with colorectal cancer. The performed statistical analysis did not reveal any correlation between Cryptosporidium sp. infection and gender, age, neoplasm advancement stage as per Astler-Coller scale, neoplasm differentiation grade, or neoplastic tumour localisation in relation to the splenic flexure. There was found high prevalence of Cryptosporidium sp. in patients with colorectal cancer. It was comparable to the prevalence reported for patients with immune deficiency.

  12. Detection of Cryptosporidium sp infection by PCR and modified acid fast staining from potassium dichromate preserved stool

    Directory of Open Access Journals (Sweden)

    Agnes Kurniawan

    2009-09-01

    Full Text Available Aim To identify the frequency of Cryptosporidium infection in children below 3 years old by examining concentrated long term preserved stool using PCR detection of 18S rRNA gene and compared with modified acid fast staining technique.Methods Hundred eighty eight stools from children ≤ 3 years old were stored for 13 months in 2.5% K2Cr2O7 solution at 40C. Cryptosporidium oocysts were isolated by water-ether concentration technique. The concentrates were smeared onto object glass and stained with modified acid fast staining, and the rest of the concentrates were DNA extracted by freezing and thawing cycles and proteinase K digestion, then direct PCR was done to detect 18S rRNA gene.Result The proportion of positive stools for Cryptosporidium sp by acid fast staining from concentrated stools and 18S rRNA PCR were 4.8% and 34.6% respectively, which showed statistically significant difference.Conclusion The frequency of Cryptosporidium infection among children ≤ 3 years old was very high and stool storage in K2Cr2O7 for 13 months did not affect the PCR result. High prevalence of Cryptosporidium infection indicated high transmission in that area and the potential to be transmitted to other individuals such as the immunocompromised. (Med J Indones 2009;18:147-52Key words: 18S rRNA, cryptosporidiosis

  13. Treatment of common warts with the immune stimulant Propionium bacterium parvum.

    Science.gov (United States)

    Nasser, Nilton

    2012-01-01

    Warts are epithelial proliferations in the skin and mucous membrane caused by various types of HPV. They can decrease spontaneously or increase in size and number according to the patient's immune status. The Propionium bacterium parvum is a strong immune stimulant and immune modulator and has important effects in the immune system and it is able to produce antibodies in the skin. To show the efficacy of the Propionium bacterium parvum in saline solution in the treatment of skin warts. A randomized double-blind study. Twenty patients with multiple warts were divided into two groups: one received 0,1 ml intradermal injection of placebo solution in just one of the warts and the other received 0,1 ml of saline solution of Propionium bacterium parvum, one dose a month, for 3 to 5 months. Among the 20 patients who participated in the study, ten received the placebo and ten received the saline solution with Propionium bacterium parvum. In 9 patients treated with the Propionium bacterium parvum solution the warts disappeared without scars and in 1 patient it decreased in size. In 9 patients who received the placebo no change to the warts was observed and in 1 it decreased in size. The immune modulator and immune stimulant Propionium bacterium parvum produced antibodies in the skin which destroyed the warts without scars, with statistically significant results (Pwarts.

  14. Detection and molecular diversity of Giardia duodenalis and Cryptosporidium spp. in sheltered dogs and cats in Northern Spain.

    Science.gov (United States)

    Gil, Horacio; Cano, Lourdes; de Lucio, Aida; Bailo, Begoña; de Mingo, Marta Hernández; Cardona, Guillermo A; Fernández-Basterra, José A; Aramburu-Aguirre, Juan; López-Molina, Nuria; Carmena, David

    2017-06-01

    Domestic dogs and cats may act as natural reservoirs of a large number of zoonotic pathogens, including the enteric parasites Giardia duodenalis and Cryptosporidium spp., the most relevant protozoan species causing gastrointestinal disease worldwide. A cross-sectional epidemiological study aiming to assess the prevalence and molecular diversity of G. duodenalis and Cryptosporidium spp. was conducted in an animal rescue centre in the province of Álava (Northern Spain). A total of 194 and 65 faecal dropping samples from individual dogs and cats, respectively, were collected between November 2013 and June 2016. G. duodenalis cysts and Cryptosporidium spp. oocysts were detected by direct fluorescence microscopy and PCR-based methods targeting the small subunit ribosomal RNA gene of these parasites. Overall, G. duodenalis and Cryptosporidium spp. were detected in 33% (63/194) and 4.1% (8/194) of dogs, and 9.2% (6/65) and 4.6% (3/65) of cats, respectively. G. duodenalis and Cryptosporidium co-infections were observed in 1.5% (3/194) of dogs, but not in cats. No significant differences in infection rates could be demonstrated among dogs or cats according to their sex, age group, status, or geographical origin. Multi-locus sequence-based genotyping of the glutamate dehydrogenase and β-giardin genes of G. duodenalis allowed the characterization of 19 canine isolates that were unambiguously assigned to sub-assemblages AII (n=7), BIII (n=1), and BIV (n=7), and assemblages C (n=3) and D (n=1). Two feline isolates were genotyped as assemblages A and F, respectively. No mixed assemblage or sub-assemblage infections were identified. C. canis (n=5) and C. hominis (n=1) were the Cryptosporidium species found in dogs, whereas C. felis (n=1) was identified in cats. The finding of G. duodenalis sub-assemblages AII, BIII, and BIV circulating in dogs (but not cats) may have zoonotic potential, although most of the AII and BIV isolates sub-genotyped corresponded to genetic variants not

  15. Cryptosporidium varanii infection in leopard geckos (Eublepharis macularius in Argentina

    Directory of Open Access Journals (Sweden)

    A. Dellarupe

    2016-06-01

    Full Text Available Cryptosporidiosis is observed in reptiles with high morbidity and considerable mortality. The objective of this study was to achieve the molecular identification of Cryptosporidium spp. in pet leopard geckos (Eublepharis macularius from a breeder colony in Buenos Aires, Argentina. Oocysts comparable to those of Cryptosporidium spp. were detected in three geckos with a history of diarrhea, anorexia and cachexia. Molecular identification methods confirmed the presence of Cryptosporidium varanii (syn. C. saurophilum. This agent was considered to be the primary cause of the observed clinical disease. This is the first description of C. varanii infection in pet reptiles in Argentina.

  16. Effects of drinking-water filtration on Cryptosporidium seroepidemiology, Scotland.

    Science.gov (United States)

    Ramsay, Colin N; Wagner, Adam P; Robertson, Chris; Smith, Huw V; Pollock, Kevin G J

    2014-01-01

    Continuous exposure to low levels of Cryptosporidium oocysts is associated with production of protective antibodies. We investigated prevalence of antibodies against the 27-kDa Cryptosporidium oocyst antigen among blood donors in 2 areas of Scotland supplied by drinking water from different sources with different filtration standards: Glasgow (not filtered) and Dundee (filtered). During 2006-2009, seroprevalence and risk factor data were collected; this period includes 2007, when enhanced filtration was introduced to the Glasgow supply. A serologic response to the 27-kDa antigen was found for ≈75% of donors in the 2 cohorts combined. Mixed regression modeling indicated a 32% step-change reduction in seroprevalence of antibodies against Cryptosporidium among persons in the Glasgow area, which was associated with introduction of enhanced filtration treatment. Removal of Cryptosporidium oocysts from water reduces the risk for waterborne exposure, sporadic infections, and outbreaks. Paradoxically, however, oocyst removal might lower immunity and increase the risk for infection from other sources.

  17. Clinical and subclinical infections with Giardia and Cryptosporidium in animals

    Science.gov (United States)

    Giardia and Cryptosporidium are frequent parasites of livestock, companion animals, and wildlife, raising questions about the clinical significance of such infections. Infections with both parasites have a wide spectrum of symptoms that can vary between asymptomatic infections to serious infection ...

  18. Cryptosporidium infection in cattle in Ogun state, Nigeria

    African Journals Online (AJOL)

    ADEYEYE

    2014-06-30

    Jun 30, 2014 ... different studies to identify Cryptosporidium spp. (Kaur et al., 2002; Mahdi .... Pakistan Journal of Biological Sciences, .... Empty rectum and caudal abdominal pain with crepitating .... 1.pdf, retrieved 25-05-2012. Anonymous ...

  19. Study on Cryptosporidium contamination in vegetable farms around Tehran.

    Science.gov (United States)

    Ranjbar-Bahadori, Sh; Mostoophi, A; Shemshadi, B

    2013-06-01

    In recent years, an increase in the number of cases of food-borne illnesses linked to fresh vegetables has been reported. One of the causative agents of these infections is Cryptosporidium and it appears that one route of transmission to humans is food-borne, so fruits and vegetables have important roles. The goal of this study was to determine the level of Cryptosporidium contamination in vegetable farms around Tehran, Iran. A total of 496 samples from 115 vegetable farms in different regions around Tehran (Capital city of Iran) were collected and different types of vegetables were investigated for the parasite in June and July, 2012. A sediment concentration method followed by modified Ziehl-Neelsen's acid-fast staining was used to determine the presence of Cryptosporidium oocysts. Our findings revealed that 6.6% of studied samples were contaminated with Cryptosporidium species. The highest rate of contamination was reported in Bagher Abad (South of Tehran) (11.1%), and green onions were more commonly contaminated (14.8%) than any other vegetables tested. Furthermore, when waste water was used to irrigate vegetable farms, the contamination rate was (33.3%). Statistical analysis showed a correlation between contamination with Cryptosporidium spp. and studied risk factors including: different regions around Tehran, type of vegetables, and type of water used for farm irrigation. Therefore, vegetables may provide a route by which Cryptosporidium can be transmitted to humans, and control strategies should be considered.

  20. Parasitismo por Giardia sp. e Cryptosporidium sp. em Coendou villosus Parasitism by Giardia sp. and Cryptosporidium sp. in Coendou villosus

    Directory of Open Access Journals (Sweden)

    João Fabio Soares

    2008-04-01

    Full Text Available O objetivo deste trabalho foi verificar o possível parasitismo por Giardia sp. e Cryptosporidium sp. em amostras de fezes de ouriço-cacheiro (Coendou villosus. As amostras foram analisadas pelo método de centrífugo-flutuação com sulfato de zinco e apresentaram elevada infecção por cistos de Giardia sp. e por oocistos de Cryptosporidium sp., embora os animais não apresentassem sinal clínico decorrente disso.This research was aimed at verifing the possible parasitism by Giardia sp. and Cryptosporidium sp. in porcupine (Coendou villosus faeces samples. Samples were analyzed by the centrifugal-flotation method with zinc sulphate and showed high infection by cysts of Giardia sp. and by oocysts of Cryptosporidium sp., although the animals did not show any associated clinical sign.

  1. Detection of Cryptosporidium hominis and novel Cryptosporidium bat genotypes in wild and captive Pteropus hosts in Australia.

    Science.gov (United States)

    Schiller, Sabine Eva; Webster, Koa Narelle; Power, Michelle

    2016-10-01

    Spillover of zoonotic pathogens from wildlife to humans has been identified as a primary threat to global health. In contrast, the process of reverse pathogen transmission (zooanthroponosis), whereby pathogens move from humans into wildlife species is still largely unexplored. Globally, increasing urbanisation and habitat loss are driving many wildlife species into urban and regional centres. In Australia, large numbers of flying foxes now live in close proximity to humans, increasing the risk of zooanthroponosis. The protozoan parasite Cryptosporidium is an emerging zoonotic parasite that infects a wide range of vertebrates yet there are limited studies on transmission potential of Cryptosporidium between humans and urban wildlife. To explore the presence of zooanthroponosis in flying foxes in Australia the occurrence and diversity of Cryptosporidium was investigated in urbanised wild and captive flying foxes. PCR screening of faecal samples (n=281) from seven wild sites and two captive facilities identified the presence of Cryptosporidium in 3.2% (95% CI 1.5% to 6.0%) of faecal samples. In faecal samples from wild sites Cryptosporidium occurrence was 1.7% (95% CI 0.3% to 4.8%) versus 5.9% (95% CI 2.2% to 12.4%) in samples from captive individuals, with no significant difference between captive and wild sites (p=0.077). Multilocus sequencing using three loci, 18s rDNA, actin and gp60 was used to identify Cryptosporidium in flying fox species. The host specific Cryptosporidium hominis was identified in faecal samples from two captive flying foxes, and one of these samples was confirmed as C. hominis at both actin and gp60. Sequencing of the 18s rDNA also revealed four novel Cryptosporidium genotypes in wild and captive individuals, actin and gp60 amplification and sequencing were unreliable for all four novel genotypes. These novel genotypes have been designated Cryptosporidium bat genotypes VIII-XI. This first report of Cryptosporidium spp. in Australian flying

  2. EVALUATING CRYPTOSPORIDIUM AND GIARDIA IN STORMWATER AS A THREAT TO DRINKING WATER SUPPLIES

    Science.gov (United States)

    Since the first identified Cryptosporidium outbreak in the United Kingdom in 1983, thepathogens Cryptosporidium and Giardia have become the subject of growing local, state, andnational concern. Both organisms have been the causative agent of many gastrointestinalilln...

  3. Molecular characterization of Cryptosporidium spp. in domestic pigeons (Columba livia domestica) in Guangdong Province, Southern China.

    Science.gov (United States)

    Li, Juan; Lin, Xuhui; Zhang, Longxian; Qi, Nanshan; Liao, Shenquan; Lv, Minna; Wu, Caiyan; Sun, Mingfei

    2015-06-01

    To investigate the prevalence and assess the zoonotic transmission burden of Cryptosporidium species in domestic pigeons in Guangdong Province, Southern China, 244 fecal samples were collected from four pigeon breeding farms between June 2012 and March 2013. Cryptosporidium oocysts were purified by Sheather's sugar flotation technique and characterized by DNA sequencing of small subunit ribosomal RNA (SSU rRNA) gene. Cryptosporidium species were determined by comparison of sequences with corresponding Cryptosporidium sequences in GenBank and phylogenetic analysis using neighbor-joining (NJ) in MEGA5.2. The overall prevalence of Cryptosporidium infection in domestic pigeons in Guangdong Province was 0.82% (2/244). Two Cryptosporidium species, namely Cryptosporidium baileyi and Cryptosporidium meleagridis, were identified in Huizhou and Chaozhou farm, respectively. These findings confirmed the existence of C. meleagridis infection in domestic pigeons in China for the first time and provided base-line information for further studies to evaluate the public health risk from pigeon to human.

  4. Giardia duodenalis and Cryptosporidium occurrence in Australian sea lions (Neophoca cinerea exposed to varied levels of human interaction

    Directory of Open Access Journals (Sweden)

    Tiffany C. Delport

    2014-12-01

    Full Text Available Giardia and Cryptosporidium are amongst the most common protozoan parasites identified as causing enteric disease in pinnipeds. A number of Giardia assemblages and Cryptosporidium species and genotypes are common in humans and terrestrial mammals and have also been identified in marine mammals. To investigate the occurrence of these parasites in an endangered marine mammal, the Australian sea lion (Neophoca cinerea, genomic DNA was extracted from faecal samples collected from wild populations (n = 271 in Southern and Western Australia and three Australian captive populations (n = 19. These were screened using PCR targeting the 18S rRNA of Giardia and Cryptosporidium. Giardia duodenalis was detected in 28 wild sea lions and in seven captive individuals. Successful sequencing of the 18S rRNA gene assigned 27 Giardia isolates to assemblage B and one to assemblage A, both assemblages commonly found in humans. Subsequent screening at the gdh and β-giardin loci resulted in amplification of only one of the 35 18S rRNA positive samples at the β-giardin locus. Sequencing at the β-giardin locus assigned the assemblage B 18S rRNA confirmed isolate to assemblage AI. The geographic distribution of sea lion populations sampled in relation to human settlements indicated that Giardia presence in sea lions was highest in populations less than 25 km from humans. Cryptosporidium was not detected by PCR screening in either wild colonies or captive sea lion populations. These data suggest that the presence of G. duodenalis in the endangered Australian sea lion is likely the result of dispersal from human sources. Multilocus molecular analyses are essential for the determination of G. duodenalis assemblages and subsequent inferences on transmission routes to endangered marine mammal populations.

  5. Ureaplasma parvum serovar 3 multiple banded antigen size variation after chronic intra-amniotic infection/colonization.

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    James W Robinson

    Full Text Available Ureaplasma species are the microorganisms most frequently associated with adverse pregnancy outcomes. The multiple banded antigen (MBA, a surface-exposed lipoprotein, is a key virulence factor of ureaplasmas. The MBA demonstrates size variation, which we have shown previously to be correlated with the severity of chorioamnion inflammation. We aimed to investigate U. parvum serovar 3 pathogenesis in vivo, using a sheep model, by investigating: MBA variation after long term (chronic and short term (acute durations of in utero ureaplasma infections, and the severity of chorioamnionitis and inflammation in other fetal tissues. Inocula of 2 × 10(7 colony-forming-units (CFU of U. parvum serovar 3 (Up or media controls (C were injected intra-amniotically into pregnant ewes at one of three time points: day 55 (69d Up, n = 8; C69, n = 4; day 117 (7d Up, n = 8; C7, n = 2; and day 121 (3d Up, n = 8; C3, n = 2 of gestation (term = 145-150d. At day 124, preterm fetuses were delivered surgically. Samples of chorioamnion, fetal lung, and umbilical cord were: (i snap frozen for subsequent ureaplasma culture, and (ii fixed, embedded, sectioned and stained by haematoxylin and eosin stain for histological analysis. Selected fetal lung clinical ureaplasma isolates were cloned and filtered to obtain cultures from a single CFU. Passage 1 and clone 2 ureaplasma cultures were tested by western blot to demonstrate MBA variation. In acute durations of ureaplasma infection no MBA variants (3d Up or very few MBA variants (7d Up were present when compared to the original inoculum. However, numerous MBA size variants were generated in vivo (alike within contiguous tissues, amniotic fluid and fetal lung, but different variants were present within chorioamnion, during chronic, 69d exposure to ureaplasma infection. For the first time we have shown that the degree of ureaplasma MBA variation in vivo increased with the duration of gestation.

  6. Ureaplasma parvum serovar 3 multiple banded antigen size variation after chronic intra-amniotic infection/colonization.

    Science.gov (United States)

    Robinson, James W; Dando, Samantha J; Nitsos, Ilias; Newnham, John; Polglase, Graeme R; Kallapur, Suhas G; Pillow, J Jane; Kramer, Boris W; Jobe, Alan H; Payton, Diane; Knox, Christine L

    2013-01-01

    Ureaplasma species are the microorganisms most frequently associated with adverse pregnancy outcomes. The multiple banded antigen (MBA), a surface-exposed lipoprotein, is a key virulence factor of ureaplasmas. The MBA demonstrates size variation, which we have shown previously to be correlated with the severity of chorioamnion inflammation. We aimed to investigate U. parvum serovar 3 pathogenesis in vivo, using a sheep model, by investigating: MBA variation after long term (chronic) and short term (acute) durations of in utero ureaplasma infections, and the severity of chorioamnionitis and inflammation in other fetal tissues. Inocula of 2 × 10(7) colony-forming-units (CFU) of U. parvum serovar 3 (Up) or media controls (C) were injected intra-amniotically into pregnant ewes at one of three time points: day 55 (69d Up, n = 8; C69, n = 4); day 117 (7d Up, n = 8; C7, n = 2); and day 121 (3d Up, n = 8; C3, n = 2) of gestation (term = 145-150d). At day 124, preterm fetuses were delivered surgically. Samples of chorioamnion, fetal lung, and umbilical cord were: (i) snap frozen for subsequent ureaplasma culture, and (ii) fixed, embedded, sectioned and stained by haematoxylin and eosin stain for histological analysis. Selected fetal lung clinical ureaplasma isolates were cloned and filtered to obtain cultures from a single CFU. Passage 1 and clone 2 ureaplasma cultures were tested by western blot to demonstrate MBA variation. In acute durations of ureaplasma infection no MBA variants (3d Up) or very few MBA variants (7d Up) were present when compared to the original inoculum. However, numerous MBA size variants were generated in vivo (alike within contiguous tissues, amniotic fluid and fetal lung, but different variants were present within chorioamnion), during chronic, 69d exposure to ureaplasma infection. For the first time we have shown that the degree of ureaplasma MBA variation in vivo increased with the duration of gestation.

  7. Survey on gastrointestinal parasites and detection of Cryptosporidium spp. on cattle in West Java, Indonesia.

    Science.gov (United States)

    Ananta, Sylvia Maharani; Suharno; Hidayat, Adi; Matsubayashi, Makoto

    2014-03-01

    To evaluate the presence of gastrointestinal parasites on cattle in Indonesia because the prevalence of parasites varies between countries depending on the terrain surrounding livestock farms and investigations in Indonesia have never been performed. Fecal samples from cattle at 35 farms in 7 districts in West Java, Indonesia, has been examined using the floatation or sedimentation methods, and a immunofluorescence assay and experimentally inoculation to mice for Cryptosporidium or Giardia.spp. 153 of 394 examined cattle (38.8%) were infected with gastrointestinal parasites. The prevalence of Eimeria spp., Nematoda spp. (including Oesophagustomum and Bunostomum-like), Fasciola gigantica and Paramphistomum spp. was 22.4%, 11.2%, 12.5% and 3.8%, respectively. Cryptosporidium andersoni (C. andersoni) was also found in two samples. One isolate of this parasite was confirmed to be transmitted to mice, in contrast to the isolates from other countries. although this survey is preliminary, the results shows that the infection of gastrointestinal parasites in Indonesia was not high, but these infected cattle could be as a potential source leading to economic losses in livestock production. Copyright © 2014 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

  8. Survey on gastrointestinal parasites and detection ofCryptosporidium spp. on cattle in West Java, Indonesia

    Institute of Scientific and Technical Information of China (English)

    Sylvia Maharani Ananta; Suharno; Adi Hidayat; Makoto Matsubayashi

    2014-01-01

    Objective:To evaluate the presence of gastrointestinal parasites on cattle inIndonesia because the prevalence of parasites varies between countries depending on the terrain surrounding livestock farms and investigations inIndonesia have never been performed.Methods:Fecal samples from cattle at35 farms in7 districts inWestJava,Indonesia, has been examined using the floatation or sedimentation methods, and a immunofluorescence assay and experimentally inoculation to mice forCryptosporidium or Giardia spp.Results:153 of394 examined cattle(38.8%) were infected with gastrointestinal parasites.The prevalence ofEimeria spp.,Nematoda spp. (includingOesophagustomum andBunostomum-like),Fasciola gigantica andParamphistomum spp. was22.4%,11.2%,12.5% and3.8%, respectively. Cryptosporidium andersoni(C. andersoni) was also found in two samples.One isolate of this parasite was confirmed to be transmitted to mice, in contrast to the isolates from other countries.Conclusions:although this survey is preliminary, the results shows that the infection of gastrointestinal parasites inIndonesia was not high, but these infected cattle could be as a potential source leading to economic losses in livestock production.

  9. Molecular evidence of Ureaplasma urealyticum and Ureaplasma parvum colonization in preterm infants during respiratory distress syndrome

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    Germani Rossella

    2006-11-01

    Full Text Available Abstract Background Ureaplasma urealyticum and U. parvum have been associated with respiratory diseases in premature newborns, but their role in the pathogenesis of the respiratory distress syndrome (RDS is unclear. The aim of this study was to detect, using molecular techniques, the role of Mycoplasma spp. and Ureaplasma spp. in respiratory secretion and blood specimens of preterm newborns with or without RDS and to evaluate the prevalence of perinatal U. urealyticum or U. parvum infection. The influence of chemotherapy on the clinical course was also evaluated. Methods Tracheal aspirate or nasopharingeal fluid samples from 50 preterm babies with (24 or without RDS (26 were analysed for detection of U. urealyticum and U. parvum by culture identification assay and PCR. Sequencing analysis of amplicons allowed us to verify the specificity of methods. Clarithromycin (10 mg kg-1 twice a day was administered in ureaplasma-positive patients who presented clinical signs of RDS. Results 15/24 neonates with RDS (p U. urealyticum or U. parvum. Culture identification assay was positive in 5/50 newborns, three of which with RDS. Sequencing analyses confirmed the specificity of these methods. Association of patent ductus arteriosus with ureaplasma colonization was more statistically significant (p = 0.0004 in patients with RDS than in those without RDS. Conclusion Colonization of the lower respiratory tract by Ureaplasma spp. and particularly by U. parvum in preterm newborns was related to RDS. The routine use of molecular methods could be useful to screen candidate babies for etiologic therapy.

  10. Emergence of Cryptosporidium hominis Monkey Genotype II and Novel Subtype Family Ik in the Squirrel Monkey (Saimiri sciureus) in China.

    Science.gov (United States)

    Liu, Xuehan; Xie, Na; Li, Wei; Zhou, Ziyao; Zhong, Zhijun; Shen, Liuhong; Cao, Suizhong; Yu, Xingming; Hu, Yanchuan; Chen, Weigang; Peng, Gangneng

    2015-01-01

    A single Cryptosporidium isolate from a squirrel monkey with no clinical symptoms was obtained from a zoo in Ya'an city, China, and was genotyped by PCR amplification and DNA sequencing of the small-subunit ribosomal RNA (SSU rRNA), 70-kDa heat shock protein (HSP70), Cryptosporidium oocyst wall protein, and actin genes. This multilocus genetic characterization determined that the isolate was Cryptosporidium hominis, but carried 2, 10, and 6 nucleotide differences in the SSU rRNA, HSP70, and actin loci, respectively, which is comparable to the variations at these loci between C. hominis and the previously reported monkey genotype (2, 3, and 3 nucleotide differences). Phylogenetic studies, based on neighbor-joining and maximum likelihood methods, showed that the isolate identified in the current study had a distinctly discordant taxonomic status, distinct from known C. hominis and also from the monkey genotype, with respect to the three loci. Restriction fragment length polymorphisms of the SSU rRNA gene obtained from this study were similar to those of known C. hominis but clearly differentiated from the monkey genotype. Further subtyping was performed by sequence analysis of the gene encoding the 60-kDa glycoprotein (gp60). Maximum homology of only 88.3% to C. hominis subtype IdA10G4 was observed for the current isolate, and phylogenetic analysis demonstrated that this particular isolate belonged to a novel C. hominis subtype family, IkA7G4. This study is the first to report C. hominis infection in the squirrel monkey and, based on the observed genetic characteristics, confirms a new C. hominis genotype, monkey genotype II. Thus, these results provide novel insights into genotypic variation in C. hominis.

  11. Between-year variation and spatial dynamics of Cryptosporidium spp. and Giardia spp. infections in naturally infected rodent populations.

    Science.gov (United States)

    Bajer, A

    2008-12-01

    Prevalence and abundance of Cryptosporidium spp. and Giardia spp. infections were studied over the 8-year period in 3 species of rodents in N.E. Poland (bank vole Myodes glareolus-1523; yellow-necked mouse Apodemus flavicollis- 638; common vole Microtus arvalis- 419). Prevalence was 53.8, 28.1 and 62.3% respectively for Cryptosporidium spp. and 58.3, 24.4 and 74.2% respectively for Giardia spp. Prevalence and abundance of infection varied markedly across 8 years of the study with 1998 and 2002 being years of higher prevalence and abundance, following changes in the densities of host species. The distribution of intestinal protozoa in forest rodents did not vary in the 3 isolated sites during the 4-year study. In the case of Cryptosporidium, fewer older animals carried infection and infections of the oldest bank and common voles were relatively milder. In the case of Giardia in yellow-necked mice, infections were more common in older age classes (2 and 3). The two species showed significant co-occurrence and in animals carrying both species there was a strong significant positive correlation between abundance of infection with each. These data are discussed in relation to the parasite genotypes identified in this region and in respect of the role of various ecological factors in shaping of intestinal protozoa communities.

  12. [Prevalence of Cyclospora sp., Cryptosporidium sp, microsporidia and fecal coliform determination in fresh fruit and vegetables consumed in Costa Rica].

    Science.gov (United States)

    Calvo, Melvin; Carazo, Melissa; Arias, Maria Laura; Chaves, Carolina; Monge, Rafael; Chinchilla, Misael

    2004-12-01

    The presence of Cyclospora sp., Cryptosporidium sp. and microsporidia and the levels of fecal coliforms were determined in lettuce, parsley, cilantro, strawberries and blackberries acquired in local agricultural markets of the Central Valley of Costa Rica, in order to establish the possible transmission risk of these microorganisms and other pathogens from the consumption of these raw products. During the second semester of 2001 and the first of 2002, 50 different samples of each product, 25 taken in the dry season and 25 in the rainy season and coming from five different local agricultural markets were evaluated. The fecal coliforms count was done according to the technique recommended by Vanderzant & Splittstoesser. The parasite determination was done using Zielh Nielsen and Weber staining techniques from a sediment obtained through the rinse of the mentioned products, using sterile peptonated water 0.1% and centrifuging at 900 G for 15 min. One hundred per cent of vegetable samples had fecal coliforms and the greatest prevalence was obtained during the rainy season. Although all vegetables presented fecal coliforms in high concentrations, lettuce and cilantro presented statistical difference between rainy and dry season, being greater during the rainy season. Fecal coliforms were not detected in strawberries and blackberries probablydue to its low pH. All products evaluated presented, at least once, Cryptosporidium sp., Cyclospora sp. and microsporidia, showing the risk they represent to Public Health. Cryptosporidium was present in all products but strawberries. Microsporidia was present in all products except blackberries and Cyclospora was only isolated from lettuce during the dry season. These results show the importance of introducing in the country Good Agricultural Practices, especially due to the resistance of Cryptosporidium and Cyclospora to disinfecting agents.

  13. Cryptosporidium and Giardia in Danish organic pig farms: Seasonal and age-related variation in prevalence, infection intensity and species/genotypes.

    Science.gov (United States)

    Petersen, Heidi H; Jianmin, Wang; Katakam, Kiran K; Mejer, Helena; Thamsborg, Stig M; Dalsgaard, Anders; Olsen, Annette; Enemark, Heidi L

    2015-11-30

    scrofarum excreted fewer oocysts (mean CPG: 54,848 ± 194,508CI: 9085-118,781) compared to pigs infected with Cryptosporidium suis (mean OPG: 351,035 ± 351,035CI: 67,953-634,117). No correlation between faecal consistency and (oo-)cyst excretion levels was observed. Of the successfully genotyped isolates, 38/56 (67.9%) were C. scrofarum and 18/56 (32.1%) were C. suis, while the livestock specific G. duodenalis Assemblage E was detected in 11/13 (84.6%) isolates and the potentially zoonotic Assemblage A was identified in 2/13 (15.4%) isolates. Piglets exclusively hosted C. suis, with one exception, while starter pigs and fatteners predominantly hosted C. scrofarum. As organic pigs are partly reared outdoors, environmental contamination with Cryptosporidium and Giardia is inevitable. Nevertheless, the present data indicate that the potential public health risk associated with both of these parasites in Danish organic pig production seems to be negligible.

  14. Improved Risk Analysis by Dual Direct Detection of Total and Infectious Cryptosporidium Oocysts on Cell Culture in Combination with Immunofluorescence Assay▿

    Science.gov (United States)

    Lalancette, Cindy; Di Giovanni, George D.; Prévost, Michèle

    2010-01-01

    The inactivation of Cryptosporidium oocysts is a main driver in the selection of water treatment disinfection strategies, and microbial risk analysis provides a sound basis for optimizing water treatment processes. U.S. Environmental Protection Agency method 1622/23 provides an estimate of the total oocyst count; however, it cannot be used directly for risk assessment, as it does not determine the fraction of infectious oocysts. Improved assessment of the risk for designated sources or in treated water requires evaluation of the total number of oocysts and an estimate of their infectivity. We developed a dual direct detection method using differential immunofluorescent staining that allows detection of both oocysts and cell culture infection foci for each sample. Using Cryptosporidium parvum oocysts, various pH levels, proteases, and gastroenteric compounds and substrates were assessed to determine their abilities to enhance the number of infection foci. The results showed that the key trigger for oocyst stimulation was acidification. Addition of a low concentration of d-glucose (50 mM) to the infection media increased rates of infectivity, while a higher dose (300 mM) was inhibitory. The total number of oocysts in each sample was determined by counting the oocysts remaining on a cell monolayer and the oocysts recovered from cell monolayer washes during processing using a simple filtration technique. With the dual direct detection on cell culture with immunofluorescence assay method, it is now possible to determine the numbers of total and infectious oocysts for a given sample in a single analysis. Direct percentages of infectivity are then calculated, which allows more accurate assessments of risk. PMID:19933339

  15. Prevalence and genetic characterization of Giardia and Cryptosporidium in cats from Italy.

    Science.gov (United States)

    Paoletti, Barbara; Otranto, Domenico; Weigl, Stefania; Giangaspero, Annunziata; Di Cesare, Angela; Traversa, Donato

    2011-12-01

    One hundred and eighty one cats living in central Italy were tested for the presence of Giardia and Cryptosporidium infection by IFAT test and specific PCRs. Overall eight (4.4%) samples were IFAT-positive for Giardia. All the IFAT-positive samples for Giardia scored positive for the PCRs, and three more samples IFAT-negative generated PCR products leading to a total 6.1% molecular positivity rate for Giardia. All the examined samples were negative for Cryptosporidium. Sequencing of samples molecularly positive to Giardia indicated that three cats harbored the zoonotic Giardia duodenalis Assemblage A, whereas all other positive animals were infected with the feline-specific G. duodenalis Assemblage F. Phylogenetic analysis carried out on the sequences obtained supported the clustering of the isolates within Assemblages A and F. The results here presented provide data on the occurrence of Giardia genotypes in cats living in close contact with humans highlighting the potential importance of this protozoan disease for the public health.

  16. Synergic activation of toll-like receptor (TLR 2/6 and 9 in response to Ureaplasma parvum & urealyticum in human amniotic epithelial cells.

    Directory of Open Access Journals (Sweden)

    Martha Triantafilou

    Full Text Available Ureaplasma species are the most frequently isolated microorganisms inside the amniotic cavity and have been associated with spontaneous abortion, chorioamnionitis, premature rupture of the membranes (PROM, preterm labour (PL pneumonia in neonates and bronchopulmonary dysplasia in neonates. The mechanisms by which Ureaplasmas cause such diseases remain unclear, but it is believed that inappropriate induction of inflammatory responses is involved, triggered by the innate immune system. As part of its mechanism of activation, the innate immune system employs germ-lined encoded receptors, called pattern recognition receptors (PRRs in order to "sense" pathogens. One such family of PRRs are the Toll like receptor family (TLR. In the current study we aimed to elucidate the role of TLRs in Ureaplasma-induced inflammation in human amniotic epithelial cells. Using silencing, as well as human embryonic kidney (HEK transfected cell lines, we demonstrate that TLR2, TLR6 and TLR9 are involved in the inflammatory responses against Ureaplasma parvum and urealyticum serovars. Ureaplasma lipoproteins, such as Multiple Banded antigen (MBA, trigger responses via TLR2/TLR6, whereas the whole bacterium is required for TLR9 activation. No major differences were observed between the different serovars. Cell activation by Ureaplasma parvum and urealyticum seem to require lipid raft function and formation of heterotypic receptor complexes comprising of TLR2 and TLR6 on the cell surface and TLR9 intracellularly.

  17. Cryptosporidium and Giardia in recreational water in Belgium.

    Science.gov (United States)

    Ehsan, Md Amimul; Casaert, Stijn; Levecke, Bruno; Van Rooy, Liesbet; Pelicaen, Joachim; Smis, Anne; De Backer, Joke; Vervaeke, Bart; De Smedt, Sandra; Schoonbaert, Filip; Lammens, Saskia; Warmoes, Thierry; Geurden, Thomas; Claerebout, Edwin

    2015-09-01

    The objective of this study was to investigate the presence of Cryptosporidium and Giardia in different recreational water bodies in Belgium and to estimate the infection risk associated with swimming and other recreational activities. Cryptosporidium oocysts and/or Giardia cysts were detected in three out of 37 swimming pools, seven out of 10 recreational lakes, two out of seven splash parks and four out of 16 water fountains. In the swimming pools no infection risk for Cryptosporidium could be calculated, since oocysts were only detected in filter backwash water. The risk of Giardia infection in the swimming pools varied from 1.13×10(-6) to 2.49×10(-6) per swim per person. In recreational lakes, the infection risk varied from 2.79×10(-5) to 5.74×10(-5) per swim per person for Cryptosporidium and from 7.04×10(-5) to 1.46×10(-4) for Giardia. For other outdoor water recreation activities the estimated infection risk was 5.71×10(-6) for Cryptosporidium and 1.47×10(-5) for Giardia. However, most positive samples in the recreational lakes belonged to species/genotypes that are either animal-specific or predominantly found in animals. No Cryptosporidium was found in splash parks and water fountains, but the presence of Giardia cysts suggests a risk for human infection. The infection risk of Giardia infection during a 3.5-minute visit to a splash park for children equalled 1.68×10(-4).

  18. Cryptosporidium cell culture infectivity assay design.

    Science.gov (United States)

    King, B J; Keegan, A R; Robinson, B S; Monis, P T

    2011-05-01

    Members of the genus Cryptosporidium, which cause the gastrointestinal disease cryptosporidiosis, still represent a significant cause of water-borne disease worldwide. While intensive efforts have been invested in the development of techniques for parasite culture, in vitro growth has been hampered by a number of factors including low levels of infectivity as well as delayed life-cycle development and poor synchronicity. In this study we examined factors affecting the timing of contact between excysted sporozoites and target host cells and the subsequent impact of this upon the establishment of infection. We demonstrate that excystation rate impacts upon establishment of infection and that in our standard assay format the majority of sporozoites are not close enough to the cell monolayer when they are released from the oocyst to successfully establish infection. However, this can be easily overcome by centrifugation of oocysts onto the cell monolayer, resulting in approximately 4-fold increases in sporozoite attachment and subsequent infection. We further demonstrate that excystation procedures can be tailored to control excystation rate to match the assay end purpose and that excystation rate can influence data interpretation. Finally, the addition of both a centrifugation and washing step post-sporozoite attachment may be appropriate when considering the design of in vitro culture experiments for developmental analysis and stage-specific gene expression as this appears to increase the synchronicity of early developmental stages.

  19. Metabolic and physiological changes in Prymnesium parvum when grown under, and grazing on prey of, variable nitrogen

    DEFF Research Database (Denmark)

    Lundgren, Veronica M; Glibert, Patricia M.; Graneli, Edna;

    2016-01-01

    the realization that mixotrophy is important ecologically. Laboratory experiments were conducted to examine changes in growth rates and physiological states of the toxic haptophyte Prymnesium parvum when fed Rhodomonas salina of varying nutritional status. Haemolytic activity of P. parvum and prey mortality of R....... Mortality rates were, however, directly related to the initial prey:predator cell ratios. On the other hand, growth of the predator was a function of nutritional status and a significant positive correlation was observed between growth rates of P. parvum and cell-specific depletion rates of N, whereas...

  20. First description of heterogeneity in 18S rRNA genes in the haploid genome of Cryptosporidium andersoni Kawatabi type.

    Science.gov (United States)

    Ikarashi, Makoto; Fukuda, Yasuhiro; Honma, Hajime; Kasai, Kenji; Kaneta, Yoshiyasu; Nakai, Yutaka

    2013-09-01

    The Apicomplexan Cryptosporidium andersoni, is a species of gastric Cryptosporidium, is frequently detected in older calves and adult cattle. Genotyping analyses based on 18S ribosomal RNA gene sequences have been performed on a novel C. andersoni genotype, namely the Kawatabi type, and the oocysts were classified into two distinct groups genotypically: Type A (the sequence in GenBank) and Type B (with a thymine nucleotide insertion not in Type A). This study analyzed 3775 cattle at a slaughterhouse and 310 cattle at a farm using microscopy and found 175 Cryptosporidium-positive animals: 171 from the slaughterhouse and four from the farm, and all infecting parasites were determined to be C. andersoni from 18S rRNA gene sequences determined from fecal DNA. In genotyping analyses with single isolated oocysts, about a half of analyzed ones were clearly classified into well known two genotypes (Type A and B). In addition to these two known genotypes, we have detected some oocysts showing mixed signals of Types A and B in the electropherogram from the automated sequencer (the Type C genotype). To determine the genotypic composition of sporozoites carried by the Type C oocysts, we analyzed their 18S rRNA gene sequences using a single sporozoite isolation procedure. Some sporozoites were classified as either Type A or Type B. However, more than half of the analyzed isolated sporozoites showed a mixed signal identical to that of Type C oocysts, and both the Type A and B signals were surely detectable from such sporozoites after a cloning procedure. In conclusion, C. andersoni carries two different genotypes heterogeneously in its haploid genome.

  1. First genotyping of Cryptosporidium spp. in pre-weaned calves, broiler chickens and children in Syria by PCR-RFLP analysis.

    Science.gov (United States)

    Kassouha, Morshed; Soukkarieh, Chadi; Alkhaled, Abdulkarim

    2016-07-30

    In this study, PCR-RFLP was used for the first time in Syria for genotyping Cryptosporidium species of man, calves and chickens. The total of 391 fecal samples included 213 from children with diarrhea (<5years), 67 from pre-weaned calves with diarrhea and 111 from broiler chicken farms. All samples were collected and examined with acid fast stain to detect the positive samples. Subsequently a nested-PCR test was performed on 35 positive samples (17 from calves, 11 from chicken, and 7 from children) targeting SSU rRNA gene, and was followed by RFLP analysis using three restriction enzymes SspI, VspI and MboII. Results showed that C. parvum was the only identified species in children and calves, on the other hand C. baileyi was identified in broilers in addition to another species with unknown RFLP profile in comparison to those which have been described in chicken. Further studies using more genes are needed to sequence and detect subtypes of this parasite.

  2. Antibody responses to the immunodominant Cryptosporidium gp15 antigen and gp15 polymorphisms in a case-control study of cryptosporidiosis in children in Bangladesh.

    Science.gov (United States)

    Allison, Genève M; Rogers, Kathleen A; Borad, Anoli; Ahmed, Sabeena; Karim, Mohammad Mahbubul; Kane, Anne V; Hibberd, Patricia L; Naumova, Elena N; Calderwood, Stephen B; Ryan, Edward T; Khan, Wasif A; Ward, Honorine D

    2011-07-01

    Although Cryptospridium hominis is the dominant Cryptosporidium species infecting humans, immune responses to cognate antigens in C. hominis-infected persons have not been reported. We investigated antibody responses to the immunodominant gp15 antigen from C. hominis and C. parvum, in C. hominis-infected Bangladeshi children less than five years of age with diarrhea (cases) and uninfected children with diarrhea (controls). We also investigated polymorphisms in the C. hominis gp15 sequence from cases. Serum IgG responses to gp15 from both species were significantly greater in cases than controls. In spite of polymorphisms in the gp15 sequence, there was a significant correlation between antibody levels to gp15 from both species, indicating cross-reactivity to conserved epitopes. Cases with acute diarrhea had a significantly greater serum IgA response to gp15 compared with those with persistent diarrhea, suggesting that this response may be associated with protection from prolonged disease. These findings support further investigation of gp15 as a vaccine candidate.

  3. ENHANCED DAPI STAINING FOR CRYPTOSPORIDIUM IN WATER SAMPLES

    Science.gov (United States)

    The U.S. Environmental Protection Agency's Method 1623 is used to detect and quantify the presence of {ital Cryptosporidium} spp. oocysts in water. The protocol consists of concentrating a sample, staining this concentrate with a fluorescent antibody, and examining the sample mi...

  4. Prevalence of Cryptosporidium in small ruminants from Veracruz, Mexico

    Science.gov (United States)

    Cryptosporidiosis is a zoonotic disease caused by the protozoan parasite Cryptosporidium spp. that can affect domestic animal and human populations. In newborn ruminants, cryptosporidiosis is characterized by outbreaks of diarrhea, which can result in high morbidity and economic impact. The aim of t...

  5. Cryptosporidium and Giardia in Africa: current and future challenges.

    Science.gov (United States)

    Squire, Sylvia Afriyie; Ryan, Una

    2017-04-20

    Cryptosporidium and Giardia are important causes of diarrhoeal illness. Adequate knowledge of the molecular diversity and geographical distribution of these parasites and the environmental and climatic variables that influence their prevalence is important for effective control of infection in at-risk populations, yet relatively little is known about the epidemiology of these parasites in Africa. Cryptosporidium is associated with moderate to severe diarrhoea and increased mortality in African countries and both parasites negatively affect child growth and development. Malnutrition and HIV status are also important contributors to the prevalence of Cryptosporidium and Giardia in African countries. Molecular typing of both parasites in humans, domestic animals and wildlife to date indicates a complex picture of both anthroponotic, zoonotic and spill-back transmission cycles that requires further investigation. For Cryptosporidium, the only available drug (nitazoxanide) is ineffective in HIV and malnourished individuals and therefore more effective drugs are a high priority. Several classes of drugs with good efficacy exist for Giardia, but dosing regimens are suboptimal and emerging resistance threatens clinical utility. Climate change and population growth are also predicted to increase both malnutrition and the prevalence of these parasites in water sources. Dedicated and co-ordinated commitments from African governments involving "One Health" initiatives with multidisciplinary teams of veterinarians, medical workers, relevant government authorities, and public health specialists working together are essential to control and prevent the burden of disease caused by these parasites.

  6. Molecular epidemiology of Cryptosporidium and Giardia in cattle

    Science.gov (United States)

    Cryptosporidium spp. and Giardia duodenalis are enteric protozoan parasites that infect a wide range of vertebrate hosts including humans. Infections with both parasites are known as one of the most common causes of diarrhea in humans and livestock. The epidemiology of cryptosporidiosis and giardias...

  7. Eliminatie van virussen, Cryptosporidium en Giardia door drinkwaterzuiveringsprocessen

    NARCIS (Netherlands)

    Medema GJ; Theunissen J; MGB

    1996-01-01

    A study on the removal efficiency of drinking water treatment processes for viruses and protozoa (Cryptosporidium/Giardia). The description is based on the best available Dutch and, if data on the Dutch situation are absent, international research data. The approach is valid for well-designed and

  8. Isolation

    DEFF Research Database (Denmark)

    Agerholm, Frank Juul

    2011-01-01

    Næringsstoffet har i dette nummer sat fokus på ”velvære i vinterkulden”, ”indendørsaktiviteter” og ”fedtafgift”. I klummen vises det, at disse tre fokusområder, der for en umiddelbar betragtning måske nok synes noget uensartede, falder sammen i ét tema: Isolation!......Næringsstoffet har i dette nummer sat fokus på ”velvære i vinterkulden”, ”indendørsaktiviteter” og ”fedtafgift”. I klummen vises det, at disse tre fokusområder, der for en umiddelbar betragtning måske nok synes noget uensartede, falder sammen i ét tema: Isolation!...

  9. Probabilistic quantitative microbial risk assessment model of farmer exposure to Cryptosporidium spp. in irrigation water within Kumasi Metropolis-Ghana

    DEFF Research Database (Denmark)

    Sampson, Angelina; Owusu-Ansah, Emmanuel de-Graft Johnson; Mills-Robertson, Felix C.

    2017-01-01

    Cryptosporidium is a protozoan parasite which can be transmitted via food and water. Some studies have shown irrigation water to be routes of transmission for Cryptosporidium into the food chain, however, little information is known about Cryptosporidium levels in wastewater used for irrigation...... causing gastroenteritis. The results indicate high positive levels of Cryptosporidium in the irrigation water, however, the levels of Cryptosporidium decreases during the rainfall seasons, risk assessment results show that, farmers face a higher risk of being infected by Cryptosporidium due to frequent...

  10. Distribution, hosts, 16S rDNA sequences and phylogenetic position of the Neotropical tick Amblyomma parvum (Acari: Ixodidae).

    Science.gov (United States)

    Nava, S; Szabó, M P J; Mangold, A J; Guglielmone, A A

    2008-07-01

    The hosts, distribution, intraspecific genetic variation and phylogenetic position of Amblyomma parvum (Acari: Ixodidae) have recently been re-assessed. Data on this tick's hosts and distribution were obtained not only from existing literature but also from unpublished records. Sequences of the ticks' mitochondrial 16S ribosomal DNA (rDNA) were used to evaluate genetic variation among specimens of A. parvum from different localities in Argentina and Brazil, and to explore the phylogenetic relationships between this tick and other Amblyomma species. Although several species of domestic and wild mammal act as hosts for adult A. parvum, most collected adults of this species have come from cattle and goats. Caviid rodents of the subfamily Caviinae appear to be the hosts for the immature stages. So far, A. parvum has been detected in 12 Neotropical biogeographical provinces (Chaco, Cerrado, Eastern Central America, Venezuelan Coast, Pantanal, Parana Forest, Caatinga, Chiapas, Venezuelan Llanos, Monte, Western Panamanian Isthmus, and Roraima) but the Chaco province has provided significantly more specimens than any other (P<0.0001). The 16S rDNA sequences showed just 0.0%-1.1% divergence among the Argentinean A. parvum investigated and no more than 0.2% divergence among the Brazilian specimens. The observed divergence between the Argentinean and Brazilian specimens was, however, greater (3.0%-3.7%). Although there is now molecular and morphological evidence to indicate that A. parvum, A. pseudoparvum, A. auricularium and A. pseudoconcolor are members of a natural group, previous subgeneric classifications do not reflect this grouping. The subgeneric status of these tick species therefore needs to be re-evaluated. The 16S-rDNA-based evaluation of divergence indicates that the gene flow between Argentinean and Brazilian 'A. parvum' is very limited and that the Argentinean 'A. parvum' may be a different species to the Brazilian.

  11. A chronicle of a killer alga in the west: ecology, assessment, and management of Prymnesium parvum blooms

    Science.gov (United States)

    Roelke, D.L.; Barkoh, Aaron; Brooks, B.W.; Grover, J.P.; Hambright, K.D.; LaClaire, John W.; Moeller, Peter D.R.; Patino, Reynaldo

    2015-01-01

    Since the mid-1980s, fish-killing blooms ofPrymnesium parvum spread throughout the USA. In the south central USA, P. parvum blooms have commonly spanned hundreds of kilometers. There is much evidence that physiological stress brought on by inorganic nutrient limitation enhances toxicity. Other factors influence toxin production as well, such as stress experienced at low salinity and temperature. A better understanding of toxin production by P. parvum remains elusive and the identities and functions of chemicals produced are unclear. This limits our understanding of factors that facilitated the spread of P. parvum blooms. In the south central USA, not only is there evidence that the spread of blooms was controlled, in part, by migration limitation. But there are also observations that suggest changed environmental conditions, primarily salinity, facilitated the spread of blooms. Other factors that might have played a role include altered hydrology and nutrient loading. Changes in water hardness, herbicide use, system pH, and the presence of toxin-resistant and/or P. parvum-inhibiting plankton may also have played a role. Management of P. parvum in natural systems has yet to be attempted, but may be guided by successes achieved in small impoundments and mesocosm experiments that employed various chemical and hydraulic control approaches.

  12. MLST subtypes and population genetic structure of Cryptosporidium andersoni from dairy cattle and beef cattle in northeastern China's Heilongjiang Province.

    Directory of Open Access Journals (Sweden)

    Wei Zhao

    Full Text Available Cattle are the main reservoir host of C. andersoni, which shows a predominance in yearlings and adults of cattle. To understand the subtypes of C. andersoni and the population genetic structure in Heilongjiang Province, fecal specimens were collected from 420 dairy cattle and 405 beef cattle at the age of 12-14 months in eight cattle farms in five areas within this province and were screened for the presence of Cryptosporidium oocysts by microscopy after Sheather's sugar flotation technique. The average prevalence of Cryptosporidium spp. was 19.15% (158/825 and all the Cryptosporidium isolates were identified as C. andersoni by the SSU rRNA gene nested PCR-RFLP using SspI, VspI and MboII restriction enzymes. A total of 50 C. andersoni isolates were randomly selected and sequenced to confirm the RFLP results before they were subtyped by multilocus sequence typing (MLST at the four microsatellite/minisatellite loci (MS1, MS2, MS3 and MS16. Four, one, two and one haplotypes were obtained at the four loci, respectively. The MLST subtype A4,A4,A4,A1 showed an absolute predominance and a wide distribution among the six MLST subtypes obtained in the investigated areas. Linkage disequilibrium analysis showed the presence of a clonal population genetic structure of C. andersoni in cattle, suggesting the absence of recombination among lineages. The finding of a clonal population genetic structure indicated that the prevalence of C. andersoni in cattle in Heilongjiang Province is not attributed to the introduction of cattle. Thus, prevention and control strategies should be focused on making stricter measures to avoid the occurrence of cross-transmission and re-infection between cattle individuals. These molecular data will also be helpful to explore the source attribution of infection/contamination of C. andersoni and to elucidate its transmission dynamics in Heilongjiang Province, even in China.

  13. Prevalence of Cryptosporidium-like infection in one-humped camels (Camelus dromedarius of northwestern Iran

    Directory of Open Access Journals (Sweden)

    Yakhchali M.

    2012-02-01

    Full Text Available Cryptosporidium is a ubiquitous enteropathogen protozoan infection affecting livestock worldwide. The present study was carried out to determine the prevalence of Cryptosporidium infection in different age groups of dromedary camels in northwestern Iran from November 2009 to July 2010. A total number of 170 fecal samples were collected and examined using modified Ziehl-Neelsen (MZN staining under light microscope. Examination of stained fecal smears revealed that 17 camels (10% were positive for Cryptosporidium-like. The prevalence of Cryptosporidium-like was significantly higher in camel calves (< 1 years old (20% than other age groups, in which the diarrhoeic calves had the prevalence of 16%. In adult camels the prevalence was 6.5%. There was no significant difference in the prevalence of Cryptosporidium-like between male and female camels. It is concluded that Cryptosporidium infection is a problem in camel husbandry and could be of public health concern in the region.

  14. Prymnesium parvum revisited: Relationship between allelopathy, ichthyotoxicity, and chemical profiles in 5 strains

    Energy Technology Data Exchange (ETDEWEB)

    Blossom, Hannah E., E-mail: hblossom@bio.ku.dk [Marine Biological Section, University of Copenhagen, Strandpromenaden 5, 3000 Helsingør (Denmark); Rasmussen, Silas A. [Department of Systems Biology, Technical University of Denmark, Søltofts Plads, Building 221, 2800 Kgs Lyngby (Denmark); Andersen, Nikolaj G. [Marine Biological Section, University of Copenhagen, Strandpromenaden 5, 3000 Helsingør (Denmark); Larsen, Thomas O.; Nielsen, Kristian F. [Department of Systems Biology, Technical University of Denmark, Søltofts Plads, Building 221, 2800 Kgs Lyngby (Denmark); Hansen, Per J. [Marine Biological Section, University of Copenhagen, Strandpromenaden 5, 3000 Helsingør (Denmark)

    2014-12-15

    Highlights: • Five strains of P. parvum were tested for toxicity towards rainbow trout and microalgae. • Toxicity towards microalgae was not correlated to toxicity towards fish. • A microalgal bioassay cannot be used as a reliable proxy for ichthyotoxicity. • Concentrations of GATs were low and not correlated to effects on fish or on algae. • P. parvum does not produce oleamide based on {sup 13}C labeling and extraction in glass. - Abstract: Bioassay-guided discovery of ichthyotoxic algal compounds using in vivo fish assays is labor intensive, costly, and highly regulated. Since the mode of action of most known algal-mediated fish-killing toxins is damage to the cell membranes in the gills, various types of cell-based bioassays are often used for bioassay guided purification of new ichthyotoxins. Here we tested the hypothesis that allelopathy is related to ichthyotoxicity and thus that a microalgal bioassay can be used as a proxy for ichthyotoxicity by comparing the toxicity of five strains of Prymnesium parvum toward rainbow trout (Oncorhynchus mykiss, 10 g) and the microalga Teleaulax acuta. No relationship between median effective concentrations (EC{sub 50}s) on fish and median lethal concentrations (LC{sub 50}s) on algae was observed in the 5 strains showing that a microalgal bioassay cannot be used as a proxy for ichthyotoxicity. Fish were more sensitive to P. parvum with EC{sub 50}s ranging from 6 × 10{sup 3} to 40 × 10{sup 3} cells ml{sup −1}, compared to the test alga where LC{sub 50}s ranged from 30 × 10{sup 3} to nearly non-toxic at 500 × 10{sup 3} cells ml{sup −1}. In addition, the cellular concentrations of two recently suggested ichthyotoxins produced by P. parvum, the “golden algae toxins”, GAT 512 and a novel GAT 510, did not show any relationship to either ichthyotoxicity or allelopathy, and are not the biologically relevant toxins, but are simply lipids found in algal chloroplasts. Finally, we demonstrate that the recently

  15. Molecular forensic profiling of Cryptosporidium species and genotypes in raw water.

    Science.gov (United States)

    Ruecker, Norma J; Bounsombath, Niravanh; Wallis, Peter; Ong, Corinne S L; Isaac-Renton, Judith L; Neumann, Norman F

    2005-12-01

    The emerging concept of host specificity of Cryptosporidium spp. was exploited to characterize sources of fecal contamination in a watershed. A method of molecular forensic profiling of Cryptosporidium oocysts on microscope slides prepared from raw water samples processed by U.S. Environmental Protection Agency Method 1623 was developed. The method was based on a repetitive nested PCR-restriction fragment length polymorphism-DNA sequencing approach that permitted the resolution of multiple species/genotypes of Cryptosporidium in a single water sample.

  16. Detection and molecular characterisation of Giardia duodenalis, Cryptosporidium spp. and Entamoeba spp. among patients with gastrointestinal symptoms in Gambo Hospital, Oromia Region, southern Ethiopia.

    Science.gov (United States)

    Flecha, María J; Benavides, Cynthia M; Tissiano, Gabriel; Tesfamariam, Abraham; Cuadros, Juan; de Lucio, Aida; Bailo, Begoña; Cano, Lourdes; Fuentes, Isabel; Carmena, David

    2015-09-01

    To assess the prevalence and genetic diversity of the enteric protozoa species G. duodenalis, Cryptosporidium spp. and Entamoeba histolytica in individuals with gastrointestinal symptoms compatible with infections by these pathogens seeking medical attention in a rural area in southern Ethiopia. A total of 92 stool samples were initially screened by direct microscopy and immunochromatography and further confirmed by molecular methods. G. duodenalis-positive samples were molecularly characterised by multilocus genotyping of the glutamate dehydrogenase and β-giardin genes of the parasite. PCR and DNA sequence analysis of the gene encoding the 60-kDa glycoprotein was used for the subtyping of Cryptosporidium isolates. Detection and differential diagnosis of E. histolytica/dispar were conducted by real-time PCR. PCR-based prevalences were 10.9% for G. duodenalis, 1.1% for Cryptosporidium spp. and 3.3% for Entamoeba spp. Seven (four novel and three known) subtypes of G. duodenalis assemblage B were identified at the GDH locus and 5 (one novel and four known) at the BG locus. A novel variant of C. hominis subtype IbA9G3 was also identified. Two Entamoeba isolates were assigned to E. dispar and an additional one to E. histolytica. Although preliminary, our results strongly suggest that giardiasis, cryptosporidiosis and amoebiasis represent a significant burden in Ethiopian rural population. © 2015 John Wiley & Sons Ltd.

  17. The severity of chorioamnionitis in pregnant sheep is associated with in vivo variation of the surface-exposed multiple-banded antigen/gene of Ureaplasma parvum.

    Science.gov (United States)

    Knox, Christine L; Dando, Samantha J; Nitsos, Ilias; Kallapur, Suhas G; Jobe, Alan H; Payton, Diane; Moss, Timothy J M; Newnham, John P

    2010-09-01

    Ureaplasma species are the bacteria most frequently isolated from human amniotic fluid in asymptomatic pregnancies and placental infections. Ureaplasma parvum serovars 3 and 6 are the most prevalent serovars isolated from men and women. We hypothesized that the effects on the fetus and chorioamnion of chronic ureaplasma infection in amniotic fluid are dependent on the serovar, dose, and variation of the ureaplasma multiple-banded antigen (MBA) and mba gene. We injected high- or low-dose U. parvum serovar 3, serovar 6, or vehicle intra-amniotically into pregnant ewes at 55 days of gestation (term = 150 days) and examined the chorioamnion, amniotic fluid, and fetal lung tissue of animals delivered by cesarean section at 125 days of gestation. Variation of the multiple banded antigen/mba generated by serovar 3 and serovar 6 ureaplasmas in vivo were compared by PCR assay and Western blot. Ureaplasma inoculums demonstrated only one (serovar 3) or two (serovar 6) MBA variants in vitro, but numerous antigenic variants were generated in vivo: serovar 6 passage 1 amniotic fluid cultures contained more MBA size variants than serovar 3 (P = 0.005), and ureaplasma titers were inversely related to the number of variants (P = 0.025). The severity of chorioamnionitis varied between animals. Low numbers of mba size variants (five or fewer) within amniotic fluid were associated with severe inflammation, whereas the chorioamnion from animals with nine or more mba variants showed little or no inflammation. These differences in chorioamnion inflammation may explain why not all women with in utero Ureaplasma spp. experience adverse pregnancy outcomes.

  18. The Severity of Chorioamnionitis in Pregnant Sheep Is Associated with In Vivo Variation of the Surface-Exposed Multiple-Banded Antigen/Gene of Ureaplasma parvum1

    Science.gov (United States)

    Knox, Christine L.; Dando, Samantha J.; Nitsos, Ilias; Kallapur, Suhas G.; Jobe, Alan H.; Payton, Diane; Moss, Timothy J.M.; Newnham, John P.

    2010-01-01

    Ureaplasma species are the bacteria most frequently isolated from human amniotic fluid in asymptomatic pregnancies and placental infections. Ureaplasma parvum serovars 3 and 6 are the most prevalent serovars isolated from men and women. We hypothesized that the effects on the fetus and chorioamnion of chronic ureaplasma infection in amniotic fluid are dependent on the serovar, dose, and variation of the ureaplasma multiple-banded antigen (MBA) and mba gene. We injected high- or low-dose U. parvum serovar 3, serovar 6, or vehicle intra-amniotically into pregnant ewes at 55 days of gestation (term = 150 days) and examined the chorioamnion, amniotic fluid, and fetal lung tissue of animals delivered by cesarean section at 125 days of gestation. Variation of the multiple banded antigen/mba generated by serovar 3 and serovar 6 ureaplasmas in vivo were compared by PCR assay and Western blot. Ureaplasma inoculums demonstrated only one (serovar 3) or two (serovar 6) MBA variants in vitro, but numerous antigenic variants were generated in vivo: serovar 6 passage 1 amniotic fluid cultures contained more MBA size variants than serovar 3 (P = 0.005), and ureaplasma titers were inversely related to the number of variants (P = 0.025). The severity of chorioamnionitis varied between animals. Low numbers of mba size variants (five or fewer) within amniotic fluid were associated with severe inflammation, whereas the chorioamnion from animals with nine or more mba variants showed little or no inflammation. These differences in chorioamnion inflammation may explain why not all women with in utero Ureaplasma spp. experience adverse pregnancy outcomes. PMID:20519696

  19. In vitro culture of Cryptosporidium parvum in HCT-8%微小隐孢子虫在HCT-8细胞内的培养

    Institute of Scientific and Technical Information of China (English)

    朱惠丽; 刘利敏; 王臣荣; 张素梅; 张龙现; 王荣军

    2016-01-01

    目的 将人回盲肠癌(HCT-8)细胞作为微小隐孢子虫Ⅱd亚型体外感染对象,观察虫体在细胞中的生长发育过程.方法 将纯化的隐孢子虫Ⅱd亚型感染HCT-8细胞,通过Giemsa染色法观察微小隐孢子虫Ⅱd亚型在HCT-8细胞中发育过程,运用PCR方法对不同感染时间点的细胞样品DNA进行检测.结果 在感染后72 h内,隐孢子虫出现连续发育阶段,包括脱囊、子孢子、滋养体、裂殖体、配子体、合子和卵囊;PCR的检测结果显示,在感染细胞96 h仍能检测到虫体DNA.结论 通过HCT-8细胞,观察到微小隐孢子虫Ⅱd亚型的完整的生长发育过程,为抗隐孢子虫药物的筛选提供理论基础,亦可成为微小隐孢子虫Ⅱd亚型卵囊体外扩增的方法.

  20. CLONING AND MOLECULAR CHARACTERIZATION OF A GENE ENCODING A CRYPTOSPORIDIUM PARVUM PUTATIVE 20S PROTEASOME B1-TYPE SUBUNIT. (R825148)

    Science.gov (United States)

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  1. Sublethal concentrations of ichthyotoxic alga Prymnesium parvum affect rainbow trout susceptibility to viral haemorrhagic septicaemia virus

    DEFF Research Database (Denmark)

    Andersen, Nikolaj Gedsted; Lorenzen, Ellen; Boutrup, Torsten Snogdal;

    2016-01-01

    concentrations of the ichthyotoxic alga Prymnesium parvum affect the susceptibility of rainbow trout Oncorhynchus mykiss to viral haemorrhagic septicaemia virus (VHSV). During exposure to sublethal algal concentrations, the fish increased production of mucus on their gills. When fish were exposed to the algae...... for 12 h prior to the addition of virus, a marginal decrease in the susceptibility to VHSV was observed compared to fish exposed to VHSV without algae. If virus and algae were added simultaneously, inclusion of the algae increased mortality by 50% compared to fish exposed to virus only, depending...... on the experimental setup. We concluded that depending on the local exposure conditions, sublethal concentrations of P. parvum could affect susceptibility of fish to infectious agents such as VHSV....

  2. Candidatus Rickettsia andeanae, a spotted fever group agent infecting Amblyomma parvum ticks in two Brazilian biomes.

    Science.gov (United States)

    Nieri-Bastos, Fernanda Aparecida; Lopes, Marcos Gomes; Cançado, Paulo Henrique Duarte; Rossa, Giselle Ayres Razera; Faccini, João Luiz Horácio; Gennari, Solange Maria; Labruna, Marcelo Bahia

    2014-04-01

    Adult ticks of the species Amblyomma parvum were collected from the vegetation in the Pantanal biome (state of Mato Grosso do Sul) and from horses in the Cerrado biome (state of Piauí) in Brazil. The ticks were individually tested for rickettsial infection via polymerase chain reaction (PCR) targeting three rickettsial genes, gltA, ompA and ompB. Overall, 63.5% (40/63) and 66.7% (2/3) of A. parvum ticks from Pantanal and Cerrado, respectively, contained rickettsial DNA, which were all confirmed by DNA sequencing to be 100% identical to the corresponding fragments of the gltA, ompA and ompB genes of Candidatus Rickettsia andeanae. This report is the first to describe Ca. R. andeanae in Brazil.

  3. Prymnesium parvum revisited: relationship between allelopathy, ichthyotoxicity, and chemical profiles in 5 strains

    DEFF Research Database (Denmark)

    Blossom, Hannah E.; Rasmussen, Silas Anselm; Andersen, Nikolaj Gedsted

    2014-01-01

    used forbioassay guided purification of new ichthyotoxins. Here we tested the hypothesis that allelopathy isrelated to ichthyotoxicity and thus that a microalgal bioassay can be used as a proxy for ichthyotoxicityby comparing the toxicity of five strains of Prymnesium parvum toward rainbow trout...... (Oncorhynchusmykiss, 10 g) and the microalga Teleaulax acuta. No relationship between median effective concentrations(EC50s) on fish and median lethal concentrations (LC50s) on algae was observed in the 5 strains showingthat a microalgal bioassay cannot be used as a proxy for ichthyotoxicity. Fish were more sensitive...... of the five P. parvum strains above the limit of detection, nor was it found in a13C-labeled strain. Instead we document thatoleamide can easily be extracted from plastic materials, which may have been the source of oleamidereported previously....

  4. Candidatus Rickettsia andeanae, a spotted fever group agent infecting Amblyomma parvum ticks in two Brazilian biomes

    Directory of Open Access Journals (Sweden)

    Fernanda Aparecida Nieri-Bastos

    2014-04-01

    Full Text Available Adult ticks of the species Amblyomma parvum were collected from the vegetation in the Pantanal biome (state of Mato Grosso do Sul and from horses in the Cerrado biome (state of Piauí in Brazil. The ticks were individually tested for rickettsial infection via polymerase chain reaction (PCR targeting three rickettsial genes, gltA, ompA and ompB. Overall, 63.5% (40/63 and 66.7% (2/3 of A. parvum ticks from Pantanal and Cerrado, respectively, contained rickettsial DNA, which were all confirmed by DNA sequencing to be 100% identical to the corresponding fragments of the gltA, ompA and ompB genes of Candidatus Rickettsia andeanae. This report is the first to describe Ca. R. andeanae in Brazil.

  5. IDENTIFICATION OF CRYPTOSPORIDIUM SPECIES AND SOURCES IN RAW WASTEWATER USING A SMALL SUBUNIT RRNA-BASED PCR-RFLP TOOL

    Science.gov (United States)

    The species composition and source of Cryptosporidium oocysts in wastewater have never been determined, even though it is widely assumed that these oocysts are from human sewage. Recent molecular characterizations of Cryptosporidium parasites make it possible to differentiate hum...

  6. Prevalence of Giardia and Cryptosporidium species in dog park attending dogs compared to non-dog park attending dogs in one region of Colorado.

    Science.gov (United States)

    Wang, Andrea; Ruch-Gallie, Rebecca; Scorza, Valeria; Lin, Philip; Lappin, Michael R

    2012-03-23

    Dog parks are very popular in urban areas, but there are no current studies attempting to correlate visits to dog parks and risk of colonization by enteric parasites. The purpose of this study was to determine whether dog park visitation is associated with an increased prevalence of enteric parasites or an increase in prevalence of gastrointestinal signs in dogs in northern Colorado. Feces from dogs owned by veterinary students or Veterinary Teaching Hospital staff members were submitted with a completed survey form detailing dog park attendance rates, fecal character scores, and other clinical information. Feces were examined microscopically for parasites after sugar centrifugation, for Giardia spp. cysts and Cryptosporidium spp. oocysts by a commercially available immunofluorescence assay (FA) and the FA positive samples were genotyped after PCR amplification. The Giardia assemblages were determined using the glutamate dehydrogenase (GDH) β-giardin and triose phosphate isomerase (TPI) genes and the Cryptosporidium species were determined using the heat shock protein-70 gene. A total of 129 fecal samples were assayed; 66 were from dog park attending dogs and 63 were from non-dog park-attending dogs. The overall parasite prevalence rate was 7.0% (9 of 129 samples). Dog park attending dogs were more likely to be positive for Giardia or Cryptosporidium than non-dog park-attending dogs (p=0.0279), but there was no association of gastrointestinal signs with dog park attendance or with fecal flotation or FA results. The five Giardia isolates were assemblage C and/or D and the one Cryptosporidium isolate was Ctenocephalides canis.

  7. Amblyomma parvum Aragão, 1908 (Acari: Ixodidae): Phylogeography and systematic considerations.

    Science.gov (United States)

    Lado, Paula; Nava, Santiago; Labruna, Marcelo B; Szabo, Matias P J; Durden, Lance A; Bermudez, Sergio; Montagna, Matteo; Sánchez Quirós, Ana C; Beati, Lorenza

    2016-07-01

    The geographical distribution of Amblyomma parvum Aragão 1908 in the New World is disjunct, with two main clusters separated from each other by the Amazon basin. The main objectives of this study were to further investigate the systematic relationships within A. parvum, to determine whether or not populations from different geographical areas might represent cryptic species, and to reconstruct the phylogeographical evolutionary history of the species. The genetic diversity of A. parvum collected throughout its distributional range was analyzed by using 6 molecular markers: 5 mitochondrial [the small and the large ribosomal subunits 12rDNA and 16SrDNA, the cytochrome oxidase I (COI) and II (COII) and the control region or d-loop (DL)], and one nuclear (ITS2, Inter transcribed spacer 2). Phylogenetic trees were inferred by using maximum parsimony and Bayesian analyses. In addition, node dating was attempted for the main lineages identified phylogenetically. Although mitochondrial and nuclear topologies were not totally congruent, they all identified at least two main supported clusters, a Central American lineage, and a Brazilian-Argentinian lineage. Clade support and divergence values strongly suggest that the two lineages correspond to different taxonomic entities. Node dating placed the split between the Central American and the Brazilian-Argentinian lineages at approximately 5.8-4.9 Mya, just after the progressive replacement of the dry areas that occupied the northern part of South America by the Amazon Basin in the early-mid Miocene. This event might be the cause of fragmentation and putative speciation within the ancestral relatively xerophilic A. parvum population.

  8. Prymnesium parvum revisited: relationship between allelopathy, ichthyotoxicity, and chemical profiles in 5 strains.

    Science.gov (United States)

    Blossom, Hannah E; Rasmussen, Silas A; Andersen, Nikolaj G; Larsen, Thomas O; Nielsen, Kristian F; Hansen, Per J

    2014-12-01

    Bioassay-guided discovery of ichthyotoxic algal compounds using in vivo fish assays is labor intensive, costly, and highly regulated. Since the mode of action of most known algal-mediated fish-killing toxins is damage to the cell membranes in the gills, various types of cell-based bioassays are often used for bioassay guided purification of new ichthyotoxins. Here we tested the hypothesis that allelopathy is related to ichthyotoxicity and thus that a microalgal bioassay can be used as a proxy for ichthyotoxicity by comparing the toxicity of five strains of Prymnesium parvum toward rainbow trout (Oncorhynchus mykiss, 10 g) and the microalga Teleaulax acuta. No relationship between median effective concentrations (EC50s) on fish and median lethal concentrations (LC50s) on algae was observed in the 5 strains showing that a microalgal bioassay cannot be used as a proxy for ichthyotoxicity. Fish were more sensitive to P. parvum with EC50s ranging from 6×10(3) to 40×10(3) cells ml(-1), compared to the test alga where LC50s ranged from 30×10(3) to nearly non-toxic at 500×10(3) cells ml(-1). In addition, the cellular concentrations of two recently suggested ichthyotoxins produced by P. parvum, the "golden algae toxins", GAT 512 and a novel GAT 510, did not show any relationship to either ichthyotoxicity or allelopathy, and are not the biologically relevant toxins, but are simply lipids found in algal chloroplasts. Finally, we demonstrate that the recently suggested ichthyotoxin, oleamide, could not be detected in any of the five P. parvum strains above the limit of detection, nor was it found in a (13)C-labeled strain. Instead we document that oleamide can easily be extracted from plastic materials, which may have been the source of oleamide reported previously. Copyright © 2014 Elsevier B.V. All rights reserved.

  9. [Occurrence of Cryptosporidium spp. infection in antillean manatee (Trichechus manatus)].

    Science.gov (United States)

    Borges, João Carlos Gomes; Alves, Leucio Câmara; Vergara-Parente, Jociery Einhardt; Faustino, Maria Aparecida da Glória; Machado, Erilane de Castro Lima

    2009-01-01

    Cryptosporidiosis is a zoonosis which can affect man and a wide range of domestic and wild animals, mainly immunodeficient individuals. The objective of this paper was reported the occurrence of a Cryptosporidium infection in Antillean manatee. After an unusual behavior of an Antillean manatee kept in captivity at the Centro Mamíferos Aquáticos, ICMBio--FMA, clinical examination and posterior fecal sampling was performed. Fecal samples were examined by the Kinyoun technique, Direct Immunofluorescence Test and also examined by 4',6'-Diamidino-2-Phenylindole (DAPI) staining. At the clinical examination, the animal showed signs of abdominal pain. The results obtained by light and fluorescence microscopy analysis showed the presence of Cryptosporidium spp. oocyst in feces of this manatee.

  10. [Cryptosporidium spp. and Giardia spp.--environmental studies in Poland].

    Science.gov (United States)

    Bajer, Anna; Bednarska, Małgorzata; Siński, Edward

    2009-01-01

    Cryptosporidium spp. and Giardia spp. are intestinal protozoan parasites of humans and many other species of mammals. The aim of this article was to summarize the last twenty years of research on the environmental distribution of these parasites, with a particular emphasis on the natural reservoir of invasion and human infections in Poland. The prevalence of Cryptosporidium and Giardia has been studied in different groups of humans, in wildlife, pets and farm animals and in environmental samples. Current knowledge on the distribution of zoonotic and non-zoonotic species/genotypes in reservoir hosts and environmental samples has been summarized. The usefulness of different methods for the detection and identification of the parasites in different types of samples has been presented. Due to the wide distribution and high prevalence of both species in a range of hosts and possible vectors involved in mechanical transmission, the overall risk of outbreaks of cryptosporidiosis and giardiosis in Poland has been assessed as relatively high.

  11. La confusa taxonomía de Cryptosporidium

    Directory of Open Access Journals (Sweden)

    Gregorio Pérez-Cordón

    2006-10-01

    Full Text Available Los últimos descubrimientos en la biología y filogenética de Cryptosporidium refuerzan la necesidad de una exhaustiva revisión del ciclo de vida y la taxonomía de este parásito. Tanto futuros estudios de cultivo in vitro e in vivo así como estudios moleculares y genéticos permitirán avanzar en el profundo conocimiento de este interesante parásito.

  12. Effective removal of Cryptosporidium by a novel bioflocculant.

    Science.gov (United States)

    Ghosh, Moushumi; Pathak, Santosh; Ganguli, Abhijit

    2009-02-01

    Extracellular biopolymer produced from Klebsiella terrigena was found to have excellent flocculating ability over a wide range of colloid particles (0.5 to 25micro). The biopolymer was thermostable, with an optimum temperature for flocculation of 30 degrees C. Analysis with Fourier transform infrared spectrophotometry (FT-IR) shows that the biopolymer mainly possesses hydroxyl, carboxyl, and methoxyl groups, with neutral sugar and uronic acid as its major and minor components, and the structure of a polysaccharide. The average molecular weight of the biopolymer was greater than 2 x 10(3) kilodalton (KDa), as determined by gel permeation chromatography. Scanning electron microscopy indicated a porous morphology of the biopolymer. At a dosage of 2 mg/L, the purified biopolymer could remove 62.3% of Cryptosporidium oocysts (1 x 10(6)) spiked in tap water samples. Calcium (5mM) was required for effective removal. The removal efficiency of Cryptosporidium oocysts by the biopolymer remained unaltered over a pH range of 6 to 8. The results of this study indicates a possible utility of the Klebsiella terrigena biopolymer as an alternative to typically used chemical flocculants for removal of Cryptosporidium oocysts from water.

  13. Prevalence and molecular characterization of Cryptosporidium in giant panda (Ailuropoda melanoleuca) in Sichuan province, China.

    Science.gov (United States)

    Wang, Tao; Chen, Zuqin; Xie, Yue; Hou, Rong; Wu, Qidun; Gu, Xiaobing; Lai, Weiming; Peng, Xuerong; Yang, Guangyou

    2015-06-25

    Cryptosporidium spp. have been extensively reported to cause significant diarrheal disease in humans and domestic animals. On the contrary, little information is available on the prevalence and characterization of Cryptosporidium in wild animals in China, especially in giant pandas. The aim of the present study was to detect Cryptosporidium infections and identify Cryptosporidium species at the molecular level in both captive and wild giant pandas in Sichuan province, China. Using a PCR approach, we amplified and sequenced the 18S rRNA gene from 322 giant pandas fecal samples (122 from 122 captive individuals and 200 collected from four habitats) in Sichuan province, China. The Cryptosporidium species/genotypes were identified via a BLAST comparison against published Cryptosporidium sequences available in GenBank followed by phylogenetic analysis. The results revealed that both captive and wild giant pandas were infected with a single Cryptosporidium species, C. andersoni, at a prevalence of 15.6% (19/122) and 0.5% (1/200) in captive and wild giant pandas, respectively. The present study revealed the existence of C. andersoni in both captive and wild giant panda fecal samples for the first time, and also provided useful fundamental data for further research on the molecular epidemiology and control of Cryptosporidium infection in giant pandas.

  14. The first large scale molecular study of Cryptosporidium and Giardia in horses

    Science.gov (United States)

    The prevalence of species and genotypes of Giardia and Cryptosporidium in horses is poorly known. The present study examined feces from 195 horses, 1 month to 17 years of age, in 4 locations in Colombia. Prevalence and age distribution of Giardia and Cryptosporidium were determined by PCR. All PCR p...

  15. INVESTIGATION AND EVALUATION OF CRYPTOSPORIDIUM AND GIARDIA IN COMBINED SEWER OVERFLOW (CSO) AND STORMWATER RUNOFF

    Science.gov (United States)

    Since the first identified Cryptosporidium outbreaks occurred in the 1980s and the massive 1993 Milwaukee, WI outbreak affected more than 400,000 people, the concern over the public health risks related to protozoan pathogens Cryptosporidium and Giardia has g...

  16. CRYPTOSPORIDIUM AND GIARDIA IN STORMWATER AS A THREAT TO DRINKING WATER SUPPLIES

    Science.gov (United States)

    Since the first identified Cryptosporidium outbreak in the United Kingdom in 1983, the pathogens Cryptosporidium and Giardia have become the subject of growing local, state, and national concern. In the last decade, these organisms have been the causative agent of many gastroint...

  17. Prevalence rates of Giardia and Cryptosporidium among diarrheic patients in the Philippines.

    Science.gov (United States)

    Natividad, Filipinas F; Buerano, Corazon C; Lago, Catherine B; Mapua, Cynthia A; de Guzman, Blanquita B; Seraspe, Ebonia B; Samentar, Lorena P; Endo, Takuro

    2008-11-01

    The prevalence of Giardia and Cryptosporidium among 3,456 diarrheic patients corrected from May 2004 to May 2005 in the Philippines was determined. Of 133 (3.8%) positive samples, 69 (2.0%) were positive for Giardia and 67 (1.9%) for Cryptosporidium. Three samples had co-infection with Giardia and Cryptosporidium. Luzon had the highest positive samples (5.0%) followed by Mindanao (4.9%), then Visayas (2.2%). Giardia was most prevalent in Mindanao (3.6%) while Cryptosporidium was most prevalent in Luzon (3.1%). The prevalence of Giardia (2.0%) among pediatric patients (0-18 years) did not significantly differ from that (1.9%) among adults (> 18 years old). However, for Cryptosporidium, the prevalence (2.9%) among pediatric patients was significantly higher compared to that (0.2%) among adult patients. In the pediatric population, the highest percentage of patients with Giardia was the 5-9 year old age group, while that of Cryptosporidium was in the 0-4 year old group. The prevalence of Giardia, but not Cryptosporidium, was significantly higher in male than female adults. Seasonality had a distinct peak in September with Cryptosporidium more prevalent in the rainy (2.6%) than dry season (0.9%).

  18. Comparing feeding and reproductive parameters of Amblyomma parvum tick populations (Acari: Ixodidae) from Brazil and Argentina on various host species.

    Science.gov (United States)

    Gerardi, Monize; Martins, Maria Marlene; Nava, Santiago; Szabó, Matias Pablo Juan

    2013-10-18

    Amblyomma parvum is a Neotropical tick that is widely spread and a potential vector of pathogens, including Rickettsiae. Genetic differences are remarkable between A. parvum populations from Brazil and Argentina. In this work, feeding and reproduction parameters of A. parvum ticks from these two populations were compared on some key host species to evaluate possible differences in host suitability between them. On the whole parameters of these tick populations were similar when fed on the same host and varied similarly on different host species. Still, bovines were more suitable host for Argentinian larvae than for Brazilian cohorts. It was observed that guinea pigs were the best host A. parvum immatures from both origins, as depicted from higher recovery rate of larvae and heavier engorged nymph weights. Canids and bovids were host species most suitable to adults of both tick populations as shown by the highest number of larvae produced by adult females that engorged on these hosts. Taken together, results showed that in spite of the genetic divergence, A. parvum from Argentina and Brazil have similar biological performance on various host species.

  19. Molecular analysis of Cryptosporidium from cattle from five states of Peninsular Malaysia.

    Science.gov (United States)

    Yap, Nan Jiun; Koehler, Anson V; Ebner, Janine; Tan, Tiong Kai; Lim, Yvonne A L; Gasser, Robin B

    2016-02-01

    Despite the importance of the cattle industry in Malaysia, there are very few studies of the diversity and public health significance of bovine cryptosporidiosis in this country. In the present study, we used a PCR-based approach to detect and genetically characterize Cryptosporidium DNA in faecal samples from a cohort of 215 asymptomatic cattle (of different ages) from six farms from five states of Peninsular Malaysia. Cattle on four of the six farms were test-positive for Cryptosporidium, with an overall prevalence of 3.2%. Cryptosporidium bovis and Cryptosporidium ryanae were detected in two (0.9%) and five (2.3%) samples tested; this low prevalence likely relates to the age of the cattle tested, as most (73%) of the samples tested originated from cattle that were ≥2 years of age. Future studies should investigate the zoonotic potential of Cryptosporidium in pre-weaned and weaned calves in rural communities of Malaysia.

  20. Rapid Quantification of the Toxic Alga Prymnesium parvum in Natural Samples by Use of a Specific Monoclonal Antibody and Solid-Phase Cytometry

    DEFF Research Database (Denmark)

    West, N. J.; Bacchieri, R.; Hansen, Gert

    2006-01-01

    alga Prymnesium parvum in natural samples, using a specific monoclonal antibody and indirect immunofluorescence. The immunoglobulin G antibody 16E4 exhibited narrow specificity in that it recognized several P. parvum strains and a Prymnesium nemamethecum strain but it did not cross-react with P. parvum...... and enumeration of cultured P. parvum cells preserved with different fixatives showed that the highest cell counts were obtained when cells were fixed with either glutaraldehyde or formaldehyde plus the cell protectant Pluronic F-68, whereas the use of formaldehyde alone resulted in significantly lower counts....... Immunofluorescence labeling and analysis with the solid-phase cytometer of fixed natural samples from a bloom of P. parvum occurring in Lake Colorado in Texas gave cell counts that were close to those obtained by the traditional method of counting using light microscopy. These results show that a solid...

  1. Transport of Cryptosporidium, Giardia, Source-specific Indicator Organisms, and Standard Water Quality Constituents During Storm Events

    Science.gov (United States)

    Sturdevant-Rees, P. L.; Bourdeau, D.; Baker, R.; Long, S. C.; Barten, P. K.

    2004-05-01

    Microbial and water-quality measurements are collected during storm events under a variety of meteorological and land-use conditions in order to 1) identify risk of Cryptosporidium oocysts, Giardia cysts and other constituents, including microbial indicator organisms, entering surface waters from various land uses during periods of surface runoff; 2) optimize storm sampling procedures for these parameters; and 3) optimize strategies for accurate determination of constituent loads. The investigation is focused on four isolated land uses: forested with free ranging wildlife, beaver influenced forested with free ranging wildlife, residential/commercial, and dairy farm grazing/pastureland using an upstream and downstream sampling strategy. Traditional water-quality analyses include pH, temperature, turbidity, conductivity, total suspended solids, total phosphorus, total Kjeldahl-nitrogen, and ammonia nitrogen, Giardia cysts and Cryptosporidium oocysts. Total coliforms and fecal coliforms are measured as industry standard microbial analyses. Sorbitol-fermenting Bifidobacteria, Rhodococcus coprophilus, Clostridium perfringens spores, and Somatic and F-specific coliphages are measured at select sites as potential alternative source-specific indicator organisms. Upon completion of the project, the final database will consist of wet weather transport data for a set of parameters during twenty-four distinct storm-events in addition to monthly baseline data. A subset of the results to date will be presented, with focus placed on demonstrating the impact of beaver on constituent loadings over a variety of hydrologic and meteorological conditions.

  2. Origin of a major infectious disease in vertebrates: The timing of Cryptosporidium evolution and its hosts.

    Science.gov (United States)

    Garcia-R, Juan C; Hayman, David T S

    2016-11-01

    Protozoan parasites of the genus Cryptosporidium infect all vertebrate groups and display some host specificity in their infections. It is therefore possible to assume that Cryptosporidium parasites evolved intimately aside with vertebrate lineages. Here we propose a scenario of Cryptosporidium-Vertebrata coevolution testing the hypothesis that the origin of Cryptosporidium parasites follows that of the origin of modern vertebrates. We use calibrated molecular clocks and cophylogeny analyses to provide and compare age estimates and patterns of association between these clades. Our study provides strong support for the evolution of parasitism of Cryptosporidium with the rise of the vertebrates about 600 million years ago (Mya). Interestingly, periods of increased diversification in Cryptosporidium coincides with diversification of crown mammalian and avian orders after the Cretaceous-Palaeogene (K-Pg) boundary, suggesting that adaptive radiation to new mammalian and avian hosts triggered the diversification of this parasite lineage. Despite evidence for ongoing host shifts we also found significant correlation between protozoan parasites and vertebrate hosts trees in the cophylogenetic analysis. These results help us to understand the underlying macroevolutionary mechanisms driving evolution in Cryptosporidium and may have important implications for the ecology, dynamics and epidemiology of cryptosporidiosis disease in humans and other animals.

  3. Cryptosporidium and Giardia in Swimming Pools, Atlanta, Georgia

    Centers for Disease Control (CDC) Podcasts

    2008-05-29

    In this podcast, Dan Rutz speaks with Dr. Joan Shields, a guest researcher with the Healthy Swimming Program at CDC, about an article in June 2008 issue of Emerging Infectious Diseases reporting on the results of a test of swimming pools in the greater Atlanta, Georgia area. Dr. Shields tested 160 pools in metro Atlanta last year for Cryptosporidium and Giardia. These germs cause most recreational water associated outbreaks.  Created: 5/29/2008 by Emerging Infectious Diseases.   Date Released: 5/29/2008.

  4. Prevalence of Cryptosporidium and Giardia in the water resources of the Kuang River catchment, Northern Thailand.

    Science.gov (United States)

    Chuah, C Joon; Mukhaidin, Nabila; Choy, Seow Huey; Smith, Gavin J D; Mendenhall, Ian H; Lim, Yvonne A L; Ziegler, Alan D

    2016-08-15

    A catchment-scale investigation of the prevalence of Cryptosporidium and Giardia in the Kuang River Basin was carried out during the dry and rainy seasons. Water samples were collected from the Kuang River and its tributaries as well as a major irrigation canal at the study site. We also investigated the prevalence of gastrointestinal parasitic infection among dairy and beef cattle hosts. Cryptosporidium and/or Giardia were detected in all the rivers considered for this study, reflecting their ubiquity within the Kuang River Basin. The high prevalence of Cryptosporidium/Giardia in the upper Kuang River and Lai River is of a particular concern as both drain into the Mae Kuang Reservoir, a vital source of drinking-water to many local towns and villages at the research area. We did not, however, detected neither Cryptosporidium nor Giardia were in the irrigation canal. The frequency of Cryptosporidium/Giardia detection nearly doubled during the rainy season compared to the dry season, highlighting the importance of water as an agent of transport. In addition to the overland transport of these protozoa from their land sources (e.g. cattle manure, cess pits), Cryptosporidium/Giardia may also be re-suspended from the streambeds (a potentially important repository) into the water column of rivers during storm events. Faecal samples from dairy and beef cattle showed high infection rates from various intestinal parasites - 97% and 94%, respectively. However, Cryptosporidium and Giardia were only detected in beef cattle. The difference in management style between beef (freeranging) and dairy cattle (confined) may account for this disparity. Finally, phylogenetic analyses revealed that the Cryptosporidium/Giardia-positive samples contained C. ryanae (non-zoonotic) as well as Giardia intestinalis assemblages B (zoonotic) and E (non-zoonotic). With only basic water treatment facilities afforded to them, the communities of the rural area relying on these water supplies are

  5. Prevalência de Cryptosporidium serpentis em serpentes de cativeiro Prevalence of Cryptosporidium serpentis in captive snakes

    Directory of Open Access Journals (Sweden)

    Paulo Custório Ruggiero

    2011-11-01

    Full Text Available Cryptosporidium é um protozoário encontrado em uma grande variedade de espécies animais como responsável por casos de gastrite e enterite, porém com epidemiologia pouco conhecida em animais silvestres. A presente investigação teve como objetivo avaliar a prevalência de Cryptosporidium serpentis em lavado gástrico de serpentes mantidas em cativeiro no serpentário do Instituto Butantan (São Paulo, Brasil. A coleta foi realizada uma semana após alimentação, evitando, assim, a regurgitação devido à manipulação. Foram realizados esfregaços do sedimento do lavado gástrico, obtido por centrifugação, corados pela técnica de coloração de Kinyoun. Parte do sedimento foi submetido à técnica de RFLP-PCR para identificação da espécie de Cryptosporidium. O serpentário é dividido em três seções, por espécie - a primeira com oito jibóias (Boa constrictor amarali, a segunda com dez jararacas (Bothropoides jararaca e a última com sete cascavéis (Caudisona durissa. A prevalência de C. serpentis encontrada neste estudo para as serpentes C. durissa, B. jararaca e Boa c. amarali, foi de 57,14% (04/07, 40% (04/10 e 37,5% (03/08, respectivamente, revelando importante ocorrência desse protozoário em serpentes de cativeiro. Apesar da alta prevalência encontrada, apenas as jiboias apresentaram sintomas como perda de peso e regurgitação, refletindo uma sensibilidade diferente da espécie para C. serpentis.Cryptosporidium is a protozoan found in a wide variety of animal species which is responsible for gastritis and enteritis, but its epidemiology is poorly known in wild animals. The present investigation aimed to evaluate the prevalence of Cryptosporidium serpentis in gastric aspirate of captive snakes from the public serpentarium of the Butantan Institute (São Paulo, Brazil. Sampling was performed preferably one week after feeding, thereby preventing regurgitation due to manipulation. Smears were done from the gastric

  6. Effect of Ureaplasma parvum co-incubation on Chlamydia trachomatis maturation in human epithelial HeLa cells treated with interferon-γ.

    Science.gov (United States)

    Yamazaki, Tomohiro; Matsuo, Junji; Nakamura, Shinji; Oguri, Satoshi; Yamaguchi, Hiroyuki

    2014-08-01

    Chlamydia trachomatis is an obligate intracellular bacterium that causes a sexually transmitted disease. Ureaplasma parvum is commensal in the human genital tract, with a minimal contribution to urogenital infection. We have recently found that U. parvum has a significant effect on the presence of C. trachomatis in the genital tract of healthy women. We therefore assessed the effect of U. parvum co-incubation on C. trachomatis maturation from reticulate bodies (RBs) to elementary bodies (EBs) in HeLa cells in the absence or presence of interferon (IFN)-γ, which is a critical host defense factor. IFN-γ stimulation of viable U. parvum significantly prompted chlamydial growth with an increase in infectious particles, EBs, in HeLa cells. IFN-γ treatment of killed U. parvum had a similar effect on C. trachomatis maturation in HeLa cells. There was no change in expression of indoleamine 2,3-dioxygenase (IDO) in cultures of viable or killed U. parvum. We concluded that U. parvum co-incubation by IFN-γ helped C. trachomatis to mature from RBs to EBs in HeLa cells, independent of IDO expression. This suggests a novel survival strategy of C. trachomatis against IFN-γ exposure, prompting secondary infection of the genital mucosa, with possible clinical implications.

  7. Removal of Cryptosporidium sized particle under different filtration temperature, flow rate and alum dosing

    Institute of Scientific and Technical Information of China (English)

    XU Guo-ren; Fitzpatrick S. B. Caroline; Gregory John; DENG Lin-yu

    2007-01-01

    Recent Cryptosporidium outbreaks have highlighted concerns about filter efficiency and in particular particle breakthrough. It is essential to ascertain the causes of Cryptosporidium sized particle breakthrough for Cryptosporidium cannot be destroyed by conventional chlorine disinfection. This research tried to investigate the influence of temperature, flow rate and chemical dosing on particle breakthrough during filtration. The results showed that higher temperatures and coagulant doses could reduce particle breakthrough. The increase of filtration rate made the residual particle counts become larger. There was an optimal dose in filtration and was well correlated to ζ potential.

  8. [Progress on the application of Cryptosporidium infected animal models and in vitro cultivation].

    Science.gov (United States)

    Yin, Jian-Hai; Shen, Yu-Juan; Cao, Jian-Ping

    2010-10-30

    The genus Cryptosporidium is composed of protozoan parasites that infect epithelial cells in the microvillus border of the gastrointestinal tract of all classes of vertebrates, and cause severe diarrheal disease in a variety of neonatal animals, children and immunocompromised persons. Establishment of Cryptosporidium infected animal models and its in vitro cultivation system have established a good foundation for characterizing life cycle stage, exploring immunological mechanism, developing vaccines, screening and evaluating potential drugs, as well as assessing oocyst inactivation techniques. This paper reviews recent development and application of the Cryptosporidium infected animal models and its in vitro cultivation.

  9. Occurrence of Cryptosporidium spp., Giardia spp. and other pathogenic intestinal parasites in the Beberibe River in the State of Pernambuco, Brazil

    Directory of Open Access Journals (Sweden)

    Dayana Andrade de Freitas

    2015-04-01

    Full Text Available INTRODUCTION: Transmission of pathogenic protozoa and helminths by water is a serious public health problem. In this study, we analyzed the presence of these organisms in the Beberibe River in Pernambuco, Brazil. METHODS: Parasite analysis was performed using the Hoffman, Pons, & Janer method followed by centrifugation and preparation of slides by staining with acetic acid and Lugol's solution. Protozoan oocysts were isolated by the modified Ziehl Neelsen method. RESULTS: Cryptosporidium spp., Giardia spp. and other parasites were found in the Beberibe River. CONCLUSIONS: Sanitation companies must assess pathogenic intestinal parasites in water basins providing public water and subsequently develop improved treatment systems for removal of such parasites.

  10. Importância de Cryptosporidium spp. como causa de diarréia em bezerros Importance of Cryptosporidium spp. as a cause of diarrhea in calves

    Directory of Open Access Journals (Sweden)

    Francisco L.F. Feitosa

    2008-10-01

    Full Text Available Avaliou-se a presença de oocistos de Cryptosporidium spp. em amostras de fezes de 14 bezerros e de suas mães até a oitava semana pós parição. A maior taxa de excreção de oocistos foi verificada em bezerros com sete dias de idade. Das vacas, 42,8% foram positivas para Cryptosporidium no período pós-parto. Em outra etapa deste estudo, foram acompanhados 57 bezerros positivos para Cryptosporidium, com até 30 dias de idade, provenientes de 32 propriedades leiteiras, e estudouse o grau de eliminação dos oocistos com a possível ocorrência de diarréia. Em todos os animais positivos para Cryptosporidium foi pesquisada a presença de bactérias enteropatogênicas, vírus (Rotavirus e Coronavirus e protozoários (Eimeria spp..The aim of this research was to evaluate the shedding of Cryptosporidium spp. oocysts in fecal samples from 14 calves from one dairy farm, from birth until 60 days old and from cows until